To estimate the prevalence of Neisseria meningitidis relatively resistant to penicillin in the United States, antimicrobial susceptibility testing was performed on all US meningococcal isolates submitted to the Centers for Disease Control and Prevention in 1991, including isolates identified through population-based surveillance for invasive meningococcal disease in selected areas of the United States. Three of the 100 isolates tested had MICs of penicillin of 0.125 microgram/mL. All were serogroup B, beta-lactamase-negative, and unique by multilocus enzyme electrophoresis subtyping. None of the 3 patients had been treated solely with penicillin; all recovered completely. About 4% of the isolates obtained from the population-based surveillance system were relatively penicillin-resistant. Given the low prevalence and uncertain clinical significance of infection with these organisms, routine susceptibility testing of meningococcal isolates is not indicated at this time; however, continued surveillance is necessary to monitor trends in antimicrobial susceptibility of meningococci in the United States.