Anopheles stephensi Liston s.l. (Diptera: Culicidae) is one of the major vectors of malaria in Pakistan, India, Iran and Afghanistan. In parts of its range this species has shown increases in both relative and absolute abundance in what is hypothesized to be a response to human-mediated environmental change resulting from extensive irrigation. We attempted to detect the molecular genetic signatures of this population instability based on three samples obtained from two villages (149/6R and 111/6R) within an irrigation zone in Punjab Province and from one village (Azakhel) outside the irrigation scheme in Northwest Frontier Province (NWFP), Pakistan, using seven microsatellite loci and 682 basepairs of the mitochondrial CO1 gene. For microsatellite loci, high levels of genetic diversity were observed within populations (mean alleles per locus 10.71-11.57; mean heterozygosity 0.703-0.733). Deviation from Hardy-Weinberg expectations was observed for only two microsatellite loci in 21 tests. No genetic differentiation was observed between populations and average pairwise F(ST) values did not differ significantly from zero for any population pair or either marker system. Tests of population expansion for both mitochondrial and microsatellite loci were inconclusive.