We studied the population structure of 3 species in the Myotis mystacinus complex (M. mystacinus, M. brandtii, and M. alcathoe) during the swarming period by sampling bats at 27 caves in the Carpathian Mountains and adjacent areas in southern Poland using 14 or 15 nuclear microsatellite loci. None of the examined species were in Hardy–Weinberg equilibrium following the global test for heterozygote deficiency. Inbreeding coefficient (F_IS) values at the species level also were statistically greater than zero. This may indicate some level of inbreeding in all examined taxa although it seems to be relatively low because significant F_IS values were recorded in only 2 swarming sites of M. mystacinus, 2 sites of M. brandtii, and 3 sites of M. alcathoe. However, almost 10% of related individuals sharing 1 parent (i.e., half siblings) were found in the same swarming sites for M. mystacinus and M. brandtii, with fewer observed in M. alcathoe. At the population level, 4.1%, 5.9%, and 8.7% of individuals of M. mystacinus, M. brandtii, and M. alcathoe, respectively, were assigned as full siblings. These figures suggest the possibility of females mating selectively with the same male in more than 1 year.