Abstract:[Objectives] This study aims to use molecular markers to investigate the genetic diversity and differentiation of the Chestnut Bulbul (Hemixos castanonotus), an omnivorous bulbul commonly found in the southern mountains of China. [Methods] In this study, the mitochondrial Cytochrome b (Cyt b) gene and seven other nuclear gene intron fragments with a concatenated length of 6 808 bp were used as molecular markers to analyze 11 populations from Guangdong, Guangxi, Hainan, Guizhou, and Jiangxi Provinces. We calculated population estimates, built haplotype networks, performed Bayesian analysis, and inspected the correlation between geographic distance and genetic divergence among populations. [Results] Based on Cyt b, 37 haplotypes were detected, of which 35.6% were presumed to be ancestral. Analysis of molecular variance showed that genetic variation was mainly within populations (79.77%). The results of Tajima’s D and Fu’s Fs neutral analysis supported H. castanonotus might have experienced population expansion (Appendix 1). The Bayesian Skyline Plot based on the nuclear genes suggested that H. castanonotus might have gone through population expansion between 5.3 to 3.7 Mya and 0.7 to 0.3 Mya (Fig. 5). Bayesian phylogenetic analysis of the Cyt b gene showed that the 11 geographic populations were divided into two clades (Fig. 2). The first clade contained only individuals from one population found in Hainan and are ascribed to H. c. castanonotus. The second clade contained individuals from the remaining 10 populations, no significant geographical structure was observed, and individuals in this clade are ascribed to H. c. canipennis. Haplotype network analysis also displayed similar results to the Bayesian phylogenetic analysis (Fig. 4). [Conclusion] Our findings suggest limited differentiation among H. castanonotus using the molecular markers studied, possibly due to insufficient isolation among populations or ancestral polymorphism in the markers. Further research is needed to better understand the demographic and evolutionary history of this species. This information is crucial to understand the underlying discrepancy between genetic and morphologic features in the avifauna evolution in southern China.