A multi-resistant clinical isolate of Citrobacter diversus, particularly resistant to penicillins and cephalosporins, has been studied. The strain produced constitutively high levels of beta-lactamase which efficiently hydrolysed cephaloridine but not ampicillin. The apparent Km value for cephaloridine (125 +/- 25 microM) was within the range usually observed for beta-lactamases in similar bacteria. The strain possessed two forms of the enzyme as indicated by isoelectric focusing patterns, HPLC analysis and two pH optima. SDS-PAGE analysis indicated a single protein responsible for both forms. The strain possessed a single plasmid, as revealed by agarose gel electrophoresis, but beta-lactamase production was not plasmid encoded. The available data indicate the beta-lactamase(s) of C. diversus ULA27 to be chromosomally encoded.