To evaluate the degree of postmating isolation and the evolutionary relationships among frog species in the genus Fejervarya from Indonesia (Lesser Sunda), Bangladesh, China, and Japan, crossing experiments and molecular phylogenetic analyses were carried out. Crossing experiments revealed that reciprocal hybrids among F. iskandari, F. verruculosa, and F. sp. large type and between F. multistriata and F. kawamurai are viable through metamorphosis, while those between the F. iskandari group and F. limnocharis group were completely or partially inviable at the tadpole stage and those between Southeast Asian and South Asian Fejervarya groups were completely inviable at the embryonic stage. The mature reciprocal hybrids between F. iskandari and F. verruculosa from Lesser Sunda, Indonesia, showed some degree of abnormality in spermatogenesis. In phylogenetic analyses based on mtDNA Cytb sequences, F. iskandari formed a sister clade with F. verruculosa from Lesser Sunda, Indonesia, with 8.1% sequence divergence. F. multistriata from China formed a clade with populations of F. limnocharis in Thailand, Malaysia, and Indonesia (topotype), and these taxa showed sister relationships to F. kawamurai from Japan with 8.9% sequence divergence. Fejervarya sp. small type from Bangladesh formed a clade with the other South Asian members of the Fejervarya group and formed a sister clade with the Southeast Asian Fejervarya group, with 23.1% sequence divergence in the Cytb gene. These results showed that the degree of postmating isolation reflects molecular phylogenetic relationships and that F. iskandari and F. verruculosa from Indonesia (Lesser Sunda) are reproductively isolated by abnormalities in spermatogenesis and show genetic differentiation.
To elucidate genetic divergence and evolutionary relationship in Fejervarya cancrivora from Indonesia and other Asian countries, allozyme and molecular analyses were carried out using 131 frogs collected from 24 populations in Indonesia, Thailand, Bangladesh, Malaysia, and the Philippines. In the allozymic survey, seventeen enzymatic loci were examined for 92 frogs from eight representative localities. The results showed that F. cancrivora is subdivided into two main groups, the mangrove type and the large- plus Pelabuhan ratu types. The average Nel's genetic distance between the two groups was 0.535. Molecular phylogenetic trees based on nucleotide sequences of the 16S rRNA and Cyt b genes and constructed with the ML, MP, NJ, and BI methods also showed that the individuals of F. cancrivora analyzed comprised two clades, the mangrove type and the large plus Pelabuhan ratu / Sulawesi types, the latter further split into two subclades, the large type and the Pelabuhan ratu / Sulawesi type. The geographical distribution of individuals of the three F. cancrivora types was examined. Ten Individuals from Bangladesh, Thailand, and the Philippines represented the mangrove type; 34 Individuals from Malaysia and Indonesia represented the large type; and 11 individuals from Indonesia represented the Pelabuhan ratu / Sulawesi type. Average sequence divergences among the three types were 5.78-10.22% for the 16S and 12.88-16.38% for Cyt b. Our results suggest that each of the three types can be regarded as a distinct species.