Detailed aerobic and anaerobic bacteriological studies on the operative specimen from a patient with antibiotic-associated pseudomembranous colitis revealed approximately 10(11) facultatively anaerobic bacteria, but less than 10(5) obligate anaerobes per g of involved colonic tissue. Fourteen isolates of Escherichia coli, three isolates of Pseudomonas aeruginosa, and two isolates of Streptococcus fecalis were identified. The majority of the organisms were resistant to most of the antibiotics that the patient had received. Examinations of E. coli for heat-labile and heat-stable enterotoxins and tissue invasiveness were negative, and enterotoxin tests on the P. aeruginosa isolates were negative. Inoculation of mice with the bacterial isolates revealed no unusual pathogenicity. These findings suggest that antibiotic-associated colitis is associated with a marked loss in anaerobic colonic flora, but the colitis could not be explained by the presence of enterotoxins, tissue invasiveness, and pathogenicity of remaining microorganisms.