Phenotypic and Genotypic Methods for Detection of Extended Spectrum β Lactamase Producing Escherichia coli and Klebsiella pneumoniae Isolated from Ventilator Associated Pneumonia

Abstract

Background: Ventilator Associated Pneumonia (VAP) is one of the common nosocomial infections associated with high morbidity due to multidrug resistant pathogens. Rapid spread of resistance to broad spectrum beta-lactams in pathogenic strains causes antibiotics ineffectiveness and increased severity of illness. The CTX-M is the most dominant Extended Spectrum β Lactamase (ESBL) among Enterobacteriaceae in many regions of the world. The aim of the study was to identify the occurrence of ESBL and detect the genes responsible for ESBL production by conventional Polymerase Chain Reaction (PCR) method. Methods: This prospective study included patients, clinically diagnosed as VAP. Endotracheal aspirates (EA) were collected and cultured by quantitative method. The bacterial isolates were identified as per standard methods. Isolates resistant to 3rd generation cephalosporins were screened for ESBL production by disk approximation method and combination disc diffusion method. Isolates confirmed as ESBL producers were subjected to genotyping by conventional PCR. Statistical Analysis: Statistical analysis was done by using MS Excel sheet. Descriptive statistics like percentage was done in the study. Results: Among the isolates from 428 patients who developed VAP, 144 isolates belonged to the Enterobacteriaceae family (Klebsiella pneumoniae 87 and Escherichia coli 57). A total of 66 isolates (28 Klebsiella pneumoniae and 38 Escherichia coli) were confirmed as ESBL producer by disc approximation method and 63 isolates by double disc combination method. In the present study by conventional PCR bla CTX-M was the common gene in 48.5% strains followed by 22.22% bla SHV and 14.81% bla TEM. Conclusion: The genotypic methods using specific PCR amplification of resistance genes seems to have 100% specificity and sensitivity in detection of ESBL when compared to phenotypic methods which lacks the constant sensitivity.

Keywords

• blatem gene
• blashv gene
• blactx-m gene
• extended spectrum beta lactamase
• polymerase chain reaction
• ventilator associated pneumonia