Official journal website:

amphibian-reptile-conservation.org

Amphibian & Reptile Conservation

11(1) [Special Section]: 1-23 (e144).

Amphibians of Haut-Ogooue Province, southeastern Gabon

^Gregory F. M. Jongsma, 2 Elie Tobi, 3 Graham P. Dixon-MacCallum, 4 Abraham Bamba-Kaya,

4 Jean-Aime Yoga, 4 Jean-Daniel Mbega, 4 Jean-Herve Mve Beh, 5 Andrea M. Emrich, and 6 David C.

Blackburn

1 New Brunswick Museum, 277 Douglas Avenue, Saint John, NB, CANADA E2K 1E5 2 Center for Conservation and Sustainability, Smithsonian

Conservation Biolog)> Institute, CNG, Gabon Biodiversity Program 3 1756 Cambridge St. Halifax, NS, CANADA, B3H 4A9 Hnstitut de Recherches

Agronomiques et Forestieres, Libreville, GABON 5 141 Wentworth Ave. Saint John, NB, CANADA, E2L 2S7 ^Florida Museum of Natural History,

University of Florida Gainesville, Florida 32611, USA

Abstract. —We provide the most complete inventory to date of amphibians for Haut-Ogooue

province in southeastern Gabon. This inventory is based on an 11-day survey conducted in 2015

around two villages, Doumaye and Mboua, near the Gabon-Congo border and a previous survey in

Bateke Plateau National Park during 2011. We report 42 species of anuran amphibians (21 genera; 11

families) for Haut-Ogooue including 26 new species records for the province and two new country

records for Gabon (Afrixalus osorioi and Hyperolius balfouri). This work brings the total known

amphibian diversity in Gabon to 98 species.

Resume. —Nous fournissons dans cet article, I’inventaire le plus complet des amphibiens de la

province du Haut-Ogooue dans le sud-est du Gabon. Cet inventaire se fonde sur des recherches

menees durant 11 jours en 2015 autour de deux villages, Doumaye et Mboua, pres de la frontiere

Gabon-Congo et cedes menees en 2011 dans le pare national des Plateaux Bateke par Zimkus &

Larson (2013). Nous rapportons 42 especes d’amphibiens (21 genres, 11 families) pour le Haut-

Ogooue dont 26 nouvelles especes pour la province et trois nouvelles mentions pour le Gabon

(Afrixalus osorioi et Hyperolius balfouri). Ce travail porte a 98 especes le nombre total connu de la

diversity des amphibiens du Gabon.

Keywords. Africa, anuran, diversity, frogs, herpetofauna, savanna, forest

Citation: Jongsma GFM, Tobi E, Dixon-MacCallum GP, Bamba-Kaya A, Yoga J-A, Mbega J-D, Mve Beh J, Emrich AM, Blackburn DC. 2017. Amphib¬

ians of Haut-Ogooue Province, southeastern Gabon. Amphibian & Reptile Conservation 11(1) [Special Section]: 1-23 (e144).

NoDerivatives 4.0 International License, which permits unrestricted use for non-commercial and education purposes only, in any medium, provided

the original author and the official and authorized publication sources are recognized and properly credited. The official and authorized publication

credit sources, which will be duly enforced, are as follows: official journal title Amphibian & Reptile Conservation ; official journal website <amphibian-

reptiie-conservation.org>.

Received: 06 December 2016; Accepted: 13 September 2017; Published: 20 November 2017

Introduction

Despite its relative small size, Gabon hosts a number of

significant biogeographic features, including the Bateke

Plateau, the Ogooue River, and two hypothesized Pleis¬

tocene forest refugia (Crystal Mountains and Chaillu

Massif). The country is dominated by lowland rainforest,

interspersed with a forest-savanna mosaic. Despite top¬

ographic diversity and expansive pristine habitat, there

is a paucity of research that directly explores the influ¬

ence and interactions of these features on biodiversity. In

part, this is a result of incomplete sampling across most

of Gabon. Our report adds to the growing inventory of

Gabonese amphibians based on surveys of a region that

is underrepresented in natural history collections.

The earliest known amphibian and reptile specimens

from Gabon were collected by Charles Eugene Aubry-

Lecomte. A French civil servant and keen naturalist,

COrrespondence. *gregor.Jongs mafpgmail.com

Amphib. Reptile Conserv.

Aubry-Lecomte made an important collection along

the Gabonese coast for the Museum national d’Histoire

naturelle in Paris (Dumeril 1856; Beolens et al. 2011).

Between 1850 and 1854, he collected the first series

of Cycloderma aubryi (Dumeril 1856) and Leptopelis

aubryi (Dumeril 1856) as well as several new species

of birds and plants (Dumeril 1856; Beolens et al. 2011).

Around the same period (1851-1855), Henry Alexander

Ford, an American M.D, was stationed at Baraka Mission

in present day Fibreville to research malarial fever (Ford

1856). During this time, he collected reptiles for the Acad¬

emy of Natural Sciences of Philadelphia (now, of Drexel

University), including the type series of Poromera fordii

(Hallowell 1857) and Gerrhosaurus nigrolineatus (Hal-

lowell 1857). In 1855, Paul Belloni Du Chaillu became

the first westerner to explore the interior of Gabon (Du

Chaillu 1861). Funded by the Academy of Natural Sci¬

ences of Philadelphia, Du Chaillu collected large series

November 2017 | Volume 11 | Number 1 | e144

Jongsma et al.

Fig. 1. Map of Haut-Ogooue province, with the capital Franceville (yellow star). Sample sites from this study are indicated with

white circles. White squares denote study site for Zimkus and Larson (2013).

of birds and mammals (including the first intact gorilla

specimens) and the type series of Amnirana albolabris

(Hallowell 1856). At the turn of the 20 th Century, Ernest

Haug, a missionary for the Societe des missions evan-

geliques de Paris, conducted two methodical herpeto-

logical inventories approximately 50 km southwest of

Fambarene, Moyen-Ogooue for the Museum national

d’histoire Naturelle de Paris. This resulted in 29 reptile

species and 23 frog species (Mocquard 1897, 1902). Her-

petological work since the early 1900s has been sporadic

but there was an upsurge of inventory work around the

beginning of the 21 st Century (Burger et al. 2004; Burger

et al. 2006; Fretey and Blanc 2000; Fretey and Dewynter

1998; Knoepffler 1966, 1974; Fotters et al. 2000; Fot-

ters et al. 2001; Pauwels et al, 2004; Pauwels and Rodel

2007; Zimkus and Farson 2013), and Gabon’s known

amphibian diversity has increased substantially through

these recent efforts.

At the turn of the millennium, the country amphib¬

ian total for Gabon was 72 species (Fretey and Blanc

2000). Today, less than two decades later, there are now

96 known amphibian species in Gabon, including 94

Fig. 2. Species rarefaction curve based on amphibians

encountered in Haut-Ogooue province between April 21 st to

May 1 st , 2015.

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Amphibians of southeastern Gabon

Fig. 3. Different habitats sampled in Haut-Ogooue during 2015 field surveys. Lentic habitat in closed forest (A), lotic habitat with

closed forest (B, C), lentic habitat at edge of forest (D), lentic habitat in savanna (E), lentic habitat in disturbed open area (F),

savanna (G), lentic habitat open forest habitat.

frogs and two caecilians (Neil and Jongsma 2016; Evans

et al. 2015; Zimkus and Larson 2013; Fretey et al. 2011;

Bell et al. 2011; Burger et al. 2006; Pauwels and Rodel

2007; Pauwels 2016). All published amphibian invento¬

ries for Gabon are restricted to just six of the 13 national

parks (see summary Pauwels and Rodel 2007; Pauwels

2016). However, several recently discovered species

were described from outside parks, including Werneria

iboundji Rodel et al. 2004 and Leptodactylodon stevarti

Rodel and Pauwels 2003. Zimkus and Larson (2013) car¬

ried out the first survey of amphibians in Haut-Ogooue

Province in Bateke National Park (BNP) and reported

18 frog species (three unidentified), including four new

country records. Our recent survey, presented below,

reveals many additional species for the province and two

additional species records for Gabon. We hope that our

study will serve as a guide to students and researchers

undertaking future herpetofaunal work in both Gabon

and Haut-Ogooue Province.

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Jongsma et al.

Materials and Methods

Gabon is a small equatorial country (267,667 km 2 ) domi¬

nated by tropical moist forest (80% total land cover; Lee

et al. 2006). The dominant hydrological feature is the

Ogooue River basin. There are four seasons: a long rainy

season from January to May; a long cold dry season from

June to September; a short rainy season from October

to December; and a short dry season from December to

January. The average annual temperature is 26 °C (Lee

et al. 2006).

Haut-Ogooue Province (36,547 km 2 ) is located in

southeastern Gabon and is composed of three major geo¬

logical formations: the Chaillu Massif in the southwest,

the Franceville Basin in the northwest, and the Bateke

Plateau to the east. The Chaillu Massif and Bateke Pla¬

teau are unique in Gabon because of their geological his¬

tories and contemporary environments. The Chaillu Mas¬

sif is an ancient formation dating to >2 billion years ago

that hosts some of the highest elevation forests in Gabon,

including forest refugia (Sosef 1994; Vande weghe 2009).

The massif is dominated by forest but also hosts small

forest-savanna mosaics around the foothills that origi¬

nate in the Haut-Ogooue Province. The Bateke Plateau

has a sandy substrate and is dominated by large swaths

of savanna that are contiguous with plains in southern

Africa (Vande weghe 2009). The border of Haut-Ogooue

represents the boundary between three major watersheds:

the Kouilou-Niari River, the Congo River, and the Ogo¬

oue River. We conducted surveys at sites within the Ogo¬

oue Basin at the foothills of the Chaillu Massif. Zimkus

and Larson (2013) work was based at the Bateke Plateau,

also within the Ogooue Basin.

We conducted visual encounter surveys around two

villages: Doumaye (02.2402°S, 013.5812°E) on the

left side of the Ogooue River, and Mboua (02.1532°S,

013.6398°E) to the right side of the river. Both sites are

located in the administrative department of Lekoko in

Haut-Ogooue province. The village of Doumaye is dom¬

inated by savanna habitat with gallery forest associated

with rivers. The habitat around the village of Mboua con¬

sists of continuous gallery forest. We spent five survey

nights in Doumaye (21-25 April 2015) and six in Mboua

(26 April-1 May 2015). We typically worked between

19h00 to OOhOO each night, targeting forested streams

and rivers, and small still bodies of water (Fig. 3). Our

research in Haut-Ogooue Province focused on six spe¬

cies (Afrixalus dorsalis , Amnirana albolabris , Hypero-

lius olivaceus , H. ocellatus , Phrynobatrachus africa-

nus , and Scotobleps gabonicus) for a comparative phylo-

geographic study around the Ogooue River. We captured

other amphibians opportunistically.

All species encountered across both sites were pho¬

tographed alive and swabbed for chytrid fungus ( Batra-

chochytrium dendrobatidis ; Bd). Voucher specimens

were euthanized using an aqueous solution of MS-222,

Amphib. Reptile Conserv.

and a sample of liver tissue was removed and stored

in RNAlater, before preserving the whole specimen in

10% neutral-buffered formalin. Specimens are depos¬

ited at the California Academy of Sciences (CAS) in San

Francisco, California, Sam Noble Museum (OMNH) in

Norman, Oklahoma, and Gabon’s national collection in

Yenzi Camp, Gamba, Gabon. We refer to specimens in

Gabon’s collection using GFMJ field numbers. To deter¬

mine the extent to which our species sampling was com¬

prehensive, we constructed a rarefaction curve using the

rare curve function in the vegan package (Oskansen et al.

2013) for R (R Core Team 2013).

Laboratory work was conducted at the Florida

Museum of Natural History (FLMNH) by GFMJ. We

extracted genomic DNAfrom tissues (liver, muscle, or toe

clips) using Qiagen DNeasy Kits following their protocol

for animals. Using polymerase chain reaction (PCR), we

amplified a -762 base pair (bp) fragment of mitochon¬

drial DNA that encodes part of the mitochondrial ribo-

somal 16S gene (94 °C 30 s, 52 °C 30 s, 72 °C one min)

using 35 cycles and the oligonucleotide primers 16Sc

and 16Sd (Moriarty and Cannatella 2004). We used Exo-

SAP-IT (Affymetrixs) to purify all amplified PCR prod¬

ucts and then shipped this product for Sanger sequencing

at Genewiz Co. All sequences are deposited in GenBank

(accession numbers: MF537671-MF537697).

Species Accounts

AMPHIBIA-Froas

ARTHROLEPTIDAE

Arthroleptis cf. poecilonotus (Peters 1863)

Material: One (1) specimen. Doumaye: CAS 258166.

Fig. 4A.

Comments: Arthroleptis poecilonotus is a leaf-litter

species that is associated with forest habitats but also

found in wet savanna and near human habitations. It is

widespread across West and Central Africa and is likely

composed of several unnamed species (Blackburn 2008).

Populations in Central Africa, including eastern Nige¬

ria, Cameroon, Gabon, and Republic of Congo, referred

to A. poecilonotus are not conspecific with those identi¬

fied as the same species in western Africa (Blackburn et

al. 2010), though no taxonomic changes have yet been

made. This species was first reported for Gabon by Moc-

quard (1902; under Arthroleptis inguinalis) near Lam-

barene in Moyen-Ogooue province. It has since been

found in several national parks including: Bateke NP

(Zimkus and Larson 2013), Crystal Mountains NP (Let¬

ters et al. 2001), Ivindo National Park (Fretey and Blanc

2000), Loango NP (Burger et al. 2006), and Lope NP

(Fretey and Blanc 2001).

4 November 2017 | Volume 11 | Number 1 | e144

Amphibians of southeastern Gabon

Table 1. Amphibian species recorded for Haute-Ogooue Province. *=New provincial record. **=New country record. Habitats

include forest (F), open disturbed areas (O), savanna (S), and edge (ED). Microhabitats include leaf litter (LL), arboreal (AR), and

aquatic (AQ). Some species lacking microhabitat information were not collected by the authors.

Doumaye

Mboua

Zimkus and Larson 2013

Habitat

Microhabitat

ARTHROLEPTIDAE

Arthroleptis cf. poecilonotus

A. cf. sylvaticus

Astylosternus batesi*

Cardioglossa gracilis *

Leptopelis aubryi *

L. aubryioides*

L. calcar at us*

L. m ills on i *

L. not at us *

L. oc ell at us*

Scotobleps gabonicus*

BUFONIDAE

Sclerophrys gracilipes*

S. superciliaris*

CONRAUIDAE

Conraua crassipes*

DICROGLOSSIDAE

Hoplobatrachus occipitalis

HYPEROLIIDAE

Afrixalus dorsalis*

A. osorioi**

A. quadrivittatus

Cryptothylax greshoffi

Hyperolius adspersus

H. balfouri**

H. bolifambae

H. kuligae*

H. ocellatus*

F/ED

H. olivaceus*

H. par dal is*

S/ED

H. phantasticus*

Kassina maculosa

Opisthothylax immaculatus *

Phlycti mantis leonardi*

PHRYNOBATRACHIDAE

Phrynobatrachus africanus*

P. horsti (P. ruthbeateae)

PIPIDAE

Hymenochirus boettgeri

Xenopus pygmaeus

PT Y CHADENIDAE

Ptychadena perreti

P. taenioscelis

P. uzungwensis

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November 2017 | Volume 11 | Number 1 | el 44

Jongsma et al.

Table 1 (continued). Amphibian species recorded for Haute-Ogooue Province. *=New provincial record. **=New country record.

Habitats include forest (F), open disturbed areas (O), savanna (S), and edge (ED). Microhabitats include leaf litter (LL), arboreal

(AR), and aquatic (AQ). Some species lacking microhabitat information were not collected by the authors.

Doumaye

Mboua

Zimkus and Larson 2013

Habitat

Microhabitat

PYXICEPHALIDAE

Aubria masako*

RANIDAE

Amnirana albolabris

A. amnicola*

A. lepus*

RHACOPHORIDAE

Chiromantis rufescens

Arthroleptis cf. sylvaticus (Laurent 1954)

Material: Six (6) specimens. Doumaye: CAS 258184;

GFMJ 1327. Mboua: CAS 258166, 258241-42; OMNH

44767. Fig. 4B.

Comments: Arthroleptis sylvaticus is a leaf-litter

species that is widespread across Central Africa, north

of the Congo River and is a complex of several unde¬

scribed species. This species is known from the follow¬

ing national parks: Bateke (Zimkus and Larson 2013),

Ivindo (Fretey and Blanc 2000), Loango (Burger et al.

2006), Lope (Fretey and Blanc 2001), and Moukalaba-

Doudou (Burger et al. 2004).

Astylosternus bate si (Boulenger 1900)

Material: Eighteen (18) specimens. Doumaye: CAS

258139, 258151, 258211-12; GFMJ 1240, 1242, 1270,

GFMJ 1322-23,1372. Mboua: CAS 258285-86; OMNH

44768-69; GFMJ 1397, 1461, 1514-15. Fig. 4C, D.

Comments: Astylosternus batesi is a leaf-litter species

that is strongly associated with forested streams. Individ¬

uals are best detected by eye-shine at night. This species

is distinguishable from the closely related and sympatric

species Scotobleps gabonicus by its smoother skin. It is

known from three national parks: Crystal Mountains NP

(Potters et al. 2001), Ivindo NP (Fretey and Blanc 2000),

and Moukalaba-Doudou NP (Burger et al. 2004).

Cardioglossa gracilis (Boulenger 1900)

Material: Ten (10) specimens. Doumaye: CAS 258182-

83, 258197, 258208, 258210; GFMJ 1326. Mboua: CAS

258227, 258251; OMNH 44770-71. Fig. 4E

Comments: Male C. gracilis are typically found call¬

ing from leaf litter within 10 meters of forested streams

with slopped sides. Their call is an insect-like click.

Males are faithful to their calling sites (GFMJ, pers.

obs.); if disturbed, they will vacate the calling site, but

then return to the same spot a short while later. This spe¬

cies was first collected in neighboring Equatorial Guinea

by the ornithologist George F. Bates along the Benito

River (Boulenger 1900). Cardioglossa gracilis is also

known from Ivindo NP (Fretey and Blanc 2000) and

Moukalaba-Doudou NP (Burger et al. 2004). We also

collected voucher specimens from Mitone village near

Fambarene (000.64375°S, 010.22071°E; CAS 258016)

and Madoukou village near Fastoursville (00.86831°S,

12.67244°E; GFMJ 1583).

Leptopelis aubryi (Dumeril 1856)

Material: Six (6) specimens. Doumaye: CAS 258202,

258260-61; GFMJ 1470-71, 1473. Fig. 4F.

Comments: Leptopelis aubryi was originally col¬

lected by Charles Eugene Aubry-Fecomte in the early

1850s and is among the first amphibians ever collected

in Gabon (Dumeril 1856). We encountered all individu¬

als in tall grass in ditches and around well pumps in Dou¬

maye. It is a disturbance specialist. In the Plaine Ouanga

Reserve in the Gamba Complex of Protected Areas

(GCPA) in the Ogooue-Maritime province, one of us (E.

Tobi) regularly finds this species on the branches of trees

at the edge of forest and in the forest. It is widespread

across Central Africa, North of the Congo River. Within

Gabon voucher specimens are known from the follow¬

ing national parks: Crystal Mountains NP (Fotters et al.

2001), Ivindo NP (Fretey and Blanc 2000), Foango NP

(Burger et al. 2006), Pope NP (Fretey and Blanc 2001),

and Moukalaba-Doudou (Burger et al. 2004).

Leptopelis aubryioides (Andersson 1907)

Material: One (1) specimen. Mboua: CAS 258234. Fig.

4G, H.

Comments: A single individual was encountered near

a small forest stream next to a foot path. This species is

easily distinguished from similar species by the distinct

spurs on its heels and its small size (Amiet 2012). The

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Amphibians of southeastern Gabon

Fig. 4. Arthroleptis cf. poecilonotus CAS 258166 (A), A. sylvaticus CAS 258184 (B), Astylosternus batesi CAS 258139, GFMJ

1240 (C, D), Cardioglossa gracilis CAS 258016 (E), Leptopelis aubryi 258261 (F), L. aubryioides CAS 258234 (G, H).

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Jongsma et al.

similar sympatric species, L. aubryi, lacks heel spurs and

is found in more disturbed habitats. This species is known

across Cameroon (Amiet 2012) but few records exist for

Gabon. Within Gabon, it is known from: Ivindo NP (Bell

et al. 2011), Loango NP (Burger et al. 2006, listed as L.

omissus ), and Lope NP (Fretey and Blanc 2001 listed as

L. omissus). We sequenced 16S rRNA of CAS 258234,

and confirmed its identification using BLAST (GenBank

accession: MF537690; nearest GenBank sequence is

KT967084.1; 96% identical).

Leptopelis calcaratus (Boulenger 1906)

Material: Seven (7) specimens. Doumaye: CAS 258148,

258190-91,258274, GFMJ 1340. Mboua: CAS 258253-

54. Fig. 5A.

Comments: Leptopelis calcaratus is an arboreal spe¬

cies that is widespread across Central Africa (Cameroon,

Gabon, Republic of Congo, and DRC). Within Gabon,

it is known from Crystal Mountains NP (Lotters et al.

2001), Ivindo NP (Fretey and Blanc 2000), and Mouka-

laba-Doudou (Burger et al., 2004). We encountered six of

the seven individuals perched in trees above four meters

from the ground adjacent to forested streams. It is easily

identified by the spur on its heel and larger size than L.

aubryioides .

Leptopelis millsoni (Boulenger 1895)

Material: Three (3) specimens. CAS 258147, OMNH

44774. Mboua: CAS 258233. Fig. 5B, C.

Comments: We found individuals 1-2.5 m above the

ground along forested streams. This species is closely

associated with streams. This arboreal species is wide¬

spread across Central Africa (Cameroon, Gabon, Repub¬

lic of Congo, and DRC). Within Gabon, it is known

from Crystal Mountains NP (Lotters et al. 2001; Bell

et al. 2011), Ivindo NP (Fretey and Blanc 2000; Bell et

al. 2011), and Moukalaba-Doudou (Burger et al. 2004).

In addition to this new Haut-Ogooue record, we also

found this species in Moyen-Ogooue (CAS 257990-91,

258049, 258076, 258119, 258126) and Ogooue-Lolo

(CAS 258303) provinces.

Leptopelis notatus (Peters 1875)

Materials: Five (5) specimens. Mboua: CAS 258230-

32, 258283; GFMJ 1405. Fig. 5D-F.

Comments: This arboreal species is widespread across

Central Africa and we found it near streams 1-2 m above

the ground. We encountered three females and two males.

The females were all uniformly green, and the males

were mottled green and light brown. Leptopelis nota¬

tus is known from the following national parks: Crystal

Mountains NP (Lotters et al. 2001), Ivindo NP (Fretey

and Blanc 2000), and Moukalaba-Doudou (Burger et al.

2004).

Leptopelis ocellatus (Mocquard 1902)

Material: Nine (9) specimens. Doumaye: CAS 258189,

258196; GFMJ 1337. Mboua: CAS 258252; GFMJ 1422,

1431, 1456-57; OMNH 13751. Fig. 5G.

Comments: Leptopelis ocellatus is associated with

slow rivers and forested swamps. They were found

between 10 cm and one m above the ground or water.

Across Gabon, this species is known from Ivindo NP

(Fretey and Blanc 2000) and Moukalaba-Doudou NP

(Burger et al. 2004). We also encountered this species

around Junkville, Moyen-Ogooue (CAS 258134-35) and

around Ogooue-cinq and Madoukou villages near Last-

oursville, Ogooue-Lolo (CAS 258287, 258306, 258316).

Scotobleps gabonicus (Boulenger 1900)

Material: Twelve (12) specimens. Doumaye: CAS

258149, 258150; GMFJ 1239, 1267-68, 1350, 1367.

Mboua: CAS 258228-29; GFMJ 1399, 1401-02. Fig.

5H.

Comments: Scotobleps gabonicus is found in leaf lit¬

ter, close to stream edges with sandy to pebbly substrates.

While found near streams, we never observed individu¬

als {n = 84 across Gabon) to leap into the water when

approached; when detected, it either remains in place or

moves in a direction other than the stream. This species

appears to prefer clear streams as we did not find it near

sections with muddy water. This may suggest that its

reproduction and life history are dependent on specific

stream qualities, though its tadpoles remain unknown.

Adults are best detected at night by eye-shine. Scoto¬

bleps gabonicus is widespread and common across the

lower Guinean forest (Cameroon, Equatorial Guinea, and

Gabon; Portik et al. 2017) In Gabon, it is known from

Crystal Mountains NP (Lotters et al. 2001), Lope NP

(Fretey and Blanc 2001), Ivindo NP (collected by Bell

and Stuart in 2011; NCSM 78914-15), and Moukalaba-

Doudou NP (Burger et al. 2004).

BUFONIDAE

Sclerophrys gracilipes (Boulenger 1899)

Material: Five (5) specimens. Doumaye: CAS 258175.

Mboua: CAS 258257, 258282; OMNH 44780; GFMJ

1506. Fig. 6A.

Comments: This is a common terrestrial species in

lowland forests. All individuals were encountered associ¬

ated with small to medium-sized, forested streams. This

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Amphibians of southeastern Gabon

Fig. 5. Leptopelis calcareous CAS 258148 (A), L. millsom CAS 257991, 257990 (B, C), L. notatus GFMJ 1495, CAS 258283,

258230 (D-F), L. ocellatus GFMJ 1431 (G), Scotobleps gabonicus GFMJ 1350 (H).

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Jongsma et al.

species is widespread across Central Africa, north of the

Congo River. In Gabon, this species is known from Ivindo

NP (Fretey and Blanc 2000), Loango NP (Burger et al.

2006), and Moukalaba-Doudou NP (Burger et al. 2004).

In 2015 we also collected vouchers in Moyen-Ogooue

province near Lambarene (CAS 257988, 257994) and

Ndjole (CAS 258051, 258096). This species was col¬

lected along Onangue Lake in 2013 (by R. Bell and B.

Stuart; CAS 254506-10).

Sclerophrys superciliaris (Boulenger 1888)

Material: One (1) specimen. Doumaye: GFMJ 1314.

Fig. 6B, C.

Comments: One individual was brought to our camp

by a local civil servant, Eric Dipanda, who found the

toad on the main dirt road running through Doumaye

(02.23373°S, 013.60008°E). This is the first provincial

record and most southern record for the species (Barej

et al. 2011), although Fretey and Blanc (2000) reports

the species from the Republic of Congo without precise

locality information. It is best detected by its bright eye-

shine in the dark (DCB, pers. obs.).

CONRAUIDAE

Conraua crassipes (Buchholz and Peters 1875)

Material: Two (2) specimens. Mboua: CAS 258277-78.

Fig. 6D, E.

Comments: Conraua crassipes is closely associated

with forested small tributaries of the Ogooue River. Indi¬

viduals were found in shallow muddy substrate near the

banks and detected by their eye-shine. This species is

known from the following national parks: Crystal Moun¬

tains (Lotters et al. 2001), Ivindo (Fretey and Blanc 2001),

Lope (Fretey and Blanc 2000), and Moukalaba-Doudou

(Burger et al. 2004). In 2015, we also encountered this

species near Lambarene (CAS 257993, 258019), Ndjole

(CAS 258084), Junkville (CAS 258122-23), and Last-

oursville (CAS 258317).

DICROGLOSSIDAE

Hoplobatrachus occipitalis (Gunther 1858)

Material: Three (3) specimens. Doumaye: CAS 258174;

OMNH 44786; GFMJ 1294. Fig. 6F.

Comments: This highly aquatic species is associated

with permanent bodies of water in savanna habitat and

tolerant of disturbance (Guibe and Lamotte 1958; Rodel

2000). We encountered this species at pristine savanna

lakes as well as disturbed permanent pools in the village

of Bafounou. This species is widely distributed across

Africa, from Senegal to Ethiopia and south to Zambia

and Angola (Rodel 2000). Within Gabon, H. occipitalis

is known from: Bateke NP (Zimkus and Larson 2013),

Crystal Mountains NP (Lotters et al. 2001), and Loango

NP (Burger et al. 2006). It is also known from the follow¬

ing areas: Ivindo, Rougier Gabon Forestry Concession

(NCSM 78971-74) and from the Ouanga Plains, Basse-

Banio in Nyanga province (USNM 580613-17).

HYPEROLIIDAE

Afrixalus dorsalis (Peters 1875)

Material: Five (5) specimens. Doumaye: CAS 258200,

258201, 258240, 258284; GFMJ 1353. Fig. 6G.

Comments: Afrixalus dorsalis is a disturbance special¬

ist, and we found individuals concentrated near village

water pumps, calling from tall grasses. This species was

found in sympatry with Leptopelis aubryi. Its identifica¬

tion is based on the key by Fretey et al. (2011), includ¬

ing the brown mediodorsal band that widens and spreads

towards the eyelids (see Fig. 6G). Afrixalus dorsalis is

known from the following national parks: Ivindo, Lope

(Fretey and Blanc 2000, 2001), Loango, Moukalaba-

Doudou (Burger et al. 2004, 2006).

Afrixalus osorioi (Ferreira 1906)

Material: Twenty one (21) specimens. Doumaye: CAS

258160, 258161, 258262-70; OMNH 44788; GFMJ

1356-58, 1475-78, 1531-32. Fig. 6H, Fig. 7A, B.

Comments: Afrixalus osorioi is similar in appearance

and habitat preference to A. dorsalis , but distinguished

from that species by its distinct advertisement call. Afrix¬

alus osorioi is known from Angola, Republic of Congo,

Democratic Republic of Congo, Kenya, and Uganda.

These specimens represent the first country records for

Gabon. This species inhabits bushland habitat. It has a

unique pattern that helps distinguish it from other Gab¬

onese Afrixalus , typically with a rectangular dark dor¬

sal spot and a narrow fight dorsal pattern extends to the

anus (Schiotz 1999). Laurent (1982) mentions that this

pattern does not vary within populations; however, we

encountered some variation, including individuals that

possessed no dark rectangle at all (Fig. 6, 7). The identi¬

fication of these specimens was confirmed by comparing

DNA sequence data for 16S ribosomal RNA from these

specimens to another identified sample (K. Charles and

D. Portik, unpubl. data; CAS 256140).

Afrixalus quadrivittatus (Werner 1908)

Material: Three (3) specimens. Doumaye: CAS 258271-

72, GFMJ 1492. Fig. 7C.

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Fig. 6. Sclerophiys gracilipes CAS 258282 (A), Sclerophiys superciliaris GFMJ 1314 (B. C), Conraua crassipes ORB 97 (D, E),

Hoplobatrachus occipitalis CAS 258278 (F), Afrixalus dorsalis ORB 140 (G) A. osorioi GFMJ 1356 (H).

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Jongsma et al.

Comments: This species is found in open grassy habi¬

tat. In Doumaye, we found individuals in a flooded area

near the football stadium (02.23300°S, 13.60305°E),

calling from within bunches of grasses between 10-50

cm above the ground or shallow water. At this site, A.

quadrivittatus is sympatric with A. osorioi, Hoplobatra-

chus occipitalis , and Phlyctimantis leonardi. This species

is known from Bateke NP (Zimkus and Larson 2013).

We sequenced 16S rRNA of CAS 258271 and GFMJ

1492, and confirmed their identification using BLAST

(GenBank accession: MF537696-97; nearest GenBank

sequence is KF 178889.1; 99% identical).

Cryptothylax greshoffi (Schilthuis 1889)

Material: Eleven (11) specimens. Mboua: CAS 258214-

216; OMNH 44789-90; GFMJ 1430, 1442^16. Fig. 7D.

Comments: We found C. greshoffi in open grassy

habitat bordered by forest at a dammed section of the

stream where locals did laundry. At night, individuals

were found calling within the vegetation surrounding

the water, ranging from 5 to 100 cm above the water.

This species is known from Ivindo NP (Fretey and Blanc

2001) and Bateke Plateau NP (Zimkus and Larson 2013).

Hyperolius adspersus (Peters 1877)

Material: Twelve (12) specimens. Doumaye: CAS

258167-69, 258332-33, 258170; GFMJ 1303-06. Fig.

7E-G.

Comments: We found H. adspersus in open, natural

savanna habitat. Individuals called in high densities from

grasses in and at the edge of shallow ponds. This spe¬

cies was previously considered a part of the Hyperolius

nasutus complex (Channing et al. 2013). This species is

known from Bateke Plateau NP, Loango NP (Burger et

al. 2006), Moukalaba-Doudou NP (Burger et al. 2004),

and Pongara NP (Pauwels 2016). It is also known from

Libreville (Gratwick et al. 2011).

Hyperolius balfouri (Werner 1908)

Material: Four (4) specimens. Doumaye: CAS 258171-

73; GFMJ 1307. Fig. 8E.

Comments: We encountered this species at one site,

a pond in savanna habitat near Doumaye (Fig. 3). It

was found in open savanna calling from the edge of a

pool. Hyperolius adspersus also occurred at this same

site. This is a new country record for Gabon. It is dis¬

tinguishable from other Gabonese species based on its

green dorsolateral lines and the presence of a vertebral

stripe (Fig. 8). The nearest known population is 715-km

north in Cameroon (Amiet 2012) and represented by the

subspecies H. b. viridistriatus. However, based on our

16S sequence data, this is more closely related to popula¬

tions in East Africa, including ~2,180 km east in Mabira,

Uganda (CAS 256187). We sequenced the 16S rRNA of

CAS 258171-73 and GFMJ 1307 (GenBank accession:

MF537676-79), and confirmed their identification using

an unpublished dataset (Portik; 100% identical to CAS

256187).

Hyperolius kuligae (Mertens 1940)

Material: Four (4) specimens. Doumaye: CAS 258238-

39, 258247, GMFJ 1418. Fig. 8F, G.

Comments: These individuals were found within

closed forest on vegetation surrounding a still section

of river that was dammed for manioc fermentation. The

males exhibited a bright yellow coloration at night, which

helps distinguish it from similar species like H. platyceps

(Amiet 2012). This species is known from Ivindo NP

(Bell et al. 2011), Loango NP (Burger et al. 2006), and

Moukalaba-Doudou (Burger et al.2004).

Hyperolius ocellatus (Gunther 1858)

Material: Ten (10) specimens. Doumaye: CAS 258142,

258176-77, GFMJ 1246, 1248. Mboua: CAS 258237,

GFMJ 1411, 1414-15, 1417. Fig. 8H, 9A-C.

Comments: This dichromatic species is found in the

forest or at the forest-edge, typically on leaves 1-2 m

above the ground and near slow sections of streams.

Males are green with light dorsolateral stripes and a

pale triangle on the snout that is diagnostic of the spe¬

cies. Females range in coloration from silvery grey to red

with black spots (Schiotz 1999; Amiet 2012). Hypero¬

lius ocellatus is widespread and common across Central

Africa, including Gabon. To date, it is known from four

national parks: Ivindo (Bell et al. 2011), Loango (Burger

et al. 2006), Moukalaba-Doudou (Burger et al. 2004), and

Crystal Mountains (Bell et al. 2011). It was also encoun¬

tered near Mitone (CAS 258083; -0.641950 10.217420)

and Ndjole (CAS 257997; -0.193950, 10.784770) in

Moyen-Ogooue, and from Basse-Bania department in

Nyanga province (USNM 558547; -3.23331, 10.619).

Hyperolius olivaceus (Peters 1876)

Material: Ten (10) specimens. Doumaye: CAS 258158-

59; GFMJ 1489-91. Mboua: CAS 258217-18; GFMJ

1380-82. Fig. 7H.

Comments: This disturbance specialist is common

across Gabon. This species was until recently consid¬

ered a subspecies of the very widespread Hyperolius

cinnamomeoventris. However, it was recently elevated

based on molecular, ecological, and phenotypic data

(Bell et al. 2017). It is known from Loango (Burger et

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Amphibians of southeastern Gabon

Fig. l.Ajrixalus osorioi CAS 258270, CAS 258161 (A, B), A. quadrivittatus GFMJ 1492 (C), Cryptothylax greshoffi OMNH 44789

(D), Hyperolius adspersus CAS 258168, 2583332, GMFJ 1304 (E-G), H. olivaceus CAS 258159 (H).

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Jongsma et al.

al. 2006), Moukalaba-Doudou (Burger et al., 2004), and

from Libreville (Gratwick et al. 2011). We encountered

individuals around artificial bodies of water in grasses

and palm fronds, but never within closed canopy for¬

ests. We sequenced 16S rRNAfrom GFMJ 1489-91 and

confirmed their identification using BLAST (GenBank

accession: MF537693-95; closest GenBank sequence is

MF376266; 100% identical).

Hyperolius pardalis (Laurent 1948)

Material: Twenty (20) specimens. Doumaye: CAS

258162, 258203-06, OMNH 44794-99, GFMJ 1281,

1363. Mboua: CAS 258219-20, GFMJ 1447-51. GFMJ

Fig. 9D-H.

Comments: We encountered H. pardalis in disturbed

areas. One female was found on a tent, where we camped

in a field in Doumaye. All other individuals were encoun¬

tered in vegetation next to a small fish pond, adjacent

to a forested stream. Of 17 males encountered, two

males demonstrated the F-phase representing 11.8% of

the male population sampled (Fig. 6 D-H). This species

can be distinguished from the similar species Hyperolius

bolifambae by its distinct vocal sac, which is pearl-white

posterior to the gular gland (Amiet 2012). In Gabon,

Hyperolius pardalis is known from Crystal Mountains

NP (Lotters et al. 2001), Rabi-Toucan (Burger et al.

2006), and Ivindo NP (Fretey and Blanc 2001).

Hyperolius phantasticus (Boulenger 1899)

Materials: Three (3) specimens. Doumaye: CAS

258163-65. Fig. 10A-C.

Comments: Hyperolius phantasticus was found in

small trees next to a large savanna pond, 2-2.5 meters

above the ground. In the Plaine Ouanga Reserve in the

Gamba Complex of Protected Areas, this species was

found in shrubs close to or next to savanna ponds. This

species is known from Crystal Mountains NP (Lotters et

al. 2001; Bell et al.2011) and Loango NP (Burger et al.

2006). We sequenced 16S rRNAof CAS 258165 and con¬

firmed its identification using BLAST (GenBank acces¬

sion: MF537674; closest BLAST sequence is FJ594099;

97% identical).

Opisthothylax immaculatus (Boulenger 1903)

Material: One (1) specimen. Mboua: CAS 258235. Fig.

10D.

Comments: This monotypic genus is distinguished

from other Hyperoliidae by the combination of its verti¬

cal pupils, rough skin, and orange color (Schiotz 1999).

The males have large gular glands and non-descendible

vocal sacs (Schiotz 1999). This arboreal species builds

foam nests (Amiet 1991). This species was encountered

at night on a stem one m above the ground, between a

forested stream and an elephant wallow. In Gabon, it is

known from Ivindo NP (Bell et al. 2011) and Rabi-Tou-

can (Burger et al. 2006). We sequenced 16S rRNA of this

individual and confirmed its identification using BLAST

(GenBank accession: MF537682; the most similar Gen¬

Bank sequence is KX492629; 98% identical).

Phlyctimantis leonardi (Boulenger 1906)

Material: Three (3) specimens. Doumaye: CAS 258209,

258273; GFMJ 1369. Fig. 10E, F.

Comments: This large hyperoliid frog was encountered

in shrubs or trees, 1-2 m above the ground and near still

bodies of water in open habitat. In Gabon, P. leonardi is

common and widespread. It is known from the following

national parks: Crystal Mountains (Lotters et al. 2001),

Ivindo, Lope (Fretey and Blanc 2000, 2001), Loango,

and Moukalaba-Doudou (Burger et al. 2004, 2006).

It is also known from Basse-banio, Nyanga (USNM

580612; -3.0876, 10.4285), Junkville (CAS 258130-31;

-0.051710, 11.166210), and nearNdjole, (CAS 258063-

65; -0.18482, 10.77727) in Moyen-Ogooue.

PHRYNOBATRACHIDAE

Phrynobatrachus africanus (Hallowell 1868)

Material: Thirteen (13) specimens. CAS 258187-88,

GFMJ 1332-34. Mboua: CAS 258222, 258243, 1425,

1427-28, 1463-65. Fig. 11 D-E.

Comments: The distantly related genera Phrynoba¬

trachus and Arthroleptis have often proved difficult for

field researchers to distinguish. The most reliable diag¬

nostic feature is the presence of a tubercle roughly in

the middle of the tarsus in Phrynobatrachus in addition

to both an inner and outer metatarsal tubercle (Zimkus

and Blackburn 2008). Phrynobatrachus africanus is a

common forest species found in the leaf litter and easily

identified by its rugose skin, yellow legs, and in males

both a large flat nuptial pad and odontoid processes. We

also encountered individuals with red legs in sympatry

with the yellow-legged individuals (Fig. 11), but these

morphotypes were confirmed as conspecific using 16S

rRNA sequences (100% similarity). It is widely distrib¬

uted across Gabon and known from Ivindo NP (Fre¬

tey and Blanc 2001), Loango NP (Burger et al. 2006),

Lope (Fretey and Blanc 2000), and Moukalaba-Doudou

(Burger et al. 2004). We sequenced 16S rRNA of CAS

258243, GFMJ 1332-34, 1425, 1427-28 and confirmed

their identifications using BLAST (GenBank accession:

MF537671-73, MF537675, MF537680-81, MF537685-

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Amphibians of southeastern Gabon

Fig. 8. Hyperolius olivaceus GFMJ 1381, CAS 258218, GFMJ 1181, CAS 258108 (A-D), H. balfouri GFMJ 1307 (E), H. kuligae

CAS 358247 (F, G), H. ocellatus CAS 258177 (H).

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Jongsma et al.

89, MF537691; most similar to GenBank sequence

GU457531; 93% identical).

Phrynobatrachus horsti (Rodel, Burger, Zassi-

Boulou, Emmrich, Penner, and Barej 2015)

Material: One (1) specimen. GFMJ 1332.

Comments: We encountered a single adult individual in

leaf litter in the forest. Rodel et al. (2015) proposed that

this species occurs in Bateke Plateau NP and that Zimkus

and Larson (2013) misidentified P. ruthbeateae , which is

endemic to Cameroon (Rodel et al. 2012). We sequenced

16S rRNA of GFMJ 1332 and confirmed their identifica¬

tions using BLAST (GenBank MF537676; most similar

sequence on GenBank is KR827534; 99% identical).

PYXICEPHALIDAE

Aubria masako (Ohler and Kazadi 1990)

Material: One (1) specimen. Mboua: CAS 258250.

Comments: We collected a single male individual from

a muddy pool adjacent to a forested stream in Mboua.

Typical of this species, the individual was skittish and

required multiple search events over two nights to cap¬

ture successfully. Aubria masako is widespread in Cen¬

tral Africa, including specifically the Congo Basin (Ohler

and Kazadi 1990). This individual was identified based

on 16S rRNA (GenBank MF537692; most similar to

GenBank sequence is KU560021; 99% identical).

RANIDAE

Amnirana albolabris (HaNowell 1856)

Material: Nine (9) specimens. Doumaye: CAS 258140,

258146, GFMJ 1260-61. Mboua: CAS 258248-49,

GFMJ 1436-37, 1504. Fig. 10G.

Comments: Amnirana albolabris is typically found

on vegetation (-0.2-1.5 m above the ground) near

still water within the forest, for example around man¬

ioc fermentation sites or elephant wallows. Individuals

are easily spotted by eye-shine. The type series of this

species was collected by Du Chaillu between 1855 or

1856 north of the Ogooue River in Gabon (Du Chaillu,

1861; Hallowell 1856). This species is common and

widespread across Central Africa, and within Gabon

it is known from Bateke Plateau NP (Zimkus and Lar¬

son 2013), Mayumba NP (USNM 2013), Loango NP

(Burger et al. 2006), Lope NP, Ivindo NP, (Fretey and

Blanc 2000, 2001), and Moukalaba-Doudou (Burger et

al. 2004). There are also records from Lac Oguemoue

(CAS 254595-98; -1.1001600, 10.02999983), Mitone

(CAS 257980; -0.641950, 10.217420), Junkville (CAS

258116; -0.062160, 11.15870), and near Ndjole (CAS

258085; -0.193950, 10.784770).

Amnirana amnicola (Perret 1977)

Material: Eight (8) specimens. Doumaye: CAS 258179-

81, 258192, GFMJ 1247, 1262. Mboua: CAS 258244.

Fig. 10H, Fig. 11 A.

Comments: We frequently encountered this species

near slow sections of forested streams on stems 1-2 m

above the ground. Amnirana amnicola is often in found

sympatry with the morphologically similar and related

species A. albolabris from which it is distinguishable by

less webbing between the toes (Perret 1977). This spe¬

cies is known from Crystal Mountains NP (Letters et al.

2001), Ivindo NP (Fretey and Blanc 2001), and Mouka¬

laba-Doudou NP (Burger et al. 2004).

Amnirana lepus (Andersson 1903)

Material: Fifteen (15) specimens. Doumaye: CAS

258143—45, GFMJ 1250-52, 1255-57. Mboua: CAS

258224-26, 258279-81, GFMJ 1394. Fig. 11B, C.

Comments: This large arboreal species is commonly

found along forested streams at night on vegetation 0.5-

1.5 meters above the ground. A single individual was

encountered on a rock in the center of a stream. Amni¬

rana lepus releases a pungent sour odor when captured,

which smells like vinegar. H-W Hermman observed

other species kept in the same container with A. lepus

from western Cameroon died (HWH, pers. comm.). This

is possibly due to the presence of peptides (Daly et al.

2004). This species is known from Ivindo NP (Fretey and

Blanc 2001), Moukalaba-Doudou (Burger et al. 2004),

and near Ndjole (CAS 258115; -0.193950, 10.784770).

RHACOPHORIDAE

Chiromantis rufescens (Gunther 1869)

Material: One (1) specimen. Mboua: CAS 258245. Fig.

11H.

Comments: Similar to C. rufescens encountered else¬

where in Gabon, we encountered this single male indi¬

vidual near small, temporary pools in the forest. This

species is typically found on branches 1-3 m above the

ground and is widespread across West and Central Africa.

Within Gabon, C. rufescens is known from six national

parks: Crystal Mountains (Bell et al. 2011), Ivindo (Fre¬

tey and Blanc 2001), Loango (Burger et al. 2006), Lope

(Fretey and Blanc 2000), Moukalaba-Doudou (Burger et

al. 2004), and Bateke Plateau NP (Zimkus and Larson

2013).

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Amphibians of southeastern Gabon

Fig. 9. Hyperolius ocellatus CAS 258142, GFMJ 1184 (A-C), H. pardalis GFMJ 1281, CAS 258204, OMNH 44797, CAS 258162

(D-H).

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Jongsma et al.

Fig. 10. Hyperoliusphantasticus CAS 258165, 358163 (A-C), Opistothylax immaculatus CAS 258235 (D), Phlyctimcmtis leonardi

CAS 258237, 2558071 (E, F), Amnirana albolabris ORB 291 (G), A. amnicola CAS 258244 (H).

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Amphibians of southeastern Gabon

Fig. 11. Amnirana anmicola CAS 258224 (A) A. lepus CAS 258144, 258224 (B, C), Phrynobatrachus africanus CAS 258223,

GFMJ 1389 (D, E, F, G), Chiromantis rufescens GFMJ 1095 (H).

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Jongsma et al.

Results and Discussion

Haut-Ogooue Province contains a rich diversity of

amphibians but remains understudied in spite of its inter¬

esting geological history (Bateke Plateau, Chaillu Mas¬

sif, Franceville Basin) and pristine savanna-forest mosaic

habitat. Further inventory work will no doubt reveal

more amphibian diversity. We encountered 34 amphib¬

ian species, 26 of which are new provincial records and

two of which are new records for Gabon (Afrixa/us oso-

rioi and Hyperolius balfouri). These results bring the

known amphibian diversity of the Haut-Ogooue province

to 42 species and 98 species for Gabon. Several of our

records also represent significant range extensions (>100

km from nearest known record). Our record of Hypero¬

lius balfouri represents a 725-km distribution expansion

south of its known distribution (nearest voucher, MHNG

1559 from Obala, Central province, Cameroon). Other

range extensions include Leptopelis calcaratus (315 km

S; nearest voucher is NCSM 77692, Ivindo NP), Scoto-

bleps gabonicus (275 km SE; nearest voucher NCSM

78915, Ogooue-Ivindo province), Sclerophrys supercili-

aris (320 km S, nearest locality is Makokou, Ogooue-

Ivindo province), Hyperolius kuligae (275 km S; near¬

est voucher is CUMV 15570, Ogooue-Ivindo province),

H. pardalis (310 km SE; nearest locality is Ivindo NP),

and Amnirana amnicola (310 km SE; nearest voucher is

MCZ A-139750, Ivindo NP). Laurent (1951) reported

Hyperolius steindaclmeri from Franceville, Gabon. If

confirmed, this record represents an 870 km extension

to the north for this species and brings the total known

diversity of Haut-Ogooue province to 43 species. Hav¬

ing not examined this specimen, we refrain from includ¬

ing it in the list presented here. The rarefaction curve for

amphibians suggest that we are nearing the true species

diversity at our sampling sites, though we anticipate that

uncommon species remain to be discovered in this area

(Fig. 2).

This article represents the first attempt to characterize

amphibian diversity in Haut-Ogooue province in south¬

eastern Gabon. The most amphibian-diverse province in

Gabon is the Ogooue-Maritime with 77 species (Burger

et al. 2004; Burger et al. 2006). Haut-Ogooue province

is comparable in overall amphibian diversity to Ogooue-

Ivindo, which has 46 known amphibian species (Burger

et al. 2004), and Moyen-Ogooue, which has 41 species

(Mocquard 1902; Pauwels 2016; specimens at the Cali¬

fornia Academy of Sciences, unpubl ). Based on GBIF

data, Estuaire province has 31 known species, Ogo-

oue-Lolo has 17, and Nyanga has 14. Other provinces

have never been surveyed for amphibians (Ngounie and

Woleu-Ntem). We believe that further surveys in Haut-

Ogooue that focus on higher elevation sites and savannas

will increase the known diversity of amphibian for this

underexplored province.

Acknowledgements. —We thank the Centre National

de la Recherche Scientifique et Technologique (CENAR-

EST, permit #AR008/15/CSAR) for providing scientific

permits and the Direction de la Faune et de la Chasse

for providing an export permit. For logistical support, we

thank The Nature Conservancy (Marie-Claire Paiz) and

the Smithsonian Institute (Lisa Korte). We are indebted

to Glen Ratel and Marie Coupe for hosting us in Libre¬

ville. Thanks to Daniel M. Portik for helping identify

hyperoliid species, including the new country record A.

osorioi. We thank Jordana Abugattas for assistance with

molecular genetics labwork. For assistance in the field

we thank Freye Pavel for always getting us safely to

our destination and keeping us fed once there. We are

indebted to Chief Dipanda Guillaume and his family

(Doumaye) and to Chief Michel Ngari (Mboua) for their

hospitality, warmth and knowledge. Marius Burger and

Olivier S.G. Pauwels provided valuable revisions on this

article. Edward L. Stanley helped make Figure 1. And

finally, we thank our local guides (Julien Yinga, Crepin

and Blaise) that never led us astray during our nocturnal

quests for frogs. This project was supported by the NSF

grant (#1202609) to DCB and funding by Shell Gabon

and the Smithsonian Conservation Biology Institute for

ET.

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Gregory F.M. Jongsma is a Ph.D. student at the Florida Museum of Natural History at the University

of Florida, USA. He received a B.S. from Acadia University (2010) and a M.S. from San Francisco

State University (2014). For his dissertation he is taking a comparative phylogeographic approach to

explore the diversification of frogs in Central Africa. He is seeking sponsorship from Regab and Vache

Qui Rit to help sustain the team during future field work in Gabon.

Elie Tobi has been working with the Smithsonian Institution’s, Gabon Biodiversity Program since

2001. He has been involved in the Monitoring and Assessment of Biodiversity in the Gamba Complex

of Protected Areas. He conducted the amphibian assessment before the Loubomo-Moungagara

National Road construction in South West Gabon and did recommendations to avoid and or mitigate

the road construction impact on amphibian populations. He is also involved in the monitoring of Nile

Crocodile nesting in the Gamba area. Elie manages an important zoological reference collection in

Africa (123,000 specimens of mammals, birds, fish, reptiles, amphibians, and arthopods). He is leading

environmental education and awareness programs with schools and workers of hydrocarbon companies

to reduce human/animals incidents, conflicts, and impacts.

Amphib. Reptile Conserv.

22 November 2017 | Volume 11 | Number 1 | e144

Amphibians of southeastern Gabon

Graham P. Dixon-MacCallum developed his love for herpetology over the summer before his third

year at Acadia University while working for a travelling snake show in Ontario. After graduation he

worked bird jobs to pay the bills but always kept an eye to the ground (and in the trees) for all things

scaled or slimy. He returned officially to the world of herpetology while completing a master’s degree

on garter snake habitat use at the University of Victoria, British Columbia. He misses wading through

elephant wallows in Gabon in search of frogs. Graham currently works in the Centre for Conservation

Research at the Calgary Zoo, in Calgary, Alberta.

Abraham Bamba-Kaya has been employed since 2015 at Institut de Recherche Agronomique

et Forestiere. His research interests include aquatic biodiversity, conservation, and investigating

amphibian declines. He has participated in expeditions of assessments and surveys of amphibians in

various localities in Gabon.

Jean-Aime Yoga is a researcher at the National Center for Scientific and Technological Research

(CENAREST) of Gabon. He has a special interest in herpetology and has been involved in several

projects since 2005. He helped with monitoring and assessing biodiversity at the Rabi site for

amphibians and reptiles (Smithsonian Institution Project). He was the first to collect Raniphotyplops

braminus (Snakes: Typlopidae) from Gabon.

Jean-Daniel Mbega is the head researcher at the Laboratory of Hydrobiology and Oenology of the

National Center based at Institut de Recherches Agronomiques et Forestieres (IRAF/CENAREST).

Jean Daniel has made several important contributions to our knowledge about the freshwater fish of

Gabon. In 2008, the National Assembly of Gabon presented Dr. Mbega the gold medal for his published

Identification Guide of the Fishes of the Lower Ogooue Basin.

Jean Herve Mve Beh is a researcher at the Laboratory of Hydrobiology and Oenology of the National

Center for Scientific and Technological Research (CENAREST) of Gabon. There are nearly twenty-

two experiments on projects on freshwater fish in Gabon and brackish fish in Gabon. He participates in

several proj ects with a focus on taxonomy, biology, and conservation. He participated in the development

of the IUCN report on marine fish in the eastern Atlantic. Jean Herve is currently working on a project

on the role of mangroves in the Akanda National Park as a nursery for species of commercial interest

landed by artisanal fisheries. It has just contributed to a project financed by TNC Gabon and to the study

of the baselines of the fish populations of the sites proposed for the hydropower project. Jean Herve is

a member of the scientific society Gilbert.

Andi Emrich fostered a love for amphibians as a small child in Canada but re-invigorated the spark

after working with Greg Jongsma on his research in Ecuador in 2007. After moving to San Francisco,

Andi got involved at the California Academy of Sciences, volunteering her time in the mammalogy

collections. She now works for the Florida Organic Growers in Gainesville, Florida but hopes to get

back in the field on another amphibian adventure soon.

David C. Blackburn is the Associate Curator of Herpetology at the Florida Museum of Natural History

at the University of Florida, USA. He received a BA from the University of Chicago (2001) and a Ph.D.

from Harvard University (2008). His research focuses on the diversity and evolution of frogs. He hopes

to one day see the following strange frogs alive in the field: CalyptocephalelJagayi, Conraua beccarii ,

Myobatrachus gouldii, and Triprion petastatus.

Amphib. Reptile Conserv.

November 2017 | Volume 11 | Number 1 | e144

Official journal website:

amphibian-reptile-conservation.org

Amphibian & Reptile Conservation

11(1) [Special Section]: 24-43 (el48).

Preliminary herpetological survey of Ngonye Falls and

surrounding regions in south-western Zambia

12 *Darren W. Pietersen, 3 Errol W. Pietersen, and 4 ’ 5 Werner Conradie

1 Department of Zoology> and Entomology\ University> of Pretoria, Private Bag X20, Hatfield, 0028, SOUTH AFRICA 2 Research Associate,

Herpetology Section, Department of Vertebrates, Ditsong National Museum of Natural History, P.O. Box 413, Pretoria, 0001, SOUTH AFRICA

3 P.O. Box 1514, Hoedspruit, 1380, SOUTH AFRICA 4 Port Elizabeth Museum (Bayworld), PO. Box 13147, Humewood, 6013, SOUTH AFRICA

5 School of Natural Resource Management, George Campus, Nelson Mandela Metropolitan University>, George, SOUTH AFRICA

Abstract. —The herpetofauna of Zambia has been relatively weil-studied, although most surveys were conducted

decades ago. In western Zambia in particular, surveys were largely restricted to a few centers, particularly

those along the Zambezi River. We here report on the herpetofauna of the Ngonye Fails and surrounding

regions in south-western Zambia. We recorded 18 amphibian, one crocodile, two chelonian, 22 lizard, and 19

snake species, although a number of additional species are expected to occur in the region based on their

known distribution and habitat preferences. We also provide three new reptile country records for Zambia

(Long-tailed Worm Lizard, Dalophia longicauda, Anchieta’s Worm Lizard, Monopeltis anchietae, and Zambezi

Rough-scaled Lizard, Ichnotropis grandiceps ), and report on the second specimen of Schmitz’s Legless Skink,

Acontias schmitzi, a species described in 2012 and until now known only from the holotype. This record also

represents a 140 km southward range extension for the species.

Keywords. Sioma Ngwezi National Park, Barotseland, Western Province, Africa, distribution, reptiles, lizards, am¬

phibians

Citation: Pietersen DW, Pietersen EW, Conradie W, 2017. Preliminary herpetological survey of Ngonye Falls and surrounding regions in south¬

western Zambia. Amphibian & Reptile Conservation 11(1) [Special Section]: 24-43 (e148).

NoDerivatives 4.0 International License, which permits unrestricted use for non-commercial and education purposes only, in any medium, provided

the original author and the official and authorized publication sources are recognized and properly credited. The official and authorized publication

credit sources, which will be duly enforced, are as follows: official journal title Amphibian & Reptile Conservation ; official journal website <amphibian-

reptiie-conservation.org>.

Received: 19 November 2017; Accepted: 13 December 2017; Published: 31 December 2017

Introduction

Despite the herpetofauna of Zambia being relatively

well-studied (e.g., Broadley 1971a), large areas of the

country remain poorly surveyed (Broadley 2000a; Poyn-

ton and Broadley 1985a). Most collecting was conducted

prior to 1970, although important subsequent contribu¬

tions were made by Broadley (1991a,b, 2000a), Branch

and Haagner (1993), Haagner et al. (2000), Chansa and

Wagner (2006) and Wagner et al. (2012a,b,c, 2013). The

last systematic review of Zambia’s herpetofauna was

undertaken by Broadley (1971a), at which time there

were 65 amphibian, two crocodile, nine chelonian, 54

lizard, and 75 snake species recorded. The amphibi¬

ans of Zambia have been dealt with in detail by Poyn-

ton and Broadley (1985a,b, 1987, 1988, 1991a), while

snakes were covered by Broadley et al. (2003). Since

these publications a number of taxonomic changes have

occurred, new species described (Broadley 2014; Wag¬

ner et al. 2012a,b), and species recorded from Zambia

COrrespondence. *pietersen.darren@gmail.com

for the first time (Broadley and van Daele 2003; Wagner

et al. 2013). Currently the Zambian herpetofauna com¬

prises 85 amphibian, two crocodile, 10 chelonian, 75 liz¬

ard, and 91 snake species (AmphibiaWeb 2016; Broadley

1971a; Broadley and van Daele 2003; Uetz et al. 2017).

Of these, 35 amphibian, one crocodile, three chelonian,

27 lizard, and 39 snake species are known from the Baro-

tse Floodplains and surroundings (Broadley 2000a).

Barotseland lies at the junction of three broad zoo¬

geographic zones, viz. mesic Angolan/Congolian Zone,

arid Kalahari Zone, and the East African coastal zone,

and the region is thus expected to support high herpe-

tofaunal diversity (Timberlake 2000). Western Zambia

remains one of the neglected regions of Zambia from a

biodiversity perspective, although it received attention

during the cross-border Zambezi Basin Wetland survey

conducted in the late 1990s (Broadley 2000a; Channing

2000; Timberlake 2000). Targeted surveys of the Barotse

Floodplains led to the description of a new frog species,

Hemisus barotseensis (Channing and Broadley 2002).

Amphib. Reptile Conserv.

December 2017 | Volume 11 | Number 1 | e148

Pietersen et al.

Fig. 1. (A) Map of western Zambia indicating major localities mentioned in the text, including Sioma Ngwezi National Park (dark

grey polygon). The light grey rectangle indicates the study site, and the dotted line indicates the Zambezi River. Inset: Map of

Zambia indicating the enlarged region. (B) Enlarged study site with locality names. The Ngonye Falls campsite and visitor’s center

are both within 1 km of Ngonye Falls and are therefore not indicated on the map.

Prior to this, small collections from western Zambia were

reported on by Roux (1907), Angel (1920, 1921, 1922)

and Broadley (1968a, 1971a), which led to the descrip¬

tions of various snakes and lizards from the region,

including Dalophia ellenbergeri (Angel, 1920); Tetra-

dactylus ellenbergeri (Angel, 1922); Typhlacontias grac¬

ilis Roux, 1907; T. rohani Angel, 1923; Acontias jappi

(Broadley, 1968); Amblyodipsas ventrimaculata (Roux,

1907); Crotaphopeltis barotseensis Broadley, 1968 and

Zygaspis nigra Broadley and Gans, 1969.

We had the opportunity over a period of three-and-a-

half-years to document the herpetofauna of the Ngonye

Falls region in western Zambia, and present here an ini¬

tial inventory.

Methods

Study area

The Ngonye Falls are located in the Western Province in

south-western Zambia (Fig. 1). A tourist attraction in its

own right (Fig. 2a), it is also the location of the admin¬

istrative headquarters of Sioma Ngwezi National Park,

which is situated to the south-west. The Ngonye Falls

form a northern extension of the Sioma Ngwezi National

Park, and as such the falls and a small area surround¬

ing it are afforded official protection. The vegetation falls

into the Flora Zambesiaca bioregion and is dominated

by Baikiaea woodland on deep Kalahari soils, although

intense settlement and subsistence agriculture prevail in

the vicinity of the Zambezi River. Rupicolous habitat is

restricted to the immediate vicinity of the Zambezi River.

Geographical coordinates for the main localities men¬

tioned in the species accounts are presented in Table 1.

Data collection

One of us (EWP) was stationed permanently at Ngonye

Falls from 8 February 2013 to 30 August 2016. During

this time reptiles and amphibians were recorded inciden¬

tally, with some active searching. Herpetofauna were

actively searched for on the western bank of the Zam¬

bezi River by DWP, John Davies, and EWP from 21 to

28 April 2013, on the eastern shore by WC from 17 to

27 October 2015, and Roger Bills from 7 to 11 October

2017. Voucher specimens were not collected in the early

stages of this survey due to the lack of collecting per¬

mits, but in these instances photographic records were

obtained, as has also been done in other surveys and

regional works (e.g., Tuberville et al. 2005; Gooley et al.

2011; Bates et al. 2014).

All surveys involved opportunistic visual encoun¬

ters. Diurnal surveys involved actively searching specific

microhabitats, particularly beneath rocks and decaying

logs. Nocturnal surveys for amphibians were undertaken

in wetlands and surrounding woodland. Two standard

Y-shape trap arrays were deployed on the eastern bank of

the Zambezi River in October 2015, with each array con¬

sisting of three drift fences (10 m long and 50 cm high),

with four pitfall traps (one at the center and at each fence

Amphib. Reptile Conserv.

December 2017 | Volume 11 | Number 1 | e148

Herpetofauna of Ngonye Falls

Fig. 2. Selection of amphibians and reptiles photographed in the vicinity of Ngonye Falls, south-western Zambia. (A) View of

Ngonye Falls from the eastern side of the Zambezi River. (B) Mapacha Grass Frog ( Ptychadena cf. mapacha, VMUS 5990),

Sioma Ngwezi National Park Headquarters. (C) Long-tailed Worm Lizard ( Dalophia longicauda), Sioma Ngwezi National Park

Headquarters. (D) Zambezi Rough-scaled Lizard ( Ichnotropis grandiceps , TM 86237), Sioma Ngwezi National Park Headquarters.

(E) Barotse Blind Legless Skink (Acontias jappi , TM 86232), Sioma Ngwezi National Park Headquarters. (F) Eastern Black-lined

Plated Lizard ( Gerrhosaurus intermedius), Sioma Ngwezi National Park Headquarters.

tip, respectively) and six one-way funnel traps placed on

opposite sides of the fences in the center of each arm.

Specimens retained for subsequent study were

humanely euthanized by injecting tricaine methanesul-

fonate (MS222) solution into the intracoelomic cavity

for reptiles (Conroy et al. 2009), and submerging frogs

in a MS222 solution, after which they were formalin-

fixed for 48 hours and then transferred to alcohol for

long-term storage. Prior to fixing, tissue samples (either

liver or muscle) were preserved in 96% ethanol for use

in genetic analyses. Voucher specimens (Appendix 1) are

held in the herpetological collections of the Port Eliza¬

beth Museum (PEM), Ditsong National Museum of Nat¬

ural History, Pretoria (TM), and South African Aquatic

Biodiversity Institute, Grahamstown (SAIAB). Reptile

and amphibian photographic records were submitted to

the Animal Demography Unit Virtual Museum (Avail¬

able: http://vmus.adu.org.za) on the platforms Rep-

tileMAP and FrogMAP, respectively. Ventral scales were

counted from the first scale posterior to the mental up to

(but excluding) the cloacal shield. Subcaudal scales were

counted from the first scale posterior to the cloacal shield

up to, but excluding, the terminal scale. For amphisbae-

nids, dorsal annuli were counted along the dorsal mid-

line from the first whole annulus posterior to the head up

to the last annulus anterior to the cloacal shield. Caudal

annuli were counted along the dorsal midline starting at

the first complete annulus posterior to the cloacal shield,

up to (but excluding) the terminal pad.

Relevant field guides (Branch 1998; Broadley 1983;

Broadley et al. 2003; Channing 2001; du Preez and

Carruthers 2009) were used for species identification.

Amphib. Reptile Conserv.

December 2017 | Volume 11 | Number 1 | e148

Pietersen et al.

Table 1 . Coordinates for the main localities at and around Ngonye Falls and Sioma Ngwezi National Park, as mentioned in the text.

Locality

Latitude

Longitude

Ngonye Falls airstrip

16° 41’40” S

23° 36’ 49” E

Ngonye Falls campsite

16° 39’ 41” S

23° 34’23”E

Park Headquarters

16° 40’08” S

23° 34’ 03” E

Visitor’s center

16°39’24”S

23° 34’ 11” E

Maziba Bay

16° 41’ 16” S

23° 38’ 12” E

East bank of Zambezi River opposite Ngonye Falls

16° 39’ 20” S

23°34’35”E

West bank of Zambezi River opposite Ngonye Falls

16° 39’ 11”S

23° 34’ 14” E

Idjobwa Pan

16° 53’ 48” S

23°35’50”E

Sioma Barge

16°38’35”S

23°33’25”E

Nomenclature was based on established online databases

(amphibians: Frost 2016; reptiles: Uetz et al. 2017),

updated where appropriate. Vernacular names follow du

Preez and Carruthers (2009) for amphibians and Branch

(1998) for reptiles, updated from Frost (2016) and Uetz

et al. (2017) for those taxa not covered by these guides.

No regional conservation assessment has been under¬

taken for Zambian amphibians and reptiles as yet, but

where global conservation assessments are available

(IUCN 2017) these are noted. Endemic (defined as spe¬

cies with ranges restricted to Zambia) and near-endemic

species (>90% of distribution within Zambia) are also

indicated.

Species accounts

Amphibia

Breviceptidae

Breviceps adspersus adspersus Peters, 1882

Bushveld Rain Frog

Photograph: VMUS 5982

Individuals were photographed in Ngonye Falls camp¬

site, and were heard calling from this area, from the visi¬

tor’s center and from the vicinity of Park Headquarters.

This species is distinguished from B. poweri on the basis

of call, having a series of pale paravertebral and dorsolat¬

eral patches, absence of a continuous pale line from the

upper lip to the forearm, and having a less intense dark

throat that is medially divided by a white line (du Preez

and Carruthers 2009; Poynton and Broadley 1985a,

1991). In Zambia, this species has been collected only in

the vicinity of Kalabo, about 200 km to the NNW (Chan-

ning 2001; Poynton and Broadley 1985a, 1991).

Breviceps poweri Parker, 1934

Power’s Rain Frog

Photograph: VMUS 5983

This species was often heard, and photographed, in the

vicinity of Ngonye Falls campsite, visitor’s center and

Park Headquarters. It is distinguished from B. adsper¬

sus on the basis of call, absence of paravertebral patches

(usually present in B. adspersus ); presence of a pale

patch above the vent (usually absent in B. adspersus );

uniformly dark throat (usually mottled in B. adsper¬

sus ); continuous pale band from upper lip to forearm;

and presence of a short, dark band between the nostrils

and mouth (usually not well developed in B. adspersus ;

du Preez and Carruthers 2009; Poynton and Broadley

1985a). At present only B. a. adspersus is known to occur

west of the Zambezi River in south-western Zambia,

with B. poweri largely restricted to east of the Zambezi

River, although both species have been recorded occur¬

ring sympatrically at Kalabo (Broadley 1971a; Channing

2001; du Preez and Carruthers 2009; Poynton and Broad¬

ley 1985a, 1991). This record extends the distribution of

the species by about 200 km SSE from Kalabo.

Bufonidae

Poyntonophrynusfenoidheti (Hewitt and Methuen, 1913)

Northern Pygmy Toad

Photograph: VMUS 5989

A single individual was photographed at Ngonye Falls

campsite. It was distinguished from P. kavangensis on

the basis of the tympanum being distinctly visible, and

the presence of small tubercles on the dorsal surface of

the snout (du Preez and Carruthers 2009). The only pre¬

vious Zambian records are from the northern shore of

Lake Kariba and the Zambian bank of the Victoria Falls

(Broadley 1971a; Channing 2001; Poynton and Broadley

1988, 1991), while the nearest locality is Katima Mulilo

in the Zambezi Region of Namibia (Channing 2001;

Poynton and Broadley 1991). This record extends the

known range of this species 110 km north-west.

Schismaderma carens (Smith, 1848)

Red Toad

Material: SAIAB 205361, 205631

Photographs: VMUS 5992, 5993

Individuals were recorded at Ngonye Falls campsite,

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Herpetofauna of Ngonye Falls

at the airstrip, and on the eastern banks of the Zambezi

River below Ngonye Falls. This species was previously

recorded at Livingstone and Kalabo in Western Province

(Broadley 1971a; Channing 2001; Poynton and Broadley

1988, 1991).

Sclerophrys gutturalis (Power, 1927)

Guttural Toad

Photograph: VMUS 5994

Seen at Ngonye Falls campsite, and common throughout

Zambia (Broadley 1971a; Channing 2001; Poynton and

Broadley 1988, 1991).

Sclerophrys poweri (Hewitt, 1935)

Western Olive Toad

Material: SAIAB 205356

Photographs: VMUS 5996, 5997

Observed and photographed at Ngonye Falls campsite,

and collected on the eastern banks of the Zambezi River

below Ngonye Falls. These records extend the known

distribution of this species about 110 km north-west from

the nearest records at Sesheke (Channing 2001; Poynton

and Broadley 1988, 1991), although Broadley (2000a)

records it from the “Barotse floodplains.”

Sclerophrys pusilla (Hallowell, 1855)

Flat-backed Toad

Material: PEM A11719, 11720; SAIAB 205360

Photographs: VMUS 5995, 5998

Individuals were seen at Ngonye Falls campsite, as well

as on the eastern bank of the Zambezi River opposite

Ngonye Falls. In Western Province it has been collected

only at Kalabo (Broadley 1971a; Channing 2001; Poyn¬

ton and Broadley 1991).

Hemisotidae

Hemisus marmoratus (Peters, 1854)

Mottled Shovel-nosed Frog

Photograph: VMUS 5985

One individual was photographed in Ngonye Falls camp¬

site. It was distinguished from H. guineensis on the basis

of coloration (dorsum mottled light and dark in H. mar¬

moratus and dark with small yellow, orange or white

spots in H. guineensis ; Channing 2001; du Preez and

Carruthers 2009). It was distinguished from H. baro-

tseensis by having the upper eyelid length exceeding

the eye-nostril distance (Channing 2001; Channing and

Broadley 2002). The only previous record of this spe¬

cies in Western Province is at Livingstone, although also

recorded from Katima Mulilo in Namibia (Poynton and

Broadley 1985a, 1991).

Hyperoliidae

Hyperolius angolensis Steindachner, 1867

Angolan Reed Frog

Photograph: VMUS 5986

A single individual, which we tentatively refer to H.

angolensis , was photographed at Ngonye Falls campsite.

This species is likely to be more common and is proba¬

bly found in pans and other temporary wetlands in Sioma

Ngwezi National Park, as well as other wetlands associ¬

ated with the Zambezi and Cuando Rivers. The only pre¬

vious Zambian records are from the western shore of the

Upper Zambezi at Sandaula Plain, Kalabo and Kalenga

(Broadley 1971a; Poynton and Broadley 1987), although

it is fairly widespread in the Okavango Swamps of

Botswana and the Zambezi Region of northern Namibia

(Poynton and Broadley 1987, 1991) as well as south¬

eastern Angola (Conradie et al. 2016). This record par¬

tially bridges the gap between the Namibian and Upper

Zambezi records. The taxonomic status of this species

remains unresolved and it is considered part of the larger

unresolved H. parallelus Gunther, 1858 group which is

widespread across Angola and adjacent countries (Frost

2016). Many regional color patterns exist, the specimen

from Ngonye Falls conforms best to that of H. angolensis

(fide Schiotz 1999).

Phrynobatrachsdae

Phrynobatrachus natalensis (Smith, 1849)

Snoring Puddle Frog

Material: SAIAB 205351

Photograph: VMUS 5987

This species was recorded on the west bank of the Zam¬

bezi River in the vicinity of Ngonye Falls and campsite,

as well as on the eastern bank of the Zambezi River oppo¬

site Ngonye Falls. It is widespread in Zambia (Broadley

1971a; Channing 2001; Poynton and Broadley 1985b,

1991).

Phrynobatrachus parvulus (Boulenger, 1905)

Small Puddle Frog

Photograph: VMUS 5988

An individual was photographed at Ngonye Falls camp¬

site. The only previous record for Western Province is

Ngambwe Rapids, about 90 km to the south-east (Chan¬

ning 2001; Poynton and Broadley 1985b, 1991).

Pipidae

Xenopus muelleri (Peters, 1844)

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Pietersen et al.

Muller’s Platanna

Material: SAIAB 202357

One male and one female were collected in sympatry

with X. poweri in an eastern tributary (16°39’07”S,

23°37’43”E) flowing into the Zambezi River. Conradie

et al. (2016) also recorded these two species in sympatry

in south-eastern Angola.

Xenopus poweri Hewitt, 1927

Power’s Platanna

Material: SAIAB 202355

One male and one female were collected in sympatry

with X muelleri in an eastern tributary of the Zambezi

River (see above).

Ptychadenidae

Ptychadena cf. mapacha Channing, 1993

Mapacha Grass Frog

Material: TM 86255

Photograph: VMUS 5990

One individual (TM 86255) was collected at Park Head¬

quarters after being killed by a vehicle, while a second

individual (VMUS 5990, Fig. 2b) was photographed at

the same site. Individuals are provisionally assigned to

this species based on external morphology and color¬

ation, however molecular analyses and/or call record¬

ings are required to unequivocally confirm these iden¬

tifications. For many years this species was only known

from the type locality, viz. Mapacha Airfield and the

area surrounding Katima Mulilo in the Zambezi Region

of Namibia (Channing 1993; du Preez and Carruthers

2009), although expected to occur in south-western Zam¬

bia, south-eastern Angola and northern Botswana as well

(Channing 2001). Haacke (1999) collected four individu¬

als along the Ojmatako River about 80 km east of Rundu

(this record has largely been overlooked in the litera¬

ture), while most recently Ceriaco et al. (2016) recorded

it from Rundu District in Kavango-East Region, north¬

ern Namibia. Conradie et al. (2016) collected a series

of Ptychadena at Jamba in neighboring south-eastern

Angola which they provisionally assigned to P. cf. mos-

sambica, although noting that their specimens may be

referable to P. mapacha. The records reported here are

the first for Zambia and the first outside Namibia. These

records extend the known distribution of this Data Defi¬

cient species 120 km NNW into Zambia, and 320 km

NNE into south-eastern Angola, which could have posi¬

tive conservation implications (IUCN SSC Amphibian

Specialist Group, SA-FRoG 2017).

Ptychadena oxyrhynchus (Smith, 1849)

Sharp-nosed Grass Frog

Material: SAIAB 205353, 205354

Photograph: VMUS 5991

An individual was photographed at Ngonye Falls camp¬

site, and individuals were also heard calling from the

western bank of the Zambezi River in the vicinity of

Ngonye Falls. Additional material was collected from

the eastern bank of the Zambezi River below Ngonye

Falls. This species is widespread in Zambia (Broadley

1971a; Channing 2001), although the only other pub¬

lished record for Western Province is Sesheke (Channing

2001; Poynton and Broadley 1985b, 1991), 110 km to

the south-east.

Ptychadena snbpimctata (Bocage, 1866)

Speckled-bellied Grass Frog

Material: PEM A11717, 11718; SAIAB 205358, 205365

This species was recorded from the eastern shore of the

Zambezi River, opposite Ngonye Falls. It is widespread

in Zambia, including Upper Zambezi Region (Broadley

1971a; Channing 2001; Poynton and Broadley 1985b,

1991).

Pyxicephalidae

Tomopterna cf. cryptotis (Boulenger, 1907)

Tremolo Sand Frog

Material: SAIAB 205362

Photographs: VMUS 5999, 6000

Recorded at Ngonye Falls campsite and on the eastern

banks of the Zambezi River below Ngonye Falls. Previ¬

ously collected in Western Province at Kalabo, Sandaula

Plain and Sesheke (Channing 2001; Poynton and Broad¬

ley 1985b, 1991), and our records partially fill the gap

between these localities. We provisionally assign our

records to T. cryptotis based on distribution, but note that

species delineation in this genus is problematic when

based solely on external morphology, and these speci¬

mens may in fact refer to the similar Tandy’s Sand Frog

T. tandyi Channing and Bogart, 1996.

Rhacophoridae

Chiromantis xerampelina Peters, 1854

Southern Foam Nest Frog

Photograph: VMUS 5984

An individual was photographed at the airstrip. Although

widespread in Zambia, the only previous records from

Western Province are Sesheke and Fukulu (Broadley

1971a; Channing 2001; Poynton and Broadley 1987,

1991).

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December 2017 | Volume 11 | Number 1 | e148

Herpetofauna of Ngonye Falls

Reptilia

Squamata

Sauria

Agamidae

Agama armata Peters, 1855

Peter’s Ground Agama

Material: PEM R22017, 22018

Photographs: VMUS 158866-158869

Recorded regularly at Ngonye Falls campsite and visi¬

tor’s center, and the vicinity of the airstrip. It is wide¬

spread in Zambia (Broadley 1971a).

Amphisbaenidae

Dalophia longicauda (Werner, 1915)

Long-tailed Wonn Lizard

Photograph: VMUS 163523

A single individual (Fig. 2c) was found on the soil surface

in the vicinity of Park Headquarters. It is uniform flesh-

pink in color with 326 dorsal and 37 caudal annuli, with

the tail ending in a calloused pad. There is no constricted

caudal autotomy site, and the dorsal caudal annuli form

a “herring-bone” pattern. The cephalic shield consists of

a single large plate, with lateral sulci. This individual is

distinguished from D. angolensis and D. ellenbergeri by

the absence of a constricted caudal autotomy site (Broad¬

ley et al. 1976). It is further distinguished from both D.

angolensis and D. pistillum by the high subcaudal counts

(usually 20-27 caudal annuli in D. angolensis and 19-33

in D. pistillum). It is distinguished from D. angolensis , D.

ellenbergeri , and D. pistillum by the dorsal caudal annuli

forming a “herring-bone” pattern (Broadley et al. 1976).

This represents the first record of this fossorial species in

Zambia (Branch 1998; Broadley 1971a; Broadley et al.

1976; Uetzetal. 2017).

Dalophia pistillum (Boettger, 1895)

Blunt-tailed Worm Lizard

Material: PEM R22925

A single individual was collected on the eastern bank

of the Zambezi River below Ngonye Falls. This species

is distinguished from other Dalophia in Zambia by the

absence of a constricted caudal autotomy annulus (pres¬

ent in D. angolensis and D. ellenbergeri), lower numbers

of caudal annuli (27 versus 33-42 in D. longicauda), and

absence of a “herring-bone” pattern on the dorsal caudal

annuli (present in D. longicauda:, Broadley et al. 1976).

It is fairly widespread in southern and western Zambia,

although records are sparse due to its predominantly fos¬

sorial habits (Broadley et al. 1976).

Monopeltis anchietae (Bocage, 1873)

Anchieta’s Worm Lizard

Material: TM 86250

One individual (Fig. 3a,b) was found beneath an ele¬

phant carcass at Idjobwa Pan in the buffer zone to the

north of Sioma Ngwezi National Park, while a juvenile

(TM 86250) was unearthed during construction at Park

Headquarters. Members of the genus Monopeltis gen¬

erally inhabit deep Kalahari sands, only coming to the

surface after their burrows have been flooded by heavy

rains (Branch 1998, DWP pers. obs.). These are the first

records of this species in Zambia. Previously known

from northern Botswana, northern Namibia and south¬

ern Angola (Broadley 1971a; Broadley et al. 1976; Uetz

etal. 2017).

Zygaspis nigra Broadley and Gans, 1969

Black Round-headed Worm Lizard

Material: TM 86209

Photograph: VMUS 158938

This small fossorial species was collected with Z. quadri-

frons in Baikiaea woodland at the airstrip. Known from

Zambia, Angola, and northern Namibia, with most Zam¬

bian specimens collected at Kalabo, the type locality,

with a subsequent record from Ndau School (ca. 25 km

south-west of Mongu on the western side of the Zam¬

bezi River; Broadley 2000a). The new record partially

fills the gap between the Ndau School, eastern Angola

and Namibia records.

Zygaspis quadrifrons (Peters, 1862)

Kalahari Round-headed Worm Lizard

Material: TM 86208

Photograph: VMUS 158939

One individual was found in Baikiaea woodland at the

airstrip, while a second was found near Ngonye Falls

campsite. This species is probably quite common and

widespread throughout the area (see also Broadley

1971a), being overlooked due to its fossorial nature.

Chamaeleonidae

Chamaeleo dilepis Leach, 1819

Flap-neck Chameleon

Photographs: VMUS 158873, 158876, 158877

Recorded regularly around Park Headquarters, visitor’s

center, and Ngonye Falls campsite.

Gekkonidae

Pachydactylus wahlbergii wahtbergii (Peters, 1869)

Kalahari Thick-toed Gecko

Photographs: VMUS 163521, 163522

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Pietersen et al.

Fig. 3. Selection of reptiles photographed in the vicinity of Sioma Ngwezi National Park and Ngonye Falls in south-western Zambia.

(A) Anchieta’s Worm Lizard ( Monopeltis anchietae ), Idjobwa Pan, northern buffer zone of Sioma Ngwezi National Park and (B)

close-up of the head, showing the double head shields. (C) Wahlberg’s Kalahari Gecko ( Pachydactylus wahlbergii wahlbergii),

Ngonye Falls campsite. (D) Schmitz’s Blind Legless Skink ( Acontias schmitzi, PEM R22015), east bank of the Zambezi River

opposite Ngonye Falls.

An individual (SVL 56.3 mm) was photographed (Fig.

3c) at Ngonye Falls campsite on 21 April 2013, while a

second was photographed at the same site on 13 February

2014. This species has been recorded only once before

in Zambia, in the extreme south at Kalamba Station on

the old Zambezi Sawmill Railway (Broadley and Daele

2003). These new records are only the second and third

confirmed records of this species in Zambia. This spe¬

cies was previously placed in the genus Colopus , but was

transferred to Pachydactylus by Heinicke et al. (2017).

Hemidactylus mabouia (Moreau De Jonnes, 1818)

Amphib. Reptile Conserv.

Common Tropical House Gecko

Material: PEM R22019, 22020; TM 86251, 86252

Photographs: VMUS 158888-158890, 158892

Seen on the walls of the visitor’s center, Park Headquar¬

ters, and Ngonye Falls campsite. This species is common

and widespread throughout Zambia (Broadley 1971a).

Lygodactylus chobiensis FitzSimons, 1932

Chobe Dwarf Gecko

Material: PEM R22026; TM 86253

December 2017 | Volume 11 | Number 1 | e148

Herpetofauna of Ngonye Falls

Photographs: VMUS 158901, 158903-158906

The most common gecko species, seen on trees and walls

in the vicinity of Ngonye Falls campsite, visitor’s center,

and Park Headquarters. Largely restricted to the Zam¬

bezi Valley (Branch 1998; Broadley 1971a), but should

be searched for along the banks of the Cuando River.

Gerrhosauridae

Gerrhosaurus auritus Boettger, 1887

Kalahari Plated Lizard

Photograph: VMUS 158885

Individuals were photographed at Ngonye Falls visi¬

tor’s center and Park Headquarters. We refer our indi¬

viduals from Ngonye Falls to G. auritus on the basis of

non-mucronate dorsal scales, the proximal caudal scales

lacking spines, the large tympanum covering a large por¬

tion of the ear opening, and coloration. The only previ¬

ous Zambian specimens are from Lealui, 170 km NNW,

although it is also recorded 130 km to the south in the

Zambezi Region of Namibia (Broadley 1971a).

Gerrhosaurus intermedins Lonnberg, 1907

Eastern Black-lined Plated Lizard

Photograph: VMUS 158887

A number of individuals suspected to be referable to G.

intermedins were seen, however only a single individual

was photographed and the scalation details later deter¬

mined from this photograph (Fig. 2f). A narrow, pale dor¬

solateral line is present, bordered on each side by a dark

line. A pale vertebral stripe, flanked on either side by a

dark stripe, originates on the nape and extends to the base

of the tail as discrete, evenly-spaced dashes. The fronto¬

nasal is divided into a large, heart-shaped anterior por¬

tion and a smaller posterior “inter-frontonasal,” which is

in contact with the frontal thus separating the prefron-

tals. The dorsal scales are arranged in 22 longitudinal and

approximately 56 transverse rows. The flank scales are

keeled and weakly mucronate and the head length is con¬

tained in the body length approximately 4.14 times.

Despite the prefrontals being separated, this individ¬

ual is distinguished from G. flavigularis by the presence

of four supraciliaries, the keeled and weakly mucronate

lateral scales, head length into snout-vent length ratio

and coloration (Bates et al. 2013; Branch 1998; FitzSi-

mons 1943). It is distinguished from G. auritus by the

number of longitudinal and transverse scale rows and the

keeled and weakly mucronate lateral scales. We refer this

individual to G. intermedins rather than G. nigrolinea-

tus sensu stricto based predominantly on coloration. In

G. nigrolineatus the dark stripes bordering the pale dor¬

solateral stripes are often ragged, while the pale verte¬

bral stripe (and flanking dark stripes) may be continu¬

ous or absent, but are usually better-developed than in G.

intermedins (Bates et al. 2013). When present, the dark

Amphib. Reptile Conserv.

markings surrounding the discontinuous vertebral stripe

appear to be more extensive (Bates et al. 2013; Hallowed

1857). This record is about 110 km NNW of the nearest

previous reported locality at Sesheke (Broadley 1971a).

Lacertidae

Ichnotropis capensis (Smith, 1838)

Cape Rough-scaled Lizard

Material: PEM R22021-22024

Photographs: VMUS 158896, 158902

Individuals were recorded in the vicinity of Park Head¬

quarters, Ngonye Falls campsite, visitor’s center, eastern

bank of the Zambezi River opposite Ngonye Falls, and

also in Sioma Ngwezi National Park. It is widespread in

western Zambia (Broadley 1971a).

Ichnotropis gmndiceps Broadley, 1967

Zambezi Rough-scaled Lizard

Material: TM 86237

A number of individuals were encountered in Baikiaea

woodland in the vicinity of Park Headquarters (Fig. 2d).

All observed individuals were judged to be adult based

on their size, and were seen in April and May. TM 86237

has a snout-vent length of 49.6 mm and a tail length of

101.8 mm. The frontonasal is entire, the occipital does

not project beyond the parietals, and there are five supra-

labials anterior to the subocular, which borders the lip.

The dorsal scales are strongly keeled and overlapping,

and arranged in 47 rows at midbody. In all these char¬

acters our specimen closely resembles the type descrip¬

tion by Broadley (1967). Furthermore, there are 14 femo¬

ral pores present on each thigh. In coloration, the speci¬

men and photographed individuals closely resemble the

holotype (USNM 163989, available: http://inaturalist.ca/

taxa/35953-Ichnotropis-grandiceps; Accessed 5 Decem¬

ber 2017). All individuals are similar in coloration with

the head and back pale grey-brown to olive-brown ante¬

riorly, becoming olive-yellow posteriorly, usually with

scattered darker spots throughout. The limbs are reddish-

orange, with small pale spots sometimes present on the

hind limbs. A rust-red lateral stripe originates just behind

the eye and extends for the entire length of the body, fad¬

ing on the base of the tail. Below the lateral stripe the

flanks are light grey, frequently with large white spots.

The tail is uniformly grey-brown to pinkish, with a series

of paired dorsolateral dark and white spots. The ventrum

is white. This is the first record of this Data Deficient

species in Zambia (Turner 2010), and these are the first

specimens reported since Haacke (1970).

Meroles squamulosus (Peters, 1854)

Common Rough-scaled Sand Lizard

Photograph: VMUS 158907

December 2017 | Volume 11 | Number 1 | e148

Pietersen et al.

An individual was photographed in a dry marsh in open

woodland between Ngonye Falls campsite and visitor’s

center, while a second individual was photographed at the

visitor’s center. This species was also observed in Sioma

Ngwezi National Park. These are the first records of this

species in Western Province (Broadley 1971a), although

it is known from northern Namibia (Branch 1998) and

was recently recorded from south-eastern Angola (Con-

radie et al. 2016).

Scincidae

Acontias jappi (Broadley, 1968)

Barotse Tegless Skink

Material: TM 86232-34, TM 86254

Individuals were collected in soil under bush clumps

near Ngonye Falls campsite, airstrip and Park Head¬

quarters (Fig. 2e). Numerous additional individuals were

unearthed during the construction of Park Headquarters.

Originally described as a subspecies of Acontias kga-

lagadi (previously Typhlosanrus lineatus [Boulenger,

1887]), Schneider and Bauer (2009) elevated this taxon

to species status on morphological grounds. Specimens

were distinguished from A. k. kgaiagadi on the basis of

being significantly more robust, having only two longi¬

tudinal dark stripes which fade out on the tail (typically

4-8 in A. k. kgaiagadi)., 24-25 subcaudal scales (26-35

in A. k. kgaiagadi), the ocular scale being longer than

high (as long as high in A. k. kgaiagadi), and only three

sublingual scales border the mental (usually four in A. k.

kgaiagadi, Broadley 1968b; Branch 1998). TM 86232-

34 have four supralabials and three infralabials on each

side, while TM 86233 has the mental longitudinally

divided into two subequal parts (undivided in the remain¬

ing specimens). The superior border of the ocular scale is

incompletely fused with the anterior supraciliary on both

sides in all four specimens, resulting in a narrow slit that

is apparently continuous with the eye (i.e., an immovable

lower eyelid). Most other members of Acontias have the

eye either completely covered by an ocular scale or have

a moveable lower eyelid (Branch 1998), with only A. rie-

pelli having an immovable lower eyelid (Branch 1998).

Acontias jappi is restricted to south-western Zambia and

adjacent Angola (Broadley 1968b, 1971a; Schneider and

Bauer 2009).

Acontias schmitzi Wagner, Broadley and Bauer, 2012

Schmitz’s Legless Skink

Material: PEM R22015

A single specimen was collected on the east bank of the

Zambezi River, opposite Ngonye Falls (Fig. 3d). It was

found under a large log in deep sand in Miombo (. Bra-

chystegia spp.) woodland. Head scalation conforms to

the type specimen. The new specimen measures 138 mm

snout-vent length and 21 mm tail length, has 14 mid¬

body scale rows, 178 ventrals and 26 subcaudals. Color

in life is light orange ventrally and blue-grey dorsally,

with the anterior two-thirds of the tail darkly pigmented

ventrally. This is only the second record of this species

and the most southerly locality. The holotype was col¬

lected in the Kataba Reserve, south of Mongu, Western

Province, Zambia (15°23’00.9”S, 23°23’43.7”E; Wagner

et al. 2012a), and this record is 140 km south of the type

locality. Both records are on the eastern side of the Zam¬

bezi River in deep Kalahari sands. Based on morphologi¬

cal features, Wagner et al. (2012a) place A. schmitzi in a

clade with A. jappi. Considering their distribution, it is

expected that these two species are most probably sister

taxa, and that the Zambezi River acts as a barrier to dis¬

persal, thus facilitating their independent evolution and

parapatric distribution. This species is endemic to Zam¬

bia.

Mochhis simdevallii (Smith, 1849)

Sundevall’s Writhing Skink

Material: PEM R22027

Photograph: VMUS 158908

This semi-fossorial species was found at Ngonye Falls

visitor’s center as well as on the east bank of the Zambezi

River opposite Ngonye Falls. It is widespread in Zambia

(Broadley 1971a).

Panaspis maculicollis Jacobsen and Broadley, 2000

Spotted-neck Snake-eyed Skink

Photograph: VMUS 158911

Photographed in the vicinity of Ngonye Falls visitor’s

center. A Panaspis seen on the east bank of the Zam¬

bezi River opposite Ngonye Falls is provisionally also

assigned to this species. These records are situated

between the previous records at Sesheke and Ndanda

(Broadley 1971a; Jacobsen and Broadley 2000).

Typhlacontias rohani Angel, 1923

Kalahari Burrowing Skink

Material: TM 86235-36, 86248

Three specimens were collected during the construction

of Park Headquarters, while additional individuals were

photographed at the same locality as well as at Ngonye

Falls campsite and the vicinity of the airstrip. All three

specimens have five supralabials on either side of the

head, with the second contacting the eye. On the right

side of TM 86235, the prefrontal is separated from the

frontoparietal by the enlarged third supraocular, while

the prefrontal and frontoparietal are in contact on the

left side of TM 86235 and on both sides of TM 86236.

In Zambia this species was previously recorded only at

December 2017 | Volume 11 | Number 1 | e148

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Herpetofauna of Ngonye Falls

Kalabo in Western Province, where it is sympatric with T.

gracilis , although it is widespread in north-western Zim¬

babwe, northern Botswana and Namibia, and south-east¬

ern Angola (Branch 1998; Broadley 2000a; Conradie et

al. 2016; Haacke 1997; Uetz et al. 2017).

Trachylepis damarana (Peters, 1870)

Damara Variable Skink

Material: PEM R22030, 22031

Photographs: VMUS 158929, 158930, 158933-158935

The most common skink in the area, seen at Ngonye

Falls campsite, vicinity of the visitor’s center, Park Head¬

quarters, vicinity of the airstrip, and on the east bank of

the Zambezi River opposite Ngonye Falls. It is common

throughout Zambia (Broadley 1971a). Recently split

from the larger Trachylepis varia complex by Weinell

and Bauer (2018).

Trachylepis wahlbergii (Peters, 1869)

Wahlberg’s Striped Skink

Material: PEM R22029

Photographs: VMUS 158931, 158936

Common throughout the area, seen at Ngonye Falls

campsite, visitor’s center, Park Headquarters, and on the

east bank of the Zambezi River opposite Ngonye Falls.

This species is common and widespread throughout

Zambia, extending into Botswana and Namibia (Branch

1998; Broadley 1971a). Castiglia et al. (2006), using

karyotypic and genetic data, suggest that T. wahlbergii is

conspecific with T. striata , despite the morphological dif¬

ferences which prompted Broadley (2000b) to treat them

as separate species.

Serpentes

Colubridae

Telescopus semiannulatus semiannulatus Smith, 1849

Eastern Tiger Snake

Photograph: VMUS 158924

Recorded on a number of occasions in the vicinity of

Ngonye Falls campsite and visitor’s center. This species

appears to be widespread throughout Zambia (Broadley

1971a).

Dispholidus typiis viridis (Smith, 1828)

Green Boomslang

Photographs: VMUS 158882, 158884

Regularly recorded at Ngonye Falls campsite, around the

visitor’s center, as well as at Idjobwa Pan in the northern

buffer zone of Sioma Ngwezi National Park. Widespread

in the southern provinces of Zambia (Broadley 1971a).

Thelotornis capensis oatesii (Gunther, 1881)

Oates’s Vine Snake

Photograph: VMUS 158925, 158926, 158928

Commonly encountered at Ngonye Falls campsite, and

widespread in Zambia (Broadley 1971a).

Philothamnus angolensis Bocage, 1882

Western Green Snake

Material: PEM R22028; TM 86231

Photograph: VMUS 158913

A common, diurnal species seen regularly along the

banks of the Zambezi River. One individual (VMUS

158913) was photographed at Ngonye Falls campsite,

one (TM 86231) had drowned in the Zambezi River and

one (PEM R22028) was collected on the eastern bank of

the Zambezi River below Ngonye Falls. It is widespread

in Western Province, as well as in the northern provinces

of Zambia (Broadley 1971a).

Natricidae

Limnophis bangweolicus (Mertens, 1936)

Eastern Striped Swamp Snake

Material: PEM R22926; TM 86203, 86249

One individual (TM 86203) was found emerging from

the Zambezi River at dusk at Ngonye Falls campsite; a

second individual (TM 86249) was found dead and par¬

tially desiccated at the same site; while a third was found

partially consumed on the east bank of the Zambezi River

just below Ngonye Falls. Near-endemic.

Elapidae

Dendroaspis polylepis Gunther, 1864

Black Mamba

Photographs: VMUS 158880, 158881, 158883

Regularly observed at Ngonye Falls campsite and visi¬

tor’s center. The species is widespread in Zambia (Broad¬

ley 1971a).

Naja nigricollis Reinhardt, 1843

Black-necked Spitting Cobra

Photographs: VMUS 158909, 158910

Individuals were encountered regularly at Ngonye Falls

campsite and near the visitor’s center. This species

occurs widely in central and northern Zambia, but is

largely replaced by N. mossambica in the south (Broad¬

ley 1971a; Broadley et al. 2003).

Lamprophiidae

Amblyodipsaspolylepis (Bocage, 1873)

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Pietersen et al.

Common Purple-glossed Snake

Photograph: VMUS 158870

Individuals were photographed at Ngonye Falls camp¬

site and Park Headquarters. This species is widespread

in Zambia, and is particularly common in sandy regions

(Branch 1998; Broadley 197la,c; Broadley et al. 2003).

Hemirhagerrhis nototaenia (Gunther, 1864)

Eastern Bark Snake

Photographs: VMUS 158891, 158893-158895, 158900

Observed frequently at Ngonye Falls campsite, around

the visitor’s center, and at Park Headquarters. The spe¬

cies is widespread in Zambia (Broadley 1971a).

Lycophidion multimaculatum Boettger, 1888

Spotted Wolf Snake

Photograph: VMUS 158898

An individual was photographed at Ngonye Falls camp¬

site. The species is widely distributed in western and

northern Zambia (Broadley 1971a; Broadley et al. 2003).

Psammophis mossambicus Peters, 1882

Olive Sand Snake

Photographs: VMUS 158914-158916

Commonly encountered at Ngonye Falls campsite, visi¬

tor’s center, Park Headquarters and on the roads in the

vicinity. The species is common and widespread through¬

out Zambia (Broadley 1971a, 2002).

Psammophis subtaeniatus Peters, 1882

Western Stripe-bellied Sand Snake

Photograph: VMUS 158917

This diurnal species was often encountered at Ngonye

Falls campsite, around the visitor’s center, at Park Head¬

quarters, Maziba Bay, and vicinity. Individuals were

identified by coloration (including presence of a broad

yellow mid-ventral band flanked on each side by a dark

longitudinal stripe) and having the preocular in contact

with the frontal (well-separated in P. mossambicus). This

species has not previously been recorded from Western

Province (Broadley 1971a, 2002; Broadley et al. 2003),

having been recorded peripherally in Southern and

Central Provinces, although more widespread in East¬

ern Province (Broadley 1971a, 2002). It is widespread

in adjacent northern Namibia (Branch 1998; Broadley

2002). These records extend the known distribution by

about 110 km to the north-west.

Pseudaspis cana (Linnaeus, 1758)

Mole Snake

Photographs: VMUS 158918, 158919, 158922, 158923

This common, diurnal snake was regularly seen in the

vicinity of Ngonye Falls campsite, visitor’s center, Park

Headquarters, and surrounding area. A number of indi¬

viduals were also killed by passing traffic on the tarred

road running parallel to the Zambezi River. It is wide¬

spread in Zambia (Broadley 1971a).

Xenocalamus mechowii Peters, 1881

Elongate Quill-snouted Snake

Material: TM 86247

Photograph: VMUS 158937

Individuals were recorded at Ngonye Falls campsite and

Park Headquarters, where a number of individuals (e.g.,

TM 86247) were unearthed during construction. Two

subspecies have been historically recognized, which

were separated on distribution and ventral and subcau-

dal scale counts (Peters 1881, Witte and Laurent 1947;

Broadley 1971c). Broadley (1971c) notes a population

of apparent intergrades in northern Zambia and this,

together with the overlap in the supposedly diagnostic

characters, lead us to not recognize these subspecies until

a thorough review of these taxa has been undertaken.

Leptotyphlopidae

Leptotyphlops scutifrons (Peters, 1854)

Peters’ Thread Snake

Material: PEM R22025

A single individual was collected in a pitfall trap on the

east bank of the Zambezi River, opposite Ngonye Falls.

The only previous Zambian locality is Kalichero in East¬

ern Province, although it has been recorded in Namibia at

Katima Mulilo (Broadley and Broadley 1999).

Pythonidae

Python natalensis Smith, 1840

Southern African Python

Photograph: VMUS 158927

A number of individuals were seen around Ngonye Falls

visitor’s center and campsite, and locals also report the

presence of this species in the vicinity. This species is

widespread throughout Zambia (Broadley 1971a).

Typhlopidae

Afrotyphlops mucruso (Peters, 1854)

Zambezi Beaked Blind Snake

Material: TM 81409

Although not recorded by ourselves, this species has

been collected at Ngonye Falls (TM 81409; Broadley and

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Herpetofauna of Ngonye Falls

Wallach 2009). This species is widespread in Zambia,

while its sister species A. schlegelii has been collected

south of the Zambezi River at Katima Mulilo (Broadley

and Wallach 2009).

Afrotyphlops schmidti (Laurent, 1956)

Schmidt’s Beaked Blind Snake

Material: PEM R22016; TM 86246

Photograph: VMUS 158865

A juvenile specimen (PEM R22016) was collected in a

pitfall trap on the eastern bank of the Zambezi River just

above Ngonye Falls, while another specimen (TM 86246)

was found dead on the dirt road leading to Sioma Barge.

Individuals were regularly encountered around Ngonye

Falls campsite and visitor’s center, especially after rains.

The nearest published locality is Kalabo (Broadley and

Wallach 2009), about 210 km to the NNW.

Viperidae

Bit is arietans arietans Merrem, 1820

Puff Adder

Photographs: VMUS 158871, 158872, 158874

This species was often encountered at Ngonye Falls

campsite and visitor’s center, and is widespread in south¬

ern and central Zambia (Broadley 1971a).

Crocodylia

Crocodylidae

Crocodylns niloticus Faurenti, 1768

Nile Crocodile

Photographs: VMUS 158875, 158878, 158879

Observed in the Zambezi River both above and below

Ngonye Falls, including at Maziba Bay and at Sioma

Barge. It is widespread in Zambia and adjacent regions

(Broadley 1971a).

Testudines

Testudinidae

Kinixys spekii Gray, 1863

Speke’s Hinged Tortoise

Photograph: VMUS 158897

Seen around Park Headquarters and visitor’s center.

This species is widespread in Zambia (Broadley 1971a).

These records are about 100 km north of the nearest

known population in the Zambezi Region of Namibia

(Branch 1998).

Stigmochelys pardalis (Bell, 1828)

Feopard Tortoise

Photograph: VMUS 158920

Seen in the vicinity of Park Headquarters, while a fresh

carapace was confiscated from a poacher in Sioma

Ngwezi National Park. Additional carapaces are on dis¬

play in the visitor’s center, and these may have been col¬

lected in the general vicinity of the study area. This spe¬

cies is most common in the Eastern Province of Zambia,

but has also been recorded from Fivingstone (Broadley

1971a).

Discussion

We report on three new reptile country records and one

potentially new amphibian country record for Zambia,

bringing the known herpetofauna to 86 amphibian, two

crocodile, 10 chelonian, 78 lizard, and 91 snake species.

The herpetofauna of Ngonye Falls and surroundings

is similar to that of south-eastern Angola (Conradie et al.

2016), with 13 amphibian and 34 reptile species in com¬

mon. There are an additional 10 reptile and one amphib¬

ian species with closely related species in Angola. This is

not surprising considering the similarity in habitat. How¬

ever, the list of species in common is largely devoid of

habitat specialists, suggesting that this apparent connec¬

tivity may not apply to all taxa. Although this may be

an artifact of the relatively small sample sizes to date,

these results also suggest that there are barriers to some

species, most likely posed by the drainage basins. This

was observed for some species (e.g., Acontias spp.: this

study; Elapsoidea spp.: Broadley 1971b), for which the

Zambezi River apparently acts as a dispersal barrier.

Fossorial taxa are also largely absent in central Zambia

(Wagner et al. 2012a), largely as a result of the Kala¬

hari sands reaching their eastern limit not far beyond the

Zambezi River. Vegetation, geology and natural barriers

therefore all appear to play a role in shaping the herpeto¬

fauna of western Zambia.

Based on our present knowledge it would appear that

members of the genus Acontias have parapatric distribu¬

tions, with A. k, kgalagadi occurring west of the Cuando

River and south of the Kavango River, A.jappi occurring

between the Cuando and Zambezi Rivers, and A. schmitzi

occurring east of the Zambezi River. The Zambezi River

is likely to pose a formidable barrier to subterranean spe¬

cies such as Acontias and it is thus likely that this river

effectively separates A. jappi and A. schmitzi. Further¬

more, considering their distribution, it is probable that

these two taxa are sister species (see also Wagner et al.

2012a). It will be informative to construct a dated phy-

logeny of these taxa to investigate whether the Zambezi

River may have played a role in their speciation. The fac¬

tors separating the distributions of A.jappi and A. k. kga¬

lagadi are less clear. Although A. k. kgalagadi has thus

far been recorded only west of the Cuando River, and A.

jappi only to the east, the Kalahari sands extend beyond

the source of the Cuando River, and this river may there-

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December 2017 | Volume 11 | Number 1 | e148

Pietersen et al.

fore only pose a local barrier between these two spe¬

cies. Additional sampling is required to more accurately

determine the distribution of these two fossorial taxa, to

investigate the probable barriers between them, and to

determine whether they do in fact occur sympatrically

anywhere.

Broadley (1967) described Ichnotropis grandiceps

from three specimens collected 40 km west of Mohembo,

Botswana, near the Botswana-Caprivi Strip (now the

Zambezi Region of Namibia) border (ca. 18°19’03”S,

21°12’03”E). Haacke (1970) subsequently collected

this species at Ndobe on the Namibia-Botswana border

(ca. 19°34’41”S, 20°59’58”E, TM 30822), the farm Deo

Volente near Grootfontein, Namibia (ca. 19 o 02 , 01 ,, S,

18°46’29”E, TM 38309 and 38310) and on the Caprivi

Strip-Botswana border 16 km east of the 21° corner bea¬

con (i.e., approximately at the type locality). To the best

of our knowledge, this species has not been seen or col¬

lected since Haacke (1970). In our experience I. grandi¬

ceps was a relatively common and active, diurnal spe¬

cies and it seems intriguing that it has not been recorded

for more than four decades. The type specimens were

collected in open woodland on Kalahari sands (Broad¬

ley 1967), while Haacke (1970) collected one specimen

on hard limy soil in Combretum-Acacia bushveld, and

three specimens on white sand in open bushveld. We

found individuals in open to relatively closed Baikiaea

woodland on pale, deep Kalahari sands. The type series

consists of two adult males and a “juvenile,” but mea¬

surements are only provided for the largest individual

(Broadley 1967). Based on measurements presented in

the article, three of the specimens collected by Haacke

(1970) were adults, while the fourth individual was a

subadult. Interestingly, all of the specimens collected to

date have been taken in either April or May (Broadley

1967; Haacke 1970; this study). These are the first pub¬

lished records of this species in Zambia, with all previous

records from south-west of the Kavango River. The pres¬

ence of this species between the Cuando and Zambezi

Rivers suggests that it may also occur in adjacent south¬

eastern Angola. However, to date, the only Ichnotropis

species collected in south-eastern Angola do not match

the description of I. grandiceps and appear to represent

undescribed taxa (Conradie et al. 2016).

Our results suggest that western Zambia, and the

region around Ngonye Falls in particular, has a suite of

taxa in common with adjoining regions of Zambia and

south-eastern Angola, as well as a suite of apparently

unique taxa. However, most surveys to data have been

restricted to the regions immediately adjacent to the Zam¬

bezi River, probably because of easy access, and it would

be insightful to conduct surveys away from the Zambezi

River to gain a better understanding of the entire herpe-

tological assemblage in this region.

Our list should be regarded as preliminary, as numer¬

ous additional species are known from the general vicin¬

ity and are likely to be recorded here in the future. There

are unconfirmed sightings of Acanthocercus atricof-

lis atricollis (Smith, 1849) from the east bank of the

Zambezi River opposite Ngonye Falls, while we also

observed an Aparallactus capensis capensis Smith, 1849

in Ngonye Falls campsite and Crotaphopeltis hotam-

boeia (Faurenti, 1768) individuals in the northern buf¬

fer zone of Sioma Ngwezi National Park at Idjobwa Pan,

as well as in Ngonye Falls campsite, but did not secure

photographic evidence or voucher specimens. Varanns

niloticus (Finnaeus, 1766) was commonly observed in

the Zambezi River in the vicinity of Ngonye Falls, while

three juvenile lacertids with a striped dorsal pattern which

were seen at Ngonye Falls campsite may be referable to

Nucras ornata (Gray, 1864), but this remains to be veri¬

fied. There are carapaces of Pelomedusa subrufa (Bonna-

terre, 1789) and Pelusios bechuanicus FitzSimons, 1932

on display in the visitor’s center, and although these are

suspected to have been sourced from the vicinity of Ngo¬

nye Falls and/or Sioma Ngwezi National Park, this could

not be verified.

Based on their occurrence in similar habitat in nearby

areas, the following additional amphibian species are

expected to occur in the region: Leptopelis bocagii (Gun¬

ther, 1865); Sderophrys lemairii (Boulenger, 1901);

Hyperolins nasntns Gunther, 1865; Kassina Senegalensis

(Dumeril and Bibron, 1841); Phrynomantis affinis Bou¬

lenger, 1901; P. bifasciatus (Smith, 1847); Hildebrand-

tia omata (Peters, 1876); Phrynobatrachus mababien-

sis FitzSimons, 1932; Ptychadena grandisonae Faurent,

1954; P. nilotica (Seetzen, 1855); P. porosissima (Stein-

dachner, 1867); P. taenioscelis Faurent, 1954; Pyxiceph-

ahis adspersus Tschudi, 1838; and Amnirana darlingi

(Boulenger, 1902) (Channing 2001; Conradie et al. 2016;

Furman et al. 2015; Poynton and Broadley 1985a,b,

1987,1988,1991).

Anumber of additional reptile species are also expected

to occur in the region based on their presence at nearby

locations, including: Pelusios rhodesianus Hewitt, 1927;

Lygodactylus angolensis Bocage, 1896; Pachydacty-

1ns punctatus Peters, 1854; Varanns albignlaris Daudin,

1802; Monopeltis mauricei Parker, 1935; Amblyodip-

sas ventrimaculata (Roux, 1907), Boaedon capensis

Dumeril, Bibron and Dumeril, 1854; Crotaphopeltis bar-

otseensis Broadley, 1968; Philothamnus semivariegatus

(Smith, 1840); Psammophis lineatus (Dumeril, Bibron

and Dumeril, 1854); P. angolensis (Bocage, 1872); Dasy-

peltis scabra (Finnaeus, 1758), Naja anchietae Bocage,

1879; Atractaspis bibronii Smith, 1849 and Causus

rhombeatus (Fichtenstein, 1823) (Bayless 2002; Branch

1998; Broadley 1968, 1971a,c, 1991b, 1995, 2000a,

2002, 2014; Broadley et al. 1976; Conradie et al. 2016;

Haagner et al. 2000; Hughes 2004; Foveridge 1958; Ras¬

mussen 1997, 2005; Roux 1907).

Acknowledgements. —We are grateful to John Davies

for field assistance, and to the Zambia Wildlife Author¬

ity (ZAWA) for granting us permission to search for rep-

Amphib. Reptile Conserv.

December 2017 | Volume 11 | Number 1 | e148

Herpetofauna of Ngonye Falls

tiles. WC thanks Michiel Jonker for field assistance and

logistics. Roger Bills (South African Aquatic Biodiver¬

sity Institute) is thanked for providing additional mate¬

rial collected from the Ngonye Falls region. We thank

William R. Branch, Michael F. Bates, and Philipp Wag¬

ner for commenting on, and greatly improving, an earlier

draft of this manuscript.

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Pietersen et al.

Appendix 1 . List of amphibian and reptile species recorded at Ngonye Falls and surrounding regions in south-western Zambia,

indicating voucher type and accession number. Museum acronyms are: PEM: Port Elizabeth Museum; SAIAB: South African

Aquatic Biodiversity Institute, Grahamstown and TM: Ditsong National Museum of Natural Elistory, Pretoria. All photographs are

accessioned into the FrogMAP and ReptileMAP platforms of the Virtual Museum, Animal Demography Unit, University of Cape

Town (available: vmus.adu.org.za).

Species

Voucher Number

Virtual Museum Number

ORDER: ANURA

FrogMAP

BREVICEPTIDAE

Breviceps adspersus adspersus Peters, 1882

5982

Breviceps poweri Parker, 1934

5983

BUFONIDAE

Poyntonophrynus fenoulheti (Hewitt and Methuen, 1913)

5989

Schismaderma carens (Smith, 1848)

SAIAB 205361,205631

5992-93

Sclerophrys gutturalis (Power, 1927)

5994

Sclerophrys poweri (Hewitt, 1935)

SAIAB 205356

5996-97

Sclerophryspusilla (Hallowell, 1855)

PEM All 719-20; SAIAB 205360

5995, 5998

HEMISOTIDAE

Hemisus marmoratus (Peters, 1854)

5985

HYPEROLIIDAE

Hyperolius angolensis Steindachner, 1867

5986

PHRYNOBATRACHIDAE

Phrynobatrachus natalensis (Smith, 1849)

SAIAB 205351

5987

Phrynobatrachus parvulus (Boulenger, 1905)

5988

PIPIDAE

Xenopus muelleri (Peters, 1844)

SAIAB 202357

Xenopus poweri Hewitt, 1927

SAIAB 202355

PTYCHADENIDAE

Ptychadena cf. mapacha Channing, 1993

TM 86255

5990

Ptychadena oxyrhynchus (Smith, 1849)

SAIAB 205353-54

5991

Ptychadena subpunctata (Bocage, 1866)

PEM Al 1717-18; SAIAB 205358,

205365

PYXICEPHALIDAE

Tomopterna cf. cryptotis (Boulenger, 1907)

SAIAB 205362

5999, 6000

RHACOPHORIDAE

Chiromantis xerampelina Peters, 1854

5984

ORDER: SQUAMATA

ReptileMAP

SAURIA - AGAMIDAE

Agama armata Peters, 1855

PEM R22017-18

158866-69

AMPHISBAENIDAE

Dalophia longicauda (Werner, 1915)

163523

Dalophia pistil!um (Boettger, 1895)

PEM R22925

Monopeltis anchietae (Bocage, 1873)

TM 86250

Zygaspis nigra Broadley and Gans, 1969

TM 86209

158938

Zygaspis quadrifrons (Peters, 1862)

TM 86208

158939

CHAMAELEONIDAE

Chamaeleo dilepis Leach, 1819

158873,158876-77

Amphib. Reptile Conserv.

December 2017 | Volume 11 | Number 1 | e148

Herpetofauna of Ngonye Falls

Appendix 1 (continued). List of amphibian and reptile species recorded at Ngonye Falls and surrounding regions in south¬

western Zambia, indicating voucher type and accession number. Museum acronyms are: PEM: Port Elizabeth Museum; SAIAB:

South African Aquatic Biodiversity Institute, Grahamstown and TM: Ditsong National Museum of Natural Flistory, Pretoria. All

photographs are accessioned into the FrogMAP and ReptileMAP platforms of the Virtual Museum, Animal Demography Unit,

University of Cape Town (available: vmus.adu.org.za).

Species

Voucher Number

Virtual Museum Number

GEKKONIDAE

Pachydactylus wahlbergii wahlbergii (Peters, 1869)

163521-22

Hemidactylus mabouia (Moreau De Jonnes, 1818)

PEM R22019-20; TM 86251-52

158888-90, 158892

Lygodactylus chobiensis FitzSimons, 1932

PEM R22026; TM 86253

158901, 158903-06

GERRHOSAUR1DAE

Gerrhosaurus auritus Boettger, 1887

158885

Gerrhosaurus intermedius Lonnberg, 1907

158887

LACERT1DAE

Ichnotropis capensis (Smith, 1838)

PEM R22021-24

158896,158902

Ichnotropis grandiceps Broadley, 1967

TM 86237

Meroles squamulosus (Peters, 1854)

158907

SCINCIDAE

Acontias jappi (Broadley, 1968)

TM 86232-34, TM 86254

Acontias schmitzi Wagner, Broadley and Bauer, 2012

PEM R22015

Mochlus sundevallii (Smith, 1849)

PEM R22027

158908

Panaspis maculicollis Jacobsen and Broadley, 2000

158911

Typhlacontias rohani Angel, 1923

TM 86235-36, 86248

Trachylepis damarana (Peters, 1870)

PEM R22030-31

158929-30, 158933-35

Trachylepis wahlbergii (Peters, 1869)

PEM R22029

158931, 158936

SERPENTES - COLUBRIDAE

Telescopus semiannulatus semiannulatus Smith, 1849

158924

Dispholidus typus viridis (Smith, 1828)

158882,158884

Thelotornis capensis oatesii (Gunther, 1881)

158925-26, 158928

Philothanmus angolensis Bocage, 1882

PEM R22028; TM 86231

158913

NATRICIDAE

Limnophis bangweolicus (Mertens, 1936)

TM 86203, 86249; PEM R22926

ELAPIDAE

Dendroaspis polylepis Gunther, 1864

158880-81, 158883

Naja nigricollis Reinhardt, 1843

158909-10

LAMPROPHIIDAE

Amblyodipsas polylepis (Bocage, 1873)

158870

Hemirhagerrhis nototaenia (Gunther, 1864)

158891, 158893-95, 158900

Lycophidion multimaculatum Boettger, 1888

158898

Psammophis mossambicus Peters, 1882

158914-16

Psammophis subtaeniatus Peters, 1882

158917

Pseudaspis cana (Linnaeus, 1758)

158918-19, 158922-23

Xenocalamus mechowii Peters, 1881

TM 86247

158937

LEPTOTYPHLOP1DAE

Leptotyphlops scutifrons (Peters, 1854)

PEM R22025

Amphib. Reptile Conserv.

December 2017 | Volume 11 | Number 1 | e148

Pietersen et al.

Appendix 1 (continued). List of amphibian and reptile species recorded at Ngonye Falls and surrounding regions in south¬

western Zambia, indicating voucher type and accession number. Museum acronyms are: PEM: Port Elizabeth Museum; SAIAB:

South African Aquatic Biodiversity Institute, Grahamstown and TM: Ditsong National Museum of Natural Ehstory, Pretoria. All

photographs are accessioned into the FrogMAP and ReptileMAP platforms of the Virtual Museum, Animal Demography Unit,

University of Cape Town (available: vmus.adu.org.za).

Species

Voucher Number

Virtual Museum Number

PYTHONIDAE

Python natalensis Smith, 1840

158927

TYPHLOPIDAE

Afrotyphlops mucruso (Peters, 1854)

TM 81409

Afrotyphlops schmidti (Laurent, 1956)

PEM R22016; TM 86246

158865

VIPERIDAE

Bitis arietans arietans Merrem, 1820

158871-72, 158874

ORDER: CROCODYLIA

CROCODYLIDAE

Crocodylus niloticus Laurenti, 1768

158875, 158878-79

ORDER: TESTUDINES

TESTUDINIDAE

Kinixys spekii Gray, 1863

158897

Stigmochelyspardalis (Bell, 1828)

158920

Darren Pietersen is a Ph.D. candidate at the University of Pretoria Department of Zoology

and Entomology, and a research associate of the Ditsong National Museum of Natural History

in Pretoria. His main interests are reptile taxonomy, as well as general reptile and amphibian

surveys and the ecology of these taxa. Darren has authored or co-authored a number of scientific

and popular articles. He has conducted herpetological surveys in a number of African countries,

including Mozambique and the Democratic Republic of the Congo.

Errol Pietersen is a conservationist with a passion for reptiles and amphibians. His work has

taken him to various interesting locations in Africa, where he has contributed to the knowledge of

the herpetofauna of these regions. Errol has co-authored various popular articles on reptiles and

amphibians, in particular on the diversity of these taxa at various sites in Mozambique.

Werner Conradie has ten years of experience in southern African herpetofauna, with his main

research interests focusing on taxonomy, conservation, and ecology of amphibians and reptiles. He

has published numerous principal and collaborative scientific papers and has served on a number

of conservation and scientific panels, including the Reptile Atlas Committee and Amphibian

IUCN Workshop. Werner has represented his field on television and in numerous field guides

and has participated in expeditions in various countries including Namibia, Botswana, Zimbabwe,

Mozambique, Angola, Malawi, Lesotho, and Zambia. He is currently the Curator of Herpetology

at the Port Elizabeth Museum (Bayworld), South Africa.

Amphib. Reptile Conserv.

December 2017 | Volume 11 | Number 1 | e148

Official journal website:

amphibian-reptile-conservation.org

Amphibian & Reptile Conservation

11(1) [General Section]: 1-16 (e132).

Cryptic multicolored lizards in the Polychrus marmoratus

Group (Squamata: Sauria: Polychrotidae) and the status of

Leiolepis auduboni Hallowell

15 John C. Murphy, 2 Richard M. Lehtinen, 3 Stevland P. Charles,

^Danielle Wasserman, 'Tom Anton, 2 Patrick J. Brennan

1 Science and Education, Field Museum of Natural History, 1400 Lake Shore Drive, Chicago, Illinois 60616 USA 2 The College of

Wooster, Department of Biology, 931 College Mall, Wooster, Ohio, 44691 USA 3 Department of Biology, Howard University, 415

College Street NW, Washington, DC 20001 USA 4 Vertebrate Museum, Department of Biology, Southeastern Louisiana University,

Hammond, Louisiana 70402 USA

Abstract.—The Neotropical genus Polychrus contains seven species of arboreal lizards. The type species for

the genus is the widespread Polychrus marmoratus. We compared a few populations of P. marmoratus using

16S and COI mitochondrial gene sequences (1,035 bp total) and found several lineages existing under the name

Polychrus marmoratus. Working backwards, using morphology we identify Polychrus marmoratus from the

Guiana Shield and resurrect the name Leiolepis auduboni Hallowell for the species present in Trinidad, Tobago,

and northern Venezuela. The number of species in the genus Polychrus is raised to eight. However, we also

discuss evidence for the existence of other cryptic species within P. marmoratus, and the likelihood that both

P. virescens Schniz and P. neovidanus Wagler are valid names.

Keywords. Guyana Shield, Venezuelan Coastal Ranges, Trinidad, Tobago, Atlantic Forest, reptiles

Citation: Murphy JC, Lehtinen RM, Charles SP, Wasserman D, Anton T, Brennan PJ. 2017. Cryptic multicolored lizards in the Polychrus marmoratus

Group (Squamata: Sauria: Polychrotidae) and the status of Leiolepis auduboni Hallowell. Amphibian & Reptile Conservation 11(1) [General Section]:

1-16 (el32).

NonCommercialNoDerivatives 4.0 International License, which permits unrestricted use for non-commercial and education purposes only, in any

medium, provided the original author and the official and authorized publication sources are recognized and properly credited. The official and

authorized publication credit sources, which will be duly enforced, are as follows: official journal title Amphibian & Reptile Conservation ; official journal

website <amphibian-reptile-conservation.org>.

Received: 26 March 2016; Accepted: 04 September 2016; Published: 16 January 2017

Introduction

The multi-colored lizards of the genus Polychrus have

traditionally been treated as part of the Iguanidae since

Gray (1845) organized lizards into 24 families. Townsend

et al. (2011) note that, while Polychrus and Anolis have

been considered sister taxa (Frost and Etheridge 1989),

all published analyses of molecular data contradict this

finding (Frost et al. 2001; Schulte et al. 2003; Schulte

and Cartwright 2009). Frost et al. (2001) defined Poly¬

chrotidae based on morphology-only in a combined mo¬

lecular and morphological analysis. Using conventional

alignment methods and a much larger molecular data set

Schulte et al. (2003) failed to recover the Polychrotidae

clade. More recently, Pyron et al. (2013) suggested Poly¬

chrus is more closely related to hoplocerids. Hoploceri-

dae contains four genera and about 24 species of medium

sized Neotropical endemic saurians with spiny tails,

pleurodont teeth, diurnal activity, arboreal life styles, and

an omnivorous diet. Members of the genus Hoplocercus

are dry forests lizards, while Enyalioides and Moruna-

saurus inhabit rain forests. Polychrus , on the other hand,

spans a range of habitats, has an exceptionally long, non-

spiny tail, and is diurnal, omnivorous and arboreal. The

slow moving, arboreal Polychrus range from Honduras

southward into continental South America on both sides

of the Andes (Avila-Pires 1995; this study), extending as

far as 25.63°S on the east side of the Andes.

Polychrus is composed of seven recognized species: P.

marmoratus Linnaeus 1758, P. acutirostris Spix 1825, P.

gutturosus Berthold 1846, P. liogaster Boulenger 1908,

P. femoral is Werner 1910, P. peruvianus Noble 1924, and

the most recently described P. jacquelinae Koch et al.

2011. Boulenger’s (1914) P. spurrelli was considered a

Correspondence. Email: 5 serpentresearch@gmail.com (Corresponding author)

Amphib. Reptile Conserv.

January 2017 | Volume 11 | Number 1 | el 32

Murphy et al.

subspecies of P. gutturosus by Peters and Donoso-Barros

(1970), however Koch et al. (2011) considers the name a

junior synonym of P. gutturosus.

Much of the distribution of Polychrus is occupied by

the type species for the genus, Polychrus marmoratus.

It occurs in South America east of the Andes, including

Guyana, French Guyana, Suriname, Venezuela, Colom¬

bia, Ecuador, Peru, Brazil, and Paraguay (Hoogmoed

1973; Avila-Pires 1995). Vanzolini (1983) noted P. mar¬

moratus ’ distribution was disjunct with a large gap in

northeastern Brazil separating the Caribbean and north¬

ern Amazonian populations from the Atlantic Forest

populations. A museum specimen (MCZ R-25135) docu¬

ments Breder’s (1946) report of its presence in southern

Panama. Other museum specimens suggest its presence

in Valparaiso, Chile (MCZ R-3369) and elsewhere on

the west side of the Andes. The species also occurs on

the Isla de Margarita (Ugueto and Rivas 2010), Trinidad

(Court 1858), Tobago (Barbour, 1916), and on the Bo-

cas Islands of Gaspar Grande, Monos, and Chacachacare

(Boos 1984a, b, 1990). This distribution covers about 37°

of latitude and 45° of longitude with populations rang¬

ing in elevation from sea level to at least 2,500 m above

sea level (asl) in Venezuela. Rivas et al. (2012) reported

Polychrus marmoratus occurs in all of the biogeographic

divisions of Venezuela except the marine, coastal, and

Pantepui divisions. Molina et al. (2004) suggests it is

present in the gallery forests and swamp forests of the

llanos. It is not likely to be present in open grasslands.

Figure 1 illustrates specimens from six different portions

of the range.

Lacerta marmorata Linnaeus (1758:208) was based

upon specimens documented in the Museum Principis

and the Amphibia Gyllenborgiana. Specimens from the

later collection were missing when Lonnberg (1896)

examined the collections. However, Holm (1957) later

found four specimens in the Amphibia Gyllenborgiana

apparently overlooked by Lonnberg (1896). Cuvier

(1817) established the genus Polychrus for Lacerta mar¬

moratai, citing Lacepede’s (1788) illustration which he

considered accurate enough for identification. Hoogmoed

(1973) restricted the type locality for Lacerta marmorata

to Paramaribo, Suriname. A specimen collected by Wied-

Neuwied at the Villa Vigosa, Bahia, Brazil was the basis

for the description of Polychrus virescens Schniz (1822).

However, when Wied-Neuwied (1822-1831) published

on this specimen, he referred to it as P. marmoratus.

Fitzinger (1826) listed the name Polychrus geomet-

ricus as patria ignota (locality unknown) and Vanzolini

(1983) considered it nomen nudum. Delaporte (1826) de¬

scribed Polychrus fasciatus based upon a mounted speci¬

men in the Museum National d'Histoire Naturelle (Paris)

and reported its type locality as the Philippines or Moluc¬

cas. This specimen was not discussed by Dumeril and Bi-

bron (1837) nor Guibe (1954) and remains nomen dubia.

Wagler (1828) described Polychrus strigiventris in a

single sentence in the same account he discusses Poly¬

chrus virescens. He states that the femoral pores and col¬

oration distinguish it from the other species but, provides

no type locality. Wagler’s Polychrus strigiventris is also

a nomen dubia.

Wagler (1833a) described Polychrus neovidanus

(based on Seba’s (1734-1765) plate 76, figure 4 and a

Spix specimen from Rio de Janeiro) on the basis that the

specimen lacked obvious femoral pores (it was a female).

Vanzolini (1983) proposed the name P. neovidanus Wa¬

gler which should be associated with Spix’s specimen

from Rio de Janerio.

Hallowell (1845) described Leiolepis auduboni col¬

lected by Samuel Ashmead at a location within 200 miles

of Caracas, Venezuela. HallowelFs specimen was identi¬

fied as Polychrus marmoratus by Roze (1958), however

its status as a junior synonym of Polychrus marmoratus

has been overlooked in more recent works (Peters and

Donoso-Barros 1970; Avila-Pires 1995).

Numerous taxonomic changes due to underestimated

diversity in other Neotropical lizards (Giugliano et al.

2013; Domingos et al. 2014; Werneck et al. 2015) sug¬

gests Polychrus marmoratus may also be a good candi¬

date for holding undescribed, cryptic species. Polychrus

marmoratus has not been examined in detail since Hoog¬

moed (1973) and Avila-Pires (1995) provided species ac¬

counts in their catalogues for Suriname and Brazil. Here,

we focus on northeastern South America, define Poly¬

chrus marmoratus , and discuss possible cryptic species

within the species.

Methods and Materials

We examined 118 alcohol preserved specimens and 17

skeletal and cleared and stained museum specimens la¬

beled Polychrus marmoratus , as well as five specimens

labeled Polychrus liogaster (listed in species accounts,

Appendix 1 lists other material examined). Localities

with precise information were plotted using Arc View. We

also used localities from VertNet and the literature to pro¬

vide an overall view of the distribution of the Polychrus

marmoratus group. Preserved specimens were fixed in

formalin and stored in 70% ethanol. Morphological data

was collected (JCM, SPC, DW, TA) and morphological

nomenclature used follows Hoogmoed (1973) and Avi¬

la-Pires (1995). Body and tail lengths were taken to the

nearest one mm with a metric ruler, and head and scale

measurements were taken with dial and digital calipers.

Values for paired head scales are given in left/right order.

Univariate analyses of morphological data were conduct¬

ed with Excel in combination with QI Macros. Abbre¬

viations used include: n: number of specimens, X: mean

value, SD: standard deviation, SVL: snout vent length,

and asl: above sea level.

Two tissue samples were collected by RML in To¬

bago while the remaining tissue samples were obtained

via loan from various museums (see Table 1 for list of

localities). Total genomic DNA was extracted from tis-

January 2017 | Volume 11 | Number 1 | el 32

Amphib. Reptile Conserv.

Cryptic multicolored lizards in the Polychrus marmoratus Group

Fig. 1. A-B: Polychrus auduboni from the Arima Valley, Trinidad ( JCM ). C-D: Polychrus marmoratus , probably from Suriname,

(Twan Leenders). E: Polychrus sp. from 4.5 km S of Cumanacoa, Venezuela at 300-400 m asl ( Walter E. Schargel). F: Polychrus

sp. from Vitoria do Xingu, Para, Brazil ( Pedro Peloso). G: Polychrus sp. Camamu, Bahia, Brazil. H: Polychrus sp. from Guarapari,

Espirito Santo, Brazil (bottom) ( Pedro Peloso ).

sues using a DNeasy blood and tissue kit (Qiagen, Inc.,

Valencia, California, USA) following the manufacturer’s

instructions. Using a Qiagen TopTaq PCR Master Mix

kit, we amplified a fragment of the cytochrome oxidase 1

(hereafter, COI) mitochondrial gene (~ 650 bp) using the

primers LCO1490 and HC02198 from Castaneda and de

Queiroz (2011). We also amplified a ~ 430 bp fragment

of the 16S mitochondrial gene using primers from Haas

et al. (1993). Thermocycler conditions for PCR followed

Castaneda and de Queiroz (2011).

Amplicons were purified using a Qiagen Min-Elute

column purification kit and sequenced on an ABI PRISM

3100x1 automated sequencer at the Molecular and Cellu¬

lar Imaging Center at the Ohio Agricultural Research and

Development Center, Ohio State University using the

PCR primers. Bases were called in Codon Code Aligner

(version 4.0.1). Sequences for each gene were aligned

in MEGA 6.06 (Tamura et al. 2013) using the Clustal W

module with the default gap opening and gap extension

penalties. Alignments were unambiguous.

We used both parsimony and maximum likelihood

analyses in MEGA to analyze the combined aligned

sequences of 16S and COI (1,035 bp total). For our

maximum likelihood analysis, we used the Hasegawa-

Kishino-Yano model of molecular evolution as it was

supported by the Bayesian Information Criterion as the

closest fit to our data using MEGA. Initial tree(s) for

the heuristic search were obtained by applying Neigh¬

bor-Joining algorithms to a matrix of pairwise distances

estimated using the Maximum Composite Likelihood

(MCL) approach, and then selecting the topology with

the best log likelihood value. A discrete Gamma distri¬

bution was used to model evolutionary rate differences

among sites (five categories [+G, parameter = 0.1811]).

One thousand bootstrap pseudoreplicates were used to

assess topological support. In the parsimony analysis,

Amphib. Reptile Conserv.

January 2017 | Volume 11 | Number 1 | el 32

Murphy et al.

Fig. 2. A: the paravertebral, lateral and ventral scales of Polychrus marmoratus. B: paravertebral scales of Polychrus auduboni,

a single keel per scale. C: paravertebral scales of Polychrus marmoratus with multiple keels per scale (multicarinate). D: ventral

scales of Polychrus auduboni with keel extending the length of the scale. E: ventral scales of Polychrus marmoratus showing short

keels, with some terminating in a small swollen gland at the apex of some scales (FMNH 3294, Demerara, Guyana).

all characters were unweighted and we used the subtree

pruning-regrafting search method to identify the most

parsimonious tree(s). Five thousand bootstrap pseudo-

replicates were used to assess topological support. In all

analyses, all codon positions were used, positions with

less than 90% site coverage were eliminated, and se¬

quences obtained from P. acutirostris were used as an

outgroup (Table 1). Uncorrected p-distances were calcu¬

lated for all pairs of sequences for each gene separately,

using default settings in MEGA.

Collected specimens were covered by Trinidad and

Tobago Forestry Division Special Game Licenses issued

to JCM and RML on June 18, 2013 and June 5, 2014.

Results

The Polychrus marmoratus Group members share: 85-

119 vertebral scales counted between the occiput to the

posterior margin of the hind legs with a single keel or

multiple keels; nasal scale contacts upper labials 1-2 or

2-3; first pair of chin shields may be in contact, or not;

vertebral crest is absent; parietal eye absent; gular crest

greatly reduced; 62-93 scales around mid-body; 23-35

lamellae on the fourth finger; 30—44 lamellae on the

fourth toe; 10-18 semicircle scales; 5-8 rows of supra-

oculars; 8-28 total pores; weak to strongly keeled ventral

scales. Scales on the snout (those anterior to semicircle

scales) differ in ornamentation and organization within

species, making it difficult to obtain consistent charac¬

ters. The canthals (scales between the preorbitals and the

supranasal) numbered two or three. The loreal is bor¬

dered by the canthals, the preorbitals, the upper labials,

the nasal, and sometimes the suborbitals. The loreal can

be single, divided into two parts, or fragmented into mul¬

tiple parts and is variable within species. Polychrus mar¬

moratus group members lack a gular crest but does have

conical shaped scales on the mid line of the gular region.

Paravertebrals (scales on the vertebral line and similarly

shaped scales on either side of it) are in 8 to 14 transverse

rows. These scales differ from lateral scales in ornamen¬

tation and size. They are often flat and may be ovate or

polygonal with a single keel or two or three keels (multi¬

carinate scales). The ornamentation of these is relatively

consistent within species. Paravertebrals may be similar

in size to lateral scales, or larger or smaller. Lateral scales

are usually convex with a keel. They may be oval or

quadrangular, and the interstitial skin contains numerous

tiny granules. Ventral scales are triangular, larger than

laterals, imbricate, and keeled. The keels may extend the

entirety of the scale or only part of its length, and keels

may end in a swollen bulb. The transition zone between

laterals and ventrals make ventral counts ambiguous. See

Figure 2 for the scale shapes and transition zones and

other scale characters. Polychrus marmoratus has been

Amphib. Reptile Conserv.

January 2017 | Volume 11 | Number 1 | el 32

Cryptic multicolored lizards in the Polychrus marmoratus Group

Table 1. Specimens examined for molecular work, their geographic origins and Genbank accession numbers.

Specimen

Locality

COI

16S

YPM H-013040

none (pet trade specimen)

KY458391

KY458408

YPM H-014659

none (pet trade specimen)

KY458392

KY458409

KU 212631

Peru, San Martin, 14 km ESE of Shapaja

KY458396

KY458413

MVZ 163071

Peru, Amazonas, vicinity of Sua (Aguaruna village), Rio Cenepa

(4°34'12.00"S, 78°13'18.01"W)

KY458406

KY458423

MVZ 230130

none (pet trade specimen, P. acutirostris)

KY458407

KY458424

LSUMZ 14270

Brazil, Para, Agropecuaria Treviso, LTDA, ca 101 km south, 18 km

east Santarem (3°09'2.4"S, 54°50 , 32.9"W)

KY458398

KY458415

LSUMZ 14271

Brazil, Para, Agropecuaria Treviso, LTDA, ca 101 km south, 18 km

east Santarem (3°09'2.4"S, 54°50'32.9"W)

KY458395

KY458412

LSUMZ 14392

Brazil, Para, Agropecuaria Treviso, LTDA, ca 101 km south, 18 km

east Santarem (3°09T0.2"S, 54°50'28.4 n W)

KY458397

KY458414

LSUMZ 4458

Trinidad and Tobago: Trinidad, San Fernando

KY458399

KY458416

AMNH 138080

Guyana: Northern Rupunum Savanna, Yupukari (on Rupununi River),

7 mi (airline) SSW Karanambo, 370 ft

KY458393

KY458410

AMNH 139787

Guyana: Southern Rupunum Savanna, Aishalton (on Kubanawau

Creek), 150 m, (2 0 28'31"N 59°19T6"W)

KY458404

KY458421

CAS 231770

Trinidad and Tobago: Trinidad, Nariva Road, Manzanilla Beach.

(10°29'25.6"N, 61° 03T6.8"W)

KY458402

KY458419

CAS 231781

Trinidad and Tobago: Trinidad, 5 km E of Laguna Mar Beach Resort,

Blanchisseuse. (10°47'39.9"N, 61°17' 46.4" W)

KY458403

KY458420

UWIZM.2012.27.61

Trinidad and Tobago: Trinidad, Arima Valley, (10° 4T 5.57"N,

61°16'54.12"W)

KY458401

KY458418

UWIZM.2012.27.47

Trinidad and Tobago: Tobago, Amos Valle Bridge Courtland River

(~11°12'21.59"N, 60°45'35.99"W)

KY458400

KY458417

UWIZM.2012.42.12

Trinidad and Tobago: Tobago, 1.5 km upstream from Bloody Bay river

bridge

KY458394

KY458411

RML Charlotteville

Trinidad and Tobago: Tobago, west side of Charlotteville (tissue col¬

lected, specimen discarded)

KY458405

KY458422

described as having three dark eye stripes: one extend¬

ing posteriorly over the supratemporals, one extending

to the rictus, and one extending ventrally to the upper

labials. All specimens examined for this study have these

markings, but in some specimens the pigment has faded

from light and chemicals. Coloration in Polychrus mar¬

moratus is highly variable and we have not emphasized

coloration descriptions in alcohol or life for this reason.

The maximum likelihood tree of our 16S and COI se¬

quence data provide evidence for a clade in Trinidad and

Tobago but with little inter-island differentiation (Fig.

3). Both sequences from Guyana group together with

high support, but are strongly divergent from all other

sequences. There is also modest support for a grouping

of the three Brazilian sequences, but the position of the

Peruvian sequences is not well resolved. The parsimony

analysis resulted in a highly similar tree with similar sup¬

port (not shown).

The molecular phylogenetic results are supported by

the p-distance data (Tables 2 and 3). There is little varia¬

tion within sequences from Trinidad and Tobago (mean

16S p-distance 0.3% mean COI p-distance 0.1%) but

a moderate amount when compared to sequences from

elsewhere (mean 16S p-distance 1.5% mean COI p-dis-

tance 1.6%). While the two Guyana sequences are nearly

identical (16S p-distance 0.5%, COI p-distance 0.3%),

they are well differentiated from all the other sequences

(mean 16S p-distance 2.1% mean COI p-distance 3.1%).

The three samples from Para, Brazil are identical in their

16S sequence (mean 16S p-distance 0.0%) but somewhat

different for COI (mean COI p-distance 1.1%). Com¬

pared to all the other sequences, the Brazil samples are

moderately differentiated (mean 16S p-distance 1.1%

mean COI p-distance 2.1%).

Polychrus head scale arrangements and counts are

quite variable and working backwards from the molecu¬

lar data we found a single species present on Trinidad

and Tobago that morphologically appears conspecihc

with some Venezuelan Caribbean Coastal Range popula¬

tions. Guyana and Suriname populations are molecularly

and morphologically distinct from the Trinidad-Tobago-

Venezuela species. The molecular results also suggest a

third lineage is present in Para, Brazil.

Amphib. Reptile Conserv.

January 2017 | Volume 11 | Number 1 | el 32

Murphy et al.

93 r LSU 14271 (Brazil)

LSU 14270 (Brazil)

I—LSU 14392 (Brazil)

83 j~

I— KU 21263157316 (Peru)

MV2 163071 (Peru)

■- 040F 009 E01 (no locality)

i— 659F 007 D01 (no locality)

RML260 (Tobago)

RMLPoly (Tobago)

CAS 231781 (Trinidad

LSU 4458 (Trinidad)

78 CAS 231770 (Trinidad)

UWIZM.2012.27.47 (Tobago)

67 L UWIZM 2012 27 61 (Trinidad)

100

r AMNH 138080 (Guyana)

— AMNH 139787 (Guyana)

130F 005 C01 (P. acutirostris)

I -1

0.005

Fig. 3. Consensus maximum likelihood tree for combined 16S and COI sequence data from seventeen Polychrus tissue samples

(1,035 bp total). Bootstrap support values are indicated at each node, if greater than 50%. Samples are indicated by their museum

accession number and country of origin, if known. The tree is drawn to scale, with branch lengths measured in the number of

substitutions per site. For details of analysis, see text.

Localities for the sources of Polychrus DNA, locali¬

ties from which collected morphological data, type local¬

ities, and localities from which P. marmoratus has been

reported are shown in Figure 4.

In a comparison of Trinidad (n = 3) and Guyana ( n =

1) Polychnis skulls, one of us (DW) found the following.

The parietal in Trinidad specimens had a slight constric¬

tion where the lateral borders give way to the posterior

processes. The dorsal crests of the posterior processes

angled medially so that, from the dorsal perspective, the

lateral surfaces are visible and the medially surfaces are

obscured. The lateral edges of the parietal table form

slight bony lips that do not overhang the lateral surfaces

of the parietal, which are supramedially sloped and un¬

depressed. Braincase: on the supraoccipital process, from

the posterior perspective, there is a median seam running

the length of the process, which extends to the rim of the

foramen magnum (Fig. 5).

The parietal in Guyana specimens had the lateral edg¬

es of the parietal table project laterally, each forming a

shelf that overhangs a depression in the adjacent lateral

surface of the parietal bone. From the dorsal view, the

shape of the parietal table is square and is un-constricted

where the lateral borders give way to the posterior pro¬

cesses. The dorsal crests of the posterior processes angle

laterally so that, from the dorsal perspective, the medial

surfaces are visible and the lateral surfaces are obscured.

Amphib. Reptile Conserv. 6

The braincase of Guyana specimens lacked a hairline

median crest on the supraoccipital process.

The combined molecular and morphological data

suggests Polychnis marmoratus is composed of multi¬

ple lineages. The morphology of the lineage present on

Trinidad, Tobago, and the Coastal Ranges of Venezuela

agrees well with the morphology of the holotype of Leio-

lepis auduboni Hallowell, including the morphology of

the specimens we sequenced from Trinidad (UWIZM

2012.27.61) and Tobago (UWIZM 2012.27.47).

Polychrus auduboni (Hallowell)

new combination

Figure 6 (holotype), la, b

Leiolepis auduboni Hallowell 1845: ANSP 8138, Type

locality: “Colombia within 200 miles of Caracas.”

Collected by Samuel Ashmead. Restricted here to La

Cumbre, Maracay, Aragua, Venezuela (-10.233333

-67.333336).

Polychnis marmoratus marmoratus - Burt and Burt

1933: 41. [in part]

Polychrus marmoratus - Roze 1958:2.

Referred specimens: Skeletal and cleared and stained

specimens examined: Trinidad . AMNH 148543 fe¬

male; FMNH 49840, FMNH 49845, FMNH 49848,

UF 18395, males, UF 18922. Venezuela . FMNH

January 2017 | Volume 11 | Number 1 | el 32

Cryptic multicolored lizards in the Polychrus marmoratus Group

Table 2. Uncorrected p-distances estimating the evolutionary divergence between 16S sequences. The proportion of nucleotides

that differ are shown, including all codons. All positions with less than 75% site coverage were eliminated. There were a total of 428

positions in the final dataset. Analyses were conducted in MEGA 6.06 (Tamura et al. 2013).

YPM 13040 (no locality)

YPM 14659 (no locality)

0.002

AMNH 138080 (Guyana)

0.019

0.021

RML260 (Tobago)

0.009

0.012

0.023

LSU 14271 (Brazil)

0.012

0.014

0.023

0.012

KU 212631 (Peru)

0.012

0.014

0.019

0.012

0.005

LSU 14392 (Brazil)

0.012

0.014

0.023

0.012

0.000

0.005

LSU 14270 (Brazil)

0.012

0.014

0.023

0.012

0.000

0.005

0.000

LSU 4458 (Trinidad)

0.007

0.009

0.021

0.002

0.009

UWIZM.2012.27.47 (Tobago)

0.009

0.012

0.023

0.005

0.012

0.002

UWIZM.2012.27.61 (Trinidad)

0.009

0.012

0.023

0.005

0.012

0.002

CAS 231770 (Trinidad)

0.007

0.009

0.021

0.002

0.009

0.000

0.002

CAS 231781 (Trinidad)

0.007

0.009

0.021

0.002

0.009

0.000

0.002

0.000

AMNH 139787 (Guyana)

0.019

0.021

0.005

0.023

0.026

0.021

0.026

0.021

0.023

0.021

RML Charlotteville (Tobago)

0.009

0.012

0.023

0.005

0.012

0.002

0.005

0.002

0.023

MVZ 163071 (Peru)

0.012

0.014

0.019

0.012

0.005

0.000

0.005

0.010

0.012

0.010

0.022

MVZ 230130 (P. acutirostris)

0.098

0.100

0.103

0.098

0.100

0.095

0.098

0.095

0.100

17791 (Sucre, Venezuela) male. Alcohol specimens

examined: Polychrus auduboni (n = 66). Tobago .

UWIZM.2012.27.47, Arnos Valle Bridge Courtland

River (-11.206 -60.760), UWIZM.2012.42.12, Bloody

Bay River Bridge (11.301070 -60.626965); St. John;

Charlotteville; Charles Turpin Estate (11.3165 -60.5499)

FMNH 217257, USNM 227928-30, UWIZM 2011.30.2

UWIMZ CAREC.R.129, UWIMZ2012.27.42 Charlot¬

teville, Tobago (11.3165 -60.5499). Trinidad . Arima

Valley (10.684883 -61.281702) UWIZM 2012.27.61,

Bush-bush (10.360255 -61.090106), Curepe (10.636930

-61.405493), UWIZM 2010.12.49; Brickfield (10.455021

-61.467952) FMNH 49839,49841,49843,49844,49846;

San Rafael (10.57174 -61.2642325) FMNH 49847,

49849; Port-Of-Spain (10.666667 -61.50579) MCZ

R-79119-79123; San Fernando (10.2833 -61.4667)

R-100484-87, R-119881; Tunapuna-Piarco (10.585543

-61.329526) CM S4846, S6520, S6534, S6539, S6543,

S6561; Trinidad (no specific locality) MCZ R-12065,

R-145299, R-145300-07, R-68888, R-69417, UMMZ

123692, UWIZM 2010.12.47a-c, BMNH 92.9.10.2,

97.7.23.17. Venezuela . Aragua (10.233333 -67.333336),

CM S7412, S7425; UMMZ 124309; Distrito Capi¬

tal (10.46786 -66.90625); CM 22797, 64748, MCZ

R-109009, Falcon (11.016667 -68.566666), R-48729-

30, R-49053; Fa Culebra, base of Duida (3.7299633

-65.80171967) R-58330; Monagas (10.2 -63.533)

R-9981; Sucre (10.147126 -63.808614), MCZ R-50202,

CM S7874, S7915, S7918, S7949; within 200 mi from

Caracas ANSP 8138; Uroma, Yaracuay (10.48.3337

-68.31668) FMNH 29189-91.

Diagnosis: Foreal usually fragmented into two scales;

supranasal not in contact with loreal; two or three in¬

ternasals; loreal contacts upper labials 2—3—4; vertebral

rows 93-112; scales around mid-body 62-80; lamellae

on fourth toe 30-43; usually three scales (2-A) between

the first canthals, and five scales (4-6) between the sec¬

ond canthals; nasal does not usually contact first upper

labial, but does contact the second and third; total femo¬

ral pores 17-28; multicarinate scales in paravertebral

rows few or none; paravertebrals much larger than later¬

als; the number of supraocular rows usually five, rarely

four or six; scales on anterior of snout finely striated with

scattered tiny tubercles; scales on anterior surface of hu¬

merus keeled.

Re-description of holotype: HallowelFs (1845) de¬

scription for this species is of little or no use in distin¬

guishing it from Polychrus marmoratus or other taxa in

this genus. Based upon photographs and the original de¬

scription: Adult female with a snout-vent-length (SVF)

of 101 mm. The head is 0.29 of the SVF; 0.65 times lon¬

ger than wide, as wide as high. Snout blunt. Neck nar¬

rower than the head, almost as wide as the anterior por¬

tion of the body. Body compressed. Tail almost round in

cross section, tapering toward the tip, 2.83 times SVF.

Rostral pentagonal, almost two times as wide as high,

visible from above, bordered posteriorly by two large

postrostral scales. Scales on snout heterogeneous in size,

irregularly polygonal, juxtaposed, rugose. Three scales

across snout between anterior canthals, five scales across

snout between posterior canthals, two canthals between

nasal and supraciliaries, anterior one largest. Supranasals

separated by three scales across the snout. Supraorbital

semicircles more or less distinct, with 6/7 scales, separat¬

ed medially by one row of scales, slightly smaller in size

than those of supraorbital semicircles. Thirty-eight scales

in supraocular region distinctly smaller than those on

snout, polygonal to rounded, juxtaposed, flat and smooth,

irregularly arranged except a row of smaller scales ad¬

jacent to supraciliaries. Supraciliaries 8/9, juxtaposed,

smooth, in a continuous series with canthals. Scales in

Amphib. Reptile Conserv.

January 2017 | Volume 11 | Number 1 | el 32

Murphy et al.

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5 Xntre j

Kl OS '

1'AUAM A

C uritiba o

SANTA

CATARINA

Saoo

Panto ° Santos

Rio de Janeiro

Flonanopolis

RIO

GRANDE DO

SUL Oporto

Alegre

/ /

GUAY /

Fig. 4. Localities of members of the Polychrus marmoratus Group. Green pushpins denote localities from which DNA was sampled.

Red stars denote locations for Polychrus auduboni, black stars denote locations for P. marmoratus. Black circular markers represent

unconfirmed locations for members of the P. marmoratus group based upon the literature and specimens reported in VertNet. The

blue star is the type locality for P. marmoratus (Linnaeus). The purple star is the type locality of P. auduboni Hallowed. The green

star is the type locality for Polychrus virescens Schniz, the orange star is the type locality for P. neovidanus Wagler.

the parietal region are irregular polygonal, juxtaposed,

flat, smooth, intermediate in size between those on snout

and on supraocular region. Scales in interparietal region

polygonal, juxtaposed, rugose, some somewhat swollen.

Parietal eye absent. Loreal region with two scales. Nos¬

tril directed laterally, in upper anterior of a single nasal,

nasal pentagonal and in contact with first and second su-

pralabial. Orbit length 0.29 times head length. Eyelids

partially fused, covered by granules of almost same size

throughout the eyelids. A continuous series of two pre¬

oculars and two suboculars, in direct contact with supra-

labials and four postoculars. Upper labials seven on both

sides, followed to rictus by a few relatively small scales.

Temporal region with seven vertical rows of polygonal or

rounded, juxtaposed, flat, and smooth scales, followed by

two or three rows of granular scales near the ear, delim¬

ited dorsally by a single row of five or six enlarged supra-

temporal scales. Ear opening vertically oval with smooth

margin, tympanum superficial. Mental bell-shaped, al¬

most 2.5 times wide as long. Lower labials taller than

Amphib. Reptile Conserv.

upper labials, six on each side, followed by several small

scales to rictus. Lateral scales on chin slightly larger than

median scales. Gular crest reduced but present, com¬

posed of about 10 enlarged, conical scales between men¬

tal and dewlap (anterior most obscured by thread hold¬

ing tag). Dewlap has scales the same size and structure

as chin scales, which are separated from each other by

an extensible skin covered with granules, reaches level

of forelimbs. Scales on anterior nape relatively small,

granular, and almost rounded, juxtaposed, convex, in

about 17 rows, posteriorly grading into dorsals. Scales

on the sides of the neck about the same size as those on

the nape but more elongated, merging ventrally with the

gulars. Paravertebrals polygonal to rounded, juxtaposed,

to some extent convex but mostly flat and keeled, 110

paravertebral scales in a mid-dorsal line between the

occiput and the posterior margin of the hind limbs. Lat¬

eral scales are of a similar size and same shape as those

of dorsum, convex, smooth, in poorly defined, oblique

rows. Ventrals larger than dorsals, flat, keeled, lanceo-

8 January 2017 | Volume 11 | Number 1 | el 32

Cryptic multicolored lizards in the Polychrus marmoratus Group

Table 3. Uncorrected p-distances estimating the evolutionary divergence between COI sequences. The proportion of nucleotides

that differ are shown, including all codons. All positions with less than 75% site coverage were eliminated. There were a total of 635

positions in the final dataset. Analyses were conducted in MEGA 6.06 (Tamura et al. 2013).

YPM 13040 (no locality)

YPM 14659 (no locality)

0.003

AMNH 138080 (Guyana)

0.028

0.032

RML260 (Tobago)

0.006

0.031

LSU 14271 (Brazil)

0.020

0.021

0.033

0.020

KU 212631 (Peru)

0.006

0.010

0.028

0.009

0.017

LSU 14392 (Brazil)

0.008

0.011

0.031

0.011

0.016

0.008

LSU 14270 (Brazil)

0.019

0.031

0.019

0.002

0.016

0.015

LSU 4458 (Trinidad)

0.003

0.031

0.003

0.024

0.009

0.011

0.022

UWIZM.2012.27.47 (Tobago)

0.003

0.031

0.003

0.024

0.009

0.011

0.022

0.000

UWIZM.2012.27.61 (Trinidad)

0.003

0.031

0.003

0.024

0.009

0.011

0.022

0.000

CAS 231770 (Trinidad)

0.003

0.031

0.003

0.024

0.009

0.011

0.022

0.000

CAS 231781 (Trinidad)

0.003

0.031

0.003

0.024

0.009

0.011

0.022

0.000

AMNH 139787 (Guyana)

0.031

0.036

0.003

0.035

0.036

0.031

0.034

0.035

RML Charlotteville (Tobago)

0.006

0.031

0.000

0.020

0.009

0.011

0.019

0.003

0.035

MVZ 163071 (Peru)

0.003

0.032

0.003

0.024

0.010

0.011

0.023

0.000

0.036

MVZ 230130 (P. acutirostris )

0.000

0.002

0.031

0.004

0.020

0.004

0.007

0.018

0.002

0.035

late, and imbricate, in poorly defined, oblique and trans¬

verse rows. Scales around mid-body about 80. Preanal

plate has scales which are slightly smaller than ventrals,

in nine rows. Preanal pores absent. Femoral pores indi¬

cated on 14/13 notched scales (female). Tail has rhom-

boidal, flat, sharply keeled scales, distinctly larger than

the dorsals, arranged in longitudinal and oblique rows;

keels aligned longitudinally; on ventral surface of the

tail, scales are slightly larger and more rectangular. Tail

not regenerated ending somewhat bluntly. Scales on fore¬

limbs slightly larger than dorsals, polygonal to rounded,

flat, keeled, mostly imbricate but more juxtaposed on up¬

per arm, slightly smaller on ventral aspect of forearms,

towards posterior aspect ventrals become smaller. Scales

on hind limbs are as large as dorsals, polygonal to round¬

ed, keeled, smooth, and imbricate on thigh and ventrally,

juxtaposed on dorsal surface of tibia, slightly larger and

slightly keeled on ventral part of tibia, and slightly lan¬

ceolate. Toward posterior aspect of thighs, both dorsally

and ventrally, scales become distinctly smaller. Subdigi¬

tal lamellae of fingers and toes single, short, with mul¬

tiple keels, 25 under fourth finger, 34 under fourth toe.

Variation . We examined 66 specimens of this species.

Females were significantly larger than males in size (p =

<0.05, 53 df). Body length: females SVL X = 116.3 (/? =

31), males SVL X = 105.1 (n = 24). Tail length: females

X = 291.9 ( n = 26), males X = 277.1 (n = 19), statisti¬

cal tests inconclusive as to significant difference. Fore¬

legs 0.30-0.48 of the SVL, X = 0.37, SD = 0.51; hind

legs 0.37-0.65 of the SVL X = 0.52, SD = 0.60. Rostral

broader than tall, contacts two postrostrals. Scales on

snout slightly imbricate. Nasal with large nare, single su-

pranasal above on the snout, separated by 1-3 internasals.

Supranasal rarely makes contact with loreal. First pair of

canthals (most anterior) separated by three (rarely two)

scales, second pair of canthals separated by 4-6 scales

(usually 5). Semicircle scales 10-16, the total bilateral

average 12.4, they were usually separated by a single

scale (or scale row) anteriorly, and a double row posteri¬

orly. Ciliaries 10-13, usually with two keels, supraocular

scales polygonal to round in five or six rows between

ciliaries and semicircle scales, first row or first two rows

of supraoculars with keels (first row sometimes appears

to be a second row of ciliaries), supraoculars highly vari¬

able in number (26^4). A prenasal scale usually sepa¬

rated the postrostral from the nasal. The nasal was mostly

in contact with second upper labial, sometimes in nar¬

row contact with the first or the third upper labial. Loreal

usually in two parts, an anterior quadrangular scale and

a posterior triangular scale; loreal usually contacts up¬

per labials 2-3—4, sometimes 2-3 or 3-4; first canthal

usually makes contact with loreal; loreal usually not in

contact with supranasal. Upper labials 6-9 per side, usu¬

ally seven and the bilateral average for total upper labials

was 14.86; fifth upper labial usually under middle of eye,

sometimes the seam of 4-5, and rarely the seam of 5-6.

Lower labials 6-9, usually seven, and the total bilateral

average for lower labials was 14.8 scales. Orbital scales:

preorbitals two or three, rarely one; suborbitals usually

two; and postorbitals usually three or four, rarely two.

Temporal scales usually in 7-8 vertical rows between

post orbitals and tympanum, separated from parietal

scales by a row of 4-5 enlarged supratemporals. Gulars

(counted between tympani) 39-60 (X = 46.4), rows of

gulars separated by numerous micro-scales that vary in

size. Dorsal scales in 62-80 rows around mid-body (X

= 71.5); 93-112 scales along the vertebral line from the

occiput to the posterior edge of the hind legs; on the ver¬

tebral line 10-14 rows of slightly enlarged, pentagonal

to hexagonal, keeled scales, which gradually become

elongate and ovate laterally, often losing the keels later¬

ally (note these may appear as a mid-dorsal stripe) tran-

Amphib. Reptile Conserv.

January 2017 | Volume 11 | Number 1 | el 32

Murphy et al.

sitioning into sharply keeled, pointed, imbricate ventrals;

scales similar to the ventrals in size and shape extend

onto the anal plate. Scales on tail pentagonal, heavily

keeled, and imbricate. Scales on limbs keeled, imbricate,

and slightly more ovate proximally than distally.

Coloration: In alcohol the specimen is a uniform

brown, see Fig. 5. In life, highly variable, may change

quickly from bright green with white lateral blotches bor¬

dered in black to almost a uniform brown. See Figure

la, b.

Distribution: Polychrus auduboni occurs in the Cor¬

dillera de Costa of Venezuela, Trinidad and Tobago. It is

also likely present on the Isla de Margarita, in the for¬

ested areas of the llanos, and Orinoco Delta. However,

it may be replaced by another species in Bolivar, Ven¬

ezuela.

Natural History: Beebe (1944) discussed the natural

history of this lizard. Unfortunately, he combines infor¬

mation from both Caripito Venezuela and Kartabo, Guy¬

ana and it is not possible to unravel the comments he

makes about P. marmoratus from Guyana, and from this

species from Venezuela. However, Test et al. (1966) ob¬

served P. auduboni in the Botanical Garden at Caracas

and above Turiamo Pass, Aragua, Venezuela. The latter

individual was kept alive and is described as moving

slowly and deliberately, unless grasshoppers were placed

in its cage, at which time it jumps or runs towards the

insects. The authors also describe tail waving as a human

approached the cage, and interpret this as defensive be¬

havior. During the day, the lizard was green in color but

changed to gray-green at night while it slept on a branch.

Its prey may also include humming birds. We have ob¬

served this lizard on Trinidad and Tobago, active during

the day in secondary forests, climbing through branches

and drinking water from the surface of leaves. Several

times we have observed pairs together in the same or

adjacent trees. Known predators include the hawk Leu-

copternis albicolis , house cats, and the parrot snake Lep-

Fig. 5. On the supraoccipital process, from the posterior

perspective, the presence of a median seam that runs the length

of the process, down to the rim of the foramen magnum (DW).

tophis coeruleodorsus (Murphy 1997; Renoir Auguste,

pers. comm.).

Comparisons: Distinguished from Polychrus marmo¬

ratus by few or no multicarinate paravertebral scales on

the dorsum (many in P. marmoratus ); paravertebral s con¬

sistently larger than laterals; loreal is one or two scales

(3-4 in P. marmoratus)', 17-28 femoral pores (8-19 in P.

marmoratus).

Polychrus marmoratus (Linnaeus)

Figures lc, d.

Lacerta marmorata Linnaeus 1758: 208. Type locality

“Hispania.” Restricted to the vicinity of Paramaribo,

Suriname by Hoogmoed (1973).

Polychrus marmoratus - Cuvier 1817: 41.

Psilocercus marmoratus - Wagler 1821: 341.

Agama marmorata - Daudin 1802: 433.

Polychrus marmoratus marmoratus - Burt and Burt,

1933: 41.

Table 4. A comparison of Polychrus marmoratus and P. auduboni morphology.

Character

P. auduboni

P. marmoratus

n =

loreal scales (usual number)

1-2

2-3

vertebral rows, range

93-112100.34

95-113108.2

mean = SD =

(6.15)

(5.17)

scales around mid-body, range mean = SD =

62-8071.52 (6.15)

63-9378.5 (8.17)

nasal contacts upper labials

2 or 2-3

1-2

total semicircle scales

10-16

12-18

total pores, range mean = SD =

17-2821.41 (2.91)

8-1914.31 (3.31)

canthals

2 or 3

multicarinate paravertebral s

few or none

many

first labial contacts the nasal

rarely

usually

Amphib. Reptile Conserv.

January 2017 | Volume 11 | Number 1 | el 32

Cryptic multicolored lizards in the Polychrus marmoratus Group

Fig. 6. ANSP 8138. The holotype for Leiolepis auduboni Hallowell. Photo credit: Ned Gilmore.

Referred specimens: Skeletal material: Suriname AMNH

141130, AMNH 148544 male, AMNH 141084, UF 56618

(Guyana or Suriname), UF 60914 female, UF 68102.

Alcohol material: (n = 32). Guyana, Demerara, (6.733

-57.983) FMNH 3294, Dunoon Demerara River UMMZ

47632, 53965, 47630, 47751, 47752, 47753, 47754;

Cabacalli Island Moruco River (6.78915 -58.182949)

UMMZ 56467, 56468; Wismar (5.9999 -58.30001)

UMMZ 76685; Kartabo (6.377459 -58.706761) UMMZ

47631, CM S4244, S4245, S5361; no specific locality;

MCZ R-24391, UMMZ 47633, 55839, 55856; Suriname,

Paramaribo (5.442523 -55.09896); MCZ R-8255, CM

44369, 52384, 52385, 44362^4368, 49531-32.

Diagnosis: Loreal usually fragmented into three or

four scales; supranasal scale frequently (about 0.50)

makes contact with loreal. One to three intemasals.

Vertebral rows 95-117; scales around mid-body 66-90.

Snout length is 0.58 of orbit length. Lamellae on fourth

toe 35—44. Usually three scales between first canthals,

and five between the second; nasal contacts first two up¬

per labials. Total femoral pores 8-19. Many multicari-

nate paravertebral scales; paravertebrals not much larger

than laterals. The number of supraocular rows usually six

(rarely 5 or 7). Scales on snout and supraoculars finely

striated to smooth with tiny tubercles. Scales on anterior

surface of humerus keeled; a reduced gular crest starts at

the level of the first or second lower labial.

Variation: Females are significantly larger than males.

Females SVL X = 114.5 (n = 29), males SVL X = 102.9

(n = 11) (p = <0.05, 29 df). Tail length: females X =

292.3 (n = 16), males X = 272 (;n = 10). Forelegs X =

0.40 of SVL; r = 0.35-0.47; hind legs X = 0.52 of SVL,

r = 0.45-0.57.

Rostral broader than tall, contacts two or three

postrostrals. Scales on snout juxtaposed. Nasal with large

nare, one or two supranasal above nasal on the snout,

separated by 3-5 scales. Supranasal frequently makes

contact with loreal. First pair of canthals (most anterior

pair) separated by three (rarely two) scales; second pair

of canthals separated by 4-5 scales (usually five). Semi¬

circle scales 5-9, the total bilateral average 14.4; usually

separated by a single scale (or scale row) anteriorly, and

a double row posteriorly. Ciliaries 10-13, usually with

two keels. Supraocular scales polygonal to round in five

or six rows between ciliaries and semicircle scales, first

row or first two rows of supraoculars with keels (first

row sometimes appears to be a second row of ciliaries);

supraoculars highly variable in number (31—44) and in

five or six rows (rarely seven). A prenasal scale usually

separates the postrostral from the nasal. The nasal is usu¬

ally square and in contact with the first two upper labials,

sometimes in narrow contact with the third upper labial.

Loreal usually in two or three parts, anterior scale quad¬

rangular and posterior triangular scale. Loreal usually

contacts upper labials 2—3—4, sometimes 2-3 or 3-4; first

canthal usually makes contact with loreal. Upper labials

numbered 5-9 per side, usually six; bilateral average for

total upper labials 14.7; upper labial under middle of eye

usually fifth, or 4-5, or 5-6. Lower labials 6-9, usually

7; total bilateral average for lower labials 14.1 scales.

Orbital scales: preorbitals two; suborbitals 2-4, usually

2-3; postorbitals usually two, sometimes 3 or 4. Tem-

Amphib. Reptile Conserv.

January 2017 | Volume 11 | Number 1 | el 32

Murphy et al.

Table 5. A comparison of eight species currently recognized in the genus Polychrus. K = keeled, r = reduced, nd = no data, both

means smooth and keeled in same species. Based on the literature for species not examined in this study.

acutirostris

auduboni

femoralis

gutturosus

jacquelinae

liogaster

marmoratus

peruvianus

vertebrals

111-126

93-112

75-105

198-215

103-125

95-113

56-70

SAB

57-73

62-80

53-73

63-82

131-186

66-95

63-93

52-74

ventral ornamentation

none

both

parietal eye

yes

vertebral crest

yes

gular crest obvious

yes

4 th finger

23-32

23-34

25-36

33-36

29-37

23-35

25-33

4 th toe

19-32

30-43

35-45

42-48

38-47

34-44

32-43

femoral pores

23-24

17-28

30-34

18-42

13-15

15-24

8-19

12-26

poral scales usually in eight vertical rows between post

orbitals and tympanum, separated from parietal scales by

a row of 4-5 enlarged supratemporals. Gulars (counted

between tympani 42-55 (X = 47.0)), rows of gulars sepa¬

rated by numerous micro-scales that vary in size. Dorsal

scales around mid-body X = 78.5 (r = 66-90, SD = 8.71).

Vertebrals and paravertebrals in 10-14 rows; slightly en¬

larged, pentagonal, and keeled; radually become elongate

and ovate laterally often losing the keels; and transition

into sharply keeled, pointed, imbricate ventrals. Scales

similar to the ventrals in size and shape extend onto the

anal plate. Vertebral scales rows X = 108.2 (r = 95-113,

SD = 5.17) between the occiput and posterior edge of

hind legs. Lamellae on fourth huger 25-35 (X = 30.5 SD

= 3.18), lamellae on fourth toe 30-43 (X = 38.69, SD =

3.01). Total pores 8-19 (X = 14.3, SD = 3.31). Scales on

tail pentagonal, heavily keeled, and imbricate. Scales on

limbs keeled, imbricate, and slightly more ovate near the

body, than distally.

Coloration: In preservative most specimens are a uni¬

form brown-tan with black pigmented eye stripes and la¬

bial seams. In life, coloration is highly variable and can

change in less than a minute from green to brown. See

Figures lc,d.

Distribution: We have only documented this species

from Guyana and Suriname, but it may be expected to

occur in French Guyana and northern Brazil.

Natural history: Hoogmoed (1973) described it as

being diurnal, arboreal, and omnivorous; females lay

4-6 eggs in July and August. It is capable of rapid color

change from bright green to brown. Comments on its

natural history in the literature are deeply entangled with

other members of the species group.

Comparisons: Distinguished from Polychrus audu-

boni by numerous multicarinate paravertebral scales (few

or none in P. auduboni ); paravertebrals not much larger

than laterals; a loreal fragmented in two or more parts

(as opposed to one or two in P. auduboni ); a nasal that

is pentagonal and usually contacts the first upper labial

(it usually does not in auduboni ); 8-19 (X = 14.32, SD

= 3.31 ) femoral pores {auduboni has 17-28, X = 21.4,

SD = 2.91); the supranasal often makes contact with the

loreal (in auduboni it rarely does so).

Discussion

We have been unable to examine specimens from the

west side of the Andes and Panama. However, we sus¬

pect the localities are in error, or the lizards have been

misidentihed. It is likely that members of the Polychrus

marmoratus group are restricted to the east side of the

Andes.

Given the high degree of morphological variation

within the P. marmoratus complex, there are likely other

cryptic species present. The P. marmoratus group will

likely also include populations currently considered

Polychrus liogaster. The two species are supposedly sep¬

arated by keeled ventrals in P. marmoratus and smooth or

weakly keeled scales in P. liogaster (Peters and Donoso-

Barrio 1970). Additionally, the latter species has a post¬

ocular stripe that extends onto the neck and body. How¬

ever, preliminary examination of museum specimens

labeled P. liogaster revealed that all had keeled ventrals,

as well as slightly higher upper labial counts than those

reported in the literature.

We cannot rule out the possibility that other taxa re¬

main to be found within Polychrus auduboni in fact, we

think it likely. Ugueto and Rivas (2010) note that the Isla

de Margarita population has a red dewlap and a white

vertebral stripe. A mainland Venezuelan specimen (CM

22797) that we have tentatively included in auduboni,

also has a red dewlap. Only recently have we observed a

red dewlap in a Trinidad Polychrus auduboni. Addition¬

ally, we have observed P. auduboni often have a broad

vertebral stripe that frequently contains white pigment.

This may be normal variation in a species capable of sig¬

nificant changes in coloration, or it may signal a cryptic

species within P. auduboni. Two specimens from Cocol¬

lar, Sucre (FMNH 17792-93) also appear to be distinct

from P. auduboni, with the lowest number of vertebral

scale counts (85) we observed in Polychrus and an ex¬

ceptionally well defined canthal ridge. A single speci¬

men from Bolivar, Venezuela (UMMZ 85232) does not

appear to be P. auduboni or P. marmoratus. It has three

postrostrals, exceptionally small nape scales that are tu-

berculate, and other traits that are uncommon or unob¬

served in our sample. Barrio-Amoros and Ortiz (2015)

Amphib. Reptile Conserv.

January 2017 | Volume 11 | Number 1 | el 32

Cryptic multicolored lizards in the Polychrus marmoratus Group

suggest an undescribed Polychrus exists in eastern Ven¬

ezuela. It is unclear if they are referring to P. auduboni

or another species. Unraveling the natural history of P.

auduboni from P. marmoratus from the extant literature

will be difficult because locality data is often missing.

Further collections with tissues for DNA work are neces¬

sary to fully document the diversity of these lizards.

Our examination of the morphology of three speci¬

mens from Para, Brazil (MCZ 2889, 5549, 92644) agrees

with the molecular results which suggest populations in

Para are neither P. auduboni nor P. marmoratus. These

specimens have a higher vertebral count, more lamellae

on the fourth finger and toe, and a loreal which contacts

upper labials 3-4-5.

Additional taxa are likely present in Brazil’s Atlantic

Forest. Polychrus virescens Schniz (1822: 65) is likely a

valid species. The type locality was not given by Schniz,

but Wied-Neuwied (1825) considered the specimens to

be Polychrus marmoratus when he published and report¬

ed the specimens as being from “Villa Vigoza am Flusse

Peruhype.” Vanzolini (1983: 119) noted this is now Nova

Vicosa, Bahia, Brazil (-17.900872 -39.371644). Myers et

al. (2011: 8) examined the history of this name and found

Wagler (1828: pi 12) contained an illustration labeled

Polychrus virescens and attributed the name to Wied-

Neuwied in Schinz, but suggest it should be attributed

to Schinz alone. They reason that Wied-Neuwied used

Polychrus virescens as a manuscript name only and when

he published referred the specimens to Polychrus mar¬

moratus. We examined photographs of two specimens

collected by Wied-Neuwied. An adult female has about

99 vertebrals which are keeled and some are multicari-

nate. Indicated femoral pores total 22, and the canthal

contacts the loreal. The loreal is in three parts; it has one

large scale between the supranasals (the largest scale on

the snout); three scales separate the first canthals and four

or five separate the second canthals; and the loreal con¬

tacts the supranasal. The semicircle scales are completely

separated by a single row of scales.

Polychrus neovidanus Wagler (1833b: 897) is also

a candidate for being a valid species. Vanzolini (1983)

noted the description was based upon a figure in Seba

(1734-1765, Volume 2, Plate 76, Figure 4) and on Spix’s

P. marmoratus. He states the name should be attached

to Spix’s specimen from Rio de Janerio. We examined

three specimens from Rio de Janerio. They had 103-107

multicarinate paravertebrals and 70-88 scales around

the mid-body. The canthal does not contact the loreal

and the loreal contacts the supranasal. These specimens

also differed from all other Polychrus marmoratus group

members by having five scales between the first pair of

canthals and seven scales between the second pair of can¬

thals.

Hoogmoed (1973) Vanzolini (1983) and Avila-Pires

(1995) suggest the Polychrus marmoratus group is dis¬

junct with a population present in north and western

South America, as well as with a second population in

the Atlantic Forest. However, Kawashita-Ribeiro and

Avila (2008) reported a specimen from Aripuana in Mato

Grosso which narrows the gap, suggesting the P. marmo¬

ratus group may be found throughout the area, and the

disjunct distribution simply reflects a lack of collecting

and knowledge.

Much remains to be learned about the Polychrus mar¬

moratus group and how it reflects the historic landscapes

of the South American continent. It seems likely that ad¬

ditional, unrecognized species remain to be discovered

and we encourage further work on these remarkable and

poorly studied lizards.

Acknowledgments. —Our sincerest thanks go to Har¬

old K. Voris, Alan Resetar, and Kathleen Kelly at the

Field Museum (FMNH); David Kizirian, and Lauren

Vonnahme of the American Museum of Natural History

(AMNH); Mike G. Rutherford at the University of the

West Indies (UWIZM) and Greg Schneider at the Uni¬

versity of Michigan, Museum of Zoology (UMMZ) for

providing logistical support, access to the museum’s col¬

lection, and data collection. For the loan of specimens

and photography services we thank Ted Daeschler and

Ned Gilmore at Academy of Natural Sciences (ANSP);

Steve Rogers at Carnegie Museum (CM); Max Nickerson

and Kenneth Krysko Florida Museum of Natural History

(FLMNH); Jose Rosado at the Museum of Comparative

Zoology; Smithsonian (USNM). We also thank Ana Pru-

dente, Pedro Peloso, Gilson A. Rivas, Gabriel Ugueto,

and Walter E. Schargel for photographs and discussions

about Polychrus. Additionally appreciation goes to Alvin

L. Braswell and Sara E. Murphy for comments on the

manuscript.

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Murphy et al.

Appendix 1. Other specimens examined.

Skeletal and cleared and stained specimens examined: Ecuador: FMNH 42501; Peru: AMNH 71170 (Upper Ucayali), AMNH 71171

(Upper Ucayali); Suriname AMNH 141130, AMNH 148544 male AMNH 141084, UF 56618 (Guyana or Surinam), UF 60914

female, UF 68102. Trinidad: AMNH 148543 female, FMNH 49848, FMNH 49845, FMNH 49840, UF 18395, male, UF 18922.

Venezuela FMNH 17791 (Sucre, Venezuela) male.

Polychrus liogaster ( n = 5). Peru: FMNH 40586, 45499, 59184, 68599, 68600.

Polychrus neovidanus {n = 3). Brazil, Rio de Janeiro (-22.9 -43.23333) MCZ R-3390, R-170011, R-170012. Polychrus virescens

Nova Vicosa, Bahai, Brazil (~I7°54’I5”S 39°22’W) AMNH R105, R1695. Polychrus sp. A (w = 3). Sucre Cocollar (10.147126

-63.808614) FMNH 17791-93.

Polychrus sp. B (w = 3). Brazil, Para (-1.45 -48.48333) MCZ R-2889, R-5549, R-92644.

Polychrus sp. C (« = 4) Bolivia: no specific locality BMNH 61.3.23.1; Buena Vista (-17.459161 -63.659221) FMNH 16163; 21510.

Ecuador: FMNH 53890. Polychrus sp D. (« = 1) Venezuela, Bolivar (1.93965 -64.716248) UMMZ 85232.

r John C. Murphy resides in Sahuarita, Arizona and is a Research Associate at the Field Museum. His research

interests focus on the herpetofauna of Trinidad and Tobago and aquatic snakes.

Richard M. Lehtinen is an Associate Professor, in the Department of Biology, The College of Wooster. His

work on amphibians has taken him to Madagascar and Trinidad and Tobago.

Stevland P. Charles completed his Ph.D. at Howard University, in Washington D.C. in 2016. His research

focused on the distribution, habitat and microhabitat use of the lizards in the genus Gonatodes native to

Trinidad and Tobago, as well as the general effects of biogeography on the diversity of reptiles in Trinidad

and Tobago. His current interests include the ecology, biogeography, behavior, systematics and conservation

biology of Neotropical amphibians and reptiles.

H Danielle Wasserman is a Ph.D. student at the City University of New York, studying trait evolution in avian

and non-avian reptiles. She does collections based research and specializes in comparative morphology of

Tom Anton is president and CEO of the Ecological Consulting Group, EEC. A Chicago-based naturalist

and historian he specializes in astacology (crayfishes), arachnology (specifically scorpions) herpetology and

ichthyology. He is a Field Research Associate at the Field Museum, and an affiliate of the Illinois Natural

History Survey.

Patrick Brennan graduated with a biology degree from The College of Wooster in 2013. He is currently

working toward his Master’s degree at the University of Toledo with a specialization in Bioinformatics.

Amphib. Reptile Conserv.

January 2017 | Volume 11 | Number 1 | el 32

Official journal website:

amphibian-reptile-conservation.org

Amphibian & Reptile Conservation

11(1) [General Section]: 17-24 (e133).

Short Communication

New distribution records and conservation status of

Atelopus seminiferus Cope, 1874: A Critically Endangered

harlequin frog from northern Peru

^uan C. Cusi, 2 Andy C. Barboza, 3 Vance T. Vredenburg, and 4 Rudolf von May

1 Deportamento de Herpetologia, Museo de Historia Natural, UniversidadNacional Mayor de San Marcos, Av. Arena! es 1256, Jesus

Marla, Lima, PERU 2 Division de Herpetologia, CORBIDI (Centro de Ornitologlay Biodiversidad), Santa Rita 117, Huertos de San

Antonio, Surco, Lima, PERU 3 Department of Biology, San Francisco State University, San Francisco, CA 94132-1722, USA ‘^De¬

partment of Ecology and Evolutionary Biology, University of Michigan, 1109 Geddes Ave, Ann Arbor, Michigan 48109-1079, USA

Abstract. —We provide information of the distribution, habitat, and conservation status of the harlequin frog

Atelopus seminiferus, a poorly known species from northern Peru. Multiple individuals of A. seminiferus were

detected inside the Alto Mayo Protected Forest, San Martin region, 87-98 km northwest from the type locality.

Additionally, we used skin swab samples to test for the prevalence of the chytrid fungus Batrachochytrium

dendrobatidis (Be/), a pathogen that has been linked with population declines of harlequin frogs throughout

tropical America. Our findings represent the first record of A. seminiferus inside a natural protected area, and

we recommend an update of the IUCN Red List geographic range map of this species. Though we did not detect

individuals infected by Bd, additional surveys are required to further assess the elevational distribution and

potential for chytrid fungus infection of this Critically Endangered species.

Keywords. Bosque de Proteccion Alto Mayo, Batrachochytrium dendrobatidis , chytrid, UICN Red List, San Martin

Citation: Cusi JC, Barboza AC, Vredenburg VT, von May R. 2017. New distribution records and conservation status of Atelopus seminiferus Cope,

1874: A Critically Endangered harlequin frog from northern Peru. Amphibian & Reptile Conservation 11(1): 17-24 (e133).

NoDerivatives 4.0 International License, which permits unrestricted use for non-commercial and education purposes only, in any medium, provided

the original author and the official and authorized publication sources are recognized and properly credited. The official and authorized publication

credit sources, which will be duly enforced, are as follows: official journal title Amphibian & Reptile Conservation ; official journal website <amphibian-

reptile-conservation.org>.

Received: 12 August 2016; Accepted: 22 November 2016; Published: 17 January 2017

Introduction

The Neotropical bufonid toad genus Atelopus contains

97 described species distributed across different habitats,

from humid forest to paramo grassland in tropical Amer¬

ica (Lotters 1996; Letters et al. 2005). Of these, 69 spe¬

cies (71%) are categorized as Critically Endangered or

Extinct under the IUCN Red List of Threatened Species

(IUCN 2015). The conservation status of at least 30 spe¬

cies is uncertain because they remain undescribed (Co-

loma et al. 2010) or because a comprehensive systematic

revision is required (La Marca et al. 2005; La Marca and

Lotters. 2008; Letters et al. 2011; Flechas et al. 2015).

Peru contains 19 nominal species of Atelopus and three

confirmed candidate new species from the Andes and

Amazon regions (Frost 2016; Rueda-Almonacid et al.

2005). Of these, A. loettersi, A. pulcher, A. spumarius ,

and A. tricolor are primarily distributed in the lowlands

whereas the remaining species are restricted to elevations

above 1,000 m. Montane areas along the eastern slopes of

the Andes are particularly important habitat because they

harbor many species of Atelopus. Although several spe¬

cies have not been seen in decades, recent field surveys

have uncovered rare species such as A. epikeisthos (R.

Santa-Cruz et al., In press). Because of this, it is essential

to continue surveying these montane areas to assess if

amphibian species, some of which have not been seen in

many decades (e.g., Lehr and von May 2004), still exist,

and to evaluate their current conservation status.

Atelopus seminiferus was described by Cope in 1874

based on a single specimen (ANSP 11383) collected by

Prof. Orton from between Balsa Puerto and Moyobamba,

San Martin department, northern Peru (Malnate 1971).

Subsequently, this species was recorded at the Quebra-

Correspondence. Emails: l jcarloscusim@gmail.com\ 4 r~vonmay@gmail.com

Amphib. Reptile Conserv.

January 2017 | Volume 11 | Number 1 | el 33

Cusi et al.

77°40’0"W 77°20'0"W 77°0’0"W 76“40'0"W

77*40"0"W 77°200"W 77*0'0"W 76'40'0"W

Fig. 1. Distribution of Atelopus seminiferus in the Mayo River basin, San Martin, Peru. Yellow polygon corresponds to geographic

range estimated by IUCN. Compare with Fig. 3, which shows proposed new polygon based on results from this study. Map by Juan

C. Cusi.

da Kevlada, close to an Awajun native village in Rioja

Province, San Martin (Schulte et al. 2004; R. Schulte,

pers. comm.). This second record was near an urban area

known as Naranjos, located along the Fernando Belaunde

road (05°44'34.05"S, 77°30'20.87"W, 959 m) and ca. 7.5

km E from the boundary of Alto Mayo Protected Forest

(AMPF). Subsequently, AMPF park rangers reported this

species in 2007 from a site between Nuevo Eden-El Car¬

men villages, Rioja, San Martin (although no geographic

coordinates available). In 2008, biologist Jorge Carrillo

conducted field research focused on harlequin frogs (At¬

elopus spp.) at Sector Serranoyacu inside the AMPF, but

no specimens were recorded in this area (J. Carrillo, pers.

comm.). Recent herpetological surveys at the AMPF in¬

dicate that this reserve has at least 35 species of amphib¬

ians and 10 species of reptiles (J. Cusi et al., unpubl.

data). Prior to this study, which we present here, no re¬

cords of A. seminiferus were available from the AMPF.

Thus, presenting new data on A. seminiferus is relevant

given that recent studies of threatened amphibians from

Peru did not include this Critically Endangered species

(e.g., von May et al. 2008; Jarvis et al. 2015).

In this report, we provide new distributional data for

A. seminiferus and recommend an update to the map of

its known geographic distribution (Fig. 1). We also tested

for the prevalence of the chytrid fungus Batrachochytri-

um dendrobatidis ( Bd ), a pathogen that has been linked

with population declines of harlequin frogs throughout

tropical America (Lampo et al. 2006; Venegas et al.

2008; Flechas et al. 2015). Additionally, given that other

factors such as habitat loss may have caused population

declines in many other amphibian species (Catenazzi and

von May 2014; Tarvin et al. 2014), we noted the type of

habitat used by A. seminiferus in the region. Although

conducting a thorough assessment of habitat change and

disturbance was not a goal of the study, we provide pre¬

liminary information about habitat change and distur¬

bance observed at some localities.

Methods and Materials

We conducted fieldwork at the Alto Mayo Protected For¬

est (AMPF) and Moyobamba, San Martin region, be¬

tween March and December 2014 (Fig. 1). The AMPF

is located along the Cordillera Oriental and is part of the

upper basin of the Mayo River, northern Peru. Addition¬

ally, we surveyed around the city of Moyobamba because

it is one of the type localities of A. seminiferus. A team of

2-3 people carried out Visual Encounter Surveys (Angu¬

lo et al. 2006; Crump and Scott 1994) during both diur¬

nal (10:00-14:00) and nocturnal periods (18:00-00:00).

Total survey effort at AMPF was 240.5 person-hours, and

16.2% of it (39 person-hours) was invested at El Carmen

village, within the Venceremos sector. We collected life

history data including sex, snout-vent length, and weight,

as well as the type of substrate used by each individual.

Additionally, we collected skin swab samples to test

for prevalence of the chytrid fungus Batrachochytrium

Amphib. Reptile Conserv.

January 2017 | Volume 11 | Number 1 | el 33

Status of Atelopus seminiferus

Table 1. Known localities for Atelopus seminiferus in the basin river Mayo, San Martin, Peru. Total survey effort around El Carmen

village (all localities combined) was 39 person-hours.

Locality

Map

locality

number

Latitude

Longitude

Elevation

Date

No. Ind.

Sex

Reference

Kevlada creek, km 393.7, near the

Naranjos village

—

1100

2004

—

Rainer Schulte

(Second record)

Trail between El Carmen and La

Esperanza villages

5°38'48.5" S

77°41'26.0" W

1641

18/01/14

S, 9

Fredi Sangama and

Florencio Leon*

El Arenal forest, near El Carmen

village

5°36'26.6" S

77°41'33.1" W

1279

23/08/14

Authors

Nueva Jordania village

5°34'51.2" S

77°40'50.7" W

1127

Nov. 2011

—

Mathieu Chouteau

El Carmen village: coffee plantations

5°34'57.7" S

77°4E38.2" W

1134

30/06/14

Authors

El Carmen village

5°35’30.4" S

77°42'05.5" W

1243

6/10/13

—

Florencio Leon*

El Carmen village

5°35'30.3" S

77°42'02.1" W

1224

26/08/14

—

Jhonny Ramos and

Elan Cachique*

El Carmen village

5°35'30.4" S

77°42'03.3" W

1222

30/06/14

Authors

El Carmen Creek

5°35'36.2" S

77°42'06.7” W

1277

22/08/14

9 gravid

Authors

El Carmen Creek

5°35'36.7" S

77°42'07.2” W

1267

22/08/14

Authors

El Carmen village

5°35'29.1" S

77°42'03.3" W

1229

1/07/14

Authors

Las Palmas village

5°37'43.6" S

77°43'57.0" W

1902

16/10/13

—

Florencio Leon*

Villa Hermosa village (Boundary

Amazonas-San Martin)

5°32'32.4" S

77°45'49.9" W

1756

17/03/14

—

Marco Ramirez*

* Park rangers’ names (Alto Mayo Protected Forest)

dendrobatidis ( Bd ). We took skin tissue samples using

MW 113-Advantage Bundling sterile cotton swabs over

the abdomen, thighs, and hind limbs of each animal for

a total of 30 strokes (Catenazzi et al. 2013). Skin swab

samples were stored in 1.5 ml tubes. DNA was extracted

from each swab and analyzed following standard pro¬

tocols (Boyle et al. 2004; Hyatt et al. 2007). Addition¬

ally, we used a handheld infrared thermometer (RayTek

MiniTemp MT6) to record the body temperature in vivo

and the temperature of the substrate used by individu¬

als of A. seminiferus. Air temperature and relative hu¬

midity were measured every 15 minutes during 24 hours

with one HOBO U23 Pro v2 data logger (Onset) at one

of the survey sites. Given that, at the time of the study,

only the holotype was available in a museum collection

( Academy of Natural Sciences, Philadelphia [ANSP] in

North America), four specimens were collected (MUSM

33328, 33327, 33662, JCM H-24) and deposited as refer¬

ence material at the Museo de Historia Natural, Univer-

sidad Nacional Mayor de San Marcos (MUSM) in Lima,

Peru. For this purpose, a research and collecting permit

(RJ N° 001 -2014-SERNANP-BPAM-JEF) was obtained

from Peru’s Ministerio del Ambiente.

Results

We found individuals of A. seminiferus at six localities

within of the Alto Mayo Protected Forest: 1) trail be¬

tween El Cannen and La Esperanza villages, 2) El Are-

nal forest near El Carmen village, 3) Nueva Jordania vil¬

lage, 4) El Carmen village, including the actual village,

a creek, and coffee plantations, 5) Las Palmas village,

and 6) Villa Hermosa village (Table 1). All of these lo¬

calities fall outside the IUCN range map polygon (http://

maps.iucnredlist.org/map.html?id=54548, accessed on 8

August 2016; Fig. 1). Most individuals observed in the

field were detected around El Carmen, a small village

inhabited primarily by coffee farmers and surrounded

by coffee plantations. We also obtained photographic re¬

cords from Las Palmas and Villa Hermosa, provided by

park rangers who found A. seminiferus during patrols and

surveillance against illegal logging of the forests. Even

though we did not visit Nueva Jordania, Mathieu Chou¬

teau (pers. comm.) informed us about the record of one

specimen in this locality (Table 1). The complete set of

these new localities was used to calculate the extent of

occurrence. Using ArcGIS, we estimated that the Extent

of Occurrence of A. seminiferus is ca. 2,520 km 2 .

We recorded 14 adult individuals at the AMPF be¬

tween March and December on 2014. Most adult in¬

dividuals were found on leaf litter between 10:00 h to

14:00 h (MUSM 33328, 33327, 33662), and some indi¬

viduals were found at night along the margins of a creek;

these individuals were sitting on top of fem leaves near

the ground. Although we did not hear vocalizations of

A. seminiferus , one mating pair was photographed on 18

January 2014 (Fig. 2A) and one gravid female (field num¬

ber JCM H-24) was found on 22 August 2014. Advertise¬

ment calls and tadpoles of this species remain unknown.

Coloration pattern coincides with the description of Let¬

ters and Schulte (2005): dorsal surface uniformly velvety

black with minute yellow or pinkish cream dots scattered

throughout the dorsum, forelimbs and hindlimbs; some

individuals possess pinkish dots on lower jaw; palmar

and plantar surfaces dark red; belly pink in males and

dark red in females, over a black background in both sex¬

es; throat with pink blotches over a black background;

iris black with yellow ring around pupil (Fig. 2 B-D).

Amphib. Reptile Conserv.

January 2017 | Volume 11 | Number 1 | el 33

Cusi et al.

Fig. 2. (A) A pair of Atelopus seminiferus in amplexus, found between El Carmen and La Esperanza [not collected]. Photo by Fredi

Sangama and Florencio Leon. (B) Dorsal coloration pattern of a female MUSM 33328. (C) Ventral coloration pattern in a male

MUSM 33327. (D) Ventral coloration pattern in a female MUSM JCM EI-24. (D) El Carmen village in Alto Mayo Protected Forest,

Rioja province, San Martin (E). Photos B~E by Juan C. Cusi.

Amphib. Reptile Conserv.

January 2017 | Volume 11 | Number 1 | el 33

Status of Atelopus seminiferus

• Atelopus seminiferus Records

• Naranjos village

Extent of occurrence in this study

Alto Mayo Protected Forest

Fig. 3. Updated distribution map of Atelopus seminiferus. Black dots indicate new localities reported in this study. Light green area

corresponds the estimated Extent of Occurrence (ca. 2,520 km 2 ) based on the new records presented here and the previously known

localities. Numbers correspond to labels in Table 1. Map by Juan C. Cusi.

Our Bd prevalence assays were negative for the pres¬

ence of the chytrid fungus ( Bd) in samples from El Car¬

men (Bd negative, n = 5). Mean body temperature was

19.9 °C, and temperatures of air and substrate were 21.2

°C and 21.3 °C (;n = 4), respectively. We recorded cli¬

matic parameters in one primary forest near El Carmen

(locality number 6 in Fig. 1 and Table 1) on 27-28 June

2014. Mean air temperature during the day was 18.48 ±

2.14 °C and relative humidity was 96.89 ± 2.67%. Mean

air temperature at night was 15.90 ± 1.19 °C and relative

humidity was 98.16 ± 2.41%. Using the new records and

the previously know localities, we created a polygon to

update the geographic distribution map of A. seminiferus

(Fig. 3).

Discussion

Our study documents the existence of populations of A.

seminiferus inside the Alto Mayo Protected Forest, and it

represents the first record of this Critically Endangered

species inside a natural protected area. The new locali¬

ties reported here represent an extension of the geograph¬

ic range of A. seminiferus by ca. 45 km west from the

western boundary of the geographic range recognized

by IUCN (Fig. 1). Specifically, the new localities are ca.

23.1 km northwest from Naranjos and 86.7 km northwest

from Moyobamba (type locality). The record from Villa

Hermosa (Table 1) represents the northernmost locality

known to date for this species. Therefore, we recommend

an update of the IUCN Red List geographic range map

of this species. Concretely, we recommend that the new

polygon generated here (Fig. 3) should be considered in

the next IUCN Red List assessment and replace the cur¬

rently available polygon. As with most harlequin frogs,

A. seminiferus is considered a rare species given that

very few specimens have been observed and collected

in the wild. The IUCN Red List assessment (Schulte et

al. 2004) states that data on population status or abun¬

dance were not available and emphasized that additional

field surveys were needed in the region. In addition to

detecting A. seminiferus at six new localities (i.e., sites

located >1 km apart from each other), our findings sug¬

gest that this species has a fragmented distribution. Using

the IUCN Red List criteria (IUCN 2016), which indicates

that if a species is known from fewer than ten threat-de¬

fined locations and the extent of occurrence is smaller

than 20,000 km 2 , it should be classified as Vulnerable or

Endangered. Atelopus seminiferus is known from eight

localities (Table 1), has an estimated Extent of Occur¬

rence (EOO) of 2,520 km 2 ; additionally, the estimated

EOO and the number of known subpopulations or loca¬

tions has varied over time (with a total of 16 individu¬

als detected in 10 years). Therefore, we suggest that A.

seminiferus might be classified as Vulnerable Blac(i,iii).

Our field surveys indicate that A. seminiferus inhabits

primary montane forests and might tolerate some level

of disturbance given that some individuals were found in

modified forested habitats. In particular, A. seminiferus

Amphib. Reptile Conserv.

January 2017 | Volume 11 | Number 1 | el 33

Cusi et al.

Fig. 3. Dorsal and ventral views of the holotype of Atelopus seminifeus (ANSP 11383), deposited in the herpetological collection at

the Academy of Natural Sciences of Drexel University, Philadelphia. Photos courtesy of Ned Gilmore.

occurs in areas surrounding El Carmen (Fig. 2E), where

native montane forests have been cleared and replaced by

subsistence agricultural plantations (coffee, pineapple,

and banana) and areas used by livestock (cattle, horses,

and mules). However, it would be premature to assume

that populations of A. seminiferus in disturbed areas will

persist on the long term, given that pesticides and fertil¬

izers used in agricultural plantations may have negative

effects on amphibians (Hayes et al. 2002). Furthermore,

habitats used by A. seminiferus appear to be impacted

by the expansion of human settlements associated with

urban development in Moyobamba and Balsapuerto in

recent years. Given the geographic proximity to Moyo-

bamba-Balsapuerto, it is possible that A. seminiferus also

occurs at Cordillera La Escalera Regional Conservation

Area (RCA), in San Martin region, and Cordillera Escal¬

era, in Loreto region; Fig. 1. However, the species has not

been detected in this protected area (Pitman et al. 2014).

The chytrid fungus ( Bd ), a pathogen associated with

massive declines of amphibians around the world (Cat-

enazzi et al. 2011; Lips et al. 2008; Vredenburg et al.

2010), has been assumed to be a possible threat for A.

seminiferus. Although we did not detect ZE/-infected in¬

dividuals, further monitoring of populations of A. semi¬

niferus and larger skin swab sample sizes are needed to

test if the pathogen is affecting any of these populations

more widely. Continuous assessment of Bd prevalence is

essential given that chytriodiomycosis has likely affected

many species of Atelopus (Bonaccorso et al. 2003; La

Marca et al. 2005; Lampo et al. 2006; Lips et al. 2008).

The new voucher specimens collected during this study

will be useful for future morphological studies, especial¬

ly because the only available type material (the holotype,

ANSP 11383) has deteriorated and has broken phalanges

on left hand and right foot (Fig. 4). In summary, our find¬

ings provide valuable insights on the conservation status

of A. seminiferus and an updated map of the known geo¬

graphic range of this species.

Acknowledgments. —We thank the Ministry for For¬

eign Affairs of Finland and Conservation International

Foundation (BioCuencas project) for financing our re¬

search. We thank Dr. Ulla Helimo for her encouragement

and valuable suggestions on our research plan; Jesus

Cordova and Betty Millan for providing access to Museo

de Historia Natural, Universidad Nacional Mayor de San

Marcos (MUSM), Peru. We also thank Rainer Schulte and

Stefan Lotters for providing information and suggestions

for the manuscript. Gustavo Montoya and Ivonne Paico

of the AMPF office kindly helped with collecting per¬

mits (RJ N°001 -2014-SERNANP-BPAM-JEF) and park

rangers (Jhonny Ramos, Florencio Leon, Fredi Sangama,

Marco Ramirez, Elan Cachique) that gently provided re¬

cords of this species of Atelopus , and we recognize their

valuable efforts for the conservation of the forests in Alto

Mayo region. We also thank Rainer Schulte and Mathieu

Chouteau for kindly providing locality information on A.

seminiferus , and Ned Gilmore (Academy of Natural Sci¬

ences of Drexel University, Philadelphia) for kindly pro¬

viding photos of the holotype of A. seminiferus. Thanks

to Mr. Bartolome (local guide) at El Carmen for its as¬

sistance in field and hospitality in his property. We thank

two anonymous reviewers for providing helpful com¬

ments on the manuscript.

Amphib. Reptile Conserv.

January 2017 | Volume 11 | Number 1 | el 33

Status of Atelopus seminiferus

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Juan Carlos Cusi is an associate researcher at the Herpetology Department at the Museum of Natural

History, Universidad Nacional Mayor de San Marcos, Peru (MUSM). His research interests include the

taxonomy and ecology of amphibian and reptiles, and he is currently completing a Master’s program

in zoology at the Universidad Nacional Mayor de San Marcos. His thesis focuses on molecular

phylogenetics and morphology of Neotropical salamanders in the genus Bolitoglossa.

Andy C. Barboza is a Pemvian biologist and scientist associated with the Herpetological Collection

of Centro de Ornitologia y Biodiversidad (CORBIDI), Peru. Her research interests focus on systematic

and evolutionary history of amphibians and reptiles of the Neotropical region.

Vance T. Vredenburg is an Associate Professor in the Department of Biology at San Francisco State

University, research associate and fellow of the California Academy of Sciences, and research associate

at the Museum of Vertebrate Zoology at UC Berkeley. His current research focuses on the impacts of

emerging infectious diseases on amphibians (e.g., chytridiomycosis) and the role of the amphibian skin

microbiome in health and disease. He is also co-founder of AmphibiaWeb (www.AmphibiaWeb.org),

an online conservation resource for amphibians.

Rudolf von May is a postdoctoral research fellow at the Department of Ecology and Evolutionary

Biology at the University of Michigan. His current research seeks to understand how amphibian and

reptile communities are structured across habitats and elevations, taking into account the phylogenetic

relatedness among species present in those communities.

Amphib. Reptile Conserv.

January 2017 | Volume 11 | Number 1 | el 33

Official journal website:

amphibian-reptile-conservation.org

Amphibian & Reptile Conservation

11(1) [General Section]: 25-35 (e134).

Restricted diet in a vulnerable native turtle, Malaclemys

terrapin (Schoepff), on the oceanic islands of Bermuda

13 ’ 4 Mark E. Outerbridge, 2 Ruth O’Riordan, 2 Thomas Quirke and 2 John Davenport

'Department of Environment and Natural Resources, 17 North Shore Road, Hamilton Parish, FL04, BERMUDA 2 School ofBiological, Environmental

and Earth Sciences, University College Cork, Distillery Fields, Cork, IRELAND

Abstract .—Diamondback Terrapins (Malaclemys terrapin) are native to Bermuda, presently

inhabiting only four small brackish-water ponds. Their foraging ecology was investigated using

direct observation, fecal analysis, and necropsy. They do not have as varied a diet as reported from

their North American range. Small gastropods (<3 mm shell height) were found in 66.7% of fecal

samples and made up 97.3% of animal material dry mass, thus dominating their diet. Scavenged

fish and other vertebrates (19% of samples overall), plus terrestrial arthropods (14.3% of samples)

were other common items. Polychaete worms and bivalves each occurred in less than 3% of fecal

samples. Pond sediment was found in 74% of the samples, probably incidentally ingested while

foraging (by oral dredging) for the gastropods. The distribution and abundance of arthropods and

molluscs within the terrapins’ brackish-water environment were assessed in three different habitats;

pond benthos, mangrove swamp, and grass-dominated marsh. These indicated that Bermuda’s

terrapins do not fully exploit the food resources present. On Bermuda M. terrapin is basically a

specialist microphagous molluscivore and mainly forages by deposit-feeding on gastropods living

in soft sediments. This dietary restriction has made them particularly vulnerable to environmental

contamination.

Keywords. Anchialine pond, Diamondback Terrapin, fecal analysis, feeding ecology, aquatic gastropod

Citation: Outerbridge ME, O’Riordan R, Quirke T, Davenport J. 2017. Restricted diet in a vulnerable native turtle, Malaclemys terrapin (Schoepff), on

the oceanic islands of Bermuda. Amphibian & Reptile Conservation 11(1): 25-35 (el 34).

cialNoDerivatives 4.0 International License, which permits unrestricted use for non-commercial and education purposes only, in any medium, provided

the original author and the official and authorized publication sources are recognized and properly credited. The official and authorized publication

credit sources, which will be duly enforced, are as follows: official journal title Amphibian & Reptile Conservation-, official journal website <amphibian-

reptiie-conservation.org>.

Received: 29 March 2016; Accepted: 08 September 2016; Published: 26 January 2017

The Diamondback Terrapin Malaclemys terrapin is one of

two emydid turtle species living in the inland pond envi¬

ronments of the oceanic islands of Bermuda. The other,

Trachemys scripta elegans, is a widely-distributed intro¬

duced freshwater pest (Outerbridge 2008). Diamondback

Terrapins are less abundant than the sliders and have a

greatly restricted local brackish distribution (Davenport

et al. 2005). Native to Bermuda (Davenport et al. 2005;

Parham et al. 2008) they fonn the only known population

outside of the USA.

Diamondback Terrapins have been identified as an

important component of the trophic dynamics of the east

coast USA salt marsh ecosystem (Silliman and Bertness

2002; Davenport 2011) and are carnivorous, feeding

mostly upon a variety of marine molluscs and crustaceans

throughout the North American range (Butler et al. 2006;

Ernst and Lovich 2009). There is, however, a growing

body of evidence to support the hypothesis that this

terrapin species may be a dietary generalist that is oppor¬

tunistic in its foraging habits (Spivey 1998; Petrochic

2009; Butler et al. 2012; Erazmus 2012). Diamondback

Terrapins show resource partitioning, whereby indi¬

viduals with wider heads (the largest females) consume

larger snails and crabs than terrapins possessing smaller

heads (Tucker et al. 1995). Diamondbacks appear to be

predators that use visual cues while foraging, showing

selectivity in the prey that they eat (Davenport et al.

1992; Tucker et al. 1995, 1997; Butler et al. 2012).

Though the diet of Diamondback Terrapins has

been studied in various regions throughout their North

American range, no studies have been conducted on

Bermuda. Analysis of fecal material is a non-destruc¬

tive and non-invasive way of examining dietary pref¬

erence and has been used on several species of small

turtles previously (Demuth and Buhlmann 1997; Lima

et al. 1997), including Diamondback Terrapins (Tucker

Correspondence. 3 mouterbridge@gov.bm. Present address: 4 P.O. Box FL 145, Flatt’s Village, FL BX, BERMUDA

Amphib. Reptile Conserv.

January 2017 | Volume 11 | Number 1 | el 34

Outerbridge et. al

Fig. 1. Benthic survey locations in Mangrove Lake (A)

and South Pond (B). Squares represent detritus sample lo¬

cations along the belt transects; triangles represent the

pond quadrat sample locations; circles represent the quad¬

rat sample locations in the adjacent wetland communi¬

ties. M = Mangrove Lake, T = Trott’s Pond, S = South Pond,

N = North Pond.

et al. 1995; Spivey 1998; Roosenburg et al. 1999; King

2007; Petrochic 2009; Butler et al. 2012; Erazmus

2012; Tulipani 2013; Tulipani and Lipcius 2014). This

method of dietary determination has the added benefit of

allowing multiple samples to be taken from a single indi¬

vidual over time. However, it is limited by the differential

digestibility of the various hard and soft-bodied dietary

components which in turn affects their representation

within the feces.

The primary objective of the current investigation was

to examine the diet and foraging ecology of Bermuda's

terrapin population, with specific aims to assess food

preferences within the land-locked, brackish-water

pond environment, as well as assess the abundance and

distribution of potential food items within the ponds and

adjacent wetland communities. It was envisaged that

detailed knowledge of terrapin diet in Bermuda would

help appropriate conservation and management efforts to

be directed towards protecting the areas in which they

forage.

Materials and Methods

Study Site

Bermudian Diamondback Terrapins occur in four neigh¬

bouring brackish-water ponds: Mangrove Lake, South

Pond, North Pond, and Trott’s Pond (Fig. 1) situated

on a private golf course located at the eastern end of

the islands (32.32858°N, 64.70547°W; WGS 84).

They move between these ponds (Outerbridge 2014).

Mangrove Lake (10 ha area) and Trott’s Pond (3 ha) are

the largest of these and both are simple, shallow, anchia-

line basins fringed by Red Mangrove Trees (Rhizophora

mangle ) with deep benthic deposits of highly organic

sediment (Thomas et al. 1992). Anchialine ponds are

relatively small land-locked brackish bodies of water

with subterranean connections to the sea (Holthuis 1973)

and they show limited tidal influence. North Pond (0.4

ha) and South Pond (0.5 ha) are considerably smaller in

area, shallower in depth, and lack mangrove vegetation.

However, both have small central marshes dominated by

grasses (Cl odium jamaicense and Paspalum vaginatum).

All ponds were incorporated into the golf course as

water hazards during the 1920s, are situated upon a

single square kilometer of land, and are only separated

from each other by, at most, 380 m of land (straight-line

distance between North Pond and Trott’s Pond).

Fecal Analyses

Juvenile, immature, and adult Diamondback Terrapins

were opportunistically captured using a long-handled

dip net from Mangrove Lake, South Pond, North Pond,

and Trott’s Pond from March-September 2010 and

January-October 2011. Maturity status was determined

following Lovich and Gibbons (1990); individuals <91

mm straight plastron length (SPL) were classified as

juveniles, males as sexually mature if SPL = 91-137 mm,

females as sexually mature if SPL >138 mm. Females

of SPL 91-137 mm were regarded as immature. After

capture, each individual was kept outside in the shade

for 48 h in covered, plastic storage bins (55 cm long x

45 cm wide x 30 cm deep). All fecal material collected

in the 48 h period was strained through a one mm mesh¬

sized sieve, oven dried at 80 °C for 48 hours, and stored

in a sealed glass vial for subsequent identification. Fecal

samples were also collected from neonate terrapins (i.e.,

individuals that were less than one year old) that were

followed as part of a radio-telemetry study (Outerbridge

2014). At the end of the tracking period, each individual

was placed in a 500 ml plastic bowl containing enough

freshwater to cover the carapace and held in a room with

an ambient temperature of 30 °C for 48 hours. All fecal

material collected in this period was strained through

47 mm filter paper to retain finer particles from smaller

prey items consumed, allowed to air dry for 48 hours,

Amphib. Reptile Conserv.

January 2017 | Volume 11 | Number 1 | el 34

Restricted diet in Malaclemys terrapin

and stored in a sealed glass vial. All terrapins captured

during the fecal analysis investigation were released at

their original capture location.

Each fecal sample was examined at magnifications

between 10* and 25x using a stereoscopic micro¬

scope with an ocular scale. Food items were identified

to the lowest possible taxonomic level, and weighed

to the nearest 0.000lg. The shells of gastropods, when

encountered whole, were counted and shell height (SH;

maximum measurement along the central axis) was

measured to the nearest 1.0 mm (note that some fecal

samples only contained broken shells, the size of which

could not be estimated). Quantification of dietary items

was accomplished by determining the percentage dry

mass of each item relative to the total dry mass of each

sample. The relative frequency of occurrence of each

dietary item was determined by calculating the percentage

of turtles containing a given food type in relation to the

total number of turtles examined.

Benthic Biotic Surveys within the Terrapins’

Wetland Environment

Assessments of mollusc and crustacean abundance and

distribution within the ponds and adjacent wetland envi¬

ronments were conducted to determine prey availability

for Bermuda’s Diamondback Terrapins. These assess¬

ments were accomplished by perfonning a series of

benthic transects within three different habitats utilized

by all size and age classes of Bermuda’s Diamondback

Terrapins; the sediment at the bottom of Mangrove Lake

and South Pond, the Red Mangrove swamp community

that surrounds Mangrove Lake, and the Saw-grass

(Cl odium jamaicense) marsh in the center of South Pond.

Pond Benthic Surveys

Two belt transect surveys of benthic biota were

performed in Mangrove Lake and one belt transect

survey was carried out in South Pond in July 2011. The

Mangrove Lake transects were straight-line and followed

an east-west direction (Transect 1) and a south-north

direction (Transect 2), whereas the survey in South Pond

was circular (Transect 3). Ten locations were haphaz¬

ardly sampled along the path of each transect (Figs. 1A,

IB). The GPS coordinates were recorded at each location

together with a brief description of the benthic charac¬

teristics. Collection consisted of sweeping a dip net with

one mm mesh and a square opening of 25 x 25 cm for

a distance of one m and a depth of approximately 2.5

cm at the surface of the sediment (thereby sampling a

linear area of 0.25 m 2 at each location). The collected

sediment was passed through a one mm mesh sieve at the

surface of the pond and the material that remained was

transferred into a one litre container. In addition to the

belt transects, four replicate 25 x 25 cm quadrat surveys

(A-D, Fig. 1A) were performed at random in sand, rock,

and gravel areas of the margins of Mangrove Lake. The

area defined by each quadrat was dredged to a depth of

2.5 cm and the contents transferred into a bucket and

sorted by hand.

Mangrove Swamp Surveys

Sixteen replicate quadrat surveys were performed within

the mangrove swamp that borders Mangrove Lake (Ql-

Q16, Fig. 1A). The sites were haphazardly chosen, using

an aerial map, at various locations around the periphery

of the pond. Upon arrival in the field, a 25 x 25 cm

quadrat was randomly placed upon the leaf litter imme¬

diately land-ward of the water-line. The area defined

by each quadrat was dug to a depth of 2.5 cm and the

contents transferred to a 3.8 liter sealable plastic bag.

The contents of each bag were gently sifted in the labora¬

tory using running water and a sieve with five mm mesh

stacked on top of a one mm mesh-sized sieve.

Saw-grass Marsh Surveys

Four replicate quadrat surveys were perfonned within the

saw-grass marsh at the center of South Pond (Q1-Q4, Fig.

IB). These sites were also haphazardly chosen using an

aerial map. Upon arrival in the field, a 25 x 25 cm sample

of saw-grass and turf was cut, to a depth of 2.5 cm, from

the marsh at each of the four sites. The saw-grass blocks

were transferred to separate 19 L buckets and taken to

the laboratory for examination. Each sample was placed

in a plastic bin (60 cm long x 40 cm wide x 14 cm deep),

carefully broken apart and gently sifted in the labora¬

tory using running water and a five mm sieve stacked on

top of a one mm sieve. Shoot bundles were counted to

determine saw-grass density.

All biological specimens from the belt transect and

quadrat surveys were kept for subsequent identifica¬

tion in the laboratory, but only living specimens were

counted and measured (i.e., empty gastropod shells were

discarded). Live gastropods were counted, measured

(total shell height mm), and frozen for eco-toxicolog-

ical analyses (Outerbridge et al. 2016). All other living

biological specimens were returned to their original

locations and released after identification. All transect

and quadrat survey results were standardized as values

nr 2 as depth was constant throughout.

Results

Fecal Analyses

A total of 54 Diamondback Terrapins were netted between

March and September 2010 (n = 21) and January and

October 2011 (n = 33), of which 42 (77.8%) produced

fecal samples during the 48-hour confinement period (30

adults, four immature females, three juveniles of unde¬

termined gender, and five neonates). Of the 54 terrapins,

30 were captured from South Pond (of which 23 (76.7%)

produced fecal samples), 20 from Mangrove Lake (of

Amphib. Reptile Conserv.

January 2017 | Volume 11 | Number 1 | el 34

Outerbridge et. al

Table 1. Malaclemys terrapin dietary items obtained from 42 fecal samples (from females, males Juveniles and neonates combined)

collected from inhabitants of four brackish ponds in Bermuda. Symbols: n = number of samples containing a given food type; % =

percentage of samples containing a given food type in relation to the total number of samples. Presence (+) and absence (-) of dietary

items’ data for the various gender/age categories are given separately.

Dietary Item

n (%)

Adult

females

Adult males

Juveniles

Neonates

Plants (grass, seeds, algae)

14 (33.3%)

Gastropoda

28 (66.7%)

Heleobops bermndensis

24(57.1%)

Melanoides tuberculata

15 (35.7%)

Melampus coffeus

2 (4.8%)

Insecta

6 (14.3%)

Polychaeta

Arenicola cristata

1 (2.4%)

Bivalvia

Isognomon alatus

1 (2.4%)

Cmstacea

Armadillidium vulgare

1 (2.4%)

Osteichthyes

Fundulus bermudae

5(11.9%)

Amphibia/Reptilia

Rhinella (syn Bufo) marinus

2 (4.8%)

Malaclemys terrapin

1 (2.4%)

Sediment

31 (73.8%)

Trash (cigarette filter)

1 (2.4%)

which 15 [75.0%] produced fecal samples), three from

North Pond (all of which produced fecal samples), and

one was captured from Trott’s Pond (which also produced

a fecal sample). Note that the small Bermudian terrapin

population meant that some terrapins were netted more

than once in this exercise; three females, one male, and

one neonate were captured twice. One of the females was

captured three times.

Of the 42 terrapins that produced fecal matter, 28

(66.7%) were classified as female (24 mature, four

immature) ranging from 126-196 mm straight carapace

length (SCL) (mean 172, SD 17.9) and six (14.3%) were

classified as male (all mature) ranging from 114-134 mm

SCL (mean 122, SD 8). Three (7.1%) were classified as

juveniles (97-107 mm SCL, mean 102, SD 5), and five

(11.9%) were classified as neonates (31-35 mm SCL,

mean 33.7, SD 1.6).

Sediment occurred in 73.8% of the fecal samples,

gastropods in 66.7%, plant material in 33.3%, fish and

other vertebrate bones in 19%, terrestrial arthropods

in 14.3%, polychaete worms, bivalves, terrestrial crus¬

taceans, and trash (each 2.4% respectively) [Table 1],

The gastropods comprised three species: an endemic

hydrobiid snail Heleobops bermndensis, the Red-rimmed

Melania (Melanoides tubercnlata), and the Coffee Bean

Snail ( Melampus coffeus). Heleobops bermndensis

occurred in 57.1% of all fecal samples and was obtained

from terrapins captured in South Pond, Mangrove Lake,

and North Pond. Melanoides tubercnlata occurred in

35.7% of the fecal samples but was only obtained from

terrapins captured in South Pond, while M. coffeus only

occurred in 4.8% of the fecal samples and was obtained

from terrapins captured in Mangrove Lake.

The plant materials consisted mostly of mown grass

fragments, saw-grass seeds, and green algae. None of

the plant material appeared to have been digested and

may have been ingested incidentally with animal prey

(cf. Erazmus 2012). The terrestrial arthropods consisted

of honey bees (Apis mellifera ) (4.8% of the samples),

small beetles (Berosns infnscatns ), an isopod (Armadil-

lidium vulgare ), a millipede (Julus sp.), a big-headed ant

(Pheidole megacephala), and an unidentified caterpillar

(each represented in 2.4% of the samples). Vertebrate

animal bones came from aquatic species and included

fish from the family Cyprinodontidae—which occurred

in 11.9% of the samples; an amphibian (the toad Rhinella

[syn Bnfo\ marinus )—which occurred in 4.8% of the

samples; and another terrapin ( Malaclemys terrapin ),

probably scavenged—which occurred in 2.4% of the

samples. The fecal samples containing arthropods and

fish and vertebrate animal bones were acquired from

terrapins captured in a variety of ponds. The samples that

contained the burrowing polychaete worm (Arenicofa

cristata) and shell fragments from the Flat Mangrove

Oyster (Isognomon alatns) all came from terrapins

captured in Mangrove Lake. The single sample that

contained a cigarette filter was obtained from a terrapin

captured in South Pond. It is worth noting that most of

the samples (n = 33 or 78.6%) that contained sediment

also contained other dietary items, whereas nine samples

Amphib. Reptile Conserv.

January 2017 | Volume 11 | Number 1 | el 34

Restricted diet in Malaclemys terrapin

Table 2. Dry mass summary of all animal food items obtained from 33 fecal samples of Diamondback Terrapins collected from four

sites combined (South Pond, Mangrove Lake, Trott’s Pond, and North Pond).

Melanoides

Heleobops

Melampus

Isognomon

Insect

Fundiiliis bone

Rhinella bone

Malaclemys bone

Polychaete

TOTAL

dry mass (g)

Proportion of

37.08

14.85

2.22

0.0595

0.117

0.139

1.17

0.0003

0.0153

55.65

total dry mass

66.6%

26.7%

3.99%

0.11%

0.21%

0.25%

2.1%

0.0005%

0.027%

100%

(21.4%) comprised only sediment. Female, male, and

juvenile terrapins were all found to have ingested

sediment, but none of the neonate terrapins produced

feces that contained sediment.

Table 2 summarises the dry mass of all animal food

items obtained from 33 terrapin fecal samples. It is

evident that the three gastropod species made up most

(97.3% of dry mass) of the collected material. Table 3

summarises their numbers and sizes. First, it can be seen

that the terrapins ate very large numbers of M. tubercu-

lata and H. bermudensis, and second that the gastropods

were predominantly small in size (M tuberculata mean

SH 3.2 mm; H. bermudensis mean SH 1.7 mm). Thirdly,

these data show that H. bermudensis had been consumed

by all age classes (i.e., adults, juveniles, and neonates),

whereas M. tuberculata had been consumed by adults

and juveniles and the larger M. coffeus were found only

in female adult samples. Most H. bermudensis measured

<2 mm SH and M. tuberculata measured <3 mm SH. The

majority (ca. 70%) of the M. coffeus snails ingested by

the females measured 9-10 mm SH.

Further statistical analysis is compromised because a)

many gastropod shells were broken, so unmeasureable,

and b) there were not matched numbers of female, male,

juvenile and neonate terrapins. However, it appears from

Table 3 that adult females consumed rather larger prey

than adult males. This is consistent with earlier studies

of this markedly sexually-dimorphic species (Tucker et

al. 1995).

Finally, it should be noted that the diet of neonate

terrapins was extremely restricted (Table 1). Four out of

five samples only contained remains of the gastropod H.

bermudensis. The last sample also contained this species

together with a little insect material. None of the neonate

fecal samples contained sediment, presumably reflecting

their terrestrial lifestyle.

Benthic Biotic Surveys within the Terrapins’

Wetland Environments

Pond Benthic Surveys

Only two species of aquatic gastropods were encountered

during the Mangrove Lake surveys; the False Horn Shell

{Batillaria minima) and H. bermudensis. Two species

of aquatic gastropods were also encountered during the

South Pond surveys; H. bermudensis and M. tuberculata.

Table 4 summarises the gastropod survey data for all

three transects in both ponds. Gastropod abundance in

Mangrove Lake varied along Transects 1 and 2. Batillaria

minima and H. bermudensis were encountered in rela¬

tively low numbers at locations that comprised sediment

only (B. minima range 0-28 snails nr 2 , mean 3.0, SD 7.2,

n = 52; H. bermudensis range 0-192 snails nr 2 , mean

27.0, SD 47.7, n = 424); however abundance increased

significantly at locations where widgeon grass ( Ruppia

maritima) was found {B. minima range 0-56 snails nr 2 ,

mean 33.0, SD 27.8, n = 132; H. bermudensis range

252-772 snails nr 2 , mean 474, SD 221.5, n = 1,896).

Shell height of H. bermudensis along both transects

ranged from 1^1 mm (mean 1.7 mm, SD 0.5, n = 580);

B. minima ranged from 6.5-11 mm (mean 8.9, SD 1.0,

n = 46). Pooling the data for each of the two separate

transects in Mangrove Lake shows that H. bermudensis

was more abundant than B. minima along the central axes

of the pond.

All of the sample locations along Transect 3 in South

Pond comprised sediment and both snail species were

encountered in low numbers ( H. bermudensis 0-4 snails

nr 2 , mean 0.4, SD 1.3, n = 4; and M. tuberculata 4-20

snails nr 2 , mean 13.2, SD 5.7, n = 132). Shell heights

of H. bermudensis encountered along Transect 3 all

measured one mm and the shell heights of M. tubercu-

Table 3. Pooled summaries of the total numbers ( n ) and sizes (shell height, SH) for whole Melanoides tuberculata, Heleobops

bermudensis, and Melampus coffeus obtained from the 28 Diamondback Terrapin fecal samples that contained gastropods.

Terrapin

samples

Melanoides tuberculata

Size Range Mean

(SH; mm) (SH; mm)

(mm)

Heleobops bermudensis

Size Range Mean

(SH; mm) (SH; mm)

(mm)

Melampus coffeus

Size Range Mean

(SH; mm) (SH; mm)

(mm)

All pooled

2224

1-18

3.2

2.1

1910

1-5

1.7

0.7

7-11

9.4

1.1

Female pooled

2112

1-18

3.3

2.1

1643

1-5

1.8

0.8

7-11

9.4

1.1

Male pooled

1-7

2.1

1.0

150

1-3

1.5

0.6

Juvenile pooled

1-3

0.6

1-3

1.2

0.4

Neonate pooled

1-2

1.2

0.4

Amphib. Reptile Conserv.

January 2017 | Volume 11 | Number 1 | el 34

Outerbridge et. al

Table 4. Summary of gastropod abundance (number of snails

0.25 nr 2 ) at each sampling site along Transects 1 and 2 in

Mangrove Lake and Transect 3 in South Pond.

Site No.

Description

Batillaria

Heleobops

Melanoides

1-1

sediment

1-2

sediment

1-3

sediment

1-4

sediment

1-5

sediment

1-6

sediment

1-7

widgeon

123

grass

1-8

widgeon

grass

1-9

sediment

1-10

sediment

2-1

sediment

2-2

sediment

2-3

sediment

2-4

sediment

2-5

sediment

2-6

widgeon

193

grass

2-7

widgeon

grass

2-8

sediment

2-9

sediment

2-10

leaf litter

3-1

sediment

3-2

sediment

3-3

sediment

3-4

sediment

3-5

sediment

3-6

sediment

3-7

sediment

3-8

sediment

3-9

sediment

3-10

sediment

lata ranged from 1-11 mm (mean 3.1 mm, SD 2.0). The

pooled data for Transect 3 shows that M. tuberculata was

more abundant than H. bermudensis within the sediment

of South Pond. Furthermore, H. bermudensis appeared to

be more abundant within Mangrove Lake than in South

Pond.

Further analyses of gastropod abundances along

the three transects were attempted. The data were non¬

normal and variance was heterogenous whether the data

were raw or square root transformed. The requirements

of parametric statistics were therefore violated. Accord¬

ingly, a non-parametric approach was adopted. First, the

abundances of B. minima were investigated. A Kruskall-

Wallis test across the three transects showed that there

were significant differences amongst the numbers of this

species (Chi-Square = 7.885, df = 2 ,p = 0.019). Post-hoc

tests using Mann-Whitney U tests were then conducted

Table 5. Summary of gastropod (Bat ill aria minima ) and

crustacean (Alpheus armillatus ) total abundance (individ. nr 2 )

at each quadrat site (/? = 4) within Mangrove Lake.

Site No.

Description

Batillaria

minima

Alpheus

armillatus

Sand and gravel

2128

Rocks

2000

Rocks

3504

Rocks

6752

to compare Transect 1 with Transect 2, Transect 1 with

Transect 3 and finally Transect 2 with Transect 3. This is

not an ideal approach as there is an attendant risk of Type

1 error (i.e., incorrect rejection of a null hypothesis),

but no better alternative is available. These post-hoc

tests indicated that there were no significant differences

in numbers of B. minima between Transects 1 and 2

(both from Mangrove Lake) (Mann-Whitney U = 36.50,

Wilcoxon W = 91.50, Z = -1.153, p = 0.315). There

were no significant differences in numbers of B. minima

between Transects 1 and 3 (Mann-Whitney U = 33.00,

Wilcoxon W = 88.00, Z = -1.302 ,p = 0.218), but there

were significant differences between Transects 2 and 3

(Mann-Whitney U = 12.00, Wilcoxon W = 67.00, Z =

-2.954, p = 0.003).

Second, the same approach was adopted for the

abundances of H. bermudensis. A Kruskall-Wallis test

across the three transects showed that there were signifi¬

cant differences amongst the abundances of this species

(Chi-Square = 12.76, df = 2 ,p = 0.002). Post-hoc Mann-

Whitney tests showed that abundances of H. bermudensis

did not differ between Transects 1 and 2 (Mann-Whitney

U = 39.00, Wilcoxon W = 94.00, Z = -2.954, p = 0.436),

but did differ significantly between Transects 1 and 3

(Mann-Whitney U = 11.00, Wilcoxon W = 66.00, Z =

-3.229 ,p = 0.002) and between Transects 2 and 3 (Mann-

Whitney U = 12.50, Wilcoxon W = 67.50, Z = -3.117, p

= 0.003). Overall these tests indicate that there is strong

(but not conclusive) support for the abundance trends

identified above.

Table 5 shows the results of the four replicate quadrat

surveys that were performed in the sandy, rocky, and

gravelly marginal areas of Mangrove Lake. Only one

species of gastropod (B. minima ) and one species of crus¬

tacean (the Snapping Shrimp, Alpheus armillatus ) were

encountered. The snails were found most often attached

to the rocky substrate, whereas the shrimp were found

either buried within the gravel or hidden beneath rocks.

The density of B. minima ranged from 2,000-6,752 snails

nr 2 (mean 3,596, SD 2,211.4) and their sizes ranged from

3.5-10 mm SH (mean 6.4); the density of A. armillatus

ranged from 0—48 shrimp nr 2 (mean 20, SD 24) and their

total lengths (TL) ranged from 10-19 mm (mean 15.6).

These data suggest that the density of B. minima surveyed

upon the rocky shoreline habitat (mean 3,596 snails nr 2 )

was nearly 400 times more than the mean density of live

Amphib. Reptile Conserv.

January 2017 | Volume 11 | Number 1 | el 34

Restricted diet in Malaclemys terrapin

Table 6. Biotic summary of the quadrat surveys (n = 16) performed within the mangrove swamp around Mangrove Lake. M.c.

= Melampus coffeus, M.m. = Myosetella myositis , L.c. = Laemodonta cunensis , M.o. = Microtralia occidentalism Pm. = Pedipes

mirabilis , Amp. = Amphipod spp., L.b. = Ligia baudiniana, A.e. = Armadilloniscus ellipticus , Av. = Armadillidium vulgare , A/. =

Bersos infuscatus, Lep. = Lepidopteran larvae, Jul. =Julus sp., Am. = Anisolabis maritima , Fun. = Fundulus eggs, Ara. = Arachnid

spp., P = Earthworm sp.

M.c.

Gastropods

M.m. L.c.

M.o.

P.m.

Amp.

Crustaceans

L.b. A.e.

A.v.

B.i.

Insects

Lep. Jul.

A.m.

Fish

Fun.

Other

Ara. P

Mean density

(indiv. nr 2 )

282

371

197

1 17

313

9 9

Size range

(mm)

2-15

1-6

1-3

6-7

2-3

Mean size

(mm)

8.8

2.8

1.8

6.3

2.3

”

” ”

”

” ”

3.2

1.2

0.8

0.6

5. minima found upon the sediment along the central

axes of Mangrove Lake (9.2 snails nr 2 ).

Mangrove Swamp Surveys

Table 6 summarises the various aquatic and terrestrial

species discovered during the quadrat surveys (;n =

16) performed within this environment. A total of five

gastropod species were encountered; all were found

within the detritus of the intertidal zone and some indi¬

viduals of M. coffeus were also encountered attached to

Red Mangrove prop roots, usually in clusters, immedi¬

ately above the water line of the pond. Melampus coffeus

were most frequently encountered. Density for this

species ranged from 0-1,168 snails nr 2 (mean 282, SD

399.3, n = 4,512), and shell height ranged from 2-15 mm

SH (mean 8.8, SD 3.2, n = 4,512). Myosetella myosotis

was the second most frequently encountered gastropod,

but only at one of the 16 locations. Sizes ranged from

1-6 mm SH (mean 2.8, SD 1.2, n = 848). Laemodonta

cubensis was encountered in densities of 80 snails nr 2

and all occurred in one location. Sizes ranged from 1-3

mm SH (mean 1.8, SD 0.8). Microtralia occidentalis

and Pedipes mirabilis were infrequently encountered.

Sizes of the fonner ranged from 6-7 mm SH (mean 6.3,

SD 0.6, n = 48), and the latter ranged from 2-3 mm SH

(mean 2.3, SD 0.6, n = 48).

In addition to the gastropods mentioned above,

four species of crustaceans were encountered among

the detritus (Table 5). The amphipods were the most

abundant crustaceans encountered, being found in 81.3%

of the quadrat locations. Densities ranged from 0-2,272

nr 2 (mean 371, SD 656.8, n = 5,936). The isopod Arma¬

dilloniscus ellipticus was the second most frequently

encountered crustacean, with densities of 0-1,008 nr 2

(mean 197, SD 311.5, n = 3,152). Ligia baudiniana and

A. vulgare were not commonly encountered.

Eggs (approx, two mm diameter) from the endemic

Bennuda Killifish (Fundulus bermudae) were encoun¬

tered in 25% of the quadrat surveys. Abundance varied

from 0-3,824 eggs nr 2 (mean 313, SD 958.5, n = 5,008).

The eggs were usually found hidden within the leaf

detritus, but also attached to the Red Mangrove prop

roots at the high water mark. A variety of primarily

terrestrial organisms were occasionally encountered

in low densities within the 16 quadrat locations; these

included millipedes, earwigs, small spiders, earthworms,

small beetles, and a lepidopteran larva.

Saw-grass Marsh Surveys

Table 7 summarizes the aquatic and terrestrial species

discovered during the quadrat surveys performed within

this environment. Only one species of gastropod was

found during the quadrat surveys (H. bermudensis).

Densities ranged from 176-272 snails nr 2 (mean 208, SD

43.3, n = 832), and shell heights ranged from 1^1 mm

SH (mean 2.3, SD 0.7). Terrestrial organisms were infre¬

quently encountered within the quadrats and consisted of

millipedes and small spiders. The number of saw-grass

shoot bundles ranged from 16—48 nr 2 .

Discussion

The anchialine ponds inhabited by Bermudian Diamond-

back Terrapins are unusual habitats for the species. In the

USA terrapins live predominantly in Spartina salt marshes

and in the Everglades mangrove swamps of west Florida.

The latter environments feature substantial allochthonous

inputs from neighbouring marine and freshwater habitats

as well as abundant autochthonous energy sources, so are

amongst the most productive natural environments in the

world, supporting diverse plant and animal communities

(Schmalzer 1995; Whitney et al. 2004).

In contrast, energy sources of anchialine pools are

Table 7. Biotic summary of the quadrat surveys (n = 4)

performed within the saw-grass marsh habitat at the center of

South Pond. Note: results standardized to values nr 2 .

Site

No.

No. of

grass shoot

bundles

Heleobops

bermudensis

Millipedes

Spiders

176

272

192

Amphib. Reptile Conserv.

January 2017 | Volume 11 | Number 1 | el 34

Outerbridge et. al

largely autochthonous. The Bennudian anchialine pools

inhabited by terrapins proved to have limited faunal

diversity. Over most of the area of Mangrove Lake (the

largest pond), only two species of benthic gastropod snails

were found; H. bermudensis and B. minima. Similarly,

two species of aquatic gastropods were encountered

during the benthic South Pond surveys; H. bermudensis

and M. tuberculata. All three species are operculate

deposit-feeders; B. minima and H. bermudensis are

native, while M. tuberculata is primarily a freshwater

(though salt-tolerant) species that is native to tropical

and sub-tropical regions of southern Asia and northern

Africa (Clench 1969), but widely-introduced to various

regions via the aquarium trade. Heleobops bermudensis

is a small endemic hydrobiid snail, limited to brackish-

water ponds in Bermuda (see Pilsbry in Vanatta 1911),

while B. minima is found also on local mudflats (Sterrer

1986).

The results of the quadrat and transect surveys

revealed that the sediment surface in Mangrove Lake and

South Pond generally showed relatively low densities

of gastropods; however B. minima and H. bermudensis

were both found to exist in higher densities in localized

patches throughout Mangrove Lake. Batillaria minima

was most often associated with sand, rock, and gravel

substrate, reaching densities ca. 6,750 snails nr 2 , whereas

H. bermudensis was more commonly found within beds

of widgeon grass in densities up to 772 snails nr 2 . Benthic

mapping of Mangrove Lake was not performed, but visual

assessments of the pond in 2011 suggested that both the

gravel/rock and widgeon grass environments comprised

a very small proportion (< 5%) of the total pond area.

Taken with the fecal sample results, it would appear that

juvenile and adult terrapins on Bermuda rely heavily on

benthic dredging of small gastropods (Outerbridge and

Davenport 2013) from the large areas of pool bottoms,

presumably because this unselective feeding behavior

provides them with plenty of food.

Gastropods were more abundant and diverse within

the mangrove and saw-grass marsh environments. Five

species of gastropods (all pulmonates of the Family

Melampidae) were encountered during the quadrat

surveys within the detritus of the mangrove swamp

intertidal zone around Mangrove Lake. Melampus

coffeus grow to 20 mm SH, but the other species rarely

exceed eight mm SH (Sterrer 1986). Thomas et al.

(1992) and Herjanto (1994) reported that M. coffeus was

frequently encountered upon the detritus and prop roots

of mangrove trees in Mangrove Lake and Troths Pond.

The present investigation showed that gastropods within

Bermuda’s saw-grass marsh and mangrove swamp

environments can reach densities of up to 1,168 snails

nr 2 (M. coffeus). However, it is evident that the adult and

juvenile terrapins rarely, if ever, use this resource and are

essentially aquatic foragers.

Crustaceans were rarely encountered within the

aquatic environment of Mangrove Lake; only one species

(Alpheus armillatus) was found in the rocky marginal

habitats; no crustaceans were encountered within South

Pond. However, crustaceans (mostly small amphipods

and isopods) were frequently encountered (87.5%) in

the quadrat surveys performed in the mangrove swamp

surrounding Mangrove Lake. The Mangrove Crab

(Goniopsis cruentata) was not encountered during

the present study though it was reported to inhabit the

intertidal zone of Mangrove Lake and Trott’s Pond two

decades ago (Thomas et al. 1992). Small numbers of

terrestrial invertebrates were also found in the vegetated

areas around the pools.

Some potential food organisms had been surveyed

before this study. The Flat Mangrove Oyster (Isognomon

alatus) grows in clumps on the submerged prop roots of

red mangrove trees in Mangrove Lake and Trott’s Pond

and has been reported to reach densities of 250 oysters

roof 1 or about 2,700 oysters nr 2 of pond (Thomas and

Dangeubun 1994); the Bermudian terrapins hardly use this

resource. Fish have also been investigated; the endemic

killifish ( Fundulus bermudae) occurs in Mangrove Lake

(estimated population about 11,000) and Trott’s Pond

(about 8,000) (Outerbridge et al. 2007). Killifish in

Mangrove Lake are benthopelagic and are omnivorous

opportunistic feeders. They are swift swimmers that form

loose schools of similarly-sized fish (Rand 1981) and are

probably difficult for terrapins to catch.

Overall, it appeared that the ponds themselves had

low faunal diversities, but abundant supplies of small

deposit-feeding gastropod snails; the neighbouring

vegetated areas had rather higher diversities, but

gastropods were again dominant. Given the small size

of the terrapin population (ca. 100 individuals > 81 mm

straight carapace length, see Outerbridge et al., In Press),

it was evident that plenty of food was available to them.

The benthic sediment in all of the terrapin ponds is

gelatinous and extremely flocculent which allows the

terrapins to both easily move through it and process it,

apparently allowing them to consume M. tuberculata ,

the most frequently encountered gastropod within the

pond’s sediment (Outerbridge and Davenport 2013).

In support of this hypothesis, fecal analyses from this

study confirm that Bermuda’s terrapins consume very

high numbers of small (<2 mm) M. tuberculata and H.

bermudensis together with large quantities of sediment.

The sediment is believed to have been incidentally rather

than deliberately ingested (as is probably the case for

plant material too). It is evident that small gastropods

form almost all of the adult and juvenile terrapins’ animal

diet (97.3% of dry mass).

All of the few insects recorded from fecal material

were probably consumed after falling into the ponds,

rather than having been ingested in the terrestrial

environment (with the exception of those consumed by

neonate terrapins which are residents of the intertidal

mangrove and grass-dominated marsh environments

Amphib. Reptile Conserv.

January 2017 | Volume 11 | Number 1 | el 34

Restricted diet in Malaclemys terrapin

adjacent to the ponds; Outerbridge 2014). The fish, toad,

and terrapin bones discovered in some fecal samples

indicate that Bermuda’s terrapins also scavenge on animal

remains. Carcasses of these species are periodically

observed floating at the surface of the study ponds and

it is likely that they are opportunistically ingested when

encountered. Scavenging has been reported for other

diamondback terrapin populations in the USA (Ehret and

Werner 2004; Petrochic 2009; Butler et al. 2012).

Plant material (mown grass fragments, saw-grass

seeds, algae) was found in small quantities in a third

of fecal samples. All appear to have been incidentally

ingested. Mown grass fragments presumably reflect

the golf course management of the terrapins’ habitat.

The presence of seeds in feces has been reported before

(Tulipani 2013; Tulipani and Lipcius 2014) from

terrapins foraging in salt marshes in Virginia; in that case

the turtles were shown to be significant in the dispersal of

Eelgrass ( Zostera marina) seeds.

It is interesting to note that Bermuda’s Diamondback

Terrapins apparently did not ingest or rarely ate some

items common in their environment. There was no

evidence that they ever ate the Snapping Shrimp Alpheus

armillatus, though substantial numbers were available in

rocky areas of the shoreline. They also ate few of the

Mangrove Oysters {Isognomon alatns ) despite the latter’s

high population densities on mangrove roots. There

was little evidence of foraging amongst the mangrove

vegetation; most of the pulmonate gastropod species

(M coffeus does not appear to be an important dietary

food item for Bermuda’s terrapins, and M. myosotis, L.

cubensis, M. occidentals, and P. mirabilis do not appear

to be consumed at all), amphipods and isopods were not

recorded in fecal samples.

The dietary specialization and restriction in Bermuda’s

terrapins carries penalties. It has been demonstrated that

they are exposed to a wide range of toxic compounds (e.g.,

trace metals, gasoline-range, and diesel-range petroleum

hydrocarbons and polycyclic aromatic hydrocarbons)

via food-chain contamination, specifically through the

ingestion of gastropods, but probably exacerbated by

the high incidence of associated sediment intake. It has

also been shown that these contaminants are transferred

to terrapins eggs, which show low hatching rates and

evidence of embryonic abnormalities (Outerbridge et al.

2016).

Conclusion

The field surveys and fecal analyses reported on here

showed that Diamondback Terrapins in Bermuda are

specialist microphagous molluscivores that do not exploit

the full range of potential prey species available to them.

The range of food items ingested is much narrower

than reported from North American populations, but

this is probably caused by the near absence of tidal

action that permits the accumulation of organic-rich

sediments browsed upon by abundant small gastropods.

The anchialine pools and surrounding vegetated areas

exhibit a low potential prey diversity in comparison with

those found in the salt marshes of the eastern seaboard

of the USA, but adult and juvenile terrapins evidently

select preferentially within this low diversity for small

gastropods of only two species (M tuberculata and H.

bermudensis).

Acknowledgments —We are grateful to the Mid

Ocean Club for granting access to the study site and

wish to express our thanks to S. Massey, M. Hoder, P.

Harris, and E. Limerick for their invaluable assistance

with field work. Funding for this study was provided

by the Atlantic Conservation Partnership, the Bermuda

Zoological Society, and the Mid Ocean golf club. This is

contribution #243 of the Bermuda Biodiversity Project

(BBP) Bermuda Aquarium, Natural History Museum and

Zoo, Department of Environment and Natural Resources.

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Mark Outerbridge works for the Bermuda Government at the Department of Environment and

Natural Resources and received his Ph.D. from the University College Cork (Ireland). He has spent

the last decade studying a wide variety of threatened and endangered species on Bermuda and also

has a professional interest in the impact that invasive, non-native species have upon Bermuda’s fragile

island-ecosystems.

Ruth O’Riordan is the head of the Graduate School and a senior lecturer at the School of Biological,

Earth and Environmental Sciences at the University Cork College (Ireland). Her research focuses on

temperate and tropical intertidal ecology, supply-side ecology of marine invertebrates, biology and

ecology of exotic aquatic species, climate change, and behavior of vertebrate animals.

Thomas Quirke is currently a lecturer within the Animal Management Department at Reaseheath

College in the United Kingdom. After completing a B.S. in Zoology at University College Cork in

Ireland, Thomas then moved on to complete his Ph.D., studying cheetahs within zoos in Ireland,

the UK, Canada, and Southern Africa. He next spent a year at the University of Pretoria and the

National Zoological Gardens of South Africa studying how animal personality influences the effects

of environmental enrichment.

John Davenport is Emeritus Professor of Zoology at University College Cork (Ireland) and holds

a D.Sc. from the University of London. A professional marine biologist since the 1970s, he has

collaborated with Bermudian scientists since the 1980s, working on fish, skinks, and turtles.

Amphib. Reptile Conserv.

January 2017 | Volume 11 | Number 1 | el 34

Official journal website:

amphibian-reptile-conservation.org

Amphibian & Reptile Conservation

11(1) [General Section]: 36-44 (e135).

Conservation status of Amphibians of Argentina:

An update and evaluation of national assessments

1 ’ 3 Marcos Vaira, 1 Laura C. Pereyra, Tviauricio S. Akmentins, and 2 Jon Bielby

1 Institute) de Ecorregiones Andinas (INECOA), CONICET, Universidad National de Jujuy, Av. Bolivia 1711 (4600), San Salvador de Jujuy,

ARGENTINA 2 Institute of Zoology), Zoological Society of London, Regent's Park, London NW1 4RY, UNITED KINGDOM

Abstract .—We present a review on the conservation status of the 177 species and subspecies

of amphibians of Argentina and compare the first national assessment, conducted in 2000, with

the most recent one, from 2012, to determine changes in conservation status over time. We also

evaluate the degree of taxonomic and geographic non-randomness in extinction risk among

these taxa. The present study shows an improvement in the knowledge of amphibian diversity in

Argentina, but also increasing evidence of population declines and species absences. Twenty-two

species showed a genuine increase in threat status between national assessments, and habitat

loss and/or degradation, chytrid fungus infection, and introduction of invasive species have been

reported as the main threats. Randomization tests showed families Telmatobiidae and Batrachylidae

to be over-threatened and Hylidae and Leptodactylidae to be significantly under-threatened. Also,

four ecoregions were shown to be significantly over-threatened (Patagonian Steepe, Patagonian

Woodlands, Puna, and Yungas Forests). This evaluation help to identify groups of species that face

similar suites and intensities of threat as a result of their overlapping geographical distributions and

shared biological susceptibility as a result of their evolutionary history. We consider that our results

highlight patterns and trends to alert policymakers and to guide priority actions.

Keywords. Batrachylidae, diversity, ecoregions, Hylidae, Leptodactylidae, threats

Citation: Vaira M, Pereyra LC, Akmentins MS, Bielby J. 2017. Conservation status of amphibians of Argentina: An update and evaluation of national

assessments. Amphibian & Reptile Conservation 11(1) [General Section]: 36-44 (el 35).

NoDerivatives 4.0 International License, which permits unrestricted use for non-commercial and education purposes only, in any medium, provided

the original author and the official and authorized publication sources are recognized and properly credited. The official and authorized publication

credit sources, which will be duly enforced, are as follows: official journal title Amphibian & Reptile Conservation / official journal website <amphibian-

reptiie-conservation.org>.

Received: 26 April 2016; Accepted: 25 November 2016; Published: 31 January 2017

Introduction

The widespread loss of biological diversity on a global

scale poses a challenge demanding effective methods to

assess the threat status of the biodiversity at a range of

spatial scales (Mace et al. 2008). Global, regional, and/

or local assessments of the status of species according to

their extinction risk are important tools for guiding the

development of conservation planning policies and regu¬

lations. Since most conservation actions are based on the

threat category assigned to the species, the implementa¬

tion of more efficient public policies and the improve¬

ment of public awareness may depend on reliable species

information and assessments (Hoffmann et al. 2010).

Argentina harbors the tenth largest amphibian fauna

among the 40 countries included in the Neotropical

Realm. The Argentine amphibian fauna is also highly

endemic, being among the twenty countries in the world

Correspondence. 3 marcos.vaira@gmail.com

in which 30% of its amphibian species are endemic

(Bolanos et al. 2008; Lavilla and Heatwole 2010). Like

in most Neotropical countries, there are major gaps of

information on the amphibian species of Argentina

including systematic, genetic, range size, natural history,

and ecology (Lavilla and Heatwole 2010). The usual

barriers faced to accurately assess the threat status of

amphibians in such countries are the many remote or

unexplored regions coupled with relatively few scientific

experts to detect, identify, and study species and/or popu¬

lations, and the limited resources available to evaluate

them (Becker and Loyola 2008; Brito 2008). Although

challenging, the sum of individual efforts by amphibian

researchers allowed, the development of the first national

Red List of amphibians of Argentina in 2000, using a

locally designed categorization method (Lavilla et al.

2000). Other contributions later summarized and updated

the information on Argentinean amphibian diversity,

Amphib. Reptile Conserv.

January 2017 | Volume 11 | Number 1 | el 35

Vaira et al.

geographic distribution, and description of the principal

threats (Lavilla and Cei 2001; Lavilla et al. 2002; Lavilla

and Heatwole 2010).

Given the substantial increase in the rate of amphibian

species description (Kohler et al. 2008), continual

updating of existing conservation assessments is

necessary in order to properly maintain that assesment’s

value (Stuart 2007). A status review requires the compi¬

lation of new information from new species descrip¬

tions, new taxonomic arrangements, and new evidence

or research, and in most cases it is necessary to reassess

the consensus of several experts about conservation

status (Lukey et al. 2010). To accomplish this task, a

new assessment of the conservation status of the amphib¬

ians of Argentina was published in 2012. The updated

assessment shows an improvement in the knowledge of

amphibian diversity in Argentina, but also increasing

evidence of population declines and species absences

(Vaira et al. 2012).

Nevertheless, none of the existing assessments or

updates on the diversity and conservation status of

Argentinean amphibians evaluate whether phylogeneti-

cally related species or those sharing similar distribu¬

tional ranges face similar kinds of threats. Taxonomic

and geographic selectivity in threats has been observed

in some vertebrates, consisting of non-random distribu¬

tions of the extinction risk of species among families or

regions (Russell et al. 1998). Species have different prob¬

abilities of extinction depending on intrinsic factors like

body size, population size, and genetic variability (Sodhi

et al. 2008). Moreover, the probability of extinction

rely also on external factors such as human disturbance,

disease, habitat loss, and other threatening processes,

as well as on the interaction between such extrinsic and

intrinsic factors (Bennett and Owens 1997), and even

stochastic events associated with small population sizes

(Schaffer 1981). It is therefore possible that species that

share some of these factors will have similar levels of

threats. Thus, an evaluation of taxonomic and geographic

patterns in the threat status of amphibians of Argentina

could be used to focus conservation practices on entire

clades or particular biogeographical regions rather than

on individual species (Mace et al. 2003; Bielby et al.

2006; Corey and Waite 2008).

Given the reported improvement in the knowledge

of amphibian diversity in Argentina (Vaira et al. 2012),

coupled with changes in the conservation status and the

intensity of threatening processes, prompted us to analyze

changes in species’ conservation status between the first

comprehensive Argentinean conservation assessment

(Lavilla et al. 2000) and the most recent one (Vaira et

al. 2012). We are interested to know if reported changes

are attributable to genuine improvement or deterioration

of their conservation status or attributable to improved

knowledge on taxonomy, ecology or distribution of the

species. We also evaluate the degree of non-random¬

ness in threat status of the species by taxonomy and

geographic distribution to analyze whether threat status

was randomly distributed across taxonomic families or

regions.

Materials and Methods

The method used to categorize threatened species in

Argentina was originally proposed by Lavilla et al.

(2000), adapted from the method of Reca et al. (1994).

More recently, Giraudo et al. (2012) reviewed the

method with the aim to improve consistency and provide

guidance on the assessment process. The categories are:

Insufficiently Known (IC), Not Threatened (NA), Vulner¬

able (VU), Threatened (AM), and Endangered (EP).

We evaluated the changes in the conservation catego¬

ries of the different taxa between the two assessments.

A status change due to reported increase or decrease

of threats was considered a “genuine” change. Those

changes attributable to improved knowledge of both

geographic distribution and taxonomy of the taxa were

considered as “non-genuine” status changes (adapted

from Hoffmann et al. 2010).

We followed the analytical methods proposed by

Bielby et al. (2006) to analyze whether threat status was

randomly distributed across taxonomic families and

regions. We combined the categories VU, AM, and EP

as Threatened and retained category NA as Not-threat-

ened. The taxa in the IC category were omitted from the

analysis, in order to remove the effects of non-random

lack of knowledge for conservation status (Bielby et

al. 2006). We then constructed two data sets ordering

threat categories by taxonomic family or region. We

assigned taxa range distributions to regions following the

ecoregion classification scheme described in Lavilla and

Heatwole (2010).

For each of the data sets, we first conducted a chi-

square test to test for deviation from the null expecta¬

tion that threatened taxa are distributed randomly among

families or ecoregions. When non-random extinction risk

was detected, we conducted further analyses to determine

which families or ecoregions deviated from the expected

level of threat. We did this by using a binomial test to

calculate the smallest family size necessary to detect a

significant deviation from the observed proportion of

threatened taxa and excluded the families represented by

an insufficient number of taxa.

For taxa in the remaining families, we generated a

null frequency distribution of the number of threatened

species from 10,000 unconstrained randomizations, by

randomly assigning the categories to all remaining taxa.

We then compared the actual number of threatened taxa

in the datasets with the null frequency distribution. The

null hypothesis (extinction risk is taxonomically and

geographically random) was rejected if this number fell

in the 2.5% at either tail of the distribution.

Amphib. Reptile Conserv.

January 2017 | Volume 11 | Number 1 | el 35

Conservation status of amphibians of Argentina

Table 1 . Taxonomic arrangements of amphibians of Argentina not included in the list of the 2012 national assessment by Vaira et

al. (2012).

Taxon name listed in Vaira et al. 2012

Changed to

Source

Alsodes gargola gargola

Alsodes gargola

Blotto et al. 2013

Alsodes gargola neuquensis

Alsodes neuquensis

Blotto et al. 2013

Not listed

Elachistocleis haroi

Pereyra et al. 2013

Not listed

Oreobates berdemenos

Pereyra et al. 2014

Pseudis limellus

Lysapsus limellum

Garda et al. 2010

Somuncuria somuncurensis

Pleurodema somuncurense

Faivovich et al. 2012

Leptodactylus diptyx

Adenomera diptyx

Pyron and Wiens 2011

Results

Update and summary of the conservation status

of the amphibians of Argentina

Based on taxonomic changes and the description of two

new species since 2012, updated list of amphibians of

Argentina consists of a total of 17 families, 42 genera,

and 177 species and subspecies (Tables 1 and 2).

Twenty taxa registered a “genuine” status change in

its threat categories when comparing the 2000 Argen¬

tinean conservation assessment with the 2012 list, (Table

2). Most status changes represent an increase in the threat

categories for the taxa due to: population decline (65%),

habitat deterioration (25% of taxa), invasive species

(10% of taxa), or by infection caused by the chytrid

fungus Batrachochytrium dendrobatidis (10% of taxa).

(Table 3). Twenty-five taxa registered “non-genuine”

status changes attributable to improved knowledge

of taxonomy or geographic distribution, while 19 taxa

maintained the same threat category as listed in the first

national assessment (Table 2).

Degree of non-randomness in threats of the

species

The family data set showed a significant deviation from

a random distribution of threatened species and subspe¬

cies among the amphibian families (x 2= 76.5, df = 9 P <

0.001). Randomization tests showed two families to be

significantly overthreatened (Telmatobiidae and Batra-

chylidae with 100% and 60% of their taxa threatened,

respectively) and two families to be significantly under¬

threatened (Hylidae and Leptodactylidae with 5% and

14% of the taxa threatened, respectively) (Tables 2 and

4).

The ecoregions data set also showed a significant

deviation from a random distribution of threatened taxa

(% 2 = 140.25, df = 14 P < 0.001). Randomization tests

showed four ecoregions overthreatened (Fig. 1): Pata¬

gonian Steepe (75% of the taxa threatened); Patagonian

Woodlands (65% of the taxa threatened); Puna (71% of

the taxa threatened), and Yungas Forests (29% of the taxa

threatened) [Table 5],

Discussion

This update shows a substantial improvement in

the knowledge of amphibian diversity in Argentina

since the first major assessment in 2000, with 11 new

species described (see Vaira et al. 2012) and improved

knowledge on taxonomy and/or geographic distribution

of several species reflected by 16 taxa decreasing their

threat categories and changing status as a consequence of

the amount of information (i.e., the “non genuine” status

change).

Unfortunately, there is also evidence of taxa

increasing their threat status in Argentina since the first

national conservation assessment. Habitat loss and/or

degradation, chytrid fungus infection, and introduction

of invasive species have been considered as principal

threats suggested for eight species changing to higher

threat categories (Vaira et al. 2012). Nonetheless, the

lack of studies that simultaneously evaluate the impor¬

tance of those threats on the species is notable.

Major concerns constitute the lack of registries verifi¬

cation of these four species in the wild for prolonged time

lapses: Telmatobius ceiorum, T. laticeps , Gastrotheca

christiani , and G. chrysosticta (Barrionuevo and Ponssa

2008; Akmentins et al. 2012) even after exhaustive

surveys conducted in recent years within their natural

geographic ranges. Whether these species are still extant

is uncertain. Nevertheless, it is generally recommended

to be extremely cautious to declare a species extinct

because of the conservation implications involved, and it

seems appropriate to encourage additional conservation

efforts until there is no reasonable doubt of its extinc¬

tion (Mace et al. 2008; Akmentins et al. 2012). However,

to better reflect the likelihood of these species becoming

extinct under prevailing circumstances we suggest that

their global conservation status should be reconsidered.

Like in most Neotropical countries, there are major

gaps of information on the amphibian species of Argentina

including genetic, geographic range size, natural history,

and ecology. The status of most of their populations is

unknown since there is simply not enough information

to estimate or infer a trend. Despite limited information,

reported declines and identified threats require quick

decisions on prioritizing conservation actions on certain

Amphib. Reptile Conserv.

January 2017 | Volume 11 | Number 1 | el 35

Vaira et al.

Fig. 1. Map of the ecoregions of Argentina from Burkart et

al. 1999. Numbers indicate ecoregions divisions as follows:

(1) Puna, (2) High Andean, (3) Yungas Forest, (4) Dry Chaco,

(5) Humid Chaco, (6) Delta and Islands of the Parana River,

(7) Esteros of Ibera, (8) Paranaen Forest, (9) Campos and

Malezales, (10) Espinal, (11) Pampas, (12) Monte of Sierras

and Bolsones, (13) Monte de Elanuras and Mesetas, (14)

Patagonian Steppe, (15) Patagonian Woodlands. Arrows

indicate the four overthreatened ecoregions.

species over others. We encourage the development and

implementation of a conservation action plan of threat¬

ened amphibian species of Argentina through specialist

consensus.

Hylidae and Leptodactylidae were found to be under¬

threatened families probably because they both contain

taxa with large geographic distributions and small

numbers of endemic species (Lavilla and Heatwole 2010).

Randomization test results suggest that Telmatobiidae

and Batrachylidae clades are overthreatened and hence

may be especially prone to extinction. Reported threats

within these families are consistent with the possibility

that shared evolutionary history per se is an important

precursor to vulnerability. The genus Telmatobius shares

ecological traits present in many amphibians that have

declined worldwide such as restricted distributions in

high mountain ranges, low fecundity, and aquatic adults

(Lips et al. 2003; Sodhi et al. 2008; Bielby et al. 2006).

Then, endemic populations of Telmatobius should be

more prone to extinction from environmental and demo¬

graphic stochasticity which prompts us to consider how

severe the human impacts on those species will be. Four

possible factors have been suggested as causes of decline

for the species of Telmatobius in Argentina: unusual

climate coupled to an increase in erosive processes and

debris flowing events in montane streams, introduction

of exotic predatory fishes in the river basins, and chytrid

fungus infection (Barrionuevo and Ponsa 2008; Vaira

et al. 2012). Similar results were reported in Ecuador,

where the most critically endangered species belonging

to Telmatobius genera occurred in regions characterized

by drier conditions and high suitability for Batrachochy-

trium dendrobatidis (Menendez-Guerrero and Graham

2013).

Species with similar life history traits and habitat use

patterns are likely to be more sensitive to environmental

instability and are less able to adapt to or recover from

environmental or ecological changes (Sodhi et al. 2008).

The evaluation of non-randomness of threat status can

help to identify groups of related species that face similar

suites of human-caused threat and biological suscepti¬

bility due to overlapping geographical distributions and

shared evolutionary history respectively. This approach

can help us to pinpoint needs for emergency action

and to alert policymakers and conservation managers.

This knowledge could be used to plan future protected

areas where threat is concentrated and to guide mitiga¬

tion measures. Telmatobius and Atelogjiathus might

be examples of such an approach, setting conservation

actions in the specific ecoregions where these genera

inhabit (Patagonian Steepe and Woodland, Puna and

Yungas Forests) and to manage their specific threats

(e.g., introduction of predatory fishes and/or mining)

that might yield better results than directing resources

towards single species or individual populations.

On the other hand, broadly distributed species

represent another challenge for setting conservation

priorities if they comprise evolutionary lineages that

may be under different levels of threat. An assessment

at country level may lead to conflicting results in the

threat category of a species complex and the possibility

of over or underestimation of their conservation status. It

is conceivable that as studies continue, species which we

now consider widespread may indeed be local endemics.

Additionally, since the effects of threats can be expected

to vary spatially, especially in heterogeneous environ¬

ments, better assessments are needed across ecoregions

and thus fully accomplish the conservation assessment of

amphibians in Argentina.

Because our knowledge to estimate the risk of extinc¬

tion of amphibians of Argentina is still rather limited,

the identification of most threatened regions may prove

useful in cases where available data limit the certainty

of the assessment outcomes. Our approach based on the

evaluation of non-randomness of threat status may help

to identify regions that are at greater risk and to capture

the attention of researchers and policymakers. Thus, our

rather broad results may be refined for more concerted

Amphib. Reptile Conserv.

January 2017 | Volume 11 | Number 1 | el 35

Conservation status of amphibians of Argentina

Table 2. Comparison of the 2000 and 2012 national assessments of threatened species of Argentina showing number of taxa with

the same, increasing, or decreasing threat categories. A status change due to reported increase or decrease of threats was considered

a “genuine” change. Those changes attributable to improved knowledge of both geographic distribution and taxonomy of the taxa

were considered as “non-genuine” status changes (adapted from Hoffmann et al. 2010).

Threatened

species and

Families subspecies

on 2000 Red

List

Siphonopidae (3)

Typhlonectidae (1)

Alsodidae (9)

Batrachylidae (15)

Brachycephalidae (1)

Bufonidae (30) b

9 a

ya,b

2 a

l a

3 a

Centrolenidae (1)

Ceratophryidae (6)

Craugastoridae (3) b ’ c

2 b

Hemiphractidae (3)

Hylidae (38)

Hylodidae (2)

Leptodactylidae (37)

Microhylidae (4) b c

Odontophrynidae (8)

Rhinodermatidae (1)

Telmatobiidae (15)

15 b

Notes: Taxonomy follows Frost (2015). See Vaira et al. (2012) for the complete list of species and subspecies considered (see also text of this contribution to account

for a few nomenclatural changes at genera and species level).

a The threatened taxa in 2000 and 2012 are not the same. See Vaira et al. (2012).

b Include new species described after 2000.

c Include one new species described after 2012 and not evaluated.

studies focused on particular regions to expand the

assessment goals not only to identify species at risk but

also threats to ecological and evolutionary processes.

Conclusion

Applications of the national assessment:

Challenges and future directions

Compiling a national threatened species lists helps

to reveal information gaps and stimulate data collec¬

tion focusing on species or areas where there may be

needed conservation actions and where more research

may be required (Gardenfors 2001). We have now a

substantial body of knowledge that can provide insights

on the conservation status of amphibians of Argentina.

A remaining task should be to objectively evaluate the

uncertainties of the national assessment. The perfor¬

mance of the national assessment may be improved by

testing and refining the accuracy of protocols and criteria

to ensure future reassessments in an objective, compa¬

rable, and repeatable manner. Also, we must foster better

linkages between national and global assessment efforts

(de Grammont and Cuaron 2006).

Much is still unknown about potential threats in most

species of amphibians of Argentina and many groups

exhibit high levels of data deficiency doing status assess¬

ments unevenly detailed across species (Vaira et al.

2012). Due to the limited number of empirical data for

most species, the national assessment can assist in the

identification of groups of species that are more prone

to future declines under common threats, due to their

shared traits and geographic distributions, constituting an

alternative approach to integrate this knowledge into the

development of coordinated strategies for data collec¬

tion or into proactive conservation programs. Data of

“genuine” changes in the status of threatened species can

then be used to measure progress of programs, and also

be used to inspire development of national policies and

legislation to protect species and particular regions they

inhabit.

An exclusive focus on species-based approaches to

conservation planning is controversial (Sastersdal and

Gjerde 2011; Nicholson et al. 2013). As better data and

methodologies become available, defining priority areas

for conservation constitute a most desirable goal (Jenkins

et al. 2013). Ideally, we must also address the complexity

of natural ecosystems including phylogenetic, ecolog-

Threatened

species and

subspecies

on 2012 Red

List

Number

taxa same

threat

categories

“Genuine”

status

change

“Non-

genuine”

status

change

Amphib. Reptile Conserv.

January 2017 | Volume 11 | Number 1 | el 35

Vaira et al.

Table 3. Species of amphibians of Argentina that increase their

threat categories between the 2000 and the 2012 national Red

List assessment. Only species with “genuine changes” were

considered (see text for explanation). Reasons of changes

follow Vaira et al. (2012). Values in parentheses indicate status

deteriorations from a lower to a higher category of threat.

Species

Threat category

in Argentina

Reason of change

Alsodes australis

VU (-1)

invasive species

Alsodes gargola

VU(-l)

habitat deterioration

Alsodes neuquensis

AM (-1)

invasive species

Alsodes pehuenche

EP (-3)

habitat deterioration

Rhinella achalensis

AM (-1)

habitat deterioration / Bd *

Ceratophys ornata

VU (-1)

habitat deterioration

Gastrotheca christiani

EP (-2)

population decline

Gastrotheca chrysosticta

EP (-2)

population decline

Gastrotheca gracilis

EP (-2)

population decline

Pleurodema somuncurense

EP(-l)

habitat deterioration

Rhinoderma darwini

AM (-1)

population decline

Telmatobius ceiorum

EP (-2)

population decline

Telmatobins contrerasi

AM (-2)

population decline

Telmatobius hauthali

AM (-1)

population decline

Telmatobius laticeps

EP (-2)

population decline

Telmatobius oxycephalus

AM (-1)

population decline

Telmatobius pisanoi

AM (-1)

population decline / Bd *

Telmatobhis schreiteri

AM (-1)

population decline

Telmatobius scrocchii

VU (-1)

population decline

Telmatobius stephani

AM (-1)

population decline

Categories: Vulnerable (VU), Threatened (AM), Endangered (EP).

* Bd: Infection caused by chytrid fungus, Batrachochytrium dendrobatidis.

Table 4. Results of the analysis of distribution of threatened

species in the amphibian families of Argentina after the omission

of Insufficiently Known (IC) species. The null hypothesis

(threat status is taxonomically random) was rejected if P values

were equal or less than 0.025% at either tail. Families under

or overthreatened are bolded. NA: families represented by an

insufficient number of species from analysis.

Families

Threatened

taxa / Total # of

taxa*

> Expected

threat-level

P-value

< Expected

threat-level

P-value

Alsodidae

5/7

0.04

Batrachylidae

9/11

0.001

Bufonidae

7/25

0.74

0.49

Ceratophryidae

1/6

0.89

Hemiphractidae

3/3

0.03

Hylidae

2/37

<0.001

Leptodactylidae

5/36

0.01

Microhylidae

0/3

Odontophrynidae

1/7

0.93

Telmatobiidae

15/15

<0.001

* IC species omitted.

ical, and evolutionary processes (Lindenmayer et al.

2007). National conservation assessment disaggregated

by habitats, ecosystems, or ecoregions can thus provide

a valuable base to support the design of priority areas

requiring us to translate assessments from country to

regional or local levels.

A common confusion introduced in some national

conservation assessment applications is to consider

conservation status and conservation priorities as equals

when they are related but different processes. Conser¬

vation status alone should not necessarily determine

conservation priorities (de Grammont and Cuaron 2006).

Assigning species to a threat category in a conservation

assessment should be an objectively scientific process to

estimate the risk of extinction of a species. By contrast,

setting conservation priorities determine which species

should be protected and will often involve political as

well as logistical considerations, so it is possible to

establish different sets of species with conservation prior¬

ities in different regions within the country. Both compo¬

nents are essential for better policy-making and for more

accurate scenarios for conservation and management.

Many national or regional conservation agencies

interpret conservation assessments as a priority¬

setting tool for conservation action (Miller et al. 2007).

Sometimes, there is a direct connection between conser¬

vation assessments and conservation policies, basing

protective legislation or conservation actions directly on

conservation categories. This can have undesired conse¬

quences, such as Data Deficient species being disregarded

when allocating resources for conservation or protec¬

tion. We must consider an increased communication and

cooperation between researchers and policy-makers for

generating and using national conservation assessments

to effective conservation actions and legislation.

Acknowledgments —This project was partially sup¬

ported by a PICTO-UNJu grant # 153 and a Secter-UNJu

grant # D-084. We specially thank two reviewers for

providing insightful comments that greatly helped us to

improve the manuscript.

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tives. Conservation Biology 21(3): 684-696.

Nicholson E, Lindenmayer DB, Frank K, Possingham

HP. 2013. Testing the focal species approach to

making conservation decisions for species persis¬

tence. Diversity and Distributions 19(5-6): 530-540.

Pereyra LC, Akmentins MS, Laufer G, Vaira M. 2013.

A new species of Elachistocleis (Anura: Microhyl-

idae) from north-western Argentina. Zootaxa 3694(6):

525-544.

Pereyra MO, Cardozo DE, Baldo J, Baldo D. 2014.

Description and phylogenetic position of a new

species of Oreobates (Anura: Craugastoridae) from

Northwestern Argentina. Journal of Herpetology

70(2): 211-227.

Pyron RA, Wiens JJ. 2011. A large-scale phylogeny of

Amphibia including over 2800 species, and a revised

classification of extant frogs, salamanders, and caeci-

lians. Molecidar Phylogenetics and Evolution 61(2):

543-583.

Reca A, Ubeda C, Grigera D. 1994. Conservacion de la

fauna de tetrapodos. I. Un indice para su evaluacion.

Mastozoologia Neotropical 1(1): 17-28.

Russell GJ, Brooks TM, McKinney MM, Anderson CG.

1998. Present and future taxonomic selectivity in

bird and mammal extinctions. Conservation Biology

12(6): 1,365-1,376.

Sastersdal M, Gjerde I. 2011. Prioritising conservation

areas using species surrogate measures: Consistent

with ecological theory? Journal of Applied Ecology

48(5): 1,236-1,240.

Shaffer ML. 1981. Minimum population sizes for species

conservation. BioScience 31(2): 131-134.

Sodhi NS, Bickford D, Diesmos AC, Lee TM, Koh

LP, Brook BW, Sekercioglu CH, Bradshaw CJA.

2008. Measuring the Meltdown: Drivers of Global

Amphibian Extinction and Decline. PLoS ONE 3(2):

el636.

Stuart SN. 2007. The continuing need for assess¬

ments: Making the Global Amphibian Assessment an

ongoing process. Pp: 43—44 In: Amphibian Conser¬

vation Action Plan. Editors, Gascon C, Collins JP,

Moore RD, Church DR, McKay JE, Mendelson III

JR. IUCN/ SSC Amphibian Specialist Group, Swit¬

zerland, USA, Gland, Cambridge. 64 p.

Vaira M, Akmentins M, Attademo M, Baldo D, Barrasso

D, Barrionuevo S, Basso N, Blotto B, Cairo S, Cajade

R, et al. 2012. Categorizacion del estado de conser¬

vacion de los anfibios de la Republica Argentina.

Cuadernos de Herpetologia 26(3): 131-159.

Amphib. Reptile Conserv.

January 2017 | Volume 11 | Number 1 | el 35

Conservation status of amphibians of Argentina

Marcos Vaira is a researcher at the Institute de Ecorregiones Andinas (CONICET - Universidad

Nacional de Jujuy). His primary area of interest is amphibian diversity. The aim of our research is

to contribute to a better understanding on the structure and functioning of amphibian communities

in the subtropical montane forest landscapes of Northwestern Argentina to provide a solid

framework for their conservation.

Laura Cecilia Pereyra is a researcher at the Institute de Ecorregiones Andinas (CONICET

- Universidad Nacional de Jujuy). Her primary area of interest is the study of multifaceted

components of diversity combining measures to assess and compare amphibian diversity in

human-modified forest landscapes of Northwestern Argentina.

Mauricio Sebastian Akmentins is a researcher at the Institute de Ecorregiones Andinas

(CONICET - Universidad Nacional de Jujuy). His primary area of interest is the ecology and

conservation of direct-developing frogs of the subtropical montane forest of Northwestern

Argentina.

Jon Bielby is a Research Fellow at the Institute of Zoology, London. He researches wildlife

disease, population decline, and extinction risk, with a particular focus on amphibians.

Amphib. Reptile Conserv.

January 2017 | Volume 11 | Number 1 | el 35

Official journal website:

amphibian-reptile-conservation.org

Amphibian & Reptile Conservation

11(1) [General Section]: 45-50 (e136).

Preliminary observations on the circadian variation in

site fidelity in Atelopus hoogmoedi (Lescure, 1974)

(Anura, Bufonidae)

Michael P.J. Nicolai, 12 Sara Porchetta, 13 Shashank Balakrishna,

14 David P. Botha, and 1 ’ 5 Philippe J.R. Kok

'Amphibian Evolution Lab, Biolog)’ Department, Vrije Universiteit Brussel, Pleinlaan 2, B-1050 Brussels, BELGIUM

Key words. Anura, Atelopus hoogmoedi , Guyana, homing behavior, Iwokrama, site fidelity

Citation: Nicolai MPJ, Porchetta S, Balakrishna S, Botha DP, Kok PJR. 2017, Preliminary observations on the circadian variation in site fidelity in

Atelopus hoogmoedi (Lescure, 1974) (Anura, Bufonidae). Amphibian & Reptile Conservation 11(1) [General Section]: 45-50 (el 36).

NoDerivatives 4.0 International License, which permits unrestricted use for non-commercial and education purposes only, in any medium, provided

the original author and the official and authorized publication sources are recognized and properly credited. The official and authorized publication

credit sources, which will be duly enforced, are as follows: official journal title Amphibian & Reptile Conservation ; official journal website <amphibian-

reptile-conservation. org>.

Received: 25 February 2016; Accepted: 08 June 2016; Published: 31 January 2017

The genus Atelopus (Bufonidae) is mostly composed

of conspicuously colored species, several of which are

known to secrete toxins (Fuhrman 1969; Yotsu-Yamashita

and Tateki 2010). The genus has recently received in¬

creased attention due to severe decline in population num¬

bers, often suggested to have revealed extinctions (La Marca

et al. 2005; Pounds et al. 2006; Wake and Vredenburg

2008; but see Luger et al. 2008 for the Guianas). This

mass decline has been attributed to multiple factors such as

habitat loss, pollution, introduced species, and the chytrid

fungus Batrachochytrium dendrobatidis (La Marca et al.

2005). One of the most widespread, and probably less

threatened species in the genus is Atelopus hoogmoedi

(Fig. 1 A-B), which is found in French Guiana, Suri¬

name, Guyana, and northern Brazil (Noonan and Gau¬

cher 2005; Kok and Kalamandeen 2008; Luger et al.

2008; Segalla et al. 2014). Two color morphs co-occur

syntopically in Iwokrama (Guyana), an orange and a

yellow morph (Fig. 1 A-B). This diurnal toad exhibits

spatio-temporal segregation of sexes, with males usually

found near streams, while females are found deeper in

the forest, away from water bodies (Luger et al. 2009).

During the breeding season, which mostly occurs in the

dry season (see below), females migrate to streams for

mating (Fig. 1 C, Luger et al. 2009). Similar reproduc¬

tive strategies are observed in other anurans and are often

characterized by site fidelity in which the males remain

in the vicinity of the same perching site for the duration

of the breeding period (e.g., Roithmair 1992; Ringler et

al. 2009).

Several studies have investigated homing behavior

and site fidelity in some Atelopus species (e.g., Crump

1986), including A. hoogmoedi (Luger et al. 2009). It has

been suggested that site fidelity increases the probabil¬

ity of finding a suitable mate by improving the detection

either by males through an increased field of vision, or

by females as a result of a more conspicuous male perch

(Himmel 2013). However, the occurrence of site fidel¬

ity outside of the reproductive season makes the mate

detection hypothesis unlikely (Crump 1986; Luger et

al. 2009). Alternatively, site fidelity may provide better

knowledge of the local microenvironment making forag¬

ing more efficient, as well as providing means of protec¬

tion from predation e.g., by good knowledge on possible

escape routes (Luger et al. 2009). Previous studies have

focused on diurnal site fidelity, and investigations per¬

taining to circadian variation in site fidelity are lacking.

Since protection from predation is a plausible hypothesis

explaining site fidelity, nocturnal behavior (when the di¬

urnal animal is most vulnerable and an optimal protective

perch likely required) deserves further attention. If pro¬

tection is indeed a major driver for site fidelity, scarcity

of optimal refuges might lead to increased site fidelity.

This hypothesis was briefly tested during fieldwork

conducted in the Iwokrama Forest Reserve, central Guy¬

ana. Iwokrama is mostly covered by tropical moist low¬

land forest (Holdridge 1967), with some of the Iwokrama

Mountains reaching ca. 900 m asl (MPFITRF 2009).

Climate in Iwokrama is tropical, with an annual mean

temperature of 25 °C and a mean rainfall of 3,000 mm

Correspondence. 1 michaelnicolai22@hotmail.com (Corresponding author); 2 saraporchetta@hotmail.com;

3 rb.shashank@gmail.com; 4 slangseim@gniail.com; ^Philippe.Kok@vub.ac.be (Corresponding author).

Amphib. Reptile Conserv.

January 2017 | Volume 11 | Number 1 | el 36

Nicolai et al.

Fig. 1. (A) Orange and (B) Yellow color morphs of Ate/opus hoogmoedi, both encountered at the studied locality in the Iwokrama

Mountains, Guyana. (C) Typical breeding habitat of A. hoogmoedi in the Iwokrama Mountains. Photos by PJRK.

in the north of the reserve and 1,400 mm in the south

of the reserve (MPFITRF 2009). Wet season usually ex¬

tends from May to August and again from November to

February (MPFITRF 2009), although this has seemed

more irregular in recent years, especially during El Nino

events (reported as particularly strong in 2015, and the

months of November and December 2015 were very dry

in Iwokrama).

Fourteen Atelopus hoogmoedi males were “marked”

(see below) and “recaptured” in order to track their circa¬

dian variation in site fidelity. All individuals were spotted

along a portion of the trail between Turn Falls and the

Linden-Lethem road (between N 4°24.74’ W 58°47.13’

and N 4°24.63’ W 58°47.30’; WGS 84; Fig. 2). Eleva¬

tion ranged between 92 and 120 m asl. The trail was

walked twice a day, once at 13h (daylight) and once at

18h (shortly after nightfall) between November 27 and

December 1, 2015. Color pattern is individual-specific in

this species (see Fig. 1 A-B), and individuals were iden¬

tified based on photographs of the dorsal pattern taken in

situ using a digital camera (Canon® Eos 7D). To limit

physical interaction, ventral patterns were not examined

(no specimen was manipulated during this study). The

use of color pattern for identification in this species is

an appropriate alternative to invasive marking techniques

such as toeclipping (Luger et al. 2009). Locations of ob¬

servations were recorded using a Garmin eTrex 30®

GPS, and marked with colored flagging tape at the site

of first detection, allowing immediate recognition of

the site. For each observation three parameters were re¬

corded: (1) height above ground; (2) perch type (catego¬

rized as leaf litter, shrub, or rock); and (3) distance from

Amphib. Reptile Conserv.

January 2017 | Volume 11 | Number 1 | el 36

Circadian variation in site fidelity in Atelopus hoogmoedi

original diurnal or nocturnal perch site. Distances were

measured using a 50-m measuring tape. In order to stan¬

dardize the procedure, an individual was considered to be

present if three investigators were able to track it within

a three-meter radius from the original perch site within

three minutes. We used independent r-tests to compare

the observation probabilities between different days and

different sampling times. Additionally, Fisher’s exact test

was used to determine whether any of the two most com¬

mon substrates (shrub or leaf litter) was used more than

the other. All statistical analyses were done using IBM

SPSS® V20.0.0.

Observation probabilities did not differ significantly

between days (/ = 1.14, P - 0.26), nor did they differ

significantly between daylight (62%) and night (69%)

{t = 0.71, P = 0.54). More individuals did, however, re¬

turn to the original perch site at night than during the

day (distance from original perch site = 0; P = 0.08). Fi¬

nally, a difference in perching substrate use was found

at different times of the day (Fig. 3). While this differ¬

ence between substrate use was not significant during

daylight (P = 0.25), a significant difference was found

at night when most frogs were present on shrubs (P =

0.00). Unlike Luger et al. (2009), who detected A. hoog¬

moedi in leaf litter only 14% of the time, in our study leaf

litter was the most used microhabitat, especially during

daylight (64%). After dark most individuals were found

perching in shrubs, at heights between 10-130 cm.

Most Atelopus species, including A. hoogmoedi , mate

close to streams, and occurrences of flashfloods have

been reported to wash away entire Atelopus populations

(Duellman and Trueb 1986). As a result, high perching

sites were explained as a strategy against such sudden

water rises (e.g., Duellman and Trueb 1986; Luger et

al. 2009). This could explain the differences in obser¬

vations between our study and previous studies (Luger

et al. 2009). Our study, in which individuals were often

found on the ground, was conducted during the dry sea¬

son (see above), in contrast with previous studies that

were conducted during the wet season (protection from

sudden water rises is likely more necessary during the

latter). Furthermore, as reproductive interactions occur in

the leaf litter (observation of amplexus during our field¬

work), male individuals are more likely to perch on the

leaf litter during breeding season to increase the chance

of inter-sexual interactions. Alternatively, differences

Legend

Iwokrama Forest Zones

Sustainable Utilisation Area (184.506.3 ha)

_| Wilderness Preserve (187.174.5 ha)

n FairView Village

0 3.75 7.5 1 5 Kilometers

I i i i I i i i 1

rtf'®*

Venezuela

Atlantic

Ocean

Suriname

Field Station

Fig. 2. Geographical overview of the study area. (A) Map of the Iwokrama Forest Reserve and its location in Guyana (top right

corner). The red line crossing Iwokrama corresponds to the Linden-Lethem Road. (B) Relief map of the Iwokrama Mountains with

Turn Falls represented by a black triangle (N 4°24.770’ W 58 o 47.06L). (C) Portion of the trail between Turn Falls camp and the

Linden-Lethem Road monitored, with dots corresponding to Atelopus individuals (from Al - N 4°24.742’, W 58°47.130’ to A14 - N

4°24.750’, W 58°47.128’). A and B from Kok et al. (2013).

Amphib. Reptile Conserv.

January 2017 | Volume 11 | Number 1 | el 36

Nicolai et al.

Sampling

time

□ Daylight

Shrub Leaf litter

Microhabitat

Fig. 3. Substrate (shrub or leaf litter) use in Atelopus hoogmoedi during the day (light grey) and night (dark grey) at Turn Falls,

Guyana. As indicated, substrate use was significantly different after dark.

with Luger et al. (2009) might be explained by differ¬

ent abiotic and biotic factors between the two study sites.

As previous studies indicated that perching on shrubs

occurred during both breeding and non-breeding sea¬

son (Crump 1986; Luger et al. 2009), it is unlikely that

this elevated perching plays a major role in mating. The

fact that the frogs leave these perch sites diumally, when

they are actively reproducing, further corroborates the

hypothesis that elevated perching is not reproduction re¬

lated. In other Atelopus species, such as A. zeteki , higher

nocturnal perches are proposed to be a safe retreat from

predators for this diurnal frog, shifting vigilance from vi¬

sual to tactile (Lindquist et al. 2007). Some perches serve

as better retreats than other perches, and difference in

perch quality could drive both diurnal and nocturnal site

fidelity. Our preliminary data indeed show that there was

no significant difference in site fidelity between night and

day as would be expected when site fidelity is linked to

lower predation pressure. Furthermore, individuals re¬

turned more to the original nocturnal perching site than

to the original diurnal perching site. This indicates that

site fidelity might actually be linked to nocturnal perch¬

ing site rather than to diurnal perching site. As both for¬

aging and breeding occur during the day, the protection

hypothesis provides a good explanation for this nocturnal

site fidelity, and site fidelity in general.

Some species of Atelopus are known to have lived

over ten years in the wild (La Marca 1984), and at least

one individual was recorded on the same boulder two

years after the previous observation (Crump 1986). Such

life history strategies make Atelopus ideal organisms for

study of long-term site fidelity. Future studies are encour¬

aged to expand our preliminary findings by increasing

the length of the study, and if possible the number of re¬

captures.

In conclusion our observations, although sparse, seem

to confirm that Atelopus hoogmoedi does indeed show

strong diurnal and nocturnal site fidelity, during breeding

and non-breeding seasons. Although several hypotheses

may explain this, the fact that perch site return rate is the

highest after dark supports the predation evasion hypoth¬

esis.

Acknowledgments. —These observations were made

in the framework of the Field Herpetology course in Guy¬

ana provided to the second year students of the Master

in Herpetology at Vrije Universiteit Brussel. We are in¬

debted to Raquel Thomas (Iwokrama, Guyana) for grant-

Amphib. Reptile Conserv.

January 2017 | Volume 11 | Number 1 | el 36

Circadian variation in site fidelity in Atelopus hoogmoedi

ing us the permission to have this training course taught

in the Iwokrama Mountains. We also thank Marcelo

Kokubum (Universidade Federal de Campina Grande,

Brazil), Stefan Lotters (Universitat Trier, Germany), and

Ross MacCulloch (Royal Ontario Museum, Canada) for

comments that improved our original manuscript, and

Ruben D.E. Culqui, Raheleh Dezfoulian, Yousri El Adak,

and Berengere Ferrier for help and companionship dur¬

ing fieldwork.

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hantgen PB, Criswell JM. 2007. Nocturnal position in

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ing. Phyllomedusa 6(1): 37-44.

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frogs (Atelopus) in the Guy anas. Studies on Neotropi¬

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quin toads (Amphibia: Atelopus hoogmoedi) from Su¬

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Forest. 2009. Available at: http://idbdocs.iadb.org/

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JA, Fogden MP, Foster PN, La Marca E, Master KL,

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Azofeifa A, Still CJ, Young BE. 2006. Widespread

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Amphib. Reptile Conserv.

January 2017 | Volume 11 | Number 1 | el 36

Nicolai et al.

Michael Nicolai' is a Ph D. student in the Amphibian Evolution Lab, Vrije Universiteit Brussel. He received

his B.S. and M.S. in Biology at the KU Leuven (Belgium) as well as an additional master in Herpetology at

the Vrije Universiteit Brussel. His main research interests are amphibian evolution, in particular the evolu¬

tion of coloration in different ecomorphs of frogs.

Sara Porchetta is a Ph.D. student at the Environmental and Applied fluid Dynamics department of the von

Karman Institute, Belgium. Both her B.S. and M.S degree were in Engineering at the KU Leuven (Belgium),

after which she obtained a research master at the von Karman Institute. Apart from her main research focus

she has a major interest in biodiversity, in particular that of amphibians.

Shashank Balakrishna is a Master’s student studying Herpetology at the Vrije Universiteit Brussel, Bel¬

gium. He is due to complete his degree in June 2017 with a thesis on the effectiveness of tail autotomy across

different landscapes. He is an active member of the Centre for ecological sciences at the Indian Institute of

Science, where he works on the local adaptations of lizards from an eco-physiology and behavioral ecology

approach within landscape levels. He also interns at the Universiteit Antwerpen where he investigates at¬

tributes influencing personality traits in a native and invasive population of Podarcis muralis.

David P. Botha is a South African trained conservationist, most recently graduating (2016) from the Vrije

Universiteit Brussel, Belgium, with a Master’s degree in biology. With a strong background in ecology and

a great passion for herpetology, he aims to use multidisciplinary approaches to solve complex questions

within these fields. He continues to equip himself with new skillsets that will aid in this endeavour. His main

interests lie with the African herpetofauna, particularly snakes and other squamates.

Philippe J. R. Kok is a Belgian evolutionary biologist and herpetologist. He obtained his Ph.D. in biology

at the Leiden University (The Netherlands) in 2013. He is currently LWO postdoctoral researcher in the Am¬

phibian Evolution Lab at the Vrije Universiteit Brussel, Belgium, where he also teaches field Herpetology to

the second year Master’s students. His interests are eclectic, the main ones being the evolution, systematics,

taxonomy, biogeography, and conservation of amphibians and reptiles in the Neotropics, more specifically

from the Guiana Shield. His work now primarily focuses on vertebrate evolution in the Pantepui region.

Amphib. Reptile Conserv.

January 2017 | Volume 11 | Number 1 | el 36

Official journal website:

amphibian-reptile-conservation.org

Amphibian & Reptile Conservation

11(1) [General Section]: 51-71 (e137).

urn:lsid:zoobank.org:pub:B35CE81 E-66F5-48C7-9049-E5C3598E4E5A

A new rupicolous species of gecko of the genus

Hemidactylus Oken, 1817 from the Satpura Hills, Central India

^Zeeshan A. Mirza and 2 David Raju

l G-18, 4th A-Cross Rd, Canara Bank Layout, Rajiv Gandhi Nagar, KodigehaUi, Bengaluru, Karnataka 560097, INDIA 2 Singinawa Jungle Lodge,

Tehsil Baihar, Balaghat, Kohka 481111, Madhya Pradesh, INDIA

Abstract .—We here describe a new species of rupicolous gecko from the Satpura Hills of central

India. The new species is a member of the Hemidactylus brookii complex, and can be distinguished

based on the following suite of characters: moderate sized species (SVL 54.3-74.2 mm); anterior

postmental width equal to first infralabial; posterior postmental width equal to second infralabial,

posterior postmental not in contact with first infralabial; enlarged, keeled, tubercles, fairly regularly

arranged in 15-16 longitudinal rows on dorsum; two angular series of seven precloacal femoral

pores separated by diastema of eight non-pored scales; non-pored scales equal to size of pored

scales; scales bordering anterior edge of pored scales half the size of pored scales; five lamellae

on digit I and seven on digit IV of manus as well as pes; lamellae on digit IV and V of pes absent on

basal 25% of the digit; legs long and slender; ventral aspect of tail with broad caudal scales covering

~80% of tail; two subconical post cloacal spurs, anterior spur slightly larger than posterior spur.

Key words. Hemidactylus brookii, complex, taxonomy, bPTP, multivariate analysis, DNA

Citation: Mirza ZA, Raju D. 2017. A new rupicolous species of gecko of the genus Hemidactylus Oken, 1817 from the Satpura Hills, Central India.

Amphibian & Reptile Conservation 11(1) [General Section]: 51-71 (e137).

NoDerivatives 4.0 International License, which permits unrestricted use for non-commercial and education purposes only, in any medium, provided

the original author and the official and authorized publication sources are recognized and properly credited. The official and authorized publication

credit sources, which will be duly enforced, are as follows: official journal title Amphibian & Reptile Conservation ; official journal website <amphibian-

reptile-conservation.org>.

Received: 27 May 2015; Accepted: 20 December 2016; Published: 24 March 2017

Introduction

The genus Hemidactylus Oken, is the second most spe-

ciose gekkonid genus in the world, with -143 species

distributed globally (Uetz and Hosek 2016), its diversity

being concentrated in the tropics. India is home to at least

29 species (including H. gleadowi Murray) and this num¬

ber is likely to increase with further sampling (Giri 2008;

Giri and Bauer 2008; Mirza and Sanap 2014).

During an expedition that led to the discovery of Eu-

blepharis satpuraensis (Mirza, Sanap, Raju, Gawai, and

Ghadekar 2014), a species of Hemidactylus was collected

from Pachmarhi town. Superficially resembling members

of the Hemidactylus brookii complex, it could be sepa¬

rated from most members of the group by the presence of

enlarged subcaudal plates on the tail and other morpho¬

logical characters. Hemidactylus brookii Gray has had a

complicated taxonomic history and nearly all attempts to

resolve the group have thus far failed largely due to the

commensal nature of the species, and inadequate sam-

Correspondence. 3 snakeszeeshan@gmail. com

pling effort in terms of specimens and collection locali¬

ties, (Bauer et al. 2010a; Mahony 2011; Rosier and Glaw

2010) and a long list of synonyms that remained unad¬

dressed until recently. Mahony (2011) made an effort to

address the taxonomic status of several synonyms and

his compilation serves as a vital resource for members

of this group. However, a few issues remain, for exam¬

ple, the obscure identity of Hemidactylus brookii sensu

stricto, with Grays specimens being the only true exem¬

plars of the species, as pointed out by Kathriner et al.

(2014). Lajmi et al. (2016) presented a comprehensive

analysis of the group in India using molecular as well

as morphological data. However, due to lack of material

from Pakistan, the resolution of the entire species com¬

plex is incomplete. Despite the controversial taxonomic

nature of members of this group, there are several dis¬

tinct morphotypes within the complex (see Kathriner et

al. 2014; Lajmi et al. 2016; Mahony 2009) that need to

be addressed to help resolve the systematics of the entire

H. brookii complex.

Amphib. Reptile Conserv.

March 2017 | Volume 11 | Number 1 | el 37

Mirza and Raju

Following the key provided by Lajmi et al. (2016), the

specimens of Hemidactylus from Satpura Hills show af¬

finity to members of clade 4 /H. cf. murrayi. Multivariate

analysis and molecular data support the distinctiveness

of the specimens of Hemidactylus obtained from Satpura,

which we herein describe as a new species. A molecular

phylogenetic analysis based on 302 bp of cytochrome b

gene suggests that the new species is sister to H. treutleri

Mahony.

Material and Methods

Morphological and meristic data: Specimens in the

field were captured by hand and euthanized, followed

by fixation in 4% formaldehyde buffer. The specimens

were later washed to remove traces of formalin, stored in

70% ethanol, and deposited in the collection of the Na¬

tional Centre for Biological Sciences, Bangalore, India

and the Bombay Natural History Society, Mumbai, India.

All measurements were taken following Giri and Bauer

(2008) with Mitutoyo™ digital calipers (to the nearest

0.1 mm): snout-vent length (SVL: from tip of snout to

vent), trunk length (TRL: distance from axilla to groin

measured from posterior edge of forelimb insertion to

anterior edge of hind limb insertion), body width (BW:

maximum width of body), crus length (CL: from base

of heel to knee); tail length (TL: from vent to tip of tail),

tail width (TW: measured at widest point of tail); head

length (HL: distance between retroarticular process of

jaw and snout-tip), head width (HW: maximum width of

head), head height (HH: maximum height of head, from

occiput to underside of jaws), forearm length (FL: from

base of palm to elbow); ear length (EL: longest dimen¬

sion of ear); orbital diameter (OD: greatest diameter of

orbit), nares to eye distance (NE: distance between ante-

riormost point of eye and nostril), snout to eye distance

(SE: distance between anteriormost point of eye and tip

of snout), eye to ear distance (EE: distance from anterior

edge of ear opening to posterior corner of eye), inter-

narial distance (IN: distance between nares), interorbital

distance (IO: shortest distance between left and right su-

praciliary scale rows) [Table 1], Morphological and mor-

Table 1. Morphometric and mensural data for Hemidactylus chipkali sp. nov.

Specimen

number

Holotype

NCBSAT107

Paratype

NCBS ATI 08

Paratype

NCBS ATI 09

Paratype

BNHS 2427

Paratype

BNHS 2426

Sex

SVL

74.2

65.6

60.1

61.7

54.3

TRL

26.7

24.1

25.8

23.7

11.7

13.9

12.0

12.5

10.4

10.8

11.2

10.5

11.7

10.1

59.4

60.4

37.6*

70.7

50*

5.4

5.9

7.7

6.6

13.4

16.7

18.4

17.5

12.5

12.2

12.4

11.5

10.9

6.5

5.9

7.1

5.5

5.4

8.9

9.2

10.3

9.4

8.5

4.0

3.5

3.2

3.3

3.6

5.5

5.8

6.3

5.0

5.3

7.2

7.9

6.4

6.8

4.4

5.0

4.9

4.8

4.3

1.2

1.3

1.2

1.4

1.5

1.2

4.1

4.5

4.3

4.0

4.1

Lamellae L manus

5-6-7-7-6

5-1-1-1-8

5-1-1-1-1

Lamellae R manus

5-1-1-1-1

5-1-1-1-8

5-1-1-1-1

Lamellae L pes

5-1-1-1-1

5-8-8-8-1

5-1-8-1-6

5-1-8-1-1

Lamellae R pes

5-7-8-7-6

5-8-8-1-1

5-1-8-1-1

5-8-8-1-1

Supralabials Left

Supralabials R

Infralabials L

Infralabials R

Pores L/R

7/7

8/8

gap between pores

Amphib. Reptile Conserv.

March 2017 | Volume 11 | Number 1 | el 37

A new gecko of the genus Hemidactylus

phometric data for Hemidactylus brookii group was ob¬

tained from, Lajmi et al. (2016), Mahony (2011), Rosier

and Glaw (2010). Principal Component Analysis (PCA)

was performed to further support the distinctiveness of

the new species. Meristic counts and external observa¬

tions of morphology were made using a LeicaTM S8A-

PO dissecting microscope. Images of the specimens were

taken with a CanonTM 70D mounted with a CanonTM

100 mm macro illuminated with two external CanonTM

430EX-II flashes, and plates were edited in Adobe® Pho¬

toshop CS5 (http://www.adobe.com/legal/permissions/

trademarks.html). Institutional acronyms used in the

manuscript are as follows: BNHS (Bombay Natural His¬

tory Society), Mumbai; NHM (Natural History Museum

London); NCBS (Collection facility, National Centre for

Biological Sciences), Bangalore; ZSI (Zoological Survey

of India), Kolkata.

Molecular methods and analysis: Genomic DNA for

a single specimen was extracted from tail tissue using

the Phenol-Chloroform-Isoamyl Alcohol method, fol¬

lowing Sambrook et al. (1989). Partial mitochondrial cy¬

tochrome b gene was amplified using primer CytbF700

(5’-CTTCCAACACCAYCAAACATCTCAGCAT-

GATGAAA-3’) and CytbR700 (5’-ACTGTAGCCCCT-

CAGAATGATATTTGTCCTCA-3 , ) published by Bauer

et al. (2007). Polymerase Chain Reaction protocols were

as followed by Mirza and Patel (2017). The PCR product

was cleaned, and sequenced with a 3730 DNA Analyzer

after cleaning. The sequence was cleaned manually in

MEGA7 (Kumar et al. 2016). In order to ascertain phylo¬

genetic position of the new species, published sequences

were retrieved from GenBank used by Lajmi et al. (2016)

listed in Appendix I. Sequences were aligned in Mega7

using ClustalW (Thompson and Gibson 2002) with de¬

fault settings. For optimal partitioning strategy and evo¬

lutionary substitution model, aligned data was analyzed

using PartitionFinder v. 1.1.1 (Lanfear et al. 2012). Maxi¬

mum Likelihood method was implemented to assess

phylogenetic relationship with RAxML (Silvestro and

Michalak 2012). Data were partitioned into three codons

and GTR+G was used as the sequence substitution mod¬

el, based on the optimal partitioning scheme suggested

by PartitionFinder. Maximum likelihood analysis was

run for 1,000 non-parametric bootstrap replicates with

rapid ML search option. Sequence divergence uncor¬

rected “p-distance” was calculated in Mega7. Sequence

for the new species has been deposited with GenBank

accession number “KX044190” for the specimen NCBS

ATI 10.

Species delimitation: Bayesian Poisson Tree Process

(bPTP) based on evolutionary placement algorithm was

implemented using the web server (http://species.h-hs.

org/ptp/) following Zhang et al. (2013) for inferring pu¬

tative species. Maximum likelihood tree was supplied for

the analysis. Outgroup, Hemidactylus frenatus , was ex¬

cluded from the analysis for optimum results. The analy¬

sis was run for 100,000 generations with three chains and

25% of the trees were discarded as burn-in. Results of the

analysis are presented in Appendix III and Appendix IV.

Systematics

Hemidactylus chipkali sp. nov.

Fig. 1-5, Table 1.

urn:lsid:zoobank.org:act:EB61DAC6-B9D6-41C7-862F-09B500778187

Holotype: NCBS AT 107, adult male, from a cliff along

the road leading to Pachmarhi town, Hoshangabad Dis¬

trict, Madhya Pradesh (22.485050°, 78.449340°, 1,092

m). Collected on 09 May 2014 by Rajesh Sanap, David

Raju, and Zeeshan Mirza.

Paratypes (four specimens): NCBS AT 109 and BNHS

2426, adult males; NCBS AT 108 and BNHS 2427, adult

females, same data as holotype.

Diagnosis: A moderate sized species of the genus mea¬

suring 54.3-74.2 mm; TRL/SVL 36^43.6%; HL/SVL

26-31%; ear opening oval; anterior postmental width

equal to first infralabial; posterior postmental width

equal to second infralabial, posterior postmental not in

contact with first infralabial; enlarged, keeled, tubercles,

fairly regularly arranged in 15-16 longitudinal rows on

dorsum; two angular series of seven precloacal femoral

pores on each side separated by diastema of eight noil-

pored scales; non-pored scales equal to size of pored

scales; scales bordering anterior edge of pored scales

half the size of pored scales; five lamellae on digit I and

seven (rarely eight) on digit IV of manus as well as pes;

lamellae on digit IV and V of pes absent on basal -25%

of the digit; limbs long and slender FL/SVL 0.15 and CL/

SVL 0.18; ventral aspect of tail with broad caudal scales

covering -80% of the tail; two subconical post cloacal

spurs, anterior spur slightly larger than the posterior spur.

Etymology: The specific epithet “ chipkali ” is the Hindi

word for gecko.

Description of holotype male NCBS AT107: Holotype

in good condition preserved in a linear manner with a

slightly curved tail. Hemipenis partly everted. Over half

of the tail regenerated (Fig. 1A, B).

A medium sized gecko (SVL 59 mm) with a fairly

large head (HL/SVL ratio 0.23), head slightly longer than

wide (HW/HL ratio 0.92), head slightly depressed (HH/

HL ratio 0.48), distinct from neck (Fig. 2A); canthus ros-

tralis slightly inflated; snout short (SE/HW ratio 0.57),

obtusely pointed from dorsal view and acutely in lateral

view (Fig. 2B), longer than eye diameter (OD/SE ratio

0.55); scales on the snout subequal, convex, those an¬

terior to the eye and on canthus rostralis, larger than the

surrounding scales; eyes large (OD/HL ratio 0.29), pupil

vertical with crenulated edges; supraciliaries smaller on

Amphib. Reptile Conserv.

March 2017 | Volume 11 | Number 1 | el 37

Mirza and Raju

Fig. 1. Hemidactylus chipkali sp. nov. male holotype NCBS AT 107, (A) dorsal view, (b) ventral view.

the anterior edge of the orbit, gradually increasing in size

as they progress towards upper surface of the head; ear¬

opening large, sub-oval, obliquely oriented, its length at

its greatest extent thrice that of the orbital diameter (EL/

OD ratio 0.30) bearing three lobules on its anterior in¬

ner wall; eye to ear distance greater than diameter of eye

(EE/OD ratio 1.11); rostral quadrangle, much wider than

deep, divided by a median suture for its entire length;

rostral in contact with nasal, first supralabial and interna¬

sals; two large and a slightly smaller internasals between

nasals; mental triangular, wider (3.2) than long (2.8); two

pairs of postmentals, anterior postmental longer (2.6)

than wide (1.8); posterior pair of postmental slightly

smaller than anterior pair, longer (2.3) than wide (1.4);

anterior postmental in contact with mental, infralabials

and posterior pair of postmental; posterior postmentals

separated by five scales; anterior postmental equal in

width to the first infralabial; posterior postmental equal

to width of second infralabials (Fig. 2C); scales on throat

circular, smaller than the ones ventral aspect of trunk;

supralabials (to midorbital position) nine on left and ten

on right side; supralabials (to angle of jaw) eleven on left

side and twelve on right side; infralabials (to angle of

jaw) nine on left and ten on right side.

Body elongate (TRL/SVL ratio 0.45) and dorsoven-

trally flattened, more so after preservation; lacking dis¬

tinct ventrolateral furrow; dorsal scalation on trunk gran¬

ular intermixed with enlarged, keeled, tubercles, fairly

regularly arranged in 15-16 longitudinal rows; dorsal

tubercles on mid-dorsum longer (1.0) than wide (0.9);

individual tubercle row separated from the adjacent by

three transverse scale rows and by preceding tubercle of

the same row, by four scale rows (Fig. 3A); ventral scales

on trunk smooth, flat, larger than dorsal scales; mid body

scales across belly 28-30 (Fig. IB); eight (left) and seven

(right) femoral pores separated at mid-pelvic region by

eight non-pored scales; non-pored scales slightly larger

than pored scales (Fig. 3B).

Limbs moderately long, slender; digits dilated, bear¬

ing slightly oblique lamellae on ventral surface; clawed,

claw nearly half the length of the lamellar region; fore¬

limbs short (FL/SVL ratio 0.15), slightly shorter than

hind limbs (CL/SVL ratio 0.18), all digits of manus and

digits I-IV of pes indistinctly webbed at the base. Termi¬

nal phalanx of all digits curved, arising angularly from

distal portion of expanded lamellar pad, free portion of

Amphib. Reptile Conserv.

March 2017 | Volume 11 | Number 1 | el 37

A new gecko of the genus Hemidactylus

Fig. 2. Hemidactylus chipkali sp. nov. male holotype N.CBS

AT 107 head, (A) dorsal view, (B) lateral view, (C) ventral view.

phalanx of all digits half to more than half long as the

dilated portion. Lamellae beneath the digits, left manus

5-6-7-7-6, right manus and left pes 5-7-7-7-7 (Fig. 4A),

right pes 5-7-S-7-6 (Fig. 4B). Lamellae not reaching the

base of the digit IV of pes. Relative lengths of digits:

III>IV=V>II>I (right manus), IV>H=III>V>I (right pes).

Tail moderately depressed, oval in cross section, longer

than snout-vent length (TL/SVL ratio 1.26), 59.4 mm of

the tail regenerated. Caudal segments distinct; pholido-

sis of original tail dorsum with small, juxtaposed scales

intermixed with large depressed keeled tubercles, scales

on regenerated portion of tail with slightly larger scales

and lacking tubercles. First tail segment with a whorl of

ten large conical, keeled tubercles, second segment on¬

wards, each segment with six tubercles. Ventral aspect

with large, broad scales covering about -80% of the tail

width from base of tail to the tip (Fig. 3C). Two subconi-

cal post cloacal spurs, anterior spur slightly larger than

the posterior spur.

Coloration in life (Fig. 5): Coloration is a shade of

pale brown with white and dark brown spots across the

dorsum. Three adjoining brown spots over the nape and

similar spots at a fairly regular distance from the preced-

Fig. 3. Hemidactylus chipkali sp. nov. male holotype NCBS

AT 107, (A) dorsal view of trunk, (B) ventral view of cloacal

showing precloacal femoral pores and hemipenis, (C) ventral

view of tail showing enlarged sub-caudal scales.

ing row all through the trunk up to the flank. Each dark

band is separated from the subsequent one by loosely

connected white spots, to form thin white bands. Limbs

pale brown with whitish spots ah over the dorsum. Tail

with alternate pale brown and cream colored bands. Col¬

oration in preservative (Fig. 6): Specimens paler than in

life with vestigial remains of dark brown spots and little

to no trace of white spots on dorsum. Underparts straw

colored.

Natural History: The new species was collected from

vertical cliffs along a road leading to Pachmarhi situ¬

ated in the Satpura Hills of central India (Fig. 7). Satpura

Hills are located south of the Narmada River, running

parallel to the river from western Gujarat through the

borders of Maharashtra and Madhya Pradesh, extending

up to northeastern Madhya Pradesh. The landscape at the

type locality is undulating with the highest peak reaching

1,350 m and is dominated by tree species like Tectona

grandis and Shorea robusta , characteristic of deciduous

forests in the region. The hills at the type locality have

steep cliffs where the new species was found (Fig. 8).

Ah the type specimens and a few additional uncollected

specimens were found actively moving on roadside rocks

(Fig. 9). The species was observed to be sympatric with

Eiiblepharis satpuraensis , which likely shares the crev¬

ices in the cliff' during the day and emerges at dusk. Most

individuals would dart towards the nearest crevice when

Amphib. Reptile Conserv.

March 2017 | Volume 11 | Number 1 | el 37

Mirza and Raju

Fig. 4. Hemidactylus chipkali sp. nov. male holotype NCBS AT 107 (A) right manus, (B) right pes.

approached with flashlights. The new species is likely re¬

stricted to these high elevation cliff's as only H. cf. glead¬

owi was recorded from near Bijakhori village, Sohagpur

ca. 30 km (aerial distance, elevation 367 m) northwest

of the type locality. One of the female paratypes (NCBS

AT 108) shows presence of two well-developed eggs

within the body cavity suggesting that the species breed

during the summer. With regards to its IUCN status, we

recommend designating this species as “Data Deficient”

in terms of its distribution, until further reports surface.

During the course of the survey, no immediate threat to

the species was observed and the entire area where the

species was observed is protected as part of the Satpura

Tiger Reserve. However, large parts of the Satpura hills

are not protected and further surveys must be conducted

to assess conservation threats to the species outside pro¬

tected areas.

Suggested common name: Central Indian Leaf-toed

Gecko.

Comparison: Hemidactylus chipkali sp. nov. differs

from most Indian congeners in bearing the following set

of differing and non-overlapping characters: SVL 54.3-

74.2 mm (vs. SVL > 80 mm in H. maculatus Dumeril

and Bibron, H. graniticolns Agarwal, Giri, and Bauer, H.

giganteus Stoliczka, H. gujaratensis Giri, Bauer, Vyas,

and Patil, H. prashadi Smith, H. acanthopholis Mirza

and Sanap, H. aaronbaueri Giri, H. yajurvedi Murthy,

Bauer, Lajmi, Agarwal, and Giri), dorsum with keeled

tubercles fairly arranged in 15-16 longitudinal rows (vs.

tubercles absent or few or irregularly arranged in H. aq-

uilonius Zug and Mcmahan, H. flaviviridis Riippell, H.

frenatus Schlegel, H. garnotii Dumeril and Bibron, H.

leschenaultii Dumeril and Bibron, H. hemchandrai Dan-

dge and Tiple), presence of seven femoral pores sepa¬

rated medially by a diastema of eight non-pored scales

(vs. precloacal pores in H. sataraensis Giri and Bauer, H.

gracilis Blanford, H. reticulatus Beddome, H. albofas-

ciatus Grandison and Soman, H. scabriceps Annandale,

H. persicus Anderson, H. robustus Heyden, H. turcicus

Linnaeus), dorsal tubercles sub-trihedral (vs. tubercles

trihedral in H. trie dr us Daudin and H. sub trie dr us Jer-

don), lacking webbing on hind feet and a fringe of skin

on lateral aspect of tail (vs. present in H. platyurus

Schneider), lamellae divided (undivided in H. anamal-

lensis Gunther).

Hemidactylus chipkali sp. nov. is most similar to

members of the Hemidactylus brookii group and is here

compared with taxa considered valid by Mahony (2011)

and Kathriner et al. (2014). Hemidactylus chipkali sp.

nov. differs from H. brookii (as defined by Mahony 2011)

in larger SVL 74 mm (vs. SVL 55.8 mm in H. brookii ,

SVL 43.1 mm in H. gleadowi Murray, SVL 45-51.4 mm

in H. kushmorensis Murray, 51.5 mm in H. parvimacu¬

latus Deraniyagala); anterior postmental width equal to

first infralabial; posterior postmental width equal to sec¬

ond infralabial, posterior postmental not in contact with

first infralabial or with narrow contact Figs. 10A-E (vs.

width of posterior postmental shorter than width of sec¬

ond infralabial in H. brookii , width of posterior postmen¬

tal shorter than width of second infralabial, posterior in

contact with first infralabial in H. gleadowi , anterior, pos¬

terior postmental in broad contact with first infralabial

in H. kushmorensis , width of anterior postmental longer

than first infralabial, and anterior postmental in contact

with first and second infralabials in H. trenderi Mahony);

seven precloacal femoral pores separated medially by a

Amphib. Reptile Conserv.

March 2017 | Volume 11 | Number 1 | el 37

A new gecko of the genus Hemidactylus

Fig. 5. Hemidactylus chipkali sp. nov. (A and B) male holotype NCBS AT 107 in life, (C) male paratype NCBS AT 108 in life.

Fig. 6. Dorsal aspect of the type series showing coloration after preservation.

Amphib. Reptile Conserv.

March 2017 | Volume 11 | Number 1 | el 37

Mirza and Raju

Fig. 7. Map of Madhya Pradesh, central India, showing the type locality Pachmarhi (indicated by a red triangle). Inset map shows

location of Madhya Pradesh in India.

Fig. 8. Biotope of Satpura hills showing characteristic rocky cliffs and forest cover where the new species was collected.

Amphib. Reptile Conserv.

March 2017 | Volume 11 | Number 1 | el 37

A new gecko of the genus Hemidactylus

Fig. 9. Rocks along the road leading to Pachmarhi town where the type specimens were collected.

Fig. 10. Ventral view of head, (A) H. cf. gleadowi NCBS HA- 107, (B) H. cf. murrayi NCBS HA- 110, (C) H. subtriedroides lecto-

type NHM 1946.8.2554, (D) H. treutleri holotype ZSI 25711, (E) H. chipkali sp. nov.

diastem of eight non-pored scales Figs. 11A-E(vs. 12-13

precloacal femoral pores separated medially by a diastem

of one non-pored scale in H. brookii and H. gleadowi,

10-11 precloacal femoral pores separated by 2-3 non-

pored scales in H. kushmorensis, 12-15 pores separated

by 2-4 non-pored scales in H. parvimaculatus, 7-8 sepa¬

rated by 5-6 non-pored scales in H. subtriedroides (An-

nandale); lamellae series on digit IV do not extend to base

Figs. 12A-C (vs. lamellae series cover the entire digit IV

in H. brookii, H. cf. murrayi Gleadow, H. subtriedroides,

H. tenkatei Lidth de Jeude, H. treutleri and H. kushmo-

rensis ); scales bordering anteriorly the precloacal pores

half the size of pored scales (vs. scales bordering anteri¬

orly the precloacal pores > the size of pored scales in H.

brookii ); keeled dorsal tubercles in 15-16 fairly longitu¬

dinal rows (vs. 19-20 in H. kushmorensis)', five lamellae

on digit I of pes (vs. 6-7 in H. treutleri)', lamellae on

digit IV of pes 7 rarely 8 (vs. 10 in H. kushmorensis, 8

in H. cf. murrayi, 9 in H treutleri, 11 in H. mahendrai

Shukla); caudal plates enlarged and cover -80% of the

underside of the tail (vs. tail plates not covering entire

tail in H. gleadowi, H. kushmorensis, H. subtriedroides,

H. tenkatei); two sub-conical postcloacal spurs, anteri¬

or one slight larger than the posterior (vs. 2-3 medium

sized with or without an additional large spur in H. sub¬

triedroides and H. cf. murrayi, two very small ones in H.

gleadowi, a single domed spur in H. kushmorensis, three

enlarged spurs in H. treutleri)', sub-caudal completely

transverse the tail width (sub-caudal not as enlarged as

in H. gleadowi, H. kushmorensis, H. subtriedroides, H.

tenkatei, H. brookii, H. cf. murrayi), FL/SVL 0.15 (vs.

0.13 in, H. kushmorensis, H. subtriedroides, H. brookii,

0.12 in H. gleadowi, H. cf. murrayi)', CF/SV1 0.18 (vs.

0.15 in H. brooki, H. gleadowi, 0.14 in//, kushmorensis,

0.16 in H. tenkatei, H. subtriedroides).

Amphib. Reptile Conserv.

March 2017 | Volume 11 | Number 1 | el 37

Mirza and Raju

Discussion and Conclusion

Phylogenetic relationships within Indian and South

Asian Hemidactylus have been the subject of recent stud¬

ies (Bansal and Karanth 2010; Bauer et al. 2010b). How¬

ever, due to lack of extensive sampling in most studies,

the H. brookii complex remained unresolved and still

does. Kathriner et al. (2014) provided new insights on

Fig. 11. Precloacal and or femoral pores, (A) H. cf. gleadowi

NCBS HA-107, (B) H. cf. murrayi NCBS HA-110, (C) H. sub-

triedroides lectotype NHM 1946.8.2554, (D) H. treutleri holo-

type ZSI 25711, (E) H. chipkali sp. nov.

the systematics of the H. brookii complex, which raises

doubts on previous assumptions and results based solely

on morphology. Lajmi et al. (2016) presented the first

ever comprehensive analysis of H. brookii complex in¬

corporating molecular as well as morphological data

which has enhanced our understanding of this group thus

enabling us to present preliminary data on phylogenetic

relationship of the new species. Based on a short frag¬

ment of -302 bp of mitochondrial cyt b gene, the new

species appears to be allied to H. treutleri (Fig. 13) and

is a member of clade 5 of Lajmi et al. (2016). It however

differs in having an uncorrected p-distance of 14-16%

(Fig. 14, Table 4). The phylogenetic analysis results pre¬

sented here are preliminary, based on data generated by

Lajmi et al. (2016). Publication by Lajmi et al. (2016)

contains 30 accession numbers, which are repeats and is

likely a typographical error. The authors have not copied

accession number correctly from previous studies and

have the same accession numbers for several sequences

of specimens from different localities. It is hoped that the

authors provide correct accession numbers and their re¬

spective voucher details in a subsequent paper.

Relationships recovered from molecular phyloge¬

netics shows discordance with morphology in the new

species, showing close affinity to H. cf. murrayi based

on morphology whereas it shows affinity to H. treutleri

based on molecular data. The new species however dif¬

fers from H. treutleri and H. cf. murrayi in having broad

sub-caudals transverse the entire width of the tail and in

bearing 7 lamellae on fourth toe vs. 8 in H. cf. murrayi

and 9 in H. treutleri. Principal Component Analysis of

data including morphometric data for the new species,

H. treutleri and H. cf. murrayi for standardized morpho¬

metric data showed PCI + PC2 explaining 80% + 16%

of variance, respectively (Fig. 14, Appendix II). Plot of

the first two principal axes resulted in two clusters; one

of H. chipkali and another one of H. treutleri and H. cf.

murrayi (Fig. 15). Results from bPTP support the dis¬

tinctiveness of the species with high support (Appendix

III and IV).

Hemidactylus murrayi Gleadow, 1887 was described

based on a series of 24 specimens from “Pimpri and Gar-

vi, in the Dangs” in southern Gujarat. The types are like-

Fig. 12. Ventral aspect of right pes, (A) H. cf. gleadowi NCBS HA-107 note lamellae on digit IV not reaching base, (B) H. cf. mur¬

rayi NCBS HA-110 lamellae covering entire digit IV, (C) H. chipkali sp. nov. Note lamellae on digit IV not reaching base.

Amphib. Reptile Conserv.

March 2017 | Volume 11 | Number 1 | el 37

A new gecko of the genus Hemidactylus

it?:

-| 1 00

r ti

H 3(1

■ 5 —

-[l 00

HM 595647

HM595646

KU720650

KU720649

KU720652

K U 72 06 4 8

KU720651

KU720640

K U 7 2 0 6 4 1

KU720644

KU720645

KU 7 2064 7

KU720646

GQ375298

GQ3753Q0

GQ3 7529 1

5 b GQ375296

3 . GQ375297

3 2 GQ375299

2 GQ37529 2

KU720637

DQ120271

0 DQ1 20272

HM595 6 45

KU720638

, GQ37529Q

*- DQ 1 20273

KU720643

KU720642

KU720639

KU720654

K U 7 2 0 6 5 3

KX044190

KU720681

KU720679

KU720677

KU720678

EU2684Q7

GQ375294

GQ375293

EU268397

KU720667

KU720666

KU720676

HM595 649

KU720674

KU720673

HM595659

EU268385

EU268386

HM595 6 42

HM595643

H M595670

HM595671

H M 5 9 5 6 6 9

EU268410

H M 5 9 5 6 7 2

HM 5 9 5 6 6 0

KU 7 20656

KU720657

KU720659

KU720660

KU720661

KU720662

KU720658

EU268391

Hemidactylus cf. kushmorensis

Hemidactylus parvimaculatus

Hemidactylus cf. kushmorensis

■ Hemidactylus chipkali sp. nov.

Hemidactylus treuHeri

Hemidactylus ct’ murrayi

| Hemidactylus imbricatus

Hemidactylus albofasciatus

Hemidactylus reticulatus

■ Hemidactylus saturaensis

■ Hemidactylus gracilis

Hemidactylus cf* g/eadowi

I Hemidactylus fremitus

Fig. 13. Maximum likelihood tree for selected members of the H. brookii group showing relationship of H. chipkali sp. nov. rooted

with H. frenatus as outgroup based on -302 bp of mitochondrial cytochrome b gene with 1,000 non-parametric bootstrap replicates.

Numbers at nodes indicate bootstrap support.

Fig. 14. PCAplot for standardized morphometric data for H. chipkali sp. nov. (black), H. cf. murrayi (red) and H. treutleri (blue).

Circles = male and squares = female.

Amphib. Reptile Conserv.

March 2017 | Volume 11 | Number 1 | el 37

Mirza and Raju

ly lost and hence the identity of the species can only be

ascertained after examination of material from both the

said localities. Lajmi et al. (2016), who considered clade

4 of their work as H. murrayi, however do not include

molecular data from either of the localities from where

types of H. murrayi were collected. Hemidactylus ten-

katei was found nested in clade 4 of Lajmi et al. (2016),

suggesting that this clade might represent more than one

species. To further support this, a sample from Nashik

(GenBank accession number KU720676) in clade 4 of

Lajmi et al. (2016) shows ~8% sequence divergence for

cytochrome b from other members of the clade which

may represent yet another undescribed taxon (Appendix

V). Results from bPTP suggests that H. trenderi likely

is a species complex (Appendix III and IV). In order to

resolve this group, a neotype for H. murrayi and H. ma-

hendrai , each, must be designated and molecular data for

the neotypes/topotypes must be incorporated in a phy-

logeny, and also molecular data from the type locality

of H. subtriedroides Annandale (suggested to be a valid

species by Kathriner et al. 2014), which will shed light

on the systematics of this clade that we refer to as H. cf.

murrayi in the present work. Considering that members

of the H. cf. murrayi clade are commensal, it may not be

an easy task to resolve this complex, largely due to mul¬

tiple back and forth colonizations in recent years through

human agencies.

Description of yet another member of the H. brookii

complex highlights the diversity of the H. brookii com¬

plex in India. Including the present description, at least

seven species of the H. brookii complex have been re¬

corded from India including H. mahendrai (see Lajmi et

al. 2016). India and Sri Lanka are home to several en¬

demic species of Hemidactylus (Bauer et al. 2010b). This

number is steadily increasing with dedicated surveys, as

well as re-examination of museum specimens (Agarwal

et al. 2011; Mirza and Sanap 2014). Explorations of iso¬

lated and/or unexplored hill ranges like the Satpura hills,

Aravalli hills, and other small hills across the country

would certainly harbor undocumented species of rep¬

tiles as demonstrated by the discoveries of Cyrtodacty-

lus srilekhae , C. rishivalleyensis, C. varadgirii (Agarwal

2016, Agarwal et al. 2016), Eublepharis satpuraensis

(Mirza et al. 2014), H. chipkali sp. nov., H. sataraensis

(Giri and Bauer 2008), and Wallaceophis gujaratensis

(Mirza et al. 2016). Our finding further attests the poor

nature of reptilian documentation in the country and the

lack of taxonomic revisions on most reptilian groups (see

Gowande et al. 2016, Mirza and Sanap 2014, Mirza et

al. 2010).

Acknowledgments .—This herpetofaunal documenta¬

tion project would not have been possible without neces¬

sary permits granted by the forest department of Madhya

Pradesh for which we are grateful to Narendra Kumar

(PCCF, Wildlife and Chief Wildlife Warden), Ravi Shriv-

astava (PCCF Wildlife), R.P Singh (APCCF Wildlife),

Anil Nagar (Field Director, Satpura Tiger Reserve), and

Dr. Suhas Kumar for necessary pennissions to carry out

research in Madhya Pradesh. We thank Tulika Kedia and

Singinawa Conservation Foundation for all their help and

support. We also thank Forsyth Fodge for logistic sup¬

port and permission to conduct surveys on their property.

Rahul Khot (BNHS, Mumbai), Kaushik Deuti (ZSI, Kol-

kata), and Patrick Campbell (NHM, Fondon) helped with

access to type specimens. Special thanks goes to Anurag

Mishra (NCBS) for reviewing the final draft of the man¬

uscript. Harshil Patel provided valuable input on H. cf.

murrayi for which we kindly acknowledge him. Rajesh

Sanap and Surya Ramachandran in addition helped with

fieldwork. Krushnamegh Kunte granted permission and

access to lab facilities at the National Centre for Biologi¬

cal Sciences. Fieldwork was supported by a generous

grant from the Rufford foundation to ZM. Special thanks

to Robin K. Abraham for constructive comments on the

final draft of this paper. Finally, I. Das and Ishan Agarwal

provided valuable comments from which the manuscript

greatly benefitted.

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Zeeshan A. Mirza is an independent researcher presently pursuing a Master’s degree in zoology from Mum¬

bai University. He has largely been interested in the taxonomy of snakes, lizards, and arachnids. Zeeshan has

published several papers on their taxonomy including descriptions of over thirty new species. He received a

Bachelor’s degree in zoology from Bhavans College, Mumbai University. He was bom in Mumbai city and

has worked largely on the city’s reptilian diversity and has worked on several projects to document reptiles

and arachnids of Western Ghats. He plans to pursue his further studies on systematics of reptiles and arach¬

nids with an integrated approach involving molecular and morphological data.

David V. Raju comes from a small village called Kuzhimattom in Kottayam District of Kerala, India. He

graduated from Baselius College Kottayam in English Literature. David has always had an interest in wild¬

life and further developed his interest at the Kottayam Nature Society. Currently he is working as a naturalist

in Central India. He has co-authored a book on the dragonflies of Kerala. He was also a part of the team

which discovered ten frogs in Western Ghats and a leopard gecko from Satpura hills. His other interests are

birds, butterflies, and mammals.

Amphib. Reptile Conserv.

March 2017 | Volume 11 | Number 1 | el 37

Mirza and Raju

Comperative material examined:

Hemidactylus aaronbaueri: Holotype BNHS 1739 (male),

Ghatghar, Taluka Junnar, Pune District, Maharashtra, India;

ZSI21648A and ZSI21648C (female), ZSI21648B (male),

Bhairavgadh Fort, Taluka Karjat, Pune District, Maharash¬

tra, India.

Hemidactylus acanthopholis: Flolotype NFIM 1946.8.23.68

(male), paratypes NHM 1946.8.2367 (male), Tirunelveli

District, Tamil Nadu.

Hemidactylus albofasciatus: Paratype ZSI 21109 (female),

Dorle village, Rajapur Taluka, Ratnagiri District, Maharash¬

tra, India; two males BNFIS 1579 and 1582 Dabhil-Ambere,

Ratnagiri District; Maharashtra, India.

Hemidactylus bengaliensis Anderson (=H. flaviviridis): Syn-

type ZSI 5780, Bengal.

Hemidactylus flaviviridis: ZSI 20963 (male) Jaipur, Rajasthan,

India; ZSI 21688 (female) Udaipur, Rajasthan, India.

Hemidactylus hunae: Type specimen NHM 1946.8.23.77 (fe¬

male), Okanda, Eastern Province, Sri Lanka.

Hemidactylus giganteus: Syntype, NHM 1877.8.6.5 (male),

Godavari valley near Bhadrachalam, Andhra Pradesh, India.

Hemidactylus cf. gleadowi: Male NCBS HA-107 Khamgaon,

Maharashtra; male NCBS HA-108 Sohagpur, Madhya

Pradesh.

Hemidactylus gracilis: Syntype, ZSI 5190 (male), from “S.E.

Berar” (in Madhya Pradesh, India; BNHS 1591 (male) and

BNHS 1592 (female), Chatushringi hills, Pune, Maharash¬

tra, India.

Hemidactylus graniticolus: Holotype BNHS 1850 (female),

hills near Harohalli, Bangalore Rural district, Karnata¬

ka, India; paratypes NHM 1946.8.23.70 (female), NHM

1946.8.23.71 (female), NHM 1946.8.23.72 (male), NHM

1946.8.23.73 (female), NHM 19.46.8.23.74 (female), NHM

1946.8.23.75 (female), Salem District, Tamil Nadu, India;

NHM 1946.8.23.76 (male), “Malabar,” India.

Hemidactylus gujaratensis: Holotype BNHS 1818 (female) Va-

gheshwari Mata Temple, Junagadh City, Junagadh District,

Gujarat, India.

Hemidactylus kelaartii: Syntypes ZSI 2617 (male) and ZSI

2618 (female), from “Ceylon” (= Sri Lanka).

Hemidactylus maculatus: NHM 1956.1.11.41 (female), Ma-

theran, Raigad District, Maharashtra, India; ZSI 25608

(male) Government rest house, Panchagani, Satara District,

Maharashtra, India; BNHS 74 (female) and BNHS 75 (fe¬

male), Mumbai, Maharashtra, India; BNHS 1086 (male),

Kanheri caves, Mumbai, Maharashtra, India.

Hemidactylus marmoratus (=Hemidactylus leschenaulti): Ho¬

lotype, ZSI 5058, from “S.E. Berar, near Chanda” Maha¬

rashtra, India.

Hemidactylus cf. murrayi: BNHS 1947-1948 (males), BNHS

1949 (female), Aarey Milk Colony, Mumbai, Maharashtra.

Hemidactylus persicus: Holotype, ZSI 5961, from “Persia” (=

Iran). The register lists the type as from “Shiraz, Persia.”

Hemidactylus platyceps (=Hemidactylus gracilis ): Holotype,

ZSI 17020, from “Bilimora, Bombay Presidency” Gujarat,

India.

Hemidactylus prashadi: BNHS 147 (male), Shiroli forest, Bel-

gaum North Kanara, Karnataka, India; BNHS 146 (male),

Gersoppa falls, North Kanara, Karnataka, India; ZSI 20123

(female) neighbourhood of Jog, North Kanara district,

Bombay Presidency’ (at present in Karnataka, India).

Hemidactylus reticulatus: Type specimens NHM 1874.4.29.410

(male) and NHM 1874.4.29.411 (female), Kollegal, Karna¬

taka, India.

Hemidactylus sataraensis: Holotype BNHS 1743 (female)

Chalakewadi, Satara District, Maharashtra, India; paratype

BNHS 1742 (female); non-type BNHS 2288 (male), BNHS

2289 (female), Chalakewadi, Satara District, Maharashtra,

India.

Hemidactylus scabriceps: Type specimens, ZSI 15353, from

“Ramanad,” Tamil Nadu, India.

Hemidactylus sp. (H. cf. maculatus/H. cf. subtriedrus ): ZSI

24155 (female), Bastar District, Chhattisgarh, India; ZSI

25866 (male) Tyda railway station, Tyda, Vishakapatnam

district, Andhra Pradesh, India; ZSI 25708 (male) Ganjam

district, Odisha, India; a large male without locality and reg¬

istration tag along with ZSI 25708.

Hemidactylus sykesii (=H. maculatus): Type specimen, NHM

XXII.20a (male), Deccan, India (Donated by Indian Mu¬

seum XXII. 20a).

Hemidactylus subtriedroides: Syntype, NHM 1946.8.25.54/

ZSI 4135, “Tsagain, Upper Burma.”

Hemidactylus treutleri: Holotype ZSI 25711 (male), paratype

ZSI 25712 (female), outer stone wall of Golconda Fort, Hy¬

derabad, Andhra Pradesh, India.

Hemidactylus triedrus: ZSI 17054 (female) Travindrum, Ker¬

ala, India; ZSI 5852, ZSI 5853 (males), Bangalore, Karna¬

taka, India; ZSI 21483, ZSI 21486 (males), Pune, Maha¬

rashtra, India.

Amphib. Reptile Conserv.

March 2017 | Volume 11 | Number 1 | el 37

A new gecko of the genus Hemidactylus

Appendix I. List of species and their sequence accession numbers for the gene cytochrome b used in the present study.

Species

Locality

Accession number

Hemidactylus albofasciatus

Dorle, Ratnagiri, Maharashtra, India

HM595642

Hemidactylus albofasciatus

Malvan, Sindhudurg, Maharashtra, India

HM595643

Hemidactylus cf. gleadowi

Chikkabellapur, Karnataka, India

KU720656

Hemidactylus cf. gleadowi

Ranebennur, Karnataka, India

KU720657

Hemidactylus cf. gleadowi

Mysore, Karnataka, India

KU720658

Hemidactylus cf. gleadowi

Bagalkot, Karnataka, India

KU720659

Hemidactylus cf. gleadowi

Dapoli, Maharashtra, India

KU720660

Hemidactylus cf. gleadowi

Ahmednagar, Maharashtra, India

KU720661

Hemidactylus cf. gleadowi

Iqbalgadh, Gujarat, India

KU720662

Hemidactylus cf. kushmorensis

Reasi, Himachal Pradesh, India

KU720648

Hemidactylus cf. kushmorensis

Dehradun, Uttarakhand, India

HM595646

Hemidactylus cf. kushmorensis

Chamba, Himachal Pradesh, India

KU720649

Hemidactylus cf. kushmorensis

Kangra-Jawalamukhi Road, Himachal

Pradesh, India

KU720650

Hemidactylus cf. kushmorensis

Ajmer, Rajasthan, India

KU720651

Hemidactylus cf. kushmorensis

Baripada, Odisha, India

KU720652

Hemidactylus cf. kushmorensis

Jammu, India

HM595647

Hemidactylus cf. kushmorensis

Chotila, Gujarat, India

KU720653

Hemidactylus cf. kushmorensis

Mt. Abu, Rajasthan, India

KU720654

Hemidactylus cf. murrayi

Badlapur, Maharashtra, India

KU720666

Hemidactylus cf. murrayi

Mumbai, Maharashtra, India

KU720667

Hemidactylus cf. murrayi

Loagan Bunut National Park, Sarawak,

Malaysia

GQ375293

Hemidactylus cf. murrayi

Mandalay Division, Myanmar

EU268407

Hemidactylus cf. murrayi

Yangon, Myanmar

GQ375294

Hemidactylus cf. murrayi

Empangon Air Hitam, Pulau Pinang, Malaysia

EU268397

Hemidactylus cf. murrayi

Palakkad, Kerala, India

HM595649

Hemidactylus cf. murrayi

Malshej Ghat, Maharashtra, India

KU720673

Hemidactylus cf. murrayi

Junagadh, Gujarat, India

KU720674

Hemidactylus cf. murrayi

Nasik, Maharashtra, India

KU720676

Hemidactylus frenatus

Sri Lanka, Rathegala

EU268391

Hemidactylus gracilis

Pune, Maharahstra, India

HM595660

Hemidactylus gracilis

Kolhapur, Maharashtra, India

HM595659

Hemidactylus imbricatus

Pakistan (captive specimen)

EU268386

Hemidactylus imbricatus

Pakistan (captive specimen)

EU268385

Hemidactylus parvimaculatus

Gandagan, Odisha, India

KU720637

Hemidactylus parvimaculatus

Polupalli, Tamil Nadu, India

DQ120272

Hemidactylus parvimaculatus

Tumkur, Karnataka, India

HM595645

Hemidactylus parvimaculatus

Mauritius

DQ 120271

Hemidactylus pannmaculatus

Mampuri, Sri Lanka

GQ3 75292

Hemidactylus parvimaculatus

Dehikindagama, Sri Lanka

GQ375296

Hemidactylus parvimaculatus

Matale, Sri Lanka

GQ375298

Hemidactylus parvimaculatus

Gonaganara, Sri Lanka

GQ375297

Hemidactylus parvimaculatus

Kartivu, Sri Lanka

GQ375291

Hemidactylus parvimaculatus

Matale, Sri Lanka

GQ3 75299

Hemidactylus parvimaculatus

Tempitiya, Sri Lanka

GQ375300

Amphib. Reptile Conserv.

March 2017 | Volume 11 | Number 1 | el 37

Mirza and Raju

Appendix I (continued). List of species and their sequence accession numbers for the gene cytochrome b used in the present

study.

Species

Locality

Accession number

Hemidactyhis pan’imaculatus

Rushikulya, Odisha, India

KU720638

Hemidactyhis parvimaculatus

Attagulipura, Karnataka, India

KU720639

Hemidactylus pan’imaculatus

Bangalore, Karnataka, India

KU720640

Hemidactyhis pan’imaculatus

Chennai, Tamil Nadu, India

KU720641

Hemidactyhis parvimaculatus

Poinguinim, Goa, India

KU720642

Hemidactyhis pan’imaculatus

Modern, Goa, India

KU720643

Hemidactylus pan’imaculatus

Kutugam, Odisha, India

KU720644

Hemidactyhis pan’imaculatus

Araku Valley, Andhra Pradesh, India

KU720645

Hemidactylus pan’imaculatus

Vizianagaram, Andhra Pradesh, India

KU720646

Hemidactyhis pan’imaculatus

Majhiguda, Odisha, India

KU720647

Hemidactyhis parvimaculatus

Kollam, Kerala, India

DQ120273

Hemidactyhis pan’imaculatus

Kandy, Sri Lanka

GQ375290

Hemidactyhis reticulatus

Pavgada, Karnataka, India

HM595669

Hemidactyhis reticulatus

Nandi Hills, Karnataka, India

HM595670

Hemidactyhis reticulatus

Nandi Hills, Karnataka, India

HM595671

Hemidactyhis reticulatus

Vellore, Tamil Nadu, India

EU268410

Hemidactyhis sataraensis

Chalakewadi, Maharashtra, India

HM595672

Hemidactyhis treutleri

Hyderabad, Telangana, India

KU720681

Hemidactyhis cf. treutleri

Rishi valley, Andra Pradesh, India

KU720679

Hemidactylus cf. treutleri

Kangudi, Tamil Nadu, India

KU720678

Hemidactyhis cf. treutleri

Chikkabellapur, Karnataka, India

KU720677

Appendix II. PCA loadings for each character

PC 1

PC 2

SVL

0.2125

0.7975

TRL

0.0613

0.3992

0.0326

0.1582

0.0462

0.1035

0.9720

-0.2202

0.0158

0.1238

0.0123

0.1818

0.0286

0.1637

-0.0010

0.1013

0.0299

0.0967

0.0161

-0.0052

0.0002

0.0720

0.0032

0.0915

0.0032

0.0565

-0.0030

0.0209

-0.0020

0.0209

-0.0276

0.0740

Amphib. Reptile Conserv.

March 2017 | Volume 11 | Number 1 | el 37

A new gecko of the genus Hemidactylus

Appendix III. Results of species delimitation using bPTP based on ML tree. Numbers above nodes/tips represent posterior

delimitation probabilities from Bayesian reconstruction.

^ KU720658

0.10

Species 11: Hemidactylus cf. kushmorensis

Species 9: Hemidactylus parvimaculatus

| Species 10: Hemidactylus cf. kushmorensis

I Species 1: Hemidactylus chipkall sp. nov.

i Species 12: Hemidactylus treutleri (topotype)

I Species 13: Hemidactylus cf. treutleri

Species 14: Hemidactylus cf. treutleri

Species 2: Hemidactylus cf. murrayi

| Species 8: Hemidactylus imbricatus

| Species 7: Hemidactylus albofasciatus

Species 5: Hemidactylus reticulatus

I Species 6: Hemidactylus satarensis

I Species 3: Hemidactylus gracilis

Species 4: Hemidactylus cf. gleadowi

Amphib. Reptile Conserv.

March 2017 | Volume 11 | Number 1 | el 37

Mirza and Raju

Appendix IV. Results of bPTP showing support to each species.

bPTP results

Accession numbers

Species

Species 1 (support = 1.000)

KX044190

Hemidactyhis chipkali sp. nov.

Species 2 (support = 0.909)

EU268407, GQ375294, GQ375293,

EU268397, KU720667, KU720666,

KU720676, HM595649, KU720674,

KU720673, HM595659

Hemidactylus cf. murrayi

Species 3 (support = 1.000)

HM595660

Hemidactyhis gracilis

Species 4 (support = 0.865)

KU720656, KU720657, KU720659,

KU720660, KU720661, KU720662,

KU720658

Hemidactyhis cf. gleadowi

Species 5 (support = 0.689)

HM595670, HM595671, HM595669,

EU268410

Hemidactyhis reticulatus

Species 6 (support = 1.000)

HM595672

Hemidactyhis sataraensis

Species 7 (support = 0.915)

HM595642, HM595643

Hemidactyhis albofasciatus

Species 8 (support = 0.860)

EU268385, EU268386

Hemidactyhis imbricatus

Species 9 (support = 0.754)

KU720640, KU720641, KU720644,

KU720645, KU720647, KU720646,

GQ375298, GQ375300, GQ375291,

GQ375296, GQ375297, GQ375299,

GQ375292, KU720637, DQ120271,

DQ120272, HM595645, KU720638,

GQ375290, DQ 120273, KU720643,

KU720642, KU720639

Hemidactyhis parvimaculatus

Species 10 (support = 0.469)

KU720654, KU720653

Hemidactyhis cf. kushmorensis

Species 11 (support = 0.539)

HM595647, HM595646, KU720650,

KU720649, KU720652, KU720648,

KU720651

Hemidactyhis cf. kushmorensis

Species 12 (support = 1.000)

KU720681

Hemidactyhis treutleri (topotype)

Species 13 (support = 0.594)

KU720679

Hemidactyhis cf. treutleri

Species 14 (support = 0.425)

KU720677, KU720678

Hemidactyhis cf. treutleri

Amphib. Reptile Conserv.

March 2017 | Volume 11 | Number 1 | el 37

A new gecko of the genus Hemidactylus

Appendix V. Un-corrected pairwise sequence divergence between selected species of the genus Hemidactylus for the gene

cytochrome b.

1 DQ 120271

2 DQ 120272 0.00

DQ 120273

0.05

EU 268385

0.16

EU 268386

0.16

0.01

EU 268391

0.17

0.16

EU 268397

0.17

0.16

0.17

0.16

EU 268407

0.18

0.17

0.18

0.19

0.17

EU 268410

0.18

0.19

0.13

0.14

0.20

GQ 375290

0.05

0.01

0.15

0.16

0.17

GQ 375291

0.02

0.05

0.15

0.16

0.17

GQ 375292

0.02

0.05

0.15

0.16

0.17

GQ 375293

0.17

0.16

0.17

0.16

GQ 375294

0.17

0.16

0.17

0.16

GQ 375296

0.02

0.05

0.15

0.16

0.17

GQ 375297

0.02

0.05

0.15

0.16

0.17

GQ 375298

0.02

0.06

0.16

0.17

GQ 375299

0.02

0.05

0.15

0.16

0.17

GQ 375300

0.02

0.05

0.15

0.16

0.17

HM 595642

0.17

0.18

0.12

0.13

0.20

HM 595643

0.17

0.18

0.12

0.13

0.20

HM 595645

0.00

0.05

0.16

0.17

HM 595646

0.13

0.11

0.14

0.15

HM 595647

0.13

0.12

0.15

0.16

HM 595649

0.16

0.17

0.16

HM 595659

0.16

0.17

0.16

HM 595660

0.17

0.10

0.11

0.17

HM 595669

0.20

0.13

0.14

0.20

HM 595670

0.19

0.13

0.19

HM 595671

0.19

0.13

0.19

HM 595672

0.17

0.18

0.11

0.17

KU 720637

0.01

0.05

0.15

0.16

0.17

KU 720638

0.02

0.05

0.14

0.16

KU 720639

0.04

0.05

0.16

0.17

KU 720640

0.04

0.05

0.16

0.17

KU 720641

0.04

0.05

0.16

0.17

KU 720642

0.05

0.07

0.17

0.18

KU 720643

0.04

0.06

0.16

0.17

KU 720644

0.05

0.07

0.18

KU 720645

0.06

0.08

0.19

0.20

0.19

KU 720646

0.06

0.05

0.15

0.16

0.18

KU 720647

0.04

0.06

0.15

0.16

0.18

KU 720648

0.13

0.11

0.14

0.15

KU 720649

0.13

0.12

0.14

0.15

0.16

KU 720650

0.13

0.11

0.14

0.15

KU 720651

0.12

0.11

0.14

0.16

KU 720652

0.12

0.11

0.14

0.15

0.16

KU 720653

0.11

0.13

0.14

0.15

KU 720654

0.11

0.13

0.14

0.15

KU 720656

0.15

0.14

0.11

0.12

0.13

KU 720657

0.15

0.14

0.12

0.13

KU 720658

0.14

0.13

KU 720659

0.15

0.13

0.14

KU 720660

0.16

0.15

0.13

0.14

0.13

KU 720661

0.14

0.15

0.12

0.13

KU 720662

0.16

0.15

0.14

0.13

KU 720666

0.18

0.19

0.17

Amphib. Reptile Conserv.

0.02

0.20 0.21

0.16 0.17 0.18

0.16

0.17

0.05

0.16

0.17

0.05

0.00

0.02

0.20

0.16

0.00

0.02

0.20

0.16

0.00

0.16

0.17

0.05

0.00

0.16

0.17

0.05

0.00

0.16

0.00

0.17

0.18

0.17

0.05

0.01

0.17

0.01

0.16

0.17

0.05

0.00

0.16

0.00

0.16

0.17

0.05

0.00

0.16

0.00

0.18

0.20

0.15

0.17

0.18

0.17

0.18

0.20

0.15

0.17

0.18

0.17

0.18

0.05

0.02

0.17

0.02

0.15

0.17

0.20

0.11

0.12

0.15

0.12

0.16

0.17

0.21

0.11

0.13

0.16

0.13

0.03

0.04

0.18

0.15

0.16

0.03

0.16

0.03

0.04

0.18

0.15

0.16

0.03

0.16

0.17

0.18

0.15

0.17

0.16

0.17

0.16

0.20

0.21

0.03

0.19

0.18

0.20

0.18

0.19

0.21

0.03

0.18

0.19

0.18

0.19

0.21

0.03

0.18

0.19

0.18

0.16

0.17

0.11

0.17

0.16

0.17

0.04

0.01

0.16

0.01

0.16

0,17

0.18

0.05

0.02

0.16

0,16

0.02

0.18

0.20

0.18

0.04

0.18

0.04

0.18

0.20

0.18

0.04

0.18

0.04

0.18

0.20

0.18

0.05

0.03

0.18

0.03

0.19

0.20

0.19

0.07

0.04

0.19

0.04

0.18

0.19

0.18

0.05

0.04

0.18

0.04

0.17

0.18

0.06

0.03

0.17

0.03

0.17

0.18

0.07

0.04

0.17

0.04

0.18

0.19

0.18

0.05

0.04

0.18

0.04

0.17

0.19

0.17

0.05

0.04

0.17

0.04

0.15

0.17

0.20

0.11

0.12

0.15

0.12

0.16

0.17

0.20

0.11

0.13

0.16

0.13

0.15

0.17

0.20

0.11

0.12

0.15

0.12

0.15

0.16

0.20

0.10

0.11

0.15

0.11

0.16

0.17

0.21

0.10

0.11

0.16

0.11

0.14

0.15

0.20

0.11

0.14

0.11

0.14

0.15

0.20

0.11

0.14

0.11

0.14

0.15

0.17

0.14

0.15

0.14

0.15

0.13

0.14

0.17

0.14

0.15

0.13

0.15

0.13

0.14

0.17

0.13

0.15

0.13

0.15

0.12

0.13

0.17

0.14

0.16

0.12

0.16

0.13

0.14

0.18

0.15

0.16

0.13

0.16

0.14

0.15

0.18

0.14

0.15

0.14

0.15

0.13

0.14

0.16

0.15

0.13

0.15

0.02

0.03

0.20

0.17

0.18

0.02

0.18

0.01

0.00 0.01

0.00 0.01 0.00

0.17

0.00

0.02

0.17

0.12

0.13

0.12

0.19

0.13

0.20

0.13

0.01

0.16

0.19

0.16

0.19

0.16

0.00

0.16

0.17

0.16

0.17

0.16

0.17

0.18

0.19

0.18

0.16

0.20

0.18

0.19

0.18

0.15

0.19

0.20

0.17

0.18

0.15

0.19

0.20

0.17

0.16

0.17

0.16

0.11

0.17

0.18

0.16

0.01

0.02

0.01

0.17

0.01

0.12

0.13

0.16

0.02

0.03

0.02

0.17

0.02

0,11

0.12

0.15

0.04

0.16

0.04

0.11

0.12

0.18

0.04

0.16

0.04

0.11

0.12

0.18

0.03

0.04

0.03

0.16

0.04

0.11

0.12

0.18

0.04

0.05

0.04

0.16

0.05

0.14

0.18

0.04

0.16

0.04

0.12

0.13

0.17

0.03

0.04

0.03

0.17

0.05

0.13

0.14

0.17

0.04

0.05

0.04

0.17

0.06

0.13

0.14

0.18

0.04

0.05

0.04

0.18

0.06

0.13

0.17

0.04

0.17

0.04

0.13

0.17

0.12

0.13

0.12

0.19

0.13

0.00

0.01

0.16

0.13

0.19

0.13

0.01

0.16

0.12

0.13

0.12

0.19

0.13

0.00

0.01

0.16

0.11

0.12

0.11

0.18

0.12

0.01

0.15

0.11

0.12

0.11

0.19

0.12

0.02

0.16

0.11

0.17

0.11

0.10

0.14

0.11

0.17

0.11

0.10

0.14

0.15

0.16

0.15

0.14

0.15

0.14

0.15

0.16

0.15

0.14

0.15

0.14

0.15

0.14

0.13

0.16

0.15

0.13

0.16

0.17

0.16

0.15

0.16

0.14

0.15

0.16

0.14

0.15

0.14

0.15

0.16

0.15

0.17

0.16

0.17

0.13

0.18

0.20

0.18

0.17

0.02

March 2017 | Volume 11 | Number 1 | el 37

Mirza and Raju

Appendix V (continued). Un-corrected pairwise sequence divergence between selected species of the genus Hemidactylus for

the gene cytochrome b.

HM 595660

0.17

HM 595669

0.18

0.15

HM 595670

0.17

0.14

0.01

HM 595671

0.17

0.14

0.01

0.00

HM 595672

0.16

0.15

0.11

0.10

KU 720637

0.16

0.19

0.18

0.16

KU 720638

0.15

0.19

0.18

0.15

KU 720639

0.18

0.16

0.20

0.18

KU 720640

0.18

0.16

0.20

0.18

KU 720641

0.18

0.16

0.20

0.18

KU 720642

0.18

0.17

0.21

0.20

0.18

KU 720643

0.17

0.16

0.20

0.19

0.18

KU 720644

0.17

0.18

0.20

0.19

0.18

KU 720645

0.18

0.20

0.19

0.18

KU 720646

0.17

0.19

0.18

0.17

KU 720647

0.17

0.19

0.18

0.17

KU 720648

0.16

0.18

0.19

0.18

KU 720649

0.16

0.18

0.19

0.18

KU 720650

0.16

0.18

0.19

0.18

KU 720651

0.15

0.18

0.17

KU 720652

0.16

0.19

0.20

0.18

KU 720653

0.14

0.16

0.21

0.20

0.16

KU 720654

0.14

0.16

0.21

0.20

0.16

KU 720656

0.14

0.15

0.12

KU 720657

0.14

0.16

0.15

0.11

KU 720658

0.13

0.15

0.12

KU 720659

0.13

0.16

0.12

KU 720660

0.14

0.16

0.13

KU 720661

0.14

0.15

0.16

0.12

KU 720662

0.13

0.15

0.16

0.15

0.13

KU 720666

0.02

0.18

0.20

0.19

0.17

KU 720658

0.01

KU 720659

0.01

0.02

KU 720660

0.01

0.02

0.01

KU 720661

0.03

0.04

0.03

KU 720662

0.04

0.05

0.04

KU 720666

0.15

0.14

0.15

0.14

0.02

0.04 0.05

0.04 0.05 0.00

0.03

0.04

0.01

0.04

0.05

0.04

0.03

0.04

0.05

0.03

0.02

0.04

0.05

0.04

0.05

0.07

0.05

0.04

0.05

0.01

0.05

0.07

0.04

0.07

0.06

0.04

0.05

0.04

0.05

0.04

0.05

0.04

0.05

0.06

0.12

0.11

0.14

0.12

0.13

0.12

0.14

0.13

0.14

0.12

0.11

0.14

0.12

0.13

0.11

0.13

0.11

0.13

0.11

0.13

0.11

0.13

0.14

0.11

0.10

0.11

0.14

0.12

0.13

0.14

0.11

0.10

0.11

0.14

0.12

0.13

0.14

0.15

0.14

0.16

0.15

0.14

0.16

0.15

0.13

0.15

0.16

0.15

0.14

0.15

0.16

0.14

0.16

0.17

0.16

0.15

0.16

0.15

0.17

0.16

0.15

0.13

0.16

0.18

0.17

0.16

0.17

0.15

0.17

0.18

0.17

0.16

0.18

0.17

0.20

0.21

0.20

0.18

0.19

0.04

0.13

0.01

0.13

0.00

0.01

0.12

0.01

0.12

0.02

0.13

0.10

0.09

0.13

0.10

0.09

0.00

0.15

0.16

0.14

0.15

0.16

0.14

0.15

0.01

0.14

0.15

0.14

0.13

0.14

0.01

0.15

0.16

0.15

0.14

0.15

0.02

0.16

0.17

0.15

0.16

0.02

0.16

0.17

0.15

0.14

0.15

0.03

0.17

0.18

0.16

0.17

0.15

0.05

0.20

0.19

0.17

0.16

0.17

0.15

Amphib. Reptile Conserv.

March 2017 | Volume 11 | Number 1 | el 37

A new gecko of the genus Hemidactylus

Appendix V (continued). Un-corrected pairwise sequence divergence between selected species of the genus Hemidactylus for

the gene cytochrome b.

KU 720667

0.17

0.16

0.17

0.16

0.00

0.02

0.20

0.16

0.00

0.16

0.17

0.16

0.18

0.17

0.15

0.16

0.03

KU 720673

0.16

0.17

0.16

0.03

0.04

0.18

0.15

0.16

0.03

0.16

0.19

0.16

0.00

KU 720674

0.16

0.17

0.16

0.03

0.04

0.18

0.15

0.16

0.03

0.16

0.19

0.16

0.00

KU 720676

0.16

0.15

0.07

0.08

0.18

0.16

0.15

0.07

0.15

0.16

0.15

0.20

0.16

0.14

0.15

0.06

KU 720677

0.17

0.16

0.14

0.15

0.17

0.11

0.13

0.17

0.15

0.16

0.11

0.16

0.17

0.16

0.18

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0.16

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0.13

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0.16

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0.16

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0.17

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0.11

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0.17

0.11

0.17

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0.16

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0.16

0.17

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0.12

0.14

0.15

0.17

0.15

0.12

0.15

0.16

0.15

0.19

0.16

0.13

KX 044190

0.22

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0.17

0.18

0.17

0.13

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0.20

0.22

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0.18

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0.16

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0.11

0.19

0.20

0.21

0.22

0.21

0.22

0.23

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0.23

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0.18

0.17

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0.16

KU 720667

0.13

0.12

0.13

0.14

0.13

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0.14

0.13

0.14

0.13

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0.03

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0.14

0.13

0.14

0.13

0.02

0.03

0.00

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0.13

0.12

0.14

0.13

0.06

0.07

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0.13

0.14

0.13

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0.13

0.11

0.12

0.14

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0.14

0.15

0.14

0.15

0.14

0.13

0.15

0.03

KU 720679

0.12

0.13

0.12

0.13

0.11

0.13

0.11

0.10

0.12

0.04

0.05

KU 720681

0.14

0.13

0.14

0.12

0.13

0.10

0.09

0.10

KX 044190

0.17

0.16

0.11

0.13

0.11

0.14

0.15

0.17

0.14 0.16

Amphib. Reptile Conserv. 71 March 2017 | Volume 11 | Number 1 | el 37

Official journal website:

amphibian-reptile-conservation.org

Amphibian & Reptile Conservation

11(1) [General Section]: 72-87 (e138).

Larval development and breeding ecology of Ziegler’s

Crocodile Newt, Tylototriton ziegleri Nishikawa, Matsui and

Nguyen, 2013 (Caudata: Salamandridae), compared to other

Tylototriton representatives

12 Marta Bernardes, ^nna Rauhaus, 2 Clara Michel, 3 ’ 8 Cuong The Pham, 3 ’ 8 Truong Quang Nguyen,

4 ’ 5 ’ 6 Minh Due Le, Trank Pasmans, 2 Michael Bonkowski, and ^Thomas Ziegler

1 Cologne Zoo, Riehler Strafe 173, 50 735, Cologne, GERMANY 2 Department of Terrestrial Ecology, Institute of Zoology, University of Cologne,

Ziilpicher Strafe 47b, 50674, Cologne, GERMANY 3 Institute of Ecology! and Biological Resources, Vietnam Academy of Science and Technology,

18 Hoang Ouoc Viet Road, Hanoi, VIETNAM ^Faculty of Environmental Sciences, Hanoi University of Science, Vietnam National University, 334

Nguyen Trai Road, Hanoi, VIETNAM 5 Central Institute for Natural Resources and Environmental Studies, Hanoi National University, 19 Le Thanh

Tong, Hanoi, VIETNAM 6 Department of Herpetology>, American Museum of Natural History, Central Park West at 79th Street, New York, New York

10024 1 Department of Pathology>, Bacteriology> and Avian Diseases, Faculty of Veterinary Medicine, Ghent University, Salisburylaan 133, B 9820

Merelbeke, BELGIUM 8 Graduate University of Science and Technology>, Vietnam Academy of Science and Technology, 18 Hoang Ouoc Viet, Can

Giay, Hanoi, Vietnam.

Abstract. —We describe for the first time the larval development and stages of the recently described

Ziegler’s Crocodile Newt (Tylototriton ziegleri), an endemic species to northern Vietnam. Diagnostic

morphological characters are provided for Grosse (1997, 2013) stages 27-32, 35-36, and 44-45, as

well as comparisons with larval stages of other Tylototriton representatives. In addition, natural

history data and an ecological assessment of the breeding niche are presented for T. ziegleri as well

as for T. vietnamensis, from whom the former species was only recently taxonomically separated.

We provide data extending the known breeding season of these two cryptic species in the North of

Vietnam, which in fact lasts from April until July. On average, the clutches of T. ziegleri consisted

of 67 ± 32 eggs, were found on rock and soil substrates with a distance of 50 ± 28 cm from water,

whereas the clutches of T. vietnamensis were significantly smaller (43 ± 19 eggs), found only on soil

and were further distant from water (80 ± 41 cm). The known maximum altitudinal distribution of T.

vietnamensis is herein increased to 980 m above sea level. Based on the examples of T. ziegleri and

T. vietnamensis, this study highlights how important it is to uncover cryptic species, define their

exact distribution range, and investigate potential differences in ecological adaptations in order

to assess the conservation status, develop proper conservation planning and provide suitable

conditions for potential ex situ breeding programs.

Keywords. Vietnam, Crocodile Newts, cryptic species, developmental biology, larval staging, microhabitat character¬

ization, conservation, captive breeding

Citation: Bernardes M, Rauhaus A, Michel C, Pham CT, Nguyen TQ, Le MD, Pasmans F, Bonkowski M, Ziegler T. 2017. Larval development and

breeding ecology of Ziegler’s Crocodile Newt, Tylototriton ziegleri Nishikawa, Matsui and Nguyen, 2013 (Caudata: Salamandridae), compared to other

Tylototriton representatives. Amphibian & Reptile Conservation 11(1) [General Section]: 72-87 (el 38).

NoDerivatives 4.0 International License, which permits unrestricted use for non-commercial and education purposes only, in any medium, provided

the original author and the official and authorized publication sources are recognized and properly credited. The official and authorized publication

credit sources, which will be duly enforced, are as follows: official journal title Amphibian & Reptile Conservation ; official journal website <amphibian-

reptiie-conservation.org>.

Received: 04 Nov 2016; Accepted: 02 March 2017; Published: 17 April 2017

Introduction

The genus Tylototriton currently consists of 22 species

with a distribution from Nepal, Bhutan, and India east¬

wards to China and southwards to Indochina (Nishikawa

et al. 2013a). Phylogenetic analyses divided the genus

into the T. asperrimus group (Fei et al. 2005) or the sub¬

genus Yaotriton (Dubois and Raffaelli 2009), which

Correspondence. Email: *ziegler@koelnerzoo.de

includes: T. asperrimus, T. broadoridgus, T. dabienicus,

T. hainanensis, T. liuyangensis, T. lizhenchangi, T. notia-

lis, T. podichthys, T. panhai, T. vietnamensis, T. wenxia-

nensis, and T. ziegleri ; and the T. verrucosus group (Fei

et al. 2005) or the subgenus Tylototriton (Dubois and

Raffaelli 2009), which includes: T. anguliceps, T. hima-

layanus, T. kweichowensis, T. pseudoverrucosus, T. shan-

jing, T. shanorum, T. taliangensis, T. uyenoi, T. verruco-

Amphib. Reptile Conserv.

April 2017 | Volume 11 | Number 1 | el 38

Bernardes et al.

Fig. 1. A. Adult male of Tylototriton ziegleri ; B. Habitat type in Bao Lac district, Cao Bang Province; C. Adult male of Tylototriton

vietnamensis ; D. Habitat type in Tay Yen Tu Nature Reserve, Bac Giang Province. Photos M. Bernardes.

sns, and T. yangi (Khatiwada et al. 2015; Le et al. 2015;

Nishikawa et al. 2014; Phimmachak et al. 2015; Yang et

al. 2014).

In Vietnam, this genus is currently represented by T.

asperrimus, T. notialis, T. angidiceps, as well as by two

endemic species, viz. T. vietnamensis, and T. ziegleri.

Specimens of Ziegler’s Crocodile Newt were previously

referred to T. asperrimus (Sparreboom et al. 2011, Yuan

et al. 2011) or T. cf. vietnamensis (Stuart et al. 2010).

Tylototriton ziegleri was subsequently described as a dis¬

tinct species by Nishikawa et al. (2013b) based on mor¬

phological and molecular differences from T. vietnamen¬

sis. The latter species has been evaluated as Endangered

in the Vietnam Red Data Book (Tran et al. 2007), and

in the IUCN Red List (IUCN SSC Amphibian Specialist

Group 2016).

Tylototriton vietnamensis inhabits secondary ever¬

green lowland forests on granite parent rock mate¬

rial consisting of hardwood, bamboo and shrubs and is

known from lower elevations in Bac Giang, Quang Ninh,

Lang Son, and Phu Tho provinces (Bernardes et al. 2017;

Nguyen et al. 2009). In contrast, T. ziegleri is known

from primary forests on limestone parent rock material at

higher elevations characterized mainly by bamboo veg¬

etation in Cao Bang and Ha Giang provinces (Nishikawa

etal. 2013b) (Fig. 1).

In-depth studies focusing on distinctive features and

thus on the taxonomic status of closely related or at least

similar, potential taxa which are either threatened and/

or have a limited range, as in the case of the species pair

T. vietnamensis and T. ziegleri , are important for proper

identification and suitable conservation actions. Since

both species are distributed at different elevations and

occupy distinct geological areas, we expected to find eco¬

logical, morphological, and developmental differences

to support their discrimination. Herein, we document

for the first time the larval development of T. ziegleri ,

in comparison with information on the development of

other Tylototriton species. We also provide data on the

ecological niche of T. ziegleri , in particular microhabitat

preferences associated with reproduction, and compare

this with our own field data for T. vietnamensis.

Materials and Methods

Field surveys. Field surveys were conducted by M.

Bernardes, C.T. Pham and H.T. An during the rainy sea¬

son between 10 April and 11 July 2010, 8 June and 7

August 2012, 13 June and 28 July 2013, and 15 May and

28 June 2014 in northern Vietnam. The surveys were con¬

ducted in Son Dong and Luc Nam districts in Bac Giang

Province, Hoanh Bo district in Quang Ninh Province,

and Mau Son district in Lang Son Province for T. viet¬

namensis ; and in Quan Ba and Bac Quang districts in Ha

Giang Province, and Bao Lac district in Cao Bang Prov¬

ince for T. ziegleri. Daytime visits to breeding sites were

conducted for an average time of 20 minutes and var¬

ied between two and eight times, sometimes in repeated

Amphib. Reptile Conserv.

April 2017 | Volume 11 | Number 1 | el 38

Larval development and breeding ecology of Ziegler’s Crocodile Newt

years. Besides coordinates and elevations recorded with

Garmin GPS MAP62, a defined set of several abiotic

variables were collected and recorded in order to classify

each study site according to their environmental condi¬

tions. The physical characteristics of each pond (area and

maximum depth) were determined by use of a measur¬

ing rope of precisely known size. A pH meter (Hanna HI

98129) calibrated for 25 °C was used to measure the pH

and record water temperature. Water chemical parame¬

ters were taken for an analysis of pollution (concentra¬

tion of nitrate [N0 3 -] and nitrite [N0 2 -]) and water hard¬

ness (carbonate [KH] and total hardness [gH], measure¬

ments following the German degree) using drop-by-drop

color tests from JBL (Testlab, Germany). Since some

pools occurred only temporary the hydroperiod was also

taken into account. Weather data consisting of tempera¬

ture, humidity, and atmospheric pressure were recorded

with a weather station (Krestel 3500) at each breeding

site. The tree canopy cover above the pond water was

assessed visually and classified in five levels ranging

from 0-100% cover.

Adult animals were searched on the bottom of the

water body using visual survey encounter techniques fol¬

lowed by subsequent capture with a sweep of a 25 cm

dip-net (JBL, Germany). The surrounding shore of the

pond was searched for the presence of egg clutches, efts,

and additional adults until a maximum distance of three

meters from the water line. All captured adults were

counted, photographed, and subsequently released. The

distance of the egg clutch to the water source was mea¬

sured and the total number of eggs present in the clutches

counted with minimum disturbance. The diameter and

the weight of seven randomly picked eggs (in the case of

T. ziegleri belonging only to Bao Lac district, Cao Bang

Province) were measured by using a digital calliper to the

nearest 0.1 mm and weighted with a digital scale to the

nearest 0.01 g.

Larval staging, morphological description and

comparisons with congeners. One clutch of T.

ziegleri was collected on the 17 th July 2014 in Bao Lac

district, Cao Bang Province, Vietnam to observe the lar¬

val development and for further morphological compari¬

sons. The clutch was collected together with associated

substrate and foliage coverage from the shore of a breed¬

ing pond. The clutch was kept inside an open-air plastic

box and regularly sprayed with water to keep up the suit¬

able humidity level. Due to the thickness of the gelati¬

nous layer (albumen) it was not possible to observe and

document the developing larvae inside; therefore we pre¬

served few eggs for morphological analysis. Every one

or two days one egg was randomly selected (in total 23),

transferred to a 4% formalin solution for fixation and

subsequently preserved in 70% ethanol. This procedure

was repeated until hatching of the first larva. The remain¬

ing larvae were later transferred to the Me Linh Station

for Biodiversity to contribute to a captive breeding pro¬

gram. In addition, seven swimming larvae were collected

at the breeding pond for a morphological description

of more advanced developing stages. The larvae were

anaesthetized with ethyl acetate, fixed in 40% ethanol,

and later transferred to 70% ethanol for preservation.

Preserved individuals subsequently were deposited in

the collections of the Institute of Ecology and Biological

Resources (IEBR), Hanoi, Vietnam, with the catalogue

numbers: IEBR A.2016.19-A.2016.31 and of the Zoolo-

gisches Forschungsmuseum Alexander Koenig (ZFMK),

Bonn, Germany, with the catalogue numbers: ZFMK

98792-ZFMK 98796.

Preserved eggs were cut open to examine the devel¬

oping larvae inside. A detailed description of the devel¬

opment and measurements of the ovum in early stages

could not be performed as the jelly layers could not be

opened without destroying the fragile content. Staging

was performed under a magnifying loupe and by support

from a digital microscope (Keyence VHX-500F) when

extra magnification, photographs, and morphological

measurements were needed. To complement the charac¬

terization process (e.g., for characterizing the coloration

in life), additional photographs of hatched larvae were

taken during field work by M. Bemardes or at the Me

Linh Station for Biodiversity by T. Ziegler by placing

single larvae into a water filled glass vessel.

Larval stages were identified according to Grosse

(1997, 2013) and the morphological terminology fol¬

lowed Nishikawa et al. (2013b). The following measure¬

ments were taken: snout-vent length (SVL), from tip of

snout to posterior edge of the vent; head length (HL),

from posterior edge of right parotid to snout tip; maxi¬

mum head width (HW); head height (HH), measured

above the eyes; snout length (SL), from right nostril

to right posterior corner of mouth; interocular distance

(IoD), from anterior comer of eyes; internostril distance

(InD); eye-nostril distance (EnD), from right nostril to

anterior corner of right eye; forelimb length (F1L), from

right anterior limb measured from point of body insertion

to tip of longest finger; hind-limb length (H1L), right pos¬

terior limb measured from point of body insertion to tip

of longest toe; axillar distance (AD), from axilla to groin

on right side; width of tail base (TW), measured at poste¬

rior edge of vent; maximum tail height (TH); tail length

(TaL), from posterior edge of vent to tail tip; total length

(TL), from tip of snout to tail tip.

For morphological comparisons, data from literature

was included for the following species: T. angnliceps, T.

asperrimns , T. broadoridgus , T. hainanensis , T. himala-

yanus, T. kweichowensis, T. liuyangensis, T. podichthys,

T. cf. sharping , T. shanorum , T. taliangensis, T. uyenoi,

and T. wemianensis (see Bourret 1942; Khatiwada et al.

2015; Kuzmin et al. 1994; Mudrack 2005; Nishikawa et

al. 2013a, 2014, 2015; Phimmachak et al. 2015; Shen et

al. 2012; Sparreboom2014; Yang etal. 2014; Zhao 1988;

Ziegler et al. 2008). For detailed comparisons with T.

vietnamensis we included our own field data and pictures

Amphib. Reptile Conserv.

April 2017 | Volume 11 | Number 1 | el 38

Bernardes et al.

and Sal_ND2_R2 (Nishikawa et al. 2013b). Tissue sam¬

ples were extracted using DNeasy blood and tissue kit,

Qiagen (California, USA). Extracted DNA from the

fresh tissue was amplified by PCR mastermix (Fermen-

tas, Canada). The PCR volume consisted of 21 pi (10 pi

of mastermix, five pi of water, two pi of each primer at 10

pmol/pl, and two pi of DNA or higher depending on the

quantity of DNA in the final extraction solution). PCR

condition was: 95 °C for five minutes to activate the taq;

with 40 cycles at 95 °C for 30 s, 50 °C for 45 s, 72 °C

for 60 s; and the final extension at 72 °C for six minutes.

PCR products were subjected to electrophoresis

through a 1% agarose gel (UltraPure™, Invitrogen). Gels

were stained for 10 minutes in IX TBE buffer at two pg/

ml of ethidium-bromide, and visualized under UV fight.

Successful amplifications were purified to eliminate PCR

components using Gene JET™ PCR Purification kit (Fer-

mentas, Canada). Purified PCR products were sent to

Macrogen Inc. (Seoul, South Korea) for sequencing.

Sequences generated in this study were aligned with

one another using the De Novo Assemble function in

the program Geneious v.7.1.8 (Kearse et al. 2012). They

were then compared with other sequences using the Basic

Local Alignment Search Tool (BLAST) in GenBank.

Results

Molecular analysis. Three sequences of 987 bps were

obtained. The sequences were almost identical, except in

two positions, and 99% to 100% similar to the sequence

with the Gen Bank’s accession number AB769542 of T.

ziegleri (voucher VNMN 3389). The results confirm the

samples collected in Cao Bang Province are conspecific

with T. ziegleri.

Distribution, ecological niche and microhabitat

use of T. ziegleri and T. vietnamensis. Tylototri-

ton ziegleri was found in Cao Bang Province at eleva¬

tions between 1,325 and 1,420 m above sea level, in Ha

Giang Province, Bac Quang district between 868 and 932

m above sea level, and in Quan Ba district between 1,080

Table 1. Results of water chemical analysis conducted during field work in the habitat of Tylototriton vietnamensis and T. ziegleri

during 2010, 2013, and 2014. Values are presented as min. - max. (mean ± standard deviation).

Species

Province

district

°KH

°gH

NOffmg 1-1)

N0 3 '(mg 1-1)

T. vietnamensis

Bac Giang

Son Dong

4.65-6.43

1-5

1-5.5

0-0.5

0-10

(5.48 ±0.48)

(1.80 ± 1.11)

(2.46 ± 1.63)

(0.04 ±0.11)

(3.39 ±2.38)

Quang Ninh

Uong Bi

7.36-7.51

4-8

5-6

0-0.4

5-15

(7.43 ±0.11)

(6 ±2.83)

(5.5 ±0.71)

(0.2 ±0.28)

(10 ±7.07)

T. ziegleri

Cao Bang

Bao Lac

7.08-7.28

6-6

7-7

0.03-0.03

0.03-20

(7.18 ±0.14)

(10.01 ± 14.12)

Ha Giang

Bac Quang

6.36-7.05

1-1

1-2

0-0.05

1-1

(6.63 ±0.37)

(1.33 ±0.58)

(0.03 ± 0.03)

Ha Giang

Quan Ba

6.41-7.94

1-8

1 -9

0.05-0.4

0-1

(7.30 ±0.61)

(5.33 ±2.88)

(5 ±3.41)

(0.13 ±0.14)

(0.79 ±0.4)

100 %

90%

80%

70%

60%

50%

40%

30%

20 %

10 %

T. vietnamensis T. ziegleri

Fig. 2. Percentage of the number of adults of Tylototriton

vietnamensis and T. ziegleri found at each interval of percentage

of canopy cover measured above the water of the breeding site.

of larvae (in stages 33, 35, 41, and 42) photographed

either in situ during our field work in Bac Giang Prov¬

ince by M. Bernardes or ex situ at the Me Linh Station for

Biodiversity by T. Ziegler.

Statistical analysis. Comparisons between T. ziegleri

and T. vietnamensis regarding the area and depth of the

different ponds, clutch sizes, and distance to water, as

well as the regression between the clutch and egg sizes

within the genus were examined with Student’s /-test

after confirming a normal distribution of the data. Analy¬

ses were performed in R version 3.2.3.

Molecular analysis. For species identification, we

sequenced a partial mitochondrial gene, the NADH dehy¬

drogenase subunit 2 (ND2), for the egg / larval tissue

samples (IEBR A.2016.19-A.2016.21) from the clutch

of T. ziegleri collected on the 17th July 2014 in Bao Lac

district, Cao Bang Province, Vietnam, which was used

for larval staging, using the primer pair, Sal_ND2_Fl

■ 75-100%

□ 50-75%

□ 25-50%

□ 0-25%

□ 0 %

Amphib. Reptile Conserv.

April 2017 | Volume 11 | Number 1 | el 38

Larval development and breeding ecology of Ziegler’s Crocodile Newt

Fig. 3. A: Typical clutch of Tylototriton ziegleri composed by

single eggs; B: an exceptional case of “stickiness” where eggs

were aggregated in groups of 2-4. Photos M. Bernardes.

and 1,369 m above sea level. Tylototriton vietnamensis

was found between 181 and 512 m above sea level in Bac

Giang and Quang Ninh provinces, and between 840 and

980 m above sea level in Lang Son Province. Spawning

sites consisted of small ponds for both species, although

in the district Quan Ba, Ha Giang Province we also found

clutches of T. ziegleri in the slopes of a slow flowing for¬

est stream, suggesting that this species can also breed in

this type of habitat. A physical evaluation of ponds dur¬

ing our field work showed that the ones inhabited by

T. ziegleri were significantly deeper (F 142 = 25.11, P <

0.001; mean 79 ± 58 cm, n = 19, range between 10 and

200 cm) than those inhabited by T. vietnamensis (mean

25 ± 14 cm, n = 81, range between 3 and 60 cm), while

the area was roughly the same (Fj 44 = 0.004, P = 0.95; T.

ziegleri: mean 84 ± 165 m 2 , range between 2.5 and 510

m 2 ; T. vietnamensis: mean 82 ± 102 m 2 , range between

one and 460 m 2 ). Most adults (61% of 82 individuals of

T. ziegleri and 72.2% of 255 individuals of T. vietnam¬

ensis) were found in breeding sites with 50% or more

canopy cover, although still 34.1% of all T. ziegleri and

12.2% of all T. vietnamensis were found in breeding sites

with no canopy cover (Fig. 2).

A comparison of water quality showed that T. ziegleri

occurred in ponds with pH values between 6.4 and 8

(mean 7 ± 0.5; throughout Cao Bang and Ha Giang prov¬

inces), while T. vietnamensis occurred in ponds with pH

values ranging from 4.7 to 7.5 (mean 5.6 ± 0.7; through-

Amphib. Reptile Conserv.

Fig. 4. Drawing of a formol-preserved larva of Tylototriton

ziegleri at stage 35. Drawing C. Michel.

out Bac Giang and Quang Ninh provinces). Follow¬

ing US Geological Survey standard for water hardness

classification both T. ziegleri and T. vietnamensis var¬

ied between soft (0-1 °KH) and hard (8 °KH), with T.

ziegleri distributed over an average of 4 ± 3 °KH and T.

vietnamensis over an average of 2 ± 2 °KH. The general

hardness was also on average higher for T. ziegleri (1-9

°gH; mean 4 ± 3 °gH) compared to T. vietnamensis (1-6

°gH; mean 3 ± 2 °gH). Concentration of nitrite ranged

from 0-0.4 mg 1-1 for T. ziegleri and from 0-0.5 mg

1-1 for T. vietnamensis, while concentrations of nitrate

ranged from 0-20 mg 1-1 and 0-15 mg 1-1, respectively

(Table 1). Environmental data revealed higher humid¬

ity levels for T. ziegleri (mean 100 ± 0%) than those for

T. vietnamensis (mean 94 ± 9%, range between 68 and

100%), and slightly higher temperature oscillations for

T. vietnamensis 24.2-34.2 °C (mean 28.6 ± 2.2 °C) than

those for T. ziegleri 26-34.4 °C (mean 27.4 ±3.3 °C).

Mating and egg deposition of T. ziegleri and T.

vietnamensis. During the breeding season of these two

species (April-July) reproductive males that were other¬

wise terrestrial, moved into the water at the breeding sites

and waited for the females. When precipitation was lack¬

ing and breeding sites dried out, adults were forced to

maintain their terrestrial life mode. However, if climatic

conditions were favorable, males preferably were found

inside the water. From a total of 547 captured adults of

T. vietnamensis , and 101 adults of T. ziegleri, only five

(0.91%) and two (1.98%), respectively, were found on

land. Females seem to join the males in the water for a

very short period, since only 12 females of T. vietnam¬

ensis and one female of T. ziegleri were captured dur¬

ing field work. After courtship and reproduction, gravid

females laid large eggs at the shore of the water body

and returned to the forest. Due to their slight stickiness,

the eggs aggregated in egg masses (Fig. 3A) which were

subsequently covered by the females with leaf litter from

the forest ground. The clutch size was significantly dif¬

ferent for the two species (F } 71 = 11.29, P < 0.001). For

T. ziegleri, it ranged between 10 and 109 eggs (mean 67

± 32 eggs, n = 10), with clutches found between rock

(83.3%) and soil (16.7%) substrates, while for T. vietna¬

mensis (n = 6 3) it ranged between 5 and 85 eggs (mean

43 ± 19 eggs) and clutches were always found on soil

substrate. Clutches of T. ziegleri and T. vietnamensis dif¬

fered significantly in their distances to the nearest water

April 2017 | Volume 11 | Number 1 | el 38

Bernardes et al.

Fig. 5. Metamorph of Tylototriton ziegleri at stage 44, with an

additional finger on left hand, collected in Ha Giang Province

in 2012 and preserved in ethanol. Photo M. Bernardes.

body (Fj 50 = 5.32, P < 0.01). Clutches of T. ziegleri were

between 10 and 100 cm away from water (mean 50 ± 28

cm, n = 11), while the ones from T. vietnamensis were

found at a distance between 17 and 188 cm (mean 80 ±

41 cm, n = 41) from the water.

In one exceptional case a clutch of T. ziegleri con¬

sisted of eggs sticking so strongly together that they were

no longer solitary but formed small aggregations of two

to four eggs (Fig. 3B).

Egg description, developmental stages and lar¬

val morphology of T. ziegleri. From a total of 80

eggs in the collected clutch of T. ziegleri , 34 (42%) did

not show normal development. The same count was done

in the field with one other clutch consisting of 107 eggs

and revealed that 23% of the eggs had ceased to develop.

We estimated that the collected egg clutch was around

two days old at the time of collection (16th June 2014).

The diameter of the preserved eggs ranged between 8.7

and 11.2 mm (mean 10.1 ± 0.8 mm, n = 23), while the

diameter of eggs measured randomly in the field (all

measurements of the eggs from Bao Lac district, Cao

Bang Province) ranged between 7.2 and 8.9 mm (mean

8.1 ± 0.8 mm, n = 28). The associated weight for the eggs

measured in the field ranged between 0.2 and 0.4 g (mean

0.3 ±0.1 g,« = 28).

In general, the egg shape was round and the sur¬

face of the jelly layers uneven. Most eggs cointained a

clear gelatinous layer, few were slightly more opaque.

The liquid inside was clear. The capsular chamber con¬

tained the embryo or in less developed stages the ovum

which was either attached or not attached to the inside of

the chamber. An outer observation of the ovum in early

stages identified a round ovum with a creamy yellow¬

ish white color. The size of three measured ovae varied

between 3.87 and 4.78 mm. Larvae in an advanced stage

of development showed a more slender shape, curved to

fit inside the capsular chamber. While some stages were

represented by multiple specimens, stages 33, 34, and

37—43 were not found and thus could not be examined.

At stage 27 (IEBR.A.2016.22) gill and forelimb buds

were discernible. Stage 28 (IEBR.A.2016.23) was char¬

acterized by further development of the forelimb buds.

Fig. 6. Tylototriton ziegleri larvae from Bao Lac district, Cao Bang Province with indication of the corresponding developmental

stage and scale. Photographs of stages 27 to 36 are from preserved eggs photographed under a digital microscope (photos C. Michel)

and photographs of stages 44 and 45 are from individuals in life kept at the Me Linh station (photos T. Ziegler).

Amphib. Reptile Conserv.

April 2017 | Volume 11 | Number 1 | el 38

Larval development and breeding ecology of Ziegler’s Crocodile Newt

This was also the case for stage 29 (IEBR.A.2016.24 and

ZFMK 98792) along with the growth of fimbriae. Stage

30 (IEBR.A.2016.25) could be determined by the dome

shaped distal tip of the forelimb. The forelimb was cone

shaped at stage 31 (IEBR.A.2016.26 and ZFMK 98793).

Two digits were formed at stage 32 (IEBR.A.2016.27).

At stage 35 (IEBR.A.2016.28 and ZFMK 98794) the

forelimb had a joint and a hand with three digits. Addi¬

tionally the hind limbs started to develop and in some

cases already showed toe buds (see Fig. 4). At this stage,

around 20 days after the assumed egg deposition date (4th

July), the first larvae hatched, while some still remained

inside the egg. At hatching time larvae had an average

total size of 14.65 ± 0.77 mm (size ranged between 13.78

and 15.22 mm, n = 3). Stage 36 (IEBR.A.2016.29 and

ZFMK 98795) was characterized by a forelimb with four

digits and a hind limb with three toes and a knee joint.

The yolk sac was prominent in stages 27-30 and was

evident until stage 35. At stages 44 (IEBR.A.2016.30)

and 45 (IEBR.A.2016.31 and ZFMK 98796), larvae were

black and had well developed limbs with four fingers and

five toes, and the gills atrophied. In one individual at

stage 44 we observed the splitting of one finger in two,

resulting in five fingers on the left hand (Fig. 5). No juve-

Table 2. Developmental stages, morphological description and coloration of Tylototriton ziegleri from stages 27 to 45; stage

diagnostic characteristics according to Grosse (1997, 2013) are italicized. Specimens from stages 27-36 originated from the egg

clutch while data on stages 44 and 45 were gathered from hatched larvae collected inside the breeding pond.

Stage

Morphology

Coloration

(n= 1)

Head trapezoidal and sloping in profile, snout short and flat, no labial fold

visible, eyes distinguishable but unobtrusive. Gills upright, shorter than

head. No balancers. Dorsal and ventral fins about the same height, higher

than head; dorsal fin starts at last third of the trunk; tail short; tail-tip round.

Forelimb-buds start developing; yolk mass twice as high as body, nearly

round. Larva clearly visible through egg jelly and can be moved inside the

capsular chamber.

Ground color white-yellowish, with yolk mass more

yellow. Dark, irregularly distributed pigmentation

on dorsum head and flanks, getting lighter towards

the ventrum until total disappearance on ventral side.

Eyes white without pigmentation.

(»= 1)

Head trapezoidal and sloping in profile, snout short and flat, no labial fold

visible; gills nearly head high. Dorsal fin higher than head, starting at last

third of the trunk; ventral fin shorter than dorsal fin; tail stretching; tail-tip

round. Forelimb-buds clearly visible, yolk mass big and round.

Pigmentation similar to stage 27, additionally

longitudinal lateral stripe on rib area without

pigmentation. Slightly pigmented rim around the

eyes forming circle; forelimb-bud base with slight

pigmentation on dorsal side.

(» = 2)

Head more pronounced, labial fold distinct at posterior half of upper jaw;

gills developing fimbriae and higher than head; tail getting longer; yolk is

less round and oval shaped; forelimb-buds longer with rounded tip; mouth

is located on ventral side of head, beneath the snout tip.

Pigmentation getting darker, particularly in the eyes,

also slight pigmentation underneath the gular fold.

in-2)

Shape similar to stage 29, forelimb-buds are slightly longer with a dome

shaped distal tip; gill rami and fimbriae more developed, dorsal and ventral

fin have become larger.

No change in pigmentation.

in-2)

Gills growth; labial folds distinct at posterior half of upper jaw; forelimb

cone shaped; tail and fins well developed; dorsal fin starts at middle of the

trunk; yolk mass receding.

Pigmentation getting darker, denser pigmentation

on dorsum behind head; eyes nearly fully black with

white pupil, small, unpigmented stripe from pupil

towards ventral side; gill rami slightly pigmented on

upper side; fimbriae without pigmentation.

in- 1)

Dorsal and ventral fin growing; gular fold clearly visible; mouth orientation

is more rostral; two digits developing as small knobs on distal edge of

forelimb with a notch forming in the middle.

Pigmentation getting denser on dorsum forming a dark

stripe with unpigmented spots; head pigmentation

less dense; eyes except for pupil fully pigmented; few

dark spots on dorsal and ventral fin and forelimb.

in- 11)

Head more depressed, sloping in profile; mouth more pronounced with nares

clearly visible; hand with three digits is visible beyond the end of the gills;

digits round at the tip; the middle digit the longest; limb with joint, bending

at the elbow; yolk mass has almost completely receded; gut getting tubular.

Hmd limb buds discernible and in some cases elongated, indentation

between first two toes in some larvae.

No change in pigmentation.

in = 3)

Lateral line organs visible on ventral side of head; mouth open with well-

developed teeth; four digits have formed on hand, forelimb turned, palm is

facing ventrally. Hind limb with three toes and a knee joint starting to form.

No change in pigmentation.

in - 4)

Skin mostly smooth with some warts starting to form; tail long and pointed;

limbs well developed with four fingers and five toes; no remains of yolk;

head trapezoidal, wide and depressed with a short and flattened snout; dorsal

and ventral fin receding; dorsal fin beginning on the first quarter of back and

ventral fin beginning above the cloaca; caudal fin higher than head; gills

higher than body, with fimbriae still clearly visible.

Pigmentation black and dense over the whole body;

lighter on underside of head and ventral side; tip of

toes and fingers and labial folds are unpigmented.

in = 3)

Skin gets less smooth and more granular and warty; teeth well developed;

fins receding; gills atrophy (only stumps left).

Similar to stage 44, but tip of toes and fingers are

colored in yellow.

Amphib. Reptile Conserv.

April 2017 | Volume 11 | Number 1 | el 38

Bernardes et al.

Table 3. Morphological measurements of larvae and respective eggs of Tylototriton ziegleri from Ha Giang sorted by stage (mean

± standard deviation, range in parenthesis, in mm). N: Number of individuals, D: diameter. For abbreviations see Material and

Methods. Internostril distance, eye-nostril distance, and snout length were not discernible for stages 27-30 and hind limb length

could only be measured from stage 25 onwards.

Stage:

(N)

(2)

(H)

(3)

(4)

(3)

SVL

7.3

5.30

5.94 ±0.21

(5.73-6.15)

5.41

5.95 ±0.14

(5.85-6.05)

7.87

7.6 ±0.56

(6.32-8.28)

6.89 ±0.18

(6.76-7.02)

21.39 ±9.37

(10.92-31.94)

34.62 ± 0.25

(3444-34.8)

Tal

2.36

1.67

3.71 ±0.19

(3.53-3.9)

3.99

3.13 ±0.73

(2.61-3.65)

5.59

6.49 ±0.85

(5.16-8.7)

5.56 ±2.54

(3.76-7.36)

14.04 ±7.04

(5.69-23.33)

24.89 ± 1.37

(23.92-25.86)

9.66

6.97

9.65 ± 0.52

(9.02-10.05)

9.39

9.08 ±0.5

(8.67-9.38)

13.46

14.09 ±1.17

(12.17-16.31)

12.45 ±3.77

(10.53-15.86)

35.43 ± 16.42

(17.25-55.11)

59.512 ±3.4

(57.82-62.62)

0.73

0.92

0.97 ± 0.03

(0.94-1)

1.20

1.23 ±0.03

(1.21-1.25)

1.95

1.57 ±0.26

(1.23-2.14)

1.82 ±0.07

(1.77-1.86)

5.15 ± 1.97

(3.07-7.64)

6.03 ±1.15

(5.22-6.84)

0.89

0.95

1.01 ±0.18

(0.83-1.19)

1.05

1.36 ±0.01

(1.35-1.37)

1.68

1.99 ±0.16

(1.78-2.24)

2.22 ±0.01

(2.21-2.22)

5.54 ±2.37

(2.8-8.36)

8.84 ±0.04

(8.81-8.87)

0.66

0.94

0.92 ± 0.06

(0.86-0.98)

1.16

1.21 ±0.01

(1.2-1.22)

1.62

1.5 ±0.22

(1.13-1.82)

1.65 ±0.1

(1.58-1.72)

2.75 ± 1.06

(1.37-4.03)

4.65 ±0.41

(4.36-4.94)

IoD

0.77

0.75

0.88 ±0.12

(0.76-0.99)

1.14

1.09 ±0.03

(1.07-1.11)

1.34

1.62 ±0.09

(1.46-1.79)

1.81 ±0.07

(1.76-1.86)

3.62 ± 1.62

(1.75-5.55)

5.67 ±0.51

(5.31-6.03)

InD

—

0.43

0.57

0.78 ±0.18

(0.57-1.28)

0.76 ± 0.06

(0.72-0.8)

1.69 ±0.76

(0.76-2.61)

2.25 ±0.48

(1.91-2.59)

EnD

—

0.38

0.44

0.68 ±0.08

(0.53-0.78)

0.84 ±0.01

(0.83-0.85)

1.63 ±0.68

(0.9-2.42)

2.2 ±0.56

(1.81-2.6)

—

0.32

0.47

0.78 ±0.21

(0.51-1.4)

1.03 ±0.08

(0.97-1.09)

2.18 ±1.8

(0.44-4.14)

3.21 ±0.53

(2.84-3.59)

0.86

1.00

1.11 ±0.03

(1.08-1.13)

1.22

1.15 ± 0.08

(1.09-1.21)

1.85

1.95 ±0.32

(1.49-2.55)

2.32 ±0.23

(2.16-2.47)

3.46 ± 1.75

(1.9-5.76)

5.29 ± 1.32

(4.36-6.23)

0.75

0.69

0.77 ± 0.04

(0.73-0.8)

0.74

0.65 ± 0.05

(0.61-0.68)

0.87

1.04 ± 0.18

(0.81-1.44)

1.1 ±0.09

(1.04-1.16)

2.57 ± 1.32

(1.05-4.08)

3.28 ± 1.51

(2.21-4.35)

F1L

0.16

0.36

0.43 ± 0.06

(0.37-0.48)

0.00

0.73 ±0.01

(0.73-0.73)

0.82

1.65 ±0.26

(1.2-2.24)

2.37 ±0.13

(2.28-2.47)

6.84 ±2.53

(3.98-9.97)

10.04 ±0.35

(9.79-10.29)

H1L

—

0.66 ±0.35

(0.24-1.41)

1.41 ±0.17

(1.29-1.54)

6.4 ±2.79

(3.69-9.69)

9.63 ±0.39

(9.35-9.91)

2.44

2.94

3.22 ±0.48

(2.73-3.7)

3.48

3.24 ±0.01

(3.23-3.25)

3.91

3.75 ±0.43

(3.19-4.75)

3.26 ±0.36

(3.01-3.52)

11.74 ±5.2

(6.25-17.53)

19.49 ±0.31

(19.27-19.71)

egg D

8.8

9.55

10.63 ±0.81

(10.05-11.2)

10.57

10.86 ± 0.14

(10.76-10.96)

10.72

9.78 ±0.77

(8.67-11.18)

10.31 ±0.65

(9.85-10.77)

—

niles of T. ziegleri were found in the field, but at stage 45

with total lengths between 57.82-62.62 mm larvae still

had not reached metamorphosis.

The detailed descriptions of the available larval stages

of T. ziegleri are given in Table 2 and respective pho¬

tographs in Fig. 6. For detailed morphological measure¬

ments of the larval stages see Table 3. The overall shape

and pigmentation of the larvae remained similar through

all stages: head wider than long (with the exception of

stages 30 and 32); interocular distance bigger than inter¬

nostril distance; eye-nostril distance very similar to inter¬

nostril distance; width of tail base smaller than tail height;

tail length smaller than snout-vent length. There was no

evidence of balancers throughout the development.

Coloration in life: Body with golden yellowish-brown

ground color; venter whitish-transparent. Fingers and

toes yellow. Golden spots scattered on dorsal head and

trunk, lateral body, tail fin and axilla to throat. Gills yel¬

low with an orange-reddish hue at the edges and on the

fimbriae. Ground color turned darker with age. Shortly

after metamorphosis the skin was totally black except for

yellow fingers, toes, and ventral ridge of tail. At this time

the skin started to become less smooth and more granu¬

lar and warty.

Developmental biology of T. vietnamensis

Eggs: The record of one egg directly after deposition had

a diameter of 11.97 mm and 0.73 g of weight, while one

Amphib. Reptile Conserv.

April 2017 | Volume 11 | Number 1 | el 38

Larval development and breeding ecology of Ziegler’s Crocodile Newt

Fig. 7. Dark and light phenotypic variations of Tylototriton

vietnamensis found at the type locality. Photo M. Bernardes.

egg ready to hatch measured 10.10 mm and weighted

0.56 g. Measurements from random eggs in the field

showed an egg diameter ranging between 6.06-13.58

mm (mean 9.73 ± 1.61 mm, n = 133) and weight ranging

from 0.19-1.15 g (mean 0.48 ±0.21 g, n = 133). Eggs

were transparent and clear shortly after egg deposition

and later changed to brownish transparent.

Body shape and size of hatched larvae: range of body

length at hatching time was 15.59-17.85 mm (mean

17.04 ± 0.85 mm, n = 5). Dorsal fin well developed and

higher than head, starting at the middle of trunk; ventral

fin shorter than dorsal fin; body long and slender; snout

short and flat; gills well developed. At stage 33 two fin¬

gers were visible in the forelimb and the hind limb bud

was already visible. Toes, fingers, and joints were fully

developed at stage 41. At stage 44 gills started to atrophy.

Efts started to move to land at a size of 44.15 mm with

0.6 g of weight.

Coloration in life: ground color light yellowish ochre;

dark pigmentation on dorsal flanks, tail and head; venter

slightly transparent to creamy white with no pigmenta¬

tion; yellow spots scattered on dorsal side of head, body

and tail; fingers and toes transparent to yellowish; gills

light orange; eyes golden with black pupils. Pigmen¬

tation got darker with age turning black shortly before

metamorphosis; toes and finger tips remained yellow as

well as ventral ridge of tail. However, during field work

at the type locality of this species we came across slight

phenotypic variations, where larvae were also totally

white at older stages (Fig. 7).

Comparison with T. ziegleri: The diameter of the

gelatinous layer of the egg was bigger in I vietnamensis,

as well as sizes of hatchlings. However, the estimated

size at metamorphosis is likely bigger in T. ziegleri. The

development and body shape of larvae of T. vietnamensis

were very similar to T. ziegleri, with the exception that in

T. vietnamensis the body is more slender and elongated

and the gills more orange than reddish.

Comparisons between the development of T.

ziegleri and its congeners. Tylototriton ziegleri

showed terrestrial oviposition, while T. taliangensis

and T. cf. shanjing showed aquatic oviposition and T.

kweichowensis and T. himalayanns showed both. In

T. podichthys and T. panhai eggs were laid adhered to

vegetation, while in T. ziegleri eggs were oviposited

on the ground. One exceptional clutch of T. ziegleri

showed eggs in small aggregations, like in T. podichthys.

T. ziegleri had similar clutch sizes compared with T.

hainanensis, but they were smaller than clutch sizes of

T. kweichowensis and T. taliangensis and bigger than

those of T. vietnamensis, T asperrimus, T. wenxianensis,

and T. himalayanns. Eggs of T. ziegleri were transparent

in coloration when young and turned to yellow-

brownish when older, like in T. vietnamensis, while in

T. himalayanns eggs were greenish-yellow in color.

The comparison between sizes of ovae showed larger

diameters for T. ziegleri in relation to T. kweichowensis,

T. asperrimus, and T. podichthys. In relation to the

diameter of the gelatinous layer, T. ziegleri had similar

diameters to those of T. linyangensis and T. wenxianensis,

which were bigger than those of T. taliangensis, T.

kweichowensis, and T. podichthys, and smaller than those

of T. asperrimus. T. vietnamensis, T. cf. shanjing, and T.

himalayanns showed a wider range of egg diameter, both

bigger and smaller than those of T. ziegleri. Furthermore

egg size was related to clutch size, as species with smaller

eggs had bigger clutches and vice-versa (y = -29.68 x +

313.64; F, s = 66.85, P < 0.001; r 2 = 91.7%). At hatching

time T. vietnamensis had the largest larvae, followed in

size by larvae of T. ziegleri, T. kweichowensis, and lastly

by T. himalayanns. Size at metamorphosis seemed the

smallest for T. shanorum and T. vietnamensis, followed

by T. cf. shanjing, T. kweichowensis, T. broadoridgns,

and T. himalayanns, while in comparison T. uyenoi and

T. taliangensis had the largest sizes at metamorphosis

(Table 4).

Generally, the larvae of T. ziegleri can be distinguished

from the described larvae of the genus Tylototriton by

having: 1) a broad head (longer in T. cf. shanjing)-, 2)

the interorbital distance wider than internostril distance

(similar distances in T. cf. shanjing)-, 3) a pointed tail tip

(round in T. uyenoi, T. taliangensis, T. cf. shanjing, and T.

linyangensis)-, 4) the absence of balancers (versus present

in T. uyenoi and T. cf shanjing)', 5) dorsal fin higher than

ventral fin (almost identical height in T. linyangensis)-, 6)

tail shorter than SVL (tail longer than SVL in T. hima¬

layanns)-, 7) reddish gills (versus orange in T. vietnam¬

ensis ); 8) advanced larval stages with dark ground color

with the exception of yellow digits and ventral fin (versus

orange digits and fin in T. broadoridgns-, yellow at head,

Amphib. Reptile Conserv.

April 2017 | Volume 11 | Number 1 | el 38

Bernardes et al.

Table 4. Reproduction data of TyJototriton representatives (after Bourret 1942; Khatiwada et al. 2015; Kuzmin et al. 1994; Mudrack

2005; Nishikawa et al. 2013a, 2014, 2015; Phimmachak et al. 2015; Shen et al. 2012; Sparreboom 2014; Yang et al. 2014; Zhao

1988; Ziegler et al. 2008; and after own data for T. vietnaniensis in comparison with T. ziegJeri). * - based on the description of a

single larva; ** - based on the description of two juveniles. All measurements in mm. For abbreviations see Material and Methods.

Clutch size

(Egg nrs.)

Oviposition

site

Jelly layer

diameter

Ovum diam¬

eter

TL hatchling

Body shape

and size of

larvae

Larvae

coloration

Dilferences to

T. ziegleri

Subgenus

Yao triton:

T. asperrinms

30-52

3.0-3.4

bigger diameter of ge¬

latinous layer; ovae di¬

ameter and clutch size

smaller

T. broador-

idgus

gills start to at¬

rophy at 62 mm

orange coloration

on digits and ven¬

tral fin

yellow coloration on

digits and ventral fin in

T. ziegleri

T. hainanensis

58-90

similar clutch sizes

T. kweichow-

ensis

121-141

in water, on

moist soil

or under

large stones

nearby

water

6.2-6.5

2.3-3.4

~ 62 mm TL at

metamorphosis

bigger clutch size; di¬

ameter of gelatinous

layer, ovae and hatch¬

ling size smaller, in T.

ziegleri only terrestrial

oviposition

T. liuyangensis

on land

7.8-8.1

dorsal and ventral

fins almost identi¬

cal in height; tail

tip rounded

dorsal fin higher and

tail tip pointed in T.

ziegleri

T. panhai

adhered

to sticks

slightly

above

the water

surface

egg deposition terres¬

trial, mostly on rock

substrate in T. ziegleri

T. podichthys

adhered to

vegetation

(individu¬

ally or in

groups of

up to three)

5.0 ±0.3

2.9 ±0.2

eggs not adhered to

vegetation and egg ag¬

gregations only in one

exceptional case in T.

ziegleri ; gelatinous

layer and ovum diam¬

eter larger

T. vietnaniensis

5-85

on land

6.06-13.58

15.59-17.85

44.15 mm at

metamorpho¬

sis; slender and

elongated body

orange colored

gills

smaller clutch sizes;

diameter of gelatinous

layer and hatchling

sizes bigger; body

more slender and elon¬

gated; estimated TL at

metamorphosis for T.

ziegleri bigger; gills

more reddish than or¬

ange in T. ziegleri.

T. wenxianensis

56-81

on land or

in transition

to water

7-8

clutch size slightly

smaller

parotids, vertebral ridge, rib nodules, limbs and tail in T.

nyenoi and T. shanorum ; brighter coloration laterally in

the rib area in T. cf. shanjing) ; and 9) being less slender

than larvae of T. vietnaniensis.

Discussion

TyJototriton ziegJeri occurred at elevations congru¬

ent with the data provided by Nishikawa et al. (2013b).

Occurrences at higher elevations were also found in Cao

Bang Province, but not as high as the 1,600 m above sea

level reported by Sparreboom (2011) at Mt. Pia Oac in

Nguyen Binh district, Cao Bang Province. TyJototriton

vietnaniensis was always found at lower elevations, how¬

ever, the population from Lang Son Province at 980 m

above sea level was found higher than previous records

for this species, setting a new elevational record.

The breeding season of T. ziegJeri in northern Viet¬

nam was previously thought to last from April to May

(Nishikawa et al. 2013b); based on our new findings

this period lasts longer, from April to July. Likewise, the

breeding season of T. vietnaniensis was recorded to last

from June to July (Bohme et al. 2010), but our records

show that it starts already in April.

We present for T. ziegJeri a broader range for the dis¬

tance of clutches to water with 10-100 cm instead of

the 50-60 cm reported by Nishikawa et al. (2013b). The

average amount of eggs in a clutch unable to produce

viable offspring is still unknown, but might strongly

increase by mycosis infection, as observed in one clutch

Amphib. Reptile Conserv.

April 2017 | Volume 11 | Number 1 | el 38

Larval development and breeding ecology of Ziegler’s Crocodile Newt

Table 4 (continued). Reproduction data of Tylototriton representatives (after Bourret 1942; Khatiwada et al. 2015; Kuzmin et al.

1994; Mudrack 2005; Nishikawa et al. 2013a, 2014, 2015; Phimmachak et al. 2015; Shen et al. 2012; Sparreboom 2014; Yang et

al. 2014; Zhao 1988; Ziegler et al. 2008; and after own data for T. vietnamensis in comparison with T. ziegleri). * - based on the

description of a single larva; ** - based on the description of two juveniles. All measurements in mm. For abbreviations see Material

and Methods.

Subgenus

Tylototriton:

T. anguliceps *

dorsal fin high¬

er than ventral

fin; tail tip

pointed

fingers and toes

yellow

very similar body

shape and coloration

of larvae

T. himalayanus

26-60

in water or

on land

6-10

(greenish-

yellow in

color)

TaL < SVL;

10.9 ±0.62 62.5 ±2.67 mm

at metamorpho¬

sis

clutch size smaller;

wider range of egg di¬

ameter; smaller hatch¬

lings; in T. ziegleri

only terrestrial ovipo¬

sition, eggs transpar¬

ent and later brownish,

and SVL > TaL

T. cf. shanjing

in water

(e.g., on

submerged

vegetation)

6-10

HL > HW; IoD

> EnD; InD

~ IoD; TaL <

SVL; rounded

tail tip; pres¬

ence of balanc¬

ers; ~ 52 mm at

metamorphosis

lack of dark pig¬

mentation in the

rib area, which in

older larvae might

generate the color¬

ful flank tubercles

wider range of egg

diameter; larvae with

presence of balanc¬

ers and lighter ground

color; in T. ziegleri

terrestrial oviposition,

HW > HL and tail tip

pointed

T. shanorwn **

34 and 43 mm

(juveniles)

bright yellow col¬

oration on dorsal

head, lips, pa¬

rotids, vertebral

ridges, rib nodules,

limbs, vent region

and whole tail

estimated TL at meta¬

morphosis bigger for

T. ziegleri', juveniles of

T. ziegleri completely

black except for yel¬

low coloration on fin¬

gers and tail fin

T. taliangensis

in water (in-

250-280 dividually

on water

plants)

2 - 2.2

dorsal and ven¬

tral fins almost

in parallel; tail

tip rounded;

larvae overwin¬

ter and meta¬

morphose the

following year

with 59-72 mm

sizes

oviposition terrestrial

in T. ziegleri, egg di¬

ameter smaller; clutch

size bigger; tail tip

pointed and dorsal and

ventral fins not parallel

in T. ziegleri

T. uyenoi

anterior head,

presence of bal- parotids, vertebral

ancers in early ridge, rib nodules,

stages; tail tip limbs and tail yel-

round low in advanced

larval stages

absence of balancers,

tail tip pointed and yel¬

low coloration only on

fingers and tail fin in T.

ziegleri

in Bao Lac district.

Aquatic breeding sites with alkaline pH values and

higher carbonate hardness (Quang Ninh, Cao Bang and

Ha Giang provinces) were associated with the geological

substrate of the areas, mainly limestone rock (Sterling

et al. 2006). These karst limestone areas are character¬

ized by thin layers of surface soils, periods of severe soil

dryness due to quick drainage of water and erosion of

subsurface rock material, resulting in extensive cave sys¬

tems with underground streams. Firstly, this explains the

significantly deeper ponds found in habitats of T. ziegleri

and secondly the deficiency of soil, leaving the parent

rock exposed and explaining the choice of substrate for

oviposition. Tylototriton vietnamensis on the other hand

occurs on soils with granite parent rock material (Ber-

nardes et al. 2017) which are more acidic and have low

cation exchange capacity (Ulrich 1991). Ponds inhabited

by T. ziegleri had higher nitrite and nitrate concentrations

than ponds inhabited by T. vietnamensis. High levels of

these nutrients have usually an anthropogenic origin, like

leaching of nitrogen from manure and mineral fertiliz¬

ers from upstream villages or agricultural fields. High

concentrations of nitrate and nitrite can have negative

effects on aquatic amphibian larvae, although there are

interspecific differences in species’ sensitivity (Marco et

al. 1999), which at this point do not seem to negatively

affect the investigated species.

The additional finger found in one individual at stage

44 can have numerous causes, as malformations in

amphibians have not yet been fully investigated (Blaus-

tein and Johnson 2003). Polydactyly was, for exam-

Amphib. Reptile Conserv.

April 2017 | Volume 11 | Number 1 | el 38

Bernardes et al.

pie, associated with ultraviolet radiation (Ankley et al.

2000, 2002), chemical contamination (Burkhart et al.

1998; Gardiner and Hoppe 1999), and parasitic infection

(Johnson et al. 1999; Sessions and Ruth 1990). However,

developmental abnormalities found in amphibian popu¬

lations have typical frequencies of 0-3% and are only

considered abnormally high when exceeding 5% (Piha et

al. 2006). Although held observations revealed that this

type of malformation is quite common among adults of

Tylototriton in Vietnam (M. Bernardes, pers. obs.), more

studies have to be conducted to conclude whether these

abnormalities are above natural levels.

The diagnostic characteristics of the different

developmental stages in T. ziegleri corresponded well

to the staging system adopted by Grosse (1997, 2013).

Noticeable differences concern the earlier development

of hind limbs (at stage 36 hind limbs had already

developed three toes, while according to Grosse the hand

development is finished before hind limb buds occur)

and the absence of balancers. Balancers are usually

present in salamander larvae that develop in lentic

habitats sustaining the hypotheses that these structures

are adaptive to still waters and non-functional in flowing

waters (Crawford and Wake 1998). This theory does

not seem to be verified in T. vietnamensis, a species that

breeds in lentic habitats, but could explain the absence of

balancers in T. ziegleri, a species able to reproduce both

in lentic as in lotic habitats.

The body length of hatchlings of T. ziegleri was on

average 30% smaller than the 21-22 mm reported by

Sparreboom et al. (2011). According to these authors

the larvae completed their metamorphosis at sizes of

43-62 mm. In our collection the biggest larva had not

completed metamorphosis at stage 45 with 60 mm total

length. However, length at metamorphosis seems to be

a variable feature in Tylototriton representatives and

apparently also depends on parameters such as feeding

regime, temperature, etc. Total lengths in eight metamor-

phlings of T. vietnamensis reared in captivity by F. Pas-

mans varied between 49.9 and 65.8 mm. Further, obser¬

vations in captive reared T. wenxianensis larvae revealed

large variation in the stages at which the larvae hatched

and consequently also in the total length at hatching (F.

Pasmans, pers. obs.).

The morphological similarity within the T. asperrimus

group in terms of body shape and coloration of adults

makes it especially difficult to tell species apart (Stuart

et al. 2010). Morphological similarity is even higher in

larvae as this study shows. The larval development of T.

ziegleri is still not completely recorded, as several stages

still are unknown. Nevertheless our data allow clear mor¬

phological comparisons of developmental stages within

this genus. Egg capsule diameter seems to be larger in

species with terrestrial oviposition compared to species

with aquatic oviposition. We therefore hypothesize that

eggs of species with terrestrial oviposition are character¬

ized by an extra thick gelatinous layer to prevent exsic¬

cation of the developing larva, and therefore this trait

might be a good indicator for the species’ behavior. Tylo¬

totriton ziegleri had the biggest clutches of all land-lay¬

ing species and amongst the sub-genus Yaotriton, while

clutches of T. vietnamensis are among the smallest. The

wide ranges in clutch sizes seen in T. vietnamensis and

T. cf. shanjing might be related to the big sample size in

the first case and associated with hidden cryptic diversity

in the second. These comparisons must be regarded with

caution, since accumulating evidence suggests that the

description of the larval development of T. cf. shanjing

by Ziegler et al. (2008) was based on a species complex

(e.g., Nishikawa et al. 2013a). More information on the

larval development in the genus Tylototriton is certainly

needed.

Conclusion

Tylototriton ziegleri is a small-ranged species known

only from four localities in the North of Vietnam, none of

them within protected areas. The species is not yet listed

in the IUCN Red List, although it is likely under threat

of extinction. Like T. vietnamensis it is in demand for

the international pet trade as well as the Chinese market

(Rowley et al. 2016). Despite morphological conserva¬

tism in particular within larval stages, our data clearly

confirm contrasting habitat requirements between these

cryptic species, both in adults and larval stages. There¬

fore, our results provide useful guidance to establish

proper captive conditions for these two species with

strongly deviating breeding requirements. This is in par¬

ticularly important as Tylototriton is known for its cryptic

diversity, as it can be seen for example with what was in

the past thought to be the single species T. shanjing. From

this morphological cryptic group several species have

been described, like T. panhai and T. nyenoi (Nishikawa,

Khonsue, Pomchote, and Matsui, 2013), T. anguliceps

(Le, Nguyen, Nishikawa, Nguyen, Pham, Matsui, Ber¬

nardes, and Nguyen, 2015), and T. podichthys (Phim-

machak, Aowphol, and Stuart, 2015), while T. v. pulcher-

rima was considered to be conspecific (Nishikawa et al.

2013a). Meanwhile the T. shanjing complex is widely

distributed in zoological gardens, but origin and specific

identification is in most cases uncertain, as well as infor¬

mation about potential captive hybridization. It is virtu¬

ally impossible to identify representatives of the T. shan¬

jing group without a comprehensive genetic screening.

This negative impact on proper conservation breeding

measures is yet aggravated by the lack of information

regarding origin, natural history data, and data on differ¬

ent habitat adaptations in the field.

Our study describes the different ecological adapta¬

tions to strongly contrasting environmental conditions of

two morphologically similar species. We highlight the

necessity to improve the knowledge on the natural his¬

tory of the Tylototriton species, not only for enhanced ex

situ measures (viz. husbandry and conservation breeding,

Amphib. Reptile Conserv.

April 2017 | Volume 11 | Number 1 | el 38

Larval development and breeding ecology of Ziegler’s Crocodile Newt

see Ziegler et al. 2016), but also for in situ approaches,

such as supporting the establishment of new reserves, or

extending the area of existing ones, as the populations of

T. ziegleri from Quan Ba and Bac Quang districts occur

in the vicinity of protected areas (Bat Dai Son Nature

Reserve in Ha Giang Province and Cham Chu Nature

Reserve in Tuyen Quang Province, respectively) but are

not included in one.

Acknowledgements. —We are grateful to the direc¬

torates and staff of Tay Yen Tu, Yen Tu, Dong Son - Ky

Thuong and Bat Dai Son nature reserves, Forest Protec¬

tion Departments of Bac Giang, Quang Ninh, Lang Son,

and Ha Giang provinces for support of our field work

and issuing the required permits. We cordially thank S.V.

Nguyen, C.X. Le (IEBR, Hanoi), T. Pagel and C. Lands-

berg (Cologne Zoo) for support of our research. We

thank M. van Schingen and H.T. An for their assistance

in the field and H.T. Ngo for laboratory assistance. This

research was partially funded by the Cologne Zoo (Ger¬

many), the European Association of Zoos and Aquaria

(EAZA), the Deutsche Gesellschaft fur Herpetologie

und Terrarienkunde (DGHT), the Amphibian Conserva¬

tion Fund of German Zoo Associations, private partici¬

pants in the German-speaking region as well as Stiftung

Artenschutz, and the University of Cologne.

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Larval development and breeding ecology of Ziegler’s Crocodile Newt

Marta Bernardes is a Ph.D. candidate at the Zoological Institute of the University of Cologne

and the Cologne Zoo, Germany. She has a M.S. degree in Conservation Biology from Lisbon

University, Portugal. Since 2007 she has been engaged in the research of amphibians and reptiles

and their natural environment with a main interest in ecology and conservation. In 2010 she joined

the working group of Thomas Ziegler and initiated ecological research projects in Southeast Asia

with a main focus on the Salamandridae family from Vietnam.

Anna Rauhaus started her career at the Aquarium/Terrarium Department of the Cologne Zoo

in May 2011 and is head keeper of the Terrarium section since 2014. Her focus of expertise is

keeping and breeding of amphibians, monitor lizards, snakes, and crocodilians as well as behavioral

training. She also trains keepers and helps to build amphibian and reptile facilities within the frame

work of Cologne Zoo’s conservation projects in Vietnam. Since 2011 she has been involved in

35 herpetological publications with a focus on zoo biology, with ten of them dealing with captive

breeding, larval development, and diversity of tropical species, in particular Vietnamese amphibians.

Clara Michel performed her bachelor thesis “Larval development and ecological niche of Ziegler’s

Crocodile Newt (Tylototriton ziegleri ),” which was submitted in February 2015 at the University of

Cologne, under the supervision of Profs. Thomas Ziegler and Michael Bonkowski, as well as co¬

supervised by Marta Bernardes and Anna Rauhaus.

Cuong The Pham is a Ph.D. candidate and researcher of the Institute of Ecology and Biological

Resources (IEBR) - Vietnam Academy of Science and Technology (VAST). He is member of the

Cologne Zoo’s Biodiversity and Nature conservation projects in Vietnam. Cuong has published

several papers, mainly dealing with Vietnams’ herpetodiversity. Cuong is very experienced in

biodiversity and field research and conducted numerous field surveys in Vietnam.

Truong Quang Nguyen is a researcher at the Institute of Ecology and Biological Resources (IEBR),

Vietnam Academy of Science and Technology (VAST) and is a member of the Biodiversity and

Nature Conservation projects of the Cologne Zoo. He finished his Ph.D. in 2011 at the Zoological

Research Museum Alexander Koenig (ZFMK) and the University of Bonn, Germany (DAAD

Fellow). From 2011 to 2014 he worked as a postdoctoral student in the Zoological Institute at the

University of Cologne. Truong has conducted numerous field surveys and is the co-author of seven

books and more than 150 papers relevant to the biodiversity research and conservation in Southeast

Asia. His research interests are systematics, ecology, and phylogeny of reptiles and amphibians from

Southeast Asia.

Minh Due Le has been working on conservation-related issues in Southeast Asia for more

than 15 years. His work focuses on biotic surveys, wildlife trade, and conservation genetics of

various wildlife groups in Indochina. He is currently working on projects which characterize

genetic diversity of highly threatened reptiles and mammals in the region. Minh has pioneered the

application of molecular tools in surveying critically endangered species in Vietnam. Minh has long

been involved in studying the impact of the wildlife trade on biodiversity conservation in Vietnam,

and is developing a multidisciplinary framework to address the issue in the country.

Frank Pasmans is a veterinarian and director of the laboratory of veterinary bacteriology and

mycology at Ghent University (Belgium). He has had a lifelong obsession for amphibians, notably

urodeles. His research currently focuses on fungal infections in amphibians. By studying fundamental

processes of host - pathogen - environment interactions, this research aims at developing long-term

sustainable measures to mitigate the impact of fungal diseases on amphibian communities.

Amphib. Reptile Conserv.

April 2017 | Volume 11 | Number 1 | el 38

Bernardes et al.

Michael Bonkowski is Professor for Terrestrial Ecology in the Institute of Zoology at the

University of Cologne. His research spans from soil biodiversity and function to sustainable soil

management, and more recently included studies on the biodiversity and function of tropical

ecosystems in Southeast Asia. One aim is to better understand the mechanisms of community

assembly of amphibians and reptiles in tropical rain forests of Vietnam and Laos. This research

is focusing on patterns of geographic genetic differentiation and attributes of the ecology and

life history of amphibians and reptiles. This work is placed in the context of ecosystem-level

consequences of biodiversity loss due to factors such as habitat degradation and destruction, and

on shifts in tolerances to changing temperatures as expected by global change.

Thomas Ziegler has been the Curator of the Aquarium/Terrarium Department of the Cologne

Zoo since 2003 and is the coordinator of the Cologne Zoo’s Biodiversity and Nature Conservation

Projects in Vietnam and Laos. Thomas studied biology at the University Bonn (Germany),

and conducted his diploma and doctoral thesis at the Zoological Research Museum Alexander

Koenig in Bonn, with focus on zoological systematics and amphibian and reptile diversity. He

has been engaged with herpetodiversity research and conservation in Vietnam since 1997. As

a zoo curator and project coordinator he tries to combine in situ and ex situ approaches, viz., to

link zoo biological aspects with diversity research and conservation, both in the Cologne Zoo, in

rescue stations and breeding facilities in Vietnam and in Indochina’s last remaining forests. He is

Professor at the Zoological Institute of Cologne University. Since 1994, Thomas has published

370 papers and books, mainly dealing with herpetodiversity.

Amphib. Reptile Conserv.

April 2017 | Volume 11 | Number 1 | el 38

Official journal website:

amphibian-reptile-conservation.org

Amphibian & Reptile Conservation

11(1) [General Section]: 88-92 (e139).

On the distribution of the Himalayan Stripe-necked Snake

Liopeltis rappi ( Gunther, 1860) (Serpentes: Colubridae) in

Nepal

^antosh Bhattarai, 2 Kul Bahadur Thapa, ^ina Chalise, hashish Gurung, 1 Chiranjibi Prasad

Pokheral, 3 Naresh Subedi, 2 Tej Bahadur Thapa, and 4 Karan Bahadur Shah

1 National Trust for Nature Conservation-Biodiversity Conservation Center (NTNC-BCC), Ratnanagar-18, Sauraha, Chitwan-44204, NEPAL

2 CentraI Department of Zoology’, Tribhuvan University, Kirtipur, Kathmandu, NEPAL 3 National Trust for Nature Conservation-Khumaltar, Lalitpur,

NEPAL 4 Natural History Museum, Tribhuvan University, Swoyambhu, Kathmandu, NEPAL

Abstract .—The distribution of the Himalayan Stripe-necked Snake (Liopeltis rappi) is poorly

documented. We summarize the distribution of this little known snake in Nepal and provide a new

locality record from Kabilas, Chitwan, Nepal. Compiled observations presented here suggest that

the species is more widely distributed and we call for additional surveys and a systematic inventory.

Keywords. Chitwan-Annapurna landscape, conservation, reptile, biodiversity, South Asia, Squamata

Citation: Bhattarai S, Thapa KB, Chalise L, Gurung A, Pokheral CP, Subedi N, Thapa TB, and Shah KB. 2017. On the distribution of the Himalayan

Stripe-necked Snake Liopeltis rappi (Gunther, 1860) (Serpentes: Colubridae) in Nepal. Amphibian & Reptile Conservation 11(1) [General Section]:

88-92 (el 39).

NoDerivatives 4.0 International License, which permits unrestricted use for non-commercial and education purposes only, in any medium, provided

the original author and the official and authorized publication sources are recognized and properly credited. The official and authorized publication

credit sources, which will be duly enforced, are as follows: official journal title Amphibian & Reptile Conservation ; official journal website <amphibian-

reptiie-conservation.org>.

Received: 24 January 2017; Accepted: 15 May 2017; Published: 30 May 2017

Introduction

The Himalayan Stripe-necked Snake Liopeltis rappi

(Gunther, 1860) is a small, slender, non venomous

snake distributed through the Himalayas of Nepal and

India (Smith 1943; Wallach et al. 2014). Gunther (1860)

described this species from Sikkim, India and originally

it was described as Ablabes rappi. Later, Wall (1921)

placed it in the genus Liopeltis. Reporting of its distri¬

bution has been sporadic both in time and space and

it is an uncommonly encountered species. Previously

it has been reported in India from Sikkim (Chhetri et

al. 2011; Gunther 1860), Darjeeling (Wall 1909), and

Himanchal Pradesh (Saikia et al. 2007; Smith 1943) in

India and Chitwan (Schleich and Kastle 2002; Shres-

tha 2001), Khotang, Terhathum, Shankhuwasabha (Rai

2003), Kaski (Shah and Tiwari 2004), and Palpa (Thapa

2016) in Nepal. The information on diversity and species

richness of the ophidian fauna in Nepal is scanty. Nepal¬

ese snakes are represented by Palearctic, Ethiopian, and

Oriental species (Schleich and Kastle 2002). Recently,

Kastle et al. (2013) listed the occurrence of 82 species in

Nepal, and questioned the occurrence of 14 snake spe¬

cies included in the list of Schleich and Kastle (2002)

and Shah and Tiwari (2004) due to several taxonomic

revisions. Most of the herpetological expeditions have

focused in eastern and central Nepal. These expeditions

usually report new taxa or new distribution records for

the country. For example, Sharma et al. (2013) and Pan-

dey (2015) recently added two new snake species record

viz Bnngarus sindanus walli (Boulenger 1897) and Oli-

godon cylcurus (Cantor 1839) respectively for Nepal. We

here add one more significant record of Liopeltis rappi

from Kabilas, Chitwan which is a part of the Chitwan-

Annapurna Landscape.

The Chitwan-Annapurna Landscape (CHAL) is

located in central Nepal and it includes all or part of 19

districts covering an area of 32,057 km 2 , with elevations

ranging from 200 m to 8,091 m asl. The landscape is

drained by eight major perennial rivers and their tributar¬

ies from the broader Gandaki River system. The CHAL

experiences a range of climates from subtropical in the

lowlands to alpine in the high mountains, and cold and

dry in the trans-Himalayan region. It is suggested that

climate change is a major cause behind changes in floral

and faunal diversity in the CHAL (MoFSC 2015). In this

regard, twelve permanent plots have been established in

the CHAL to study the vulnerability of species due to cli¬

mate change or other factors. These plots lie in Barand-

abhar, Kaule, and Kabilas of Chitwan district, Tilakpur

and Asardi of Palpa district, Panchase of Syangja district

and in Mustang district.

Amphib. Reptile Conserv.

May 2017 | Volume 11 | Number 1 | el 39

Bhattarai et al.

Fig. 1. Liopeltis rappi from Dhodeni, Kabilas, Chitwan. Photo Santosh Bhattarai.

Materials and Methods

We examined a recently dead specimen of Liopeltis rappi

found at Kabilas-09, study site at Dhodeni, Chitwan

(GPS: 27.78418°N 84.51605°E, elevation, 978 m, WGS:

1984) during survey work monitoring climate effects on

one of the permanent plots. It was found dead on a trail

in an abandoned cultivated field at 12:19 h on 16, July

2016. The cause of death was not readily apparent.

We recorded morphometric and meristic data for this

specimen: dorsal scale rows at three points, approxi¬

mately one head length posterior to the head, midbody,

and one head length anterior to the vent. Ventral scales

were counted following Dowling (1951), subcaudals and

dorsal scales. Snout Vent Length (SVL) and Total Length

(TTL) were measured with a thread, later scaling it to a

metallic ruler. Digital camera Canon-65 X optical zoom

was used for photographic record, description of col¬

ors, and patterns. The broad habitat type and plant spe¬

cies were recorded at the place where the specimen was

located.

Results

The small snake measured 462 mm SVL, 572 mm TTL

and was identified as Liopeltis rappi (Fig. 1) based on the

following combination of characteristics: head short and

not distinct from the neck, round pupil, nostrils large and

between two nasals, dorsal scales 15:15:15, all smooth;

ventrals 176; subcaudals 60; all paired. There were six

supralabials, with the 3 rd -4 th contacting the eye, 5 th larg¬

est, a single preocular, and two post oculars. The dorsal

Table 1. Locality records of Liopeltis rappi in Nepal.

S.N.

Locality

Habitat

Coordinates

Elevation

District

Source

Lasune

Small town

27.14586°N, 87.46302°E

2,250 m

Tehrathum

Rai 2003

Chisapani /Nagdhunga

Paddy field

26.96709°N, 86.88333°E

1,600 m

Khotang

Rai 2003

Makalu Barun NP

—

27.66266°N, 87.10458°E

2,200 m

Sankhuwasabha

Rai 2003

Ghandruk

—

28.46638°N, 83.71421°E

2,972 m

Kaski

Shah and Tiwari 2004

Khaliban

Waste land

27.85624°N, 83.84418°E

813m

Palpa

Thapa 2016

Chappani, Jhirbhanjyang

Waste land

27.89579°N, 83.56964°E

1,056 m

Palpa

Thapa 2016

Bandipokhara, Lipindevi

Waste land

27.86725°N, 83.50192°E

1,498 m

Palpa

Thapa 2016

Tansen, Bhusaldanda

Small town

27.87159°N, 83.55696°E

1,292 m

Palpa

Thapa 2016

Khanigaun, Khiluadada

Waste land

27.91685°N, 83.55087°E

1,139 m

Palpa

Thapa 2016

Dhodeni/Kabilas

Cultivated land

27.78418°N, 84.51605°E

978 m

Chitwan

This study

Amphib. Reptile Conserv.

May 2017 | Volume 11 | Number 1 | el 39

Distribution of Liopeltis rappi in Nepal

80WE

81°0'0"E

82°0’0"E

83°0 , 0"E

84°0'0"E

85°0’0"E

86°0'0"E

87"0'0"E

88°0'0"E

Fig. 2. Distribution of Liopeltis rappi in Nepal.

color was uniform coffee brown and the venter was yel¬

low. The specimen represents the 10 th locality record of

L. rappi for Nepal. Figure 2 provides an overview of dis¬

tribution of L. rappi in Nepal and Table 1 summarizes the

locality records.

Habitat and herpetological community : The speci¬

men was recorded in a seasonally abandoned cultivated

land (Fig. 3). The broad habitat type of the locality was

broadleaf mixed forest. The plant species recorded at the

site were Lantana camara, Eupatorium sp., and Agera-

tum sp. Other herpetofauna recorded at the same local¬

ity comprised of anurans: Duttaphrynus melanostic-

tus (Schneider 1799), Microhyla ornata (Dumeril and

Bibron 1841), Sphaerotheca maskeyi (Schleich and

Anders 1998), S. breviceps (Schneider 1799), Polyped-

ates maculatus (Gray 1830), Amolops marmoratus (Blyth

1855), and Fejervarya sp. (Bolkay 1915); lizards: Calo-

tes versicolor (Daudin 1802), Eutropis carinata (Schnei¬

der 1801), E. macularia (Blyth 1853), and Laudakia

tuberculata (Gray 1827); and snakes: Amphiesma sto-

latum (Linnaeus 1758), Dendrelaphis tristis (Daudin,

1803), and Sibynophis collaris (Gray 1853).

Discussion

Distribution: The closest published records to the CHAL

for Liopeltis rappi are Ghandruk, Kaski 100 NW (Shah

and Tiwari 2004) and Palpa 95 km W (Thapa 2016). Ear-

Amphib. Reptile Conserv.

Her published records of L. rappi (Gunther 1860; Ander¬

son 1871; Boulenger 1890; Smith 1943; Krammer 1977;

Shrestha 2001) did not mention data on locality records

for Nepal. Shah (1995) and Schleich and Kastle (2002)

reported the occurrence of L. rappi in Sauraha, Chitwan.

Later, Shah, and Tiwari (2004) verified the record from

Chitwan as a wrongly identified specimen and mentioned

the new occurrence locality of this species from Ghan¬

druk, Kaski. Captain (2010) also questioned the occur¬

rence of L. rappi in Sauraha, Chitwan as this species is

thought to be distributed at higher elevations. We agree

with Shah and Tiwari (2004) and Captain (2010), hence,

remove the occurrence of L. rappi from Sauraha and

report our observation locality, Kabilas-09, Dhodeni as

the first confirmed record from Chitwan. Our observa¬

tion locality is 978 m asl and ranges within the elevation

record of Thapa (2016). The elevational records range

from 813 m to 2,972 m, demonstrating that the species

probably exhibits a wider distribution in Nepal.

Conservation status: The IUCN (2016) has assessed L.

rappi as a Data Deficient (DD) species and its popula¬

tion trend unknown. Of ten distribution localities, two

localities Ghandruk and Makalu Barun National Park are

within protected areas. Other localities are designated as

either cultivated land, waste land, or small towns, and

these sites were identified as important for the conserva¬

tion of the species in Nepal, as the cultivated lands are

being mechanized and villages are growing larger and

May 2017 | Volume 11 | Number 1 | el 39

Bhattarai et al.

Fig. 3. Habitat of Liopeltis rappi at Dhodeni, Kabilas, Chitwan.

Photo Santosh Bhattarai.

into towns. The present record of the dead specimen

from cultivated land tends to show the possible igno¬

rance of local people in the survival of the species. Thapa

(2016) recorded five specimens from Palpa, of which

four were found killed by local people and a single live

specimen from Khaliban. People in this area kill snakes

at the moment they encounter them as standard practice

in the culture. This rampant killing of snakes, includ¬

ing L. rappi by local people, is an observed threat in the

CHAL. All snakes are believed by the local people to be

venomous despite the fact that only 17 species of snakes

in Nepal are venomous (Sharma et al. 2013). Outreach

activities among farmers, local communities, in schools,

and colleges should focus on the good ecosystem func¬

tion of snakes and basic identification tools of snakes

would be instrumental in better protecting the snake

fauna of the CHAL. Our finding indicates that a coun¬

trywide detailed herpetological survey would be benefi¬

cial to better understand the ecology, distribution pattern,

threats, and conservation status of L. rappi in Nepal.

Acknowledgements. —This is an offshoot of the

Project “ Climate Change Monitoring on Permanent

Plots in Chitwan-Annapurna Landscape ” funded by

USAID-Hariyo Ban Program/NTNC. We thank Harka-

man, Kapil, Pratigya, Biraj, Trishna, Pramod, Deepu,

Tika, Om, Binod, and Ramesh for field support. NTNC-

BCC provided logistic support and field coordination.

We acknowledge Mark O’Shea and George Zug for com¬

ments on earlier drafts and Frank Tillack for related lit¬

erature and discussion which benefitted the manuscript.

We thank Pabitra Gotame for her help in the field and

map preparation.

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ML and Tambe S. Biodiversity of Sikkim: Exploring

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scape_nepal.pdf [Accessed: 26 May 2017],

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evance in Nepal: Study on Species, Epidemiol¬

ogy of Snake Bite and Assessment of Risk Factors

of Envenoming and Death. Ph.D. Dissertation, der

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of Nepal. Koenigstein: Koeltz Scientific Books, Ger-

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USA. 1,237 p.

Santosh Bhattarai is currently working as a Conservation Officer at the National Trust for Nature

Conservation- Biodiversity Conservation Center (NTNC-BCC), Sauraha, Chitwan, Nepal. He is

particularly interested in understanding evolutionary and ecological drivers of amphibians and reptiles

for which species diversify and evolve through time and space.

KB Thapa is a graduate in zoology from Tribhuvan University of Nepal. He is impassioned about

wildlife research, conservation, and particularly herpetology. His study on amphibians and reptiles

in his M.Sc. thesis was guided by this same passion. In that study he reported a total of forty eight

species of herpetofauna of which sixteen species had national and/or global altitudinal/latitudinal and/

or longitudinal distribution range extension. At present, he is engaged with one of the leading national

non-government organizations working in the field of nature conservation and research.

Lina Chalise holds a M.Sc. in Biodiversity and Eenvironmental Mmanagement. She is currently

working as a conservation officer at the National Trust for Nature Conservation- Biodiversity

Conservation Center, Sauraha, Chitwan, Nepal.

Aashish Gurung graduated in Environmental Science and Natural Resource Management from

Kathmandu University. Currently, he works as a Conservation Officer at the National Tmst for Nature

Conservation- Biodiversity Conservation Center, Sauraha, Chitwan, Nepal.

Chiranjibi Prasad Pokheral currently works as a Project Manager at the National Tmst for Nature

Conservation-Biodiversity Conservation Center, Sauraha, Chitwan, Nepal. He completed his Ph.D. in

2012 and has more than two decades of experience in species conservation and management in Nepal.

He is focused on managing biodiversity projects in Chitwan-Parsa complex.

Naresh Subedi completed his Ph.D. in 2012 and is currently based at the NTNC-central office,

Kathamandu and works as a Conservation Program Manager. His earlier research was focused on the

impact of invasive species on native wild animals and subsequent conservation measures.

Dr. T.B. Thapa is a professor of wildlife at Central Department of Zoology, Tribhuvan University in

Kathmandu, Nepal. He has made significant contribution in wildlife research and in the formulation of

national conservation strategies of many species. He has also been instrumental in guiding many new

researchers fascinated in wildlife.

Karan Bahadur Shah is a Professor of Zoology at Tribhuvan University, Natural History Museum,

Swoyambhu, Kathamadu. He has described several reptiles in Nepal and authored a book ‘Herpetofauna

of Nepal: Conservation Companion ” published by IUCN Nepal.

Amphib. Reptile Conserv.

May 2017 | Volume 11 | Number 1 | el 39

Official journal website:

amphibian-reptile-conservation.org

Amphibian & Reptile Conservation

11(1) [General Section]: 93-107 (e140).

The herpetofauna of central Uzbekistan

1A *Thomas Edward Martin, 12 Mathieu Guillemin, 12 Valentin Nivet-Mazerolles, 12 Cecile Landsmann,

^Jerome Dubos, 1>2 Remy Eudeline, and 3 James T. Stroud

1 Emirates Centre for the Conservation of the Houbara, Urtachol massif Karmana Shirkat farm, Navoi Region, REPUBLIC OF UZBEKISTAN

2 Reneco for Wildlife Presentation, PO Box 61 741, Abu Dhabi, UAE. 3 Department of Biological Sciences, Florida International University, Miami,

Florida, USA

Abstract .—The diverse habitats of central Uzbekistan support a rich herpetofaunal community, but

distributions and relative abundances of the species comprising this community remain poorly

known. Here, we present an annotated species inventory of this under-explored area, with detailed

notes on distributions and population statuses. Fieldwork was concentrated in southern Navoi and

western Samarkand provinces, although some records were also made in the far north of Navoi

province, near the city of Uchkuduk. Data were collected between March and May/June in 2011,

2012, and 2013, with herpetofaunal records being made opportunistically throughout this period.

Survey effort was concentrated in semi-desert steppe habitats, especially the Karnabchul steppe

area located to the south of the city of Navoi and an expanse of unnamed steppe located to the north

of Navoi. Further records were made in a range of other habitat types, notably wetlands, sand dune

fields, and low rocky mountains. Total fieldwork equated to approximately 8,680 person-hours of

opportunistic survey effort. In total, we detected two amphibian and 26 reptile species in our study

area, including one species classified as Globally Vulnerable by the IUCN. We present distributional

data supporting the first record of regional range extensions of five species from within our study

area. Our results represent the most detailed data concerning reptile and amphibian diversity and

distributions produced from Uzbekistan in recent years. We conclude by recommending that further,

systemized survey work needs to be conducted within the area to supplement our findings with

more robust estimates of species abundances supported by more detailed information on species-

habitat relationships.

Keywords. Central Asia, faunistics, inventory, steppe, distribution, survey

Citation: Martin T, Guillemin M, Nivet-Mazerolles V, Landsmann C, Dubos J, Eudeline R, and Stroud J. 2017. The herpetofauna of central Uzbekistan.

Amphibian & Reptile Conservation 11(1) [General Section]: 93-107 (el40).

NoDerivatives 4.0 International License, which permits unrestricted use for non-commercial and education purposes only, in any medium, provided

the original author and the official and authorized publication sources are recognized and properly credited. The official and authorized publication

credit sources, which will be duly enforced, are as follows: official journal title Amphibian & Reptile Conservation ; official journal website <amphibian-

reptiie-conservation.org>.

Received: 08 April 2016; Accepted: 01 Aug 2016; Published: 14 June 2017

Introduction

Central Asia (defined in this study as the five Central Asian

Republics and Afghanistan) encompasses a wide range of

habitats, which in turn support a rich and, in some areas,

highly endemic biodiversity. The region’s ecological

importance is reflected by it encompassing five “global

200” terrestrial ecosystems (Olson and Dinerstein 1998)

and two biodiversity hotspots (Myers 2003). Despite this

importance, the region’s fauna remains poorly explored

(Aye et al. 2012). Increased research interest in Central

Asia in recent years has resulted in a significant increase

in information regarding some taxonomic groups, nota¬

bly birds (Aye et al. 2012; Wassink 2015), although little

contemporary field-based work has examined the diver¬

sity and distributions of other taxa, including reptiles and

amphibians (herpetofauna).

Correspondence. *tom martin 2010@yahoo. co. uk

Recent information regarding regional-scale distribu¬

tions and habitat associations of Central Asian amphib¬

ian and reptile communities is scarce, with the limited

available data focusing on particular countries and habi¬

tats. Large-scale herpetofaunal distribution studies have

been completed for Turkmenistan (Schkammakov et

al. 1993; Tuniyev et al. 1999) and parts of Kazakhstan

(Lambert 2002). Trans-national biogeographical patterns

for lizard communities in the region’s mountains have

also been examined (Bobrov 2005). Detailed descrip¬

tions, however, remain largely lacking for entire habi¬

tat types and countries within Central Asia. Very little

community-level information exists on the semi-desert

steppe habitats that predominate in non-montane areas of

southern Central Asia, and recent outputs from Uzbeki¬

stan—the most populous country in the region—are

restricted to a small number of species-specific ecology

Amphib. Reptile Conserv. 93 June 2017 | Volume 11 | Number 1 | e140

Martin et al.

Fig. 1. Map of our study areas within central Uzbekistan. Inset shows the study area within the entirety of Uzbekistan. Notations

represent the following locations: K = Karnabchul steppe, N = northern steppe, S = Sarmysh nature park, T = Lake Tudakul, A =

Lake Aydarkul, U = Uchkuduk study area.

papers (Lagarde et al. 2002, 2003; Ikramov and Azimov

2004; Clemann et al. 2008). Prior to these, the only exist¬

ing herpetology resources from Uzbekistan are a number

of regional-scale Russian-language texts dating back to

the Soviet era, which remain largely inaccessible to the

international scientific community (e.g., Bannikov 1971;

Rustamov 1981; Rustamov and Shcherbak 1986).

As well as a lack of community-level research,

knowledge relating to the statuses of individual spe¬

cies in Uzbekistan is also restricted to a limited range of

resources. These include IUCN (2016) species distribu¬

tion maps (which are lacking for the majority of Central

Asian species), coarse-grained spatial range maps pro¬

vided by The Reptile Database (2016), and a Soviet-era

Russian-language text (Bannikov 1971), and regional-

scale atlas maps provided in guidebooks to the Western

Palearctic as a whole (Sindaco and Jeremcenko 2008;

Sindaco et al. 2013). This general lack of zoological

knowledge appears to have had an impact on regional

conservation strategies, with steppe and semi-deserts in

Central Asia having been noted as being poorly-repre¬

sented in existing protected area networks (Chemonics

International 2001).

In this study we attempt to address this knowledge-gap

by providing an annotated checklist of the herpetofauna

community of central Uzbekistan, based on opportunis¬

tic records made while conducting surveys of the Asian

Houbara Bustard (Chlamydotis macqueenii). These

records represent the first recent data regarding herpeto-

faunal community composition in this part of Uzbekistan

and from the Central Asian semi-desert steppe habitats

where survey work was focused. We also provide records

from a number of other habitats occurring in the region,

notably sand dunes, low mountains, and wetlands.

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The herpetofauna of central Uzbekistan

Materials and Methods

Study site: Fieldwork was concentrated principally in

semi-desert grassland habitats (Plate 1) which predomi¬

nate in central Uzbekistan (Aye et al. 2012; World Wild¬

life Fund 2013). This habitat is invariably referred to

locally as “steppe.” While true steppe is a less arid eco¬

system found in higher latitudes, for ease of reference

we henceforth use this term for the semi-desert habitats

in our study area. The principal purpose of the authors’

work in these habitats was to monitor populations of

Asian Houbara Bustard (Chlamydotis macqueenii). Bus¬

tard surveys were concentrated in two large expanses of

steppe; the Karnabchul steppe region (BirdLife Interna¬

tional 2016a) located in southern Navoi province and

far western Samarkand province, and a large expanse

of steppe located to the north of Navoi city (Fig. 1).

While completing this survey work we opportunistically

recorded herpetofauna wherever possible (see below).

When not committed to completing bustard surveys in

the steppe, we surveyed a number of other habitats, most

notably the extensive sand dune fields which occur spo¬

radically within our two main steppe study areas, the low

mountains of the Sarmysh nature park (BirdLife Interna¬

tional 2016c) and the wetland habitats found along the

western shore of Lake Aydarkul (BirdLife International

2016b) and eastern shore of Lake Tudakul (BirdLife

International 2016d) (Fig. 1). A single five-day visit was

also made from 29 April to 03 May 2013 to a large area

of steppe and dunes in northern Navoi province, near the

city of Uchkuduk, on the fringes of the Kyzylkum des¬

ert (Fig. 1).

Altitude throughout this spatially extensive study

area ranges from 230 m along the shore of Lake Tuda¬

kul to 1,993 m at the highest peak of the mountainous

Sarmysh area. Most of the area consists of slightly undu¬

lating steppe, however, where elevation typically var¬

ies between 300-380 m. The area possesses a continen¬

tal climate characterized by hot, dry summers, and cold

winters with frequent thaws (Glazirin et al. 1999). Mean

temperatures vary from 33 °C in July to 1.9 °C in Janu¬

ary. Average annual rainfall is approximately 126 mm,

with an average of 32 mm falling in February (the wet¬

test month) and <1 mm falling in July (the driest month)

(Emirates Centre for the Conservation of Houbara 2013,

unpublished data). The area possesses a highly complex

geology—the result of its location on a tectonic collision

zone (Hendrix and Davis 2001).

Vegetation in steppe habitats is dominated by hardy

shrubs of the Artemesia genus, interspersed with other

shrub assemblages, while sand dune habitats are dom¬

inated by a variety of psammophytic plant species

(Makhmudovich 2006). Mountainous areas within the

Sarmysh Nature Park area possess an Irano-Turanian

vegetation assemblage characterized by small, hardy

shrubs and trees, notably those of the genera Primus and

Pistacia (Aye et al. 2012).

Fieldwork: Herpetofaunal records were made by the

authors over the course of three fieldwork seasons span¬

ning the spring (and in one case the early summer) months

of 2011, 2012, and 2013. Fieldwork dates ran from 13

March to 27 May in 2011, 04 March to 31 May in 2012,

and 15 March to 25 June in 2013. These spring and early

summer months represent the optimal time for complet¬

ing herpetofauna surveys in Central Asia given that most

species hibernate during the cold winter months, and that

some species return to hibernation prior to the hottest

summer months and do not resume activity until the fol¬

lowing spring (Lagarde et al. 2003). All records were col¬

lected opportunistically, rather than via formalized sur¬

vey work. These opportunistic records were made in a

number of ways. During formal Bustard survey hours

within steppe habitats, records were kept of all herpe¬

tofaunal species observed while driving between estab¬

lished survey sites during the day, or encountered on foot

at these survey sites. Records within steppe habitats were

also made driving along roads at night, and from casual

exploration during the middle of the day when conditions

were not suitable for formal survey work. Exploration

was also conducted in sand dune habitats, the low moun¬

tains of Sarmysh Nature Park, and the shores of Lake

Aydarkul and Tudakul outside of formal survey work.

This exploration involved extensive driving and walking

on foot in these habitats, both in the day and at night, and

noting any records made, as well as targeted searching in

microhabitats likely to support specialized herpetofaunal

species, such as dune crests, rocky gullies, and well-veg¬

etated river banks. While this opportunistic record mak¬

ing did not follow a systematic survey methodology, her¬

petofauna species were still actively searched for by the

authors, except when formal bustard survey work was

being conducted. We estimated the approximate person-

hours of survey effort represented by our opportunistic

records by calculating the number of days each surveyor

spent in the field multiplied by eight (the average number

of hours per day each surveyor spent in the field, exclud¬

ing hours spent conducted formalized bustard counts).

Data analysis: After the completion of survey work

we carefully identified all species detected by our sur¬

vey effort, using all existing field guides and distribu¬

tion atlases encompassing Central Asia (Bannikov 1971;

Sindaco and Jeremcenko 2008; Sindaco et al. 2013). All

species identifications were then independently verified

by JS. We also sought additional species verifications

from Dr. Tatjana Dujsebayeva at the Kazakhstan Institute

of Zoology for all records of Eiyx Sand boas—a group

that can be particularly difficult to separate in the field.

We then compiled an inventory of all identified species

following the taxonomy provided by Frost (2014) for

amphibians and the Reptile Database (2016) for reptiles.

We recorded the conservation status of each species in

our inventory following the most recent IUCN Red List

database (IUCN 2016). We also noted whether each spe-

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June 2017 | Volume 11 | Number 1 | e140

Martin et al.

cies was endemic to Central Asia as defined by species

descriptions given by the IUCN (2016).

Categorical abundance estimates for each species

were then assigned based on frequency of records. The

following categories were used: abundant (typically

recorded multiple times every day in suitable habitat);

common (typically recorded around once per day); fairly

common (typically recorded once per week); uncommon

(recorded less than five times per season on average) and

rare (known from less than five records overall). A data

deficient (DD) category was used for species exclusively

found in the Sarmysh mountains which were too briefly

explored to provide meaningful abundance estimates.

Finally, we attempted to determine whether our

records for each species in our inventory represented

an extension to their known spatial range. We assessed

two different magnitudes of range extensions: national

range extensions for species which had not been pre¬

viously reported from Uzbekistan, and regional range

extensions for species previously reported as occurring

in Uzbekistan, but not within our study area, by exist¬

ing distribution maps. Potential range extensions were

assessed by comparing each species in our inventory

with records reported in herpetological papers from the

region (Lagarde et al. 2002, 2003; Ikramov and Azimov

2004; Clemann et al. 2008), biological summaries of

sites of special scientific interest (Birdlife International

2016a,b,c,d), existing distribution maps provided by the

IUCN (2016), the Reptile Database (2016), and the two

volumes of the only recent species distribution atlas cov¬

ering Central Asia (Sindaco and Jeremcenko 2008; Sind-

aco et al. 2013). We also consulted distribution maps pro¬

vided in Bannikov (1971), although this fairly inaccessi¬

ble Russian-language text does not provide range maps

for all species in the region.

Results

Opportunistic survey effort in our study area constituted

approximately 8,680 person-hours. A total of 28 species

were detected by this survey effort—two amphibian spe¬

cies and 26 reptile species. All these species are native

to the study area, with no current records of introduced

or invasive species in the region. Only a single species

(Testudo horsfieldii ) is considered Globally Threatened

or Near-Threatened by the IUCN (2016). However, only

five species (17.9%) are assigned a definitive threat cat¬

egory—all other species in our inventory remain unas¬

sessed or are considered data deficient. No species in

our inventory is restricted exclusively to Central Asia,

although several, such as the Turkestan Agama ( Para-

laudakia lehmanni ), are almost entirely confined to the

region and therefore considered “near-endemic.” All

species we detected were previously known to occur

in Uzbekistan (IUCN 2016; Reptile Database 2016),

thus we report no national range extensions. However,

records for five species ( Eremias scripta , Eryx miliaris,

Hemorrhois ravergieri, Natrix tessellata , and Echis cari-

Amphib. Reptile Conserv.

natus) represent regional range extensions within central

Uzbekistan. A full summary of species detected in our

study area is provided in Table 1, with the descriptions

below providing more detailed information for each spe¬

cies in our inventory.

AMPHIBIANS

Green Toad: Bufotes viridis (Laurenti 1768) (Bufoni-

dae) (Plate 5)

A widespread species found from western Europe

to Kazakhstan. The taxonomy of species appears to be

unclear, with some sources splitting Central Asian popu¬

lations as B. variabilis , and others separating the popula¬

tions of the southern and eastern Central Asian steppes

still further as B. pewzowi (Ficetola and Stock 2016),

although we retain the Frost (2013) nomenclature. A noc¬

turnal species, it is locally abundant in our study area,

and is typically found around permanent and ephem¬

eral water sources throughout the area’s steppe habitats.

However, individuals were also occasionally encoun¬

tered in the open steppe at least one km from any known

water sources. The species has been previously reported

as occurring in the study area (Bannikov 1971; BirdLife

International 2016c).

Marsh Frog: Pelophylax ridibundns (Pallas 1771) (Ran-

idae) (Plate 6)

A widely distributed species found throughout much

of Europe and western Asia. It is abundant in unpolluted,

non-saline water sources, particularly in agricultural

fields and drainage ditches located on the peripheries of

steppe habitats and the two large lake ecosystems, and

small mountain streams in the Sarmysh region. Unlike

B. viridis , this species was never observed far away from

water. Pelophylax ridibundus is previously known to

occur within the study area (Bannikov 1971; BirdLife

International 2016c).

REPTILES

Russian Tortoise: Testudo horsfieldii (Gray 1844) (Tes-

tudinidae) (Plate 7)

The only Chelonian species found in our study area,

T. horsfieldii is restricted to Central Asia and Iran, and is

listed as Vulnerable by the IUCN (2016). The species is

abundant in the steppe habitats of our study area, with

multiple individuals typically being seen every day in

this habitat from between early March when they emerge

from hibernation to mid-June, when their hibernation

resumes (Lagarde et al. 2003). It was by a large margin

the most frequently encountered herpetofaunal species

during the course of our fieldwork. The species has been

previously mapped as occurring in central Uzbekistan

(Bannikov 1971; Sindaco and Jeremcenko 2008; Bird-

Life International 2016a-d) although the atlas map in

Sindaco and Jeremcenko (2008) does not note its pres-

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The herpetofauna of central Uzbekistan

Table 1. Summary of amphibian and reptile species recorded in our central Uzbekistan study area between 2011 and 2013. New

range extensions are marked in bold. Species marked f are assessed as Threatened or Near Threatened by the 1UCN (2016).

Abundance estimates are denoted as follows: A = abundant; C = common; Fc = fairly common; U = uncommon; R = rare; DD = data

deficient. Broad locales in the “locations” column are denoted as follows: A = Lake Aydarkul; K = Karnabchul steppe; N = northern

steppe area; S = Sarmysh nature park; T = Lake Tudakul; U = Uchkuduk area. All taxonomy follows Frost (2014) for amphibians

and the Reptile Database (2016) for reptiles.

Class Order Family

Common name

Scientific name

Abundance Locations

Amphibia Anura Bufoidae

Green Toad

Bufotes viridis

K, N

Ranidae

Marsh Frog

Pelophylax ridibundus

K, S

Reptilia Testudines Testudinidae

f Russian Tortoise

Testudo horsfieldii

K, N, U

Squamata (Sauria) Agamidae

Turkestan Agama

Paralaudakia lehmanni

K, S

Brilliant Ground Agama

Trapelus agilis

K, N

Sunwatcher Toadhead Agama Phrynocephahis helioscopus

K, N

Secret Toadhead Agama

Phrymocephalus mystaceus

N, U

Lichtenstein’s Toadhead Agama Phiynocephahis interscapularis Fc

Geckkonidae

Common Wonder Gecko

Teratoscincus scincus

Russian Bent-toed Gecko

Tenuidactylus fedtschenkoi

Caspian Bent-toed Gecko

Tenuidactylus caspius

K, N

Lacertidae

Striped Racerunner

Eremias lineolata

N, U

Rapid Fringe-toed Lizard

Eremias velox

K, N

Reticulate Racerunner

Eremias grammica

N, U

Sand Racerunner

Eremias scripta

N, U

Scincidae

Berber Skink

Eumeces schneideri

Anguidae

European Glass Lizard

Pseudopus apodus

Varanidae

Desert Monitor

Varan us griseus

K, N, U

Squamata (Serpentes) Boidae

Tartar Sand Boa

Eryx tataricus

K, N

Dwarf Sand Boa

Ervx miliaris

K, N

Colubridae

Spotted Desert Racer

Platyceps karelini

K, N

Wadi Racer

Platyceps rhodorachis

Diadem Snake

Spalerosophis diadema

Spotted Whipsnake

Hemorrhois ravergieri

Lamprophiidae

Steppe Ribbon Racer

Psammophis lineolatus

K, N

Natricidae

Dice Snake

Natrix tessellata

A, T

Elapidae

Central Asian Cobra

Naja oxiana

K, S

Viperidae

Saw-scaled Viper

Echis carinatus

K, N

ence in the Uchkuduk area, where our records indicate

the species to occur.

Turkestan Agama: Paralaudakia lehmanni (Nikolsky

1896) (Agamidae) (Plate 8)

Found only in southern Central Asia and Pakistan.

Paralaudakia lehmanni is a fairly common species in

suitable habitats within our study area, being regularly

sighted on boulders and crevasses in rock faces within

low mountain habitats on the periphery of the Karnab¬

chul steppe and in the Sarmysh Nature Park. The species

has been previously mapped as occurring in the study

area (Sindaco and Jeremcenko 2008).

Brilliant Ground Agama: Trapelus agilis (Olivier

1807) (Agamidae) (Plate 9)

A widespread species found in the Caucasus, much of

Central Asia, Iran, and the Indian subcontinent. Trape¬

lus agilis is abundant in our study area, being typically

observed multiple times per day in open steppe habitats

and in the low mountains on the edge of the Karnabchul

area, once the species has emerged from hibernation in

mid-late March. It is frequently observed basking on top

of Artemisia shrubs. Trapelus agilis has been previously

mapped as occurring in our study area (Sindaco and Jer¬

emcenko 2008; BirdLife International 2016a-d).

Sunwatcher Toadhead Agama: Phrynocephahis

helioscopus (Pallas 1771) (Agamidae) (Plate 10)

This species is found in southern European Russia and

much of Central Asia. It is fairly common in the open

steppe habitats of our study area, and has occasionally

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Martin et al.

Plate 1. Semi-arid “steppe” habitats (Photograph - CL).

Plate 3. Low rocky mountains (Photograph - JD).

Plate 5. Bufotes viridis (Photograph - TM).

Plate 7. Testudo horsfieldi (Photograph - TM).

Plate 2. Vegetated sand-dune habitats (Photograph - CL).

Plate 6. Pelophylax ridibundus (Photograph - TM).

Plate 8. Paralaudakia lehmanni (Photograph - TM).

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The herpetofauna of central Uzbekistan

been observed in sand dunes. It is previously recorded

as occurring throughout central Uzbekistan (Bannikov

1971; Sindaco and Jeremcenko 2008; IUCN 2016; Bird-

Life International 2016a,c).

Secret Toadhead Agama: Phrynocephalus mystaceus

(Pallas 1776) (Agamidae) (Plate 11)

This distinctive, large-bodied agama possesses a wide

but patchy distribution across southern European Rus¬

sia, much of Central Asia, and eastern Iran. A strict sand

dune species, it is uncommon in our study area, typically

being observed only once or twice per research season

in very specific areas, such as the dune fields within the

northern steppe area and in the vicinity of Uchkuduk

where we observed both single individuals and apparent

pairs. The species has been previously mapped as occur¬

ring throughout our study area (Sindaco and Jeremcenko

2008).

Lichtenstein’s Toadhead Agama: Phrynocephalus

interscapularis (Lichtenstein 1856) (Agamidae) (Plate

12 )

A small agamid found in all the Central Asian Repub¬

lics and the northern border areas of Iran. Phrynocepha¬

lus interscapularis is fairly common in our study area. As

with P. mystaceus it is restricted to sand dunes, although

the two species display strong niche separation within

this habitat. Clemann et al. (2008) describe how P. mys¬

taceus occupies dune crest microhabitats, while P. inter¬

scapularis occupies mid-dune microhabitats and swales

between dunes—a pattern our observations corrobo¬

rate. The species has been previously mapped as occur¬

ring throughout our study area (Sindaco and Jeremcenko

2008).

Common Wonder Gecko: Teratoscincus scincus (Schle-

gel 1858) (Gekkonidae) (Plate 13)

A widespread species found throughout Central Asia,

Iran, Pakistan, and parts of the Arabian Peninsula. Ter¬

atoscincus scincus is a strictly nocturnal species which

appears to be fairly common within our study area

(although this abundance estimate could be a product of

our sampling effort—see Discussion). We only detected

T. scincus in the northern steppe habitats of our study

area, where it was encountered regularly on roads while

driving at night, and occasionally on foot when walking

in the open steppe after dusk. The species is indicated

to occur throughout our study area by Bannikov (1971),

although our records represent a slight range extension

to the atlas maps produced by Sindaco and Jeremcenko

(2008).

Russian Bent-toed Gecko: Tenuidactylus fedtschenkoi

(Strauch 1887) (Gekkonidae) (Plate 14)

Restricted to Central Asia and northern border areas

of Pakistan and Iran. Tenuidactylus fedtschenkoi is fairly

common in rocky mountain habitats on the peripheries of

our two main steppe study areas, as well as on isolated

rocky outcrops within the steppe. The species was almost

always observed within a short distance of holes and cre¬

vasses in the rock face, where they retreated when dis¬

turbed. Tenuidactylus fedtschenkoi has been previously

mapped as occurring in our study area (Bannikov 1971;

Sindaco and Jeremcenko 2008; BirdLife International

2016a,c).

Caspian Bent-toed Gecko: Tenuidactylus caspius

(Eichwald 1831) (Gekkonidae) (Plate 15)

A widespread species found throughout southern Cen¬

tral Asia and around the basin of the Caspian Sea. Tenu¬

idactylus caspius is common in our study area, being

found in a variety of habitats including cliff faces, iso¬

lated rocky outcrops in the steppe, abandoned ruins, and

within inhabited buildings. It is mapped as occurring

throughout our study area by the IUCN (2016), although

our records represent modest extensions to the distribu¬

tion maps provided by Bannikov (1971) and Sindaco and

Jeremcenko (2008).

Striped Racerunner: Eremias lineolata (Nikolsky

1897) (Lacertidae) (Plate 16)

This species is found throughout southern Central

Asia and in north-eastern Iran. It is uncommon in our

study area, being found only in extensive areas of veg¬

etated sand dunes, often alongside populations of E.

scripta and E. grammica. It has been previously mapped

as occurring throughout our study area (Sindaco and Jer¬

emcenko 2008; BirdLife International 2016d).

Rapid Fringe-toed Lizard: Eremias velox (Pallas 1771)

(Lacertidae) (Plate 17)

A widespread species found throughout Central Asia,

southern European Russia, and Iran. Eremias velox is

abundant in our study area and was by far the most fre¬

quently observed Eremias species, occurring at high

densities throughout the region’s open steppe habitats.

The species has been previously mapped as occurring

throughout our study area (Sindaco and Jeremcenko

2008).

Reticulate Racerunner: Eremias grammica (Lichten¬

stein 1823) (Lacertidae) (Plate 18)

Distributed across Central Asia, western China, and

north-eastern Iran, E. grammica is an uncommon species

in our study area. It was only observed in vegetated sand

dune habitats similar to those inhabited by E. lineolata

and E. scripta. It has been previously reported as occur¬

ring throughout the study area (Bannikov 1971; Sindaco

and Jeremcenko 2008).

Sand Racerunner: Eremias scripta (Strauch 1867)

(Lacertidae) (Plate 19) * Regional range extension

A widely, although patchily, distributed species found

throughout Central Asia, western China, Iran, and Paki-

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Martin et al.

Plate 9. Trapelus agilis {Photograph - TM).

Plate 11. Phrynocephalus mystaceus {Photograph - RE).

Plate 13. Teratoscincus scincus {Photograph - TM).

Plate 15. Tenuidactylus caspius {Photograph - TM).

Plate 10. Phrynocephalus helioscopus {Photograph - VNM).

Plate 12. Phrynocephalus interscapularis {Photograph - VNM).

Plate 14. Tenuidactylus fedtschenkoi {Photograph - TM).

Plate 16. Ere mi as lineolata {Photograph - MG).

stan. It is rare within our study area, having been recorded

a total of four times. It appears to inhabit similar habi¬

tats to E. lineolata and E. grammica, being found locally

within well-vegetated sand dune fields. While E. scripta

has been noted as occurring at Lake Tudakul (BirdLife

International 2016d), our records of this species in the

northern steppe and Uchkuduk areas represent regional

range extensions to its known distribution, with none of

our consulted sources noting its occurrence in these areas

of central Uzbekistan.

Amphib. Reptile Conserv.

100

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The herpetofauna of central Uzbekistan

Berber Skink: Enmeces schneideri (Daudin 1802)

(Scincidae) (Plate 20)

A widespread species found from North Africa to the

Indian sub-continent. Within our study area, the species

is known from a single individual found on rocky scree

in a canyon within the mountains of the Sarmysh Nature

Park on 26 May 2012. As we only spent a few days pros¬

pecting for herpetofauna in the Sarmysh area, it is hard

to ascertain whether this species is genuinely rare here,

or whether it is more common than our limited survey

effort in appropriate habitats suggests. At the very least,

it appears to be very localized and habitat-specific within

central Uzbekistan. We never encountered the species in

the steppe or in mountainous ecosystems immediately

adjacent to the steppe (unlike other rocky habitat special¬

ists such as Paralaudakia lehmanni and Tenuidactylus

fedtschenkoi ). The species has been previously mapped

as occurring in our study area (Sindaco and Jeremcenko

2008; BirdLife International 2016c).

European Glass Lizard: Pseudopus apodus (Pallas

1775) (Anguidae) (Plate 21)

A widespread species found from eastern Europe to

eastern Kazakhstan. Within our study area it is known

only from three records of live individuals in tall grass

meadows in close proximity to streams within the Sar¬

mysh Nature Park, and four dead individuals seen in vil¬

lages on the outskirts of Sarmysh (two having been hit

by vehicles, one being carried in a plastic bag by a local,

and one found washed up on the bank of a small river).

As with Enmeces schneideri , the species appears to be

very localized within our study area, but our low survey

effort within the Sarmysh mountains makes it difficult to

accurately estimate its relative abundance within its lim¬

ited range. It has been previously mapped as occurring in

our study area (Bannikov 1971; Sindaco and Jeremcenko

2008; BirdLife International 2016c).

Desert Monitor: Varanus griseus (Daudin 1803) (Vara-

nidae) (Plate 22)

This large-bodied monitor lizard is widely distributed

in arid habitats from north Africa to the Indian Subconti¬

nent. It is a common species in our study area, typically

being observed at least once per day in open steppe habi¬

tat and sand dunes from mid-April onwards, when the

species emerges from hibernation. It has been previously

recorded as occurring throughout our study area by Ban¬

nikov (1971) although Sindaco and Jeremcenko (2008)

do not map its occurrence in the Uchkuduk area, where

we observed the species several times.

Tartar Sand Boa: Eryx tataricus (Lichtenstein 1823)

(Boidae) (Plate 23)

A widespread Asian species found from Iran through

southern Central Asia to western China and Mongo¬

lia. It is an uncommon inhabitant of open steppes and

sand dunes in our study area. Its appearance seems to

be highly variable in central Uzbekistan. The species has

been previously mapped as occurring throughout central

Uzbekistan (Sindaco et al. 2011).

Dwarf Sand Boa: Eryx miliaris (Pallas 1773) (Boidae)

(Plate 24) * Regional range extension

A less widely-distributed species than E. tataricus.

Eryx miliaris is largely confined to Central Asia, with

its range extending slightly into Iran and southern Euro¬

pean Russia. An uncommon species in our study area, it

was detected about as frequently, and in similar habitats

to, E. tataricus. Our records of this species constitute a

regional range extension. BirdLife International (2016d)

noted its presence at Lake Tudakul, but none of our con¬

sulted sources indicate the species to occur in Karnab-

chul or the northern steppe areas where we have detected

it.

Spotted Desert Racer: Platyceps karelini (Brandt 1838)

(Colubridae) (Plate 25)

This species is found in all the Central Asian Repub¬

lics, Iran, and Pakistan. It is uncommon in our study area,

being occasionally recorded in the open steppe and sand

dune habitats of Karnabchul and the northern steppe

areas. It has been previously mapped as occurring in our

study area (Bannikov 1971; Sindaco et al. 2011; BirdLife

International 2016d).

Wadi Racer: Platyceps rhodorachis (Jan 1865) (Colub¬

ridae) (Plate 26)

A widely but disjunctively distributed species, found

in east Africa, Arabia, Iran, Central Asia, and the north¬

ern Indian sub-continent. It appears to be rare in our study

area, being known from a single predated and partially

consumed individual (see Plate 26) found on the north¬

ern border of the Karnabchul steppe, close to a range of

rocky foothills, on 17 May 2012. The species has been

previously mapped as occurring in our study area (Ban¬

nikov 1971; Sindaco et al. 2011).

Diadem Snake: Spalerosophis diadema (Schlegel 1837)

(Colubridae) (Plate 27)

A widely distributed species occurring from west and

north Africa throughout the Middle East to Central Asia.

This fairly large snake species is uncommon in our study

area, typically being observed two or three times per sea¬

son in open steppe and in close proximity to inhabited

areas. The species is noted as occurring throughout the

study area by Bannikov (1971), although Sindaco et al.

(2011) do not map its occurrence in central Uzbekistan.

Spotted Whipsnake: Hemorrhois ravergieri (Menetries

1832) (Colubridae) (Plate 28) * Regional range extension

Distributed in Turkey, the Caucasus, Iran, Cen¬

tral Asia, and western China. It is a rare species in our

study area, known from a single record of an individ¬

ual observed in open steppe habitat on 11 May 2011 in

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Plate 17. Eremias velox (Photograph - TM).

Plate 18. Eremias grammica (Photograph - RE).

Plate 19. Eremias script a (Photograph - MG).

Plate 20. Eumeces schneideri (Photograph - TM).

Plate 22. ITar anus griseus (Photograph - RE).

Plate 23. Eiyx tataricus (Photograph - TM).

Plate 24. Eryx miliaris (Photograph - MG).

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The herpetofauna of central Uzbekistan

our northern steppe study area. This record represents a

regional range extension for H. ravergieri. The species is

not mapped as occurring in central Uzbekistan by Ban¬

nikov (1971) and Sindaco et al. (2011), and while its

presence has been noted in the Lake Tudakul area (Bird-

Life 2016d), it does not appear to have been previously

reported as occurring further north in Uzbekistan.

Steppe Ribbon Racer: Psammophis lineolatus (Brandt

1838) (Lamprophiidae) (Plate 29)

Found in all the Central Asian Republics, western

China, and Iran. Psammophis lineolatus is fairly common

in the steppe and sand dune habitats of our study area,

being observed on average about once per week. The

species was, overall, the most frequently observed snake

species during the course of our fieldwork. The species

has been previously mapped as occurring throughout our

study area (Bannikov 1971; Sindaco et al. 2011).

Dice Snake: Natrix tessellata (Laurenti 1768) (Natric-

idae) (Plate 30) * Regional range extension

A widespread species ranging from central Europe to

Egypt, the Middle East, and western China. It was uncom¬

mon within our study area, typically being observed a few

times per season in freshwater habitats along the shores

of Lake Aydarkul and Lake Tudakul, as well as in irriga¬

tion canals in cultivated land in the vicinity of Tudakul.

Our records for this species represent a regional range

extension to its known distribution. Bannikov (1971) and

Sindaco et al. (2011) do not map its presence anywhere

in our study area. BirdLife International (2016d) notes its

presence at Lake Tudakul, but our records from the west¬

ern shore of Lake Aydarkul appear to be entirely new.

Central Asian Cobra: Naja oxiana (Eichwald 1831)

(Elapidae) (Plate 31)

Restricted to southern Central Asia and borders areas

of Iran and the Indian sub-continent. Naja oxiana is an

uncommon species in our study area. It was typically

observed about once per season in steppe habitats close

to low foothills in the Karnabchul area (including one

dead individual found on a road measuring 156 cm), and

in the low mountains of Sarmysh. The species has been

previously noted as occurring throughout the study area

(Bannikov 1971; Sindaco et al. 2011; BirdLife Interna¬

tional 2016a,c).

Saw-scaled Viper: Echis carinatus (Schneider 1801)

(Viperidae) (Plate 32) * Regional range extension

A widespread species found in the Middle East, Iran,

Central Asia, and the Indian sub-continent. This highly

venomous viper is common in suitable habitats within

our study area, being observed almost daily within a

few specific areas of human habitation on the edge of

the Karnabchul area, and occasionally in the open hab¬

itats of Karnabchul and the northern steppe areas. Our

records represent a regional range extension for E. cari¬

natus. Bannikov (1971) only maps the species as occur¬

ring along Uzbekistan’s western border with Turkmeni¬

stan, while Sindaco et al. (2011) note its occurrence only

in the south-west and extreme north of the country.

Discussion

The results presented in this paper represent by far the

most extensive recent account of Uzbekistan’s herpeto¬

fauna, resulting from nearly 8,700 person-hours of obser¬

vational sampling. Our results are not only based on sig¬

nificant survey effort, but are also derived from extensive

surveys encompassing a broad and representative range

of Central Uzbekistan’s habitats, in contrast to other

recent papers from the country (i.e., BirdLife Interna¬

tional 2016a-d; Lagarde et al. 2002, 2003; Ikramov and

Azimov 2004; Clemann et al. 2008). Thus, this study pro¬

vides a much-needed update to the understanding of the

diversity and distributions of the region’s understudied

herpetofauna, especially with regards to the five reported

regional range extensions. The results of this study allow

for some appreciation of the relative diversity of the cen¬

tral Uzbekistan herpetofauna compared to that found in

surrounding countries. While differences in sampling

effort and scope need to be considered, the 28 species

detected in our study area do not appear to represent as

diverse a species assemblage as steppe/foothill mosaic

landscapes found further south in Turkmenistan. In Turk¬

menistan, 49 species have been recorded from a broadly

comparable site (Tuniyev et al. 1999) but do seem to sup¬

port higher overall richness compared to a steppe/foot¬

hills site in southern Kazakhstan, further to the north

(Lambert 2002), where just 17 species were recorded.

This tentatively suggests that a latitudinal diversity gra¬

dient exists across the Turian Plain—the biogeographi-

cal zone encompassing most non-mountainous areas

of Central Asia (Djamali et al. 2010). Determining the

precise delimitations of such a gradient may represent

an interesting avenue for future regional research. The

results of this study also highlight the extent to which

the conservation status of the Central Asian herpetofauna

remains heavily neglected. For example, our review of

IUCN (2016) classifications revealed that only 18% of

species in the area possess definitive threat evaluations.

An improved understanding of Central Asia’s herpeto¬

fauna—and biodiversity in general—is therefore cru¬

cial in order to both better understand the consequences

of, and to mitigate, the heavy environmental pressures

facing the region. Regionally, key threats to the Central

Asian herpetofauna include climate change and habitat

degradation due to overgrazing and other unsustainable

land uses (Christensen et al. 2004; Lioubimtseva et al.

2005), as well as unsustainable collection for the inter¬

national pet trade (Kuzmin 1994; Cheung and Dudgeon

2006; Robinson et al. 2015). With regards to our cen¬

tral Uzbekistan study sites specifically, habitats are also

threatened by extensive mining operations seeking to

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Martin et al

Plate 25. Platyceps karelini (Photograph - MG).

Plate 26. Platyceps rhodorachis (Photograph - CL).

Plate 29. Psammophis lineolatus (Photograph - MG). Plate 30. Natrix tessellata (Photograph - TAP).

Plate 31. Naja oxiana (Photograph - TM).

Plate 32. Echis carinatus (Photograph - TM).

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The herpetofauna of central Uzbekistan

exploit the abundant mineral resources (including gold,

uranium, and natural gas) found throughout much of the

area (F. Andrianova, pers. comm.). The relative impacts

of these threats are expected to differ between habitats.

Species well adapted to human-modified landscapes

(notably T. caspius and E. carinatus) or rocky hills (such

as T. fedtschenkoi , P. lehmanni, and E. schneideri) are in

likelihood less vulnerable to habitat loss and degradation

than species occurring exclusively in the steppe, where

grazing and mining activities are concentrated.

While the results of this study represent a valuable

contribution to regional herpetological knowledge, these

records exhibit a strong bias towards the steppe ecosys¬

tems forming the focus of our simultaneous Bustard sur¬

vey work. While considerable observational effort was

directed towards central Uzbekistan’s other major habi¬

tats (with the exception of the mountains of Sarmysh—

see Materials and Methods), they were not explored to

the same extent as the steppe. This could mean that some

of the relative abundance values assigned for non-steppe

species in Table 1 are influenced by reduced survey

effort in dune, wetland, and mountain habitats, and thus

underestimate true relative abundances. As the bulk of

our observations were made during daylight hours, this

could also be true for strictly nocturnal species, such as

Teratoscincus scincus or Echis carinatus. Concentrating

survey effort in the spring may also have led to underesti¬

mating the relative abundance of any species possessing

peak activity periods in the summer months.

Reduced survey efforts in non-steppe habitats may

have led to some species in these areas being unrecorded

due to a simple lack of detection. For example, the Blunt-

nosed Viper ( Macrovipera lebetina ) was not conclusively

observed during our fieldwork, but has been previously

recorded as occurring in central Uzbekistan (Bannikov

1971; BirdLife International 2016c). A long (50 cm>),

fat-bodied snake carcass observed along a roadside in

low hills near the village of Kyzulkuduk in the north¬

ern steppe area in May 2011 may have been this spe¬

cies. However, this specimen was not closely examined

and no photograph was taken to verify its identity. Other

species indicated to potentially occur in the region, but

were not detected by our survey effort, include Eremias

arguta, Crossobamon eversmanni, and Gloydius halys

(Bannikov 1971; Sindaco et al. 2011).

Conclusions

This study provides a valuable overview of the diverse

herpetofaunal community of central Uzbekistan. How¬

ever, further work in the area is required to extend

the region’s species inventory, and subsequently pro¬

vide more accurate species abundance estimates, while

improving knowledge of species-habitat relationships.

Further field surveys are encouraged to focus on the

region’s non-steppe habitats (especially montane ecosys¬

tems), which were under-represented by survey effort in

this study, and to employ more systematic survey meth¬

ods than were possible here. This study highlights the

significant lack of information regarding the conserva¬

tion status of most species occurring in the steppes and

other habitats of central Uzbekistan, and we strongly rec¬

ommend that IUCN threat status auditors utilize all avail¬

able resources to address the apparent knowledge gap

occurring in this part of Central Asia.

Acknowledgements. —This project was completed

under the supervision of the Emirates Centre for the

Conservation of the Houbara (ECCH), which is managed

by Reneco for Wildlife Preservation (www. reneco.org).

We greatly thank H.H. Sheikh Mohammed Bin Rashid

Al Maktum, funder of the ECCH, for his support. We

are also grateful for the support of Frederic Lacroix,

Mohamed Beljafla, Adeline Cadet, and Yves Hingrat,

respectively general manager of, director of ECCH, proj¬

ect manager of ECCH, and head of Reneco’s ecology and

conservation department. We also thank staff involved in

methods design and data collection—particularly ecol¬

ogy coordinators: Eric Le Nuz and Cedric Ferlat, and

field workers: Andy Simpkin, Olga Lukshyts, Vladimir

Bezmelnitsyn, Alfonso Godino, Yury Bakur, Jesse Gab¬

bard, Edward Mongin, Maksim Tarantovich, and Val¬

erie Dombrovski. Finally, we extend thanks to Dr Tat-

jana Dujsebayeva of the Kazakhstan Institute of Zoology

for assistance with verifying our species records of Eryx

Sand Boas, and two anonymous reviewers for their use¬

ful and constructive comments.

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The herpetofauna of central Uzbekistan

Thomas Edward Martin is a conservation biologist based with Reneco for Wildlife Preservation,

UAE. He has an interest in the biogeography and ecology of species inhabiting poorly-explored parts

of the tropics and sub-tropics, in particular the steppes of Central Asia and the rainforests of Indonesia.

Mathieu Guillemin is a field biologist based with Reneco for Wildlife Preservation, UAE. He has a

long-standing interest in the herpetofauna of the arid zones of North Africa and Central Asia, having

spent nearly a decade completing fieldwork in these ecosystems. He currently spends a large part of the

year as a project manager in the Betpak-Dala steppe, Kazakhstan.

Valentin Nivet-Mazerolles is a conservation biologist with a wide range of interests, having recently

completed varied forms of fieldwork in Morocco, Uzbekistan, Kazakhstan, and Crozet Island in the

Southern Indian Ocean. He currently works for the French National Reserve service in the Jura region.

Cecile Landsmann is a field biologist based with Reneco for Wildlife Preservation, UAE. She has

worked for many years in the arid steppe ecosystems of North Africa and Central Asia, and has

developed a strong broad interest in the ecology of these regions. She currently spends much of her

year working as a project manager in Central Uzbekistan.

Jerome Dubos is an experienced conservation biologist who first worked as a field technician in the

Central Asian steppes in 2009, and has returned to the region most years since. He also has extensive

experience conducting field surveys in France, North Africa, and the Middle East. He is currently

working on the LIFE+ Petrels pro ject on Reunion Island, working towards the conservation of the two

endemic Petrel species found there.

Remy Eudeline is an enthusiastic herpetologist and a biology teacher, currently based on Mayotte

Island in the Indian Ocean. He has developed a strong interest in the herpetofauna of Mayotte, in

particular its poorly-studied endemic blind snakes.

James T. Stroud is a Ph.D. candidate at Florida International University. He has a broad range of

research interests, often revolving around investigating how ecological processes may explain

evolutionary patterns, most commonly using herpetofauna as model species and study systems. He has

extensive herpetological fieldwork experience in Europe, the Neotropics, and South-East Asia.

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