Phenotypic and Genotypic Detection of Plasmid-Mediated AmpC β-lactamases among Klebsiella Isolates

Abstract

AmpC β-lactamases have gained importance since the late 1970s as one of the mediators of antimicrobial resistance in Gram negative bacilli. These enzymes are typically produced by isolates of E. coli, Klebsiella spp., Proteus, and Salmonella spp. and are associated with multiple antibiotic resistance that leaves few therapeutic options. Enterobacteriaceae producing AmpC β-lactamases (AmpC) have become a major therapeutic challenge. The detection of AmpC-producing Klebsiella spp. is of significant clinical relevance since AmpC producers may appear susceptible to expanded-spectrum cephalosporins when initially tested. This may lead to inappropriate antimicrobial regimens and therapeutic failure. Thus, a simple and reliable detection for AmpC producers is needed. Cefoxtin screening test is reported as useful in screening for AmpC. Modified Three Dimensional Test (M3D) is reported as simple and sensitive phenotypic detection method. Many studies have validated the use of cloxacillin (as AmpC inhibitor) to detect AmpC β-lactamases among Gram-negative bacteria. Also, many studies have validated the use of imipenem and cefoxitin as inducing agents to detect the presence of inducible AmpC enzymes among enterobacterial strains. Even though multiplex-PCR is a gold standard method to detect AmpC β-lactamase genes (differentiating between chromosomal and plasmid-mediated AmpC β-lactamases and the different types or families of PMABLs), but the test is still costly and time consuming and equipment availability is limited to few laboratories, hence looking for specific and sensitive phenotypic tests have always been a challenge. Many clinical laboratories show interests in performing phenotypic tests as these are cost effective, simple to perform, easy to implement in the diagnostic laboratory to avoid inappropriate therapy and for better infection control, distinguishing between the cefoxitin-resistant AmpC producers and cefoxitin-resistant non AmpC producers which may be due to reduced membrane permeability and also, differentiation of organisms expressing ESBLs from organisms expressing plasmid-mediated AmpC β-lactamases in order to address surveillance and epidemiology as well as hospital infection control issues associated with these resistance mechanisms. The aim of this work is to detect of the occurrence of PMABLs in Klebsiella spp. isolates by phenotypic tests and detect its gene types by multiplex-PCR. This study was conducted on a total number of one hundred clinical isolates of Klebsiella spp. collected from different clinical specimens referred to the Central Microbiological Laboratory of Ain Shams University Hospitals