Discovery of a deeply divergent and highly endemic frog lineage from Borneo: A taxonomic revision of Kalophrynus nubicola Dring, 1983 with descriptions of two new species (Amphibia: Anura: Microhylidae)
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The genus Kalophrynus is represented by tiny to medium terrestrial or subfossorial frogs and is widely distributed in Southeast Asia. The diversity centre of this genus is Borneo, where almost half of all nominal species are distributed and all of these are endemic to this and the peripheral islands. Kalophrynus nubicola is the smallest species within this genus; it is found only at high elevations in Gunung Mulu National Park, northeastern Sarawak, Malaysian Borneo. Three groups of populations were preliminary reported, but taxonomic decision on the groups was not yet given. In this study, we investigated the morphological, acoustic, and genetic differences among these three populations to reassess their taxonomic status. Morphological analyses confirmed differences in body size and colouration, molecular analyses indicated that each group was monophyletic, and differences in the acoustic characteristics of each group were also diagnostic. Therefore, herein, we describe them as distinct taxa. Phylogenetic analyses revealed that the K. nubicola group forms a highly divergent clade from other species within the genus. Our findings reveal that seven Kalophrynus species occur in Gunung Mulu National Park, with non-overlapping distributions according to elevation or forest type. We further discuss the biogeography and evolutionary history of this species group.
A genomic study by Chan and colleagues demonstrated that Pulchrana picturata from Peninsular Malaysia and Sumatra (and southern Thailand by implication) are distinct from the name-bearing lineage from Borneo. In this study, we present additional morphological and bioacoustic data to support the recognition of the new species, which we describe herein. The new species, P. sundabarat, can be distinguished from its sister species, P. picturata, by having a more conspicuous and defined dorsolateral stripe, brighter coloured dorsal markings, smaller male body size, and marked differences in various aspects of the male advertisement call. We also provide an updated synthesis on the current knowledge of the P. picturata group with regard to taxonomy, distribution, evolutionary relationships, and ecology.
Frogs of the genus Microhyla include some of the world's smallest amphibians and represent the largest radiation of Asian microhylids, currently encompassing 50 species, distributed across the Oriental biogeographic region. The genus Microhyla remains one of the taxonomically most challenging groups of Asian frogs and was found to be paraphyletic with respect to large-sized fossorial Glyphoglossus. In this study we present a time-calibrated phylogeny for frogs in the genus Microhyla, and discuss taxonomy, historical biogeography, and morphological evolution of these frogs. Our updated phylogeny of the genus with nearly complete taxon sampling includes 48 nominal Microhyla species and several undescribed candidate species. Phylogenetic analyses of 3,207 bp of combined mtDNA and nuDNA data recovered three well-supported groups: the Glyphoglossus clade, Southeast Asian Microhyla II clade (includes M. annectens species group), and a diverse Microhyla I clade including all other species. Within the largest major clade of Microhyla are seven well-supported subclades that we identify as the M. achatina, M. fissipes, M. berdmorei, M. superciliaris, M. ornata, M. butleri, and M. palmipes species groups. The phylogenetic position of 12 poorly known Microhyla species is clarified for the first time. These phylogenetic results, along with molecular clock and ancestral area analyses, show the Microhyla-Glyphoglossus assemblage to have originated in Southeast Asia in the middle Eocene just after the first hypothesized land connections between the Indian Plate and the Asian mainland. While Glyphoglossus and Microhyla II remained within their ancestral ranges, Microhyla I expanded its distribution generally east to west, colonizing and diversifying through the Cenozoic. The Indian Subcontinent was colonized by members of five Microhyla species groups independently, starting with the end Oligocene-early Miocene that coincides with an onset of seasonally dry climates in South Asia. Body size evolution modeling suggests that four groups of Microhyla have independently achieved extreme miniaturization with adult body size below 15 mm. Three of the five smallest Microhyla species are obligate phytotelm-breeders and we argue that their peculiar reproductive biology may be a factor involved in miniaturization. Body size increases in Microhyla-Glyphoglossus seem to be associated with a burrowing adaptation to seasonally dry habitats. Species delimitation analyses suggest a vast underestimation of species richness and diversity in Microhyla and reveal 15-33 undescribed species. We revalidate M. nepenthicola, synonymize M. pulverata with M. marmorata, and provide insights on taxonomic statuses of a number of poorly known species. Further integrative studies, combining evidence from phylogeny, morphology, advertisement calls, and behavior will result in a better systematic understanding of this morphologically cryptic radiation of Asian frogs.
An analysis of the elevational distributions of Southeast Asian birds over a 28-year period provides evidence for a potential upward shift for 94 common resident species. These species might have shifted their lower, upper, or both lower and upper boundaries toward a higher elevation in response to climate warming. These upward shifts occurred regardless of habitat specificity, further implicating climate warming, in addition to habitat loss, as a potentially important factor affecting the already imperiled biotas of Southeast Asia.
We provide an integrative taxonomic analysis of the Lipinia vittigera species complex from mainland Southeast Asia. Based on examination of external morphology, color pattern, and 681 base pairs of the cytochrome oxidase subunit I (COI) mitochondrial gene, we demonstrate the presence of four morphologically distinct lineages of Lipinia in Vietnam, Cambodia, Thailand, and Malaysia, showing a sequence divergence ranging 15.5%–20.4%. All discovered lineages are discretely diagnosable from one another by a combination of scalation traits and color patterns. A review of the published distribution data and a re-examination of available type material revealed the following
(1) distribution of L. vittigera (Boulenger, 1894) sensu stricto is restricted to Sundaland and the Thai-Malay Peninsula south of the Isthmus of Kra; (2) L. microcercus (Boettger, 1901) stat. nov. is elevated to full species rank; the species has a wide distribution from central and southern Vietnam across Cambodia to eastern Thailand; we regard Lygosoma vittigerum kronfanum Smith, 1922 and Leiolopisma pranensis Cochran, 1930 as its junior synonyms; (3) Lipinia trivittata sp. nov. occurs in hilly areas of southern Vietnam, Cambodia, and eastern Thailand; and (4) Lipinia vassilievi sp. nov. is currently known only from a narrow area along the Vietnamese-Cambodian border in the foothills of the central Annamite Mountain Range. We further provide an identification key for Lipinia occurring in mainland Southeast Asia.
Anurans in Peninsular India exhibit close biogeographical links with Gondwana as well as Laurasia, often explainable by the geological history of the Indian subcontinent; its breakup from Gondwanan landmasses followed by long isolation that resulted in diversification of endemic lineages, and subsequent land connections with Asia that enabled dispersal of widespread groups. Although widely distributed, the frog subfamily Microhylinae mostly comprises of geographically restricted genera found either in Southeast and East Asia or Peninsular India and Sri Lanka. Here we report a previously unknown microhylid from the Western Ghats in Peninsular India with closest relatives found over 2,000 km away in Southeast Asia. Based on integrated evidence from mitochondrial and nuclear DNA, adult and tadpole morphology, hand musculature, male advertisement call, and geographical distance, we recognize this enigmatic frog as a distinct new species and genus endemic to the Western Ghats. The discovery of Mysticellus franki gen. et sp. nov. and its close evolutionary relationship with the Southeast Asian genus Micryletta also provide insights on the biogeography of Microhylinae. Genus-level divergences within the subfamily suggest multiple Cenozoic biotic exchange events between India and Eurasia, particularly through postulated Eocene land bridges via Southeast Asia prior to accretion of the two landmasses.
An integrative taxonomic analysis reveals a remarkable degree of cryptic speciation between sympatric, specialized, leaf-litter species in an upland clade of the scincid genus Tytthoscincus across the sky-island archipelago of Peninsular Malaysia. Bayesian inference and maximum likelihood phylogenies based on the mitochondrial gene ND2 indicate that this clade is composed of a lineage of limb, digit and tympanum-reduced, semi-fossorial species and a lineage of leaf-litter generalist bearing longer, more robust limbs and digits, and well-developed tympanae. Behavioural differences inferred from differing ecomorphologies form the basis of hypotheses proposed to account for resource partitioning at Fraser’s Hill, Pahang between the sympatric leaf-litter generalist T. bukitensis and the new semi-fossorial species T. kakikecil sp. nov. However, this hypothesis does not account for the extreme case of cryptic speciation and syntopy between T. bukitensis and a new leaf-litter generalist T. martae sp. nov. at Genting Highlands, Pahang. The phylogenetic analyses also indicate there is an additional new, upland, semi-fossorial species, T. jaripendek sp. nov., from Cameron Highlands and a new lowland, leaf-litter generalist, Tytthoscincus sp., from Hulu Terengganu, Terengganu. The specific identities of the new upland populations are corroborated by morphological analyses using combinations of principal component analysis, discriminant analysis of principal components, analysis of variance, Tukey HSD (honest significant difference) tests and Welch two-way t-tests. The Hulu Terengganu specimen was too damaged to provide a reliable description. A biogeographic scenario inferred from a time-calibrated BEAST analysis suggests that speciation in the upland clade has been ongoing since the Upper Pliocene following repeated episodes of secondary contact associated with glacier-driven climatic oscillations generating upland forest expansion and retraction. These analyses highlight the increasing levels of herpetological diversity and endemism being discovered in the montane regions of Peninsular Malaysia and bring to light the understudied ecological complexity of Peninsular Malaysia’s leaf-litter ecosystems and the growing need for its conservation.
The current account presents the results of a 14-day amphibian survey at Maliau Basin Conservation Area (MBCA). With a total of approximately 170 man-hrs, 44 species were detected at four study sites during the field period; four more species were later discovered outside the two-week campaign. The results are compared to the results of previous surveys. Apart from adults, we present the first photographic documentation of the larval stages of Chiromantis inexpectatus and Bornean Phrynoidis juxtaspera, along with a brief tadpole description; the better-known tadpoles of four more species were recorded. The results of our expedition suggest that nine more species are present at MBCA than reported by previous studies. We present an updated list of known species in the MBCA, comprising 61 species. The species accumulation curve over the 14 days period of the core survey did not show signs of asymptotic saturation. We conclude that the definitive species number for MBCA amphibians has the potential to increase with more thorough surveys in the future. Key Words regional inventory rapid assessment anuran biodiversity visual encounter genetic barcode
Southeast Asia and southern China (SEA-SC) harbor a highly diverse and endemic flora and fauna that is under increasing threat. An understanding of the biogeographical history and drivers of this diversity is lacking, especially in some of the most diverse and threatened groups. The Asian leaf-litter frog genus Leptolalax Dubois 1980 is a forest-dependent genus distributed throughout SEA-SC, making it an ideal study group to examine specific biogeographic hypotheses. In addition, the diversity of this genus remains poorly understood, and the phylogenetic relationships among species of Leptolalax and closely related Leptobrachella Smith 1928 remain unclear. Herein, we evaluate species-level diversity based on 48 of the 53 described species from throughout the distribution of Leptolalax. Molecular analyses reveal many undescribed species, mostly in southern China and Indochina. Our well-resolved phylogeny based on multiple nuclear DNA markers shows that Leptolalax is not monophyletic with respect to Leptobrachella and, thus, we assign the former to being a junior synonym of the latter. Similarly, analyses reject monophyly of the two subgenera of Leptolalax. The diversification pattern of the group is complex, involving a high degree of sympatry and prevalence of microendemic species. Northern Sundaland (Borneo) and eastern Indochina (Vietnam) appear to have played pivotal roles as geographical centers of diversification, and paleoclimatic changes and tectonic movements seem to have driven the major divergence of clades. Analyses fail to reject an "upstream" colonization hypothesis, and, thus, the genus appears to have originated in Sundaland and then colonized mainland Asia. Our results reveal that both vicariance and dispersal are responsible for current distribution patterns in the genus.
The presence of an adhesive abdominal sucker (gastromyzophory) allows tadpoles of certain species of anurans to live in fast-flowing streams. Gastromyzophorous tadpoles are rare among anurans, known only in certain American bufonids and Asian ranids. To date, Huia sumatrana , which inhabits cascading streams, has been the only Sumatran ranid known to possess gastromyzophorous tadpoles. In the absence of thorough sampling and molecular barcoding of adults and larvae, it has remained to be confirmed whether other Sumatran ranid species living in similar habitats, i.e., Chalcorana crassiovis , possesses this larval type. Moreover, the taxonomic status of this species has long been uncertain and its taxonomic position within the Ranidae, previously based exclusively on morphological characters, has remained unresolved. To study the diversity and relationships of these frogs and to establish the identity of newly collected gastromyzophorous tadpoles from Sumatra, we compared genetic sequences of C. crassiovis -like taxa from a wide range of sites on Sumatra. We conducted bayesian and maximum likelihood phylogenetic analyses on a concatenated dataset of mitochondrial (12S rRNA, 16S rRNA, and tRNA val ) and nuclear (RAG1 and TYR) gene fragments. Our analyses recovered C. crassiovis to be related to Clinotarsus , Huia , and Meristogenys . The DNA barcodes of the gastromyzophorous tadpoles matched adults from the same sites. Herein, we provide a re-description of adult C. crassiovis and propose “ C. kampeni ” as a synonym of this species. The molecular evidence, morphological features, and distribution suggest the presence of two related new species. The two new species and C. crassiovis together represent a distinct phylogenetic clade possessing unique molecular and morphological synapomorphies, thus warranting a new genus.
We report on a discovery of Siamophryne troglodytesGen. et sp. nov., a new troglophilous genus and species of microhylid frog from a limestone cave in the tropical forests of western Thailand. To assess its phylogenetic relationships we studied the 12S rRNA–16S rRNA mtDNA fragment with final alignment comprising up to 2,591 bp for 56 microhylid species. Morphological characterization of the new genus is based on examination of external morphology and analysis of osteological characteristics using microCT-scanning. Phylogenetic analyses place the new genus into the mainly Australasian subfamily Asterophryinae as a sister taxon to the genus Gastrophrynoides , the only member of the subfamily known from Sundaland. The new genus markedly differs from all other Asterophryinae members by a number of diagnostic morphological characters and demonstrates significant mtDNA sequence divergence. We provide a preliminary description of a tadpole of the new genus. Thus, it represents the only asterophryine taxon with documented free-living larval stage and troglophilous life style. Our work demonstrates that S. troglodytesGen. et sp. nov. represents an old lineage of the initial radiation of Asterophryinae which took place in the mainland Southeast Asia. Our results strongly support the “out of Indo-Eurasia” biogeographic scenario for this group of frogs. To date, the new frog is only known from a single limestone cave system in Sai Yok District of Kanchanaburi Province of Thailand; its habitat is affected by illegal bat guano mining and other human activities. As such, S. troglodytesGen. et sp. nov. is likely to be at high risk of habitat loss. Considering high ecological specialization and a small known range of the new taxon, we propose a IUCN Red List status of endangered for it.
We describe a new genus and two new species of toads from the Sumatran volcanoes Gunung Sorikmarapi and G. Kunyit, in the provinces of Sumatera Utara and Jambi, respectively. The new taxa can be distinguished from other genera, and each other, based on genetic differentiation, morphology, and advertisement call structure. We employ both nuclear and mitochondrial data to provide a phylogenetic hypothesis of relationships for the bufonid genera of the Sunda Shelf. While broadly corroborating previous studies, our results also shed light on the phylogenetic position of Pseudobufo. The new genus, Duttaphrynus, and Pseudobufo are basal to other Sunda Shelf genera in our phylogenies.
Molecular dating studies typically need fossils to calibrate the analyses. Unfortunately, the fossil record is extremely poor or presently non-existent for many species groups, rendering such dating analysis difficult. One such group is the Asian horned frogs (Megophryinae). Sampling all generic nomina, we combined a novel ∼5kb dataset composed of four nuclear and three mitochondrial gene fragments to produce a robust phylogeny, with an extensive external morphological study to produce a working taxonomy for the group. Expanding the molecular dataset to include out-groups of fossil represented ancestral anuran families, we compared the priorless RelTime dating method with the widely used prior based Bayesian timetree method, MCMCtree, utilising a novel combination of fossil priors for anuran phylogenetic dating. The phylogeny was then subjected to ancestral phylogeographic analyses, and dating estimates were compared with likely biogeographic vicariant events. Phylogenetic analyses demonstrated that previously proposed systematic hypotheses were incorrect due to paraphyly of genera. Molecular phylogenetic, morphological and timetree results support the recognition of Megophryinae as a single genus, Megophrys, with a subgenus level classification. Timetree results using RelTime better corresponded with the known fossil record for the outgroup anuran tree. For the priorless in-group, it also outperformed MCMCtree when node date estimates were compared with likely influential historical biogeographic events, providing novel insights into the evolutionary history of this pan-Asian anuran group. Given a relatively small molecular dataset, and limited prior knowledge, this study demonstrates that the computationally rapid RelTime dating tool may outperform more popular and complex prior reliant timetree methodologies.
As sources of data for global forest monitoring grow larger, more complex and numerous, data analysis and interpretation become critical bottlenecks for effectively using them to inform land use policy discussions. Here in this paper, we present a method that combines big data analytical tools with Emerging Hot Spot Analysis (ArcGIS) to identify statistically significant spatiotemporal trends of forest loss in Brazil, Indonesia and the Democratic Republic of Congo (DRC) between 2000 and 2014. Results indicate that while the overall rate of forest loss in Brazil declined over the 14-year time period, spatiotemporal patterns of loss shifted, with forest loss significantly diminishing within the Amazonian states of Mato Grosso and Rondônia and intensifying within the cerrado biome. In Indonesia, forest loss intensified in Riau province in Sumatra and in Sukamara and West Kotawaringin regencies in Central Kalimantan. Substantial portions of West Kalimantan became new and statistically significant hot spots of forest loss in the years 2013 and 2014. Similarly, vast areas of DRC emerged as significant new hot spots of forest loss, with intensified loss radiating out from city centers such as Beni and Kisangani. While our results focus on identifying significant trends at the national scale, we also demonstrate the scalability of our approach to smaller or larger regions depending on the area of interest and specific research question involved. When combined with other contextual information, these statistical data models can help isolate the most significant clusters of loss occurring over dynamic forest landscapes and provide more coherent guidance for the allocation of resources for forest monitoring and enforcement efforts.
Subsequent to the Miocene (approximately 35 Mya), Borneo has served as an insular refuge and a source of colonization for a broad range of species emigrating to others parts of Sundaland. A phylogeny-based historical biogeographical hypothesis for the Stream Toad genus Ansonia supports multiple instances of an out-of-Borneo scenario. An ancestral range estimation indicates that in situ speciation of Ansonia on the island of Borneo during the Late Miocene and Pliocene (approximately 2–13 Mya) eventually resulted in an invasion of the Philippines, Sumatra, and two independent invasions of the Thai-Malay Peninsula. When collecting material for the biogeographical analysis, a new species of Ansonia, Ansonia khaochangensis sp. nov. was discovered in a limestone cave from the Khao Chang karst tower in Phangnga Province, in southern Thailand. Ansonia khaochangensis sp. nov. can be differentiated from all other species of Ansonia by having a unique combination of morphological and colour pattern characteristics. Phylogenetic evidence based on the mitochondrial genes 12S and 16S indicates that it is nested within a clade of other species distributed north of the Isthmus of Kra. The cave lifestyle of this new species is a unique and a significant departure from lotic environments common to most other species of Ansonia. The reproductive biology of this species is unknown.
We describe two new species of skinks from Gunung Penrissen, Sarawak, Malaysia, in northern Borneo, Tytthoscincus batupanggah sp. nov. and T. leproauricularis sp. nov. Morphological and molecular analyses both corroborate the two new species as unique compared to all other Tytthoscincus and additional Sphenomorphus that are candidates for taxonomic placement in the genus Tytthoscincus. Despite their phenotypic similarity and sympatric distribution, a molecular analysis shows that the new species are not sister taxa and exhibit a deep genetic divergence between each of their respective sister taxa. We discuss how historical climatic and geographic processes may have led to the co-distribution of two relatively distantly related phenotypically similar species. In light of these discoveries, we also emphasize the importance of conserving primary montane tropical rainforest for maintaining species diversity.
A new insular, endemic species of microhylid frog of the genus Kalophrynus is described from Tioman Island, off the southeastern coast of Pahang, Peninsular Malaysia. Kalophrynus tiomanensis sp nov. can be differentiated from its congeners by the following combination of characters: SVL 21.4-26.3 mm; reduced webbing on toes; outer metatarsal tubercle absent; large, black inguinal spot and unique markings on dorsum. This discovery increases the number of endemic species of amphibians on Tioman Island to at least three.
A new microhylid frog is described from Bario, Kelabit Highlands of the State of Sarawak, in the Eastern Malaysia of Borneo Island. Morphologically, the new species differs from all known congeners in the combination of small body size; short fourth finger without subarticular tubercle; absence of subarticular tubercles on fifth toe, and usually on first finger; presence of light lateral stripe and dark inguinal spot; absence of nuptial pads and outer metatarsal tubercles. Acoustically, the new species differs from all congeners whose calls have been reported, except for K. baluensis and K. yongi, with short unpulsed notes emitted intermittently.
The tree frogs of the taxon Rhacophoridae are known for their impressive diversity of reproductive strategies. Direct development on land has been described in the Old World Bush Frogs belonging to the genera Philautus, Pseudophilautus, and Raorchestes. However, in numerous species especially within the Bornean Philautus, breeding behaviours remain unknown. In this paper, we match a clutch of eggs found on Gunung Mulu National Park, Sarawak, Malaysia (Borneo), using genetic barcoding to syntopically occurring adults of Philautus acutus. This species is known only from its type locality in the montane forests at high elevations on Gunung Mulu. The eggs were found on leaf litter of the forest floor and are characterised by a protective, compact, outer jelly capsule. The froglets inside the eggs were at advanced stages of development and showed a bifurcating dorsal pattern similar to adults of P. acutus. Beside the discovery of its breeding behaviour, we add a description of the habitat of this rare species. Furthermore, this account of aerial direct development in a Philautus species from Borneo contributes to our understanding of the evolution of reproductive strategies within the lineage. Finally, we present a review of observations of the breeding behaviour in Bornean Philautus species available in the literature.
Sticky Frogs, Kalophrynus Tschudi, are a small group of Southeast Asian species with their greatest diversity in Borneo. Two species, K. interlineatus (Blyth) and K. pleu-rostigma Tschudi, were proposed as residents of Myanmar (Burma), northern and southern populations respectively. An analysis of morphological variation in Burmese specimens and comparison with small samples from throughout the distribution of the interlineatus-pleurostigma group of species demonstrates variable levels of regional differentiation, which I interpret as evidence of speciation. This interpretation recommends the restriction of K. pleurostigma to Sumatra populations and K. interlineatus to peninsular Myanmar and adjacent mainland Southeast Asian populations. The northern Borneo populations have the largest body size of any members of this group and represent a new species, K. meizon. The Philippine populations , K. sinensis Peters, are confirmed as unique as proposed recently by Ohler and Grosjean (2005). The populations from northern Myanmar also represent a distinct taxon, K. anya, and differ from K. orangensis (India and Bangladesh) and K. interlin-eatus (peninsular Myanmar and Southeast Asia). To assist the ongoing discovery of new species of this cryptic frog group, I provide a taxonomic resumé of all currently recognized species of the genus Kalophrynus and a diagnostic key to all species of Kalophrynus. Tschudi (1838) recognized the uniqueness of the Sticky Frogs with the erection of a new genus, Kalophrynus. Simultaneously he described K. pleurostigma for a specimen from Sumatra, thereby establishing this taxon as the type species of Kalophrynus. The Sumatran origin has not been questioned, although few Sumatran specimens are available to provide a thorough examination of variation of topotypic K. pleurostigma in the broadest sense of all Sumatran populations. The scarcity of Sumatran specimens and the widespread occurrence of presumably similar appearing frogs from southern Myanmar through Southeast Asia to Borneo and the Philippines led to the name pleurostigma being applied to the larger-bodied Sticky Frogs in this area. This concept of K. pleurostigma had its foundation in Parker's review (1934) of the Microhylidae. Therein, he recognized four of the eight species of Kalophrynus described prior to his review. He considered the broadly distributed K. pleurostigma as consisting of two subspecies (nominate and interlineatus). This concept persisted for sixty years until Matsui et al. (1996) recognized the two PROCEEDINGS OF THE CALIFORNIA ACADEMY OF SCIENCES
Despite considerable progress in unravelling the phylogenetic relationships of microhylid frogs, relationships among subfamilies remain largely unstable and many genera are not demonstrably monophyletic. Here, we used five alternative combinations of DNA sequence data (ranging from seven loci for 48 taxa to up to 73 loci for as many as 142 taxa) generated using the anchored phylogenomics sequencing method (66 loci, derived from conserved genome regions, for 48 taxa) and Sanger sequencing (seven loci for up to 142 taxa) to tackle this problem. We assess the effects of character sampling, taxon sampling, analytical methods and assumptions in phylogenetic inference of microhylid frogs. The phylogeny of microhylids shows high susceptibility to different analytical methods and datasets used for the analyses. Clades inferred from maximum-likelihood are generally more stable across datasets than those inferred from parsimony. Parsimony trees inferred within a tree-alignment framework are generally better resolved and better supported than those inferred within a similarity-alignment framework, even under the same cost matrix (equally weighted) and same treatment of gaps (as a fifth nucleotide state). We discuss potential causes for these differences in resolution and clade stability among discovery operations. We also highlight the problem that commonly used algorithms for model-based analyses do not explicitly model insertion and deletion events (i.e. gaps are treated as missing data). Our results corroborate the monophyly of Microhylidae and most currently recognized subfamilies but fail to provide support for relationships among subfamilies. Several taxonomic updates are provided, including naming of two new subfamilies, both monotypic.
The first molecular phylogeny for mountain reed snakes (genus Macrocalamus) based on the mitochondrial gene cytochrome b is not entirely consistent with the previous taxonomy based on morphology and colour pattern. Macrocalamus chanardi is shown to be a species complex composed of three different allopatric lineages distributed across different upland areas in Peninsular Malaysia that are morphologically conserved but genetically distinct. A new and morphologically different species, Macrocalamus emas sp. nov., is described from the Cameron Highlands, Pahang, Peninsular Malaysia. It occurs in sympatry with four other ecologically equivalent species of Macrocalamus and one other species of Collorhabdium. The phylogeographical pattern of sympatric genetically distinct species of Macrocalamus endemic to upland areas is attributed to the fossorial nature of these snakes and the montane forest expansion and retraction resulting from cyclical, glacioeustatically driven climatic processes that have reconstructed the geography of Sundaland continuously over the last 25 Myr.
Narrow-mouthed frogs (Anura: Microhylidae) are globally distributed and molecular data suggest the rapid evolution of multiple subfamilies shortly after their origin. Despite recent progress, several subfamilial relationships remain unexplored using phylogenomic data. We analysed 1,796 nuclear ultraconserved elements, a total matrix of 400,664 nucleotides, from representatives of most microhylid subfamilies. Summary method species-tree and maximum likelihood analyses unambiguously supported Hoplophryninae, as the earliest diverging microhylid and confirm Chaperininae as a junior synonym of Microhylinae. Given the emerging consensus that subfamilies from mainland Africa diverged early, microhylids have likely occupied the continent for more than 66 million years.
An integrative taxonomic analysis of the Ptychozoon lionotum group across its range in Indochina and Sundaland recovers P. lionotum sensu lato Annandale, 1905 as paraphyletic with respect to P. popaense Grismer, Wood, Thura, Grismer, Brown, & Stuart, 2018a and composed of four allopatric, genetically divergent, ND2 mitochondrial lineages. Multivariate and univariate analyses of continuous and discrete morphological and color pattern characters statistically and discretely diagnose each lineage from one another and together, with maximum likelihood and Bayesian inference analyses, provide the foundation for the recognition of each lineage as a new species—hypotheses corroborated with a Generalized Mixed Yule Coalescent species delimitation analysis. Ptychozoon cicakterbang sp. nov. ranges throughout Peninsular Malaysia to Pulau Natuna Besar, Indonesia; P. kabkaebin sp. nov. is endemic to northern and central Laos; and P. tokehos sp. nov. ranges from southern Thailand south of the Isthmus of Kra northward to Chiang Mai, fringing the Chao Phraya Basin and ranging southward through Cambodia to southern Vietnam. Ptychozoon lionotum sensu stricto ranges from northwestern Laos through southern Myanmar to eastern India. The phylogeographic structure within each species varies considerably with P. lionotum s.s. showing no genetic divergence across its 1,100 km range compared to P. cicakterbang sp. nov. showing upwards of 8.2% sequence divergence between syntopic individuals. Significant phylogeographic structure exists within P. tokehos sp. nov. and increased sampling throughout Thailand may require additional taxonomic changes within this species.
A new diminutive species of microhylid frog (genus Microhyla) is described from the Matang Range, Sarawak, Malaysian Borneo. The new species is an obligate of the pitcher plant, Nepenthes ampullaria, breeding in senescent or mature pitchers, and is Old World’s smallest frog and one of the world's tiniest: adult males range between SVL 10.6– 12.8 mm (n = 8). The new species is diagnosable from congeners in showing dorsum with low tubercles that are relatively more distinct on flanks; a weak, broken, mid-vertebral ridge, starting from forehead and continuing along body; no dermal fold across forehead; tympanic membrane and tympanic annulus absent; Finger I reduced to a nub proximal to Finger II in males; toe tips weakly dilated; phalanges with longitudinal grooves, forming two scale-like structures; webbing on toe IV basal; toes with narrow dermal fringes; inner and outer metatarsal tubercles present; and dorsum brown with an hour-glass shaped mark on scapular region. Miniaturization and reduced webbing may be the result of navigation on the slippery zone of pitchers, situated below the peristome.
Gives a description of the adult and tadpole stages of this microphylic frog discovered in the Pasoh Forest Reserve in 1981, with brief notes on ecology and behaviour and a comparison with closely related species.-R.Land
We report a new microhylid frog of the genus Kalophrynus from Peninsular Malaysia and describe it as a new species Kalophrynus kiewi based on results of molecular and morphological analyses. The new species is a large-sized Kalophrynus (snout-vent length 34-47 mm) and is morphologically very similar to K. pleurostigma, in which it has long been unrecognized, and K. meizon, but is distinguished from them molecularly and by its body size, dorsal skin texture, and relative length of hindlimb. The new species is also clearly differentiated from all the other members of the genus by molecular and morphological differences. Taxonomic and distributional problems of the genus Kalophrynus in Peninsular Malaysia are briefly discussed.
A new highland species of the dwarf litter frog, Leptobrachella itiokai, is described from Gunung Mulu National Park, northern Sarawak, East Malaysia. It occurs syntopically with another highland species, L. brevicrus, but differs from it by a unique call that may be described as a chirp, with dominant frequency of ca. 10 kHz. The new species is morphologically distinguishable from congeners by the combination of the following traits: rounded ventrolateral glands usually not in series; side of body with small black spots; ventrum entirely dark-coloured with tiny light spots; male with thread-like lineae masculinae.
While the island of Borneo is considered a global
biodiversity hotspot, the species richness in many groups remains
unknown and appears underestimated. During herpetological
surveys carried out in the interior of Sarawak, East
Malaysia, several individuals of a small species of the genus
Ansonia Stoliczka 1870 were collected on the Usun Apau
plateau and in the Gunung Hose mountain range (Ansonia
sp. Usun Apau). An integrative taxonomic approach comprising
phylogenetic (2.4 kb mitochondrial rDNA fragment,
Bayesian Inference and Maximum Likelihood, >5.1 % to its
closest relative) and morphometric analyses (25 measurements,
multivariate ratio analysis and linear discriminant analysis),
as well as morphological comparisons support the status
of this operational taxonomic unit as a separate taxon at species
level. The obtained phylogenetic hypothesis corroborates
the two major clades within Ansonia found in previous studies.
Within Clade One Ansonia sp. Usun Apau and the enigmatic
Ansonia torrentis are part of a monophyletic group of
the Bornean species Ansonia hanitschi, Ansonia minuta,
Ansonia platysoma, Ansonia spinulifer, Ansonia vidua, and
two additional undescribed taxa. This subclade must be considered
as the result of an on-island radiation in the complex
evolution of Ansonia. The new species is formally described
including the identification of diagnostic morphometric traits.
Ansonia sp. Usun Apau is endemic to two isolated mountain
ridges in central Sarawak and must be considered as a new
element of the unique diversity of the Bornean amphibian
fauna that is potentially threatened by habitat loss at least in
parts of its range.
An integrative taxonomic analysis is used to delimit and describe three new species of Pseudocalotoes from the sky island archipelago of the Banjaran (=mountain range) Titiwangsa of Peninsular Malaysia. Pseudocalotes drogon sp. nov., from Fraser’s Hill, Pahang is basal to the sister species P. larutensis from Bukit Larut, Perak in the Banjaran Bintang and the new species P. rhaegal sp. nov. from Cameron Highlands, Pahang. Pseudocalotes drogon sp. nov. is differentiated from all other species of Psuedocalotes by having the combination of a flat rostrum; seven postrostrals; an interparietal; 11 cir- cumorbitals; five canthals; 7–10 superciliaries; one scale between the rostral and nasal; nine supralabials; eight infralabi- als; 10 postnasal-suborbital scales; four postmentals; five or six sublabials; five or six chinshields; 47 smooth, wide, gular scales; weak transverse gular and antehumeral folds; two enlarged scales between the ear and eye; enlarged upper and low- er posttemporals; a single enlarged supratympanic; no enlarged postrictals; three large scales bordering the dorsal margin of the ear opening; large pretympanic scales; eight scales in the nuchal crest not separated by a gap; enlarged vertebral scales extending to the tip of the tail; keeled and non-plate-like scales on flanks; 51 midbody scales; midventrals smaller than dorsals; 19 subdigital lamellae on the fourth finger; 23 subdigital lamellae on the fourth toe; preaxial scales on third toe enlarged and spinose; subdigital lamellae not unicarinate; HW/HL 0.52; HL/SVL 0.31; no elbow or knee patches; and a male dewlap color of lime-green bearing a central yellow spot. Pseudocalotes rhaegal sp. nov. is differentiated from all other Psuedocalotes by having the combination of a convex rostrum; 6–8 postrostrals; an interparietal; nine or 10 circu- morbitals; five canthals; 7–10 superciliaries; one or two scales between the rostral and nasal scales; eight or nine suprala- bials; seven or eight infralabials; 11 or 12 postnasal-suborbital scales; four postmentals; four or five chinshields; 40–45 smooth, wide, gular scales; no transverse gular fold; a weak antehumeral fold; three or four enlarged scales between the ear and eye; an enlarged upper and lower posttemporal; an enlarged supratympanic; no enlarged postrictals; no large scales bordering the upper margin of the ear opening or in the pretympanic region; 6–8 enlarged nuchal crest scales not separated by a gap; enlarged vertebral scales extending to the base of the tail; weakly keeled, non-plate-like scales on the flanks; 52– 58 midbody scales; midventrals smaller than dorsals; 19–21 subdigital lamellae on the fourth finger; 22–26 subdigital la- mellae on the fourth toe; preaxial scales on the third enlarged and rounded; subdigital lamellae not unicarinate; HW/HL 0.50–0.54; HL/SVL 0.28–0.30; no elbow or knee patches; and female dewlap color yellow bearing a purple base. The analyses also indicated that the new species, P. viserion sp. nov. from Genting Highlands, Pahang in the southern section of the Banjaran Titiwangsa is the sister species of P. flavigula from Cameron Highlands 121 km to the north and can be separated from all other species of Psuedocalotes by having the combination of three postrostrals; 10 circumorbitals; four or five canthals; 5–7 superciliaries; rostral and nasals in contact; supralabials contacting the nasal; six or seven suprala- bials; six or seven infralabials; two or three postmentals; 47 or 48 smooth, flat, gular scales; three chinshields; weak trans- verse gular and antehumeral folds; two enlarged scales between the ear and eye; an enlarged upper and lower posttemporal; an enlarged supratympanic; no enlarged postrictals; 7–9 nuchal crest scales lacking gaps and not extending beyond midbody; weakly keeled and plate-like scales on the flanks; 35–38 midbody scales; ventrals smaller than dorsals; 22 or 23 subdigital lamellae on the fourth finger; 26 or 27 subdigital lamellae on the fourth toe; preaxial scales on the third toe not modified; subdigital scales not unicarinate; HW/HL 0.62; no white marking below the eye; dewlap in males yel- low; and no elbow or knee patches. Pseudocalotes rhaegal sp. nov. most likely occurs in syntopy with P. flavigula in Ta- nah Rata at Cameron Highlands and its discovery adds to a growing body of literature detailing the recent descriptions of several new, upland, closely related, sympatric species in Peninsular Malaysia. Another new population referred to here as Pseudocalotes sp. nov. from the Hala-Bala Wildlife Sanctuary, Betong District, Yala Province, Thailand is discussed. The discovery and description of these three new Pseudocalotes from the upland regions of Peninsular Malaysia continues to underscore the remarkably high herpetological diversity and ecological complexity in this sky island archipelago that is still underestimated, unappreciated, and unprotected.
We present the latest version of the Molecular Evolutionary Genetics Analysis (MEGA) software, which contains many sophisticated
methods and tools for phylogenomics and phylomedicine. In this major upgrade, MEGA has been optimized for use on 64-bit computing
systems for analyzing bigger datasets. Researchers can now explore and analyze tens of thousands of sequences in MEGA. The
new version also provides an advanced wizard for building timetrees and includes a new functionality to automatically predict
gene duplication events in gene family trees. The 64-bit MEGA is made available in two interfaces: graphical and command line.
The graphical user interface (GUI) is a native Microsoft Windows application that can also be used on Mac OSX. The command
line MEGA is available as native applications for Windows, Linux, and Mac OSX. They are intended for use in high-throughput
and scripted analysis. Both versions are available from www.megasoftware.net free of charge.
The endangered frog, Odorrana ishikawae (Anura, Ranidae), is a species endemic to the mami and Okinawa Islands ofthe Ryukyu Archipelago, Japan. Segmentation of these islands has een considered to occur middle or upper Pleistocene. Our morphometric analyses revealed bvious differences between the Amami and Okinawa populations. Two distinct morphotypes were also recognized from the Amami Island (Amami common and Amami large types). Furthermore, the Amami and Okinawa populations could be distinguished clearly by coloration and dorsal uberculation. Based on 16SrRNA gene data, the Okinawa and Amami populations were phylogenetically separated but the genetic divergence (1.44-2.16%) was lower than the value suggested as species threshold in anurans (> 3% in 16S). Individuals of the Amami common and large types were nested within a single clade. Artificial hybridization experiments revealed normal hybrid viability between the two Amami types, with one exception. By contrast, between Okinawa females and two Amami type males, complete hybrid inviability was observed at early embryonic stages in the hybrids contrary to expectations from their low divergence in 16S. The reciprocal hybrids between two Amami type females and Okinawa males were viable, but spermatogenesis in the hybrid males showed some degree of abnormality. These results strongly indicate specific separation of the Amami population from the Okinawa population of O. ishikawae. Thus, we describe the Amami population as a new species, which is readily distinguishable from O. ishikawae by smaller ruggedly edged dorsal spots and an immaculate ventral surface.
Extensive clearing of Indonesian primary forests results in increased greenhouse gas emissions and biodiversity loss. However, there is no consensus on the areal extent and temporal trends of primary forest clearing in Indonesia. Here we report a spatially and temporally explicit quantification of Indonesian primary forest loss, which totalled over 6.02 Mha from 2000 to 2012 and increased on average by 47,600 ha per year. By 2012, annual primary forest loss in Indonesia was estimated to be higher than in Brazil (0.84 Mha and 0.46 Mha, respectively). Proportional loss of primary forests in wetland landforms increased and almost all clearing of primary forests occurred within degraded types, meaning logging preceded conversion processes. Loss within official forest land uses that restrict or prohibit clearing totalled 40% of all loss within national forest land. The increasing loss of Indonesian primary forests has significant implications for climate change mitigation and biodiversity conservation efforts.
Haplotype networks are an intuitive method for visualising relationships between individual genotypes at the population level. Here, we present popart, an integrated software package that provides a comprehensive implementation of haplotype network methods, phylogeographic visualisation tools and standard statistical tests, together with publication-ready figure production. popart also provides a platform for the implementation and distribution of new network-based methods – we describe one such new method, integer neighbour-joining. The software is open source and freely available for all major operating systems.
Four species of endemic Leptobrachium are known from Borneo, two lowland species L. kanowitense and L. abbotti, a montane species L. montanum, and a highland species L. gunungense. Of these, both L. montanum and L. abotti were found to contain several cryptic species by recent molecular studies. The population from Bario, Kelabit Highland of Sarawak, is one such cryptic species and was once called Lineage 2 of L. abbotti. Our morphological survey on this population proved that it has characteristics distinct from all other congeners, and therefore, we describe the Bario population as a new species, Leptobrachium kantonishikawai sp. nov. The new species is distinguished from putative topotypes of L. montanum and L. abbotti, as well as from L. gunungense by having a grayish brown abdomen, usually vermiculated, although sometimes spotted or blotched with white, and some unique morphometric characteristics.
The ubiquitous Mexican and Central American stream frogs allied to Eleutherodactylus rugulosus form a confusing spectrum of distinctive to subtly different populations. The condition of the male secondary sexual features: presence or absence of vocal slits and presence or absence of nuptial pads on the thumb, combined with the geographically consistent color (white, pale yellow, gold, orange, red or chestnut) of the venter of adults in life, provide the key to untangling the species problem in this group. For purposes of analysis the populations were grouped by the male secondary sexual features and compared in detail on the basis of 15 other characters of morphology and coloration. Twelve species are recognized within the rugulosus group, and may be placed in four series based on the presence (+) or absence (-) of vocal slits and nuptial pads in adult males. ++: Eleutherodactylus milesi of northern Honduras; E. merendonensis of northwestern Honduras; E. punctariolus of southern Costa Rica and western Panama; E. fleischmanni of Costa Rica; and E. escoces (sp. nov.) a bright red-bellied new species from the slopes (1100-2100 m) of Volcan Barba, Volcan Irazu and Volcan Turrialba of Costa Rica; +-: E. vocalis of northwestern Mexico; a new species, E. azueroensis (sp. nov.) from the Peninsula Azuero of western Panama; and E. taurus of the Golfo Dulce lowlands of southwestern Costa Rica and adjacent Panama; -+: E. matudai from the Pacific slopes of extreme southern Mexico and adjacent Guatemala; and a new species, E. angelicus, from the Cordillera de Tilaran and Volcan Poas in Costa Rica; --: E. brocchi of Alta and Baja Verapaz, Guatemala; and the wideranging lowland and slope species known from Mexico to western Panama, E. rugulosus. The Atlantic versant Mexican populations of this species are distinctive and have been variously recognized as a subspecies of E. rugulosus or as a separate species by previous authors. The earliest name for this population is Hylodes berkenbuschii Peters and E. natator Taylor and E. vulcani Shannon and Werler are strict synonyms. The seemingly allpatrically isolated southern populations of E. rugulosus in eastern and southwestern Nicaragua, Costa Rica and Panama are slightly distinct from the main population system of E. rugulosus. The earliest name for the southern stock is Lithodytes ranoides Cope with Liohyla pittieri Günther a strict synonym. Neither of these populations is recognized as separate from E. rugulosus. E. chiquito Lynch placed by its describer in the rugulosus group is a synonym of E. greggi of extreme southern Pacific slope Mexico and Guatemala, which is a member of the distantly related mexicanus group. Members of the rugulosus group fall into four geographic and ecologic distribution patterns: a) lowland and slope species, centered on the distribution of the wide-ranging E. rugulosus population system, with the allopatric E. vocalis on the northwest Mexican periphery, E. azueroensis on the southwest Panama periphery, the small population of E. merendonensis in northwestern Honduras and E. taurus occupying the Golfo Dulce lowlands of Costa Rica and western Panama, where E. rugulosus occurs only along the Pacific slope (600-1200 m) of the Talamanca-Chiriqui massif; b) E. brocchi and E. matudai in the highlands of southern Mexico and Guatemala; c) E. milesi in the uplands of northern Honduras; and d) E. angelicus, E. escoces, E. fleischmanni and E. punctariolus in the mountains of Costa Rica and western Panama. Verified cases of sympatry are known for E. rugulosus with E. matudai, with E. punctariolus and E. fleischmanni and E. punctariolus. An analysis of relationships and evolutionary trends indicates that the rugulosus group consists of four subgroups: the E. rugulosus subgroup in which males lack nuptial pads (E. azueroensis, E. taurus and E. vocalis, with vocal slits and E. brocchi and E. rugulosus, without vocal slits); the E. fleischmanni subgroup (E. angelicus, E. escoces, E. fleischmanni and E. punctariolus); the monotypic E. merendonensis subgroup; and the E. milesi subgroup (E. matudai and E. milesi). The latter three subgroups have nuptial pads in males. Within these lines vocal slits have been lost secondarily in E. matudai and E. angelicus. Of living forms E. vocalis most resembles the presumed ancestral stock of the group, that must have had a wide lowland range in Miocene. Evolution within the E. rugulosus subgroup involved fragmentation, modification and replacement in the lowlands during the remainder of Cenozoic, with E. brocchi evolving at a fairly late date in the Guatemala highland. Two other stocks seem to have diverged from the ancestral lineage by evolving nuptial pads in adult males, among other features: one in northern Central America to give rise to the specialized isolated E. merendonensis and the tuberculate highland E. milesi subgroup; a second in the mountains of Costa Rica and Panama to evolve into the E. fleischmanni group. Convergent evolution is found in each stock toward a highly specialized stream adapted form with increased webbing (E. merendonensis, E. punctariolus and E. taurus) and large toe disks in the former two, as modifications for life on boulders amid torrential racing water and splashing waterfalls. Three independent invasions of the uplands of Central America by members of the group, the E. milesi subgroup in northern Central America, the E. fleischmanni subgroup in Costa Rica and Panama and E. brocchi in Guatemala are responsible in large part for the species diversity within the rugulosus group.
The Thai population of Kalophrynus pleurostigma interlineatus is distinct from the Bornean population of K. p. pleurostigma in advertisement call structure, i.e., notes pulsed in the former but unpulsed in the latter. This difference suggests that the Thai population is reproductively isolated from, or uses different recognition signals from, the nominal subspecies and should be regarded as a full species, K. interlineatus (Blyth, 1855). Taxonomic status of the population from Java requires further investigation.
Advertisement call characteristics of Leptolalax heteropus from Peninsular Malaysia and L. gracilis, L. dringi, and two undescribed forms from Borneo are described. Leptolalax heteropus differs from the others in call characteristics, suggesting phylogenetic divergence; the call is short, lasting only 207-513 ms, and consists of a short series of 3-6 notes, the first note being longer in duration and higher in dominant frequency than subsequent ones. Calls of the remaining taxa are much longer, lasting 1.8-38 sec, and composed of a long series of 21-345 notes, each similar in duration and frequency within a call. However, dominant frequencies differ dramatically between species. Two new Bornean species, L. hamidi from Sarawak and L. arayai from Sabah, are described using acoustic and morphological characteristics.
The original description of the Bornean megophryid Leptolalax dringi was not sufficient to differentiate the species from its Bornean congeners. The species was later re-described but the redescription included characters from both type specimens and Leptolalax from other sites, including undescribed species. Analyses of the advertisement call of specimens assigned to L. dringi have been published but call descriptions differ markedly from each other. Moreover, published pictures of L. dringi are dubious in identification. Thus, the identity of L. dringi is enigmatic, hindering taxonomic work on Bornean Leptolalax. We herein provide detailed descriptions of the type series as well as recently obtained topotypic specimens. We also present the results of an analysis of the advertisement call of a male paratype. These data will facilitate future taxonomic work on Bornean Leptolalax.