The microflora of the bile and wall of the gallbladder was prospectively investigated in 104 nonselected consecutive patients treated with early cholecystectomy for acute cholecystitis after an average hospital stay of 1.8 days. The chief purpose was to relate the findings of cultures to duration of the illness. Special attention was paid to anaerobic isolation techniques. The cultures yielded 107 strains, representing 36 species, with overall agreement between four different sampling procedures. Aerobic gram-negative rods predominated, followed by streptococci and anaerobes (48, 31 and 15 per cent, respectively). The incidence of positive culture results (always greater than or equal to 10(6) colon forming units per milliliter) was 81 per cent among the patients who underwent operation within two days from the onset of symptoms and 50 to 65 per cent after longer preoperative intervals. The shorter interval was significantly more often associated with growth solely of anaerobes (p = 0.03). Postoperative sepsis was caused by biliary bacteria and not related to preoperative duration of illness. Appropriate perioperative antibiotic coverage significantly reduced sepsis--3 versus 20 per cent (p = 0.05).