Multiresistant plasmids from Pseudomonas aeruginosa highly resistant to either or both gentamicin and carbenicillin

High level resistance to gentamycin and carbenicillin was found in 30 and 10.7%, respectively, of Pseudomonas aeruginosa strains, especially in isolates from urine. In 23 out of 25 strains tested, these resistances were R mediated and linked to multiresistant plasmids, carrying genes for resistances to 5 other aminoglycosides, tobramycin, kanamycin, neomycin, streptomycin, and spectinomycin, and for resistances to chloramphenicol, tetracycline, sulfonamides, and mercury chloride. Carbenicillin resistance was unstable in Pseudomonas, and in its presence the multiresistant plasmids had a host range extended to the Enterobacteriaceae (group I plasmids). Otherwise they were transferable intragenerically only (group II plasmids). The extended host range plasmids were, as a rule, in fi- incompatibility class A-C. Segregants incompatible with both class A-C and P plasmids were detected. The β lactamase specified by the carbenicillin marker was of the TEMlike type. Multiple linkages of resistance determinants to the aminoglycosides were concomitantly present in most of the plasmids. Results from the bioassay indicated the presence of at least two aminoglycoside inactivating enzymes

Purification and properties of two gentamicin-modifying enzymes, coded by a single plasmid ppk237 originating from pseudomonas aeruginosa

A broad host range multiresistance plasmid pPK237, originating from Pseudomonas aeruginosa mediates high-level resistance to gentamicin and tobramycin. It was found to code for two gentamicin modifying enzymes, which from theirsubstrate profile by radioenzymatic assay were characterized as aminoglycoside acetyltransferase AAC(3)-I and aminoglycoside adenylyltransferase AAD(2”). The two enzymes were studied after purification from an Escherichia coli K12 host. The two gentamicin-modifying enzymes coded by pPK237 were completely separated by DEAE chromatography. The purification (126 fold) of the acetyltransferase was achieved by (NH4)2SO4precipitation, DEAE chromatography and affinity chromatography. The purification of the adenylyltransferase was performed by affinity chromatography directly after (NH4)2SO4precipitation. Both purified enzyme preparations showed a single protein band on disc electrophoresis. The Km for gentamicin C1of the acetyltransferase was 0.066 mM. The amino acid analysis of the acetyltransferase coded by pPK237 showed a different aminoacid composition than that of the gentamicin acetyltransferase AAC(3)-I purified by williams and northrop17). Theacetyltransferase after DEAE chromatography is stable for many months at — 20°C, while the adenylyltransferase after purification is highly unstable; it shows enzymatic activity only in the presence of Mg++. © 1982, JAPAN ANTIBIOTICS RESEARCH ASSOCIATION. All rights reserved