Males in polygyne populations of Solenopsis invicta are primarily sterile diploids and thought to not express the Gp-9 gene coding for a pheromone-binding protein affecting complex social behavior. We examined an aspect of the breeding system hitherto not considered-male Gp-9 genotypes in relation to sperm stored in queens. Four sites with varying frequencies of sympatric monogyne and polygyne colonies were sampled, including sexuals, workers, and broods from four colonies. Most queens were heterozygotes storing B sperm. Although predicted to be common, only 14 of 504 males were B or BB genotypes, suggesting strong selection. Increased frequency of polygyne colonies at each site paralleled increases in queens with b sperm (1.9-32.8%) and of noninseminated queens. The presence of both B and b sperm in 1.9-18.9% of queens, genotype profiles of colonies, and genotypes of offspring from individual queens suggest some frequency of multiple mating. The bb genotype, rather than an obligate, developmental lethal, was present in some queens and common in alates, workers, and brood. Selective mortality of sexuals may affect multiple aspects of the breeding system, including female-mediated dispersal, mating success, and gene flow. E USOCIAL Hymenoptera with multiple queens present a particular challenge to kin selection theory and the evolution of social systems insofar as an additional layer of relatedness dynamics (e.g., queens to queens, workers to queens) must be considered in formulating models of sociality, estimating costs and benefits, and predicting particular social outcomes (Ross and Keller 1995; Hannonen and Sundströ m 2003). The effect of polyandry has similar theoretical implications regarding the balance between reproductive cooperation and conflict (Ratnieks 1990;Keller 1995;Crozier and Pamilo 1996), although obligate polyandry is relatively rare in the social Hymenoptera (Strassmann 2001).Outside of honeybees, the red imported fire ant, Solenopsis invicta Buren, is perhaps the best-known and most intensely studied species of eusocial insect. Over the past 20 years, studies have provided a detailed description of the population genetic structure and breeding system of this species, demonstrating that complex social behavior can have a simple genetic basis and be affected by novel environmental/genetic contexts due