CHARACTERIZATION OF A NOVEL EXTENDED-SPECTRUM BETA-LACTAMASE FROM PSEUDOMONAS-AERUGINOSA

A clinical isolate of Pseudomonas aeruginosa RNL-1 showed resistance t o extended-spectrum cephalosporins which was inhibited by clavulanic a cid. Although this strain contained three plasmids ca. 80, 20, and 4 k b long, the resistance could not be transferred by mating-out assays w ith P. aeruginosa or Escherichia coli. Cloning of a 2.1-kb Sau3A fragm ent from P. aeruginosa RNL-1 into plasmid pACYC184 produced pPZ1, a re combinant plasmid that encodes a beta-lactamase. This beta-lactamase ( PER-1) had a relative molecular mass of 29 kDa and a pI of 5.4 and was biosynthesized by P. aeruginosa RNL-1 along with a likely cephalospor inase with a pI of 8.7. PER-1 showed a broad substrate profile by hydr olyzing benzylpenicillin, amoxicillin, ticarcillin cephalothin, cefope razone, cefuroxime, HR 221, ceftriaxone, ceftazidime, and (moderately) aztreonam but not oxacillin, imipenem, or cephamycins. V(max) values for extended-spectrum cephalosporins were uncommonly high, and the aff inity of the enzyme for most compounds was relatively low (i.e., high K(m)). PER-1 activity was inhibited by clavulanic acid, sulbactam, imi penem, and cephamycins but not by EDTA. A 1.1-kb SnaBI fragment from p PZ1 failed to hybridize with plasmids that encode TEM-, SHV-, OXA-, or CARB/PSE-type beta-lactamase or with the ampC gene of P. aeruginosa. However, the same probe appeared to hybridize with chromosomal but not plasmid DNA from P. aeruginosa RNL-1. This study reports the properti es of a novel extended-spectrum beta-lactamase in P. aeruginosa which may not be derived by point mutations from previously known enzymes of this species.