The multiple stages of derepression of the type I chromosomal beta-lactamase in Pseudomonas aeruginosa were examined. Mutants partially and fully derepressed for beta-lactamase were selected from a wild-type clinical isolate. An analysis of the beta-lactamase produced by these mutants and the induced wild type revealed significant differences in the products of derepression at each stage. Beta-lactamase produced by the fully derepressed mutant showed a lower affinity (Ki,m 0.113 mM) for cephalothin than that produced by the partially derepressed mutant (Km, 0.049 mM). However, due to a very large Vmax, the former possessed a much greater hydrolytic efficiency. Differences in substrate profile were also noted. Only beta-lactamase from the fully derepressed mutant hydrolyzed cefamandole, cefoperazone, and cefonicid. The partially derepressed mutant possessed a single beta-lactamase band with a pl of 8.4. The fully derepressed mutant possessed this band and an additional major band with a pl of 7.5. Induction of the wild type with cefoxitin produced both bands. The changes in physiologic parameters of the enzymes produced in the different stages of derepression suggest a complex system for beta-lactamase expression in P. aeruginosa. This may involve at least two distinct structural regions, each of which is under control of the same repressor.

Expression of chromosomal ,B-lactamase was examined in 85 clinical isolates of Pseudomonas aeruginosa.P-Lactamase assays with and without cefoxitin induction revealed four phenotypes of enzyme expressioll: low basal, inducible; moderate basal, inducible; moderate basal, constitutive; and high basal, constitutive. The isoelectric points of the major ,-lactamase bands were 9.4, 9.2, and 8.4. These results indicate that there is a limited heterogeneity in expression of chromosomal P-lactamase of P. aeruginosa.Virtually every strain of Pseudomonas aeruginosa produces a chromosomal P-lactamase that belongs to Richmond-Sykes class I (10). Under normal conditions, this enzyme is expressed at a very low basal level; however, it can be induced to much higher levels with such compounds as benzylpenicillin, cefoxitin, or imipenem (1, 6, 9-11, 13). Mutants of P. aeruginosa altered in their expression of class I 3-lactamase have also been described. These include mutants partially or fully derepressed for enzyme expression, those with little or no enzyme, and mutants expressing moderate levels of ,-lactamase constitutively (2-6, 11, 14). These altered states of enzyme expression are often associated with changes in susceptibility to various 3-lactam antibiotics. To date, most studies concerning the production single patient during a course of antimicrobial therapy. A second isolate fromn the same patient was included only if it was altered in its expression of ,-lactamase in comparison to the first isolate or was a different strain. Strain identity was based upon serotype (serotyping was kindly performed by Charles Zierdt, National Institutes of Health, Bethesda, Md.).,-Lactamase expression in each of the 85 isolates was examined in sonic extracts prepared with or without 2 h of induction with 100 ,ug of cefoxitin per ml (3). The isoelectric point (pI) of each enzyme. was determined before and after induction as described previously (12). In each isoelectric focusing run, controls were included to mnonitor run-to-iruri variations. These controls consisted of sonic extracts pre-