title: Genotypic Identification of AmpC β-Lactamases Production in Gram-Negative Bacilli Isolates.
abstract: BACKGROUND
  AmpC type β-lactamases are commonly isolated from extended-spectrum Cephalosporin-resistant Gram-negative bacteria. Also, resistance appeared in bacterial species not naturally producing AmpC enzymes. Therefore, a standard test for the detection of the plasmid-mediated AmpC enzyme and new breakpoints for extended spectrum Cephalosporins are urgently necessary.
  
  
  OBJECTIVE
  To detect plasmid and chromosomal mediated AmpC-β-lactamases in Gram negative bacteria in community and hospital acquired infections.
  
  
  METHODS
  1073 Gram negative clinical isolates were identified by the conventional methods and were screened for AmpC production using Cefoxitin discs. Confirmatory phenotypic identifications were done for the Cefoxitin-resistant isolates using Boronic Acid for combined and double disc synergy tests, Cloxacillin based double disc synergy test, and induction tests. The genotypic identification of plasmid-mediated AmpC was done using multiplex PCR. ESBL production was also screened by discs of Ceftazidime and Cefotaxime with and without Clavulanic Acid (10 μg).
  
  
  RESULTS
  The AmpC-producing isolates among all identified Gram negative bacilli were 5.8% (62/1073) as detected by screening disc diffusion methods, where 72% were positive for AmpC by combined disc method (Cefotetan and Boronic Acid), 56.5% were positive by each of Boronic Acid and Cloxacillin double disc synergy tests, 35.5% were positive by the induction test, and 25.8% were plasmid-mediated AmpC β-lactamase producers by the multiplex PCR. Plasmid-mediated AmpC genes retrieved, belonged to the families (MOX, FOX, EBC and CIT). ESBL producers were found in 26 (41.9%) isolates, 15 (57%) of which also produced AmpC. Isolates caused hospital acquired infections were (53/62); of which (39/62) were AmpC producers. While only (8/62) of the isolates caused community-acquired infections, were AmpC producers, and (1.6%) (1/62) were non AmpC producer.
  
  
  CONCLUSIONS
  The AmpC β-lactamases detection tests had to be included in the routine microbiology workup of Gram negative bacteria, namely Cefoxitin as a screening test, combined Boronic Acid disc test with Cefotetan, followed by synergy tests and finally by the induction test for phenotypic identifications. Multiplex PCR can successfully detect the plasmid AmpC genes.
authors.affiliation: Department of Clinical and Chemical Pathology, Faculty of Medicine, Cairo University, Giza, Egypt.
authors.forename: Mona
authors.initials: M
authors.lastname: Wassef
authors.affiliation: Department of Clinical and Chemical Pathology, Faculty of Medicine, Cairo University, Giza, Egypt.
authors.forename: Iman
authors.initials: I
authors.lastname: Behiry
authors.affiliation: Department of Clinical and Chemical Pathology, Faculty of Medicine, Cairo University, Giza, Egypt.
authors.forename: Mariam
authors.initials: M
authors.lastname: Younan
authors.affiliation: Department of Clinical and Chemical Pathology, Faculty of Medicine, Cairo University, Giza, Egypt.
authors.forename: Nancy
authors.initials: N
authors.lastname: El Guindy
authors.affiliation: Department of Clinical and Chemical Pathology, Faculty of Medicine, Cairo University, Giza, Egypt.
authors.forename: Sally
authors.initials: S
authors.lastname: Mostafa
authors.affiliation: Department of Botany and Microbiology, Faculty of science, Helwan University, Cairo, Egypt ; Department of Biology, Faculty of Science, Jazan University, Jazan, KSA.
authors.forename: Emad
authors.initials: E
authors.lastname: Abada
keywords: AmpC Genes; AmpC β-Lactamase; Boronic Acid; Cloxacillin; Multiplex PCR