Colonization of the vaginal introitus with Enterobacteriaceae was cleared dramatically in women receiving trimethoprim-sulfamethoxazole for recurrent urinary tract infections. Vaginal fluid was collected from seven women; five of the seven had two vaginal fluid collections each while on full-dose therapy (total, 10 samples), and five had two or more collections each while on prophylactic, low-dose therapy (total, 14 samples). Bactericidal concentrations of trimethoprim were found in all samples of vaginal fluid, often exceeding the serum level by severalfold. Sulfamethoxazole was either undetectable or present only in fractional concentrations.