Partial sequences of mitochondrial ATP6 and ND4 gene were determined by PCR direct sequencing from 8 wild Hemibarbus labeo populations. The length of the amplified fragments of the two genes was 1 629 bp in total, with mutation rate of 4.97% and nucleotide diversity of 0.007 5. The results indicated that the wild Hemibarbus labeo populations maintained a high genetic diversity. Twenty one haplotypes were detected from 8 populations among which 45.24% of the individuals belong to HQ1 and HT2 haplotypes. Network of haplotypes showed that HQ1 and HT2 were centered by other haplotypes, which indicated that HQ1 and HT2 were the ancestor haplotypes and other specific haplotypes were formed during the evolution. AMOVA, FST and genetic distance analysis indicated that the difference of 8 populations from different river systems reached a significant level. Phylogenetic tree based on haplotypes revealed that most of the Hemibarbus labeo populations from Wusulijiang River and Heilongjiang River clustered together, and the populations from Yalujiang River clustered alone while populations from Yangtze River and Jiayin region of Heilongjiang River clustered together.