The susceptibility of Chlamydia trachomatis to antibiotics was studied in vitro and in patients with nongonococcal urethritis. The minimal inhibitory concentrations of cloxacillin, erythromycin, pivampicillin, oxytetracycline, and doxycycline, which were the most effective drugs, were 10, 0.5, 0.25, 0.1, and 0.5 microgram/ml, respectively. The clinical observations suggested that of the antibiotics studied, doxycycline, erythromycin, and trimethoprim-sulfamethoxazole were effective for treatment of chlamydial infection and nongonococcal urethritis. Trimethoprim-sulfamethoxazole seemed less efficient for treatment of patients with nongonococcal urethritis whose cultures were chlamydia-negative. Pivampicillin, although it was active in vitro against C. trachomatis, did not produce a satisfactory clinical response. Untreated patients had symptoms of urethritis and chlamydial infection for three weeks. Incubation periods of a week or less were recorded for about half of the patients who had nongonococcal urethritis, both those whose cultures were chlamydia-positive and those whose cultures were chlamydia-negative. However, a few patients in both groups reported incubation periods of four weeks or more.