title: Detection of Amp C genes encoding for beta-lactamases in Escherichia coli and Klebsiella pneumoniae.
abstract: OBJECTIVE
  Amp C beta-lactamase are Ambler class C enzymes that confer resistance to extended spectrum cephalosporins and are not inhibited by beta-lactamase inhibitors. Their detection is crucial, since the phenotypic tests are not standardised leading to ambiguity in interpretation of results. This study was done to detect the types of Amp C prevalent in Escherichia coli and Klebsiella pneumoniae by multiplex polymerase chain reaction (PCR).
  
  
  METHODS
  Seventy-seven consecutive cefoxitin resistant clinical isolates of E. coli (n = 25) and K. pneumoniae (n = 52) were included in the study. Antibiotic susceptibility testing to various classes of antibiotics was performed by disc diffusion using Clinical Laboratory Standards Institute (CLSI) guidelines. Minimum inhibitory concentration (MIC) to cefoxitin, imipenem and meropenem were determined by broth microdilution method. Isolates were screened for production of Extended Spectrum Beta-Lactamase (ESBL). Multiplex PCR was performed for the detection of Amp C genes after phenotypic testing (Hodge test and inhibitor based test).
  
  
  RESULTS
  Cefoxitin Hodge test was positive in 40 isolates which included 20 E. coli and 20 K. pneumoniae. There was zone enhancement with boronic acid in 55 isolates, of which 36 were K. pneumoniae and 19 were E. coli. Multiplex PCR detected Amp C in 11/25 E. coli and 12/52 K. pneumoniae isolates. The Amp C genes detected were CIT (Amp C origin - Citrobacter freundii), DHA (Dhahran Hospital, Saudi Arabia), ACC (Ambler class C), EBC (Amp C origin - Enterobacter cloacae) groups. ESBL was co-produced in 54 isolates.
  
  
  CONCLUSIONS
  Amp C was detected in 29.87% of the study isolates. Majority of them co-produced ESBL. The most common Amp C was the CIT family. Screen tests for cefoxitin resistance may be falsely positive due to production of carbapenamases.
authors.affiliation: Department of Microbiology, Sri Ramachandra Medical College and Research Institute, Sri Ramachandra University, Porur, India.
authors.forename: M
authors.initials: M
authors.lastname: Shanthi
authors.forename: U
authors.initials: U
authors.lastname: Sekar
authors.forename: K
authors.initials: K
authors.lastname: Arunagiri
authors.forename: B
authors.initials: B
authors.lastname: Sekar
mesh_terms: D000900:Anti-Bacterial Agents / Q000494:pharmacology; D001426:Bacterial Proteins / Q000235:genetics*; D002440:Cefoxitin / Q000494:pharmacology; D004269:DNA, Bacterial / Q000235:genetics; D004926:Escherichia coli / Q000187:drug effects / Q000201:enzymology* / Q000235:genetics* / Q000302:isolation & purification; D004927:Escherichia coli Infections / Q000382:microbiology; D005838:Genotype; D006801:Humans; D007194:India; D007710:Klebsiella Infections / Q000382:microbiology; D007711:Klebsiella pneumoniae / Q000187:drug effects / Q000201:enzymology* / Q000235:genetics* / Q000302:isolation & purification; D008826:Microbial Sensitivity Tests; D060885:Multiplex Polymerase Chain Reaction; D010641:Phenotype; D018440:beta-Lactam Resistance; D001618:beta-Lactamases / Q000235:genetics*
chemical_list: D000900:Anti-Bacterial Agents; D001426:Bacterial Proteins; D004269:DNA, Bacterial; D002440:Cefoxitin; C048795:AmpC beta-lactamases; D001618:beta-Lactamases