Plasmid-Encoded Metallo-β-Lactamase (IMP-6) Conferring Resistance to Carbapenems, Especially Meropenem

Abstract

In 1996, Serratia marcescens KU3838 was isolated from the urine of a patient with a urinary tract infection at a hospital in northern Japan and was found to contain the plasmid pKU501. Previously, we determined that pKU501 carries bla IMP and the genes for TEM-1-type ␤-lactamases as well as producing both types of ␤-lactamases (H. Yano, A. Kuga, K. Irinoda, R. Okamoto, T. Kobayashi, and M. Inoue, J. Antibiot. 52:1135-1139, 1999). pKU502 is a recombinant plasmid that contains a 1.5-kb DNA fragment, including the metallo-␤-lactamase gene, and is obtained by PCR amplification of pKU501. The sequence of the metallo-␤-lactamase gene in pKU502 was determined and revealed that this metallo-␤-lactamase gene differed from the gene encoding IMP-1 by one point mutation, leading to one amino acid substitution: 640-A in the base sequence of the IMP-1 gene was replaced by G, and Ser-196 was replaced by Gly in the mature enzyme. This enzyme was designated IMP-6. The strains that produced IMP-6 were resistant to carbapenems. The MICs of panipenem and especially meropenem were higher than the MIC of imipenem for these strains. The k cat /K m value of IMP-6 was about sevenfold higher against meropenem than against imipenem, although the MIC of meropenem for KU1917, which produced IMP-1, was lower than that of imipenem, and the MIC of panipenem was equal to that of imipenem. These results support the hypothesis that IMP-6 has extended substrate profiles against carbapenems. However, the activity of IMP-6 was very low against penicillin G and piperacillin. These results suggest that IMP-6 acquired high activity against carbapenems, especially meropenem, via the point mutation but in the process lost activity against penicillins. Although IMP-6 has reduced activity against penicillins due to this point mutation, pKU501 confers resistance to a variety of antimicrobial agents because it also produces TEM-1-type enzyme.Carbapenems such as imipenem, panipenem, and meropenem are the most potent agents for treatment of gramnegative infections due to the stability of these agents against the majority of ␤-lactamases and their high rate of permeation through bacterial outer membranes. In addition, imipenem, panipenem, and meropenem have a high affinity for penicillinbinding protein (PBP) 2 of gram-negative bacteria except for Pseudomonas aeruginosa. In P. aeruginosa, imipenem and panipenem have a high affinity for PBP 2 and meropenem has an affinity for PBP 3 (6, 19, 40).However, some clinical isolates have been reported to be resistant to carbapenems due to production of metallo-␤-lactamases. There have been increasing reports, especially from Japan (7,14,21,27,35), of gram-negative bacteria that carry the transferable carbapenem resistance gene bla IMP , including isolates of P. aeruginosa and Serratia marcescens. The metallo-␤-lactamases are class B enzymes in Ambler's molecular classification (1) and belong to Bush group 3 (3,31). Most of these metallo-␤-lactamases confer resistance not only to carbapenems but also to other ␤-lactams an...

Regional outbreak of CTX-M-2 β-lactamase-producing Proteus mirabilis in Japan

“…The ESBL types were determined by PCR using specific primers for bla TEM , bla SHV and bla CTX-M , using methods described elsewhere (Nagano et al, 2003). PCR experiments were also performed to detect the plasmid-encoded AmpC b-lactamases and metallo-b-lactamases according to a previously described method (Yano et al, 2001;Pérez-Pérez & Hanson, 2002). PCR was performed with HotStarTaq DNA polymerase (Qiagen) according to the manufacturer's instructions.…”

Prevalence of antibiotic resistance among clinical isolates of Kleibsiella pneumoniaeisolated from a Tertiary Care Hospital in Pakistan.

“…No doubt this represents very critical situation as compared to investigations from other regions of the world reporting resistance towards first, second and third generation antibiotics (Murray et al, 1990;Fish and Piscitell, 1995), however there are reports covering high levels of resistance of K. pneumoniae towards these antibiotics in many countries (Subha and Ananthan, 2002). This may be due to the production of ESBLs which cause the hydrolysis of β-lactam ring resulting in inactivation of cephalosporin and penicillin antibiotics (Yano et al, 2001) and overuse of cephalosporin antibiotics in developing nations (i.e. Pakistan) results in high levels of resistance.…”

Metalo-beta-lactamases

“…Essa enzima foi detectada em uma cepa clínica de Serratia marcescens isolada no Japão, a qual apresentava fenótipo de resistência a imipenem e cefalosporinas de espectro ampliado. Durante muitos anos a ocorrência de isolados produtores de IMP-1 foi restrita a esse país, entretanto, atualmente, a IMP-1 tem sido detectada em diferentes microrganismos, como Acinetobacter spp., Pseudomonas aeruginosa, Klebsiella pneumoniae isolados de diferentes regiões geográficas (11,20,21,28,29,43,61,69,82) . …”