Melanoplus lakinus (Scudder) is one of the most abundant grasshopper species in the Mexican state of Durango. This state covers a wide semi-desert region of large grassland areas with much biotic diversity, specific orography, and contrasting environmental factors that may affect the genetic structure of the species. Genetics of Melanoplus lakinus populations from seven Durango locations with different biotic and environmental characteristics were assessed. The grassland locations had different plant diversities (ranging from an H' of 1.34 to 3.04) and different proportions of grass. They also have different environmental and orographic conditions. Sixty M. lakinus specimens collected among the locations were analyzed using microsatellites. Three microsatellite markers (OD4, OD7, and OD12) were selected from 11 reported primers. The characterization of loci revealed an average of 28 alleles per locus (range 74–390 bp). The probability of combined non-exclusion was 0.9999, with a mean null allele frequency of 0.1498 and mean expected heterozygosity of He = 0.754 ± 0.0283 (range 0.300–0.962). Genetic diversity among the sampled locations was assessed using Fisher's exact probability test, revealing large values (F_st = 0.263, p = 0.022) that indicated differences in allele frequency among populations. Cluster analysis revealed the presence of three grasshopper groups without an indication of isolation because of distance among them (Mantel test p = 0.160). Canonical analysis demonstrated that differences among environmental variables were correlated with genetics of the M. lakinus population (R = 0.989, p = 0.020). The genetic diversity, as indicated by allelic frequencies in the M. lakinus population, correlated with weather variables. These observations may help elucidate M. lakinus population dynamics in Durango State, Mexico.