To determine whether prolonged retention of diaphragms alters Staphylococcus aureus colonization of the lower genital tract, 183 nonpregnant, asymptomatic women were screened for the presence of this pathogen in the cervix and upper vagina. Nineteen (10.4%) had positive cultures: 5 from the vagina, 8 from the cervix, and 6 from both sites. A second examination with quantitation revealed the presence of S. aureus in 7 of the 19 patients. In five of the six available patients, the number of staphylococci increased significantly after prolonged (24 hour) retention of diaphragms and decreased in one subject only. In two additional patients S. aureus was recovered after, but not before, the use of diaphragms. An increase in the number of other microorganisms also was noted after prolonged retention of diaphragms. In vitro study of the spermicide nonoxynol-9 revealed that it had no inhibitory activity on staphylococci or group B streptococci but was highly effective against Streptococcus pneumoniae.