Adult mosquitoes of the Anopheles quadrimaculatus complex were collected from Montgomery County, AL (MON) and Alachua County, FL, (KBG) and laboratory stocks of species A and B were established through a selection procedure employing isofemale lines. Progeny from a cross of species B females to ORL males were usually semisterile females and sterile males. Progeny of the reciprocal cross were also semisterile females and sterile males, but the sex ratio was variable and ranged from normal to no males because of male mortality during the pupal stage. Conspecific crosses between strains from the two locations resulted in fertile offspring. Crosses between the sibling species from the two locations invariably gave semisterile females and sterile males (or lethal effects). This evidence confirmed previous data from hybridization and electrophoretic analyses of field populations indicating that Anopheles quadrimaculatus is a species complex.