Relatedness is a central parameter in the evolution of sociality, because kin selection theory assumes that individuals involved in altruistic interactions are related. At least three reproductive characteristics are known to profoundly affect colony kin structure in social insects: the number of reproductive queens per colony, the relatedness among breeding queens and queen mating frequency. Both the occurrence of multiple queens (polygyny) and multiple mating (polyandry) decrease within-colony relatedness, while mating among sibs increases relatedness between the workers and the brood they rear. Using DNA microsatellites, we performed a detailed genetic analysis of the colony kin structure and breeding system in three ant species belonging to the genus Plagiolepis: P. schmitzii, P. taurica and P. maura. Our data show that queens of the three species mate multiply: queens of P. maura mate with 1–2 males, queens of P. taurica with 3–11 males and queens of P. schmitzii may have 1–14 different mates. Moreover, colonies are headed by multiple queens: P. taurica and P. maura are facultatively polygynous, while P. schmitzii is obligately polygynous. Despite polyandry and polygyny, relatedness within colonies remains high because all species are characterized by sib-mating, with a fixation index Fit = 0.25 in P. taurica, 0.24 in P. schmitzii and 0.26 in P. maura, and because the male mates of a queen are on average closely related.