In the house fly, Musca domestica L. (Diptera: Muscidae), sex is usually determined by a dominant factor, M, located on the Y chromosome. However, there are autosomal male (A^M) populations in which the M factor is located on one or more of the five autosomes (I-V), most commonly on the third chromosome. Herein we report the use of isogenic strains to determine the relative fitness of Y^M versus III^M males in three different experiments. First, cages were started with 50% Y^M and 50% III^M males, and the frequencies of Y^M and III^M males were evaluated across generations. Second, mating competition studies were preformed with these isogenic strains. Third, the relative emergence rates of III^M versus Y^M male pupae held at three temperatures for 3 d were examined. All three studies indicate that III^M males have a greater fitness than Y^M males. In the cage competition studies, >90% of the males were III^M after seven generations. III^M males were more likely to mate than Y^M males, and a higher percent of III^M males emerged after being held as pupae at 4, 16, or 28°C for 3 d. The implications of these studies to the distribution of III^M and Y^M males in field populations are discussed.