Eutropiichthys vacha were collected from two rivers namely Ganga (Patna, India) and Kosi (Madhepura,India) for population genetic and phylogenetic studies. Five OPA primers were used to generate the fragment patterns from the samples collected. Polymorphisms within and between populations were assayed using 5 random primers, and 45 loci were amplified ranging from 250 to 2,000 bp. The percentage of polymorphic loci was found 51.1% and 55.6% for Ganga and Kosi populations, respectively. Total genetic diversity was 0.2173,and the average coefficient of genetic differentiation was 0.0958. The highest level of genetic diversities within population as well as lower between populations suggested that lower differentiation rate between populations. Gene flow between Ganga and Kosi populations was 4.7. Nei's unbiased measure of genetic identity and genetic distances of two populations were found 0.9606 and 0.0402, respectively. Phylogenetic analysis by RAPD showed one common cluster between two wild populations (Patna and Kosi) though they are quite distant from each other but belongs to same drainage system.