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

Abstract

Frogs of the genus <i>Microhyla</i> 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 <i>Microhyla</i> remains one of the taxonomically most challenging groups of Asian frogs and was found to be paraphyletic with respect to large-sized fossorial <i>Glyphoglossus</i>. In this study we present a time-calibrated phylogeny for frogs in the genus <i>Microhyla</i>, 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 <i>Microhyla</i> species and several undescribed candidate species. Phylogenetic analyses of 3,207 bp of combined mtDNA and nuDNA data recovered three well-supported groups: the <i>Glyphoglossus</i> clade, Southeast Asian <i>Microhyla</i> II clade (includes <i>M. annectens</i> species group), and a diverse <i>Microhyla</i> I clade including all other species. Within the largest major clade of <i>Microhyla</i> are seven well-supported subclades that we identify as the <i>M. achatina</i>, <i>M. fissipes</i>, <i>M. berdmorei</i>, <i>M. superciliaris</i>, <i>M. ornata</i>, <i>M. butleri</i>, and <i>M. palmipes</i> species groups. The phylogenetic position of 12 poorly known <i>Microhyla</i> species is clarified for the first time. These phylogenetic results, along with molecular clock and ancestral area analyses, show the <i>Microhyla-Glyphoglossus</i> 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 <i>Glyphoglossus</i> and <i>Microhyla</i> II remained within their ancestral ranges, <i>Microhyla</i> I expanded its distribution generally east to west, colonizing and diversifying through the Cenozoic. The Indian Subcontinent was colonized by members of five <i>Microhyla</i> 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 <i>Microhyla</i> have independently achieved extreme miniaturization with adult body size below 15 mm. Three of the five smallest <i>Microhyla</i> species are obligate phytotelm-breeders and we argue that their peculiar reproductive biology may be a factor involved in miniaturization. Body size increases in <i>Microhyla-Glyphoglossus</i> 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 <i>Microhyla</i> and reveal 15-33 undescribed species. We revalidate <i>M. nepenthicola</i>, synonymize <i>M. pulverata</i> with <i>M. marmorata</i>, 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.