Parapenaeopsis coromandelica shrimp populations along the western coast of Sri Lanka have supported coastal trawling for the last hundreds of years. Two non-overlapping adjacent fishing grounds (Hendala and Negombo) sustain different fishing intensities. In order to obtain information on the population structure and genetic diversity of P. coromandelica in these two regions, differences in length–weight relationships, growth, spawning seasons, sex-ratios, gonadosomatic index, length at 50% maturity (L50) and sequence variation of the mitochondrial cytochrome oxidase subunit I were examined. Significant differences in population biology and genetic diversity were revealed from the two fishing grounds. Samples of Hendala showed lower L50 and genetic diversity which are considered as potential effects and symptoms of extensive selective harvesting. Further, the shrimps' behaviour seems to be triggering the separation through low mixing of individuals at the two fishing grounds resulting in significant divergence based on haplotype frequencies. Management of P. coromandelica should consider the revealed biological and genetic evidences on existence of two sub-populations/stocks together with a routine monitoring of genetic effects due to harvesting.