4446 (4): 477 500 Article …...A time-calibrated Bayesian Inference analysis was implemented in...

ZOOTAXA

ISSN 1175-5326 (print edition)

ISSN 1175-5334 (online edition)Copyright © 2018 Magnolia Press

Zootaxa 4446 (4): 477–500

http://www.mapress.com/j/zt/Article

https://doi.org/10.11646/zootaxa.4446.4.4

http://zoobank.org/urn:lsid:zoobank.org:pub:6EA8D4BA-2A4A-40C2-8D4D-CA5D8A36D37B

A new Cyrtodactylus Gray, 1827 (Squamata, Gekkonidae) from the Shan Hills

and the biogeography of Bent-toed Geckos from eastern Myanmar

L. LEE GRISMER1,8, PERRY L. WOOD, JR.2, MYINT KYAW THURA3, EVAN S. H. QUAH4,

MARTA S. GRISMER1, MATTHEW L. MURDOCH5, ROBERT E. ESPINOZA6 & AUNG LIN7

1Herpetology Laboratory, Department of Biology, La Sierra University, 4500 Riverwalk Parkway, Riverside, California 92515, USA. 2Department of Ecology and Evolutionary Biology and Biodiversity Institute, University of Kansas, Dyche Hall, 1345 Jayhawk Blvd,

Lawrence, Kansas 66045-7561, USA. 3Myanmar Environment Sustainable Conservation, Yangon, Myanmar.4School of Biological Sciences, Universiti Sains Malaysia, 11800 Minden, Penang, Malaysia.5Department of Biology, Villanova University, 800 Lancaster Avenue, Villanova, PA 19085, USA.6Department of Biology, California State University, Northridge, CA 91330-8303, USA.7Fauna and Flora International, No(35), 3rd Floor, Shan Gone Condo, Myay Ni Gone Market Street, Sanchaung Township, Yangon,

Myanmar.8Corresponding author. E-mail: [email protected]

Abstract

A phylogenetic taxonomic analysis indicates that a newly discovered population of Cyrtodactylus from the vicinity of

Ywangan Town in the Shan Hills, Shan State, Myanmar is a new species (C. ywanganensis sp. nov.) and the earliest di-

verging member of the linnwayensis group within the previously defined Indochinese clade. The DIVALIKE+J model of

a BioGeoBEARS biogeographic analysis indicates that the Indochinese clade evolved in the Shan Hills and Salween Basin

of eastern Myanmar and dispersed into Indochina on at least three separate occasions from 18.6–13.4 mya. Once there,

uplift of the Tenasserim Mountains and Thai Highlands created the intermedius group, the oldhami group, and C. tigroides

of western and southern Thailand which form sister lineages to the linnwayensis group, yathepyanensis group, and the

sinyineensis group, respectively, of eastern Myanmar. Diverging lineages within the Indochinese clade highlight the im-

portance of the Thai Highlands and Tenasserim Mountains in that group’s evolution and speciation. The discovery of C.

ywanganensis sp. nov. in karstic habitats in the Shan Hills continues to underscore the unrealized karst-associated herpe-

tological diversity of this vast, relatively unexplored, upland region and the need for additional field studies.

Key words: Phylogenetic taxonomy, Cyrtodactylus, biogeography, Shan Hills, Thai Highlands, Tenasserim Mountains;

Salween Basin, Myanmar

Introduction

The Shan Hills of eastern Myanmar compose a massive (~185,000 km2) circumscribed upland region that rises

steeply from the eastern edge of the Ayeyarwady Basin and extends eastward into southwestern China and western

Thailand. This topographically complex terrain ranges from 1,000–2,700 m in elevation and is an amalgam of

karstic hills and ranges, steep river valleys, and intermontane basins that are dissected by steep, north-south tending

river gorges emptying into the Chao Phraya, Ayeyarwady, Sittaung, and Salween River basins. Myanmar’s vast

size and ongoing political unrest—coupled with the difficulty of accessing its more remote river valleys and

ranges—have severely constrained recent efforts to conduct long term or repetitive field surveys throughout much

of the region. However, since October 2016, our team has been granted access (with military escort) into some of

the more remote, restricted areas in both the Shan Hills of Shan State and the more southern Salween Basin of

Kayin and Mon states in order to conduct herpetological surveys in their karstic regions. These surveys thus far

have resulted in the discovery of 18 new species of gekkonid lizards of the genera Cyrtodactylus Gray (15) and

Hemiphyllodactyus Bleeker (3) (Grismer et al. 2017a,b; Grismer et al. 2018) with two more species of

Accepted by M. Heinicke: 23 May 2018; published: 19 Jul. 2018 477

Hemiphyllodactylus and at least five new species of Hemidactylus Oken, 1817 in the process of being described

(Grismer in prep.; Murdoch in prep.).

In October 2017, we surveyed upland areas in the vicinity of Ywangan Town in the Shan Hills of Shan State

(Fig. 1) which resulted in the discovery of additional new species of geckos (Hemiphyllodactylus, Hemidactylus,

Cyrtodactylus) and a newt of the genus Tylototriton (Grismer et al. in prep.). Molecular phylogenetic and

morphological analyses of the new Cyrtodactylus distinguish it from all other Indomalayan Cyrtodactylus and

indicate it is the closest relative of the Shan Hills species C. linnwayensis Grismer, Wood, Thura, Zin, Quah,

Murdoch, Grismer, Lin, Kyaw, & Lwin, 2017 of the linnwayesis group (sec. Grismer et al. 2017a). Together, with

its sister lineage the intermedius group from Thailand, Cambodia, and Vietnam, the linnwayensis group is most

closely related to a larger lineage containing the remaining species groups of the Indochinese clade (sec. Grismer et

al. 2017a; 2018) from the Salween Basin and southern and western Thailand. This new species is described below

and a re-analysis of the biogeographic history of Cyrtodactylus from eastern Myanmar is revisited.

FIGURE 1. Distribution and relationships among species of the linnwayensis group in the Shan Hills, Shan State, Myanmar.

Branch lengths are arbitrary and do not reflect genetic distances. Starred localities represent type localities.

GRISMER ET AL.478 · Zootaxa 4446 (4) © 2018 Magnolia Press

Materials and methods

The primary aim of this study was to investigate the taxonomy, phylogenetic relationships, and biogeography of the

newly discovered species of Cyrtodactylus of the linnwayensis group from the Ywangan area of Shan State based

on 1497 bp of ND2 and its flanking tRNAs (WANCY). The data set of Grismer et al. (2018), which included

exemplars of all the major Cyrtodactylus clades in Wood et al. (2012) and Agarwal et al. (2014), was augmented

with four samples from the Ywangan area totaling 167 ingroup samples. Hemidactylus angulatus Hallowell, H.

frenatus Duméril & Bibron, H. garnotii Duméril & Bibron, H. mabouia (Moreau de Jonnès), and H. turcicus

(Linnaeus) served as outgroups following Grismer et al. (2018). All new sequences are deposited in GenBank

(Table 1).

TABLE 1. GenBank accession numbers for the newly recorded specimens used for the molecular phylogenetic analyses.

Accession numbers for outgroups are in Agarwal et al. (2014) and for the other specimens of Cyrtodactylus of the

Indochina clade see Grismer et al. (2017a).

Molecular data. Genomic DNA was isolated from liver or skeletal muscle specimens stored in 95% ethanol

using a Maxwell® RSC Tissue DNA kit on the Promega Maxwell® RSC extraction robot. ND2 was amplified

using a double-stranded Polymerase Chain Reaction (PCR) under the following conditions: 1.0 µl genomic DNA

(10–30 µg), 1.0 µl light strand primer (concentration 10 µM), 1.0 µl heavy strand primer (concentration 10 µM),

1.0 µl dinucleotide pairs (1.5 µM), 2.0 µl 5x buffer (1.5 µM), MgCl 10x buffer (1.5 µM), 0.1 µl Taq polymerase

(5u/µl), and 6.4 µl ultra-pure H2O. PCR reactions were executed on Bio-Rad gradient thermocycler under the

following conditions: initial denaturation at 95°C for 2 min, followed by a second denaturation at 95°C for 35 s,

annealing at 48°C for 35 s, followed by a cycle extension at 72°C for 35 s, for 31 cycles. All PCR products were

visualized on a 1.0 % agarose gel electrophoresis. Successful PCR products were sent to GENEWIZ® for PCR

purification, cycle sequencing, sequencing purification, and sequencing using the same primers as in the

amplification step (Table 2). Sequences were analyzed from both the 3' and the 5' ends separately to confirm

congruence between reads. Forward and reverse sequences were uploaded and edited in GeneiousTM version v6.1.8.

Following sequence editing we aligned the protein-coding region and the flanking tRNAs using the MAFTT

v7.017 (Katoh & Kuma 2002) plugin under the default settings in GeneiousTM (Kearse et al. 2012). Mesquite v3.04

(Maddison & Maddison 2015) was used to calculate the correct amino acid reading frame and to confirm the lack

of premature stop codons in the ND2 portion of the DNA fragment.

TABLE 2. Primer sequences used in this study for amplification and sequencing the ND2 gene and the flanking tRNAs.

Taxon Catalog no. Locality GenBank no.

Cyrtodactylus

ywanganensis sp. nov.

LSUHC 13711 2.7 km southwest of Ywangan, Ywangan Township, Taunggyi

District, Shan State, Myanmar (21.14643°N, 96.42178°E; 1157 m

in elevation).

MH607608

Cyrtodactylus




in elevation).

MH607610

Cyrtodactylus




in elevation).

MH607607

Cyrtodactylus




in elevation).

MH607609

Primer name Primer reference Sequence

L4437b (Macey et al. 1997) External 5’-AAGCAGTTGGGCCCATACC-3’

H5934 (Macey et al. 1997) External 5' -AGRGTGCCAATGTCTTTGTGRTT-3'

Zootaxa 4446 (4) © 2018 Magnolia Press · 479NEW CYRTODACTYLUS FROM THE SHAN HILLS

Three different phylogenetic analyses were employed. A Maximum Likelihood (ML) analysis was

implemented in IQ-TREE (Nguyen et al. 2015) using a Bayesian Information Criterion (BIC) which calculated

K3P+I+G4 to be the best-fit model of evolution for the tRNA, TVM+I+G4 for the first codon position, and

TIM3+G4 for the second and third codon positions. One-thousand bootstrap pseudoreplicates via the ultrafast

bootstrap approximation algorithm were employed and nodes having ML UFboot values (ML) of 95 and above

were considered significantly supported (Minh et al. 2013). A Bayesian inference (BI) analysis was carried out in

MrBayes 3.2.3. on XSEDE (Ronquist et al. 2012) using CIPRES (Cyberinfrastructure for Phylogenetic Research;

Miller et al. 2010) employing default priors and HKY + Gamma model of evolution for the tRNA and the second

codon position and GTR + G to the first and third codon positions. Two simultaneous runs were performed with

four chains, three hot and one cold. The analysis ran for 100 million generations, was sampled every 10 thousand

generations using the Markov Chain Monte Carlo (MCMC), and the first 25% of each run was discarded as burn-

in. Stationarity and .p files from each run were checked in Tracer v1.6 (Rambaut et al. 2014) to ensure effective

sample sizes (ESS) were above 200 for all parameters. Nodes with Bayesian posterior probabilities (BI) of 0.95 and

above were considered well-supported (Hulsenbeck et al. 2001; Wilcox et al. 2002). After removing outgroup taxa,

MEGA7 (Kumar et al. 2016) was used to calculate uncorrected pairwise sequence divergence among the ingroup

species.

A time-calibrated Bayesian Inference analysis was implemented in BEAUti version 2.4.7 (Bayesian

Evolutionary Analysis Utility) and run with BEAST version 2.4.6 (Bayesian Evolutionary Analysis Sampling

Trees; Drummond et al. 2012) in CIPRES employing a lognormal relaxed clock with unlinked substitution and

clock models and an HKY substitution model selected for each codon position. MCMC chains were run using a

coalescent exponential population prior for 100 million generations and logged every 10 thousand generations. It

has been demonstrated that the third codon position is susceptible to substitution saturation (Zamudio et al. 1997;

Carranza et al. 2000; Brandley et al. 2011; Grismer et al. 2015) and could contribute to overestimating node ages.

However, Grismer et al. (2015) noted that although third codon position saturation was evident in their study of the

gekkonid Hemiphyllodactylus, it was not a significant factor in estimating node ages across various codon and gene

partition schemes and was consistent with similar node age estimates using nuclear genes on the same taxa as

Heinicke et al. (2011). Fossil calibrations from Agarwal et al. (2014) on essentially the same data set used herein,

placed a mean date of 31.4 million years (95% HPD 36.2–27.2 Myr; Agarwal in litt. 2016) on the node between the

Cyrtodactylus pulchellus complex and taxa of the Indochinese clade taxa (C. intermedius and C. hontreensis—the

intermedius group). This node age was used herein as an internal constraint prior to date the same node with a

standard deviation of 2.3%. The BEAST log file was visualized and checked in Tracer 1.6.0 (Rambaut et al. 2014)

to ensure ESS values were above 200 for all parameters and a maximum clade credibility tree using mean heights

at the nodes was generated using TreeAnnotator v.1.8.0 (Rambaut & Drummond 2013) with a burnin of 1000 trees.

The time-calibrated BEAST tree was pruned to include one individual per ingroup species and one individual of

each major outgroup clade using custom R script. The tree was converted to newick format and used to estimate the

ancestral range at the nodes using the R package BioGeoBEARS (Matzke 2013). This program allows for both

probabilistic inference of ancestral geographic ranges and statistical comparisons of range expansion using

different models in a likelihood framework so that standard statistical model selection procedures using the Akaike

information criterion (AIC) can be applied to allow the data to choose the best fit model. Available models in

BioGeoBEARS include a likelihood version of the parsimony-based Dispersal Vicariance Analysis DIVA

(“DIVALIKE”) model (Ronquist 1997), the likelihood-based Dispersal-Extinction Cladogenesis (DEC) model of

the LAGRANGE program (Ree & Smith 2008), and the Bayesian-based BayArea (“BAYAREALIKE”) model

(Landis et al. 2013). Additionally, each model also incorporates founder-effect speciation +J (jump-dispersal)

which was purported to be particularly important when reconstructing biogeographic scenarios of insular lineages

(Matzke 2014). However, the DEC and the DEC +J models have come under scrutiny in that they are not time-

dependent, thus precluding anagenesis in the speciation process and deferring all speciation to nearly instantaneous

cladogenic events at the nodes (Ree & Sanmartín 2018). A presence or absence species geography file was

constructed utilizing four geographically discrete areas across which we determined a posteriori there is turnover

among Cyrtodactylus lineages. (1) Indo-Burma—including India, Sri Lanka, and the Himalayan Mountains north

of the Indo-Gangetic flood plain and extending to the eastern edge of the Ayeyerwady Basin to the border of the

West Burma and Sibumasa Blocks along the Saigang Fault that align themselves with the western edge of the Shan

Hills (Hutchison 2007; Ridd 2009). We note that the Indian-Himalayan portion of this region is very complex and


could be subdivided further based on geography and clade turnover among a number of undescribed endemic

species (Agarwal et al. 2014) but doing so would be premature at this point and has no bearing on the

biogeographic history of the Indo-Chinese clade that is the focus of this study. (2) Eastern Myanmar—extending

from Saigang Fault (i.e. the western edge of the Shan Hills and eastern edge of the Ayeyerwady Basin) and the Gulf

of Martaban in the west to the Inthanon Zone of the Indochina Block in the east along the Mae Yuan, Mae Ping, and

Three Pagoda Faults (Barr & MacDonald 1991; Sone & Metcalfe 2008; i.e. the Tenasserim Mountains and Thai

Highlands), thus delimiting the Shan Hills and the Salween Basin. (3) Indochina—extending from the Mae Yuan,

Mae Ping, and Three Pagoda Faults on the western edge of the Tenasserim Mountains and Thai Highlands along

the Salween River to the east coast of Vietnam and south to the Isthmus of Kra. And (4) a portion of Southeast Asia

extending from the Isthmus of Kra south through the Thai-Malay Peninsula and Indo-Australian Archipelago to

New Guinea (Fig 2). Each species occupied only a single area.

FIGURE 2. Discrete geographic regions used in the BioGeoBEARS analysis (map) and the ancestral ranges recovered by the

DIVALIKE+J model (tree) of that analysis for selected outgroups and all species belonging to the four species groups of the

Indochinese clade (sec. Grismer et al. 2017a) of Cyrtodactylus. Numbers at the nodes represent time of divergence and

correspond to the arrows on the map which indicate the direction of dispersal.


Morphological analysis. Color notes were taken from living specimens and digital images of living specimens

of all possible age classes prior to preservation. Measurements were taken on the left side of the body when

possible to the nearest 0.1 mm by MSG using Mitutoyo dial calipers under a Nikon SMZ 1500 dissecting

microscope. Measurements taken were: snout-vent length (SVL), taken from the tip of snout to the vent; tail length

(TL), taken from the vent to the tip of the tail, original or regenerated; tail width (TW), taken at the base of the tail

immediately posterior to the postcloacal swelling; forearm length (FL), taken on the dorsal surface from the

posterior margin of the elbow while flexed 90° to the inflection of the flexed wrist; tibia length (TBL), taken on the

ventral surface from the posterior surface of the knee while flexed 90° to the base of the heel; axilla to groin length

(AGL), taken from the posterior margin of the forelimb at its insertion point on the body to the anterior margin of

the hind limb at its insertion point on the body; head length (HL), the distance from the posterior margin of the

retroarticular process of the lower jaw to the tip of the snout; head width (HW), measured at the angle of the jaws;

head depth (HD), the maximum height of head measured from the occiput to the throat; eye diameter (ED), the

greatest horizontal diameter of the eye-ball; eye to ear distance (EE), measured from the anterior edge of the ear

opening to the posterior edge of the eye-ball; eye to snout distance (ES), measured from anterior-most margin of

the eye-ball to the tip of snout; eye to nostril distance (EN), measured from the anterior margin of the eye ball to the

posterior margin of the external nares; inter orbital distance (IO), measured between the anterior edges of the orbit;

ear diameter (EL), the greatest vertical distance of the ear opening; and internarial distance (IN), measured between

the nares across the rostrum.

Meristic characters taken by MSG were the numbers of supralabial (SL) and infralabial (IL) scales counted

from the largest scale immediately below the middle of the eyeball to the rostral and mental scales, respectively;

the number of paravertebral tubercles (PVT) between limb insertions counted in a straight line immediately left of

the vertebral column; the number of longitudinal rows of body tubercles (LRT) counted transversely across the

center of the dorsum from one ventrolateral fold to the other; the number of longitudinal rows of ventral scales

(VS) counted transversely across the center of the abdomen from one ventrolateral fold to the other; the number of

expanded subdigital lamellae proximal to the digital inflection on the fourth toe (TLE) counted from the base of the

first phalanx where it contacts the body of the foot to the largest scale on the digital inflection (see Grismer et al.

2017a: Fig. 3), large continuous scales on the palmar and plantar surfaces were not counted; the number of small,

unmodified subdigital lamellae distal to the digital inflection on the fourth toe (TLN) counted from the digital

inflection to the claw (see Grismer et al. 2017a: Fig. 3); and the total number of subdigital lamellae (TL) beneath

the fourth toe (i.e. TLE + TLN = TL). The total number of enlarged femoral scales (FS) from each thigh were

combined as a single metric. In some species, only the distalmost scales are greatly enlarged and the proximal

scales are smaller whereas in others the greatly enlarged scales are continuous with the enlarged precloacal scales

and the separation of the two scales rows was determined to be at a point even with the lateral body margin (see

Grismer et al. 2017a: Fig. 4). The total number of femoral pores (FP) in males (i.e., the sum of the number of

enlarged pore-bearing femoral scales from each leg combined as a single metric—not all enlarged femoral scales

have pores). The number of enlarged precloacal scales (PS); the number of precloacal pores in (PP) in males; the

number of continuous femoroprecloacal pores (FPP) in males (see Grismer et al. 2017a: Fig. 4); and the number of

rows of post-precloacal scales (PPS) on the midline between the enlarged precloacal scales and the vent (see

Grismer et al. 2017a:Fig. 4); number of body bands (BB) between the nuchal loop (dark band running from eye to

eye) and the hind limb insertions not including the sacral or postsacral bands, although the dark band on the nape—

which is variably present—is considered as part of this count (see Grismer et al. 2017a: Fig. 5); the number of light

caudal bands (LCB) on an original tail; and the number of dark caudal bands (DCB) on an original tail.

Non-meristic morphological characters evaluated were the degree of body tuberculation—weak tuberculation

referring to dorsal body tubercles that are relatively low, small, less densely packed, and weakly keeled whereas

prominent tuberculation refers to tubercles that are larger, higher (raised), and prominently keeled (see Grismer et

al. 2017a: Fig. 6); body tubercles extending past the base of the tail or not (see Grismer et al. 2017a: Fig. 7);

enlarged femoral scales and precloacal scales contiguous or separated by a diastema at the base of the femora (see

Grismer et al. 2017a: Fig. 4); and the relative length to width ratio of the transversely expanded, median subcaudal

scales and whether or not they extend onto the lateral surface of the tail (see Grismer et al. 2017a: Fig. 8).

Color pattern characters (see Grismer et al. 2017a: Fig. 5) evaluated were the nuchal loop being continuous

from eye to eye, separated medially into paravertebral halves, bearing an anterior azygous notch or not, and the

posterior border being straight (smooth), sinuous, or jagged; dorsal body bands bearing paired, paravertebral


elements or not; dark dorsal body bands wider than light interspaces, with or without lightened centers, edged with

light tubercles or not, jagged shaped or more regular (straight or even); dark markings present or absent in the

dorsal interspaces; light tubercles dispersed throughout the dorsal interspaces or not; ventrolateral body fold white,

appearing as a wide or narrow stripe; top of head bearing combinations of dark diffuse mottling or dark, distinct

blotches overlain with a light-colored reticulating network or not; anterodorsal margin of thighs and brachia

whitish due to a lack of dark pigment; light caudal bands bearing dark markings or immaculate; light caudal bands

encircle tail or not; dark caudal bands wider than light caudal bands; and regenerated tail bearing a pattern of

distinct, dark spots or not.

All statistical analyses were performed using the platform R v 3.2.1 (R Core Team 2015). Prior to an analysis

of variance (ANOVA), a Levene’s test was conducted to test for homogeneity of variances among the meristic

characters (p > 0.05) with the variables centered about the mean. An ANOVA was conducted on characters with

similar variances (i.e. high p values in the Levene’s test) to test for the presence of statistically significant mean

differences (p ≤ 0.05) in the data set. Characters containing statistical differences were subjected to a TukeyHSD

test to ascertain which population pairs differed significantly from each other for those characters. Boxplots were

generated using custom R script in order to visualize the range and degree of differences of characters bearing

statistical differences between the new species and/or C. linnwayensis and C. shwetaungorum. Principal

Component Analysis (PCA) and Discriminant Analysis of Principal Components (DAPC) using the ADEGENET

package in R (Jombart et al. 2010) were used to determine if species of the linnwayensis group occupied unique

positions in morphospace and the degree to which their variation in morphospace coincided with species

boundaries delimited by the molecular phylogenetic and univariate analyses. PCA, implemented by the prcomp

command in R, searches for the best overall low-dimensional representation of significant morphological variation

in the data. Femoral and precloacal pore counts were excluded from the PCA due to their presence in only males.

All PCA data were log-transformed prior to analysis and scaled to their standard deviation in order to normalize

their distribution so as to ensure characters with very large and very low values did not over-leverage the results

owing to intervariable nonlinearity. A biplot analysis implemented by the ggbiplot command in R was overlain on

the PCA plot in order to visualize the degree to which certain characters co-varied and contributed to the overall

variation in the data set. To characterize clustering and separation in morphospace, a DAPC was performed to

search for linear combinations of morphological variables having the greatest between-group variance and the

smallest within-group variance (Jombart et al. 2010). DAPC relies on log transformed and standardized data from

the PCA as a prior step to ensure that variables analyzed are not correlated and number fewer than the sample size.

DAPC principal components with eigenvalues accounting for 80–90% of the variation in the data set were retained

for the DAPC analysis according to the criterion of Jombart et al. (2010). Museum abbreviations follow Sabaj

(2016) except for LSUHC referring to the La Sierra University Herpetological Collection, La Sierra University,

Riverside, California, 92505, USA; MS referring to Montri Sumontha, Ranong Marine Fisheries Station, Ranong

85000, Thailand; and PLWJ referring to Perry L. Wood, Jr. field series, Department of Ecology and Evolutionary

Biology and Biodiversity Institute, University of Kansas, Lawrence, Kansas 66045-7561, USA.

Results

The BI and ML analyses recovered strongly supported trees with identical topologies indicating that the newly

discovered population of Cyrtodactylus from the Ywangan area is the sister species of C. linnwayensis (Fig. 3),

differing from it by an uncorrected pairwise sequence divergence of 6.5% and from the other member of the

linnwayensis species group, C. shwetaungorum, by a sequence divergence of 13.4%. The morphological analyses

corroborate the molecular analyses in that each species in the linnwayensis group can be discretely and

significantly differentiated from each other (Tables 3, 4) and occupies unique, non-overlapping positions in

morphospace (Fig. 4). The PCA biplot indicates a relatively sharp dichotomy in character variation with the

number of body bands (BB), paravertebral tubercles (PVT), longitudinal rows of tubercles (LRT), and femoral

scales (FS) co-varying by having relatively high values among the new species and C. shwetaungorum while labial

scales (SL, IL), toe lamellae (TLE, TLN, TL), ventral scales (VS), and post-precloacal scales (PPS) co-vary in the

opposite direction in C. linnwayensis. Statistically different variation between combinations of the new species

and/or C. linnwayensis and C. shwetaungorum can be visualized in Figure 5. As a result of these data, the new

species is described below.


FIGURE 3. Bayesian inference topology of the species belonging to the four species groups of the Indochinese clade (sec.

Grismer et al. 2017a) and the phylogenetic relationships of Cyrtodactylus ywanganensis sp. nov. of the linnewayensis group.

Black circles denote nodes with BI and ML values of 0.95 and 95 or above, respectively. Grey circles denote nodes supported

only by ML values of 95 or higher.


FIGURE 4. Principle component analysis (PCA) with biplot overlay, discriminant analysis of principles components (DAPC),

and three-dimensional representation of the first three principle components (PC1, PC2, PC3) illustrating the morphospatial

separation of the three species of the linnwayensis group.


FIGURE 5. Box plots of the characters having statistically significant mean differences between Cyrtodactylus ywanganensis

sp. nov. and/or C. linnwayesnsis and C. shwetaungorum illustrating the lack of overall intermediacy of C. ywanganensis sp.

nov. between C. linnwayensis and C. shwetaungorum. Colored boxes represent the interquartile range, the black horizontal bar

is the median, and the light-blue spot is the mean.


TABLE 3. Summary statistics and diagnostic characters of the species from the linnwayensis species groups. SD =

standard deviation, and N = sample size.

Summary Statistics shwetaungorum linnwayensis ywanganensis sp. nov.

supralabial scales (SL)

Mean 8.3 8 7

SD ±0.71 ±0.89 0

Range 7–9 7–9 7

N 8 11 4

infralabial scales (IL)

Mean 6.9 7 6

SD ±0.64 ±0.63 0

Range 6–8 6–8 6

N 8 11 4

paravertebral tubercles (PVT)

Mean 33.1 29.4 33.3

SD ±1.36 ±1.86 1.26

Range 31–35 25–33 32–35

N 8 11 4

longitudunal rows of body tubercles (LRT)

Mean 18.9 16.4 22.6

SD ±0.10 ±1.36 ±1.7

Range 18–21 13–18 21–25

N 8 11 4

ventral scales (VS)

Mean 37.3 39.7 35.0

SD ±2.71 ±2.37 ±0.82

Range 33–40 34–42 34–36

N 8 11 4

expanded 4th toe lamellae (TLE)

Mean 8.3 8.8 7.5

SD ±0.46 ±0.6 ±0.58

Range 8 or 9 8–10 7 or 8

N 8 11 4

unmodified 4th toe lamellae (TLN)

Mean 12.4 13.1 13

SD ±0.92 ±0.54 ±0.82

Range 11–14 12–14 12–14

N 8 11 4

total 4th toe lamellae (TL)

Mean 20.6 21.9 20.5

SD ±0.92 ±0.70 ±1.00

Range 20–22 21–23 19–21

N 8 11 4

......continued on the next page


TABLE 3. (Continued)


enlarged femoral scales (FS)

Mean 28.1 27.9 29

SD ±2.59 ±2.26 ±1.15

Range 24–32 24–32 28–30

N 8 11 4

femoral pores (FP)

Mean 15.7 17.6 20

SD ±1.15 ±4.28 0

Range 15–17 10–22 20

N 3 7 1

enlarged precloacal scales (PS)

Mean 9.8 10.5 8.8

SD ±0.89 ±1.13 ±0.50

Range 8–11 9–12 8–9

N 8 11 4

precloacal pores (PP)

Mean 10 8 8.5

SD ±1.73 ±2.12 0.71

Range 8–10 6–10 8 or 9

N 3 9 2

post-precloacal scale rows (PPS)

Mean 3 4 3

SD 0 0 0.00

Range 3 4 3

N 3 11 4

body bands (BB)

Mean 4 3.5 4

SD 0 ±0.52 0.00

Range 4 3 or 4 4

N 8 11 4

light caudal bands (LCB)

Mean 8.3 8.2 7

SD ±0.58 ±0.45 0

Range 8 or 9 8 or 9 7

N 3 9 1

dark caudal bands (DCB)

Mean 8.3 8.4 7

SD ±1.15 ±0.55 0

Range 7–9 8 or 9 7

N 3 9 1

Morphology

body tubercles low, weakly keeled yes yes no



Taxonomy

Cyrtodactylus ywanganensis sp. nov.

Suggested common name: Ywangan Bent-toed Gecko

Fig. 6

Holotype. Adult female LSUHC 13758 collected on 24 October 2017 at 2300 hrs by L. Lee Grismer, Robert E.

Espinoza, Matthew L. Murdoch, Myint Kyaw Thura, Evan S. H. Quah, and Tun Oo from 2.7 km southwest of

Ywangan, Ywangan Township, Taunggyi District, Shan State, Myanmar (21.14643°N, 96.42178°E; 1157 m in

elevation).

Paratypes. Adult female paratype LSUHC 13757 bears the same collection data as the holotype. Adult male

paratype LSUHC 13712 and subadult male paratype LSUHC 13711 bear the same collection data as the holotype

except they were collected on 23 October 2017 at 2200 hrs.

Diagnosis. Cyrtodactylus ywanganensis sp. nov. differs from all congeners by having the unique combination

of a maximum SVL of 96.1 mm; seven supralabials; six infralabials; 32–35 paravertebral tubercles; 21–25



body tubercles raised, moderately to strongly keeled no no yes

tubercles extend beyond base of tail no no no

enlarged femoral and precloacal scales continuous yes yes yes

pore-bearing femoral and precloacal scales continuous no no no

enlarged proximal femoral scales ~1/2 size of distal femorals or

nearly equal in size

equal equal equal

medial subcaudal scales 2 or 3 times wider than long 2 2 2

medial subcaudal extend onto lateral surface of tail no no no

Color Pattern

nuchal loop divided medially no 1(yes),10(no) no

nuchal loop with anterior azygous notch no yes yes

posterior border of nuchal loop straight usually straight straight

band on nape no no no

dorsal banding with paravertebrtal elements no no no

dorsal bands wider than interspaces yes variable yes

dorsal bands bearing lightened centers yes yes no

dorsal bands edged with light-colored tubercles yes yes yes

shape of dorsal bands regular variable jagged

dark markings in dorsal interspaces yes yes yes

ventrolateral body fold whitish no no no

top of head diffusely mottled, blotched, or patternless blotched blotched blotched

light reticulum on top of head yes yes no

anterodorsal margin of thighs darkly pigmented yes yes yes

anterodorsal margin of brachia darkly pigmented yes yes yes

light caudal bands bearing dark markings no 5(yes)1(no) yes

light caudal bands encircle tail yes variable yes

dark caudal bands wider than light caudal bands yes yes yes

mature regenerated tail spotted no yes yes

maximum SVL (mm) 102.2 101.7 96.1


longitudinal rows of body tubercles; 34–36 ventral scales; relatively long digits with seven or eight expanded

subdigital lamellae proximal to the digital inflection on the fourth toe, 12–14 unmodified distal subdigital lamellae,

19–21 total subdigital lamellae; raised, strongly keeled, dorsal body tubercles; tubercles not extending beyond base

of tail; enlarged femoral and precloacal scales continuous; enlarged proximal femoral scales equal in size to the

enlarged distal femoral scales; 28–30 enlarged femoral scales; 20 femoral pores in male; eight or nine precloacal

pores in male; femoral and precloacal pore-bearing scales not continuous; eight or nine enlarged precloacal scales;

three rows of enlarged post-precloacal scales; medial subcaudal scales twice as wide as long, not extending onto

lateral surface of tail; top of head darkly blotched in adults, no yellow reticulum; nuchal loop not divided medially,

pronounced anterior azygous notch, posterior border straight; four jagged, dark, dorsal bands lacking paravertebral

elements, wider than interspaces with no lightened centers, edged with yellow tubercles; no band on nape; dark

markings in dorsal interspaces; ventrolateral folds not whitish; anterodrosal margins of thighs and brachia darkly

pigmented; seven light caudal bands bearing dark markings, encircling tail; seven dark caudal bands wider than

light caudal bands; and mature regenerated tail spotted. These characters are scored against all other species in the

linnwayensis group (Table 3) and all other Burmese species of the Indochinese clade in Grismer et al. (2017a:Table

8).

Description of holotype. Adult female SVL 90.5 mm; head moderate in length (HL/SVL 0.29), wide (HW/HL

0.69), flat (HD/HL 0.38), distinct from neck, triangular in dorsal profile; lores inflated, prefrontal region concave,

canthus rostralis rounded; snout elongate (ES/HL 0.43), rounded in dorsal profile, broad in lateral profile; eye large

(ED/HL 0.25); ear opening oval, small (EL/HL 0.09); eye to ear distance greater than diameter of eye; rostral

rectangular, partially divided dorsally, bordered posteriorly by supranasals and azygous postnasal, laterally by first

supralabials; external nares bordered anteriorly by rostral, dorsally by supranasals, posteriorly by two postnasals,

and ventrally by first supralabials; 7(R,L) rectangular supralabials extending to below midpoint of eye; 6(R,L)

infralabials tapering posteriorly to below orbit; scales of rostrum and lores slightly raised, larger than granular

scales on top of head and occiput; scales on top of head and occiput intermixed with small tubercles; dorsal

superciliaries weakly pointed and directed posteriorly; mental triangular, bordered laterally by first infralabials and

posteriorly by large left and right trapezoidal postmentals which contact medially for 50% of their length posterior

to mental; two rows of variably enlarged chinshields bordering all infralabials; gular and throat scales granular,

grading posteriorly into larger, subimbricate pectoral and ventral scales.

Body relatively short (AGL/SVL 0.43) with well-defined ventrolateral folds; dorsal scales small, raised and

interspersed with large, raised, semi-regularly arranged, keeled tubercles; tubercles extend from nape onto base of

tail but no farther; tubercles on nape smaller than those on posterior portion of body; 32 paravertebral tubercles;

approximately 25 longitudinal rows of dorsal tubercles; 35 flat, subimbricate, ventral scales larger than dorsal

scales; nine enlarged precloacal scales; three rows of large, post-precloacal scales; and no deep precloacal groove

or depression.

Forelimbs moderate in stature, relatively short (FL/SVL 0.17); slightly raised, juxtaposed scales of forearm

larger than those on body, intermixed with tubercles; palmar scales slightly raised; digits well-developed, relatively

long, inflected at basal, interphalangeal joints; digits much more narrow distal to inflections; widened proximal

subdigital lamellae do not extend onto palm; webbing at base of digit; claws well-developed, sheathed by a dorsal

and ventral scale at base; hind limbs more robust than forelimbs, moderate in length (TBL/SVL 0.20), covered

dorsally by small, raised, juxtaposed scales intermixed with large tubercles and bearing flat, slightly larger scales

anteriorly; ventral femoral scales imbricate, larger than dorsals; one row of 15(R,L) enlarged femoral scales in

contact with enlarged precloacal scales; enlarged femoral scales nearly equal in size; small, postfemoral scales

form an abrupt union with larger, flat ventral scales of posteroventral margin of thigh; subtibial scales flat,

imbricate; plantar scales raised; digits relatively long, well-developed, inflected at basal, interphalangeal joints;

eight (R,L) transversely expanded subdigital lamellae on fourth toe proximal to joint inflection that do not extend

onto sole, 13 (R,L) unmodified subdigital lamellae distal to inflection; and claws well-developed, base of claw

sheathed by a dorsal and ventral scale.

Tail complete, original, moderate in proportions, 110.0 mm in length, 8.0 mm in width at base, tapering to a

point, TL/SVL (1.22); dorsal scales of tail flat; median row of transversely expanded subcaudal scales twice as

wide as long, not extending onto lateral subcaudal region; three enlarged postcloacal tubercles at base of tail on

hemipenal swellings; and postcloacal scales flat.

Coloration in life (Fig. 6). Dorsal ground color of head, body, and limbs brown; top of head and rostrum


bearing large dark-brown blotches edged in yellow; superciliary scales yellow; wide, dark-brown, nuchal band

edged in yellow, bearing a deep azygous anterior notch and a straight posterior margin; no band on nape; four

jagged body bands bearing dark centers, wider than interspaces, not bearing paravertebral elements and edged with

yellow tubercles; one dark postsacral band edged with yellow tubercles; interspaces bearing faint, dark markings;

thin, yellow, irregularly shaped bands and small spots on limbs; dorsal margins of brachia and thighs darkly

pigmented; ventrolateral body folds darkly pigmented; ventral surfaces beige, dusky in appearance; and tail

original, bearing seven dark wide caudal bands and seven much narrower white caudal bands with dark markings

that encircle tail.

Variation (Fig. 6). The paratypes closely resemble the holotype in all aspects of coloration and pattern.

Paratypes LSUHC13712 and 13757 have fully regenerated spotted tails. Subadult male LSUHC 13711 has a

broken tail with two very small regenerating sections, is lacking femoral pores, and has less yellow in its color

pattern. Additional variation in meristic and mensural characters are presented in Table 4.

TABLE 4. Meristic, mensural, and color pattern data from the type series of Cyrtodactylus ywanganensis sp. nov.

Abbreviations are listed in the Materials and methods. R = right, L = left, / = data unobtainable or not applicable.

LSUHC LSUHC LSUHC LSUHC

13758 13757 13712 13711

holotype paratype paratype paratype

Ywangan Ywangan Ywangan Ywangan

sex f f m m

supralabials 7 7 7 7

infralabials 6 6 6 6

body tubercles low, weakly keeled no no no no

body tubercles raised, moderately to strongly keeled yes yes yes yes

paravertebral tubercles 32 35 33 35

longitudinal rows of body tubercles 25 23 21 22

tubercles extend beyond base of tail no / / /

ventral scales 35 34 35 36

expanded subdigital lamellae on 4th toe 8 8 7 7

unmodified subdigital lamellae on 4th toe 13 13 14 12

total subdigital lamellae on 4th toe 21 21 21 19

enlarged femoral scales (R/L) 15R15L 15R15L 14R14L 14R14L

femoral pores (R/L) / / 10R10L /

enlarged precolacal scales 9 9 8 9

precloacal pores / / 8 9

post-precloacal scales rows 3 3 3 3

enlarged femoral and precloacal scales continuous yes yes yes yes

pore-bearing femoral and precloacal scales continuous / / no /

enlarged proximal femoral scales ~1/2 size of distal femorals or

nearly equal in size

equal equal equal equal

medial subcaudals 2 or 3 times wider than long 2 / / /

medial subcaudals extend onto lateral surface of tail no / / /

nuchal loop divided medially no no no no

nuchal loop with anterior azygous notch yes yes yes yes

posterior border of nuchal loop straight straight straight straight

band on nape no no no no

dorsal banding with paravertebral elements no no no no



Distribution. Cyrtodactylus ywaganensis sp. nov. is known only from the type locality 2.7 km southwest of

Ywangan, Ywangan Township, Taunggyi District, Shan State, Myanmar but is likely to occur in other nearby

limestone outcroppings to the west up to the crest of the Shan Plateau before it descends steeply down to the

eastern edge of the Ayeyerwady Basin (Fig. 1).

Etymology. The specific epithet, ywanganensis, is a noun in apposition in reference to the type locality being

near the town of Ywangan, Shan State.


LSUHC LSUHC LSUHC LSUHC

13758 13757 13712 13711

holotype paratype paratype paratype

Ywangan Ywangan Ywangan Ywangan

number of body bands 4 4 4 4

dorsal bands wider than interspaces yes yes yes yes

dorsal bands bearing lightened centers no no no no

dorsal bands edged with light-colored tubercles yes yes yes yes

shape of dorsal bands jagged jagged jagged jagged

dark markings in dorsal interspaces yes yes yes yes

ventrolateral body fold whitish no no no no

top of head diffusely mottled, blotched, or patternless blotched blotched blotched blotched

light reticulum on top of head no no no no

anterodorsal margin of thighs darkly pigmented yes yes yes yes

anterodorsal margin of brachia darkly pigmented yes yes yes yes

light caudal bands bearing dark markings yes / / /

light caudal bands encircle tail yes / / /

number of light caudal bands 7 / / /

number of dark caudal bands 7 / / /

dark caudal bands wider than light caudal bands yes / / /

mature regenerated tail spotted / yes yes /

SVL 90.5 94.6 96.1 80.1

TL 110 83 77 25

TW 8 9.1 9.5 7.1

FL 15.8 16 16.1 14.4

TBL 18.4 18.3 18.6 14.7

AGL 38.6 43.4 40.5 37.8

HL 26.1 26.5 27.7 22.8

HW 17.9 19.5 20 16.3

HD 10 11.7 13.1 9.3

ED 6.4 6.9 6.6 5.5

EE 7.3 7.9 8.5 6.4

ES 11.3 11.3 12.2 9.4

EN 8.5 8.4 9.3 7.3

IO 7.7 7.7 7.7 5.8

EL 2.4 2.3 2.5 2.2

IN 3.1 2.8 3.1 2.6


FIGURE 6. Cyrtodactylus ywanganensis sp. nov. from the type locality 2.7 km southwest of Ywangan, Ywangan Township,

Taunggyi District, Shan State, Myanmar: (A) juvenile male paratype LSUHC 13711, (B) adult male paratype LSUHC 13712,

and (C) adult female holotype LSUHC 13758. (D) Cyrtodactylus cf. linnwayensis. LSUDPC 10754 from Kyauk Hnat Cave in

the Shan Hills, Shan State, approximately 4.75 km soutwest of the Linn Way depression (photograph by Alice Hughs).

Natural history. The region surrounding the type locality of Cyrtodactylus ywanganensis sp. nov. is a

generally open, flat, and disturbed landscape with low, short, vegetated karstic ridges and small, isolated, karst

outcrops scattered throughout the area (Fig. 7). At the base of some of the karstic ridges are narrow (0.05–1 m

across), vertical shafts that extend underground for unknown distances. All specimens collected and observed were

near the openings of these shafts. The subadult male LSUHC 13711, was found at night on the face of a karst wall

beneath a large overhang immediately above a shaft opening. It retreated into a crack but was coaxed out into the

open and captured. Approximately 50 m away, adult male LSUHC 13712 was found the same night at the entrance

to another shaft and was captured 2 m below the opening while attempting to escape. Adult female LSUHC 13757

was found in the same place as LSUHC 13711 the next day but lower on the wall below the shaft opening where it

was relatively dark. It too was captured as it descended to approximately 3 m down the shaft in an attempt to

escape. Another specimen was seen later that day at another shaft opening but was able to escape. The adult female

LSUHC 13758 was found at night 2 m above ground level on the narrow branch of a tree growing out of a shaft. It

is clear C. ywanganensis sp. nov. is closely associated with these small, karstic, cave-like microhabitats. The

depths to which these shafts extend and the extent (if any) of their subterranean connectivity is unknown but clearly

these geckos have to be interacting with each other and it is unlikely they are doing so above ground off the karst.

From 1800–2400 hrs on two consecutive nights, seven people surveyed several isolated karst outcrops in the open

areas that contained cracks and holes suitable for Cyrtodactylus but only Hemidactylus and Hemiphyllodacylus

were found.

Comparisons. The phylogenetic relationships indicate that Cyrtodactylus ywanganensis sp. nov. is nested

within the linnwayensis group. The PCA and DAPC analyses indicate that C. ywanganensis sp. nov. is

morphospatially separate from other species of the linnwayensis group along the first three principal components

(PC) with PC1 accounting for 41% of the total variation in the data set (Fig. 4; Table 5) and loading most heavily

for numbers of paravertebral tubercles (PVT), expanded and total numbers of fourth toe lamellae (TL and TEL),

longitudinal rows of tubercles (LRT), body bands (BB), and post-precloacal scale rows (PPS). PC2, which

accounts for 18% of the total variation loads most heavily for infralabials (IL) and supralabials (SL). PC 3 accounts


for an additional 11% of the total variation and loads most heavily for the number of unmodified toe lamellae

(TLN). The ANOVA and Tukey HSD analyses indicate C. ywanganensis sp. nov. has several meristic characters

whose mean values differ significantly (p ≤ 0.05) and discreetly (i.e. no overlap in the range of variation) from the

other members of the linnwayensis group (Tables 3, 6).

FIGURE 7. A and B orientation of the vertical shafts (white arrows) at the base of karst boulders at the type locality of

Cyrtodactylus ywanganensis sp. nov. 2.7 km southwest of Ywangan, Ywangan Township, Taunggyi District, Shan State,

Myanmar. C. General habitat structure of the type locality.

Remarks. Grismer et al. (2017a) left open the possibility that Cyrtodactylus shwetaungorum and C.

linnwayensis were conspecific and that their morphological dissimilarities and uncorrected pairwise genetic

distance of 10.2% was a result of sampling error. They noted that additional populations from the intervening area

would bear significantly on this question. Cyrtodactylus ywanganensis sp. nov. from the intervening area (Fig. 1) is

not morphologically intermediate between C. shwetaungorum and C. linnwayensis in any of the significantly

different characters except for the number of post-precloacal scale rows (Fig. 5). Additionally, C. ywanganensis sp.

nov. is morphospacially unique and does not cluster between the other two species (Fig. 4). Furthermore, C.

ywanganensis sp. nov. is more strongly tuberculated, has dorsal bands lacking lightened centers, lacks a light-

colored reticulum on the top of the head, and has seven versus eight or nine light-colored caudal bands. These,

along with its 6.5% sequence divergence from its sister species C. linnwayensis, are taken as evidence that these

three populations constitute independent, non-reticulating lineages on independent evolutionary trajectories (i.e.

they are different species). We believe that based on the color pattern of a fourth population of Cyrtodactylus

discovered by Alice Hughes from Kyauk Hnat Cave in the Shan Hills approximately 4.75 km southwest of the

Linn Way depression (Fig. 1), that it too is a member of the linnwayensis group and most likely C. linnwayensis.

Like all other members of this group, it has the unique combination of an anterior azygous notch in the nuchal loop,

no band on the nape, four jagged body bands wider than the interspaces edged with light-colored tubercles, and the

body tubercles do not extend past the base of the tail (Fig. 6).


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TABLE 6. Pairwise matrix of statistically significant mean differences of characters among species of the linnwayensis

species group. Abbreviations are in the Materials and methods.

Biogeography

BioGeoBEARS model comparisons favored the DIVALIKE + J model as recovering the most likely ancestral areas

of the nodes within the pruned phylogeny (Fig. 2) in that it had the lowest AIC value among the models (Table 7).

It is noteworthy, however, that every model generated in BioGeoBEARS recovered the same ancestral range for

each respective node on the tree, thus converging on the same biogeographical scenario. Although the focus of this

analysis is on the biogeography of the Indochinese clade, the DIVALIKE +J model corroborates the biogeographic

scenarios of Wood et al. (2012) and Agarwal et al. (2014) by supporting their hypothesis that Cyrtodactylus

originated in Indo-Burma and gave rise to a monophyletic group that eventually dispersed eastward to occupy

Indochina and Southeast Asia. The scenario proposed herein departs from that of Wood et al. (2012) and Agarwal

et al. (2014) in that our analysis suggests there was no over-water back-dispersal into Indo-Burma by a clade

composed of C. triedrus (Günther), C. deccanensis (Günther), and C. collegalensis (Beddome). Instead, we posit

two independent, nearly simultaneous colonizations of Sundaland by Indo-Burmese lineages at approximately 31.5

and 30.9 mya (hpd 38.5–26.9 mya and 35.4–26.4 mya, respectively; Fig. 2). The same biogeographic pattern of

faunal exchange between Sundaic and Indo-Burmese draconine lizards occurred approximately 50–30 mya (J.

Grismer et al. 2016) when the Indian subcontinent was farther south and east of its current position and formed a

land-positive connection with Sumatra and the Thai-Malay Peninsula prior to its collision with Central Asia (Ali &

Aitchison 2008). One of the invading Sundaic Cyrtodactylus lineages subsequently dispersed across an exposed

Sunda Shelf at approximately 18.0 mya (hpd 27.7–9.2 mya) to give rise to a diverse clade of Indochinese species of

which C. irregularis is a part (Wood et al. 2012; Agarwal et al. 2014; Sang et al. 2014). The other lineage dispersed

northward approximately 25.4 mya (hpd 31.4–19.3 mya) through a broad lowland area along the west side of the

emerging Tennaserim Mountains (Hall 2013), colonizing Eastern Myanmar. At approximately 23.3 mya (hpd 29.1–

16.8 mya), this Eastern Myanmar clade diverged into an ancestral lineage in the Salween Basin and an ancestral

lineage in the Shan Hills. The Shan Hills lineage invaded Indochina and diverged on opposite sides of the

Tenasserim Mountains and Thai Highlands at approximately 18.6 mya (hpd 26.5–10.6 mya), giving rise to the

intermedius group of Thailand, Cambodia, and Vietnam and the karst-adapted species of the linnwayensis group of

the Shan Hills. The ancestor of the Salween Basin lineage radiated into three monophyletic groups that now occupy

karstic habitat-islands in the region (Grismer et al. 2018). Two of these groups, the yathepyanensis and sinyeensis

groups independently invaded Indochina and also diverged on opposite sides of the Tenasserim Mountains with the

former giving rise to the oldhami group of western and southern Thailand approximately 16.0 mya (hpd 21.2–10.6

mya) and the latter giving rise to C. tigroides (and several unnamed species, Grismer et al. in prep.) in western

Thailand at 13.4 mya (hpd, 18.7–8.4 mya). The ancestor of the sadanensis group remained and radiated in the

karstic habitat-islands of the Salween Basin.

In a previous biogeographic interpretation of the exact same phylogeny save for the addition of Cyrtodacylus

ywanganensis sp. nov., Grismer et al. (2017a) posited a completely opposite scenario for the biogeography of the

Indochinese clade stating that the Salween Basin and Shan Hills lineages were the result of four independent

invasions from Indochina. This analysis, however was based on the biogeographic model of Wood et al. (2012) and

Agarwal et al. (2014) where Southeast Asia was not partitioned to separate Indochina from Eastern Myanmar.

However, by considering these areas as separate biogeographic regions along well-established tectonic boundaries

(Upton et al. 1995, 1997) and current geophysical features that correspond geographically with lineage turnovers

(Fig. 2), we can propose here a more fine-grained analysis. Furthermore, having the 16 additional species

composing the Indochinese clade of Eastern Myanmar that were not available to Wood et al. (2012) and Agarwal et

al. (2014), we can show that the Thai Highlands and the Tenasserim Mountains were a significant factor in

linnwayensis shwetaungorum ywanganensis sp. nov.

linnwayensis *** *** ***

shwetaungorum BB,LRT,PPS,PVT,4TL *** ***

ywanganensis sp. nov. IL,LRT,PS, PVT,VS,TL,TLE LRT,PPS,SL ***


cladogenesis within the Indochinese clade. Upton et al. (1995, 1997) indicate that this mountainous tectonic block

was uplifting from 21–16 mya which correlates with the independent divergences of the three pairs of sister

lineages on opposite sides of the uplift from approximately 18.6–13.4 mya. Other sister lineages occurring on

opposite sides of the Thai Highlands and Tenasserim Mountains include an endemic lineage composed of three

species of gekkonid lizards of the genus Hemiphyllodactylus from the Shan Hills and H. jinpingensis of southern

China (Grismer et al. 2017b) and well-differentiated subspecies of the White-browed Piculet (Sasia ochracea;

Fuchs et al. 2008). As more fine-scaled analyses from a broad range of taxa unfold across this poorly sampled area

of the Indomalayan Region, we predict more taxa will bear out this same phylogeographic structure.

TABLE 7. Biogeographic model selection with and without founder-event speciation(+j) based on the lowest AIC

values. d=rate of dispersal, e=rate of extinction, and j=relative probability of founder-event speciation at cladogenesis.

Discussion

It’s clear that karstic habitats in Southeast Asia are not only important areas for maintaining biodiversity that may

have otherwise been extirpated from the surrounding forested areas but they also serve as a substrate on which new

species can evolve (Grismer et al. 2017a; Grismer et al. 2018). The latter underscores Cyrtodactylus evolution in

Eastern Myanmar where 16 new karst-associated species have been discovered in only the last two years. In the

Salween Basin, 13 of these are found in karstic habitat-islands within a narrow zone extending only 95 km with

some species even being syntopic (Grismer et al. 2018). Unraveling the taxonomy and biogeography of these taxa

is a critical first step in conserving and protecting this unprecedented degree of site-specific endemism. To that end,

biogeographical scenarios should be based on appropriately scaled a posteriori constructed models that correspond

with the species diversity of the group under study, their phylogenetic relationships, and the geography of the

regions of concern. Such analyses should be ongoing and re-evaluated each time new species are added to the data

set. The inaccessibility of major tracts of karstic regions in the Shan Hills still leaves this vast landscape largely

unprotected and based on past experience, we believe there are dozens of karst-associated species yet to be

discovered. With assistance of Fauna & Flora International, the Forestry Department of Myanmar, and local

villagers, we are beginning to systematically access many of these remote areas.

Acknowledgements

We wish to thank Mr. Win Naing Thaw of the Ministry of natural Resources and Environmental Conservation

Forest Department for the collection and export permits. We thank the staff of the Shwe Gue Gu Hotel and Genious

Coffee for their hospitality. We are indebted to Mr. Thar Thar for letting us survey his property where the new

species was discovered. LLG thanks the College of Arts and Sciences of La Sierra University and Fauna & Flora

International for partial funding. Fieldwork for PLWJ was supported the Monte L. Bean Life Science Museum at

Brigham Young University and generation of molecular data was supported by the NSF grant EF-1241885 issued

to Jack W. Sites.

Model Max number of areas LnL No. of parameters d e J AIC

DEC 1 -30.8 2 1.629650e-03 1.000000e-12 0.00000000 65.64

DEC+J 1 -20.0 3 1.000000e-12 1.000000e-12 0.02781581 45.90

DIVALIKE 1 -28.6 2 2.065915-03 1.000000e-12 0.00000000 61.12

DIVALIKE+J 1 -19.2 3 1.000000e-12 1.000000e-12 0.02757908 44.43

BAYAREALIKE 1 -42.2 2 8.380688e-04 1.913213e-02 0.00000000 88.48

BAYAREALIKE+J 1 -21.5 3 1.000000e-07 1.000000e-12 0.03336197 48.96


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