Ohler et al. (2011)

Accepted by J. Padial: 17 Oct. 2011; published: 23 Dec. 2011

ZOOTAXAISSN 1175-5326 (print edition)

ISSN 1175-5334 (online edition)Copyright © 2011 · Magnolia Press

Zootaxa 3147: 1–83 (2011) www.mapress.com/zootaxa/ Monograph

ZOOTAXA

Sorting out Lalos: description of new species and additional taxonomic data on megophryid frogs from northern Indochina

(genus Leptolalax, Megophryidae, Anura)

ANNEMARIE OHLER1 *, KATHARINA C. WOLLENBERG2 , STÉPHANE GROSJEAN1, RALF HENDRIX3, 4 , MIGUEL VENCES5 , THOMAS ZIEGLER6 & ALAIN DUBOIS1

1 UMR 7205 OSEB, Département de Systématique et Evolution, Muséum National d’Histoire Naturelle, 25 rue Cuvier, CP 30, 75005 Paris, France.

2 Department of Organismic and Evolutionary Biology & Museum of Comparative Zoology, Harvard University, Cambridge, MA 02138, USA.

3 Department of Animal Behavior, University of Bielefeld, Morgenbreede 45, 33615 Bielefeld, Germany. 4 Zoologisches Forschungsmuseum Alexander Koenig, Adenauerallee 160, 53113 Bonn, Germany.

5 Zoological Institute, Division of Evolutionary Biology, Technical University of Braunschweig, Spielmannstraße 8, 38106 Braunschweig, Germany.

6 Cologne Zoo, Riehler Str. 173, 50735 Köln, Germany* Corresponding author

Magnolia PressAuckland, New Zealand

3147

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OHLER ET AL.2 · Zootaxa 3147 © 2011 Magnolia Press

ANNEMARIE OHLER, KATHARINA C. WOLLENBERG, STÉPHANE GROSJEAN, RALF HENDRIX,, MIGUEL VENCES, THOMAS ZIEGLER & ALAIN DUBOISSorting out Lalos: description of new species and additional taxonomic data on megophryid frogs from northern Indochina (genus Leptolalax, Megophryidae, Anura)(Zootaxa 3147)

83 pp.; 30 cm.

23 Dec. 2011

ISBN 978-1-86977-847-7 (paperback)

ISBN 978-1-86977-848-4 (Online edition)

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ISSN 1175-5326 (Print edition)

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Zootaxa 3147 © 2011 Magnolia Press · 3TAXONOMIC REVISION OF FROGS OF THE SUBGENUS LALOS

Table of contents

Abstract . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 4Material and methods . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 6

Field surveys and specimen collection . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 6Abbreviations used for collections . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 6Molecular analysis. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 7Morphometry of adult specimens . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 10Analysis of diagnostic characters in adult Leptolalax. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 11Colour pattern of adult Leptolalax . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 18Tadpole sampling and morphology . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 18Terminology . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 19

Results . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 20Molecular analysis. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 20Evidence from morphology and colour pattern. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 23Morphometric analysis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 25Systematics . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 27

Genus Leptolalax, subgenus Lalos Dubois, Grosjean, Ohler, Adler & Zhao, 2010. . . . . . . . . . . . . . . . . . . . . . . . . . . . 27Leptolalax (Lalos) pelodytoides (Boulenger, 1893) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 28Leptolalax (Lalos) oshanensis (Liu, 1950) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 31Leptolalax (Lalos) bourreti Dubois, 1983. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 32Leptolalax (Lalos) eos n. sp. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 39Leptolalax (Lalos) pluvialis Ohler, Marquis, Swan & Grosjean, 2000 . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 44Leptolalax (Lalos) nyx n. sp. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 46Leptolalax (Lalos) ventripunctatus Fei, Ye & Li, 1991 . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 50Leptolalax (Lalos) aereus Rowley, Stuart, Richards, Phimmachak & Sivongxay, 2010. . . . . . . . . . . . . . . . . . 56Leptolalax (Lalos) minimus (Taylor, 1962) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 63Key to species . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 71

Discussion . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 72Generic classification . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 72Species discrimination. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 73Natural history. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 76Taxonomic diversity and biogeographical considerations . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 77

Acknowledgements . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 78Literature cited. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 79Appendix 1. Additional specimens examined . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 83

Abstract

Frogs in the subgenus Lalos of the genus Leptolalax (Megophryidae) are highly diversified in continental Asia and consist ofabout 17 nominal species. These frogs are small, inconspicuous, and of high superficial morphological similarity.

We here formulate a hypothesis of phylogenetic relationships and assess the amount of genetic variation among genealog-ical lineages on the basis of 536bp of mitochondrial 16S rDNA sequences. Combining molecular data with a study of morpho-logy, morphometric divergence and geographical proximity, we tested hypotheses of species identity. We (1) used character-based and morphometric analyses to assign the onymophoronts (type specimens) of species in Lalos available to us to respec-tively one of the main clades, in order to propose the best potential correct taxonomic and nomenclatural allocation for the indi-viduals included in the molecular study, and (2) tried to also assign the historical museum specimens to these moleculartaxonomic units and to reclassify them whenever necessary.

We also used the molecular data to match tadpoles with adults and provide tadpole descriptions for species the larvae ofwhich were previously unknown. Specimens, that could neither be allocated to a molecularly characterised species (on the basisof their DNA “barcode”) nor to a morphologically defined species named on the basis of a type specimen, are described here asnew species. Based on this integrative set of data and analyses we describe two new species, Leptolalax eos n. sp. and Leptola-lax nyx n. sp., we resurrect Leptolalax minimus, and reassess the distribution of the species studied. We propose changes in theRed List status of L. pelodytoides and L. ventripunctatus and suggest a conservation status for the new species described herein.

Key words: biodiversity—molecular phylogenetics—morphology—taxonomy—type specimens —tadpoles



Introduction

The morphology of anurans shows a rather limited number of main phenotypes and a high degree of homoplasyamong major taxa (Roelants & Bossuyt 2005; Guayasamin et al. 2009). In many groups the variation among spe-cies is small and similarity among them is high (Ohler 1996). Frog taxonomy is further complicated by the exis-tence of numerous lineages of small and inconspicuous species that are difficult to recognize morphologically. Inaddition, some of these do not have a prominent calling behaviour, so that bioacoustic data can be scarce orunavailable as taxonomic evidence. Referring to ornithologists who use the term “Little Brown Bird” or LBB (e.g.,Campbell 1991) to refer to inconspicuous species difficult to discern by morphology, Schick et al. (2010) coinedthe term LBF “Little Brown Frogs” to refer to groups of inconspicuous anurans with high cryptic diversity.

The problems that taxonomists encounter in revising such groups of organisms, and the high rate of discoveryof populations and specimens that possibly represent new species, have led to a renewed discussion of concepts andwork protocols for the discovery and description of new species, in the framework of integrative taxonomy (Dayrat2005; Will et al. 2005). Vences et al. (2005), Padial & de la Riva (2007) and Fouquet et al. (2007) proposed DNAbarcoding as a suitable initial step to flag genetically divergent specimens and populations for further study. Vieiteset al. (2009) demonstrated the prospect of this approach to discover a large number of such lineages which theyrefer to as candidate species. Dubois (1977b), Padial et al. (2009, 2010), Padial & de la Riva (2009) and Glaw et al.(2010) discussed how different lines of evidence should be integrated in frog taxonomy and argued that, a priori,no particular taxonomic character—or set of characters—should be seen as superior in serving to recognize a lin-eage as evolutionary independent—and thus as a separate species in the sense of de Queiroz (2007).

One example of a taxonomically complex group of “little brown frogs” are the species in the megophryidgenus Leptolalax Dubois, 1980. The Megophryidae are a family of exclusively Oriental species of basal phyloge-netic position relative to the Neobatrachia (e.g., Frost et al. 2006; Roelants et al. 2007). Among these, they show acomparatively high species diversity. Currently 10 genera are recognized with a total of 156 species (Frost 2011).All species are forest floor inhabitants of toad-like morphology breeding in streams. Megophrys, Xenophrys,Brachytarsophrys, Borneophrys and Ophryophryne show palpebral appendages and cryptic coloration that mimicsleaf litter. The tadpoles of megophryids show three morphotypes (Grosjean 2004; Handrigan et al. 2007): a gener-alized type in Scutiger, Oreolalax and Leptobrachium, a suspension feeding tadpole in Xenophrys, Megophrys,Borneophrys, Brachytarsophrys and Ophryophryne and a stream-dwelling tadpole with elongated body-shape inLeptolalax and Leptobrachella (although the tadpole of Leptolalax kajangensis is quite similar in body shape tothose of Leptobrachium; Grismer, pers. comm.). All 30 species of Leptolalax have a very cryptic colour pattern andno obvious morphological characters useful in systematic studies (Rowley & Cao 2009, Rowley et al. 2010a,b,c).There is no published evidence for the monophyly of Leptolalax. Only Zheng et al. (2004) included more than onespecies of this genus (three specimens allocated to two species) in their analysis.

Delorme et al. (2006) recognized two subgenera in this genus based on a cladistic analysis of morphologicaldata: the nominal subgenus Leptolalax and the newly proposed subgenus Lalax. This nomen, however, was twicepreoccupied, by Lalax Hamilton, 1990 (Insecta, Hemiptera) and Lalax Holloway & Lane, 1998 (Trilobitomorpha),and was therefore replaced by Lalos Dubois, Grosjean, Ohler, Adler & Zhao, 2010. This subgenus, with 16 nomi-nal species (Dubois et al. 2010), is defined by the presence of a lateral glandular ridge on the belly, which is consid-ered to be an apomorphy, whereas the subgenus Leptolalax, with 12 species, is defined by the absence of thisstructure only and thus its monophyly remains to be confirmed. This classification is also reflected by the distribu-tion of species assigned to the two subgenera: Leptolalax is mainly distributed in the southern Oriental region,Lalos in the northern Oriental region. The following species were allocated to the subgenus Lalos (Dubois et al.2010): Leptolalax alpinus Fei, Ye & Li in Fei, Ye & Huang 1990, Leptolalax applebyi Rowley & Cao, 2009, Lepto-lalax bourreti Dubois, 1983, Leptolalax fuliginosus Matsui, 2006, Leptolalax khasiorum Das, Tron, Rangad &Hooroo, 2010, Leptolalax lateralis (Anderson, 1871), Leptolalax liui Fei & Ye in Fei, Ye & Huang 1991, Leptola-lax melanoleucus Matsui, 2006, Leptolalax nahangensis Lathrop, Murphy, Orlov & Ho, 1998, Leptolalax oshanen-sis (Liu, 1950), Leptolalax pelodytoides (Boulenger, 1893), Leptolalax pluvialis Ohler, Marquis, Swan & Grosjean,2000, Leptolalax sungi Lathrop, Murphy, Orlov & Ho, 1998, Leptolalax tamdil Sengupta, Sailo, Lalremsanga, Das& Das, 2010, Leptolalax tuberosus Inger, Orlov & Darevsky, 1999, Leptolalax ventripunctatus Fei, Ye & Li in Fei,Ye & Huang 1990. Additionally, the recently described species Leptolalax aereus Rowley, Stuart, Richards, Phim-machak & Sivongxay, 2010 and Leptolalax bidoupensis Rowley, Le, Tran & Hoang, 2011 show a lateroventral



gland and can therefore be allocated to Lalos as well. The other species Leptolalax croceus Rowley, Hoang, Le,Dau & Cao, 2010 and Leptolalax melicus Rowley, Stuart, Thy & Emmett, 2010, do not have glands on the side ofbelly and are tentatively allocated to subgenus Leptolalax. Nevertheless figure 2A of L. croceus in Rowley et al.(2010b) shows a series of glands in line on the side of belly which might be homologous to the lateroventral gland.Allocation of L. tuberosus to Lalos will need further evidence as in the original description absence of lateroventralgland is clearly stated (Inger et al. 1999). But for L. sungi, also described as having no such lateroventral gland, thestudy of the type specimens confirmed presence of such a structure, thus requiring allocation to the subgenus. As asingle character is used for this allocation, and as the lateroventral glands are not always very prominent and dis-tinct and can be overlooked, especially in preserved specimens, further evidence is needed to confirm this classifi-cation.

In this paper we deal with a series of species from northern Laos, northern Thailand and northern Vietnam. Thehistorically first described species that can be assigned to the subgenus Lalos is Ixalus lateralis Anderson, 1871.The status of this species remained unclear for more than a century. It was first assigned to the Rhacophoridae, thenconsidered by numerous authors as a synonym of Megalophrys major Boulenger, 1908 before being considered asa valid name and allocated to a precise population (for details see Humtsoe et al. 2008). Until the second half of20th century only a single other species assignable to Lalos had been described on morphological evidence: Lepto-brachium pelodytoides Boulenger, 1893 from Myanmar. Liu (1950) described populations from Sichuan as Mego-phrys oshanensis on the basis of morphology. Taylor (1962) recognized two species, that now are considered Lalos,from northern Thailand and described Lalos minimus (as Leptobrachium minimum) on the basis of morphology andcoloration. In fact, the description of the genus Leptolalax by Dubois (1980) and the recognition of this group byChinese researchers (first under the names “Paramegophrys” or “Carpophrys” which are unavailable nomina; seeDubois et al. 2010) were the starting point for the description of numerous species within the genus, most of whichhave restricted distributions. Leptolalax alpinus and L. ventripunctatus were recognized on the basis of morpholog-ical, cytological and osteological evidence and morphometric data have since been added to the original descrip-tions (Fei et al. 1991). The Fujian population originally identified as L. pelodytoides by Pope (1931) was separatedusing morphological and osteological evidence, and data on larval stages were used to confirm the identity of thenew species L. liui described in Fei et al. (1991). As the morphological definition of this species was “goodenough”, the identity of L. liui was not questioned by other authors and the name has been applied to geographi-cally close populations and did not create any taxonomic imbroglio. Recent species descriptions have been basedeither on morphological and coloration evidence (L. tuberosus, L. pluvialis, L. khasiorum, L. tamdil) or alsoincluded evidence from advertisement call (L. melanoleucus, L. fuliginosus, L. applebyi, L. kecil, L. aereus, L. cro-ceus). Rowley et al. (2010b) added evidence from DNA data to define the new species L. melicus. Das et al. (2010)published a comprehensive table listing morphological characters used for species diagnosis.

As one of the Principles of nomenclature is priority, the identity of the first described species should beresolved before description of new ones. To confirm the identity of L. pelodytoides known only from the onymo-phoronts (type specimens) would need study of recent topotypical material. The onymotope (type locality) of thisspecies lies in a region of Myanmar forbidden to non-natives, and which has suffered from civil war for manyyears. Specimens from Myanmar, Thailand, Laos, Vietnam and China have been allocated to L. pelodytoides(Bourret 1942; Taylor 1962; Chan-ard et al. 1999; Ziegler 2002). Several other nomina are available for Lalos fromthe Northern Mountain region as defined by Inger (1999). L. oshanensis (Liu, 1950), originally described fromSichuan, has been considered a synonym of L. minimus (Taylor, 1962) from Thailand by Dubois (1981). Anotherspecies, L. bourreti Dubois, 1983, was described on material collected more than 50 years before and not redefined;recently collected specimens have been allocated to this species mainly on the basis of origin and size. Someanswers to resolve these taxonomic problems may come from new collections which hold numerous specimens ofLalos from various parts of Laos, Thailand and Vietnam, some of which show sympatric differentiation.

According to our present knowledge, the distribution patterns of the species of Leptolalax are of two kinds:numerous species have been reported from very small areas and a few species have large areas of occurrence. Sofar, no comprehensive studies have been published to assess whether these are true biological patterns, or the wide-spread species are complexes of cryptic, undescribed species of restricted ranges. Clearly, to address such ques-tions, data sets complementary to morphology are necessary (e.g., Matsui 1997, 2006).

In this study, molecular data are used to test subgeneric classification. We use a combination of morphologicaland molecular data to assess patterns of differentiation among specimens of Leptolalax that were so far subsumed



under the names pelodytoides or oshanensis, and assess the limits of evolutionary lineages within this group. Wethen allocate available nomina to these evolutionary lineages. We include detailed data and figures of onymo-phoronts (name-bearing type specimens) and onymotopic (topotypical) specimens, and identify two species-levellineages for which no nomina are available, and describe these as new species: Leptolalax (Lalos) eos n. sp. andLeptolalax (Lalos) nyx n. sp.

Material & methods

Field surveys and specimen collection

Field surveys and collection of specimens were carried out during field trips in Laos, Thailand and Vietnam from1997 to 2006. Specimens were mostly collected at night by locating calling or silent specimens with head lights.Etho-ecological variables such as location of specimens, distance from ground, distance to water or habitat descrip-tion were taken for individual frogs. Photographs were taken either of living or freshly preserved specimens torecord colour pattern in life. Specimens were fixed in 5 % formalin for 24 hours or for some hours in ascendingorder in 40–60 % ethanol, and were subsequently transferred to 70 % ethanol. Samples for molecular analyses weretaken from muscle tissue, and were preserved in 70–98 % ethanol.

During the collection of some specimens (by RH), air temperature and relative humidity were measured with astandard digital thermo-hygrometer (accuracy: ± 1°C). Water temperature was measured using a standard digitalthermometer with sensing element (Alcron AC-100; accuracy: ± 1°C). Measurements of the pH-values of waterbodies were carried out by using pH-Fix 2.0–9.0 test strips (Macherey-Nagel; pH gradiation: 0.5). Altitudes weremeasured with an analog altimeter (Sunartis BKT 381; altitude gradation: 20 m) calibrated at sea level of the Gulfof Tonkin.

Specimens studied are deposited at various collections as indicated in Table 1.

TABLE 1. Abbreviation for institutions housing specimens studied in this work.

Abbreviation Name of institution

AMNH American Museum of Natural History, New York, USA

BMNH Natural History Museum, London, United Kingdom

CIB Chengdu Institute of Biology, Chengdu, China

FMNH Field Museum of Natural History, Chicago, USA

IEBR Institute of Ecology and Biological Resources, Hanoi, Vietnam

MNHN Muséum National d’Histoire Naturelle, Paris, France

MSNG Museo Civico di Storia Naturale di Genova Giacomo Doria, Genova, Italy

MCZ Museum of Comparative Zoology, Harvard, USA

PNNP Collection of the Phong Nha – Ke Bang National Park, Vietnam

RH Ralf Hendrix collection, Hannover, Germany

THNHM National Science Museum, Thailand Natural History Museum, Bangkok, Thailand

TNRC Thai National Research Council, Bangkok, Thailand

TZ Thomas Ziegler collection, Bonn, Germany

VNUH Collection of the Vietnam National University, Hanoi, Vietnam

ZFMK Zoologisches Forschungsmuseum Alexander Koenig, Bonn, Germany

ZMB Zoologisches Museum Berlin, Berlin, Germany

ZMH Zoologisches Museum Hamburg, Hamburg, Germany



Molecular analysis

Total genomic DNA was extracted using proteinase K (final concentration 1 mg/mL), and isolated by a standardsalt extraction protocol (Bruford et al. 1992). A fragment of the mitochondrial large ribosomal subunit (16S rRNA)was amplified via polymerase chain reaction (PCR) using the primers 16Sar-L and 16Sbr-H (Palumbi et al. 1991).Sequence data collection and visualization were performed on ABI 3130 automated sequencers (AppliedBiosystems). Sequences were deposited in GenBank (accession number JN848339 to JN848544). Our finalalignment contained 536bp of 16SrRNA and was submitted to Dryad (package identifier doi:10.5061/dryad.cc454m37). Pelobates fuscus was used as an outgroup taxon, and sequences of Pelodytes punctatus, variousXenophrys, Brachytarsophrys and Leptobrachium species were included in phylogeny reconstruction to obtainhierarchical outgroups (for GenBank accession numbers see Table 2). Several sequences of Leptolalax fromGenBank were added to our dataset (Table 2). We performed both Bayesian and maximum-likelihood (ML)analyses using the software RAxML (Stamatakis et al. 2008, accessed via the CIPRES portal, Miller et al. 2009)and Mrbayes 3.1 (Ronquist & Huelsenbeck 2003). ML analyses were performed with rapid bootstrapping, theoptimal number of bootstraps was estimated by RaxML as 400 (Stamatakis et al. 2008). For the Bayesian analysis,we used FINDMODEL at the HIV databases (http://hcv.lanl.gov/content/sequence/findmodel/findmodel.html,accessed 4 Feburary 2009) to compute the best-fit model of evolution using AIC. FINDMODEL selected the GTR+ G model of evolution. Analyses consisted of four Markov chains that ran for 3 × 106 generations, sampled every1000 generations, with a random starting tree and default priors. The burn-in was empirically estimated by plotting–lnL against the generation number, and the trees corresponding to the first 500,000 generations discarded. Forinterpreting the phylogenetic tree, we considered bootstrap values of 70 % and higher, and Bayesian support valuesof 0.95 and higher, as relevant support. Uncorrected pairwise genetic distances (p-distances) were calculated usingMEGA (V4.1, Tamura et al. 2007) and manually transformed into percentages.

For two clades of Leptolalax that showed high genetic intra-clade divergence and geographic variation, wereplaced the clades found by MrBayes with minimum-spanning haplotype networks that were manually connectedto the tree topology at their respective basal nodes. These networks inferred using statistical parsimony were con-structed to infer geographical aspects of gene genealogy using the software TCS (Clement et al. 2000).

TABLE 2. Mitochondrial DNA sequences used in this study.

Species Collection Number GenBank Accession No. Country Locality

Brachytarsophrys carinensis no voucher specimen JN848360 Myanmar undisclosed localityBrachytarsophrys carinensis no voucher specimen JN848361 Myanmar undisclosed localityLeptobrachium chapaense no voucher specimen JN848353 Vietnam Ben En NP, Thanh Hoa ProvinceLeptobrachium pullum MNHN 2000.2411 JN848342 Thailand Phang Nga, Phang Nga ProvinceLeptobrachium pullum P925 JN848340 Thailand Phang Nga, Phang Nga ProvinceLeptobrachium pullum P926 JN848341 Thailand Phang Nga, Phang Nga ProvinceLeptobrachium pullum P981 JN848343 Thailand Phang Nga, Phang Nga ProvinceLeptobrachium smithi MNHN 2006.2303 JN848346 Laos Ban Keng Koung, Luang Prabang ProvinceLeptobrachium smithi MNHN 2006.2304 JN848345 Laos Ban Sop Khao, Luang Prabang ProvinceLeptobrachium smithi MNHN 2006.2305 JN848344 Laos Ban Sop Khao, Luang Prabang ProvinceLeptobrachium smithi MNHN 2006.2306 JN848350 Laos Ban Vang Thong, Luang Prabang ProvinceLeptobrachium smithi MNHN 2006.2307 JN848347 Laos Ban Vang Thong, Luang Prabang ProvinceLeptobrachium smithi MNHN 2006.2555 JN848349 Laos Luang Prabang, Luang Prabang ProvinceLeptobrachium smithi MNHN 2006.2556 JN848351 Laos Luang Prabang, Luang Prabang ProvinceLeptobrachium sp. no voucher specimen JN848348 India Guwahati, AssamLeptobrachium sp. no voucher specimen JN848352 Thailand Doi Chiang Dao, Chiangmai Province

Leptolalax aereus no voucher specimen JN848435 Vietnam Cha Noi region, Phong Nha - Ke Bang: Quang Binh Province

......continued on the next page



TABLE 2. (Continued)







Leptolalax aereus no voucher specimen JN848405 Vietnam U Bo, Quang Binh ProvinceLeptolalax aereus no voucher specimen JN848406 Vietnam U Bo, Quang Binh ProvinceLeptolalax aereus no voucher specimen JN848407 Vietnam U Bo, Quang Binh ProvinceLeptolalax aereus no voucher specimen JN848408 Vietnam U Bo, Quang Binh ProvinceLeptolalax aereus no voucher specimen JN848443 Vietnam U Bo, Quang Binh Province Leptolalax aereus ZFMK 86362 JN848409 Vietnam U Bo, Quang Binh Province

Leptolalax aereus ZFMK 86393 JN848442 Vietnam Cha Noi region, Phong Nha - Ke Bang: Quang Binh Province

Leptolalax aereus ZFMK 86394 JN848439 Vietnam Cha Noi region, Phong Nha - Ke Bang: Quang Binh Province

Leptolalax bourreti MNHN 1999.5659 JN848453 Vietnam Sa Pa, Lao Cai ProvinceLeptolalax bourreti MNHN 1999.5660 JN848454 Vietnam Sa Pa, Lao Cai ProvinceLeptolalax eos MNHN 2004.0274 JN848452 Laos Long Nai, Phongsaly ProvinceLeptolalax eos MNHN 2004.0275 JN848451 Laos Long Nai, Phongsaly ProvinceLeptolalax eos MNHN 2004.0276 JN848446 Laos Long Nai, Phongsaly ProvinceLeptolalax eos MNHN 2004.0277 JN848448 Laos Long Nai, Phongsaly ProvinceLeptolalax eos MNHN 2004.0278 JN848450 Laos Long Nai, Phongsaly ProvinceLeptolalax eos MNHN 2004.0279 JN848447 Laos Long Nai, Phongsaly ProvinceLeptolalax eos MNHN 2005.0147 JN848449 Laos Long Nai, Phongsaly ProvinceLeptolalax heteropus MNHN 2000.2412 JN848356 Thailand Long Nai, Long Nai ProvinceLeptolalax heteropus MNHN 2000.2414 JN848357 Thailand Long Nai, Long Nai ProvinceLeptolalax heteropus no voucher specimen JN848354 Thailand Long Nai, Long Nai ProvinceLeptolalax heteropus no voucher specimen JN848355 Thailand Long Nai, Long Nai ProvinceLeptolalax heteropus no voucher specimen JN848358 Thailand Long Nai, Long Nai ProvinceLeptolalax heteropus no voucher specimen JN848359 Thailand Long Nai, Long Nai ProvinceLeptolalax minimus MNHN 2006.2308 JN848379 Laos Ban Dong Khan, Luang Prabang ProvinceLeptolalax minimus MNHN 2006.2309 JN848378 Laos Ban Dong Khan, Luang Prabang ProvinceLeptolalax minimus MNHN 2006.2310 JN848377 Laos Ban Dong Khan, Luang Prabang ProvinceLeptolalax minimus MNHN 2006.2311 JN848376 Laos Ban Dong Khan, Luang Prabang ProvinceLeptolalax minimus MNHN 2006.2312 JN848382 Laos Ban Dong Khan, Luang Prabang ProvinceLeptolalax minimus MNHN 2006.2313 JN848385 Laos Ban Dong Khan, Luang Prabang ProvinceLeptolalax minimus MNHN 2006.2314 JN848383 Laos Ban Dong Khan, Luang Prabang ProvinceLeptolalax minimus MNHN 2006.2315 JN848373 Laos Ban Dong Khan, Luang Prabang ProvinceLeptolalax minimus MNHN 2006.2316 JN848384 Laos Ban Dong Khan, Luang Prabang ProvinceLeptolalax minimus MNHN 2006.2317 JN848375 Laos Ban Nong Di, Luang Prabang ProvinceLeptolalax minimus MNHN 2006.2318 JN848380 Laos Ban Nong Di, Luang Prabang Province






Leptolalax minimus MNHN 2006.2319 JN848397 Laos Ban Nong Di, Luang Prabang ProvinceLeptolalax minimus MNHN 2006.2320 JN848371 Laos Ban Nong Di, Luang Prabang ProvinceLeptolalax minimus MNHN 2006.2321 JN848396 Laos Ban Nong Di, Luang Prabang ProvinceLeptolalax minimus MNHN 2006.2322 JN848372 Laos Ban Nong Di, Luang Prabang ProvinceLeptolalax minimus MNHN 2006.2323 JN848394 Laos Ban Nong Di, Luang Prabang ProvinceLeptolalax minimus MNHN 2006.2324 JN848395 Laos Ban Nong Di, Luang Prabang ProvinceLeptolalax minimus MNHN 2006.2325 JN848393 Laos Ban Nong Di, Luang Prabang ProvinceLeptolalax minimus MNHN 2006.2326 JN848392 Laos Ban Nong Di, Luang Prabang ProvinceLeptolalax minimus MNHN 2006.2328 JN848381 Laos Ban Vang Thong, Luang Prabang ProvinceLeptolalax minimus MNHN 2006.2553 JN848374 Laos Huey Thao, Luang Prabang ProvinceLeptolalax minimus MNHN 2006.2554 JN848367 Laos Huey Thao, Luang Prabang ProvinceLeptolalax minimus MNHN 2006.2599 JN848386 Laos Ban Nong Di, Luang Prabang ProvinceLeptolalax minimus MNHN 2006.2614 JN848387 Laos Ban Nong Di, Luang Prabang ProvinceLeptolalax minimus no voucher specimen JN848366 Thailand Mae Lao Mae Sae, Chiangmai ProvinceLeptolalax minimus no voucher specimen JN848368 Thailand Mae Lao Mae Sae, Chiangmai ProvinceLeptolalax minimus no voucher specimen JN848369 Thailand Mae Lao Mae Sae, Chiangmai ProvinceLeptolalax minimus no voucher specimen JN848370 Thailand Mae Lao Mae Sae, Chiangmai ProvinceLeptolalax minimus no voucher specimen JN848403 Thailand Mae Lao Mae Sae, Chiangmai ProvinceLeptolalax minimus no voucher specimen JN848404 Thailand Doi Chiang Dao, Chiangmai ProvinceLeptolalax minimus THNHM 07417 JN848398 Thailand Doi Chiang Dao, Chiangmai ProvinceLeptolalax minimus THNHM 07418 JN848402 Thailand Doi Chiang Dao, Chiangmai ProvinceLeptolalax minimus THNHM 07419 JN848401 Thailand Doi Chiang Dao, Chiangmai ProvinceLeptolalax minimus THNHM 07420 JN848399 Thailand Doi Chiang Dao, Chiangmai ProvinceLeptolalax minimus THNHM 07421 JN848398 Thailand Doi Chiang Dao, Chiangmai ProvinceLeptolalax minimus THNHM 07422 JN848400 Thailand Doi Chiang Dao, Chiangmai ProvinceLeptolalax pluvialis MNHN 1999.5674 JN848389 Vietnam Sa Pa, Lao Cai ProvinceLeptolalax pluvialis MNHN 1999.5675 JN848391 Vietnam Sa Pa, Lao Cai ProvinceLeptolalax pluvialis MNHN 1999.5676 JN848390 Vietnam Sa Pa, Lao Cai ProvinceLeptolalax sp. no voucher specimen JN848455 Thailand Doi Chiang Dao, Chiangmai ProvinceLeptolalax ventripunctatus MNHN 2004.0270 JN848419 Laos Long Nai, Phongsaly ProvinceLeptolalax ventripunctatus MNHN 2004.0272 JN848414 Laos Long Nai, Phongsaly ProvinceLeptolalax ventripunctatus MNHN 2004.0273 JN848420 Laos Long Nai, Phongsaly ProvinceLeptolalax ventripunctatus MNHN 2005.0117 JN848429 Laos Long Nai, Phongsaly ProvinceLeptolalax ventripunctatus MNHN 2005.0118 JN848428 Laos Long Nai, Phongsaly ProvinceLeptolalax ventripunctatus MNHN 2005.0119 JN848430 Laos Long Nai, Phongsaly ProvinceLeptolalax ventripunctatus MNHN 2005.0120 JN848427 Laos Long Nai, Phongsaly ProvinceLeptolalax ventripunctatus MNHN 2005.0121 JN848426 Laos Long Nai, Phongsaly ProvinceLeptolalax ventripunctatus MNHN 2005.0122 JN848421 Laos Long Nai, Phongsaly ProvinceLeptolalax ventripunctatus MNHN 2005.0123 JN848418 Laos Long Nai, Phongsaly ProvinceLeptolalax ventripunctatus MNHN 2005.0124 JN848417 Laos Long Nai, Phongsaly ProvinceLeptolalax ventripunctatus MNHN 2005.0125 JN848425 Laos Long Nai, Phongsaly ProvinceLeptolalax ventripunctatus MNHN 2005.0126 JN848424 Laos Long Nai, Phongsaly ProvinceLeptolalax ventripunctatus MNHN 2005.0111 JN848433 Laos Phongsaly, Phongsaly Province Leptolalax ventripunctatus MNHN 2005.0112 JN848431 Laos Phongsaly, Phongsaly Province Leptolalax ventripunctatus MNHN 2005.0113 JN848432 Laos Phongsaly, Phongsaly Province




Morphometry of adult specimens

Measurements were taken by the second author on adult specimens using a digital slide caliper, or for values below5 mm, with an ocular micrometer. All measurements are given with a precision to the nearest 0.1 mm as demandedby the journal although this is much below the resolution of measurements taken with ocular micrometer.

Body. SVL snout-vent length. Head. EL eye length; EN distance from anterior corner of eye to posterior edgeof nostril; HL head length from posterior corner of mandible to tip of snout; HW head width, at the angle of jaws;IBE distance between posterior corners of eyes; IFE distance between anterior corners of eyes; IN internarial dis-tance; IUE minimum distance between upper eyelids; MBE distance from posterior corner of mandible to posteriorcorner of eye; MFE distance from posterior corner of mandible to anterior corner of eye; MN distance from poste-rior corner of mandible to posterior edge of nostril; NS distance from anterior edge of nostril to tip of snout; SL dis-tance from anterior corner of eye to tip of snout; TYD maximum tympanum diameter; TYE distance betweenanterior margin of tympanum and posterior corner of eye; UEW maximum width of upper eyelid. Forelimb. FLLforearm length, from elbow to base of outer palmar tubercle; HAL hand length, from base of outer palmar tubercleto tip of third finger; TFL third finger length, from articulation of proximal and intermediate phalange. Hindlimb.FFTF distance from maximum incurvation of web between fourth and fifth toe to tip of fourth toe, toes beingspread; FL thigh length, from vent to knee; FOL foot length, from base of inner metatarsal tubercle to tip of fourth toe;FTL fourth toe length, from articulation of proximal and intermediate phalange; IMT length of inner metatarsal tuber-cle; ITL inner toe length; MTFF distance from distal edge of metatarsal tubercle to maximum incurvation of webbetween fourth and fifth toe, toes being spread; MTTF distance from distal edge of metatarsal tubercle to maximumincurvation of web between third and fourth toe, toes being spread; TFOL length of tarsus and foot from base oftarsus to tip of fourth toe; TFTF distance from maximum incurvation of web between third and fourth toe to tip offourth toe, toes being spread; TL shank length; TW maximum shank width.

The sample consisted of 363 items composed of 353 specimens (184 males, 39 females and 130 larvae) ofwhich 83 samples where available for molecular analysis, plus 14 tissue samples of specimens not available; thus atotal of 97. Samples were analysed by separating into groups by sex, age (adult, juvenile) and populations. As num-ber of female specimens per species and population was too low for statistical analysis, and the samples could notbeen tested for sexual differentiation, only adult male specimens were included in further tests. For every analysis,composition of sample is indicated in the text and legends of figures and tables.



Leptolalax ventripunctatus MNHN 2005.0114 JN848434 Laos Phongsaly, Phongsaly Province Leptolalax ventripunctatus MNHN 2005.0115 JN848444 Laos Phongsaly, Phongsaly Province Leptolalax ventripunctatus MNHN 2005.0116 JN848410 Laos Phongsaly, Phongsaly Province Leptolalax ventripunctatus MNHN 2005.0137 JN848422 Laos Phongsaly, Phongsaly Province Leptolalax ventripunctatus MNHN 2005.0138 JN848416 Laos Phongsaly, Phongsaly Province Leptolalax ventripunctatus MNHN 2005.0139 JN848339 Laos Phongsaly, Phongsaly Province Leptolalax ventripunctatus MNHN 2005.0140 JN848388 Laos Phongsaly, Phongsaly Province Leptolalax ventripunctatus MNHN 2005.0142 JN848412 Laos Phongsaly, Phongsaly Province Leptolalax ventripunctatus MNHN 2005.0143 JN848413 Laos Phongsaly, Phongsaly Province Leptolalax ventripunctatus MNHN 2005.0144 JN848411 Laos Phongsaly, Phongsaly Province Leptolalax ventripunctatus MNHN 2005.0271 JN848445 Laos Phongsaly, Phongsaly ProvinceLeptolalax ventripunctatus MNHN 2005.0145 JN848423 Laos Houey Phihet, Phongsaly ProvinceXenophrys parva MNHN 1999.5694 JN848362 Vietnam Sa Pa, Lao Cai ProvinceXenophrys sp. K3359 JN848364 Laos Ban Keng Koung, Luang Prabang ProvinceXenophrys sp. MNHN 2006.2329 JN848365 Laos Ban Keng Koung, Luang Prabang ProvinceXenophrys sp. no voucher specimen JN848363 Thailand Doi Chiang Dao, Chiangmai Province



For univariate comparisons between samples, all measurements except snout-vent length (SVL) were trans-formed into their ratio to SVL, expressed in per thousands (‰). For multivariate analysis, data were transformedinto their base 10 logarithm to correct for size.

Calculations and statistical analyses were performed using the program SPSS 16.0 (Norusis 1992). Mean, stan-dard deviation, minimum and maximum were obtained for all variables. Principal Component Analysis (PCA) wasperformed on log-transformed variables. All genetically identified individuals were included in this analysis. PCAfactors 1 to 3 were plotted as a scatterplot indicating allocation of individuals without molecular data to MolecularOperational Taxonomic Units (MOTU; Floyd et al. 2002) as given by genetic data. To examine effects of speciesdelimitation on the principal component scores, factorial ANOVA were performed in SPSS for subsamples largerthan 1. Discriminant analysis was performed to find discriminant variables for genetically identified groups ofmore than one specimen that show no significant differences in ANOVA.

Analysis of diagnostic characters in adult Leptolalax

Descriptions of onymophoronts follow standards of Ohler (1996). The following characters were noted by the firstauthor for all specimens studied and in combination are useful to diagnose species of Leptolalax. For species thathave not been studied by AMO, the original descriptions or other available descriptions have been used for charac-ter definition. This poses the problem of character definition by different authors which are not standardized. Tohomogenize expressions used by different authors, a comparison of descriptions of characters used in tables 3–5was done and is discussed below. Most of the character states show continuous variation, and cutting them to dif-ferent states is an issue of description of existing phenomena, which involves subjectivity and never can replace theobserved object. As often as possible, measurements of comparative descriptions were used in order to limit sub-jectivity. All morphological studies were done by a single observer in order to avoid interobserver differences.

Snout-vent length: Species vary in snout-vent length of adult males and adult females. In large species, malescan reach about 40 mm in SVL, whereas in small species they reach only about 22 mm. Although intraspecificvariation is high, particularly due to sexual dimorphism and age groups, the size of adult specimens of either sex isuseful in species recognition.

Tympanum: Most species of Leptolalax have a tympanum visible by external examination. Some authorsdescribe the presence of a tympanum visible by eye as “distinct”, a few as “visible” (Grismer et al. 2004, Matsui2006). When the tympanum cannot be seen, it is described as “obscured” (Inger et al. 1999) or “indistinct” (Row-ley et al. 2010b). Most authors indicate tympanum size in relation to eye size and distance of tympanum to eye inrelation to tympanum size. In some papers the shape of the tympanic annulus or rim is described as either “tym-panic annulus not elevated relative to skin of temporal region” (Rowley & Cao 2009, Rowley et al. 2010b), “rimslightly elevated relative to skin of temporal region” (Fei & Ye 1992), “tympanic rim narrow but prominent” (Daset al. 2010), “rim not visible” (Inger et al. 1999, Rowley et al. 2010b). In this work, the tympanum is described asdistinct or indistinct and its size relative to eye as its distance from eye is given as simple fraction of eye size for thefirst, or tympanum diameter for the second. Tympanic annulus is indicated as “visible” or “not visible”, whenpresent its elevation relative to skin of temporal region is indicated. Thus, presence or absence of externally visibletympanum can be used in species recognition (Fig. 1a).

Tongue shape: In all species for which this character was observed, tongue is moderate to large in size. It has aterminal notch of various dimensions thus giving it a rounded or cordate (heart-shaped) aspect.

Vocal sacs: In anurans vocal sacs can be absent or present which can be seen by the presence of openings onthe mouth floor. When present, the vocal sacs can be visible externally on throat or not. Vocal sacs that areexternally visible are called “external”, those that cannot be distinguished by external observation are called“internal”. External vocal sacs can be single or paired and the skin of various parts of the throat can be modified. InLeptolalax, vocal sac openings seem to be present in all species where adult males have been studied. Interspecificvariation is observed in the presence of external vocal sacs. These vocal sacs have been described as paired inLeptolalax and can be observed as small sacs formed by transparent skin on sides of throat near the jawarticulations.



Tips of digits: Even though the tips of digits in Leptolalax do not bear differentiated structures such as digitalpads, shape and width of the digit tips is variable among species and can be used in diagnoses. Shape of tips of dig-its has been described as “truncate” (Taylor 1962), “blunt” (Inger et al. 1999, Rowley et al. 2010a) or “rounded”(Liu 1950, Inger et al. 1999, Matsui 1997, Grismer et al. 2004, Humtsoe et al. 2008, Rowley & Cao 2009, Das etal. 2010, Rowley et al. 2010a,c, Sengupta et al. 2010). The states here recognized for tips of digits are: rounded,blunt and truncate. Differences between these shapes are subtle. Fingers tips may be enlarged in comparison to ter-minal phalanges (Rowley et al. 2010b,c).

Fringes on toes and fingers: Toes and fingers of Leptolalax species can bear lateral fringes. These fringes havebeen described as “absent” (Liu 1950, Fei & Ye 1992, Lathrop et al. 1998, Rowley et al. 2010b), “no lateralfringes” (Fei & Ye 1992, Rowley et al. 2010a) or “without lateral fringes” (Inger et al. 1999, Matsui 2006) when nosuch structure is observable. The state of a poorly developed fringe was called “narrow” (Dubois 1987), “poorlydeveloped” (Humtsoe et al. 2008), “weak” (Lathrop et al. 1998), “weakly fringed” (Matsui 2006) or “with veryweak lateral fringing” (Rowley & Cao 2009), whereas the state of a well developed fringe was described as “large”(Fei & Ye 1992). The different states of fringes present on toes in Leptolalax are described in this work as: absent,when no fringes can be observed; weak, when fringes are poorly developed; large, when fringes are distinctlydeveloped (Fig. 1b).

Ridges under fingers and toes: Specimens of Leptolalax do not bear subarticular tubercles but longitudinalridges extending on phalanges and interrupted at the articulations. Presence or absence and shape of these ridgesare variable (Fig. 1b).

Webbing: The terms describing webbing are not consistent between authors. In L. pelodytoides, the webbingwas described as “small” which is consistent when compared to other anuran species even though in Leptolalax theextension is the largest in this species: less than 4 phalanges of toe IV are free. The other extreme of extension ofwebbing is when webbing is described as “absent” or “no web” (Liu 1950). In this state no web is observablebetween toes. A vestige of web is present in species with web described as “rudimentary” (Matsui 1997, Rowley etal. 2010a,b). Numerous expressions have been used to describe the state of webbing when a distinct fringe isobservable between the toes, such as “with a web-reminant at base” (Taylor 1962), “at bases of toes” (Matsui1997), “confined to bases” (Inger 1999, Matsui 2006), “basal between toes” (Lathrop et al. 1998) “basal” or “up tobase of phalanges” (Das et al. 2010, Sengupta et al. 2010). Here we use the following terms to describe webbing inLeptolalax: absent, no free web observable; rudimentary, a vestige of web observable; basal, a distinct fringe ofweb observable; small, less than 4 phalanges of toe IV free of web.

Dorsal skin: On the dorsum of Leptolalax, skin has been described as “smooth” (Boulenger 1900, Rowley &Cao 2009), “roughly shagreened” (Matsui 1997), or with “glandular ridges” (Liu 1950), “tubercles” (Taylor 1962,Malkmus 1992, Lathrop et al. 1998, Inger et al. 1999, Matsui 1997, 2006, Matsui et al. 2009, Das et al. 2010, Row-ley et al. 2010a,b,c, Sengupta et al. 2010), “lumps” (Inger & Stubbing 2005), “peepled” (Grismer et al. 2004). Herewe refer as tubercles to rounded structures on skin and to ridges to such elongate structures. The size, extension anddensity of such ridges and tubercles can be used to diagnose species (Fig. 2). There is a need to be careful withusing skin texture as a character. Texture is often lost after some time in preservative, and varies between sexes insome species of Leptolalax.



TABL

E 3.

Mea

sure

men

ts of

spec

imen

s of L

epto

lala

x (L

alos

). D

ata

are

eith

er fr

om o

bser

vatio

n of

spec

imen

s men

tione

d in

the

rele

vant

cha

pter

s on

spe

cies

or f

rom

lite

ratu

re. I

n sq

uare

d br

acke

ts: if

not

ava

ilabl

e as

mea

sure

s in

mm

ratio

s of

SV

L ex

pres

sed

as p

er th

ousa

nds

are

give

n. F

or

refe

renc

es se

e Ta

ble

5.

Mal

es

Fe

mal

es

Spec

ies

Prov

ince

of

onym

otop

e Co

untry

SV

L H

W

HL

TL

SVL

HW

H

L TL

Lept

olal

ax a

ereu

s H

a Ti

nh

Vie

tnam

29.8

–29.

8 10

.5–1

1.0

11.1

–11.

5 14

.5–1

4.6

– –

– –

Lept

olal

ax a

lpin

us

Yun

nan

Chin

a 24

.0–2

6.4

– –

– 31

.7–3

2.5

– –

–

Lept

olal

ax b

ourr

eti

Lao

Cai

Vie

tnam

28.0

–36.

2 10

.1–1

3.4

10.8

–14.

1 14

.5–1

7.8

42.0

–45.

0 15

.8–1

6.1

17.1

–17.

1 21

.4–2

1.7

Lept

olal

ax e

os

Phon

gsal

y La

os

33.1

–34.

7 11

.5–1

2.7

13.0

–13.

6 16

.4–1

7.9

40.7

14

.2

16.6

20

.6

Lept

olal

ax fu

ligin

osus

Pr

achu

ap K

hiri

Kha

n Th

aila

nd28

.2—

30.0

[3

44—

380]

[3

77—

393]

[4

67—

504]

- -

- -

Lept

olal

ax k

hasio

rum

M

egha

laya

In

dia

24.5—

27.3

9.

3—10

.1

6.0—

8.3

12.2—

12.9

21

.2—

33.4

10

.8—

11.7

7.

8—8.

8 14

.3—

14.8

Lept

olal

ax la

tera

lis

Nag

alan

d In

dia

26.9

0–28

.25

8.8–

9.1

8.4–

8.8

12.1

–12.

9 36

.6

12.2

11

.2

15.8

Lept

olal

ax li

ui

Fujia

n Ch

ina

22.5

–26.

5 8.

0–9.

6 8.

4–9.

8 11

.3–1

2.8

24.5

–27.

8 7.

9–10

.1

9.9–

10.6

11

.8–1

3.5

Lept

olal

ax m

elan

oleu

cus

Sura

t Tha

ni

Thai

land

26.6—

28.8

[3

40—

382]

[3

79—

411]

[4

58—

511]

32.7

[3

67]

[388

] [4

83]

Lept

olal

ax m

inim

us

Chia

ng M

ai

Thai

land

25.7

–31.

4 9.

0–10

.6

9.7–

11.5

11

.6–1

4.0

31.6

–37.

3 11

.3–1

2.8

11.9

–14.

5 14

.1–1

8.8

Lept

olal

ax m

inim

us fo

rm 2

Ch

iang

Mai

Th

aila

nd25

.6–2

8.2

8.7–

9.6

10.2

–11.

1 12

.2–1

3.5

36.4

12

.6

13.4

17

.1

Lept

olal

ax n

ahan

gens

is Tu

yen

Qua

ng

Vie

tnam

40.8

15

.6

15.0

20

.1

– –

– –

Lept

olal

ax n

yx

Ha

Gia

ng

Vie

tnam

22.4

–24.

2 7.

6–8.

7 8.

3–9.

9 10

.4–1

2.2

28.3

9.

8 10

.4

13.4

Lept

olal

ax o

shan

ensis

Si

chua

n Ch

ina

26.6

–30.

7 –

– –

31.6

–

– –

Lept

olal

ax p

elod

ytoi

des

Kar

in B

ia-p

o M

yanm

ar27

.5–3

2.3

9.8–

11.2

10

.4–1

2.6

13.7

–14.

7 35

.5–3

7.8

12.2

–13.

2 12

.7–1

4.0

15.6

–16.

7

Lept

olal

ax p

luvi

alis

Lao

Cai

Vie

tnam

22.0

–23.

0 8.

1–8.

8 8.

6–9.

1 11

.4–1

2.0

– –

– –

Lept

olal

ax su

ngi

Vin

h Ph

u V

ietn

am48

.3–5

2.7

18.9

–20.

3 19

.3–2

1.0

22.8

–23.

1 56

.7–5

8.9

21.2

–21.

9 23

.0–2

4.1

23.6

–26.

1

Lept

olal

ax ta

mdi

l M

izor

am

Indi

a 32

.3

12.0

8.

7 16

.0

31.8

12

.0

8.8

15.7

Lept

olal

ax tu

bero

sus

Gia

-Lai

V

ietn

am24

.4–2

9.5

9.9

10.0

13

.5

30.2

–

– –

Lept

olal

ax v

entri

punc

tatu

s Y

unna

n Ch

ina

23.7

–27.

7 7.

6–9.

9 8.

2–11

.0

10.7

–13.

4 31

.5–3

5.0

10.4

–11.

9 11

.5–1

3.9

14.6

–15.

4





TABL

E 5.

Col

orat

ion

patte

rns a

nd e

ye c

olou

r of s

peci

men

s of L

epto

lala

x (La

los)

. Dat

a ar

e ei

ther

from

obs

erva

tion

of sp

ecim

ens m

entio

ned

in

the

rele

vant

cha

pter

s on

spec

ies o

r fro

m li

tera

ture

. Sp

ecie

s D

orsa

l pat

tern

La

tera

l pat

tern

V

entra

l pat

tern

Iri

s col

our

Refe

renc

es

Lept

olal

ax a

ereu

s G

rey

brow

n w

ith d

ark

patte

rn

Whi

te a

nd d

ark

spot

s W

hitis

h w

ith in

disti

nct b

row

n ve

rmic

ulat

ion

Gol

d co

pper

Th

is pa

per

Lept

olal

ax a

lpin

us

Gre

y br

own

with

spot

s Fe

w d

ark

spot

sSp

ots o

n sid

es o

f che

st an

d be

lly

Ligh

t ora

nge

Feie

t al.1

992;

Fei

19

99

Lept

olal

ax b

ourr

eti

Redd

ish, g

reen

ish, b

row

n w

ith d

ark

spot

s M

oder

ate

dark

sp

ots

Whi

tish

Var

iabl

e, c

lear

er a

bove

, co

pper

y, g

reen

, bro

wni

sh

Bour

ret 1

942;

this

pape

r

Lept

olal

ax e

os

Brow

n gr

ey w

ith d

arke

r pa

ttern

N

o sp

ots

Whi

te

Ora

nge

abov

e, li

ght g

olde

n be

low

Th

is pa

per

Lept

olal

ax fu

ligin

osus

Li

ght b

row

n sc

atte

red

with

fa

int d

ark

spot

s Se

vera

l lar

ge

dark

spot

s W

hite

dus

ted

with

fine

bro

wn

netw

ork

late

rally

Redd

ish o

rang

e in

upp

er h

alf,

gold

en re

ticul

ated

with

bla

ck

belo

w

Mat

sui 2

006

Lept

olal

ax k

hasio

rum

Br

owni

sh-g

rey

with

irr

egul

ar m

id-b

row

n or

da

rk g

rey

blot

ches

Irreg

ular

dar

k sp

ots

Crea

m

Upp

er th

ird b

right

ora

nge,

rest

yello

wish

-cre

am

Das

et a

l. 20

10

Lept

olal

ax la

tera

lis

One

tria

ngul

ar o

r som

e irr

egul

ar d

ark

spot

s Fe

w sm

all d

ark

spot

s Pa

le y

ello

wish

mar

bled

with

br

own

- H

umtso

e et

al.

2008

Lept

olal

ax li

ui

Gre

y br

own

with

spot

s In

disti

nct s

pots

Imm

acul

ate

Brow

nish

Fe

i & Y

e199

2; F

ei

1999

Lept

olal

ax m

elan

oleu

cus

Oliv

e br

own

with

indi

stinc

t da

rk b

row

n m

arki

ngs

Smal

l dar

k sp

ots

Whi

te m

ottle

d w

ith la

rge,

co

ntin

uous

bla

ck m

arki

ngs

Ora

nge

in u

pper

hal

f, sil

ver

retic

ulat

ed w

ith b

lack

on

low

er

half

Mat

sui 2

006

Lept

olal

ax m

inim

us

Brow

n

Dar

k sp

ots

Whi

tish

Dar

k go

ld a

bove

, gre

y be

low

Th

is pa

per

Lept

olal

ax m

inim

us fo

rm 2

Dar

k br

own

with

poo

rly

disti

nct p

atte

rn

Smal

l bla

ckish

sp

ots

Whi

tish

Ora

nge

abov

e, d

ark

grey

be

low

Th

is pa

per

Lept

olal

ax n

ahan

gens

is La

vend

er-b

row

n w

ith

larg

e, ir

regu

lar m

ottli

ng

Larg

e sp

ots

Ligh

t spe

cklin

g on

thro

at a

nd

ches

t G

old

with

min

ute

blac

k re

ticul

atio

ns

Lath

rop

et a

l. 19

98

Lept

olal

ax n

yx

Gre

yish

-bro

wn

with

dar

k pa

ttern

Po

orly

dist

inct

sp

ots

Whi

tish

-

This

pape

r

Lept

olal

ax o

shan

ensis

Re

ddish

bro

wn

with

spot

s Fe

w la

rge

dark

sp

ots

Thro

at w

ith d

ots,

belly

im

mac

ulat

e Co

pper

abo

ve, g

olde

n be

low

Li

u 19

50; F

ei 1

999

Lept

olal

ax p

elod

ytoi

des

Brow

n w

ith in

disti

nct

dark

er o

utlin

e on

war

ts an

d fo

ldin

gs

Larg

e da

rk

blac

kish

spot

s W

hite

-

Boul

enge

r 189

3; th

is pa

per

Lept

olal

ax p

luvi

alis

Gre

yish

bro

wn

with

dar

k pa

ttern

Fe

w b

lack

spot

s Br

own

spot

s D

ark

gold

abo

ve

Ohl

er e

t al.

2000

Lept

olal

ax su

ngi

Uni

form

or w

ith li

ght s

pots

Smal

l dar

k sp

ots

Imm

acul

ate

Iride

scen

t gol

d-gr

een

Lath

rop

et a

l. 19

98

Lept

olal

ax ta

mdi

l M

id-g

rey

with

irre

gula

r da

rk g

rey

blot

ches

Bl

acki

sh-g

rey

blot

ches

Cr

eam

U

pper

third

of i

ris b

right

or

ange

, res

t gre

yish

-cre

am

Seng

upta

et a

l. 20

10

Lept

olal

ax tu

bero

sus

Dar

k gr

ey o

r bla

ck w

ith

obsc

ure

light

spec

klin

g Li

ght s

peck

lings

Th

in b

lack

retic

ulat

ion

encl

osin

g ro

und,

whi

te sp

ots

- In

ger e

t al.

1999

Lept

olal

ax v

entri

punc

tatu

s G

rey

brow

n w

ith sp

ots

Rath

er la

rge

spot

sD

istin

ct d

ark

spot

s on

ches

t an

d be

lly

Copp

er a

bove

, gre

y br

own

belo

w

Feie

t al.

1992

; thi

s pa

per



FIGURE 1. Morphological character states observed in Leptolalax. Tympanum (a): on top L. sungi, ROM 28474, holophoront:tympanum indistinct, below L. eos, MNHN 2004.0274, holophoront: tympanum distinct; feet showing extension of webbingand fringes along toes and presence of subdigital ridges (b): left L. eos, MNHN 2004.0274, holophoront: small webbing, dis-tinct fringes on toes, right L. minimus form 2, THNHM 07417: rudimentary webbing, fringes on toes absent; lateroventral glan-dular ridge (c): on top L. nyx, AMNH A163810, holophoront: poorly distinct isolated glands aligned in a row, below L. minimusform 2, THNHM 07417: distinct glands forming interrupted line; femoral glands (d): on top L. minimus, MNHN 2006.2553:small femoral glands, below L. eos, MNHN 2004.0274, holophoront: large femoral glands.



FIGURE 2. Morphological character states observed in Leptolalax. Dorsal skin structures. Left, L. pelodytoides, MSNG29845.A, lectophoront, smooth skin on dorsum with few flat tubercles and presence of glandular folds (arrows). Right, L. nyx,AMNH A163810, holophoront, with rounded granular tubercles and absence of glandular folds.

Lateroventral glandular ridge: This ridge is a macrogland of linear shape present on the side of belly. It showsvariation in aspect, formed sometimes by a continuous ridge, sometimes by a series of rounded glands. For Lepto-lalax species, this macrogland has been described in many disparate ways. Either its simple presence (Rowley et al.2010c) or absence (Fei & Ye 1992, Inger et al. 1999, Matsui et al. 2009, Rowley et al. 2010a,b) is mentioned, orvarious details on completeness (Matsui 2006, Humtsoe et al. 2008), position, extension, colour and shape (Taylor1962, Das et al. 2010, Sengupta et al. 2010) of line are indicated to describe the line. In this work variation in pres-ence or absence and shape, and continuity of this ridge is given (Fig. 1c).

Axillary glands: In Leptolalax, a pair of glands is usually present ventrally on the sides of chest at the level offorelimbs. These glands are oval or round in outline, always flat so that they are poorly distinct, and in some indi-viduals indistinct. We prefer this term to the term “absent”. It is very difficult to confirm absence of a glandularstructure in amphibian skin. Many glands are embedded in the skin and macroglands can be recognized by theaccumulation of glands in certain region of the skin. When very small they cannot be distinguished from otherglands. Another point is modifications due to preservation that might flatten glands and render them invisible byexternal observation (Rowley et al. 2010b). Absence means that it has not been observed. Inger (1954: 207) createdthe term axillary glands for this structure. Matsui et al. (2009) and Das et al. (2010) called them “pectoral glands”but Dubois (1980) and Delorme (1999) considered the glands, present either in axillary or pectoral region in mego-phryid frogs, to be homologous as either but never both structures together occur, thus different shape or position ofthe glands are different states of the same character.

Suprabrachial gland: In Leptolalax, a gland is present in the continuity of the supratympanic fold, just dorsalto the forelimb insertion. This gland is present in all species of Leptolalax studied as a prominent gland, usually oflight colour, thus clearly visible. In some specimens, the gland is brownish and covered by dark spots. Das et al.(2010) called these glands preaxillary glands; Delorme (1999) use the word “rictal glands”. Both terms are not fullycorrect to describe these glands as they are not anterior to axilla nor in the continuation of the mouth opening orstructures linked to the mouth. The presence and position (Liu 1950, Taylor 1962), relative size compared to differ-ent species (Dubois 1987) or size, shape and height (Rowley et al. 2010a,b,c) of this gland are here described.



Size of femoral gland: Femoral glands are macroglands present on the ventral side of thigh. The gland isalways single and oval or rounded in Leptolalax, and occurs in males as well as in females. Boulenger (1900)described it as “a round whitish spot on the back of each thigh” but already Liu (1950) and Taylor (1962) recog-nized this spot as gland. Rowley et al. (2010a,b,c) described size and position of the gland on shanks. The size ofthis gland shows interspecific variation (Fig. 1d).

Colour pattern of adult Leptolalax

The dorsum, head and ventral surfaces show different colorations and colour patterns. Coloration due to yellow andred pigments is modified in fixed specimens and after a short period of storage, but pattern, mainly due to presenceof melanophores, is rather persistent. Only colour pattern and presence of pigmentation were used for analysis.These patterns are defined by presence or absence of various spots, linkages between such spots and size of dots.Figure 3 represents schematic view of dorsal and lateral pattern that can be observed in Leptolalax species and thepotential linkages between the various spots. The following 22 characters were observed on up to 10 specimens perspecies. There was no sexual dimorphism for coloration as male and females from the same population show simi-lar patterns. Thus male and females were analysed together:

(1) Spot on snout (present/absent); (2) triangle between eyes (present/absent); (3) Spot on shoulder region(present/absent; shape of spot); (4) spot on upper part of iliac region (present/absent; single/pair); (5) triangle onlower part of iliac region (present/absent); (6) linkage between snout spot and triangle (present/absent); (7) linkagebetween triangle and shoulder spot (present/absent); (8) linkage between shoulder and upper iliac spot (present/absent); (9) linkage between iliac spots (present/absent); (10) outline of dark dorsal pattern (present/absent; narrowlines/broadly extended); (11) brown spots around dorsal warts (present/absent); (12) whitish warts on dorsum(present/absent); (13) dorsal background pattern (uniform/marbling); (14) blackish spots on flanks (present/absent); (15) blackish spot posterior to nostril (present/absent); (16) upper half of tympanum (same colour aslower/darker); (17) throat (presence of pigmentation/whitish); (18) border of throat (presence of pigmentation/whitish); (19) chest (presence of pigmentation/whitish); (20) belly (presence of pigmentation/whitish); (21) side ofbelly (presence of pigmentation/whitish); (22) ventral side of shanks (presence of pigmentation/whitish).

Character 18 did not show significant interspecific variation and was therefore not informative and notincluded in analysis. The other characters were coded and used in discriminant analysis.

Tadpole sampling and morphology

Tadpoles were collected in the field from streams surveyed and were fixed as quickly as possible in an aqueoussolution composed of 50 % ethanol (in 70 % concentration) and 50 % formaldehyde (in 4 % solution). A piece ofcaudal muscle or the tip of the tail of one tadpole of each series (e.g., of each collected in one stream) was removedand conserved in 99 % alcohol for molecular analysis purposes. Tadpoles of L. aereus, L. minimus and L.ventripunctatus were determined by mean of DNA barcoding whereas those of L. bourreti were determinedthrough comparison of a froglet (MNHN 1998.8769) from developmental series, with syntopic adults. Becausecertain species are found in sympatry, a series from a stream can theoretically contain specimens of several species.For this reason, the descriptions are based only on DNA voucher tadpole specimens identified by the DNAbarcoding method (except for L. bourreti). Specimens of Leptolalax bourreti, L. minimus and L. ventripunctatuswere deposited in the herpetological collection of the MNHN. Larval specimens of Leptolalax aereus weredeposited in the collection of the ZFMK.

Morphological terminology follows Altig and McDiarmid (1999a), keratodont row formula is given accordingto Dubois (1995), terminology of lateral line system follows Lannoo (1987) and developmental stages were deter-mined according to Gosner (1960). All measurements were taken with a graduated ocular attached to a stereomi-croscope to the nearest 0.01 mm except TOL and some SU values (see abbreviations below) which were taken witha digital vernier calliper to the nearest 0.1 mm. The landmarks are those shown in Altig and McDiarmid (1999a, p.26: Fig. 3. 1) and Grosjean (2006). Drawings were made with the aid of a camera lucida.



FIGURE 3. Scheme of dorsal patterns potentially present in Leptolalax specimens observed in this work. 1—spot on snout;2—triangle between eyes; 3—spots on shoulder region; 4—large blackish spots on flanks; 5—spots on upper part of iliacregion; 6—triangle on lower part of iliac region; 7—small blackish spots on flanks. The various linkages that can exist betweenthese spots are represented by chain dotted outlines.

The abbreviations used in the descriptions are the following: BH maximum body height; BL body length; BWmaximum body width; ED maximum diameter of eye; KRF keratodont row formula; LF maximum height of lowertail fin; MTH maximum tail height; NN internarial distance; NP nario-pupilar distance; ODW oral disc width; PPinterpupilar distance; RN rostro-narial distance; SS distance from tip of snout to opening of spiracle; SU distancefrom snout to beginning of upper tail fin; TAL tail length; TOL total length; TMH maximum tail muscle height;TMW maximum tail muscle width; UF maximum height of upper tail fin.

All the tadpoles of Leptolalax described here are morphologically very similar. A detailed description is there-fore given only for one species, Leptolalax bourreti, and only the differences respective to this reference speciesare given for the other species.

Terminology

Technical nomenclature terms used here follow Dubois (2000, 2005, 2010). The rationale behind this new termi-nology is to avoid the term “type” which originated from and reminds of a typological concept of species. Wethereby wish to express that those specimens are not “typical” but are name bearers in their function. In thoseinstances where the mentioning of old terms is mandatory by the Code (e.g., when defining the holophoront = holo-type) we also mention the old term in parentheses.



Exonymophoronts. Designation of a specimen as lectophoront among symphoronts results in all other originalsymphoronts becoming exonymophoronts (paralectotypes in the Code), which are just a particular category of pro-taxonts.

Holophoront. Holotype, single specimen being name-bearing type.Hypodigm. Term created by Simpson in 1940 for a sample of specimens from which the characters of a popu-

lation are to be inferred. Includes what is called paratypes in the Code, as well as other specimens included by anauthor. Paratypes have no nomenclatural status as onomatophore but inform us of the content an author gives to ataxon. All members of the original hypodigm of a taxon can be called protaxonts (Dubois 2010).

Lectophoront. Lectotype, name-bearing type specimen chosen among symphoronts.Neophoront. Neotype, name-bearing type specimen designated to replace a lost holophoront, lectophoront or

symphoronts.Nomen. Scientific name as recognized by the International Code of Zoological Nomenclature.Onomatophore. Name-bearing type, which is a specimen (onymophoront) for taxa of rank species and subspe-

cies, or a nominal taxon for higher taxa.Onymophoront. Name-bearing type specimen (holophoront, symphoront, lectophoront or neophoront).Onymotope. Place of collection of the onomatophore which is called type-locality in the International Code of

Zoological Nomenclature. Onymotopic. Specimen coming from the same place as the onomatophore.Protaxont. Any specimen member of the original hypodigm of a new taxon. Symphoronts. Syntypes, two or more specimens being together name-bearing types.

Similar to what has been done in other extensive and collaborative works (e.g., Frost et al. 2006), we wish to men-tion here that not all authors of the present work had concordant opinions on all details of methods and terminologyto be used. As a team we accepted to use the terminology of Dubois (2000, 2005) and the description scheme ofOhler (1996), but this does not imply that all authors explicitly recommend using these schemes or will use them intheir own future publications. Similarly, the last author (AD) does not agree with the use of “threshold values” inDNA divergence for species recognition (see Dubois 1977b, 1998).

Results

Molecular analysis

Bayesian and ML analyses found the genus Leptolalax to be monophyletic with high statistical support values (Fig-ure 4). L. arayai from Malaysia and L. heteropus from Phang Nga, Thailand were sister taxa. These two specieswere the only species in our analysis that Dubois et al. (2010) had allocated to the subgenus Leptolalax. In ouranalysis this clade was sister to a clade comprising all remaining lineages, including several nominal species thatDubois et al. (2010) grouped in the subgenus Lalos.

While the current manuscript was in preparation and revision, several additional species of Leptolalax weredescribed, among them L. aereus, L. applebyi, L. bidoupensis, L. croceus and L. melicus. DNA sequences of thesespecies were only partly available from Genbank but were kindly made available by J. Rowley for comparativepurposes. Since these sequences will be published in a forthcoming phylogenetic analysis by J. Rowley, werefrained to include them in our phylogenetic tree but only run a preliminary analysis to understand if any of themmight be similar to the populations studied by us. We found L. applebyi, L. bidoupensis, L. croceus and L. melicusclustering outside the Lalos clade but L. aereus within the Lalos clade (see below). The species outside the Lalosclade did not form a well-supported monophyletic group; the monophyly of the subgenus Leptolalax sensu Dubois(2010) is therefore in need of confirmation by further analysis with more comprehensive sets of molecular markers.In the following we will analyse more in detail the relationships and differentiation among sequences within theLalos clade as shown in Fig. 4.



FIGURE 4. Combined 16SrRNA Bayesian phylogeny and haplotype networks of 113 Leptolalax specimens with hierarchicaloutgroups (schematic representation in the grey box) The tree topology corresponds to the Bayesian topology obtained withthree million generations, values on nodes indicate posterior probabilities (asterisks > 95 %, double asterisks > 99 %), andMaximum Likelihood bootstrap support values (> 70 %). Red numbers are the percentage of uncorrected p - sequence diver-gence between selected sister clades. Minimum spanning haplotype networks for two Leptolalax-clades (connected to phyloge-netic tree nodes marked grey) were obtained with TCS. Numbers on haplotype connections represent number of nucleotidesubstitutions. Numbers of specimens included in haplotype networks are: B5-1(12); B5-2(17); B5-3(6); B5-4(1); B4-1(8); B4-2,3,4,7,8(1); B4-5(2); B4-6(3). Sampling localities are provided as pie charts within haplotype circles, circle size correspondsto the number of sampled individuals. Abbreviations for country of origin are as follows: L—Laos, M—Malaysia, T—Thai-land, V—Vietnam. Newly described species and haplotype networks are coloured.



Within Lalos, a highly supported clade, here named clade A (Fig. 4) was recovered consisting of specimens apriori allocated to L. liui (China; Zheng et al. 2008; a single sequence with GenBank accession numberEF544238), two specimens ascribed to L. oshanensis (China, Zheng et al. 2004; GenBank accession numbersAY561306 and AY526215) which did not form a monophyletic group, a tadpole from Doi Chiang Dao (Thailand),two specimens from the onymotope of L. bourreti (Sa Pa, Vietnam) with identical sequences, and a subclade ofseveral specimens collected by us in Long Nai and Phongsaly (Laos) which we propose as a new species, L. eos(described below). The uncorrected p-distance between L. oshanensis (AY526215) and L. eos was 4.6 %, and thusabove the 3 % sequence divergence in 16S rRNA suggested as threshold to identify candidate species (Fouquet etal. 2007, Vieites et al. 2009); due to the congruence with morphological differences (see below) we concluded thatL. eos is a distinct species.

Sister to clade A was a highly supported clade B containing all other representatives of Lalos, with specimensoriginating from Vietnam, Laos and Thailand. The main relationships within clade B are not well supported in ouranalysis (Fig. 4). There are five well supported subclades contained in this clade, named B1-5 in the following:

(B1) L. pluvialis from Sa Pa, Vietnam, with two sequences forming a clade. (B2) A sequence obtained from GenBank (DQ283381, Frost et al. 2006) that was ascribed to L. bourreti from

Ha Giang Province (Vietnam). The uncorrected p-distance between this specimen and L. bourreti (MNHN1999.5659–5660) is ~10.9 %, to L. ventripunctatus ~5.6 %, and to L. minimus ~5.1 %. Morphological differencesto L. bourreti (see below) and to other species allocated to Leptolalax furthermore warrant recognition of a newspecies for this specimen, described below as L. nyx.

(B3) Specimens ascribed on the basis of morphological and coloration characters to L. ventripunctatus col-lected from Phongsaly Province (Laos) showed very little intra-clade genetic differentiation, and formed a mono-phyletic group with two specimens ascribed to L. pelodytoides (Tam Dao, Vietnam, Garcia-Paris et al. 2003,AY236797 and AY236798). Uncorrected p-distance between these two taxa was 1.4 %. Because specimens of Lep-tolalax usually are allocated to L. pelodytoides, the allocation of the specimens from Tam Dao to this species maybe subject of discussion. We propose here to allocate them to L. ventripunctatus, in favour over the opposite sce-nario. Vice versa, that means that our analysis does not include a sequence of the species L. pelodytoides.

(B4) A well-supported clade consisting of specimens collected from Vietnam showed high intra-clade diver-gences (two subclades separated by an uncorrected p-distance of 2.4 %). The minimum-spanning network (Fig. 4)revealed that three sampled localities in Vietnam contained eight different haplotypes, each of the haplotypes wereunique to a site except B4-1 and up to five haplotypes occurred at one site. The largest distance between sites (UBo, Quang Binh Province to Ky Anh – Ke Go Nature Reserve, Ha Tinh Province) was 70 km, which was mirroredby in total 18 nucleotide site differences between the haplotypes present at these sites. The uncorrected p-distanceto L. ventripunctatus and to clade B5 (see below) was ~6.4 % and 6.2 %. Preliminary comparison with an unpub-lished DNA sequence of the recently described L. aereus provided by J. Rowley showed that this sequence clus-tered within clade B4 and represents yet another genealogical lineage within this clade. We cannot rule out that B4might include more than one species, but conservatively assign all species within this clade to L. aereus based onthis sequence similarity plus a high similarity of morphological characters of all included specimens, including thetype series of L. aereus according to the original description (Rowley et al. 2010c).

(B5) The remaining Leptolalax specimens from disjunct localities in Laos and Thailand formed a further clade,again with a split into two well-supported subclades. The minimum-spanning network shows that although thegenetic divergences between the two subclades are higher than between the subclades of the species in Vietnam(uncorrected p-distance 3.46 %), both lineages are sympatric at several sites: of the four different haplotypes intotal (B5-1 and B5-2 of subclade 1 and B5-3 and B5-4 of subclade 2), all four occur in Chiang Mai Province (Thai-land), with B5-3 and B5-4 only occurring in this region. Haplotypes B5-1 and B5-2 also occur at several sites inLaos, which are geographically proximate to each other but not to Doi Chiang Dao (distance of 340 km). The cladecontaining haplotypes B5-1 and B5-2 can be allocated to the species L. minimus on the basis of morphologicalcharacters (see below). The relatively deep genetic divergences of the two subclades but weak morphological dif-ferentiation (see below) do not give enough evidence that the clade containing haplotypes B5-3 and B5-4 from DoiChiang Dao (Thailand) constitute a new species.



Evidence from morphology and colour pattern

Leptolalax species show variation in a series of morphological characters (Table 4–5), albeit the differences some-times are weak. We will discuss this insofar as it provides a crucial confirmation that the mitochondrial lineagesidentified by molecular analysis in fact are evolutionary independent lineages and therefore merit status as separatespecies. Furthermore, these characters are obviously of high importance to separate the new lineages from speciespreviously recognized by morphology only, such as L. pelodytoides.

Most clades obtained by our molecular study had been allocated to species through morphological charactersprior to our analysis. The differences among species are weak, i.e., the character states defined do not show muchdifferentiation. For example, the webbing in Leptolalax is always partial, but can be reduced to a border of skinlinking the bases of two toes. In the few species in which the web is well developed for the genus, it always leavesat least three phalanges of the fourth toe free of web. The character definitions are similarly difficult concerningmost of the other morphological characters studied, such as finger tip shape, extension of fringes on toes or struc-ture of dorsal skin. Nevertheless, the combination of these characters allows differentiating most of the groups sup-ported by molecular evidence.

Colour patterns and coloration constitute a series of characters that are complex to use in anurans. Discriminantanalysis using 22 characters of the colour pattern allows recognizing several groups that agree with clades obtainedby molecular analysis (Fig. 5; table 6). Among the well-supported clades are specimens of L. eos which show nodark spots on the lateral body and specimens of L. ventripunctatus and L. pluvialis which both have dark spots onthe belly.

Delorme et al. (2006) proposed that the presence of a lateroventral glandular ridge is an apomorphy for thesubgenus Lalos. In an early description (Anderson 1871) it was mentioned as a white line, used as a basis for theepithet lateralis. A lateroventral glandular ridge is present at least as a series of small glands on the sides of thebelly in all lineages of Leptolalax identified in the present study except L. heteropus and L. arayai. This glandularridge is not always distinct. In some specimens it is a continuous ridge, in others a series of small glandular wartsforming a more or less interrupted line on the side of the belly (Fig. 1c). In some species, this glandular ridge is ren-dered more visible by the dark coloration of the flank and lateral belly, whereas in specimens with a dense coveringof warts on the flanks the warts forming the glandular ridge do not stand out.

Two species, L. bourreti and L. eos, have been considered as potentially conspecific because they share largebody size, but their extension of webbing and dorsal skin structure give support to their taxonomic distinctness. L.eos shows a rather uniform dorsal pattern and no black spots on the side of the body, whereas L. bourreti has dis-tinct dorsal patches and a series of black spots on the side of the body. Among the clades recognized by moleculardata, two species can be diagnosed by their small adult body length. L. pluvialis (SVL 22–23 mm) is the smallestspecies of Leptolalax included in this study and is sister group to a clade formed by L. aereus, L. minimus, L. ven-tripunctatus and L. nyx (Fig. 4). The clade formed by L. aereus and L. minimus shows poor morphological differen-tiation. Webbing is more developed in L. minimus and fingers slightly swollen in L. aereus. These species alsoshow some differentiation in coloration. The dorsal colour of L. aereus as shown in life photographs has a greyishaspect, whereas the other species have brown dorsal colour. Only in L. aereus indistinct brown vermiculations arepresent on the belly and white spots on the side of the body. Iris colour is dark gold in typical form of L. minimus,orange in form 2 of this species and copper gold in L. aereus. Finally, L. pelodytoides from Myanmar as repre-sented by at least 4 specimens collected by Fea, a species for which no recent samples are available, is character-ized by a complex dorsal pattern that forms a unique spot on the snout, head and shoulders, by having extensivewell-developed webbing on feet and by a relatively large body size.



TABLE 6 Results of canonical discriminant analysis including data of nine OTU concerning colour pattern and morpho-logical characters. A total of 59 specimens have been included into the analysis (numbers for species given in brackets):Leptolalax bourreti: protaxont (1); onymotopic specimens (5); L. eos: holophoront (1); protaxonts (6); L. liui (1); L. min-imus (10); L. minimus form 2 (10); L. nyx (1); L. pelodytoides: holophoront (1); L. pluvialis (3); L. aereus (10); L. ven-tripunctatus (10).

A. Description of Variance

B. Statistical significance

C. Standardized canonical discriminant function coefficients

Function Eigenvalue % of Variance Cumulative % Canonical Correlation1 13.289 30.4 30.4 0.9642 8.268 18.9 49.4 0.9453 6.330 14.5 63.9 0.9294 5.508 12.6 76.5 0.9205 3.537 8.1 84.6 0.8836 2.888 6.6 91.2 0.8627 2.380 5.5 96.7 0.8398 1.038 2.4 99.0 0.7149 0.419 1.0 100.0 0.543

Test of Function(s) Wilks’ Lambda Chi-square df Significance1 through 9 0.000 556.025 198 0.0002 through 9 0.000 449.646 168 0.0003 through 9 0.000 360.583 140 0.0004 through 9 0.001 280.905 114 0.0005 through 9 0.006 205.981 90 0.0006 through 9 0.026 145.494 68 0.0007 through 9 0.102 91.176 48 0.0008 through 9 0.346 42.464 30 0.065

9 0.705 13.995 14 0.450

Function1 2 3 4 5 6 7 8 9

Triangle between the eyes 0.317 0.034 0.455 -0.109 0.237 0.021 0.021 -0.427 -0.298Butterfly on the shoulder region 0.090 0.754 -0.594 0.323 0.620 -0.265 0.263 -0.045 0.384Spot on upper part of iliac region 0.633 -0.657 0.270 0.446 -0.252 0.060 0.156 -0.521 -0.203Triangle on lower part of iliac region -0.616 0.109 -0.457 -0.047 -0.133 -0.128 -0.339 0.471 -0.269Spot on snout 0.038 0.256 0.139 0.616 0.140 0.107 0.095 -0.050 -0.230Triangle_snout spot -0.056 -0.450 0.236 -0.345 0.489 -0.338 -0.667 0.239 0.158Triangle_butterfly -0.467 -0.080 -0.098 0.232 0.659 -0.103 0.113 0.157 0.015Outline of dark dorsal pattern -0.013 -0.726 0.398 -0.202 0.003 -0.350 0.406 0.508 -0.130Spots on flanks 0.544 -0.683 -0.180 -0.602 0.007 0.141 0.389 -0.022 0.055Brown spots / dorsal warts -0.255 0.632 0.008 0.167 -0.251 0.347 0.511 0.443 -0.068Warts whitish -0.193 -0.876 0.530 0.699 -0.138 0.107 0.123 -0.041 -0.092Marbling -0.013 0.356 0.073 -0.844 0.214 0.033 0.004 0.006 -0.268Throat 0.974 -0.009 -0.231 0.261 -0.302 0.075 -0.423 0.346 -0.179Chest 0.813 -0.137 -0.147 0.255 -0.550 -0.259 -0.180 0.195 -0.210Belly 0.120 0.159 0.202 -0.167 0.577 0.357 0.077 -0.018 0.288Side of belly 0.085 -0.022 -0.070 0.197 -0.491 -0.783 0.326 0.050 0.067Shanks -0.320 0.400 0.588 -0.312 0.356 -0.338 0.344 0.124 0.593Warts -0.342 0.287 0.107 -0.156 0.113 0.732 -0.018 -0.278 0.239Ridges -0.252 0.452 -0.080 -0.162 0.567 0.487 0.348 -0.044 -0.155Femoral glands size 0.594 0.377 0.203 0.399 -0.102 0.352 -0.007 0.159 -0.247Size of finger tips -0.068 -0.324 0.176 0.657 0.061 0.087 -0.059 -0.022 0.661Size of toe tips -0.455 0.337 -0.243 -0.347 0.141 0.217 0.300 0.280 0.020



FIGURE 5. Plot of functions one to four of discriminant analysis based on 22 colour pattern characters including specimens ofsubgenus Lalos. A total of 59 specimens have been included into the analysis: Leptolalax bourreti: protaxont (1); onymotopicspecimens (5); L. eos: holophoront (1); protaxonts (6); L. liui (1); L. minimus (10); L. minimus form 2 (10); L. nyx (1); L. pelo-dytoides: lectophoront (1); L. pluvialis (3); L. aereus (10); L. ventripunctatus (10).

Morphometric analysis

Morphometric analysis was applied to obtain further evidence for the distinctiveness of the lineages identified bymolecular studies and to find characters that allow the assignment of onymophoronts to the lineages identified byDNA.

Log transformed variables for all adult male specimens were included in the principal component analysiswhich leads to five principal components with a value higher than 1 (Table 7, Fig. 6). These components accountfor 74.1 % of overall variance. ANOVA analysis of these principal components shows significant variation accord-ing to the groups recognized by molecular studies for the first three variables (Table 8). Post hoc tests (whichinclude only groups of more than one specimen) show significant differences for at least one principal componentfor all group pairs but any of the combinations between the following species: L. aereus and L. minimus. Thesegroups are considered morphological siblings as they cannot be distinguished morphometrically.



TABLE 7. Principal component analyses using 24 log-transformed variables and their factor loadings after varimax-rotation.

TABLE 8. Results of ANOVA for the first five principal components (PC) of the standardized morphometric variables.

Component Extraction Sums of Squared Loadings Total % of Variance Cumulative %1 10.717 44.656 44.6562 2.999 12.494 57.1503 1.705 7.102 64.2534 1.326 5.523 69.7765 1.043 4.346 74.122

Component1 2 3 4 5

lsvl 0.794 0.344 -0.167 -0.205 -0.220lhw 0.813 -0.047 -0.288 0.058 -0.025lhl 0.676 0.274 -0.295 0.023 0.057lmn 0.759 0.442 -0.150 -0.040 -0.246lmfe 0.351 0.560 0.538 0.216 0.137lmbe 0.567 0.526 0.360 -0.007 0.134life 0.461 0.402 0.401 0.339 0.275libe 0.399 0.025 -0.118 0.707 -0.222lin 0.772 0.026 -0.298 0.195 0.268len 0.605 0.579 -0.247 -0.128 -0.000ltyd 0.439 0.346 -0.465 0.061 0.416ltye 0.598 0.337 -0.240 -0.412 -0.027lfll 0.383 0.322 0.105 0.283 -0.500lhal 0.765 -0.245 0.294 -0.225 -0.003ltfl 0.788 -0.204 0.139 -0.046 -0.116ltl 0.680 -0.349 0.103 0.134 0.115lfol 0.713 -0.523 -0.027 0.102 0.011lftl 0.729 -0.439 0.018 0.167 -0.282limt 0.649 -0.024 0.161 -0.364 -0.261litl 0.898 -0.190 0.068 0.001 -0.058lmttf 0.723 -0.187 0.382 -0.198 0.145lmtff 0.698 -0.088 0.317 -0.146 0.155ltftf 0.693 -0.451 -0.225 0.072 0.160lfftf 0.698 -0.457 -0.099 0.017 0.159

clade16S errorSS df F Sig. SS df

PC 1 16.816 6 11.725 0.000 10.518 44PC 2 15.038 6 4.790 0.001 23.024 44PC 3 20.361 6 6.284 0.000 23.761 44PC 4 7.254 6 1.626 0.163 32.713 44PC 5 4.858 6 0.775 0.594 45.943 44



FIGURE 6. Plot of first three principal components of 25 morphometric log standardized variables of Leptolalax specimens.Each symbol represents a clade as recognized in phylogenetic analysis of molecular data. A total of 60 specimens wereincluded into the analysis: L. eos (6); L. minimus (21); L. minimus form 2 (5); L. nyx (1); L. pluvialis (3); L. aereus (2); L. ven-tripunctatus (22).

Systematics

Genus Leptolalax Dubois, 1980

Subgenus Lalos Dubois, Grosjean, Ohler, Adler & Zhao, 2010

The characters listed below are those shared by all Leptolalax (Lalos) species studied here. They refer to thedescription scheme of Ohler (1996) and are the complement of the characters given for each species. We list themin order not to repeat them for every single species. Characters listed in species descriptions allow to distinguishthem from the other species studied.

Subgeneric diagnosis. (3) Snout rounded. (6) Nostrils without skin flap. (7) Pupil oval, vertical. (9) Pineal ocellusabsent. (10) Vomerine ridge absent. (11) Tongue emarginate; median lingual process absent. (14) Relative length offingers, shortest to longest: I < II < IV < III or I < II = IV < III. (15) Tips of fingers rounded. (16) Fingers withoutwebbing. (17) Subarticular tubercles absent. (18) Prepollex absent; two distinct palmar tubercles, median rounded,outer oval; supernumerary tubercles absent. (20) Toes long and thin. (21) Relative length of toes, shortest to lon-gest: I < II < V < III < IV. (22) Tips of toes rounded. (25) Subarticular tubercles below toes absent. (27) Tarsal foldabsent. (28) Outer metatarsal tubercle, supernumerary tubercles, tarsal tubercle absent. (33) A pair of often poorlydistinct whitish axillary glands present; a pair of distinct femoral and suprabrachial glands present; a pair of lat-eroventral glandular ridges, at least as a short alignement of individual glandular warts, but usually as a distinct lineof glandular warts extending from axilla to near groin, present.

Male secondary sexual characters. (37) Nuptial spines absent. (38) Vocal sac openings present. (39) No othermale secondary sexual characters.

Comments. For nomenclatural issues, we describe first the morphology of Leptolalax pelodytoides, for whichno molecular data are available, but the nomen of which has been largely applied to specimens of this group. Theother species will be presented in phylogenetic order as proposed by the phylogram in figure 4. The two species of



the subgenus Leptolalax, L. heteropus and L. arayai, included in the molecular phylogeny, are not considered fur-ther. For comparison, Tables 3–5 give morphometrical, morphological and coloration characters of species allo-cated to Lalos.

In the original description of L. sungi (Lathrop et al. 1998), the authors stated that the suprabrachial and lat-eroventral glands are absent, but examination of the hypodigm and specimens from IEBR showed that these glandsare present but relatively much smaller and finer than in other species of the subgenus.

Leptolalax (Lalos) pelodytoides (Boulenger, 1893)(Fig. 7–8)

Leptobrachium pelodytoides Boulenger, 1893: 345, pl. 11. Onymophoront: Lectophoront (lectotype) by subsequent designation of Capocaccia (1957: 212), MSNG 29845.A, adult

female. Onymotope: Thao [Thamo] (17°08’ N, 98°01’ E), Kayah State, Myanmar. Collected by Leonardo Fea, 1885.

Material examined. Myanmar: Thao, Karin Hills: BMNH 1947.2.25.14, exonymophoront, adult male; MSNG29845.A, lectophoront, adult female, coll. Leonardo Fea, 1885; — Karin Bia-po (800–1000 m): MSNG 29845.B,adult female, exonymophoront, coll. Leonardo Fea, 1885; — Carin Ghecú [Caim Gheena, near Thao] (1300–1400m): MNHN 1893.519, adult male; ZMB 11588, adult male, coll. Leonardo Fea, 1885; ZMH A02413, adult male,coll. Leonardo Fea, 1885.

Comment. Boulenger (1893) described the species based on three specimens: two from Thao and one fromKarin Bia-po (near Leiktho: 19°13’ N, 96°34’ E; 800–1000 m; Kayin State), Myanmar. He presented a figure ofspecimen BMNH 1947.2.25.14, adult male, which is now an exonymophoront. Capocaccia (1957: 212) designatedthe female specimen MSNG 29845.A from Thao as lectophoront of this nominal taxon. The two specimens showsome difference in colour pattern and morphology. The lectophoront designation of Capocaccia created some taxo-nomic problems as she did not choose the figured specimen as lectophoront as recommended by the InternationalCode of Zoological Nomenclature: most taxonomists only have access to this figure and the description of Bou-lenger. But the species is now defined by the lectotype, and the following description of the lectophoront and thefigure of the specimen will make the characters of the species available to the community of taxonomists.

The first described species nomen available for the genus Leptolalax, Ixalus lateralis Anderson, 1871, is allo-cated to a species which is known from its onymotope in Nagaland (India) only (Humtsoe et al. 2008), and whichis different from all other known species of the genus. In particular, it is distinct from Leptobrachium pelodytoidesBoulenger, 1893 as defined by the hypodigm by several consistent differences, including a smaller head in L. later-alis than in L. pelodytoides. These species can also be distinguished by the webbing, which is quite distinct in L.pelodytoides but rudimentary in L. lateralis (Humtsoe et al. 2008).

L. pelodytoides was described from Myanmar and no onymotopic material has been collected recently as theonymotope is in a part of the country which has been closed to scientists for many years. Thus only study of theoriginal symphoronts can provide information on the identity of this species.

The name pelodytoides has been used for many populations of Leptolalax from China, Vietnam, Laos, Thai-land and Myanmar. Our molecular data show that several clades can be recognized within the populations calledpelodytoides. Morphological analysis (tables 3−5) and analysis of colour pattern (see below) give evidence that thenomen Leptobrachium pelodytoides should be applied to specimens from the Karin Hills only. Thus all other spec-imens mentioned as L. pelodytoides in the literature should be allocated to other taxa.

Diagnosis. Leptolalax pelodytoides is a member of the subgenus Lalos based on the presence of a lateroventralgland and distinguished from other species included in this subgenus by the following combination of characters:large-sized species (males 27.5–32.3 mm; females 35.5–37.8 mm) (Table 3) with small webbing on feet and narrowfringes on toes; distinct tympanum; dermal ridges under toes poorly distinct; finger tips slightly dilated; dorsumwith glandular warts and short elongate ridges (Table 4); dorsal pattern distinct including dark outlines on wartsand foldings; large dark spots on flanks present; ventral side whitish; iris colour in life not known (Table 5).

Description of lectophoront. Size and general aspect. (1) Specimen of moderate size (SVL 37.8 mm), bodyrather slender.



Head. (2) Head of medium size, narrower (HW 12.2 mm) than long (HL 13.0 mm; MN 11.3 mm; MFE 8.5mm; MBE 4.5 mm), flat. (3) Snout slightly protruding, its length (SL 5.7 mm), longer than horizontal diameter ofeye (EL 4.3 mm). (4) Canthus rostralis distinct, loreal region concave, vertical. (5) Interorbital space flat, about aslarge (IUE 3.4 mm) as upper eyelid (UEW 3.5 mm) and internarial distance (IN 3.2 mm); distance between front ofeyes (IFE 5.7 mm) more than half of distance between back of eyes (IBE 9.7 mm). (6) Nostrils oval, about as closeto tip of snout (NS 2.6 mm) as to eye (EN 2.5 mm). (8) Tympanum (TYD 2.1 mm) distinct, rounded; about equal tohalf eye diameter, tympanum-eye distance (TYE 1.6 mm) three-fourths of its diameter. (11) Tongue not observed.

FIGURE 7. Leptolalax pelodytoides, MSNG 29845.A, lectophoront, adult female, SVL 37.8 mm. (A) Dorsal and (B) ventralview.

Forelimbs. (12) Forearm rather long, thin (FLL 9.0 mm), about length of hand (HAL 9.1 mm), not enlarged.(13) Fingers rather long and thin (TFL 4.7 mm). (15) Tips of fingers slightly enlarged. (16) Fingers without dermalfringe.

Hindlimbs. (19) Shanks about four times longer (TL 16.4 mm) than wide (TW 3.8 mm), about as long as thigh(FL 16.9 mm) and distance from base of internal metatarsal tubercle to tip of toe IV (FOL 16.2 mm). (20) Toe IV(FTL 7.8 mm) about one-third of distance from base of tarsus to tip of toe IV (TFOL 23.6 mm). (22) Tips of toesnot enlarged. (23) Webbing small: I 2 – 2 ½ I 1 ½ – 3 III 2 ½ − 3 ¾ IV 4 – 3 V (MTTF 6.5 mm; MTFF 6.2 mm;TFTF 8.8 mm; FFTF 9.9 mm). (24) Narrow fringe along toes present. (26) Inner metatarsal tubercle short, distinct;its length (IMT 1.5 mm) 2.3 times in length of toe I (ITL 3.5 mm).

Skin. (29) Dorsal and lateral parts of head and body: snout and region between the eyes with small glandularwarts, side of head smooth with horny spinules; back with glandular folds on shoulder and side of back formingdiscontinuous ridges, flanks with glandular warts. (30) Supratympanic fold distinct, from eye to above shoulder.(31) Dorsal parts of forelimb, thigh and tarsus smooth, leg with few glandular warts. (32) Ventral parts of head,body and limbs: throat, chest, belly and thigh smooth. (33) Presence of macroglands: lateroventral gland present asshort continuous glandular ridge in anterior part of flank continuing as separate glands in line; large femoral andaxillary glands; relatively smaller suprabrachial glands.

Coloration. In alcohol. (34) Dorsal and lateral parts of head and body: brown with indistinct darker outline onwarts and ridges; lower part of flanks greyish brown with 8 on the left, 10 on the right, large brown spots; tympanicregion brown with blackish brown line on tympanic fold; upper lip light brown with dark brown bands. (35) Dorsalparts of limbs: proximal part of forelimbs light beige, distal part slightly darker with brown crossbands; dorsal partof thigh, of shank and of foot light brown with indistinct, incomplete darker brown crossbands; posterior part of



thigh beige with large distinct dark brown spots. (36) Ventral parts of head, body and limbs: dirty white with finebrown spots on margin of throat; webbing light brown.

Sexual characters. (40) Oviduct convoluted. (41) Ovary with large creamy-whitish oocytes.

FIGURE 8. Leptolalax pelodytoides, MSNG 29845.A, lectophoront, adult female. (A) Lateral view of head, ventral view of(B) right foot and of (C) right hand. Scale bar = 5 mm (3 + 2 mm).

Variation. The female lectophoront MSNG 29845. A shows a poorly distinct darker zone on snout, an indis-tinct triangle between eyes and spots and lines outlining the warts and ridges on dorsum. The basic dorsal colour isbrown. The flanks are covered by a series of small black spots. The venter is whitish with brown spots on the bor-der of throat and the shanks are brown coloured in their posterior part. The male exonymophoront BMNH1947.2.25.14 figured by Boulenger (1893) shows snout spots and a triangle between the eyes linked to the shoulderspots; a single spot is present in the anterior and another spot in the posterior part of the iliac region; these spotsshow a fine outline. The flanks show a series of dark rather large-sized distinct spots. The warts appear lighter thanthe back but their dark outline is not visible in drawing. Boulenger (1893) mentioned that the throat of the male wasdark-coloured but the chest and belly were white. The presence of femoral glands was mentioned as presence of “around white dark-edged spot”. The lectophoront and the exonymophoront show many differences in dorsal pattern.The colour pattern of the male is unique among all specimens of Leptolalax studied. Beside these two specimens, asingle specimen of L. ventripunctatus and two specimens of L. aereus from Vietnam also show a spot on the snoutbut in these specimens the snout spot is not confluent with the triangle between eyes and with the shoulder spot.

Boulenger (1893) described the webbing as “one third webbed”. As Boulenger did not explain the significanceof terms used, the descriptions in Boulenger (1882) were screened to find species that have similar qualifiers todescribe their webbing. Rana tuberculosa Boulenger, 1882, now Tomopterna tuberculosa, a South African anuranspecies, is “one third webbed”. In this species 3 ½ to 4 phalanges of toe IV are free of webbing (du Preez & Carru-thers 2009). As for a species with very small webbing, L. gracilis, as Leptobrachium gracile in Boulenger (1882) is



described as “slightly webbed” by Boulenger (1882), as having “a very short basal membrane” by Günther (1872),and as “webbing only at base of toes” by Inger & Stuebbing (2005). Thus our finding of 3 ¾ and 4 phalanges of toeIV free of web in L. pelodytoides is consistent with Boulenger’s description.

Distribution. Myanmar: “Karin Hills” (Fig. 9).Natural history. No habitat information was given in the original description, nor other observations on natu-

ral history. The species has not been collected since its original description. Etymology. Resembling a Pelodytes, the small European pelodytid toad. Invariable adjective in apposition to

generic substantive. Placement in molecular phylogeny. No samples available for molecular study. Conservation status. This species as understood here is only known from the type locality and adjacent local-

ities in Myanmar based on the original description. There are no recent data on L. pelodytoides and on its habitatavailable. We propose therefore to change its Red List status from Least Concern to Data Deficient.

FIGURE 9. Distribution of localities of specimens of subgenus Lalos studied in this work. Onymotopes are indicted as stars.Olive green—pelodytoides; blue—L. minimus form 2; orange yellow—L. minimus; red—L. aereus; grey—L. eos; light green –L. ventripunctatus; white—L. bourreti; black—L. pluvialis; yellow—L. nyx.

Leptolalax (Lalos) oshanensis (Liu, 1950)

Megophrys oshanensis Liu, 1950: 13, 197. Onymophoront, by original designation as “type”: holophoront (holotype), Liu Collection 1000 (now CIB 1000), adult male,

SVL 27.0 mm (Liu 1950: 197–198).Onymotope: Emei Shan (29°35’ N, 103°11’ E; alt. 1070 m), Sichuan, China. Collected by Liu Ch’eng-chao, 10 June 1945.

Comment. Leptobrachium minimum was considered a synonym of Megophrys oshanensis by Dubois (1981), butin the genetic analysis, onymotopic specimens of both species appear in distinct clades (Fig. 4). Thus we recognizethem as a distinct species. The name L. oshanensis should be restricted to Chinese populations. Here we give datafrom the original description (Liu 1950), from Fei & Ye (1992) and Fei (1999) for morphological diagnosis. Molec-ular evidence from sequences deposited in GenBank would indicate that the species as currently defined is notmonophyletic. Further studies should be conducted to confirm these results as the sequences here included are fromGenBank without indication of voucher specimens.

Diagnosis. Member of the subgenus Lalos based on the presence of lateroventral glands and molecular phylo-genetic relationships (Fig. 4), distinguished from other species included in this subgenus by the following combina-tion of characters: moderate-sized species (adult males 26.6–30.7 mm) (Table 3) without webbing between toes,



without lateral fringes on toes, with distinct tympanum; thin irregular glandular ridges on back (Table 4); back red-dish brown with pattern including a triangular spot between eyes and lighter glandular spots near vent; a few largedark spots on flanks; throat with dots, belly uniformly white (Table 5).

Description of holophoront (rearranged from original description of Liu 1950; numerous characters missing):Size and general aspect. (1) Specimen of small size (SVL 27.0 mm), body moderately stout.

Head. (2) Head of medium size, as broad (HW 9.5 mm) as long (HL 9.5 mm). (3) Snout slightly protruding;(eye length EL 4.0 mm). (4) Canthus rostralis well defined, loreal region concave, oblique. (6) Nostrils closer to tipof snout than to eyes. (8) Tympanum visible (TYD 1.9 mm). (11) Tongue large, deeply notched.

Forelimbs. (12) Forearm moderately slender (FLL 13.0 mm; HAL 7.0 mm). (14) Relative length of fingers,shortest to longest: I < II = IV < III. (15) Tips of fingers rounded, slightly enlarged. (17) Subarticular tuberclesabsent, but third finger with pad-like elongated elevations. (18) Inner metacarpal tubercle very large, outer one verysmall.

Hindlimbs. (19) Hindlimbs thin (TL 12.0 mm); (FOL 11.5 mm); (TFOL 18.0 mm). (23) Webbing absent. (25)Toes ventrally with elongated pad-like elevations. (26) Inner metatarsal tubercle well developed. (28) Outer meta-tarsal tubercle absent.

Skin. (29) Dorsal and lateral parts of head and body: snout, region between eyes and side of head smooth; backwith glandular ridges; a few light-coloured warts distributed around vent. (30) Supratympanic fold well-developed,from eye extending postero-ventral to shoulder region. (32) Ventral parts of head, body and limbs: throat, chest,belly and thigh very smooth. (33) Presence of macroglands: lateroventral gland composed of white glands arrangedin rows between axilla and groin; large oval suprabrachial and femoral glands present.

Coloration. In life: (34) Dorsal and lateral parts of head and body: Reddish brown all over body with a blacktriangular mark on back of head, with a few black bars on mouth, and scattered black spots on body. (35) Dorsalparts of limbs: black bars distinctly developed. In alcohol: after preservation, the reddish brown becomes greyishbrown. (36) Ventral parts of head, body and limbs: throat with irregular dots, belly without spots.

Male secondary sexual characters: (38) Vocal sacs: no outer modification of skin; paired internal subgularvocal sacs.

Distribution. China: Guizhou Province, Hubei Province, Sichuan Province.Natural history. As described by Liu (1950: 199): “L. oshanensis was found under stones in corn fields on

hillsides at about 3500 feet. Adults are difficult to find, as I got only two specimens of adult males in three sum-mer’s work. Tadpoles are very abundant in small mountain streams.” The species is known to occur in mountainareas from 700 to 1800 m. Tadpoles were collected from side pools, in pools beneath cascades, on the bottom ofpools, in cracks between stones, hiding in vegetation or roots of bamboos and trees at the margin of the water. Theystay in shallow running water with the head directed up-stream and the tail curved like an eel (Fei & Ye 1992).

Etymology. Adjective as epithet of generic nomen (same grammatical gender), meaning “from O Shan”, i. e.,from Emei Shan, a famous mountain in Sichuan.

Placement in molecular phylogeny. Specimens allocated to Leptolalax “oshanensis” do not form a mono-phyletic group. They are members of a highly supported clade that groups L. eos, L. bourreti and Leptolalax sp.from Doi Chiang Dao. L. “oshanensis” AY561306, from the onymotope, and thus probably “true” oshanensis, issister to L. eos whereas L. “oshanensis” AY526215 is sister to Leptolalax sp. from Doi Chiang Dao. Further studiesare needed to reliably allocate the name oshanensis to probably one of these mitochondrial lineages and to describethe other of these mitochondrial lineages as new species.

Conservation status. We confirm the Red List status of this species as Least Concern. In the future, L. oshan-ensis might be dismantled into various species and then a re-evaluation will be needed.

Leptolalax (Lalos) bourreti Dubois, 1983(Fig. 10–13)

Megophrys pelodytoides — Bourret 1937: 17; Bourret 1942: 208, Fig. 45 (a2).Leptolalax bourreti Dubois, 1983: 150. Onymophoront, by original designation as “holotype”: holophoront, MNHN 1938.0094, subadult male, SVL 36.2 mm. Onymotope: Chapa [Sa Pa] (22°21’ N, 103°52’ E), Lao Cai Province, Vietnam. Collected by René Bourret, before 1937.



FIGURE 10. Leptolalax bourreti, MNHN 1938.0094, holophoront, subadult male, SVL 36.2 mm. (A) Dorsal and (B) ventralview.

Material examined. Vietnam. Sa Pa, Hoang Lien Nature Reserve (22°19’N, 103°47’E): MNHN 1938.0094, sub-adult male, holophoront, coll. René Bourret, before 1937; MNHN 1938.0093, MNHN 1938.0095, adult females,other protaxonts (paratypes); MNHN 1999.5659–5660, 1999.5665, MNHN 1999.5669–5671, adult males, coll.Annemarie Ohler, Stéphane Grosjean & Steven Swan, 8 November 1997, 20 January 1998, 19 February 1998 and28 July 1998; MNHN 1998.8702–8742, tadpoles, 19 and 21 July 1998; MNHN 1998.8743–8763, tadpoles, 28 July1998, in the quiet part of a small torrent; MNHN 1998.8764–8769, tadpoles, 02 August 1998; MNHN 1998.8770–8774, tadpoles, 5 August 1998, at 1860 m a.s.l. in a large torrent more than 5 m wide with a strong current, coll.Stéphane Grosjean.

Comments. Dubois (1983) described this large species from a series of frogs collected by Bourret in 1937 andconsidered by the collector as Megophrys pelodytoides (Bourret 1937: 17; 1942: 208). Subsequent collections ofLeptolalax in Sa Pa (Ohler et al. 2000: 74) consisted of specimens of slightly smaller body size but which can beallocated to L. bourreti by coloration and morphological characters. A specimen from Ha Giang, Vi Xuyen, Cao Bo(AMNH A163810) mentioned as L. bourreti by Frost et al. (2006), sequence available on Genbank (accessionnumber DQ283381), is morphologically quite different from the L. bourreti hypodigm (Dubois 1983) and groupsin a distinct clade from L. bourreti onymotopic specimens (see description below).

As described below under Variation, this species apparently exhibits variation in iris colour and other chro-matic characters. Especially iris colour is often species-specific in frogs and we cannot exclude that these differ-ences point to the existence of more than one species currently subsumed under bourreti. However, for the time



being we have no evidence that there is more than one species as the iris colour differences in Sa Pa cannot be cor-related to other morphological or colour differences, and insufficient molecular data are available to analyse thisquestion further.

Diagnosis. Member of the subgenus Lalos based on the presence of a lateroventral gland and molecular phylo-genetic relationships (Fig. 4), distinguished from other species included in this subgenus by the following combina-tion of characters: large-sized species (males 28.0–36.2 mm; females 42.0–45.0 mm) (Table 3) with small webbingand distinct fringes on toes; tympanum distinct; dermal ridges under toes poorly distinct; finger tips slightlyenlarged; dorsal skin smooth (Table 4); dorsum reddish, greenish or brown with dark spots; moderate sized darkspots on flanks; ventral side whitish; iris colour variable, clearer above, copper, green or brownish (Table 5).

FIGURE 11. Leptolalax bourreti, MNHN 1938.0094, holophoront, subadult male. (A) Lateral view of head, ventral view of(B) right foot and of (C) right hand. Scale bar = 5 mm (3 + 2 mm).

Description of the holophoront, MNHN 1938.0094. Size and general aspect. (1) Specimen of moderate size(SVL 36.2 mm), body slender.

Head. (2) Head of large size, narrower (HW 13.4 mm) than long (HL 14.1 mm; MN 12.1 mm; MFE 9.5 mm;MBE 4.9 mm), flat. (3) Snout not protruding, its length (SL 5.5 mm) longer than horizontal diameter of eye (EL 4.7mm). (4) Canthus rostralis rounded, loreal region slightly concave, obtuse. (5) Interorbital space flat, as large (IUE3.4 mm) as upper eyelid (UEW 3.4 mm) and internarial distance (IN 3.5 mm); distance between front of eyes (IFE5.8 mm) about half of distance between back of eyes (IBE 10.1 mm). (6) Nostrils closer to eye (EN 2.7 mm) than totip of snout (NS 2.9 mm). (8) Tympanum (TYD 1.9 mm) distinct, rounded; about two-fifths of eye diameter, tym-panum-eye distance (TYE 1.6 mm) slightly smaller than its diameter. (11) Tongue large, rounded.



Forelimbs. (12) Forearm short, thin (FLL 9.5 mm), longer than hand (HAL 8.7 mm), not enlarged. (13) Fingerslong and thin (TFL 5.7 mm). (15) Tips of fingers slightly enlarged. (16) Fingers II and III with dermal fringe.

Hindlimbs. (19) Shanks about four times longer (TL 17.8 mm) than wide (TW 3.7 mm), longer than thigh (FL16.0 mm) and distance from base of internal metatarsal tubercle to tip of toe IV (FOL 16.8 mm). (20) Toes rathershort and rather strong; toe IV (FTL 9.9 mm) longer than one-third of distance from base of tarsus to tip of toe IV(TFOL 26.8 mm). (22) Tips of toes not enlarged. (23) Webbing present, very small; fringes present, distinct: I 2 – 3II 2 – 3 III 2 ½ – 4 IV 4 – 2 ½ V (MTTF 6.5 mm; MTFF 6.3 mm; TFTF 9.6 mm; FFTF 10.3 mm). (24) Dermalridge along toe V present, from tip of toe to base of finger, poorly developed. (26) Inner metatarsal tubercle short,distinct; its length (IMT 1.6 mm) 2.4 times in length of toe I (ITL 3.8 mm).

Skin. (29) Dorsal and lateral parts of head and body: snout, region between eyes, side of head, back and flanksmooth (possibly due to preservation). (30) Supratympanic fold poorly prominent, from eye to above upper fore-limb. (31) Dorsal parts of limbs: smooth. (32) Ventral parts of head, body and limbs: smooth. (33) Presence of mac-roglands: a suprabrachial and a large oval axillary gland present; femoral glands and lateroventral gland indistinct.

FIGURE 12. Leptolalax bourreti, MNHN 1999.5659, adult male, SVL 29.8 mm. (A) Dorsal and (B) ventral view.

Coloration. In alcohol (pattern and coloration modified due to preservation): (34) Dorsal and lateral parts ofhead and body: light grey brown. (35) Dorsal parts of limbs: light grey brown with indistinct brown crossbands;posterior part of thigh light grey brown. (36) Ventral parts of head, body and limbs: greyish; webbing greyish.

Male secondary sexual characters: (38) Vocal sacs: no opening on mouth floor.



Variation. Bourret (1937) described the coloration in life of the Sa Pa specimens mentioned as Megophrys pe-lodytoides as follows (translation): “Patterns and colour very variable; back grey brown verging more or less to red,dark grey green, more or less clear or dark brown, dirty green or blackish; transversal spots on lips and hindlimbsdark. Underside whitish. Dark spots more or less distinct, marbling, or symmetrical markings sometimes with lightoutline. Often fine light dots, greenish white above, yellow on the flanks and on the shanks. A light vertical spot onthe tip of snout seems to be constant; an orange tympanic spot occurs frequently. The iris is also variable, dottedwith dark green-brown, a little clearer above and below than on the sides; yellow above, grey brown below; lightyellowish brown; light brown-yellow reticulated with dark brown; coppery, green or brownish green, finely reticu-lated with black, spotted with red in its upper part.” The first author noted life coloration of MNHN 1999.5659 inthe field: Iris copper on upper part, golden in lower part separated by a black horizontal band. Dorsum and upperpart of flank brown with darker spots, lower part lighter brown with black brown spots; loreal and tympanic regionbrown with a dark black brown band on canthus rostralis and tympanic fold; tympanum black brown; upper lipbrown with black brown bands; forelimbs and hindlimbs brown with darker crossbands; posterior part of thighbrown with darker spots; throat ivory white, transparent; margin of throat brownish with white spots; chest lightreddish; belly yellow getting orange in posterior part; webbing light brown; macroglands light orange.

All the seven males with large testes collected from Sa Pa in 1998 considered as adults, have vocal sac open-ings in the mouth. As in all other species of Leptolalax the adult males have internal vocal sacs, we consider thatthe male holophoront of Leptolalax bourreti, not exhibiting such structures, is a subadult.

Females have large (diameter 2.01 mm), yellowish white oocytes in ovary. The specimen MNHN 1938.0093,adult female, part of the hypodigm, exhibits a still distinct colour pattern: Dorsum fawn with dark brown spots onback and upper flank. A dark brown line on canthus and supratympanic fold; dark bands on upper lip, forelimbsand hindlimbs. The specimens originally mentioned by Bourret (1937) vary in coloration, but show spots andmarkings on back, limbs and head.

The snout-vent length of nine adult females varies from 41 to 54 mm. Six adult males collected in 1998 aresmaller and measure 29.5–34.0 mm and are smaller than the two males measured by Bourret (36.2 in the ho-lophoront). The only adult female in the hypodigm (Dubois 1983) measures 43.8 mm and falls into the range givenby Bourret (1937).

All recently collected specimens of L. bourreti exhibit macroglands. The lateroventral glandular ridge is com-posed of a series of glands that form a continuous line. Femoral and axillary glands are rounded or oval; whereasthe femoral glands are always clearly distinct, the axillary glands are poorly distinct in a few specimens. Thesuprabrachial glands are of round or oval shape and smaller than the femoral and axillary glands. In specimensreaching metamorphosis, the lateroventral, femoral and suprabrachial glands are distinct, but the axillary glandbecomes distinct only when metamorphosis is completed.

Tadpole. Presently two species of Leptolalax are known to occur in the Hoang Lien mountain range: L. bour-reti and L. pluvialis. They can be distinguished based on extent of webbing and on colour pattern as these charac-ters are already differentiated when tadpoles enter metamorphosis. On this basis we ascribed the tadpoles collectedfrom Sa Pa to L. bourreti although more species could be found in this area. The tadpoles described below are com-mon in the Hoang Lien Nature Reserve (Sa Pa) and were collected in three different places. On one occasion, noadult Leptolalax was observed in the area, on the second occasion adult of L. bourreti were collected with the tad-poles and in the third occasion tadpoles were collected in the onymotope of L. pluvialis (the two species are knownto occur syntopically in this area).

The external morphology is based on a specimen in stage 31 (MNHN 1998.8737; TOL 59.2 mm and BL 18.9mm).

In dorsal view (Fig. 13A), body elliptical, widest at the middle of the intestinal coil, snout semi-circular, a pairof large lateral lymphatic sacs extending from spiracle to end of body. In lateral view (Fig. 13B), body depressed,BW 143 % of BH, highest just behind the spiracle; snout rounded. Eyes moderately small, ED 7.5 % of BL,slightly bulging and not visible in ventral view, positioned almost dorsally and directed more laterally than antero-dorsolaterally. Pineal ocellus not visible; nasolacrymal ducts not visible. Nares round with an oval opening, moder-ately large, rimmed with four small lobes, positioned almost dorsally but directed anterolaterally with an openingdirection dorsolateral, closer to snout than to pupils, RN 40 % of NP; NN 82 % of PP. Spiracle sinistral, square,



moderately small, internal wall fused with body wall except its tip, positioned just below the lateral axis and situ-ated at equal distance from snout to end of body, SS 52 % of BL, oriented posterodorsally; its opening closedagainst body wall, directed posterodorsally, at the height of the apex of myotomes of the caudal muscle. Tail long,TAL 209 % of BL; tail musculature well developed, TMH 71 % of BH and 61 % of MTH, TMW 55 % of BW, thelower part slightly bulging in the proximal quarter then gradually tapering, reaching tail tip. Upper fin shallow in itsproximal third then moderately shallow, UF 30 % of MTH, not extending onto body, SU 97 % of BL, almost non-existent in its proximal sixth then increasing rather abruptly, the remaining part of about equal height, slightly con-vex and following caudal muscle; lower fin shallow in proximal third then moderately shallow, LF 26 % of MTH,straight and so increasing in size anteroposteriorly; point of maximum height of tail located just before the proxi-mal third, MTH 116 % of BH, tail tip finely rounded. Anal tube moderately large, as a short tube, dextral anddirected posteriorly, entirely attached to ventral fin; opening large, lateral, bevelled, posterolaterally directed. Lat-eral line system consisting of a dorsal body line which begins at the rear of eye and passes just above the lateral sac,a middle body line which joins the dorsal body line at midway of tail; on the body, the angular line curves behindthe eye and passes on the upper surface of the lateral sac, the ventral body line, parallel and posterior to the latter,passes laterally on the lateral sac, an unnamed line in the terminology of Lannoo (1987) starts from the spiracle andpasses ventrally on the lateral sac; anteriorly, anterior oral line, longitudinal oral line, supraorbital line and infraor-bital line present, a short line joinning the supraorbital and the infraorbital lines anterior to eye and another joiningthese two lines anterior to naris; a short unnamed line lying ventroposteriorly almost parallel to body; the system issymmetrical on both sides of the tadpole. Lateral line organs dash-shaped, all longitudinally oriented except thoseof the dorsal line which are perpendicular to the longitudinal axis. A pair of small axillary glands ventrally at thelevel of the gills; another one on the posterior part of flanks, at a short distance before the insertion of tail, halfwaybetween the insertion of hind limbs and the apex of the myotomes of the caudal muscle; a unilateral gland on theleft side just before the spiracle.

Oral disc (Fig. 13C) cup-like, transversely elongate at rest, positioned and directed ventrally, not emarginatedlaterally, moderately large, ODW 27 % of BL and 45 % of BW. Lower labium divided by a medial notch, upperlabium with four lobes, the median two separated by a deeper fold forming a median fold. A continuous row ofmarginal papillae all around the oral disc, papillae of small size, conical and pointed, the papillae of the upperlabium with a larger base (triangular) than those of the lower labium, finer; eight submarginal papillae present lat-erally to keratodont rows of the lower labium and anteromedially to row P3. Denticulate papillae present on thelower labium. KRF 1:3+4/2+2:1, keratodont rows short, A3 > A4 > A2 > A5 > A1, A1 very short, A2 shortly inter-rupted in the middle, the other lower rows separated by the upper beak; P1>P2>P3; A1 very short, one-third of P3;P3 half of P2. Keratodonts scythe blade-shaped. Jaw sheaths strong, coarsely serrated, black with a brown blurringjust above the serrations; upper sheath a large arch with a stronger and bigger median serration; lower sheath as anopen V.

Coloration in life: body and caudal muscle brown olive with lighter spots on fore part of head; numerous spotsformed by iridiophores. Fins light olive with a few darker spots on distal part.

Colour in preservative: Upper part of body and upper flanks grey brown, extension of caudal muscle onto bodylight brown; lower flanks in gradation from brown to grey. Ventral part grey transparent. Caudal muscle lightbrown mottled with ochre brown, especially in proximal third of upper part, ventral side lighter. Fins transparentgrey, upper fin brownish proximally. Nares and hindlimbs white.

The variation is based on 10 other specimens, except where noted (MNHN 1998.8726–8727, 1998.8731,1998.8738–8739, 1998.8742, 1998.8747, 1998.8762, 1998.8772, 1998.8774, in stages 25–36, TOL 38.2–56.0 mmand BL 10.9–20.8 mm). Only the ratio values different from those of the described specimen are given: BW 115–138 % of BH (n = 9); ED 6.3–10.1 % of BL; RN 41–61 % of NP; NN 75–88 % of PP; SS 45–53 % of BL; TAL157–261 % of BL; TMH 59–74 % of BH; TMH 54–67 % of MTH; TMW 52–59 % of BW (n = 8); UF 29–35 % ofMTH; LF 24–28 % of MTH; SU 101–133 % of BL; MTH 102–120 % of BH; ODW 22–28 % of BL; ODW 44–48% of BW. The more the tadpole is in an advanced stage, the more the upper fin begins close to body end. The KRFvary from 1:3+3/2+2:1 to 1:4+4/3+3:1, the additional keratodont rows appears between stages 27 and 29.

Distribution. Vietnam: Lao Cai Province (Fig. 9).



FIGURE 13. Tadpole of Leptolalax bourreti. (A) Dorsal and (B) lateral views of a specimen in stage 31 (MNHN 1998.8737),scale bar = 10 mm; (C) oral disc of a specimen in stage 29 (MNHN 1998.8739), scale bar = 1 mm.

Natural history. One of the female specimens of hypodigm (MNHN 1938.0095) has large oocytes, but thoseof the other females are half size. Adult males from Sa Pa (MNHN 1999.5659–5660, 1999.5665–5666, 1999.5669–5671) were collected in January, February, July and November and all have vocal sac openings on the mouth floor.

Specimens collected in 1998 were found in tropical montane deciduous forest at altitudes from 1150 to 1815 mnear streams, sitting on leaves, on tree trunks or on the ground on dead leaves at a height of 0–0.5 m and at a dis-tance of 0–50 m from the stream. When breeding, these frogs were usually in or near (0.4 m) the water. This spe-cies was abundant as compared to the other species observed in Sa Pa at the study period (more than 26 records).Few were observed in agricultural land, scrub and savannah, most were observed in submontane and montane for-est near rivers and streams.

The tadpoles live in mountain streams of variable size and current strength, but in the low-flow parts, in poolsoften created before small dams formed by dead vegetation lying across streams, but always in parts directly con-nected to the main water and where current is present. Other tadpoles found in the same streams belong to Lepto-brachium ailaonicum, Xenophrys sp., Quasipaa verrucospinosa and Amolops sp.

Etymology. Dedicated to René Bourret (1884–1957). This nomen is a patronym in the genitive. Placement in molecular phylogeny. Specimens allocated to L. bourreti apear in a highly supported clade that

groups L. eos, L. “oshanensis” and Leptolalax sp. from Doi Chiang Dao. They are included in a poorly supportedsubclade with L. “oshanensis” AY561306 and Leptolalax sp. From Doi Chiang Dao which is sister group of L. eosand L. “oshanensis” AY526215. The molecular differentiation within this cluster of forms is not high, and the rela-tionships and status of the various lineages included in this clade require further study.

Conservation status. Recent observations show that the species is abundant in its habitat. It has been observed



in a rather restricted area, but this is clearly due to lack of exploration and difficulty in identifying the species. Weassume that this species should be qualified as Least Concern but suggest to keep it as Data Deficient until theexpected additional locality records become available and its taxonomic status confirmed.

Leptolalax (Lalos) eos n. sp.(Fig. 14–16a)

Megophrys pelodytoides—Liu & Hu 1961: 48.Leptolalax pelodytoides—Yang 1991; Fei & Ye 1992: 246–253; Stuart 1999: 44.Leptolalax bourreti—Stuart 2005: 474.Onymophoront, by present designation: holophoront (holotype), MNHN 2004.0274, adult male, SVL 34.7 mm.Onymotope: Long Nai (21°14’ N, 101°56’ E; 800 m), Phongsaly Province, Laos. Collected by Annemarie Ohler, 14 July 2004.Other protaxonts (paratypes): Laos. Phongsaly: MNHN 2004.0275–0279, 5 adult males, MNHN 2005.0147, adult female

(paratypes), coll. Annemarie Ohler, 14 and 26 July 2004, and 4 February 2005.Other protaxonts (non type-specimens): Laos. Bolikhamxay Province, Khankent District, Nape border area (18°18’ N, 105°4’

E): FMNH 258035–258040, 6 adult males, coll. David Davenport, 15–18 March 1997.

Comment. Stuart (2005) reported this species as L. bourreti from northern Laos. Liu & Hu (1961), Yang (1991)and Fei & Ye (1992) mentioned a large-sized Leptolalax from southern Yunnan under the epithet pelodytoides.Figure 55 of Yang (1991) shows a frog with indistinct spots on dorsum and flanks, corresponding coarsely to thecolour pattern of L. eos. Based on this colour pattern and the body size mentioned by these authors, these Chinesefrogs might be related to or conspecific with L. eos, but this hypothesis requires testing. We suggest to treat theseChinese frogs as Leptolalax cf. eos since they clearly are more similar in size and colour pattern to this species thanto L. pelodytoides or other Leptolalax species currently known from this subregion.

The comparison of L. eos with the hypodigm of L. bourreti is difficult as the latter specimens are in rather poorpreservation condition. In particular their colour pattern has faded out. Nevertheless the holophoront of L. bourretishows distinct dark spots on flanks. Also figure 45 a2 of Bourret (1942) shows numerous distinct black spots on theflanks. Such spots are absent in all specimens of L. eos.

The species is slightly smaller than L. bourreti. It shows differences in several measurements on the head, par-ticularly concerning the position of eyes and nostril (MN, MFE, IFE, IBE, EL), size of tympanum (TYD), andlength of digits (TFL, FTL) and metatarsal tubercle (IMT) (Table 9).

Diagnosis. Member of the subgenus Lalos based on the presence of a lateroventral gland and molecular phylo-genetic relationships (Fig. 4), distinguished from other species included in this subgenus by the following combina-tion of characters: large-sized species (males 33.1–34.7 mm; female 40.7 mm) (Table 3) with poorly developedwebbing and well developed fringes on toes; tympanum distinct; dermal ridges under toes well developed; fingertips enlarged; dorsal skin shagreened (Table 4); dorsal pattern poorly distinct; blackish spots on flanks absent; ven-tral side whitish, iris orange above, light golden below (Table 5).

Description of the holophoront. Size and general aspect. (1) Specimen of moderate size (SVL 34.7 mm),body rather stout.

Head. (2) Head of rather large size, longer (HL 13.1 mm) than wide (HW 11.8 mm; MN 10.8 mm; MFE 8.5mm; MBE 4.2 mm), flat. (3) Snout slightly protruding, its length (SL 5.7 mm) longer than horizontal diameter ofeye (EL 4.7 mm). (4) Canthus rostralis rounded, loreal region concave, obtuse in cross section. (5) Interorbitalspace flat, larger (IUE 5.4 mm) than upper eyelid (UEW 4.5 mm) and than internarial distance (IN 3.6 mm). (6)Nostrils closer to eye (EN 2.6 mm) than to tip of snout (NS 3.1 mm). (8) Tympanum (TYD 2.3 mm) distinct,rounded; about half eye diameter, tympanum-eye distance (TYE 1.5 mm) smaller than its diameter. (11) Tonguelarge, cordate.

Forelimbs. (12) Forearm short, thin (FLL 8.5 mm), shorter than hand (HAL 9.3 mm), not enlarged. (13) Fin-gers long and thin (TFL 4.7 mm). (15) Tips of fingers slightly enlarged. (16) Finger II with distinct dermal fringeon inner side.

Hindlimbs. (19) Shanks about four times longer (TL 16.7 mm) than wide (TW 4.6 mm), longer than thigh (FL15.5 mm) and than distance from base of internal metatarsal tubercle to tip of toe IV (FOL 15.9 mm). (20) Toe IV(FTL 8.6) about one-third of distance from base of tarsus to tip of toe IV (TFOL 25.2 mm). (22) Tips of toes notenlarged. (23) Webbing present, small, fringes well developed: I 1 – 2 II 1 ½ – 2 ½ III 2 – 3 ½ IV 4 – 2 ½ V (MTTF



6.8 mm; MTFF 6.1 mm; TFTF 9.0 mm; FFTF 10.1 mm). (24) Dermal finge along toe V absent. (26) Inner metatar-sal tubercle distinct, short; its length (IMT 1.8 mm) 2.0 times in length of toe I (ITL 3.6 mm).

FIGURE 14. Leptolalax eos, MNHN 2004.0274, holophoront, adult male, SVL 34.7 mm. (A) Dorsal and (B) ventral view.

FIGURE 15. Leptolalax eos, MNHN 2004.0274, holophoront, adult male. (A) Lateral view of head, ventral view of (B) rightfoot and of (C) right hand. Scale bar = 5 mm (3 + 2 mm).



FIGURE 16. Specimens of Leptolalax in life. (a) L. eos, MNHN 2004.0274, holophoront, adult male, SVL 34.7 mm. (b) L.minimus form 2, THNHM 07417, reference specimen, adult male, 25.6 mm. (c) L. minimus form 2, THNHM 07478, adultmale, 25.8 mm. (d) Leptolalax minimus, MNHN 2006.2553, adult male, SVL 28.1 mm. (e) L. minimus, adult male (not col-lected).



TABLE 9. Comparison of Leptolalax bourreti (holophoront and other protaxonts) and L. eos (holophoront and other pro-taxonts). Values in fields filled with grey do not overlap when comparing males and females respectively.

L. bourreti L. eosMaleN=1

FemalesN=3

MalesN=6

FemaleN=1

SVL 36.2 42.9 ± 1.841.8–45.0

33.8 ± 0.633.1–34.7 40.7

HW/SVL 370 350 ± 30.9316–376

355 ± 17.8337–384 349

HL/SVL 390 385 ± 19.8369–407

392 ± 12.4378–408 407

MN/SVL 334 326 ± 23.5300–345

321 ± 11.1308–333 352

MFE/SVL 262 250 ± 25.6222–271

251 ± 6.5245–260 277

MBE/SVL 135 135 ± 18.2115–150

134 ± 12.1121–150 153

IFE/SVL 160 155 ± 10.9142–162

192 ± 5.5185–199 183

IBE/SVL 279 269 ± 6.2262–274

310 ± 14.2294–332 273

IN/SVL 97 97 ± 11.587–110

104 ± 4.599–110 97

EN/SVL 75 73 ± 4.269–78

75 ± 3.970–81 95

EL/SVL 130 127 ± 3.4124–131

138 ± 4.7133–146 119

TYD/SVL 52 58 ± 5.454–64

64 ± 7.256–73 58

TYE/SVL 44 43 ± 4.539–48

47 ± 4.543–54 40

FLL/SVL 254 257 ± 10.6249–269

257 ± 18.7232–286 258

HAL/SVL 262 248 ± 13.7235–262

277 ± 12.4261–296 259

TFL/SVL 117117–117

148 ± 11.9135–168 138

TL/SVL 492 481 ± 28.8452–510

504 ± 21.1481–533 506

FOL/SVL 464 457 ± 29.9423–478

477 ± 22.4455–514 466

FTL/SVL 216216–216

262 ± 11.5248–278 239

IMT/SVL 44 42 ± 6.138–49

52 ± 2.848–56 33

ITL/SVL 105 88 ± 4.184–91

107 ± 7.197–118 94

MTTF/SVL 180 183 ± 7.9174–188

191 ± 9.3174–200 177

MTFF/SVL 174 180 ± 6.9172–184

175 ± 5.1166–181 171

TFTF/SVL 265 242 ± 14.8226–255

274 ± 16.4258–294 255

TFFTF/SVL 285 261 ± 19.0240–274

308 ± 17.1292–330 268



Skin. (29) Dorsal and lateral parts of head and body: snout, region between eyes, side of head and dorsum sha-greened; flank with glandular warts. (30) Supratympanic fold prominent, from eye to above upper forelimb. (31)Dorsal parts of limbs: forelimbs shagreened; thigh and shank with glandular warts; tarsus smooth. (32) Ventralparts of head, body and limbs smooth. (33) Presence of macroglands: lateroventral ridges formed by a series ofclose but separate fine glands; small suprabrachial and larger femoral glands present; axillary glands poorly dis-tinct.

Coloration. In alcohol. (34) Dorsal and lateral parts of head and body: Brown grey with darker pattern, in par-ticular interorbital triangle; small white spots on back; lower part of flanks dark grey with white spots correspond-ing to glands; loreal and tympanic region brown grey; tympanum brown, lower part beige. (35) Dorsal parts oflimbs: upper forelimb beige; forearm, thigh, shank and foot dark grey with crossbands; posterior part of thighbrown finely spotted with beige. (36) Ventral parts of head, body and limbs: throat creamy white; margin of throatbrown with creamy white dots; chest and belly creamy white; thigh creamy white and dark brown; webbing darkbrown; macroglands creamy white.

In life (Fig. 16A): Upper part of iris orange, lower part light golden, separated by a dark horizontal band. Dor-sal parts of head and body and upper parts of flanks brown with indistinct darker spots; lower part of flanks slightlyclearer brown; a light pinkish line (corresponding to glands) separating a brown band on side of body; lorealregion, tympanic region and tympanum dark brown; upper lip brown with blackish bands; forearm, thigh, shankand foot brown with blackish crossbands; posterior part of thigh dark brown with an orange zone near vent and alight beige femoral gland; throat white with orange reflections; border of throat brown with pinkish dots; vocal sacreddish transparent; chest and belly orangish pearly; ventral part of thigh dark brown with pinkish dots; webbingdark brown. Description of coloration was made the day after capture. At night, when captured, dorsal surface ofspecimen was of light beige coloration of pinkish aspect.

Comparison. Among the species with ventrolateral glands, L. eos shows usually almost uniform dorsal color-ation, rarely faint spots and can thus be distinguished from L. aereus, L. alpinus, L. bourreti, L. khasiorum, L. later-alis, L. liui, L. melanoleucus, L. nyx, L. oshanensis, L. pelydytoides, L. pluvialis, L. tamdil, L. tuberosus and L.ventripuncatatus which all exhibit distinct darker spots on dorsum. The absence of dark brown spots on flanks in L.eos allows to distinguish it from all species with ventrolateral glands. Its white throat, chest and belly distinguish itfrom L. lateralis, L. nahangensis, L. oshanensis, L. pluvialis, L. aereus, L. tuberosus and L. ventripunctatus. L. eosshares with L. pelodytoides relatively large webbing (“small”, means less than 4 phalanges free of web) whereasthe other species show less developed web (“rudimentary” or “basal”). The presence of large fringes on toes distin-guish this species from L. alpinus, L. aereus, L. bourreti, L. fuliginosus, L. lateralis, L. minimus, L. nahangensis, L.nyx, L.oshanensis, L. pelodytoides, L. pluvialis, L. sungi, L. tamdil, L. tuberosus and L. ventripunctatus.With a SVLof 33–35 mm in adult males and more than 40 mm in adult females, L. eos is among the large-sized species ofLalos. L. sungi (males SVL 48–53 mm; females 57–59 mm) is the only known larger species. In L. bourreti, malesmight reach SVL over 33 mm and females 42–45 mm, whereas in L. nyx only females might reach similar sizes asknown for L. eos. For all other species adults are smaller.

Distribution. Laos: Phongsaly Province, Bolikhamxay Province (Fig. 9). China: Yunnan Province (tenta-tively).

Natural history. We collected this large-sized Leptolalax from northern Laos near Chinese border in sympatrywith a small-sized form, allocated to L. ventripunctatus. Specimens were collected in a small patch of primary for-est with a small stream (Fig. 36A). They were sitting on the ground or at a height up to 0.4 m at a distance of 0.2–0.4 mm from the water on rocks and, a single individual was on a leaf. In July and August specimens were not call-ing.

Etymology. Greek [eos], dawn of day, daybreak, morning, east [aurora]. is the personified Aurora,“the rosy fingered dawn” as Homer called her, the goddess of the Morning Star in ancient Greece. The nomenrefers to the dorsal colour of the specimens in life. Nomen used as a noun in apposition to generic substantive.

Placement in molecular phylogeny. Specimens allocated to L. eos form a highly supported monophyleticclade which includes all specimens assigned to this taxon by morphology. This clade is part of a highly supportedclade that groups L. bourreti, L. “oshanensis” and Leptolalax sp. from Doi Chiang Dao. The eos clade is sister to L.“oshanensis” AY526215.

Conservation status. This species has been observed by several authors and is morphologically quite easy torecognize, so its range should be rather precisely known. Its extent of occurrence is estimated to be smaller than20 000 km². As populations observed are small and threats like deforestation for shifting agriculture lay on habitat,we propose a Red List status for L. eos as Vulnerable (criterion B1).



Leptolalax (Lalos) pluvialis Ohler, Marquis, Swan & Grosjean, 2000(Fig. 17–18)

Leptolalax pluvialis Ohler, Marquis, Swan & Grosjean, 2000: 74. Onymophoront: holophoront, by original designation as “holotype”, MNHN 1999.5675, adult male, SVL 22.0 mm. Other protaxonts: MNHN 1999.5674 and MNHN 1999.5676, 2 adult males (paratypes), SVL 21.3–22.3 mm. Onymotope: Fan Si Pan mountain range (22°19’ N, 103°47’ E; 1900 m), Lao Cai Province, Vietnam. Collected by Annemarie

Ohler and Stéphane Grosjean, 28 July 1998.

Material examined. Vietnam. Lao Cai Province, Fan Si Pan mountain range (22°19’ N, 103°47’ E; 1900 m):MNHN 1999.5674–5676, adult males (hypodigm). Coll. Annemarie Ohler and Stéphane Grosjean, 28 July 1998.

Comment. All three specimens of the hypodigm have been included in the molecular analysis. They form thesister group of specimens allocated to a clade that includes L. aereus, L. minimus, L. nyx and L. ventripunctatus.They can be distinguished morphologically from the other representatives of this clade by their smaller size and thedense brown spotting on belly and chest. The holophoront is redescribed to allow easier comparison and in orderthat all characters used for the new species are described in the same way.

Diagnosis. Member of the subgenus Lalos based on the presence of lateroventral glands and molecular phylo-genetic relationships (Fig. 4), distinguished from other species included in this subgenus by the following combina-tion of characters: very small-sized species (males 21.0–22.0 mm) (Table 3) with rudimentary webbing and nofringes on toes; tympanum distinct; dermal ridges under toes distinct; finger tips not dilated; dorsal skin smooth(Table 4); dorsum greyish brown with dark pattern; few black spots on flanks; ventral side with dense brown spots;iris dark golden above (Table 5).

Description of the holophoront, adult male MNHN 1999.5675. Size and general aspect. (1) Specimen ofsmall size (SVL 22.0 mm), body moderately slender.

Head. (2) Head of medium size, narrower (HW 8.1 mm) than long (HL 8.5 mm; MN 6.7 mm; MFE 5.5 mm;MBE 2.2 mm), convex. (3) Snout slightly protruding, its length (SL 4.0 mm) longer than horizontal diameter of eye(EL 3.3 mm). (4) Canthus rostralis angular, loreal region concave, vertical. (5) Interorbital space convex, slightlylarger (IUE 2.8 mm) than upper eyelid (UEW 2.7 mm) but narrower than internarial distance (IN 3.0 mm); distancebetween front of eyes (IFE 4.6 mm) about two-thirds of the distance between back of eyes (IBE 7.5 mm). (6) Nos-trils oval, closer to eye (EN 1.4 mm) than to tip of snout (NS 2.0 mm). (8) Tympanum (TYD 1.7 mm) distinct,rounded; about half of eye diameter, tympanum-eye distance (TYE 0.7 mm) two-fifths of its diameter. (11) Tonguemoderate, cordate.

Forelimbs. (12) Forearm short, thin (FLL 6.2 mm), longer than hand (HAL 6.0 mm), not enlarged. (13) FingersI–II and IV rather short and rather strong; finger III long and rather strong (TFL 4.0 mm). (15) Tips of fingers notenlarged. (16) Finger II with dermal fringe.

Hindlimbs. (19) Shanks about four times longer (TL 11.4 mm) than wide (TW 3.1 mm), longer than thigh (FL10.9 mm) and than distance from base of internal metatarsal tubercle to tip of toe IV (FOL 10.8 mm); heel reachingnare when limbs folded forward. (20) Toe IV (FTL 6.35) about one-third of distance from base of tarsus to tip oftoe IV (TFOL 17.2 mm). (22) Tips of toes not enlarged. (23) Webbing absent (MTTF 3.9 mm; MTFF 3.7 mm;TFTF 6.6 mm; FFTF 6.7 mm). (24) Dermal fringe along toe V absent. (26) Inner metatarsal tubercle rather longand prominent; its length (IMT 1.7 mm) 1.9 times in length of toe I (ITL 2.2 mm).

Skin. (29) Dorsal and lateral parts of head and body: snout, region between eyes, side of head and backsmooth; flank with flat glandular warts. (30) Supratympanic fold distinct, from eye to shoulder. (31) Dorsal parts oflimbs: forelimbs, thigh, shank and tarsus smooth. (32) Ventral parts of head, body and limbs: throat, chest, bellyand thigh smooth. (33) Presence of macroglands: lateroventral ridges present as continous ridges of small glands;large, distinct rounded femoral and oval axillary glands; smaller distinct oval suprabrachial glands.

Coloration. In alcohol: (34) Dorsal and lateral parts of head and body: dorsal parts of head and dorsum andupper part of flanks greyish-brown with black band between the eyes, a black W-shaped pattern in the shoulderregion and a black V in the sacral region; lower part of flanks dark greyish-brown; loreal region blackish-brown;tympanic region including tympanum dark brown with a white band under the tympanum; upper lip dark brownwith white and blackish spots. (35) Dorsal parts of limbs: dorsal part of forelimbs, of thigh, of shank and of footgrey-brown with blackish crossbands; posterior part of thigh blackish with small white spots. (36) Ventral parts ofhead, body and limbs: throat dirty white; margin of throat dark brown with small white spots; chest and belly dirtywhite with dark brown marbling; thigh brown with some white spots; macroglands white.



FIGURE 17. Leptolalax pluvialis, MNHN 1999.5675, holophoront, adult male, SVL 22.0 mm. (A) Dorsal and (B) ventralview.

FIGURE 18. Leptolalax pluvialis, MNHN 1999.5675, holophoront, adult male. (A) Lateral view of head, ventral view of (B)right hand and of (C) right foot. Scale bar = 5 mm (3 + 2 mm).



In life: Iris dark golden above. Upper parts of head and dorsum, flanks and tympanic region grey brown withdark brown pattern, particularly in tympanum and loreal region; upper lip grey-brown with dark brown and whit-ish-grey bands. Upper part of forelimbs ochre, forearms grey-brown and dark brown; hindlimbs with dark brownand grey-brown crossbands; heels ochre; posterior part of thigh grey-brown with ventrally a dark brown zone.Throat transparent light grey-brown with dark brown margin bearing white spots; chest and belly grey with darkgrey marbling; lower part of thigh transparent dark brown with whitish spots. Macroglands whitish grey.

Male secondary sexual characters: (38) Vocal sacs inconspicuous, skin of gular region not modified; inneropenings of vocal sacs present, located laterally and posteriorly in the floor of the mouth, distinct; rounded.

Variation. The two other male protaxonts resemble strongly the holophoront in coloration and morphology.Female sexual characters: Not observed.

Distribution. Known only from onymotope (Fig. 9). Natural history. Three adult male specimens of this species were collected calling during heavy rain, while

sitting on branches and leaves near a small forest stream. They can be distinguished from sympatric Leptolalaxbourreti by their smaller size, ventral coloration and absence of dark spots on flanks. L. bourreti was very abundantin October and November 1997, and was collected in the precise locality of L. pluvialis, but was rather rare in July,when L. pluvialis was breeding.

Etymology. From Latin pluvialis: created by the rain, commanding the rain, rainy.Placement in molecular phylogeny. Leptolalax pluvialis is part of a clade that groups with high support

L. aereus, L. minimus, L. nyx and L. ventripunctatus. The specimens allocated to L. pluvialis form a well-supportedclade which, with high support, is the sister taxon of a poorly supported clade including all the other taxa men-tioned.

Conservation status. L. pluvialis is still only known from the original description. Its conservation statusremains unchanged: Data Deficient.

Leptolalax (Lalos) nyx n. sp.(Fig. 19–20)

Leptolalax bourreti — Bain & Nguyen 2004: 5; Frost et al. 2006: 50. Onymophoront, by present designation: holophoront (holotype), AMNH A163810, adult male, SVL 26.9 mm. Other protaxonts (paratypes): AMNH A163814-6, A163820–1, A163825–7, A163831, IEBR 1121–1122, 1124, 1129–1130,

1134, 1137, 1142–1143, 1145, adult males and adult females. Onymotope: Cao Bo Commune, Vi Xuyen District, Ha Giang Province, Vietnam. Collected by Nguyen Quang Truong & Raoul

H. Bain, 8 May 2000.

Comment. A specimen for which DNA sequences were available from Vietnam was tentatively allocated to L.bourreti (Bain & Nguyen 2004; Frost et al. 2006). It is morphologically quite distinct from the latter species, inparticular in adult size and colour pattern. In the phylogram, bourreti appears polyphyletic. This member of the lin-eage is here described as a new species, which is the sister taxon of an unsupported clade composed of L. aereus, L.minimus and L. ventripunctatus.

Diagnosis. Member of the subgenus Lalos based on the presence of a lateroventral gland and molecular phylo-genetic relationships (Fig. 4), distinguished from other species included in this subgenus by the following combina-tion of characters: moderate-sized species (males 26.7–32.6 mm; females 37.0–41.0 mm) (Table 3) withrudimentary webbing and no fringes on toes; tympanum distinct; lateroventral glandular ridge poorly distinct; der-mal ridges under toes rather poorly distinct; finger tips enlarged; dorsal skin with flat glandular warts (Table 4);dorsum greyish-brown with dark regularly set rounded spots; poorly distinct spots on flanks; ventral side whitishwith brown borders; iris colour not known (Table 5).

Description of the holophoront, adult male AMNH A163810. Size and general aspect: (1) Specimen of rathersmall size (SVL 26.9 mm), body rather robust.

Head. (2) Head of medium size, narrower (HW 9.0 mm) than long (HL 9.5 mm; MN 9.2 mm; MFE 7.6 mm;MBE 4.0 mm), flat. (3) Snout slightly protruding, its length (SL 4.2 mm) as long as horizontal diameter of eye (EL4.2 mm). (4) Canthus rostralis rounded, loreal region concave, virtually vertical. (5) Interorbital space convex, aslarge (IUE 3.1 mm) as upper eyelid (UEW 3.1 mm) but larger than internarial distance (IN 2.6 mm); distance



between front of eyes (IFE 5.1 mm) about three-fifths of the distance between back of eyes (IBE 8.8 mm). (6) Nos-trils rounded, closer to tip of snout (NS 1.6 mm) than to eye (EN 1.8 mm). (8) Tympanum (TYD 1.7 mm) distinct,rounded; less than half eye diameter, tympanum-eye distance (TYE 1.0 mm) two-thirds of its diameter. (11)Tongue large, cordate.

Forelimbs. (12) Forearm short, thin (FLL 7.5 mm), longer than hand (HAL 7.1 mm), not enlarged. (13) Fingerslong and thin (TFL 4.0 mm). (15) Tips of fingers rounded, enlarged. (16) Fingers without dermal fringe.

Hindlimbs. (19) Shanks about four times longer (TL 12.4 mm) than wide (TW 3.1 mm), longer than thigh (FL12.0 mm) but shorter than distance from base of internal metatarsal tubercle to tip of toe IV (FOL 12.6 mm). (20)Toe IV (FTL 7.1) longer than third of distance from base of tarsus to tip of toe IV (TFOL 19.3 mm). (22) Tips oftoes not enlarged. (23) Webbing rudimentary: I 2 – 2 ¼ II 2 – 3 III 2 2/3 – 4 IV 4 – 2 ¾ V (MTTF 4.5 mm; MTFF 4.2mm; TFTF 7.6 mm; FFTF 8.3 mm). (24) Dermal fringe along toe V absent. (26) Inner metatarsal tubercle ratherlong and prominent; its length (IMT 1.5 mm) 1.5 times in length of toe I (ITL 2.3 mm). (27) Tarsal fold absent.

Skin. (29) Dorsal and lateral parts of head and body: snout and side of head smooth; region between eyes, backand flanks with flat glandular warts, quite indistinct on dorsal part. (30) Supratympanic fold prominent, from eye toabove shoulder. (31) Dorsal parts of limbs: forelimbs, thigh, shank and tarsus smooth with sparse indistinct glandu-lar warts. (32) Ventral parts of head, body and limbs: throat, chest, belly and thigh smooth; posterior part of thighwith glandular warts. (33) Presence of macroglands: lateroventral glands present as a short chaplet of poorly dis-tinct, separate glands on both sides of vent; distinct rounded rather small femoral, axillary and suprabrachialglands.

Coloration. In alcohol: (34) Dorsal and lateral parts of head and body: dorsal parts of head and dorsum, andupper part of flanks greyish-brown with a brown W-shaped pattern in the shoulder region continued as a poorlydefined band on middorsum, all back covered with sparsely set white spots (corresponding to warts) with darkbrown outline; lower part of flanks light brown with poorly distinct dark brown spots; loreal region brown; tym-panic region including tympanum beige with a brown line under supratympanic fold covering upper half of tympa-num; upper lip beige. (35) Dorsal parts of limbs: dorsal part of forelimbs, of thigh, of shank and of foot light brownwith dark brown crossbands; posterior part of thigh light brown with darker region near vent and fine brown spots.(36) Ventral parts of head, body and limbs: throat dirty white with isolated dark melanophores; margin of throatbrown with white spots; chest and belly white with brown sides; thigh brown with some white spots; macroglandswhite; webbing brown.

Male secondary sexual characters: (38) Vocal sacs present; indistinct on throat; paired, slit-like openings poste-rior on the floor of the mouth.

Variation. In most of the specimens, the dorsal pattern formed by small dark round spots regularly distributedon back is much more distinct than in the holophoront. The larger sized males and the females exhibit spiny wartswhich are very dense on head and limbs. Female sexual characters: Large creamy white ova visible through skin.Lateroventral glandular ridges are distinct in most specimens formed of individual glands. Femoral, axillary andsuprabrachial glands are relatively small in most individuals but distinct. The femoral glands are a little larger infemales than in males.

Comparison. L. nyx shows usually numerous regularly arranged small dark spots on dorsum, whereas theother species either show relatively uniform dorsal coloration (L. eos, L. minimus) or various larger sized spotsforming triangular or W-shaped patterns. Its white throat, chest and belly distinguish it from L. lateralis, L. nahan-gensis, L. oshanensis, L. pluvialis, L. aereus, L. tuberosus and L. ventripunctatus. The webbing in L. nyx is lessdeveloped than in L. eos and L. pelodytoides but more than in L. aereus, L. alpinus, L. bourreti, L. lateralis, L. liui,L. pluvialis, L. sungi and L. ventripunctatus. The absence of fringes on toes distinguish this species from L. aereus,L. alpinus, L. bourreti, L. eos, L. fuliginosus, L. khasiorum, L. lateralis, L. liui, L. minimus, L. pelodytoides, L.sungi and L. tamdil. With a SVL of 26–30 mm in adult males and 36–42 mm in adult females, L. nyx is among themoderat-sized species of Lalos. L. sungi (males SVL 48–53 mm; females 57–59 mm) is the only known larger spe-cies. In L. bourreti males might reach SVL over 33 mm and females 42–45 mm, whereas in L. eos males (33–35mm) are larger than known for L. nyx but female size (41 mm) is quite similar. L. pluvialis males are smaller (22–23 mm) than known for L. nyx. The minimum size of L. alpinus (24–26 mm), L. khasiorum (25–27 mm), L. liui(23–27 mm) and L. ventripunctatus (24–28 mm) is below the minimum size of L. nyx.

Distribution. Known only from onymotope (Fig. 9).



FIGURE 19. Leptolalax nyx, AMNH A163810, holophoront, adult male, SVL 26.9 mm. (A) Dorsal and (B) ventral view.

FIGURE 20. Leptolalax nyx, AMNH A163810, holophoront, adult male. (A) Lateral view of head, ventral view of (B) righthand and of (C) right foot. Scale bar = 5 mm (3 + 2 mm).



TABLE 10. Measurements and ratios to SVL of adult males and females of Leptolalax nyx from the onymotope.

Males (n=7) Females (n=3)

SVL 27 ± 1.226–30

40 ± 3.136–42

HW/SVL 366 ± 18.6335–382

367 ± 16.7355–386

HL/SVL 388 ± 21.7353–427

381 ± 16.8367–399

MN/SVL 342 ± 9.4329–354

327 ± 17.3316–347

MFE/SVL 269 ± 13.4251–286

255 ± 10.7248–267

MBE/SVL 123 ± 15.8105–147

132 ± 13.2120–146

IFE/SVL 201 ± 17.3178–235

193 ± 1.7192–195

IBE/SVL 322 ± 15303–346

314 ± 13.1302–328

IN/SVL 104 ± 4.996–110

97 ± 5.593–104

EN/SVL 72 ± 5.463–78

82 ± 9.275–92

EL/SVL 164 ± 12.2154–185

143 ± 5.8137–148

TYD/SVL 57 ± 4.948–63

53 ± 0.952–53

TYE/SVL 42 ± 6.335–51

51 ± 0.950–52

FLL/SVL 281 ± 17.6247–304

270 ± 28.6252–303

HAL/SVL 268 ± 9.1258–284

249 ± 24.9222–270

TFL/SVL 152 ± 5.7147–163

133 ± 18.5113–150

TL/SVL 500 ± 20.2461–523

476 ± 41.6439–521

FOL/SVL 487 ± 14.3470–507

466 ± 48414–510

FTL/SVL 291 ± 35.3264–369

265 ± 23.2239–283

IMT/SVL 47 ± 5.840–55

41 ± 2.239–43

ITL/SVL 95 ± 10.284–115

87 ± 8.478–95

MTTF/SVL 177 ± 13.1161–200

171 ± 14.9161–188

MTFF/SVL 162 ± 12.6147–185

170 ± 5.2164–174

TFTF/SVL 296 ± 9.9284–312

291 ± 24.9263–312

TFFTF/SVL 318 ± 7.5308–329

308 ± 23282–326



Natural history. Specimens of L. nyx were collected in April and May along forest streams (up to 5 m wide),in riparian forest floor up to 1.5 m distance from the water. Males were calling in and near the streams. The speciesranges from 800 to 1700 m (Bain & Nguyen 2004).

Etymology. From the Greek , nux, night. Transcribed as nyx. In Greek mythology Nyx was the primordialgoddess of the night, a shadowy figure. The name refers to the nocturnal activity of Leptolalax species. Nomenused as a noun in apposition to the generic substantive.

Placement in molecular phylogeny. Leptolalax nyx is part of a clade that also includes with high support L.minimus, L. aereus, L. ventripunctatus and L. pluvialis. The specimen allocated to L. nyx is recovered with poorsupport as the sister taxon of the clade that includes L. minimus, L. aereus and L. ventripunctatus.

Conservation status. A large series of this species has been collected from the onymotope (Bain & Nguyen2004). According to these authors, the species was locally very abundant, but was observed in forest only. As thereare no data on the extent of occurence, we propose the status Data Deficient for L. nyx.

Leptolalax (Lalos) ventripunctatus Fei, Ye & Li, 1991(Fig. 21–24)

Leptolalax ventripunctatus Fei, Ye & Li in: Fei et al. 1991: 95, 97, 213, 274. Onymophoront, by original designation: holophoront (holotype), CIB 890063, adult male, SVL 27.0 mm (Fei et al. 1992: 51).

Collected by Luo Jiarui, 17 May 1989.Onymotope: Mengla (21°29’ N, 101°15’ E; alt. 850 m), Yunnan Province, China.

Material examined. Laos. Phongsaly Province, Phongsaly water supply river (21°40’ N, 102°06’ E; 1000 m):MNHN 2005.0111–0116, adult males; MNHN 2006.2576, 1 tadpole (DNA voucher), coll. Annemarie Ohler, 31January 2005; — Nathen (21°24’N, 101°58’ E; 600 m): MNHN 2005.0137–0140, adult males, MNHN 2006.2577,1 tadpole (DNA voucher), coll. Annemarie Ohler, 9 February 2005; — Long Nai (21°14' N, 101°56' E; 800 m),MNHN 2004.0270–0273, MNHN 2005.0117–0136, adult males, coll. Annemarie Ohler, 14 and 15 July 2004 and 4February 2005; — Nam Ou (22°03’ N, 102°16’ E; 520 m), MNHN 2005.0142–0144, adult males, MNHN2006.2578–2588, 11 tadpoles, coll. Annemarie Ohler, 13 February 2005; — Houey Phihet (21°60’ N, 102° 17’ E;500 m): MNHN 2005.0145, adult male, coll. Annemarie Ohler, 16 February 2005; Vietnam. Vinh Yen District,Tam Dao (21°27’ N, 105°38’ E): MVZ 223641–42, coll. Theodore J. Papenfuss, 22 April 1996.

Comment. This species was described from extreme southern Yunnan, near the Lao border. We had the oppor-tunity to do fieldwork on the Lao side and discovered this small-sized Leptolalax which is sympatric with the newspecies L. eos described above. Specimens allocated to the present species resemble the onymophoront in morphol-ogy and are therefore assigned to ventripunctatus (no molecular data exist so far from the Yunnan locality). Thecollection locality in Laos is less than a hundred kilometres from the onymotope. Most of the Lao specimens wereincluded in the molecular analysis and show great overall genetic similarity. They form a clade with specimensfrom Tam Dao (MVZ 223641–42) which are genetically very similar. In consequence, these specimens areincluded in L. ventripunctatus.

Diagnosis. (Based on specimens from Laos, not on onomophoront). Member of the subgenus Lalos based onthe presence of lateroventral glands and molecular phylogenetic relationships (Fig. 4), distinguished from otherspecies included in this subgenus by the following combination of characters: rather small-sized species (males23.7–27.7 mm; females 31.5–35.0 mm) (Table 3) with rudimentary webbing and no fringes on toes; tympanum dis-tinct; dermal ridges under toes distinct; finger tips slightly enlarged; dorsal skin with indistinct glandular warts(Table 4); dorsum grey brown with spots, rather large spots on flanks; ventral side with distinct dark spots on chestand belly; iris copper above, grey brown below (Table 5).

Description of the adult male MNHN 2004.0270. Size and general aspect: (1) Specimen of rather small size(SVL 25.2 mm), body moderately slender.

Head. (2) Head of medium size, narrower (HW 8.8 mm) than long (HL 10.0 mm; MN 8.4 mm; MFE 6.7 mm;MBE 3.3 mm), rather flat. (3) Snout slightly protruding, its length (SL 4.4 mm) longer than horizontal diameter ofeye (EL 3.6 mm). (4) Canthus rostralis distinct, rounded, loreal region concave, vertical. (5) Interorbital space flat,slightly narrower (IUE 2.9 mm) than upper eyelid (UEW 3.1 mm) but larger than internarial distance (IN 2.7 mm);distance between front of eyes (IFE 5.2 mm) about two-thirds of the distance between back of eyes (IBE 7.8 mm).



(6) Nostrils rounded, closer to eye (EN 1.9 mm) than to tip of snout (NS 2.1 mm). (8) Tympanum (TYD 1.9 mm)distinct, rounded; about half eye diameter, tympanum-eye distance (TYE 0.9 mm) half the diameter of tympanum.(11) Tongue moderate, cordate.

FIGURE 21. Leptolalax ventripunctatus, MNHN 2004.0270, adult male, SVL 25.2 mm. (A) Dorsal and (B) ventral view.

Forelimbs. (12) Forearm rather short, thin (FLL 7.3 mm), longer than hand (HAL 7.0 mm), not enlarged. (13)Fingers I, II and IV rather short and rather strong; finger III long and rather strong (TFL 3.9 mm). (15) Tip of fin-gers rounded, slightly enlarged. (16) Fingers without dermal fringe.

Hindlimbs. (19) Shanks about four times longer (TL 12.0 mm) than wide (TW 3.2 mm), shorter than thigh (FL12.2 mm) and than distance from base of internal metatarsal tubercle to tip of toe IV (FOL 12.1 mm); heel reachingmid-eye when limbs folded forward. (20) Toe IV (FTL 6.4 mm) about one-third of the distance from base of tarsusto tip of toe IV (TFOL 19.4 mm). (22) Tip of toes rounded, slightly enlarged. (23) Webbing rudimentary, toes with-out fringes (MTTF 4.9 mm; MTFF 4.9 mm; TFTF 7.3 mm; FFTF 7.7 mm). (24) Dermal fringe along toe V absent.(26) Inner metatarsal tubercle short and very prominent (conical); its length (IMT 1.3 mm) 2.1 times in length oftoe I (ITL 2.7 mm). (27) Tarsal fold absent.

Skin. (29) Dorsal and lateral parts of head and body: snout, region between eyes and side of head smooth; backwith indistinct glandular warts; flanks with glandular warts. (30) Supratympanic fold distinct, from eye to aboveshoulder. (31) Dorsal parts of limbs: forelimbs with few indistinct glandular warts; thigh, shank and tarsus smooth.(32) Ventral parts of head, body and limbs: throat, chest and belly smooth; thigh with glandular warts. (33) Pres-ence of macroglands: lateroventral glands present as series of glands forming continous glandular ridge; ratherlarge, distinct rounded or oval femoral and axillary glands, and oval suprabrachial glands.



FIGURE 22. Leptolalax ventripunctatus, MNHN 2004.0270, adult male. (A) Lateral view of head, ventral view of (B) rightfoot and of (C) right hand. Scale bar = 5 mm (3 + 2 mm).

Coloration. In alcohol: (34) Dorsal and lateral parts of head and body: dorsal parts of head and dorsum greyish-brown with whitish zones and two black spots between eyes; flanks grey brown with rather large blackish spots;loreal region light brown with distinct dark brown spot; tympanic region including tympanum light brown withblackish line at supratympanic fold; upper lip light brown with brown bands. (35) Dorsal parts of limbs: dorsal partof forelimbs beige with brown crossbands on lower part; dorsal part of thigh, of shank and of foot beige with browncrossbands; posterior part of thigh brown with a few small white spots. (36) Ventral parts of head, body and limbs:throat whitish with traces of brown; margin of throat light brown with whitish spots; chest whitish with few brownspots; belly whitish with dense brown spots; thigh light brown with whitish spots; webbing brown; macroglandswhite.

In life (Fig. 23E–F): Iris copper in upper third, grey brown in lower part. Dorsal parts of head, of back and ofupper part of flanks grey brown with darker spots and black spots between eyes; lower part of flanks greyish withblack spots; loreal and tympanic region brown with black canthal and supratympanic band; tympanum brown;upper lip brown with blackish bands; upper part of forelimb orange; lower part greyish brown with blackish bands;thigh, shank and foot greyish brown with black crossbands; posterior part of thigh greyish brown with dark brownbands; throat light grey, transparent; border of throat grey with white spots; chest and belly creamy white withbrown spots; ventral part of thigh dark grey brown with white dots; webbing grey brown. Macroglands white.

Male secondary sexual characters: (38) Vocal sacs present; indistinct on throat; pair, slit-like openings poste-rior on mouth floor.



FIGURE 23. Specimens of Leptolalax in life. (A) L. aereus, MNHN 2005.0235, adult male, SVL 25.8 mm; (B) L. aereus,ZFMK 71346, adult male in its natural habitat, SVL 29.8 mm; (C) L. aereus, ZFMK 71347, adult male, SVL 29.8 mm; (D) L.aereus, ZFMK 79861, subadult male, SVL 23.4 mm; (E) L. ventripunctatus, MNHN 2005.121, adult male, SVL 24.6 mm; (F)L. ventripunctatus, adult male, not collected, in dorsal view.

Variation. Specimens are rather homogenous in colour pattern and show only minor variation in extent of darkpattern on ventral side and presence of spots on dorsum and lateral side. The specimens also show some variationin presence of warts and skin ridges. Female sexual characters: oviduct convoluted, translucent; ovary with smallcreamy whitish oocytes.

The lateroventral glandular ridges are continous in all specimens. There is some variation in size and shape offemoral and axillary glands that are smaller in some specimens and more or less oval or round in shape.

Tadpole. The external morphology is based on a DNA voucher specimen in stage 33 (MNHN 2006.2576; BL16.1 mm). The description of the tail which is missing in the DNA voucher is based on a specimen in stage 36(MNHN 2006.2578).

In dorsal view (Fig. 24A), body elliptical elongate. In lateral view (Fig. 24B), BW 134 % of BH. Eyes moder-ately sized, ED 9.6 % of BL, bulging. Nares moderately sized, RN 40 % of NP; NN 77 % of PP. Spiracle is a short



tube, small, SS 50 % of BL; opening crescent-shaped, situated on an axis just below apex of myotomes of caudalmuscle. Tail long; tail musculature, TMH 82 % of BH, TMW 62 % BW, slightly bulging on less than the proximalthird, almost reaching tail tip. Upper fin, SU 108 % of BL, almost nonexistent in its proximal third then increasingrather abruptly; lower fin convex, following largely the caudal muscle; point of maximum height of tail locatedbefore its proximal third. Anal tube moderately sized. Lateral line system as in L. bourreti but with an additionalline running vertically on the lateral sac from the ventral line until the beginning of the ventral part. Glands presentbut faintly visible.

FIGURE 24. Tadpole of Leptolalax ventripunctatus. (A) Dorsal and (B) lateral views of a specimen in stage 33 (MNHN2006.2576), scale bar = 10 mm; (C) oral disc of a specimen in stage 29 artificially expanded (MNHN 2006.2577), scale bar = 1mm. Tip of tail cut for tissue sampling.



Oral disc (Fig. 24C) moderately sized, ODW 28 % of BL and 52 % of BW. Marginal papillae of lower labiumsmaller than those of upper labium and more or less round to short, cylindrical, decreasing in size towards centrethen increasing again at the extremity of indentation with four larger papillae; five submarginal papillae on left sideand two on right side on lower labium at commissure and in front of keratodont rows P1 and P2. No denticulatepapillae. KRF 1:4+4/3+3:1, A4 > A5 > A3 > A2 > A1; P2 = P1 > P3 > P4; A1 about half of P4. Details of kerato-donts not studied. Jaw sheaths black; upper sheath a large arch with a median convexity formed by the threemedian serrations, the median one bigger and stronger than the others; lower sheath hidden, arranged deeply underthe upper jaw sheath.

Coloration in life: Tadpole MNHN 2006.2578 reddish dorsally with indistinct spots; lateral body light greywith distinct grey spots; belly greyish white. Tadpole MNHN 2006.2577 uniformly greyish brown, its tail beigewith dark spots; belly bluish white; oral region whitish grey; iris golden.

Coloration in preservative: upper part of body sandy orange, the colour fading from before eyes to snout.Flanks (lymphatic sacs) grey ochre. Ventral side of body dirty white. Caudal muscle from sandy for the upper partto ivory for the lower part in gradation; small light brown spots more numerous on the upper part than on the lowerpart, giving a mottled aspect. Fins translucent with the same small spots as those on the caudal muscle but formingsmall blotches isolated from each other on the upper fin; lower fin spotless. Rims of nares white.

Tadpole variation is described on the basis of 12 other specimens, except when noted (MNHN 2006.2577,2006.2579–2588, in stages 25–36, TOL 31.0–55.9 mm and BL 9.0–16.1 mm). Only the ratio values different fromthose of the described specimen are given: BW 129–144 % of BH; ED 8.4–10.7 % of BL; RN 35–58 % NP; NN70–82 % of PP; SS 42–53 % of BL; TAL 212–251 % of BL (n = 11); TMH 63–85 % of BH; TMH 53–70 % ofMTH (n = 11); TMW 48–61 % of BW; UF 28–36 % of MTH (n = 11); SU 92–116 % of BL; LF 22–29 % of MTH(n = 11); MTH 115–126 % of BH (n = 11); ODW 25–32 % of BL (n = 11); ODW 43–49 % of BW (n = 11). Themore the tadpoles are in advanced stages, the more their upper fin begins close to body end. The upper fin increasesat the first quarter and the lower fin can be straight and so increasing in height anteroposteriorly in a few speci-mens. The KRF varies from 1:2+2/2+2:1 (stage 25) to 1:4+4/3+3:1 (stages 29–36) through the intermediate KRF1:3+3/2+2:1 (stages 25–34) and 1:3+3/3+3:1 (stages 27–28). The coloration of the tail is more extended on most ofthe other specimens: proximal half of caudal muscle tan, this coloration grouping in blotches on the distal half withthe ground colour beige, similar blotches on the distal third of the lower fin, upper fin smoked with tan pigments.

The lateral spots in the tadpole series described herein are less pronounced than in those figured by Fei et al.(2009). We assume that this might be due to the strong contrast between black and white in the drawing of Fei et al.(2009).

Distribution. China: Yunnan Province (Mengla County); Laos: Phongsaly Province (Phongsaly District);Vietnam: Vinh Phuc Province (Vinh Yen District) (Fig. 9).

Natural history. Specimens were found (July 2004 and January–February 2005) in forests at a distance up to1 m from small rivulets (Fig. 36A). Most specimens were very close to the water at a distance of 0.2 to 0.5 m. Theywere sitting either on the ground (stones, sand, earth) or above the ground on leaves or rocks, usually covered bylow vegetation. Only a few specimens were sitting in the water. Most of the specimens were in breeding condition. The tadpoles were collected in a puddle of 40 × 30 cm along a stream (but connected to the stream) with a weakcurrent. They were usually motionless on the bottom, hidden within the gravel. Another specimen was collected ina stream of a width of 10 m and a maximum depth of 50 cm, at a depth of 10 cm. The bottom was covered of stonesand dead leaves under which the tadpoles were hidden. In these streams, adults of Limnonectes kuhlii and Polype-dates “leucomystax” were also observed.

Placement in molecular phylogeny. Leptolalax ventripunctatus is part of a highly supported clade that groupsL. minimus, L. aereus, L. pluvialis and L. nyx. The specimens allocated to L. ventripunctatus form a well supportedclade which, with no support, is sister to the clade that goups L. minimus and L. aereus.

Conservation status. This work adds more data on distribution and extent of occurence of this species. It isrelatively abundant in convenient habitat, within its relatively small range. It might encounter threats by habitatloss through shift and burning agriculture. We propose therefore to change the status of L. ventripunctatus fromData Deficient to Near Threatened.



Leptolalax (Lalos) aereus Rowley, Stuart, Richards, Phimmachak & Sivongxay, 2010(Fig. 23, 25–29)

Leptolalax pelodytoides —Ziegler 2002: 47–50.Leptolalax sp. —Ziegler et al. 2004: 22–23.Leptolalax cf. pelodytoides —Ziegler et al. 2006: 261. — Hendrix et al. 2008: 27. — Ziegler & Vu 2009: 121.Leptolalax aereus Rowley, Stuart, Richards, Phimmachak & Sivongxay, 2010c: 36.Onymophoront, by original designation: holophoront (holotype) SAMA R64236, adult male, collected on 23 November 2008 by B. L.

Stuart, S. J. Richards, S. Phimmachak and N. Sivongxay.Onymotope: Rocky stream in semi-evergreen forest in Sepon Mining Tenement (16.96317º N, 106.0466º E, 326 m), Vilabouli District,

Savannakhet Province, Laos.Other protaxonts (paratypes): NCSM 76038–76057, NCSM 76061–76071, SAMA R64234–64235, SAMA R64238–63242. Collec-

tion data see Rowley et al. (2010c).

Specimens referred to the form 1: Vietnam. Quang Binh Province, Phong Nha – Ke Bang, nearby mountain stream (400–500 m a.s.l.):ZMFK 71346, adult male, SVL 29.8 mm, collected by Thomas Ziegler, 3 September 1998; ZFMK 71341, 4 tadpoles, coll. T.Ziegler, 3 September 1998; —Phong Nha – Ke Bang NP: small stream above a “Waterfall”, 30 m next to Route 20 (km 6) nearborder: ZFMK 86362, juvenile, 17 June 2006; — stream downhill from Ho Chi Minh Highway (km 54) in U Bo (530 m a.s.l.):RH06149 (PNNP), juvenile, 30 July 2006; — pond next to Ho Chi Minh Highway at km 51: ZFMK 87012–16, 5 tadpoles, 20June 2006; — stream downhill from Ho Chi Minh Highway (km 54) in U Bo (500 m a.s.l.): ZFMK 87017–19, 3 tadpoles, 30 July2006.

Specimens referred to form 2. Vietnam. Ha Tinh Province: ZMFK 79861 (TZ 819), subadult male, 3 September 1997, coll. T. Ziegler,locality at 360 m a.s.l; — Quang Binh Province: 50 m below a small mountain stream in Cha Noi: ZFMK 86393, juvenile, 5August 2006; — river bed below the mountain stream: RH06166 (PNNP), juvenile, 6 August 2006; — river bed below the moun-tain stream in Cha Noi: RH06167 (VNUH), adult male, 7 August 2006; ZFMK 86394, adult female, 5 August 2006; — mountainstream in Cha Noi: RH06172 (VNUH), imago, 7 August 2006; ZFMK 87009, 9 tadpoles, 5 August 2006; ZFMK 87010, 6 tad-poles, 7 August 2006.

Specimens referred to L. aereus, without genetic identification. Vietnam. Ha Tinh Province: MNHN 2010.1040–1045, 6 tadpoles, 11July 1997, coll. T. Ziegler; — locality at 160 m a.s.l; MNHN 2010.1037–1039, 3 tadpoles, 24 August 1997; — 160 m a.s.l., coll.T. Ziegler; — Quang Binh Province: Phong Nha – Ke Bang, nearby mountain stream (400–500 m a.s.l.): ZFMK 71347, adultmale, September 1998, coll. T. Ziegler; — Phong Nha – Ke Bang, Cha Noi: MNHN 2005.0235, young female, 1 July 2004, coll.T. Ziegler; — mountain stream in Cha Noi: ZFMK 87011, 4 tadpoles, 10 August 2006; — Phong Nha – Ke Bang National Park:PNNP0022, 0031, 0048, 0040, one without label, 4 males, 1 female, coll. Dang Ngoc Kien et al. — Nghe An Province: TuongDuong District: Nga My Commune: Pu Huong Nature Reserve (19°21’385N, 104°52’552E, 380 m a.s.l.): PHO8 007, PHO8 008,2 males, 26 March 2008, coll. Hoang Ngoc Thao and Dau Quang Vinh.

FIGURE 25. Leptolalax aereus, ZMFK 71346, adult male, SVL 29.8 mm. (A) Dorsal and (B) ventral view. Right shank andfoot were taken off for tissue sampling.



Comment. The specimens studied by us and allocated to L. aereus form a well supported clade with two subclades,with good support but weak differentiation between these sister clades. In the following text the specimens,depending on their situation in the subclades, are allocated to “form 1” or “form 2”. Note that according to prelim-inary data (see molecular results above), the original series of L. aereus forms a third genetic lineage. The adultsand juveniles belonging to the two forms included here show a low differentiation in morphology and coloration,but the tadpoles can be distinguished by morphology (see details below). We chose to describe these two tadpoleforms independently as “form 1” and “form 2” because it is uncertain whether L. aereus as here defined might infact be a species complex.

Diagnosis. Member of the subgenus Lalos based on the presence of lateroventral glands and molecular phylo-genetic relationships (Fig. 4), distinguished from other species included in this subgenus by the following combina-tion of characters: moderate–sized species (males 29.8 mm) (Table 3) with rudimentary webbing and narrowfringes on toes; tympanum distinct; dermal ridges under toes distinct; finger tips enlarged; dorsal skin granular(Table 4); dorsal pattern showing dark markings and light speckling; few small black and white spots on flanks,ventral side whitish with indistinct brown vermiculation; iris yellowish orange (Table 5).

Description of the reference specimen ZMFK 71346. Size and general aspect: (1) Specimen of rather largesize (SVL 29.8 mm), body rather stout.

Head. (2) Head of medium size, narrower (HW 10.0 mm) than long (HL 11.5 mm; MN 10.0 mm; MFE 7.9mm; MBE 3.1 mm), flat. (3) Snout protruding, its length (SL 4.6 mm) shorter than horizontal diameter of eye (EL4.9 mm). (4) Canthus rostralis rounded, loreal region concave, rather vertical. (5) Interorbital space flat, larger(IUE 3.6 mm) than upper eyelid (UEW 3.4 mm) and than internarial distance (IN 2.9 mm); distance between frontof eyes (IFE 5.2 mm) about half of the distance between back of eyes (IBE 9.2 mm). (6) Nostrils oval, as close toeye (EN 2.1 mm) as to tip of snout (NS 2.1 mm). (8) Tympanum (TYD 2.3 mm) distinct, oval, horizontal; abouthalf of eye diameter, tympanum-eye distance (TYE 1.0 mm) less than half its diameter. (11) Tongue large, cordate,emarginate.

FIGURE 26. Leptolalax aereus, ZMFK 71346, adult male. (A) Lateral view of head, ventral view of (B) right foot and of (C)right hand. Scale bar = 5 mm (3 + 2 mm).



Forelimbs. (12) Forearm rather long, thin (FLL 8.5 mm), longer than hand (HAL 8.1 mm), not enlarged. (13)Fingers long and thin (TFL 4.6 mm). (15) Tips of fingers rounded, slightly enlarged. (16) Finger II with narrowdermal fringe.

Hindlimbs. (19) Shanks about three times longer (TL 14.5 mm) than wide (TW 4.02 mm), longer than thigh(FL 13.5 mm) and than distance from base of internal metatarsal tubercle to tip of toe IV (FOL 13.9 mm). (20) ToeIV (FTL 7.5 mm) about one-third of distance from base of tarsus to tip of toe IV (TFOL 22.1 mm). (22) Tips of toesrounded, not enlarged. (23) Webbing present, rudimentary, narrow fringes on toes: I 2 – 2 II 1 ½ – 3 III 2 ½ – 4IV 4 ¼ – 2 ½ V (MTTF 5.3 mm; MTFF 5.1 mm; TFTF 8.0 mm; FFTF 9.0 mm). (24) Fringe along toes narrow. (26)Inner metatarsal tubercle rather long, prominent; its length (IMT 1.7 mm) 1.7 times in length of toe I (ITL 2.9 mm).

Skin. (29) Dorsal and lateral parts of head and body: snout, region between eyes, side of head and back granu-lar; flanks with small glandular warts, more densely set on lower part of flanks. (30) Supratympanic fold distinct,from eye to above upper forelimb. (31) Dorsal parts of limbs: smooth with few granules. (32) Ventral parts of head,body and limbs: throat, chest and belly smooth; thigh with a few glandular warts. (33) Presence of macroglands:lateroventral gland present, formed by a few relatively large glands positioned in line; oval, relatively large-sizedfemoral and axillary glands and round smaller suprabrachial glands.

FIGURE 27. Leptolalax aereus, ZMFK 71346, adult male, SVL 29.8 mm. Life photograph.

Coloration. In alcohol: (34) Dorsal and lateral parts of head and body: Grey brown with a dark brown interor-bital band and a W-like spot and light grey spots on shoulders; two dark brown spots on rear part of back; lowerpart of flanks light greyish brown with white and dark brown spots; loreal region light grey brown; tympanic regionwhitish grey with blackish line below tympanic fold; tympanum light brown; upper lip greyish white with brownspots. (35) Dorsal parts of limbs: light brown with darker brown crossbands; posterior part of thigh brown withwhite dots corresponding to glands. (36) Ventral parts of head, body and limbs: throat yellowish white; margin ofthroat brown with white dots; chest and belly white with indistinct brown vermiculation, mainly laterally; thighbrown with white dots corresponding to glands; webbing brown; macroglands white.

In life (form 1). Based both on specimens collected by TZ and RH. Iris of eye gold copper. Dorsum greyishbrown to reddish brown. Upper part of forearms lighter, yellowish to orange-brown. Lower part of forelimbs andhindlimbs completely with dark transversal bars. Upper lip with dark bars, with most prominent being under eye. A



more or less distinct dark stripe underlining tympanic fold, tympanum in part dark. Suprabrachial gland cream toyellow, may be surrounded by dark. Dark triangle-shaped pattern between eyes, pointing backwards, followed byan equally backwards-pointing dark V-shaped structure that may be surrounded by cream. Another dark transversalelement visible in rear part of dorsum, as well as dark blotches towards flanks. Flanks lighter with some lightblotches, more or less distinctly surrounded by dark. Venter light, with small light blotches, some surrounded bydark in jaw region. Underside of fore and hindlimbs somewhat darker with small light blotches in rear region.

Male secondary sexual characters: (38) Vocal sacs externally not differentiated; openings present, rounded,positioned posterior on side mouth floor.

Variation. Female sexual characters of forms 1 and 2 of this species are unknown as no adult female was col-lected although female morphology is known from the accounts of Rowley et al. (2010c). Variation in the follow-ing only refers to our own data (of forms 1 to 2) and not to the series studied by Rowley et al. (2010c) in theoriginal description. Young and adults of Leptolalax aereus were collected from several localities at several dates.They are not of the same ontogenetic stage as some have reached adult stage whereas the others are only subadultsor young. They differ slightly in dorsal and ventral coloration when preserved in alcohol: specimens of form 1,which includes the described reference specimen, are brown grey on back and light beige on ventral side withdarker edges; the glands are white. Specimens of form 2 are light brown grey on back and creamy white on belly,with greyish beige on sides. Another sample collected and deposited in Phong Nha-Ke Bang NP is brown orangeon the dorsum and whitish on the ventral side. Specimens of a series from Nghe An Province are dark brown withwhite belly. The difference between these series is more in the shade of coloration than in the colour pattern as inboth form 1 and 2, specimens have a W-shaped pattern in shoulder region and an inverted V-shaped pattern on pos-terior part of back. In some specimens of either form a dark interoccular band is visible. All specimens have dis-tinct crossbands on forearms, shank, thigh and foot. All show numerous dark spots on flanks. Variation is not onlybetween the two forms but also within the forms. Among specimens of form 2, specimen ZMFK 79861 is unique inhaving broad greyish white outlines in interorbital and shoulder spots. Form 2 has a generally very distinct colourpattern including W-shaped shoulder spots. In form 1, the two adult frogs (ZFMK 71346–71347) and two justmetamorphosed young have a more uniformly coloured dorsum. Ventral colour is uniformly whitish in all speci-mens but in the two adults which have slightly brownish throats and sides of belly covered with rather dense brownmottling.

Description of the tadpole of Leptolalax aereus form 1. The results of the molecular analysis were alsoreflected in the allocation of tadpoles to the corresponding adults. The comparison of the uncorrected p-distances ofDNA vouchers of the lineage described above as L. aereus showed a similarity of 99.1–99.4 % between tadpoles(ZFMK 87009, ZFMK 87010) and syntopic adults (VNUH uncatalogued: field number RH06167, RH06172) col-lected in Cha Noi, Quang Binh Province and the adult specimen (ZMFK 79861; field number TZ 819) from KyAnh – Ke Go, Ha Tinh Province. The tadpoles (ZFMK 87012, ZFMK 87016–87017, ZFMK 87019) from U Bomatched the syntopic juvenile (PNNP uncatalogued: field number RH06149) and the subadult and adult specimens(ZFMK 86362, ZFMK 71346) from nearby areas by a congruence of 100 %. However, the genetic comparisonrevealed constant and notable differences of 2.4 % between sampled specimens from the northern area (Cha Noi,PNKB, Quang Binh Province and Ky Anh – Ke Go, Ha Tinh Province) and the eastern and southeastern area ofPhong Nha − Ke Bang (Waterfall and U Bo).

The external morphology is based on a specimen in stage 34 (from batch ZFMK 87012; includes field numbersRH06019, RH06035; TOL 59.3 mm and BL 16.9 mm).

In dorsal view (Fig. 28A), body elliptical elongate. In lateral view, (Fig. 28B), BW 138 % of BH, highest in themiddle of the abdomen. Eyes moderately sized, ED 9.6 % of BL, bulging. Nares elliptical, moderately sized, RN53 % of NP; NN 72 % of PP. Spiracle flattened tubular, positioned laterally, SS 52 % of BL, oriented slightly moreposteriorly than posterodorsally; its opening directed posterodorsally, situated on an axis which should go throughthe apex of myotomes of caudal muscle. Tail long, TAL 249 % of BL; tail musculature, TMH 83 % of BH and 59% of MTH, TMW 63 % BW, the lower part slightly bulging in the proximal third. UF 33 % of MTH, LF 26 % ofMTH, SU 113 % of BL, upper fin almost nonexistent in its first third then increasing rather abruptly; point of max-imum height of tail located at its first third, MTH 142 % of BH. Anal tube moderately sized. Lateral line identicalto L. bourreti.

Oral disc (Fig. 28C) moderately sized, ODW 21 % of BL and 40 % of BW. Marginal papillae fine, conical andpointed; three submarginal papillae on each side in front of the extremities of the rows P2 to P4. No denticulate



papillae. KRF 1:4+4/3+2:1, A4 > A5 > A3 > A2 >> A1; P2 > P1 = P3 > P4; A1 about one-third of P4, P4 morethan half of P3. Details of keratodonts not studied. Jaw sheaths black; lower sheath hidden, deeply fitted under theupper jaw sheath.

FIGURE 28. Tadpole of Leptolalax aereus form 1. (A) Dorsal and (B) lateral views of a tadpole in stage 34 (ZFMK 87012),scale bar = 10 mm; (C) oral disc of a specimen in stage 31 (ZFMK 87015), scale bar = 1 mm.

The coloration in life of L. aereus form 1 extensively corresponds to that of L. aereus form 2 but golden pig-mentation is more distinct in form 1. In preservative, upper part of body uniformly brown. Flank (occupied by thelymphatic sacs) from brown in the upper part to grey in the lower part in gradation. Ventral side dirty white. Eachlobe of nares tinted in grey in its central part. Caudal muscle beige on slightly more than its half, the rest grey, cov-ered with densely arranged small dots of melanophores leaving some small immaculate areas; these small dotsmore numerous in the distal part of the muscle; the whole muscle speckled, but the spots more distinct in the prox-imal half. Fins translucent grey with the same spots and dots than on the caudal muscle, more dense on the upperfin, beginning from the distal half on lower fin and from the proximal third on upper fin.

The description of tadpole variation is based on 10 other specimens, except when noted (stages 25–38, TOL22.6–47.5 mm (n = 5) and BL 10.8–18.5 mm). Only the ratio values different from those of the described specimenare given: BW 119–136 % of BH; ED 8.2–10.5 % of BL; RN 42–55 % of NP; NN 61–75 % of PP; SS 46–51 % ofBL; TAL 193–256 % of BL (n = 5); TMH 60–85 % of BH; TMH 61–69 % of MTH (n = 8); TMW 60–93 % of BW



(n = 9); UF 30–32 % of MTH (n = 4); SU 109–143 % of BL (n = 4); LF 23–27 % of MTH (n = 4); MTH 87–129 %of BH (n = 8); ODW 18–26 % of BL; ODW 35–45 % of BW. More the tadpoles are in advanced stages, more theirupper fins begin close to body end. The KRF vary from 1:3+3/2+2:1 to 1:5+5/3+3:1 without obvious addition pat-tern of keratodont rows through ontogeny (1:3+3/2+2:1 one stage 26; 1:3+3/3+3:1 one stage 31; 1:4+4/2+3:1 onestage 25 and one stage 34; 1:4+4/3+3:1 one stage 28, one stage 29, one stage 33, and two stages 35; 1:5+5/2+2:1one stage 36; 1:5+5/3+3:1 one stage 38); certain specimens have denticulate papillae.

Description of the tadpole of Leptolalax aereus form 2. The external morphology is based on a specimen instage 26 (from batch ZFMK 87009 includes field numbers RH06161 and RH06164; TOL 43.7 mm and BL 12.9mm).

In dorsal view (Fig. 29A), body elliptical elongate, widest just after the spiracle. In lateral view (Fig. 29B), BW137 % of BH. Eyes bulging, ED 8.9 % of BL. Nares elliptical, moderately-sized, RN 50 % of NP; NN 77 % of PP.Spiracle flattened rectangular, positioned laterally, SS 53 % of BL, oriented at an angle of 30° with the horizontal;its opening directed posteriorly, situated on an axis which should go through the apex of myotomes of the caudalmuscle. Tail long, TAL 235 % of BL; tail musculature, TMH 72 % of BH and 59 % of MTH, TMW 57 % of BW,the lower part slightly bulging in its proximal third. Upper fin, UF 31 % of MTH, SU 104 % of BL, almost non-existent on its first third then increasing rather abruptly; lower fin, LF 23 % of MTH; MTH 122 % of BH. Analtube moderately sized. Lateral line system well visible, as in L. bourreti.

Oral disc (Fig. 29C), ODW 23 % of BL and 48 % of BW. Marginal papillae of the upper labium with largerbases than those of the lower labium but finer than those of the four other tadpoles described herein; three submar-ginal papillae on each side in front of the extremities of the rows P1 to P3. No denticulate papillae. KRF 1:4+4/3+3:1, A3 the longest, A3 and A4 subequal > A5 > A2 > A1; P1 > P2 > P3 > P4; A1 about half of P4. Details ofkeratodonts not studied. Jaw sheaths dark brown with black serrations; lower sheath hidden, deeply fitted under theupper jaw sheath.

Coloration in life: body mottled brown with golden and dark brown pigments. Tail with dark brown blotcheswhich are getting smaller towards the tip of the tail. Upper and lower fins and ventral side of body greyish translu-cent. Nares conspicuous and grey coloured. Inner gills gleam reddish through skin. Intestinal loop visible.

Colour in preservative: body and tail dorsally and laterally greyish to light brown. Distinct speckled pattern atend of body and at basis of tail. Ventral body yellowish white and fins translucent. Intestinal coils visible throughskin.

The variation is based on 10 other specimens, except where noted (stages 26–41, TOL 29.4–47.8 mm (n = 9)and BL 9.1–15.1 mm). Only the ratio values different from those of the described specimen are given: BW 124–149 % of BH; ED 9.4–12.4 % of BL; RN 44–64 % of NP; NN 59–87 % of PP; SS 50–55 % of BL; TAL 201–229% of BL (n = 9); TMH 63–98 % of BH; TMH 54–61 % of MTH; TMW 52–67 % of BW; UF 29–32 % of MTH (n= 9); SU 97–124 % of BL; LF 24–26 % of MTH (n = 9); MTH 106–160 % of BH; ODW 24–27 % of BL; ODW46–60 % of BW. Maximum body width can be reached in middle of abdomen. Spiracle opening direction at anangle of 30° with horizontal. More the tadpole is in an advanced stage, more the upper fin begins close to body end.The posterolateral glands not visible on all specimens. The KRF vary from 1:3+3/2+2:1 to 1:4+4/3+3:1 withoutclear ontogenetically driven pattern. The early stages (stage 25) have much smaller dots, sometimes even absent.

Distribution. Vietnam: Based on our data: Ha Tinh Province, Nghe An Province, Quang Binh Province (Fig.9). Laos (based on Rowley et al. 2010c): Savannakhet Province.

Natural history. One adult assigned to form 1 was collected at night of 17 June 2006 at a small stream of thesecondary forest (135 m a.s.l.) next to Route 20 near the border of the NP, sitting on a larger stone along a gravelbank. A juvenile specimen with an almost resorbed tail was found at 09:30 p.m. on 30 July 2006 (24°C, 86 % rela-tive humidity) on a small bush inside a primary forest stream in U Bo (530 m a.s.l.). Male specimens ZFMK71346–71347 (form 1) were collected at night in Phong Nha – Ke Bang NP. It was in early September, at the begin-ning of the rainy season. Specimens were on low vegetation and on a stone in the vicinity of a nearly completelydried stream bed in primary forest. In their stomachs and guts were a few worms (nematodes; presumably endopar-asites), and in specimen ZFMK 71347 we found remains of a 5 mm large spider.

The tadpoles of form 1 in different developmental stages (stages 29–37) were regularly sighted syntopicallywith Leptobrachium chapaense and Limnonectes “kuhlii” inside a small stream (water temperature: 22–25°C, pH5.0–6.0) without any vegetation in U Bo area of the Phong Nha - Ke Bang NP (480–530 m a.s.l.). All larvae werefound exposed on sparse substrate like the tadpoles from Cha Noi.

The subadult male from Ha Tinh Province (form 2) was collected at the beginning of the rainy season on a



steep primary lowland forest hillside (Fig. 36C). The specimen was found in the afternoon between leaf litter belowa root. Temperature at 16:00 was 26°C at that site, relative humidity was 73 %. The specimen MNHN 2005.0235(form 2) was found during the evening on the forest ground in leaf litter.

FIGURE 29. Tadpole of Leptolalax aereus form 2. (A) Dorsal and (B) lateral views, and (C) oral disc of a specimen in stage 26(ZFMK 87009), scale bars = 10 mm (a, b) and 1 mm (c).

Tadpoles of form 2, in different stages, were collected at night (24–27°C; 92–94 % relative humidity) betweenthe 05th and 10th August 2006 in terraces with backwater (water temperature 25°C, pH 7.3) of a rocky mountainstream in Cha Noi, northern Phong Nha - Ke Bang NP. The larvae were found exposed on sandy and silty butsparse substrate where they often were sighted syntopically with larvae of Leptobrachium chapaense.

Placement in molecular phylogeny. Leptolalax aereus is part of a clade that groups with high support theclades of L. minimus, L. ventripunctatus, L. nyx and L. pluvialis.

Conservation status. Known from Savannakhet Province, the south of Ha Tinh Province (within Ky Anh —Ke Go Reserve, 24,800 ha after Ziegler 2002), from Nghe An Province (Pu Huong Nature Reserve), and from the

west of the neighbouring province Quang Binh (within Phong Nha - Ke Bang National Park, 85,754 ha afterZiegler et al. 2004); most probably also in forested areas in between these locations and around. Thus, given thisrather large extent of occurrence we propose the status Least Concern.



Leptolalax (Lalos) minimus (Taylor, 1962)—New combination(Fig. 16D–E, 30–36)

Leptobrachium minimum Taylor, 1962: 320.Onymophoront, by designation as “type”: holophoront (holotype), EHT 34020 (now FMNH 178228) (Marx 1958: 57; exam-

ined by Dubois & Ohler in 1995); adult male, SVL 27.6 mm. Collected by Edward H. Taylor, 21 December 1957.Onymotope: Doi Suthep (18°46’ N, 98°58’ E; alt. about 1000 m), Chiang Mai Province, Thailand.

Specimens referred to L. minimus. Thailand. Phu Kradung National Park: TNRC S1-3918–3924, 6 adult males and 1 adultfemale; — Doi Inthanon: FMNH 216087, TNRC S1-3925, FMNH 216088, adult males and adult female; — Doi Suthep:FMNH 178230, protaxont (paratype); BMNH 1979.437, BMNH 1979.436, FMNH 178228, holophoront; TNRC S1-3926,TNRC S1-3927, FMNH 213933–213938, MNHN 1997.1450, 11 adult males and 2 adult females; — Doi Chiang DaoWatershed: FMNH 216077–216080, TNRC S1-3928–3931, 8 adult females; — Khao Pleung: BMNH 1972.1473, adultfemale; — Doi Nga Chang: BMNH 1972.1475, BMNH 1972.1476, BMNH 1972.1477, BMNH 1972.1474, adult malesand adult female; — Samkrae, Phu Kradung: MNHN 1987.2194, adult male.

Laos. Khammouan Province: Huey Kun Luang (17°40’ N, 104°32’ E): BMNH 1972.1467, adult male; — Xaignabouli Prov-ince: Muang Liep (18°28’ N, 101°40’ E; alt. 290 m): BMNH 1972.1478, BMNH 1974.3067, adult males; — Savannah-khet Province: Pak Mat: BMNH 1972.1471, BMNH 1972.1470, FMNH 121844, MCZ 8762, adult males; — Huey Sapan[Houei Xepon] (16°45’ N, 106°09’ E), Pak Mat, Mekong: BMNH 1972.1468, BMNH 1972.1469, adult male and adultfemale; — Luang Prabang Province, Nam Kham River tributaries: Ban Dong Khan (19°49’ N, 102°49’ E; alt. 500 m):MNHN 2006.2308–2316, adult males, collected 3 and 4 August 2006; — Ban Vang Thong (19°50’ N, 102°59’ E; alt. 550m): MNHN 2006.2328, adult male, collected 25 July 2006; MNHN 2006.2592, one tadpole (DNA voucher) collected 26July 2006; — Ban Kengkoung (19°41’ N, 102°18’ E; alt. 360 m): MNHN 2006.2593–2596, four tadpoles (all DNAvouchers), and MNHN 2006.2597–2598 collected the 28 July 2006; — Ban Nongdi (19°41’ N, 102°22’ E; alt. 400 m):MNHN 2006.2318–2326, adult males, collected 6 August 2006; MNHN 2006.2599, 2006.2614, two tadpoles (DNAvoucher), and two tadpoles in the batch MNHN 2006.2615–2616 collected the 6 August 2006; — Houey Thao (19°48’ N,102°11’ E; alt. 300 m): MNHN 2006.2553–2554.

Specimens referred to L. minimus form 2. Thailand. Chiangmai Province, Doi Chiang Dao, Houey Ban: THNHM 07417–07422, 5 adult males, coll. Yodchaiy Chuarnkern, Stéphane Grosjean, Chantip Inthara, Annemarie Ohler, 13 July 2005.

FIGURE 30. Leptolalax minimus, MNHN 2006.2553, adult male, SVL 28.1 mm. (A) Dorsal and (B) ventral view.



Diagnosis. Member of the subgenus Lalos based on the presence of lateroventral glands and molecular phyloge-netic relationships (Fig. 4), distinguished from other species included in this subgenus by the following combina-tion of characters: moderate-sized species (males 25.7–31.4 mm; females 31.6–37.3 mm) (Table 3) withrudimentary to small webbing and no fringes on toes; tympanum distinct; dermal ridges under toes distinct; fingertips not dilated; dorsal skin smooth (Table 4); brown with darker pattern; dark spots on flanks present; ventral sidewhitish; iris dark gold above, grey below (Table 5).

Description of holophoront. Size and general aspect. (1) Specimen of small size (SVL 27.6 mm), body ratherstout.

Head. (2) Head of medium size, narrower (HW 9.8 mm) than long (HL 10.2 mm; MN 8.6 mm; MFE 6.7 mm;MBE 3.6 mm), flat. (3) Snout slightly protruding, its length (SL 3.4 mm) as long as horizontal diameter of eye (EL3.4 mm). (4) Canthus rostralis angular, loreal region concave, vertical. (5) Interorbital space flat, larger (IUE 2.8mm) than upper eyelid (UEW 2.5 mm) and internarial distance (IN 2.6 mm); distance between front of eyes (IFE4.3 mm) about half of distance between back of eyes (IBE 8.2 mm). (6) Nostrils rounded, closer to tip of snout (NS1.3 mm) than to eye (EN 1.7 mm). (8) Tympanum (TYD 1.8 mm) distinct, rounded; about equal to half of eyediameter, tympanum-eye distance (TYE 1.2 mm) three-fourths of its diameter. (11) Tongue not observed.

Forelimbs. (12) Forearm rather long, rather thin (FLL 7.1 mm), longer than hand (HAL 6.8 mm), not enlarged.(13) Fingers rather short and thin (TFL 3.7 mm). (15) Tips of fingers not enlarged. (16) Fingers without dermalfringe.

Hindlimbs. (19) Shanks about three times longer (TL 12.2 mm) than wide (TW 3.8 mm), shorter than thigh(FL 13.0 mm) and longer than distance from base of internal metatarsal tubercle to tip of toe IV (FOL 11.8 mm).(20) Toe IV (FTL 6.1 mm) about one-third of distance from base of tarsus to tip of toe IV (TFOL 18.3 mm). (22)Tips of toes not enlarged. (23) Webbing rudimentary: I 2 – 2 ½ II 2 – 3 ½ III 2 – 4 IV 4 – 3 V (MTTF 4.3 mm;MTFF 4.1 mm; TFTF 6.1 mm; FFTF 6.6 mm). (24) Fringes on toes absent. (26) Inner metatarsal tubercle moder-ately long, poorly distinct; its length (IMT 1.2 mm) 1.9 times in length of toe I (ITL 2.2 mm).

FIGURE 31. Leptolalax minimus, MNHN 2006.2553, adult male. (A) Lateral view of head, ventral view of (B) right foot andof (C) right hand. Scale bar = 5 mm (3 + 2 mm).



Skin. (29) Dorsal and lateral parts of head and body: snout, region between eyes and side of head smooth; backand flanks smooth with tubercles and glandular folds. (30) Supratympanic fold distinct, extending from eye toabove shoulder. (31) Dorsal parts of limbs: forelimbs, thigh, shank and tarsus smooth with tubercles and glandularfolds. (32) Ventral parts of head, body and limbs: throat, chest, belly and thigh smooth. (33) Presence of mac-roglands: lateroventral ridges present, distinct but regularly interrupted; relatively small distinct femoral glands;indistinct axillary glands and distinct relatively large suprabrachial glands.

Coloration. In alcohol: (34) Dorsal and lateral parts of head and body: uniformly brown with dark spots onflanks, in particular two dark spots near thigh; tympanic fold with dark line. (35) Dorsal parts of limbs: dorsal partof forelimbs, of thigh, of shank and of foot brown with blackish crossbands; posterior part of thigh blackish withsmall white spots. (36) Ventral parts of head, body and limbs: uniformly light coloured.

In life (based on MNHN 2006.2553, not on the holophoront; Fig. 16D): Iris dark golden in upper part, grey inlower part. Dorsal parts of head and body dark brown grey with two round brown spots near shoulder and a fewgrey flecks in middle of back; upper part of flanks dark brown grey, lower part grey with black spots; loreal region,tympanic region and tympanum brown grey; upper lip brown grey with black bands; proximal part of forelimb withorange shade, distal part brown grey with dark grey crossbands; thigh, shank and foot brown grey with dark greycrossbands; posterior part of thigh dark brown with white spots; throat and chest transparent grey; border of throatgrey with white spots; belly white with lemon yellow reflections; shank pinkish grey; macroglands white.

Variation of L. minimus (specimens allocated to group including onymophoront). Specimens show arather high variation in size: the smallest males allocated to L. minimus measure 22.4 mm, whereas the largest malemeasures 30.5 mm (mean 27.2 ± 1.02 mm, n = 20). In the populations from Laos and Thailand allocated to L. min-imus specimens of small and large size are genetically identical. Size variation within populations may be due toage of specimens. A Mann-Whitney U-test does not show significant differences between Lao and Thai popula-tions (U = 379.5; p = 0.539). Thus the two peaks observed in the histograms may reflect the presence of two agegroups in the populations (Fig. 32).

FIGURE 32. Histogram of size (SVL) variation in adult male specimens allocated to Leptolalax minimus for populations fromLaos and Thailand.



Tadpole. The external morphology is based on a DNA voucher specimen in stage 37 (MNHN 2006.2593; TOL50.8 mm and BL 14.4 mm; genetically allocated to L. minimus).

In dorsal view (Fig. 33A), body elliptical elongate, widest at the level of the gills. In lateral view (Fig. 33B),BW 122 % of BH, highest in the middle of the abdomen. Eyes moderately sized, ED 9.6 % of BL. Nares, RN 51 %of NP; NN 70 % of PP. Spiracle tubular, SS 51 % of BL; its opening on an axis which should go through just belowthe apex of myotomes of the caudal muscle. Tail long, TAL 253 % of BL; tail musculature, TMW 75 % BW, thelower part slightly bulging in its proximal third. Upper fin, UF 31 % of MTH, almost non-existent on its first quar-ter; lower fin shallow, LF 21 % of MTH; MTH 112 % of BH, tail tip obtusely pointed. Anal tube moderate-sized,slightly conical elongate. Lateral line system identical but much less visible than in L. bourreti.

FIGURE 33. Tadpole of Leptolalax minimus. (A) Dorsal and (B) lateral views of a specimen in stage 37 (MNHN 2006.2593),scale bar = 10 mm; (C) oral disc of a specimen in stage 26 (MNHN 2006.2595), scale bar = 1 mm. Part of tail cut for tissuesampling.

Oral disc (Fig. 33C), ODW 25 % of BL and 54 % of BW. Marginal papillae of the lower labium cylindricalwith a rounded tip, those of upper labium faint with a very large base, especially medially; two submarginal papil-lae on the left side and three on the right side laterally to P1 and P2. No denticulate papillae. KRF 1:4+4/2+3:1, A1very short, A2 the longest, A3, A4 and A5 subequal; A1 half of P4, P3 more than half of P2. Details of keratodontsnot studied. Jaw sheaths black; lower sheath hidden, arranged deeply under the upper jaw sheath.

Coloration in life: Upper part of body and upper part of flanks grey olive, darker behind the eyes than before;upper part of caudal muscle of the same tint than the fore part of body with nine well defined black spots. Lowerpart of flanks quite transparent in gradation, gills red, coiled gut grey. In profile caudal muscle of the same tint thanits upper part, the black spots being smaller and more numerous towards its extremity. Lower fin immaculate,upper fin with a few small black spots on its distal part. Ventral part of body transparent, thorax immaculate, gillsand heart red, coiled gut brown, caudal muscle with some rare dots.



Coloration in preservative: Most part of upper part of body occupied by extension of the caudal muscle ontobody (muscle dorsalis trunci) beige orange, the rest (mostly the head) translucent grey; internal organs of the headvisible coloured in beige orange but by deeper layers, snout translucent grey, lobes of nares grey. Flanks (lymphaticsacs) grey tinted with light orange brown. Ventral side of body transparent, vicera visible (muscle rectus abdominis,coiled gut, heart). Caudal muscle beige mottled with scarce, medium-sized, light brown spots on proximal part,smaller and more numerous on distal half. Fins grey translucent, upper fin with diffuse light brown pigments form-ing some small indistinct spots, lower fin immaculate. Upper side of hindlimbs pigmented with stellate melano-phores.

Variation of tadpole morphology was assessed on the basis of 10 other specimens, except where noted (MNHN2006.2592, 2006.2594–2599, 2006.2614-2616, in stages 25–37, TOL 44.7–55.7 mm (n = 6) and BL 12.0–17.0mm). Only the ratio values different from those of the described specimen are given: BW 121–137 % of BH; ED8.5–10.0 % of BL; RN 34–65 % of NP; NN 68–80 % of PP; SS 48–58 % of BL; TAL 225–254 % of BL (n = 6);TMH 70–98 % of BH (n = 8); TMH 65–75 % of MTH (n = 4); TMW 63–78 % of BW; UF 29–33 % of MTH; SU106–152 % of BL (n = 8); LF 22–26 % of MTH (n = 6); MTH 113–140 % of BH; ODW 26–29 % of BL; ODW 45–55 % of BW. The maximum body width can be reached in the middle of the abdomen. The more the tadpoles are inadvanced stages, the more their upper fins begin close to body end. The number of submarginal papillae is variableand may reach the number of five on each side. Most of specimens examined have a KRF 1:4+4/3+3:1. In life cer-tain individuals show relatively large spots on dorsum and upper flanks that are formed by iridiophores. In preser-vative, the coloration can be more extended: some specimens have a marbled caudal muscle, the distal half ofupper fin smoked, and irregular blotches on lateral sacs and lateral part of head; other with more dots on caudalmuscle and upper fin, and on the distal half of the lower fin.

Description of reference specimen THNHM 07417, adult male, L. minimus form 2 (fig. 34−35). Size andgeneral aspect: (1) Specimen of small size (SVL 25.6 mm), body rather stout.

Head. (2) Head of medium size, narrower (HW 8.7 mm) than long (HL 10.2 mm; MN 8.7 mm; MFE 7.6 mm;MBE 4.2 mm), rather flat. (3) Snout slightly protruding, its length (SL 5.0 mm) longer than horizontal diameter ofeye (EL 3.5 mm). (4) Canthus rostralis distinct, loreal region concave, rather vertical. (5) Interorbital space flat,larger (IUE 4.0 mm) than upper eyelid (UEW 3.2 mm) and than internarial distance (IN 3.1 mm); distance betweenfront of eyes (IFE 5.9 mm) about two-thirds of the distance between back of eyes (IBE 7.8 mm). (6) Nostrils oval,closer to tip of snout (NS 2.1 mm) than to eye (EN 2.3 mm). (8) Tympanum (TYD 2.0 mm) distinct, rounded; two-thirds of eye diameter, tympanum-eye distance (TYE 1.3 mm) three-fifths of its diameter. (11) Tongue large, cor-date.

Forelimbs. (12) Forearm long, thin (FLL 7.7 mm), longer than hand (HAL 6.6 mm), not enlarged. (13) Fingersshort and thin, but finger III, long and thin (TFL 3.7 mm). (15) Tips of fingers not enlarged. (16) Fingers withoutdermal fringe. (18) Palmar tubercles very prominent, spine like.

Hindlimbs. (19) Shanks about four times longer (TL 13.2 mm) than wide (TW 2.9 mm), longer than thigh (FL12.1 mm) and distance from base of internal metatarsal tubercle to tip of toe IV (FOL 12.1 mm). (20) Toes rathershort and thin; toe IV (FTL 6.1 mm) about one-third of the distance from base of tarsus to tip of toe IV (TFOL 18.9mm). (22) Tips of toes not enlarged. (23) Webbing rudimentary, narrow ridges on toes: I 2 – 2 ½ II 2 – 3 III 2 ½ – 3¾ IV 4 – 3 V (MTTF 4.5 mm; MTFF 4.5 mm; TFTF 6.6 mm; FFTF 7.1 mm). (24) Dermal ridge along toe Vabsent. (26) Inner metatarsal tubercle rather long, very prominent, spine like; its length (IMT 1.3 mm) 1.8 times inlength of toe I (ITL 2.3 mm). (27) Tarsal fold absent.

Skin. (29) Dorsal and lateral parts of head and body: snout and region between eyes with glandular warts; sideof head smooth; back and upper part of flanks with glandular warts forming longitudinal ridges; lower part offlanks with glandular warts. (30) Supratympanic fold distinct, from eye to above forelimb. (31) Dorsal parts oflimbs: forelimbs, thigh, shank and tarsus with a few glandular warts. (32) Ventral parts of head, body and limbs:smooth. (33) Presence of macroglands: lateroventral glands present, continuous; relatively small distinct roundedfemoral and surpabrachial glands; indistinct axillary glands.

Coloration. In alcohol: (34) Dorsal and lateral parts of head and body: Dark greyish brown with poorly distinctpatterns including an interorbital triangle and longitudinal lines, blackish small spots on flanks; loreal and tympanicregion light greyish brown; tympanum dark brown; upper lip light greyish brown with dark brown bands. (35) Dor-sal parts of limbs: forelimb beige with dark brown crossbands on distal part; thigh, shank and foot light brown with



dark brown crossbands; posterior part of thigh brown with a blackish brown triangular region near vent, all speck-led with small white dots. (36) Ventral parts of head, body and limbs: throat whitish; margin of throat light brownwith whitish spots; chest and belly whitish; posterior part of thigh brown, anterior part whitish; webbing brown;macroglands white.

FIGURE 34. Leptolalax minimus form 2, THNHM 07417, reference specimen, adult male, SVL 25.6 mm. (A) Dorsal and (B)ventral view.

In life (Fig. 16B–C): Iris copper orange in upper part, dark grey in lower part. Dorsal parts of head and body,and upper flanks greyish brown with dark brown pattern showing whitish outline; lower flanks greyish brown withblack spots and orange dots; loreal and tympanic region greyish brown; tympanum blackish brown; forelimb in itsproximal part orange, distal part greyish brown with black crossbands; shank and foot greyish brown with blackcrossbands; posterior part of thigh dark brown with white spots; throat light grey, transparent; border of throat grey-ish brown with white spots; chest and belly ivory white with light yellow reflections; lower part of shank pinkishgrey; femoral glands white.

Male secondary sexual characters: (38) Vocal sacs present, indistinct on throat; slit-like paired openings poste-rior on mouth floor. (39) Other male secondary sexual characters not observed.

Distribution. Thailand: Chiang Mai Province; Laos: Khammouan Province, Luang Prabang Province, Savan-nahkhet Province (Fig. 9).

Natural history. Specimens of L. minimus collected in July and August 2006 were in breeding condition. Themales were found on the edges of small or moderate secondary streams (Fig. 36C) sitting on leaves, branches,stones and rocks, on or near the ground (0–0.2 m) at short to moderate distances (0–6 m) from the water. Most wereon the edge of the water (0–0.2 m distance). Two breeding males, MNHN 2006.2322 and 2006.2323, wereobserved touching one another with their head region and then separating. This is interpretated as a territorialbehavior. In several of these streams Sylvirana nigrovittata, Limnonectes cf. kuhlii and L. gyldenstolpei, and rarelyXenophrys parva, Kurixalus odontotarsus, Microhyla berdmorei and Chirixalus doriae, were observed.



FIGURE 35. Leptolalax minimus form 2, THNHM 07417, reference specimen, adult male. (A) Lateral view of head, ventralview of (B) right foot and of (C) right hand. Scale bar = 5 mm (3 + 2 mm).

Tadpoles of L. minimus were hidden under dead leaves during daytime. At night, they were found eitheruncovered in the side pools of the streams or in pools connected to the streams but devoid of current, or betweenthe gravels where the water only outcrops. They were found in the same streams as the tadpoles of Xenophrysparva although not in the same microhabitats. No other tadpole species were found in association.

Specimens of form 2 were collected at night in the forest near a small stream (Fig. 36B) and artificial pools.The specimens were sitting on the ground (maximum 0.2 m high) either close to the water or at distances up to 4 m.They were mainly on branches and leaves, dead leaves, or rocks. Adult males took a very particular calling positionexposing the suprabrachial gland and the orange coloured upper part of their forelimbs which gave them an insect-like appearance (Fig. 16C). The adult female THNHM 07421 had large (diameter 2.0 mm) whitish oocytes inovary.

Placement in molecular phylogeny. Leptolalax minimus is part of a highly supported clade that groups theclades of L. aereus, L. ventripunctatus, L. nyx and L. pluvialis. The specimens allocated to L. minimus form 1 forma well supported clade which is the sister group to L. minimus form 2 both together forming a well-supported clade.

Etymology. From latin minimus, small. Adjective, agreeing in gender with generic nomen.Comment. In the molecular analysis, specimens originally identified as Leptolalax minimus from Thailand

and Laos form a clade with two distinct branches, one including specimens from Thailand only, and a secondbranch including specimens from Thailand and Laos. These two branches show a p distance of 3.46 %. The amountof molecular divergence (3.46 %) is above the threshold of 3 % recently proposed to recognize candidate species infrogs (Fouquet et al. 2007; Vieites et al. 2009). High pairwise sequence divergences do not suggest a species statusof two specimens or lineages but are simply a probabilistic support: the more a divergence is above the empiricallycalculated threshold value, the less likely is the alternative explanation of deep haplotype sharing. The discriminantanalysis based on morphometric ratios of measurements on specimens allocated by 16S sequences to distinctclades allows separating the two forms and also gives allocation for the holophoront of Leptobrachium minimum tothe clade then named L. minimus (Table 11). To support species discrimination we searched for morphologicalcharacters that would correlate with the groups defined by DNA sequences but failed. Morphologically these twogroups cannot be distinguished. The specimens of form 2 fall within the variation observed in those specimensallocated to L. minimus. There is no evidence for morphological divergence. Despite the presence of molecular dif-



ferentiation and in absence of morphological differentiation we tentatively keep the two forms in a single specieslooking for further evidence. Nevertheless we give here a description of a voucher specimen of L. minimus form 2.

Another (larval) specimen from the same locality in Thailand, based on the molecular data, is grouped with L.oshanensis AY561306 and L. bourreti, and might represent another distinct species but this lineage is so far knownfrom this tadpole only. More data, in particular on adult specimens, are necessary for sound taxonomic conclusionson this third form.

Conservation status. This species was considered a synonym of L. oshanensis and thus has not been evalu-ated previously. It is known from a large range. Though its habitat encounters threats from deforestation, we pro-pose to allocate the status Least Concern to L. minimus on the basis of its large extent of occurrence.

FIGURE 36. Habitat of species of Leptolalax. (A) stream near Long Nai, habitat of L. eos and L. ventripunctatus; (B) stream atHouey Ban, habitat of L. minimus form 2; (C) stream near Ban Kengkoung, habitat of L. minimus; (D) stream at Ha Tinh,habitat of L. aereus.



TABLE 11. Results of canonical discriminant analysis including data of minimus1 and minimus2. The holophoront ofLeptobrachium minimum was included in the analysis without group membership.

A. Description of Variance

B. Statistical significance

C. Standardized canonical discriminant function coefficients

D. Classification results

Key to species

We present here a key to the species of subgenus Lalos of Leptolalax based on morphological characters andcoloration pattern of postmetamorphic specimens.

1 Tympanum indistinct externally . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2Tympanum distinct . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3

2 Ventral body immaculate; large-sized species (males 48.3–52.7 mm, females 56.7–58.9 mm). . . . . . . . . . . . . . . . . . . . L. sungiVentral body with black; moderate-sized species (males 24.4–29.5, females 30.2 mm) . . . . . . . . . . . . . . . . . . . . . . L. tuberosus

3 Ventral body immaculate, dark pattern not extending beyond axillary glands . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 4Ventral body with dark pattern, at least extending on side of belly . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 13

Function Eigenvalue % of Variance Cumulative % Canonical Correlation1 9.942 100.0 100.0 0.953

Test of Function(s) Wilks' Lambda Chi-square df Sig.1 .091 20.338 17 0.257

Function 1SVL 4.831rhw 2.238rhl 4.013rmn -4.234rmfe 0.762rmbe 2.679rife -0.650ribe -0.269rin 4.826ren -0.661rel -0.986rtyd 3.455rtye -1.258rhal 0.007rtl -6.609rfol 2.908rimt -1.651

OTU Predicted Group Membership Total

minimus 1 minimus 2

Count minimus 1 14 0 14minimus 2 0 5 5

Holophoront of L. minimum 0 1 1

% minimus 1 100 0 100minimus 2 0 100 100

Holophoront of L. minimum 0 100 100



4 No distinct dark spots on side of body . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5Distinct dark spots on side of body . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 6

5 Dorsal pattern indistinct; well developed fringes on toes; large-sized species (males 33.1-34.7 mm). . . . . . . . . . . . . . . . . L. eosDorsal pattern includes distinct spots surrounding lighter coloured warts; fringes on toes absent; moderate-sized species (males26.7-32.6 mm) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . L. nyx

6 Fringes on toes wide . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . L. liuiFringes on toes narrow or absent. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 7

7 Webbing on feet small . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 8Webbing on feet rudimentary or absent . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 11

8 Skin of dorsum smooth, without prominent warts or folds. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 9Skin of dorsum with prominent warts and/or folds. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 10

9 Dorsum with distinct dark brown pattern including a triangular spot between eyes; large-sized species (males 28.0-36.2 mm) .. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . L. bourretiDorsum of more uniform aspect; pattern poorly distinct; moderate-sized species (males 25.7-31.4 mm) . . . . . . . . . L. minimus

10 Dorsum with roughly granular aspect; moderate-sized species (males 26.9-28.3 mm) . . . . . . . . . . . . . . . . . . . . . . . . L. lateralisSkin of dorsum with glandular warts and folds; moderate to large-sized species (males 27.5-32.3 mm) . . . . . . .L. pelodytoides

11 Throat immaculate, only border pigmented . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .L. tamdilThroat with speckling or dots . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 12

12 Tip of toes swollen; large-sized species (male 40.8 mm) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .L. nahangensisTip of toes not dilated; moderate-sized species (males 26.6-30.7 mm) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . L. oshanensis

13 Dark pattern on belly distinct . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 14Pattern on belly incomplete or indistinct. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 16

14 Throat as belly, dark brown with distinct large confluent white spots; moderate-sized species (males 26.6-28.8 mm) . . . . . . . .. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .L. melanoleucusThroat in central part uniformly coloured . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 15

15 Throat dirty white, uniformly coloured; dorsal pattern distinct but without light outlines; small-sized species (males 22.0-23.0mm) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . L. pluvialisThroat with dark sparkling on border; dorsal pattern usually with light grey outlines; rather small-sized species (males 23.7-27.7 mm) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . L. ventripunctatus

16 Throat with fine mottling . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 17Throat of uniform colour . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 18

17 Throat entirely dusky; dorsum scattered with faint dark spots . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . L. fuliginosusThroat light grey with white mottling; dorsum with irregular darker blotches. . . . . . . . . . . . . . . . . . . . . . . . . . . . . L. khasiorum

18 Spots on side of chest and belly only; dorsal skin smooth . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . L. alpinusBelly with speckling covering sides or whole belly; dorsal skin with regularly set granules. . . . . . . . . . . . . . . . . . . . . L. aereus

Discussion

Generic classification

Any classification is based on a hypothesis of phylogenetic relationships and one of the first qualifications is itsinstability. Taxonomy is a science in progress and stability is always limited in time. Sometimes for a long periodclassifications do not change but when new methods are used and exploration uncovers new data, classificationsmove. No classification can be considered a final one, nevertheless we have to express our knowledge in formalclassifications as an effort of synthesis of knowledge at a certain point. This work on Leptolalax started in 2005when only 19 species were considered valid. Until the end of 2010 ten more species have been described. Only 6species have been so far included in phylogenetic analysis based on morphology and for 2 species molecular datahave been analysed in a phylogeny. For most species, the classification is based on morphological similarity only.

This work deals with more than 363 specimens of the Oriental frog genus Leptolalax (Megophryidae). For 97of them molecular data were available from China, Laos, Thailand and Vietnam. The analysis of molecular dataincluded 10 putative species allocated to the subgenus Lalos and two species of the nominative subgenus Leptola-lax. This is a higher number than in any previous study, but still only includes about one third of the 33 describedspecies in the genus (Dubois et al. 2010). Of three more species (L. lateralis, L. nahangenis, L. sungi), the ono-matophores and hypodigms of the original descriptions were studied and included in species comparison (appendix1). Our analysis provides evidence for monophyly of a group made up by the 12 species studied, which makes itlikely that the genus Leptolalax is monophyletic (holophyletic) although this remains to be tested by a more ampledataset. Cladistic analysis using morphological data on adults and larvae (Delorme unpublished), based on a sam-ple of six species, proposed two apomorphies for the genus (presence of a suprabrachial gland and small size of



outer metatarsal tubercle). Zheng et al. (2004) obtained a sister group relationship between L. oshanensis and L.ventripunctatus (L. pelodytoides in their work). Our study is one of the first phylogenetic approaches that includemore than one species of the subgenus Leptolalax and thus allows testing monophyly of this taxon. The two speciesadded on morphological evidence to the subgenus Leptolalax (L. arayai and L. heteropus) are sister taxa with highsupport. These results are the first indication of monophyly for this subgenus which had no support from morphol-ogy. The monophyly for the subgenus Lalos was first stated with a single synapomorphy (presence of lateroventralglands) by Delorme et al. (2006). All species with lateroventral glands from our study form a clade based on ourmolecular data. The recognition of two subgenera within the genus Leptolalax is therefore supported although aconfirmation by more extensive data sets is required. Delorme et al. (2006) included L. tuberosus and L. sungi inthe subgenus Lalos. Both have been described as not showing lateroventral glands. Examination of the hypodigmof L. sungi showed the presence of a lateroventral gland which is composed of fine small glands. The holophorontand type series of L. tuberosus has not been studied and it being a member of Lalos needs confirmation. This taxo-nomic decision was taken mainly on biogeographical reasons. In L. croceus, the lateroventral glands are describedas absent, but figure 2C shows distinct glands in a line on side of belly (Rowley et al. 2010b) which might behomologous by the criteria of position and transition with the lateroventral glands. L. applebyi and L. melicus donot show lateroventral glands and thus might be members of the subgenus Leptolalax. This discussion clearlyshows that morphology alone is not enough for classification of these frogs because of similarity, homoplasy andpoor character state definition due to overlapping intra-taxon variation. Nevertheless the search of morphologicalcharacters for classification is important in two ways: it allows to link historical data on specimens to recent find-ings and it is helpful for workers in the field which do not have access to molecular methods. At local level speciesrecognition without higher classification should be possible in most cases as sympatric species are rare in manyanuran groups. But a hierarchical approach of taxonomy allows to store and retrieve knowledge on a more generallevel.

The tadpoles of the subgenus Leptolalax studied so far are slightly but consistently different from those ofLalos. According to Inger (1983), in Leptolalax (Leptolalax) gracilis, the nares are rimmed with a mediodorsal pro-jection, and the serrations of the beak are fine without a medial bigger serration forming a medial cusp. In contrast,the nares of the tadpoles of Lalos are rimmed with four lobes, and the serrations of their beak are coarse and usuallywith a medial cusp (Delorme et al. 2006). Furthermore, there might be differences in the keratodont formula: theKRF of Leptolalax (Leptolalax) kajangensis seems to be similar to that prevalent in Lalos but with more rows(1:5+5/4+4:1; Grismer et al. 2004), whereas that of Leptolalax (Leptolalax) arayai is very different (1+1/0 withsubmaginal papillae scattered all over the oral disc; Malkmus & Kosuch 1999). The coloration of this latter speciesis also very different from that of all other known tadpoles of the genus, being clear grey with white spots on tail.The tadpoles reported by Inger (1966) under the nomen Leptolalax gracilis mixed tadpoles of Leptobrachellamjobergi (toothless form) and tadpoles of L. gracilis with a KRF 1:(2+2)–(6+6)/(2+2)–(3+3):1 (Inger 1983). Thecoloration of this tadpole is reported as pinkish grey without pattern, which is quite different from the panther pat-tern of the tadpoles described herein (with the exception of L. bourreti). There seems to exist correlation of differ-ences in tadpole morphology and colour pattern with differences in advertisement calls (Matsui 1997, 2006) whichmight provide some evidence for the subgeneric arrangement proposed by Delorme et al. (2006); at least, they sup-port the monophyly of Lalos as a subclade of Leptolalax, even if the subgenus Leptolalax may turn out to be non-monophyletic.

Species discrimination

Previous authors have pointed to the cryptic habits and high morphological similarity among Leptolalax species(Rowley & Cao 2009; Matsui et al. 2009). These species are all forest living frogs that require running water forreproduction and thus showing little ecological differentiation. Even though some species, like L. pluvialis, occuronly in high mountains, their habitat and ecology does not appear to differ from those of species living in loweraltitudes. Generally, Leptolalax have only occasionally been found in secondary or open habitat. Colour pattern andmorphology in Leptolalax are probably highly constrained by habitat. Most species are brownish in life with moreor less distinct colour patterns on the dorsal side. This kind of pattern is clearly cryptic on forest floor. Interspecificdifferentiation of these colour patterns is very low and intraspecific variation may be rather high. Variation con-



cerning webbing, relative length of various parts of limbs, head, size of tympanum and skin structure gives cluesfor taxonomic discrimination, but the states of single characters are often overlapping between species. Further-more, intrapopulational and intraspecific variation especially in colour characters is high (variation in intensity,presence of colour pattern particularities, overall colour shade) (Bourret 1942; this paper). Thus, even though ahigh number of characters was used in this study, only a few of them have diagnostic value and can be used for dis-crimination within a few species pairs. Thus most species have overlapping size ranges but a few are larger orsmaller than the bulk of the species (Table 3). Webbing and fringes on toes seem to form distinct character states.Also the presence or absence of tympanum is clearly defined (Table 4). The extent of ridges under fingers and toesand the skin structure on dorsum are probably dependent on condition of fixation and storage of specimens andneed further study to test their utility in taxonomic discrimination. Colour patterns (spots on dorsum, ventral pat-tern, spots on flanks showed to be useful to a certain extent in species discrimination (Table 5). Eye colour which isvery useful in taxonomy of many anuran groups does not seem discriminating in Leptolalax. Despite these con-straints, most species were described on the basis of morphological data and coloration. Only recently, evidencefrom advertisement calls has been used for species discrimination (Matsui 2006; Rowley & Cao 2009; Rowley etal. 2010b). Molecular data from DNA sequences were used for species discrimination by Rowley et al. (2010b) todistinguish L. applebyi and L. melicus.

Comparison with the tadpoles of other Leptolalax (Lalos) species is not easy because the published descrip-tions are typically not very detailed when they exist. A rather complete description was made by Liu (1940) for tad-poles of L. oshanensis (as L. pelodytoides; taxonomic status corrected by Liu 1950). The KRF of all knowntadpoles of Lalos vary only within a narrow range: 1:3+3/2+2:1 for L. alpinus (Fei et al. 1991, 2005; Fei 1999),1:(3+3)–(4+4)/(2+2)–(3+3):1 for L. bourreti (Grosjean 2004; this paper), 1:3+3/2+2:1 for L. liui (Fei et al. 1991,2005; Fei & Ye 1992; Fei 1999), 1:4+4/(2+2)–(3+3):1 for L. minimus (this paper), 1:3+3/2+2:1 for L. oshanensis(Liu 1940, 1950; Inger et al. 1990; Fei et al. 1991, 2005; Fei 1999), 1:(3+3)–(5+5)/(2+2)–(3+3):1 for L. aereusform 1 (this paper), 1:4+4/3+3:1 for L. aereus form 2 (this paper), 1:(3+3)–(4+4)/3+3:1 for L. ventripunctatus (Feiet al. 1991, 2005; Fei 1999; this paper), 1:(3+3)–(4+4)/(2+2)–(3+3):1 for Leptolalax sp. (Grosjean 2004) as sum-marized in table 12. It should be kept in mind that an ontogenetic variation in row number could exist and that theKRF 1:4+4/3+3:1 appears to be the most prevalent formula for prometamorphic (sensu Etkin 1932, 1968) tadpolesof this group. Other data are available in literature (Smith 1917; Pope 1931 although his Fujian population isknown to be L. liui; Inger 1983; Fei et al. 1991; Fei 1999; Inthara et al. 2005; Meewattana 2005) but as their spe-cific assignation is dubious, we do not consider them at this time.

TABLE 12. Summary of keratodont row formula (KRF) of the known tadpoles of the subgenus Lalos.

The main problem of taxonomic revisions is the necessity to resolve taxonomic allocations of nomina based onmorphological characters, often poorly described and based on single specimens or small series of specimens.Availability of onymotopic material would allow sampling of new data, but is not always possible for reasonsexternal to science. This is the situation with the nomina Leptobrachium pelodytoides and Ixalus lateralis. Thenomenclatural situation of I. lateralis has been recently resolved by neophoront designation (Humtsoe et al. 2008),whereas the identity of L. pelodytoides requires further investigation. Here we use morphological and colour evi-

Keratodont row formula References

L. alpinus 1:3+3/2+2:1 Fei et al. 1991, 2005; Fei 1999

L. bourreti 1:(3+3)–(4+4)/(2+2)–(3+3):1 Grosjean 2004; this paper

L. liui 1:3+3/2+2:1 Fei et al. 1991, 2005; Fei & Ye, 1992; Fei 1999

L. minimus 1:4+4/(2+2)–(3+3):1 This paper

L. oshanensis 1:3+3/2+2:1 Liu, 1940, 1950; Inger et al. 1990; Fei et al. 1991, 2005; Fei 1999

L. aereus form 1 1:(3+3)–(5+5)/(2+2)–(3+3):1 This paper

L. aereus form 2 1:4+4/3+3:1 This paper

L. ventripuncatus 1:(3+3)–(4+4)/3+3:1 Fei et al. 1991, 2005; Fei 1999; this paper

L. sp. 1:(3+3)–(4+4)/(2+2)–(3+3):1 Grosjean 2004



dence to propose that specimens from eastern Myanmar belong in a particular taxon distinct from all other speci-mens studied by us and by other authors who have recently worked on Leptolalax.

Leptolalax oshanensis was described on morphological evidence. Later, L. minimus from Thailand was consid-ered to constitute a junior synonym of this species based on morphological evidence (Dubois 1981). Our analysisincluded material allocated to L. oshanensis from Sichuan and topotypical material of L. minimus. Analysis ofmolecular data clearly shows that the specimens from China and Thailand belong to different clades and do notform a monophyletic group. Thus their synonymy cannot be maintained. Morphological evidence for separation ofL. oshanensis from the Thai species is poor as we did not have sufficient Chinese material for a in-depth analysis.From the illustrations of Liu (1950) and Fei (1999), the tadpole of L. oshanensis seems to be uniformly brown(similar to that of L. bourreti with which it forms a clade) and so is clearly different from that of L. minimus whichshows a panther-like pattern. This particularity is an additional evidence for the recognition of L. minimus and L.oshanensis as two good and distinct species (Dubois 1981; Fei et al. 2009). Moreover their KRF might be different(see above). Data sequences of L. oshanensis available from GenBank show important variation and this nomen islikely to have been applied to several species. The sequence AY561306 from the specimen from Emei Shan, theonymotope, probably applies to the name L. oshanensis. And the second sequence might apply to a new species.Further studies of material from the various Chinese populations might allow to understand their evolutionary rela-tionships and to clarify their taxonomy.

Leptolalax bourreti was described on poorly preserved specimens, based mainly on its large adult size (Dubois1983). This is not sufficient evidence to separate it from specimens collected in northern Laos and southern Yun-nan. Onymotopic material of L. bourreti was collected, and thus data on coloration are available. These specimenswere also included in our molecular analysis which showed that the specimens from Sa Pa form a distinct lineage.The tadpoles of L. bourreti are distinguished from the other herein described tadpoles by their uniform browncolour (vs. the “panther” pattern on the tail in the three other species) and smaller eyes (ED 6.3–10.1 % of BL) thanin the other species (8.2–12.4 %). Furthermore, the tadpoles of this species have denticulate papillae (a charactershared with the form 2 of L. aereus) unlike the tadpoles of the other species. Thus this species is supported by evi-dence from three independent data sets. Nevertheless it seems to show high variation concerning coloration, in par-ticular of eye colour (Bourret 1942; this study). Eye colour is a quite stable character in numerous anuran species(Glaw & Vences 1997), thus this observation is quite striking. Does this reflect presence of cryptic species or genu-ine intraspecific variation? More detailed studies of these populations are needed to settle the evolutionary signifi-cance of this observation.

Several specimens preliminarily allocated in this work to the new species L. eos have been allocated by mor-phological evidence to L. pelodytoides (Liu & Hu 1961; Yang 1991; Fei & Ye 1992). From the size of adult malesand females, L. eos might be considered close to L. bourreti, another large sized species of Lalos. The new specieshas support from threefold evidence (molecules, morphometry and coloration). It does not form a monophyleticclade with L. bourreti specimens and thus represents an independent lineage. Its coloration pattern is clearly dis-tinct both from L. pelodytoides and L. bourreti and it is diagnosable by morphometry. This species lives in sympa-try with L. ventripunctatus. In northern Laos these two species, members of distinct clades of Lalos, were observedat the surroundings of the same stream within a few meter distance (Ohler, personal observations).

The species Leptolalax pluvialis is known only from high altitude in northern Vietnam. It was described onmorphological and colour evidence, and analysis of molecular data is concordant with this proposal. It is sistergroup to a clade of morphologically distinct species.

A specimen from northern Vietnam was allocated to Leptolalax bourreti on the basis of morphology, but theDNA data show that it is a member of a distinct clade. Here we described it as a new species, L. nyx, on morpholog-ical, colour and molecular evidence.

Leptolalax ventripunctatus from southern Yunnan was described on distinct morphology and ventral colourpattern. Its range was later shown to extend to northern Yunnan. We here assign specimens from nearby areas inLaos to this species based on their morphological similarity. These specimens form a morphologically and molecu-larly homogenous group. We further conclude that specimens from northern Vietnam probably belong to this spe-cies on the basis of their great morphological similarity to Lao specimens.

Leptolalax aereus was separated from the other species on the basis of morphological and molecular evidence.It shows a very high intra-specific variation, in molecular sequences, in larval morphology and in adult coloration.The two lineages sampled by us differ by an uncorrected p-distance of 2.4%. Within L. aereus, two tadpole formsare here recognized. The differences in larval characters between these two forms are the following: form 1 is much



larger than form 2, all the morphometric ratios are larger in form 2 although largely overlapping the values of form1 except the ratios involving UF and LF which are equal in the two forms and TMH/MTH which shows the reversetendency (61–69 % in form 1 vs. 54–61 % in form 2), the oral disc is larger in form 2 (ODW/BW 35–45 % in form1 vs. 46–60 % in form 2), the older specimens of form 1 have an additional keratodont row on the upper labium anddenticulate papillae, and the golden pigmentation of form 1 is less distinct in life than in form 2. Furthermore, thenumber of keratodonts per 0.5 mm is higher in form 1 than in form 2 (41–49 vs. 38–41; RH personal observation).As sampling includes few adult specimens, we could not make conclusions on morphological variation of adults.For the time being we keep the two subgroups as a single species. More data, particularly evidence about presenceor absence of gene flow between the two groups would be necessary for further assessment of this taxonomicdilemma.

Leptolalax minimus is a morphologically quite similar assembly from different localities showing minor varia-tion in extension of webbing, skin structures and coloration of iris which cannot be correlated to a particular geno-type, but from the molecular evidence there clearly appear to be two distinct lineages. These two sibling lineagesare a good example for the poor morphological discrimination in this genus, molecular differentiation precedingmorphological differentiation.

In this paper, we used evidence from three independent data sets to distinguish species of Leptolalax. Weconclude that morphology and colour pattern alone are not sufficient to recognize all species as they do not provideunambiguous discriminant character states for all species pairs. The L. minimus forms had not been recognized asdistinct when observed in the field, nor can they be discriminated by morphometric methods, but molecular dataallowed to separate them and this separation is possibly reflected by very subtle morphometric differences as bothforms were neatly separated along the first factor of a discriminant analysis (not shown). Nevertheless, evidence istoo poor to separate these two forms and recognize them as different species. The situation is reversed for the twoforms of L. aereus that show some morphological discrimination, particularly in coloration of adults andmorphology of larvae, but molecular divergence is somehat lower (below the candidate species threshold) and thenumber of adult specimens is too small for a statistical analysis. It is possible that L. aereus as defined herein in factis a species complex, but we deem the available data to be insufficient for describing any species at this stage,especially as for this paper we have not studied the specimens used for the original description of L. aereus(Rowley et al. 2010c), published while this paper was being prepared for submission, and because no tadpoles areknown belonging to the genealogical lineage of the holophoront of L. aereus (which is yet different from our forms1 and 2).

Previous authors have described new Leptolalax species which show major differences, such as L. sungi, thelargest species of the genus, or L. fuliginosus exihibiting distinct colour pattern on ventral side. On the other hand,until now, specimens with an uniformly coloured belly and moderate body size from Indochinese subregion haveusually been allocated to only three species, L. pelodytoides, L. bourreti and L. oshanensis. The present analysisallows to add several species to these three species. So-called L. pelodytoides from Vietnam (Ziegler 2002) repre-sent a distinct species, L. aereus. Populations named L. pelodytoides from Yunnan (Yang 1991) and L. bourretifrom Laos (Stuart 2005) are here tentatively allocated to a distinct species described herein, L. eos. Specimens allo-cated to L. oshanensis do not form a monophyletic group; in particular specimens from Thailand and Laos can bereferred to L. minimus. The few specimens of L. oshanensis from China available in this study also seem to belongto distinct lineages and might to be allocated to more than a single species.

Natural history

Frogs from various genera (Limnonectes, Arthroleptis, Phrynobatrachus, Eleutherodactylus, Craugastor) live onthe forest floor and exhibit a brownish dorsal colour pattern. Some of these frogs show a much more prominentadaptation of dorsal coloration and skin structures to this forest habitat than Leptolalax. This is the case of Mego-phrys that not only has brown dorsal colour, but also dorsal skin ridges and palpebral appendages that resembleleaves (Manthey & Grossmann 1997). Leptolalax do not show any particular pattern or structures. As the functionof colour pattern mainly seems to protect against predators, no intraspecific differentiation due to sexual selectioncan be observed. This situation is comparable to the genera Arthroleptis, Eleutherodactylus and Craugastor, wheresimilar habits and colour patterns hide molecular differentiation (Hedges et al. 2008; Blackburn 2009).



Some specimens of Leptolalax were collected outside of forest. Liu (1950) stated that L. oshanensis was foundunder stones in corn fields. A few specimens of L. bourreti were observed in secondary habitat such as agriculturalland, scrub and savannah. Agriculture in the mountain regions of Sichuan and Vietnam is shifting cultivation withsmall-sized fields close to natural habitats (Tordoff et al. 1999). The presence of Leptolalax in secondary habitatmight be underestimated as few surveys have made equivalent collecting efforts in primary and secondary habitats.Another source of bias might come from the fact that breeding males while calling are more easily located and aremore densely grouped near streams. More studies are needed to understand the habits and ecology of Leptolalaxspecies.

The tadpoles of the subgenus Leptolalax (Lalos) are clearly adapted to a life in swift water, and even tointerstitial life. Though they are listed in the lotic suctorial and clasping ecomorphological guild by Altig &McDiarmid (1999b), they rather belong to the fossorial guild (McDiarmid & Altig 1999) as they do not use theiroral disc for fixation on the substrate, have no anterior gap in marginal papillae and a KRF higher than 2/3. Theyare very elongate, with almost dorsally positioned eyes and nares, a long tail with a strong caudal musculature andmoderately shallow and thick fins, especially the upper one which is nearly nonexistent in the proximal third of thetail, and a ventral mouth. Besides these characters, the tadpoles of this group possess several unique andconspicuous characters in the family (contrary to the statement of Lathrop 1997): the presence of well delimitedlateral lymphatic sacs (otherwise also present in the tadpoles of the unrelated subfamily Astylosterninae generaAstylosternus, Leptodactylodon, Nyctibates and Trichobatrachus: Amiet 1970, 1971; Grosjean 2004), the foursmall lobes around the nare openings, and the peculiar oral disc with an upper labium bearing four small lobes, amedially deeply notched lower labium and the typical arrangement of short keratodont rows. Despite these veryconspicuous peculiarities, the tadpoles described herein are very similar morphologically and virtuallyindistinguishable from each other. The most divergent one is the tadpole of L. bourreti which has an uniform browncolour and denticulate papillae (the last character is shared with the form 2 of L. aereus) unlike the other tadpolespecies.

Taxonomic diversity and biogeographical considerations

Previously to this paper, 30 species of Leptolalax were known but the number has increased in an important man-ner recently as nine of them (33.3 %) were validated or described from 2004 to 2010. Here we validate one morespecies which was considered a synonym (Leptolalax minimus) and we describe two more new species, whichresults in a total of 33 species. The species of this genus are known from the Oriental region going west as far asMeghalaya (they are not known from the eastern Himalayas), north to Sichuan and southern Hubei province, eastto Fujian province. In the south, Leptolalax species are present to Malaysia and Borneo, but have so far not beenreported from Sumatra, Java and the Philippines.

Most of the recently described species have small distribution areas and do not show sympatry with other Lep-tolalax species (Matsui 2006). In our work we report several pairs of sympatric occurrence which are of differentevolutionary origins. In northern Laos and northern Vietnam, the members of pairs of sympatric species (L. eos andL. ventrimaculatus; L. pluvialis and L. bourreti) belong to two distinct evolutionary lineages. In the central part ofnorthern Thailand and Laos, differentiation in geographically close locations leads to genotypically differentiatiedforms (L. minimus and form 2) belonging to a single lineage. Similarily, in L. aereus of Ha Tinh and Quang Binhprovinces of Vietnam, interpopulational variation leading to a certain degree of differentiation between the lineagescan be observed.

Whereas several species of Leptolalax appear to be confined to very small distribution areas, others appear tobe more widespread. Quaternary changes in temperature and in monsoon intensity in south-east Asia have beensuggested by palaeoclimatic studies (Cosford et al. 2008) and probably led to periodic expansion of open, unfor-ested habitats (Pushkina et al. 2010). It is uncertain to which extent such changes have affected the area at earlierages when speciation of these frogs has occurred. However, such changes in vegetation cover might have left somefrogs isolated in particular forest isolates and allowed others rapid expansions into new areas once forests wereconnected again. Such expansions might have led to the sympatry of Leptolalax eos and L. ventripunctatus, giventhat L. ventripunctatus is part of a clade present in northern Laos, Thailand and Vietnam, whereas L. eos is a mem-ber of a clade of more northern distribution in China. However, the existence of at least one sympatric pair of sister



forms (L. minimus and form 2) indicates that adaptive modes of speciation, in sympatry or parapatry, should not bea-priori disregarded when testing hypotheses of species diversification in tropical biotas (e.g., Vences et al. 2009).

When only a few species were recognized in the genus, distinguishing them by morphological charactersappeared to be sufficient. Species were distinguished by body size, webbing extension, presence of fringes on toes,particular coloration pattern. Even though we could not include all known species in our phylogenetic analysis, it isclear that none of these characters is non-homoplastic, thus they cannot be used as unique synapomorphies and thusdiagnostic characters for any species-level lineage. For most species taxa in this genus, no morphological apomor-phies can be found and only diagnoses using a combination of characters allow to recognize species on the basis ofmorphology alone.

Leptolalax pelodytoides and L. bourreti, as understood in this paper, cannot be considered as widely distributedspecies anymore. Thus the conservation status of the newly described and revised species must be assessed. Thesespecies are not uncommon in forest habitats but seem to depend strictly on this habitat for their survival. Anotherfactor of threat is the use of water from small streams for domestic needs and agriculture that destroys the breedinghabitat of the species. Therefore, evolution and conservation of Leptolalax species are closely linked to the fate offorests in south-east Asia and the high diversity of species reveals a high past and present structural diversity ofhabitats and the complex evolutionary history in this group.

Other species groups that inhabit similar habitats in the Oriental region might have had a similar evolutionaryhistory reflected by similar relationships among species. Stuart et al. (2010) published hypotheses of relationshipsamong members of the Amolops monticola group (stream living Ranidae species) and could not find any sympatricspecies. In Ansonia, bufonid toadlets with stream dwelling tadpoles, Matsui et al. (2010) uncovered high diversifi-cation in Borneo where syntopic distribution of two species can be commonly observed. These syntopic speciesbelong then to different major clades. A similar situation has been detected in the Limnonectes kuhlii group (Dicro-glossidae) where recently 22 distinct evolutionary lineages were recognized (McLeod 2010). In several cases, sym-patric lineages, none closely related, were detected.

In conclusion, despite the long taxonomic history and the important recent effort in describing biodiversity,south-east Asia still holds many uncovered species, in particular among rheophilous species (adapted to stream liv-ing). This kind of habitat is important for the breeding and living of numerous anuran species of this realm. Streamsare in fact separated from others by habitats which are not appropriate for rheophilous species. Often streams areseparated by mountains with important altitudinal amplitudes. Many Oriental species of frogs are limited to a rathernarrow altitudinal range (Dubois 1976) and thus might be unable to migrate outside the valleys. Stream living alsohas consequences on interactions between specimens and presumably on the genetic pattern of populations (whichare not really panmictic) because it imposes a linear spatial organization for breeding behavior (Inger et al. 1974;Dubois 1977a). Comparative studies of the patterns of species diversity and of phylogeography among rheophilousand non-rheophilous species emerges as an interesting field for future studies.

Acknowledgements

The fieldwork of AO and SG was supported by PPF “Faune et Flore du sud-est asiatique” and PPF “Etat et struc-ture phylogénétique de la biodiversité actuelle et fossile”; Frontier Vietnam; Maison du Patrimoine, Luang Pra-bang, Laos, and the Phongsaly Forest Conservation and Rural Development Project, Laos. Thanks are addressedfor their help in the field to Thomas Calame, Yodchaiy Chuaynkern, Sandra Dos Santos, Pierre Guédant, Chris Hat-ten, Richard Humphry, Chantip Inthara, Bounhom Phothimath, Steven Swan, Alexandre Teynié, Jack Tordoff, KimValkone and Yannis Varelides.

For the loan of specimens and visit to their collections we thank Darrel Frost (AMNH), Barry T. Clarke(BMNH), Alan Resetar (FMNH), Nguyen Thien Tao (VNMN, Vietnam National Museum of Nature) and NguyenQuang Truong (IEBR), Giuliano Doria (MSNG), José Rosado (MCZ), Yodchaiy Chuaynkern (THNHM), Wolf-gang Böhme (ZFMK), Mark-Oliver Rödel (ZMB), Jakob Hallermann (ZMH).

The drawings prepared by Julien Norwood and photographies arranged by Delphine Zigoni were funded by theDépartement de Systématique et Evolution of the Muséum National d’Histoire Naturelle.

M. Kondermann and G. Keunecke helped with the molecular lab work. J. Rowley kindly made unpublishedDNA sequences available for a preliminary comparison.



Fieldwork of RH in 2006 was funded by the Wilhelm-Peters-Fond (Deutsche Gesellschaft für Herpetologieund Terrarienkunde e.V. (DGHT), Rheinbach, Germany; the Alexander Koenig Stiftung, Bonn, Germany; theAlexander Koenig Gesellschaft, Bonn, and the European Union of Aquarium Curators (E.U.A.C). KCW was sup-ported by SYNTHESYS grant FR-TAF-4000, funded by the European Union. We thank the Phong Nha – Ke BangNational Park directorate, Dinh Huy Tri (Science Research Centre, Phong Nha – Ke Bang) and Sladjana Miskovic(Cologne Zoo’s Nature Conservation Project, Phong Nha – Ke Bang) for supporting the loan of specimens from theNational Park’s scientifc collection.

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Appendix 1. Additional specimens examined

Leptolalax lateralis: India, Nagaland, Wokha district, Nhyatsutchu stream: IASST 1-4, IASST A68 (neophoront), 4 males, 1female.

L. liui: China, Fujian, Chong’An: AMNH A30130, A30138, 2 males; BMNH 1931.2.10.1, female; FMNH 24415-19, 4 males,1 female; Yenping: AMNH A28413, male; Hongkong, Lantau Peak: BMNH 1983.323, female; FMNH 211230, female;Sunset Peak: BMNH 1983.1112-1113, 2 males; Tai Mo Shan: BMNH 1983.324, male.

L. nahangensis: Vietnam, Tuyen Quang Province, Na Hang Nature Reserve, entrance of a cave, 7 km east of the village of PacBan (22°19’94” N; 105°25’79” E) elevation 314 m: ROM 28715, holophoront, adult male.

L. sungi: Vietnam, Vinh Phu Province, from a stream on the east side of the village of Tam Dao (21°27’31”N; 105°38’61”E),elevation 925 m: ROM 28474, holophoront adult male; ROM 28471-73, 31152-53, other protaxonts, 2 males, 2 females, 1young.
