Mahmudul Hasan

Genetic Divergence and Reproductive Isolation in the Genus Fejervarya (Amphibia: Anura) from Bangladesh Inferred from Morphological Observations, Crossing Experiments, and Molecular Analyses

Abstract In the present study, morphological examinations, crossing experiments and molecular analyses were performed to elucidate the degree of genetic divergence and phylogenetic relationships within the genus Fejervarya from Bangladesh and other Asian countries. Morphological characteristics revealed that Fejervarya species from Bangladesh were divided into four distinct groups: large, medium, small, and mangrove types. Crossing experiments indicated the involvement of three reproductive isolating mechanisms: gametic isolation between the large type and mangrove type, hybrid inviability between the large type and two other types, and hybrid sterility between the medium and small types. Experimental results also indicated that these four types of frogs merit the status of individual species of Fejervarya. Molecular analyses based on mtDNA gene sequences showed that the Bangladesh Fejervarya species were largely divided into three groups: the mangrove type, large type, and others, with the last further subdivided into the medium and small types. Comparison with other Asian Fejervarya species revealed that the Bangladesh mangrove type (which resembled F. cancrivora in morphology) was closely related to F. cancrivora from India, Thailand, and the Philippines; the large type belonged to the F. iskandari group and closely resembled F. orissaensis; the small type was included in the South Asian or Indian group, and was closest to F. syhadrensis from India and Sri Lanka, whereas the medium type was most closely related to F. limnocharis from Myanmar among all described species of this genus.

Cryptic Anuran Biodiversity in Bangladesh Revealed by Mitochondrial 16S rRNA Gene Sequences

To survey the diversity of anuran species in Bangladesh, we compared mitochondrial 16S rRNA gene sequences (approximately 1.4 kbp) from 107 Bangladesh frog specimens. The results of genetic divergence and phylogenetic analyses incorporating data from related species revealed the occurrence of at least eight cryptic species. Hoplobatrachus tigerinus from two districts diverged considerably, indicating the involvement of a cryptic species. Two Fejervarya sp. (large and medium types) and Hylarana cf. taipehensis formed lineages distinct from related species and are probably new species. Microhyla cf. ornata differed from M. ornata with respect to type locality area and involved two distinct species. In addition, we found that Hylarana sp. and Microhyla sp. did not match congeners examined to date in either morphology or 16S rRNA sequence. The occurrence of M. fissipes was tentatively suggested. Consequently, at least, 19 species were found from Bangladesh in this study. These findings revealed a rich anuran biodiversity in Bangladesh, which is unexpected considering the rather simple topographic features of the country.

A new species of genus Hoplobatrachus (Anura, Dicroglossidae) from the coastal belt of Bangladesh

A new cryptic species of the genus Hoplobatrachus from Cox’s Bazar district of Bangladesh is described and compared with its relevant congeners both in morphology and mitochondrial gene sequences. The new species differs from its close relative H. tigerinus in having a distinct broad black band from the eye, through the nostrils, to the anterior edge of the upper jaw, another black band along the lateral margin of the upper jaw, and a narrow inter-orbital distance relative to eyelid width and inter-nostril distance. Advertisement calls of the new species are similar to those of H. tigerinus but differ in dominant frequency and number of pulses. Based on mitochondrial DNA sequence data, this species was proved to genetically divergent from H. tigerinus at 3.2% for the 16S rRNA gene and 14.2% for the Cytb gene. The known distribution range of the new species is restricted to the southeastern corner of Bangladesh and it seems to be endemic in this coastal belt.

Genetic divergences of South and Southeast Asian frogs: a case study of several taxa based on 16S ribosomal RNA gene data with notes on the generic name Fejervarya

Mahmudul HASAN, Mohammed Mafizul ISLAM, Md. Mukhlesur Rahman KHAN, Takeshi IGAWA, Mohammad Shafiqul ALAM, Hon Tjong DJONG, Nia KURNIAWAN, Hareesh JOSHY, Yong Hoi SEN, Daicus M. BELABUT, Atsushi KURABAYASHI, Mitsuru KURAMOTO, Masayuki SUMIDA* Institute for Amphibian Biology, Graduate School of Science, Hiroshima University, Higashihiroshima, Japan Department of Fisheries Biology and Genetics, Faculty of Fisheries, Bangladesh Agricultural University, Mymensingh, Bangladesh Department of Genetics and Fish Breeding, Faculty of Fisheries, Bangabandhu Sheikh Mujibur Rahman Agricultural University, Gazipur 1706, Bangladesh Department of Biology, Faculty of Mathematics and Science, Andalas University, Padang, West Sumatra, Indonesia Department of Biology, Faculty of Mathematics and Science, Brawijawa University, Malang, East Java, Indonesia Laboratory of Applied Biology, St. Aloysius College, Mangalore, Karnataka, India Institute of Biological Sciences, Faculty of Science, University of Malaya, Kuala Lumpur, Malaysia 3-6-15 Hikarigaoka, Munakata, Fukuoka, Japan

Postmating Isolation in Six Species of Three Genera (Hoplobatrachus, Euphlyctis and Fejervarya) from Family Dicroglossidae (Anura), with Special Reference to Spontaneous Production of Allotriploids

In light of reproductive isolation being a fundamental aspect of the biological species concept, we performed crossing experiments using six species from three genera (Hoplobatrachus, Euphlyctis and Fejervarya) of family Dicroglossidae to explore postmating isolation in dicroglossid frogs. Our results revealed gametic isolation among these genera, although the intergeneric hybrids between female E. cyanophlyctis and male H. chinensis were not viable at the tadpole stage, while the hybrids between female E. cyanophlyctis and male H. tigerinus were inviable at the hatching stage. These results showed complete hybrid inviability between the two genera. Almost all interspecific hybrids between female H. tigerinus and male H. chinensis died of underdevelopment at the tadpole stage, whereas several hybrids developed normally and survived to maturity. Chromosomal observations and mtDNA and allozyme analyses confirmed that these mature hybrids were allotriploid, with two maternal genomes and one paternal genome. The present results suggest that the allotriploids were produced spontaneously, and histological observations confirmed their sex as sterile males. We also investigated the molecular relationships between H. tigerinus, H. chinensis, and the interspecific allotriploids by mitochondrial Cytb, 12S and 16S rRNA gene analyses. The maternal inheritance mode of mitochondrial genomes was retained in the hybrids. Finally, the present results suggest that the degree of postmating isolation reflects phylogenetic relationship. In addition, we speculate that allotriploids may be produced via hybridization among cryptic species.