Characterization of the metallo-beta-lactamase determinant of Acinetobacter baumannii AC-54/97 reveals the existence of bla(IMP) allelic variants carried by gene cassettes of different phylogeny

The metallo-beta-lactamase determinant of Acinetobacter baumannii AC-54/97, a clinical isolate from Italy that was previously shown to produce an enzy me related to IMP-1, was isolated by means of a PCR methodology which targe ts amplification of gene cassette arrays inserted into class 1 integrons. S equencing revealed that this determinant was an allelic variant (named bla( IMP-2)) of bla(IMP) found in Japanese isolates and that it was divergent fr om the latter by 12% of its nucleotide sequence, which evidently had been a cquired independently. Similar to bla(IMP), bla(IMP-2) was also carried by an integron-borne gene cassette. However, the 59-base element of the bla(IM P-2) cassette was unrelated to those of the bla(IMP) cassettes found in Jap anese isolates, indicating a different phylogeny for the gene cassettes car rying the two allelic variants. Expression of the integron-borne bla(IMP-2) gene in Escherichia coli resulted in a significant decrease in susceptibil ity to a broad array of beta-lactams (ampicillin, carbenicillin, cephalothi n, cefoxitin, ceftazidime, cefepime, and carbapenems). The IMP-2 enzyme was purified from an Escherichia coli strain carrying the cloned determinant, and kinetic parameters were determined with several beta-lactam substrates. Compared to IMP-1, the kinetic parameters of IMP-2 were similar overall wi th some beta-Lactam substrates (cefoxitin, ceftazidime, cefepime, and imipe nem) but remarkably different with others (ampicillin, carbenicillin, cepha loridine, and meropenem), revealing a functional significance of at least s ome of the mutations that differentiate the tyro IMP variants. Present find ings suggest that the environmental reservoir of bla(IMP) alleles could be widespread and raise a question about the global risk of their transfer to clinically relevant species.