Resistance to  -lactam antibiotics in Bacteroides species

Nord, Carl Erik; Olsson‐Liljequist, Barbro

doi:10.1093/jac/8.suppl_d.33

Cited by 26 publications

(12 citation statements)

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“…The gene appears to be of Bacteroides origin, on the basis of its moles percent G+C content. We have also shown at the DNA level that cepA is specific to B. fragilis species, extending conclusions drawn by others on the basis of the presence of 1-lactamases with common isoelectric points (40).…”

Section: Discussionsupporting

confidence: 88%

Cloning and characterization of the endogenous cephalosporinase gene, cepA, from Bacteroides fragilis reveals a new subgroup of Ambler class A beta-lactamases

Rogers

Parker

Smith

1993

Antimicrob Agents Chemother

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Bacteroides frgigli& CS30 is a clinical isolate resistant to high concentrations of benzylpenicillin and cephaloridine but not to cephamycin or penem antibiotics. fl-Lactam resistance is mediated by a chromosomally encoded cephalosporinase produced at a high level. The gene encoding this 13-lactamase was cloned from genomic libraries constructed in Escherichia coli and then mated with B. fragilis 638 for identification of ampicillin-resistant (Apr) strains. Apr transconjugants contained a nitrocefin-reactive protein with the physical and enzymatic properties of the original CS30 isolate. The (3-lactamase gene (cepA) was localized by deletion analysis and subcloned, and its nucleotide sequence was determined. The 903-bp cepA open reading frame encoded a 300-amino-acid precursor protein (predicted molecular mass, 34,070 Da). A 13-lactamase-deficient mutant strain of B. fiugilis 638 was constructed by insertional inactivation with the cepA gene of CS30, demonstrating strict functional homology between these chromosomal 13-lactamase genes. An extensive comparison of the CepA protein sequence by alignment with other 13-lactamases revealed the strict conservation of at least four elements common to Ambler class A. A further comparison of the CepA protein sequence with protein sequences of (-lactamases from two other Bacteroides species indicated that they constitute their own distinct subgroup of class A 13-lactamases.Bacteroides fragilis is responsible for approximately half of all human anaerobic infections and is the most common anaerobe recovered from clinical specimens (12). Numerous reports of B. fragilis isolates resistant to a variety of P-lactam antibiotics indicate that these organisms are becoming increasingly refractory to treatment with these drugs. The primary mechanism of 1-lactam resistance in Bacteroides species is the production of ,B-lactamase (40). At least four types of P-lactamase have been described for members of the B. fragilis group, but the most common type is a constitutively produced, chromosomally encoded cephalosporinase having no activity against cefoxitin or imipenem.This "endogenous" P-lactamase is present in over 90% of clinical isolates tested (10). Unlike the class C chromosomally encoded P-lactamases of members of the family Enterobacteriaceae, the B. fragilis P-lactamase has an isoelectric point in the acid range and is susceptible to inhibition by clavulanic acid and sulbactam, placing it in group 2e in the Bush classification scheme (8).Regulation of the endogenous B. fragilis j-lactamase has not been extensively studied, but this enzyme may be growth rate regulated, with maximal activity occurring 3 h into the stationary phase (7). With regard to the production of the endogenous cephalosporinase, others have grouped B.fragilis clinical isolates into three expression classes (18). Low-level j-lactamase producers are susceptible to all 3-lactams, the MICs of benzylpenicillin and cephaloridine being <2 and <16 ,ug/ml, respectively. For intermediate-level 3-lactamase producers, th...

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Section: Discussionsupporting

confidence: 88%

Cloning and characterization of the endogenous cephalosporinase gene, cepA, from Bacteroides fragilis reveals a new subgroup of Ambler class A beta-lactamases

Rogers

Parker

Smith

1993

Antimicrob Agents Chemother

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“…These organisms are generally susceptible to some of the newer ,B-lactams such as cephamycins (cefoxitin) and carbapenems (imipenem), although newly acquired 3-lactamases capable of degrading these antibiotics have been described (18,21,38). The indigenous 3-lactamase is present in between 90 and 99% of B. fragilis strains and at the biochemical level, this P-lactamase has been shown to be species specific (8,16). Recently, the gene for this enzyme, cepA, was cloned and the nucleotide sequence was determined (27).…”

mentioning

confidence: 99%

“…The indigenous 3-lactamase is present in between 90 and 99% of B. fragilis strains and at the biochemical level, this P-lactamase has been shown to be species specific (8,16). Recently, the gene for this enzyme, cepA, was cloned and the nucleotide sequence was determined (27).…”

mentioning

confidence: 99%

Insertional activation of cepA leads to high-level beta-lactamase expression in Bacteroides fragilis clinical isolates

et al. 1994

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Bacteroides fragilis is an important opportunistic pathogen of humans and is resistant to many drugs commonly used to treat anaerobic infections, including P-lactams. A strain set comprised of B. fragilis isolates producing either low or high levels of the endogenous cephalosporinase activity, CepA, has been described previously (M. B. Rogers, A. C. Parker, and C. J. Smith, Antimicrob. Agents Chemother. 37:2391-2400.Clones containing cepA genes from each of seven representative strains were isolated, and the DNA sequences were determined. Nucleotide sequence comparisons revealed that there were few differences between the cepA coding sequences of the low-and high-activity strains. The cepA coding sequences were cloned into an expression vector, pFD340, and analyzed in a B.fragilis 638 cepA mutant. The results of j-lactamase assays and ampicillin MICs showed that there was no significant difference in the enzymatic activity of structural genes from the high-or low-activity strains. Comparison of sequences upstream of the cepA coding region revealed that 50 bp prior to the translation start codon, the sequence for high-activity strains change dramatically. This region of the high-activity strains shared extensive homology with IS21, suggesting that an insertion was responsible for the increased expression of cepA in these isolates. Northern (RNA) blot analysis of total RNA by using cepA-specific DNA probes supported the idea that differential cepA expression in low-and high-activity strains was controlled at the level of transcription. However, the insertion did not alter the cepA transcription start site, which occurred 27 bp upstream of the ATG translation start codon in both expression classes. Possible mechanisms of cepA activation are discussed.The anaerobe Bacteroides fragilis contains an endogenous, chromosomally encoded P-lactamase which preferentially hydrolyzes cephalosporins and is responsible for the intrinsic resistance to most penicillins and cephalosporins (6, 15). These organisms are generally susceptible to some of the newer ,B-lactams such as cephamycins (cefoxitin) and carbapenems (imipenem), although newly acquired 3-lactamases capable of degrading these antibiotics have been described (18,21,38).The indigenous 3-lactamase is present in between 90 and 99% of B. fragilis strains and at the biochemical level, this P-lactamase has been shown to be species specific (8,16). Recently, the gene for this enzyme, cepA, was cloned and the nucleotide sequence was determined (27). Southern hybridization analyses with a cepA probe showed that there was homology only with other B. fragilis strains, and construction of a cepA mutant provided evidence that this gene did in fact encode for the endogenous ,-lactamase. Comparison of the predicted CepA amino acid sequence with other 3-lactamase sequences indicated that it was not in the Ambler molecular class C like the chromosomal ,-lactamases of most other gram-negative bacteria, but rather CepA belonged to the class A 3-lactamases. The CepA enzyme together with...

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“…When the B-lactams were combined with the inhibitors, their geometric mean MICs against the remaining 78 strains were reduced from 4.2 to 150.2 ,ug/ml to 0.2 to 12.9 ,ug/ml. The activity of the P-lactams combined with the P-lactamase inhibitors was significantly greater than that of the ,-lactams alone against all groups except B. fragilis homology group II, with 76 to 100% of the strains susceptible to ampicillin plus inhibitor and .90% susceptible to the other combinations.Although the antimicrobial susceptibility spectrum of anaerobes has remained relatively stable, P-lactamasemediated resistance to penicillin and other P-lactams has been described for species in the Bacteroides fragilis group and for other Bacteroides spp., Fusobacterium nucleatum, and Clostridium butyricum (3,7,15,18,19). Several studies have demonstrated that the ,-lactamase inhibitors clavulanate and sulbactam act synergistically in vitro with ,-lactams against P-lactamase-positive, but not P-lactamase-negative, anaerobic organisms (2,4,5,16,20,21).…”

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confidence: 99%

Comparative activity of beta-lactamase inhibitors YTR 830, clavulanate, and sulbactam combined with beta-lactams against beta-lactamase-producing anaerobes

Appelbaum¹,

Jacobs²,

Spangler³

et al. 1986

Antimicrob Agents Chemother

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The in vitro activities of the P-lactamase inhibitors YTR 830, clavulanate, and sulbactam combined with six P-lactams against 88 ,-lactamase-producing anaerobes were determined. When combined with the ,-lactams, the three ,-lactamase inhibitors showed no synergy against the 10 Bacteroides fragilis homology group II strains. When the B-lactams were combined with the inhibitors, their geometric mean MICs against the remaining 78 strains were reduced from 4.2 to 150.2 ,ug/ml to 0.2 to 12.9 ,ug/ml. The activity of the P-lactams combined with the P-lactamase inhibitors was significantly greater than that of the ,-lactams alone against all groups except B. fragilis homology group II, with 76 to 100% of the strains susceptible to ampicillin plus inhibitor and .90% susceptible to the other combinations.Although the antimicrobial susceptibility spectrum of anaerobes has remained relatively stable, P-lactamasemediated resistance to penicillin and other P-lactams has been described for species in the Bacteroides fragilis group and for other Bacteroides spp., Fusobacterium nucleatum, and Clostridium butyricum (3,7,15,18,19). Several studies have demonstrated that the ,-lactamase inhibitors clavulanate and sulbactam act synergistically in vitro with ,-lactams against P-lactamase-positive, but not P-lactamase-negative, anaerobic organisms (2,4,5,16,20,21). YTR 830 is a new ,-lactamase inhibitor (1) with activity comparable to that of clavulanate and superior to that of sulbactam against ,1-lactamase-producing members of the family Enterobacteriaceae (6, 10). The purpose of this study was to compare the in vitro activities of six P-lactams (ampicillin, ticarcillin, mezlocillin, piperacillin, apalcillin, and cefoperazone) The presence of 3-lactamase was detected by a chromogenic cephalosporin method on growth from enriched blood agar plates by using nitrocefin disks (BBL Microbiology Systems, Cockeysville, Md.), in accordance with standard practice (4, 10). Organisms were stored at -70°C in thioglycolateglycerol (85:15, vol/vol) and were subcultured on enriched blood agar plates before testing.Laboratory reference powders of the following agents were used: ticarcillin and clavulanate (Beecham Laboratories, Bristol, Tenn.); mezlocillin (Miles Laboratories, Inc., West Haven, Conn.); piperacillin (Lederle Laboratories, Pearl River, N.Y.); apalcillin (Wyeth Laboratories, Philadelphia, Pa.); cefoperazone and sulbactam (Pfizer Inc., New York, N.Y.); ampicillin (Bristol-Myers Laboratories, Wallingford, Conn.); cefoxitin (Merck Sharpe & Dohme, West Point, Pa.); and YTR 830 (Taiho Pharmaceuticals, Tokyo, Japan). The powders were dissolved and diluted in accordance with the instructions of the manufacturers, and the potency of the agents was checked by determination of the MICs for quality control strains.Susceptibility testing of the isolates was performed by agar dilution on Wilkins-Chalgren agar (Difco Laboratories, Detroit, Mich.) (14,17). Plates contained serial doubling dilutions of the 3-lactams (ampicillin, 0.125 to 256 ,ug/...

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Resistance to -lactam antibiotics in Bacteroides species

Cited by 26 publications

References 0 publications

Cloning and characterization of the endogenous cephalosporinase gene, cepA, from Bacteroides fragilis reveals a new subgroup of Ambler class A beta-lactamases

Cloning and characterization of the endogenous cephalosporinase gene, cepA, from Bacteroides fragilis reveals a new subgroup of Ambler class A beta-lactamases

Insertional activation of cepA leads to high-level beta-lactamase expression in Bacteroides fragilis clinical isolates

Comparative activity of beta-lactamase inhibitors YTR 830, clavulanate, and sulbactam combined with beta-lactams against beta-lactamase-producing anaerobes

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