Seasonal variations of insect population sizes are often dramatic, particularly in temperate regions and at altitudes where the climatic conditions are unfavorable to insect development during the winter. Decline of population size (or bottlenecks) and founder events may reduce the genetic variability and may create genetic differentiation between populations by drift and founder effects, but this reduction of genetic diversity is strongly influenced by gene flow between populations. In this study, we determined the population genetic structure for two stomoxyine species (Diptera: Muscidae), Stomoxys calcitrans (L.) and Stomoxys niger niger Macquart, which co-occur in dairy barns along an altitudinal gradient on La Réunion island. Using microsatellite markers, we quantified the genetic variation within and among populations for different altitudes. This study displays that, contrary to expectations, genetic diversity is not correlated with altitude and that genetic differentiation is not larger among high-altitude populations than among low-altitude populations. These results attest to the small drift and founder effects in high-altitude populations despite drastic decreases in population size during the winter. Furthermore, at the island scale, the populations of S. calcitrans were slightly differentiated, but those of S. niger niger were not. Together, the results revealed large levels of gene flow on La Réunion Island despite the dramatic geographic barriers, and they emphasize the importance of considering agricultural practices to restrict the dispersal of stomoxyines.