Conservation genomics of the endangered Burmese roofed turtle.

The Burmese roofed turtle (Batagur trivittata) is one of the world's most endangered turtles. Only one wild population remains in Myanmar. There are thought to be 12 breeding turtles in the wild. Conservation efforts for the species have raised >700 captive turtles since 2002, predominantly from eggs collected in the wild. We collected tissue samples from 445 individuals (approximately 40% of the turtles' remaining global population), applied double-digest restriction-site associated DNA sequencing (ddRAD-Seq), and obtained approximately 1500 unlinked genome-wide single nucleotide polymorphisms. Individuals fell into 5 distinct genetic clusters, 4 of which represented full-sib families. We inferred a low effective population size (≤10 individuals) but did not detect signs of severe inbreeding, possibly because the population bottleneck occurred recently. Two groups of 30 individuals from the captive pool that were the most genetically diverse were reintroduced to the wild, leading to an increase in the number of fertile eggs (n = 27) in the wild. Another 25 individuals, selected based on the same criteria, were transferred to the Singapore Zoo as an assurance colony. Our study demonstrates that the research-to-application gap in conservation can be bridged through application of cutting-edge genomic methods.

A little frog leaps a long way: compounded colonizations of the Indian Subcontinent discovered in the tiny Oriental frog genus Microhyla (Amphibia: Microhylidae).

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.

Last exit before the brink: Conservation genomics of the Cambodian population of the critically endangered southern river terrapin.

The southern river terrapin, Batagur affinis is one of the world's 25 most endangered freshwater turtle species. The major portion of the global population is currently found in peninsular Malaysia, with the only remnant Indochinese population in southern Cambodia. For more than a decade, wild nests in this remnant Cambodian population have been fenced and hatchlings reared in captivity. Here we amplified 10 microsatellite markers from all 136 captive individuals, obtained 2,658 presumably unlinked and neutral single nucleotide polymorphisms from 72 samples with ddRAD-seq, and amplified 784 bp of mtDNA from 50 samples. Our results reveal that the last Indochinese population comprised only four kinship groups as of 2012, with all offspring sired from <10 individuals in the wild. We demonstrate an obvious decrease in genetic contributions of breeders in the wild from 2006-2012 and identify high-value breeders instrumental for ex-situ management of the contemporary genetic stock of the species.

Near-complete extinction of native small mammal fauna 25 years after forest fragmentation.

Tropical forests continue to be felled and fragmented around the world. A key question is how rapidly species disappear from forest fragments and how quickly humans must restore forest connectivity to minimize extinctions. We surveyed small mammals on forest islands in Chiew Larn Reservoir in Thailand 5 to 7 and 25 to 26 years after isolation and observed the near-total loss of native small mammals within 5 years from <10-hectare (ha) fragments and within 25 years from 10- to 56-ha fragments. Based on our results, we developed an island biogeographic model and estimated mean extinction half-life (50% of resident species disappearing) to be 13.9 years. These catastrophic extinctions were probably partly driven by an invasive rat species; such biotic invasions are becoming increasingly common in human-modified landscapes. Our results are thus particularly relevant to other fragmented forest landscapes and suggest that small fragments are potentially even more vulnerable to biodiversity loss than previously thought.

An ancient origin for the enigmatic flat-headed frogs (Bombinatoridae: Barbourula) from the islands of Southeast Asia.

BACKGROUND: The complex history of Southeast Asian islands has long been of interest to biogeographers. Dispersal and vicariance events in the Pleistocene have received the most attention, though recent studies suggest a potentially more ancient history to components of the terrestrial fauna. Among this fauna is the enigmatic archaeobatrachian frog genus Barbourula, which only occurs on the islands of Borneo and Palawan. We utilize this lineage to gain unique insight into the temporal history of lineage diversification in Southeast Asian islands. METHODOLOGY/PRINCIPAL FINDINGS: Using mitochondrial and nuclear genetic data, multiple fossil calibration points, and likelihood and Bayesian methods, we estimate phylogenetic relationships and divergence times for Barbourula. We determine the sensitivity of focal divergence times to specific calibration points by jackknife approach in which each calibration point is excluded from analysis. We find that relevant divergence time estimates are robust to the exclusion of specific calibration points. Barbourula is recovered as a monophyletic lineage nested within a monophyletic Costata. Barbourula diverged from its sister taxon Bombina in the Paleogene and the two species of Barbourula diverged in the Late Miocene. CONCLUSIONS/SIGNIFICANCE: The divergences within Barbourula and between it and Bombina are surprisingly old and represent the oldest estimates for a cladogenetic event resulting in living taxa endemic to Southeast Asian islands. Moreover, these divergence time estimates are consistent with a new biogeographic scenario: the Palawan Ark Hypothesis. We suggest that components of Palawan's terrestrial fauna might have "rafted" on emergent portions of the North Palawan Block during its migration from the Asian mainland to its present-day position near Borneo. Further, dispersal from Palawan to Borneo (rather than Borneo to Palawan) may explain the current day disjunct distribution of this ancient lineage.