Six Groups of the OXY β-Lactamase Evolved over Millions of Years in Klebsiella oxytoca

Abstract

The diversity and evolution of the class A OXY ␤-lactamase from Klebsiella oxytoca were investigated and compared to housekeeping gene diversity. The entire bla OXY coding region was sequenced in 18 clinical isolates representative of the four K. oxytoca ␤-lactamase gene groups bla OXY-1 to bla OXY-4 and of two new groups identified here, bla OXY-5 (with four isolates with pI 7.2 and one with pI 7.7) and bla OXY-6 (with four isolates with pI 7.75 and three with pI 8.1). Genes bla OXY-5 and bla OXY-6 showed 99.8% within-group nucleotide similarity but differed from each other by 4.2% and from bla OXY-1 , their closest relative, by 2.5% and 2.9%, respectively. Antimicrobial susceptibility to ␤-lactams was similar among OXY groups. Nucleotide sequence diversity of the 16S rRNA (1,454 bp), rpoB (940 bp), gyrA (383 bp), and gapDH (573 bp) genes was in agreement with the ␤-lactamase gene phylogeny. Strains with bla OXY-1 , bla OXY-2 , bla OXY-3 , bla OXY-4 , and bla OXY-6 genes formed five phylogenetic groups, named KoI, KoII, KoIII, KoIV, and KoVI, respectively. Isolates harboring bla OXY-5 appeared to represent an emerging lineage within KoI. We estimated that the bla OXY gene has been evolving within K. oxytoca for approximately 100 million years, using as calibration the 140-million-year estimation of the Escherichia coli-Salmonella enterica split. These results show that the bla OXY gene has diversified along K. oxytoca phylogenetic lines over long periods of time without concomitant evolution of the antimicrobial resistance phenotype.Klebsiella oxytoca is an important opportunistic pathogen causing serious infections in hospitalized patients, including neonates (16,19,28). K. oxytoca is naturally resistant to aminoand carboxy-penicillins (21), a phenotype due to the constitutive expression of a chromosomal class A ␤-lactamase (1), first called K1 (7,20) or KOXY (26) and now OXY (12). Due to the hyperproduction of the chromosomal ␤-lactamase, up to 10 to 20% of K. oxytoca strains (22) can show high-level resistance to certain expanded-spectrum cephalosporins (ceftriaxone and cefotaxime) and aztreonam (8). Up mutations in the promoter sequence of the genes are responsible for this phenotype (9-11). Overproducers of OXY enzymes are commonly resistant to all combinations of ␤-lactams with ␤-lactam inhibitors (21).Sequence diversity of the K. oxytoca chromosomal ␤-lactamase gene and the existence of discrete groups of OXY enzymes have been described. Fournier et al. (12) found a variant (bla OXY-2 ) of the chromosomal ␤-lactamase gene that differed from bla OXY-1 (1) by 12.7% in nucleotide sequence. It was suggested, based on colony hybridization, that the ␤-lactamase genes of K. oxytoca could be classified into two groups (OXY-1 and OXY-2), each representing approximately half of the clinical isolates (12). Recently, Granier et al. (14) identified two additional sequence variants, defined as bla OXY-3 and bla , each found in a single strain so far. Gene bla shares 85 and 84% similarity with bla OXY-1 and bla OXY...

In vitro transference and molecular characterization of bla TEM genes in bacteria isolated from Portuguese ready-to-eat foods

“…The five amplicons all from K. oxytoca corresponded to bla OXY genes. In fact, in this wild-type species, this gene is chromosomally located and constitutively expressed at low levels, which usually confers low-level resistance to amino-penicillins but no significant resistance to other b-lactams (Livermore 1995;Fevre et al 2005). This may explain the resistance phenotype obtained in this study only for AMP, an amino-penicillin (data not shown).…”

“…As expected, the bla TEM was not detected in the unique Klebsiella oxytoca isolated from cheese (isolate 138Q), as K. oxytoca usually carries the bla OXY (Fevre et al 2005). Eighty-five percent (17/20) of the meat isolates transferred the bla TEM gene through transconjugation.…”

Identification of Clinical Isolates of Indole-Positive and Indole-Negative Klebsiella spp

“…Based on nucleotide variations of the gyrA, parC, and rpoB genes, clinical isolates of Klebsiella pneumoniae fall into four phylogenetic groups, named KpI, KpII-A, KpII-B, and KpIII (6,7,17), with the newly described species K. variicola appearing to correspond to KpIII (40). In addition, Klebsiella oxytoca has been shown to include five distinct phylogenetic lines (KoI, KoII, KoIII, KoIV, and KoVI), which can be identified by 16S rRNA gene, rpoB, gyrA, gapDH, and bla OXY sequencing (18). The correct identification of Klebsiella species is not easily accomplished in most clinical microbiology laboratories, because several species share a similar biochemical profile.…”

Pesquisa de genes de resistência a antimicrobianos em filés de tilápia comercializados no município de São Paulo-SP

“…Na aquicultura o uso de antibióticos, principalmente de forma profilática, (FEVRE et al, 2005). Apesar disso, a resistência conferida por esse gene é cromossômica, reduzindo desta forma a possibilidade de transferência de material genético (FEVRE et al, 2005).…”

“…Apesar disso, a resistência conferida por esse gene é cromossômica, reduzindo desta forma a possibilidade de transferência de material genético (FEVRE et al, 2005).…”