Kurixalus is a rhacophorid genus of tree frogs that are similar in morphology but vary in reproductive behavior. We investigated the cytogenetic features and genome size using conventional G-banding, C-banding and silver-staining techniques, fluorescence in situ hybridization (FISH), and flow cytometry in two representatives of Kurixalus (K. eiffingeri Boettger, 1895 and K. idiootocus Kuramoto and Wang, 1987) and compared the data between species and sex. The two Kurixalus species share a diploid chromosome number 2n = 26 and fundamental number FN = 52. Prominent differences between species were noted in the distribution of secondary constriction (SC)/nucleolus organizer region (NOR) and dense heterochromatin. Other interspecies differences including variations in the number of metacentric and submetacentric chromosomes and staining intensity of heterochromatin were also found. The cytogenetic results are consistent with the observed differences in their genome sizes. FISH with telomeric motif (TTAGGG)n for both species detected signals in the terminal regions. Intersex comparisons revealed no differences in terms of cytogenetic features and genome size in the two species. Despite the apparent highly conserved diploid chromosome number, data on the karyotype microstructure characterize the cytogenetic profile of the two Kurixalus species that contribute to clarification of the chromosomal homologies and the rearrangement mechanisms occurring during the karyotype evolution of Kurixalus. No heteromorphic chromosome pair in both species is consistent with the view that homomorphic sex chromosome is common in amphibians.