Evaluation of various methods for detection of metallo-β-lactamase (mbl) production in gram negative bacilli

Introduction: The emergence of metallo-β-lactamase (MBL) in gram negative bacilli (GNB) is becoming a therapeutic challenge worldwide. Detection of MBL is also a challenge for routine microbiology laboratories, since there are no standardized methods for MBL detection. The aims of this study were to know prevalence of MBL production in various gram negative bacilli, to evaluate different phenotypic methods to detect MBL production and to find out antibiotic sensitivity profile of MBL producing gram negative bacilli. Material and methods: Total 450 clinical isolates of GNB including E. coli, Pseudomonas, Klebsiella, Acinetobacter and Other GNB were subjected to antibiotic susceptibility testing. Imipenem, ertapenam, meropenam and third generation cephalosporins resistant clinical isolates were taken as positive for MBL screening. Four different methods using EDTA as MBL inhibitor were evaluated: (i) Combined disk synergy test with imipenem (CDST-IPM), (ii) Double-disk synergy test with imipenem (DDST-IPM), (iii) CDST with ceftazidime (CDST-CAZ) and (iv) DDST with ceftazidime (DDST-CAZ). Result: Out of 450 clinical isolates of GNB, 27 isolates (6.00%) were resistant to imipenem, ertapenam, meropenam and third generation cephalosporins. These 27 isolates were considered screening positive and further tested for MBL production by four different methods. 26 isolates (96.30%) were MBL positive by CDST-IPM and 22 isolates (81.48%) were MBL positive by DDST-IPM. 23 isolates (85.19%) were MBL positive by CDST-CAZ and 12 isolates (44.44%) were MBL positive by DDST-CAZ. Prevalence of MBL production was highest in Pseudomonas (9.92%), followed by Klebsiella pneumoniae (7.26%), Acinetobacter spp. (7.14%) and E. coli (2.87%). Conclusion: The detection of MBL-producing isolates is of crucial importance in GNB isolates especially in Pseudomonas. CDST-CAZ is the most sensitive method for MBL detection.