When fresh, epididymal mouse spermatozoa were incubated with viable Mycoplasma pulmonis, the mycoplasma adhered to the heads and tails of spermatozoa. Spermatozoa that had been preincubated with M. pulmonis in phosphate-buffered saline (PBS) and spermatozoa incubated in PBS alone were inseminated into the uterine horns of mice induced to superovulate. The mycoplasma treatment resulted in a reduced rate of fertilization and a decrease in the number of spermatozoa associated with eggs recovered from the oviduct at 5--18 h after ovulation. The percentage of spermatozoa recovered from the oviduct with adherent mycoplasmas, as determined by fluorescence microscopy, was lower than that seen in the uterus. Mycoplasma treatment of spermatozoa inseminated directly into the ovarian bursa did not result in a reduced fertilization rate. The results suggest that M. pulmonis adversely affects sperm transport through the female reproductive tract in the mouse.