Comparative Evaluation of Phenotypic and Genotypic Methods for Detection of Carbapenemases in Clinically Significant Klebsiella Pneumoniae Isolates

Abstract

Background: Carbapenems are the most active and reliable treatment options against ever more prevalent Klebsiella pneumoniae (K. pneumoniae) that produce extended-spectrum β-lactamases, but their efficacy is threatened worldwide by carbapenemase production which is frequently associated with serious infections and higher mortality. Objectives: The aims of this study were to detect occurrence of carbapenemase producing K. pneumoniae and to evaluate the use of Multiplex PCR for rapid detection of carbapenemase genes in comparison with phenotypic methods as modified Hodge test (MHT) and combined disc test (CDT). Methodology: Different clinical samples were obtained from 430 patients admitted to Zagazig University Hospitals over the period from June 2014-June 2015. They were cultured and K. pneumoniae isolates were identified. The antimicrobial susceptibility patterns were determined by disc diffusion test .Carbapenem-resistant isolates were selected and subjected to Modified Hodge test (MHT) for carbapenemase detection, combined disc test (CDT) for differentiation of Ambler classes of carbapenemases and multiplex PCR for detection of bla KPC , bla NDM-1 , bla VIM , bla IMP and bla OXA-48 genes. Results: A total of non-duplicate 100 K. pneumoniae clinical isolates were collected from 430 clinical samples. By disc diffusion test, 42 Carbapenem-resistant K. pneumoniae isolates were detected. Thirty-four of the 42 isolates were MHT positive while 27 isolates were positive by CDT. MDT showed sensitivity & specificity of 100% & 47.06% respectively, while CDT showed sensitivity & specificity of 100% & 88.89% respectively. By PCR, it was found that out of 42 carbapenem-resistant isolates, 25 isolates harboring one gene. The most common resistance gene was bla OXA-48 (12/42) followed by bla KPC (8/42) while the bla VIM gene (4/42) while bla NDM-1 was detected only in one isolate. bla IMP had not been detected in our isolates. Conclusion: Multiplex PCR is an accurate method for detection of carbapenemase production genes which overcomes the limitations of the phenotypic methods.