Prevalence of ESBL and MBL antibiotic resistance genes in Klebsiella pneumoniae in Pretoria Academic Hospital Maningi NE, Ehlers MM, Hoosen AA, Makgothlo E, Omar S and Kock MM Department of Medical Microbiology, Faculty of Health Sciences, University of Pretoria/NHLS Figure 1:Multiplex PCR simultaneously amplified the bla CTXM (593 bp), bla SHV (747 bp) and bla TEM (445 bp). In lanes 2, 5, 6 7, 8, 9 10, 11, 12,13 and 14 ESBL genes were detected while lanes 1, 3, 4, and 15 were negative for all three genes. Negative control was included in lane C and lanes L represent the molecular weight marker (100 bp DNA ladder, Promega, Medison, USA) REFERENCES Aranzazu Valverde, Teresa MC, Lucia GM, Fernando B and Rafael C (2008) Complex molecular epidemiology of extended-spectrum β-lactamases in Klebsiella pneumoniae: a long-term perspective from a single intitution in Madrid. Journal of antimicrobial Chemotherepy 61:64-72 Franceco L, Jean-Denis D, Celine C, Adrea E, Gianluigi L, Gianfranco A, Gian M and Antoniolo (2004) Emergence in K. pneumoniae and E. cloacae clinical isolates of the VIM-4 Mettallo-β-Lactamase encoded by a conjugative plasmid. Antimicrobial agents and chemothetapy 48: Michel A, Pfaller and John S (2006) Overview of the epidemiological profile and laboratory detection of extended- spectrum β-lactamases. Clinical ifectiouse diseases 42: RESULTS AND DISCUSSION Figure 2: Pie chart representation of Multiplex PCR results of three ESBL genes bla CTX-M, bla SHV and bla TEM detected in bacterial isolates L C L 747 bp bla SHV 445 pb bla TEM 593 bp bla CTX-M 100 bp DNA ladder Multiplex PCR results obtained in this study of K. pneumoniae isolates revealed that 89% (87/97) were positive for ESBL genes This prevalence of ESBL-producing K. pneumoniae genes is alarming compared to the 22% prevalence of these gene in SA (Michel et al., 2006) Carbapenems are the preferred antibiotic for the treatment of serious infections caused by ESBLs, since they are resistant to the newer expanded-spectrum cephalosporins Extended spectrum-β-lactamase (ESBL) are group of β- lactamases that are plasmid mediated that have the ability to hydrolyse the newer cephalosporins and aztreonam and are inhibited by clavulanic acid Extended spectrum-β-lactamase producing K. pneumoniae is the cause of most of the multi-drug resistant nosocomial infections The ESBL-producing K. pneumoniae can be identified by the presence of the genes blaCTX-M, blaSHV and blaTEM coding for the β-lactamases cef enzymes The transmission of ESBLs is due to clonal spread, plasmid dissemination or both mechanisms among or between species Metallo-β-lactamases has recently emerged as a resistant class of broad-spectrum β-lactamases in K. pneumoniae and it can be identified by the presence of blaVIM gene These ESBLs and MBL enzymes can be detected simultaneously using a multiplex PCR assay INTRODUCTION This study investigated the prevalence of ESBL and MBL antibiotic resistance genes in K. pneumoniae isolates from Pretoria Academic Hospital AIM This study was a prospective descriptive study in which ninety seven K. pneumoniae isolates were obtained from clinical specimens sent for analysis to the Microbiology laboratory, National Health Laboratory Services, Pretoria Academic Hospital The VITEK 2 (bioMérieux, France) was used for phenotypic identification of the isolates as ESBL-producing K. pneumoniae The MagnaPure LC Compact (Roche Applied Science, Germany) was used for the automated extraction of DNA according to manufacturer’s instructions The multiplex PCR assay amplifying the blaCTX-M, blaSHV, blaTEM and blaVIM genes was performed using the QIAGEN Multiplex PCR kit (QIAGEN, USA) following the manufacture’s instructions MATERIALS AND METHODS The ESBL genes were detected in different combinations as only one gene, two genes or all three genes in one isolate (Figure 2) Most of the ESBL-producing K. pneumoniae isolates harboured three ESBL genes (Figure 2) Multiplex PCR detected the presence of CTX-M, SHV and TEM genes in the K. pneumoniae isolates from the Pretoria Academic Hospital (Figure 1) No VIM genes were detected with the multiplex PCR in the K. pneumoniae isolates included in this study There is a high prevalence of ESBL-producing K. pneumonia isolates in Pretoria Academic Hospital This leads to a need for improved, adequate infection control measures to be instituted and more rational use of antibiotics in Pretoria Academic Hospital CONCLUSION