title: Genetic analysis of mechanisms of multidrug resistance in a clinical isolate of Bacteroides fragilis.
abstract: This study investigated the mechanisms of multidrug resistance (MDR) in an isolate of Bacteroides fragilis (WI1) from a patient with anaerobic sepsis. The MDR of WI1 affected susceptibility to beta-lactams, clindamycin, fluoroquinolones, metronidazole and tetracycline. In addition to its 5.31-Mb chromosome, WI1 possessed two low-copy-number plasmids, pHagl (5.6 kb) and pHag2 (9.9 kb), that were absent from B. fragilis NCTC 9343. Restriction digestion with EcoRV, HindIII and SstI, combined with DNA sequencing, revealed that pHAG2 contained a tet(Q) gene at base position 3689 that resided on the conjugative transposon CTn341. Genes cfiA (encoding a metallo-beta-lactamase) and erm(F) (encoding a macrolide-lincosamide-streptogramin B resistance determinant) were also found in WI1, but were absent from B. fragilis NCTC 9343. Nitrocefin hydrolysis revealed that WI1 had high beta-lactamase activity. Sequencing of the gyrA quinolone resistance-determining region revealed a mutation causing a Ser82 --> Phe substitution, and comparative quantitative real-time RT-PCR revealed that the cfiA, erm(F) and tet(Q) genes were all expressed in WI1. In addition, the resistance-nodulation-division efflux pump genes bmeB9 and bmeB15 were significantly over-expressed (12.30 +/- 0.42-fold and 3541.1 +/- 95.4-fold, respectively), and the efflux pump inhibitors carbonyl cyanide m-chlorophenylhydrazone and reserpine significantly increased the susceptibility of the isolate to several unrelated antibiotics (p <0.005). These data suggested that WI1 was highly multidrug-resistant because of the additive effects of chromosome- and plasmid-encoded resistance determinants.
authors.affiliation: Greater Los Angeles Veterans Administration Healthcare Systems, Los Angeles, CA, USA; Department of Medicine, University of California, Los Angeles, CA, USA.
authors.forename: L
authors.initials: L
authors.lastname: Pumbwe
authors.affiliation: Department of Microbiology, Barts and The London NHS Trust; Centre for Infectious Disease, Institute of Cell and Molecular Science, Queen Mary's School of Medicine and Dentistry, University of London, London.
authors.forename: D W
authors.initials: DW
authors.lastname: Wareham
authors.affiliation: Centre for Infectious Disease, Institute of Cell and Molecular Science, Queen Mary's School of Medicine and Dentistry, University of London, London.
authors.forename: J
authors.initials: J
authors.lastname: Aduse-Opoku
authors.affiliation: Anaerobe Reference Laboratory, National Public Health Service Microbiology Cardiff, University Hospital of Wales, Cardiff, UK.
authors.forename: J S
authors.initials: JS
authors.lastname: Brazier
authors.affiliation: Greater Los Angeles Veterans Administration Healthcare Systems, Los Angeles, CA, USA; Department of Medicine, University of California, Los Angeles, CA, USA. Electronic address: hwexler@ucla.edu.
authors.forename: H M
authors.initials: HM
authors.lastname: Wexler
mesh_terms: D019943:Amino Acid Substitution; D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D000890:Anti-Infective Agents / Q000494:pharmacology*; D001442:Bacteroides Infections / Q000382:microbiology; D001441:Bacteroides fragilis / Q000187:drug effects / Q000235:genetics / Q000302:isolation & purification; D002876:Chromosomes, Bacterial; D002981:Clindamycin / Q000494:pharmacology; D004251:DNA Transposable Elements; D004269:DNA, Bacterial; D024901:Drug Resistance, Multiple, Bacterial / Q000235:genetics*; D024841:Fluoroquinolones / Q000494:pharmacology; D005798:Genes, Bacterial; D006801:Humans; D008795:Metronidazole / Q000494:pharmacology; D008826:Microbial Sensitivity Tests; D010649:Phenylalanine / Q000378:metabolism; D010957:Plasmids; D017422:Sequence Analysis, DNA; D013752:Tetracycline / Q000494:pharmacology; D001618:beta-Lactamases / Q000494:pharmacology
chemical_list: D000900:Anti-Bacterial Agents; D000890:Anti-Infective Agents; D004251:DNA Transposable Elements; D004269:DNA, Bacterial; D024841:Fluoroquinolones; D008795:Metronidazole; D002981:Clindamycin; D010649:Phenylalanine; D001618:beta-Lactamases; D013752:Tetracycline