Phenotypic detection of metallo-beta-lactamase production in clinical isolates of Escherichia coli and Klebsiella pneumoniae in a tertiary care hospital

Introduction: Carbapenem resistance due to metallo-beta-lactamase (MBL)-producing bacteria is an emerging threat worldwide. This study aimed to detect the MBL production in clinical isolates of E. coli and Klebsiella pneumoniae species in our hospital setting and to evaluate the efficiency of two phenotypic methods for the detection of MBL production. Materials and Methods: The present study was carried out in the Department of Microbiology, MGM Medical College and Hospital, Aurangabad, Maharashtra, for a period of 2 years from April 2018 to March 2020. From a total of 12,324 various clinical specimens, 393 isolates of E. coli and Klebsiella pneumoniae species were tested for MBL production. MBL was detected by two different phenotypic methods, i.e., combined disc test and E-test. Results: Out of 393 isolates, 130 (33.07%) isolates were resistant to imipenem on screening of which 71 (18.06%) were Klebsiella pneumoniae and 59 (15.01%) were E. coli. About 43.66% Klebsiella pneumoniae isolates and 40.67% E. coli isolates were MBL-positive by the combined disc test. Using the E-test, MBL production was found to be 46.47% and 45.76% in Klebsiella pneumoniae and E. coli, respectively. Conclusion: Routine screening of MBL-producing organisms should be performed in diagnostic laboratories to control the spread of resistance and for the proper management of antibiotic therapy. E-test is better than the combined disc test for the detection of MBL-producing gram-negative bacilli.

Detection and characterization of metallo-β-lactamases in Pseudomonas aeruginosa by phenotypic and molecular methods from clinical samples in a tertiary care hospital / Detección y caracterización de la metallo-betalactamasa en la Pseudomonas aeruginosa mediante métodos fenotípicos y moleculares a partir de muestras clínicas en un hospital de atención terciaria

AIMS: The aim of this study was to detect and characterize the presence of metallo-β-lactamase (MBL) production in multidrug resistant (MDR) P aeruginosa collected from clinical samples in a tertiary care hospital. METHODS AND MATERIALS: A total of 67 non-repetitive isolates of MDR P aeruginosa recovered from various clinical specimens were screened for MBL production by IPM/MEM-EDTA combined disc test. Polymerase chain reaction was performed on all isolates using blaIMP and blaVIM consensus primers to characterize them genotypically. RESULTS: Among 67 P aeruginosa isolates, 62.7% (42/67) and 70.1% (47/67) were resistant to imipenem and meropenem respectively and 47 (70.1%) were found to be MBL producers. Among this 47 MBL-producing isolates, 41 (61.1%) strains carried the blaVIM gene and 2 (3%) strains carried the blaIMP gene. Three strains were phenotypically negative but positive genotypically for blaVIM gene. One strain was resistant to both imipenem and meropenem but did not show phenotypic positivity. CONCLUSION: This study confirms the dissemination of blaVIM genes among MDR Pseudomonas aeruginosa and hence it is indispensible to identify and aptly control the threat of horizontal and vertical transfer.

OBJETIVO: El objetivo de este estudio es descubrir y caracterizar la presencia de producción de metallo-betalactamasa (MBL) en P aeruginosa resistente a los multifármacos (RMF), recogida de muestras clínicas de un hospital de atención terciaria. MÉTODO: Un total de 67 aislados no repetitivos de P aeruginosa RMF obtenidos de varios specímenes clínicos, fueron tamizados en busca de producción de MBL, mediante una prueba de disco combinado IPM/MEM-EDTA. Se efectuó una reacción en cadena de la polimerasa sobre todos los aislados, usando iniciadores de consenso blaIMP y blaVIM para la caracterización genotípica. RESULTADOS: Entre los aislados de P aeruginosa, 62.7% (42/67) y 70.1% (47/67) fueron resistentes al Imipenem y al Meropenem respectivamente, mientras que se halló que 47 (70.1%) eran productores de MBL. De los 47 aislados productores de MBL, 41 (61.1%) cepas eran portadoras del gen blaVIM en tanto que 2 (3%) cepas eran portadoras del gen blaIMP. Tres cepas fueron fenotípicamente negativas, pero genotípicamente positivas con respecto al gen blaVIM. Una cepa fue resistente tanto al Imipenem como al Meropenem, pero no mostró positividad fenotípicamente. CONCLUSIÓN: El presente estudio confirma la diseminación de los genes blaVIM entre las Pseudomonas aeruginosa RMF. Es importante identificar así como controlar adecuadamente la amenaza de la transferencia horizontal y vertical.

Evaluation of combined disc method for the detection of metallo-β-lactamase producing Gram negative bacilli

Aims: Infections due to metallo-β-lactamase (MBL) producing Gram negative rods are a cause of high mortality and morbidity. Early detection by an economical and accurate method may improve patient outcome. This study was aimed to evaluate the diagnostic accuracy of combined disc method for MBL detection by comparing it with MBL-Etest. Methodology and Results: This cross-sectional, validation study was carried out in the Department of Microbiology, Army Medical College, National University of Sciences and Technology, Rawalpindi, over a period of six months. A total of 52 non-duplicate Gram-negative rods isolated from the routine clinical specimens and found resistant to meropenem/imipenem on Kirby Bauer Disc Diffusion method were subjected to two tests for metallo-β-lactamase detection. One was combined Disc test using imipenem with Ethylene Diamine Tetraacetic Acid (EDTA), where a strain showing an increase in zone of inhibition of combined disc of ≥ 7 mm as compared to imipenem alone, was considered as MBL producer and the other one was MBL-Etest for which results were interpreted as per manufacturer’s guidelines. Combined disc method for MBL detection was found to have a sensitivity, specificity, positive predictive value, negative predictive value and accuracy of 97.5%, 100%, 100%, 92% and 98%. Conclusion, Significance and Impact of study: Combined disc method is an economical and reliable method for metallo-β-lactamase detection which can be used routinely in any laboratory.