Early Detection of Carbapenemases Producing Gram Negative Isolates by NG-Test CARBA 5: A Cross-sectional Study

Introduction: Rapid and accurate detection of carbapenemase producing Gram negative bacteria is prerequisite for effective clinical management and infection control strategies. The limited susceptibility of carbapenemase producing bacteria to antibiotics leads to serious global health issues. Currently various molecular and phenotypic test are available for early screening and identification of carbapenamase producing organisms. Aim: The aim of the present study was to detect various types of carbapeneamase produced by multidrug resistant Gram negative bacterial isolates obtained from clinical samples during the study period and to compare the effectiveness of NG-Test CARBA 5 assay with TaqMan based Real-Time Polymerase chain reaction (RT-PCR) for the early detection of carbapenmase producing Gram negative isolates. Materials and Methods: The present cross-sectional study was conducted at NKP Salve Institute of Medical Sciences & Research Centre and Lata Mangeshkar Hospital, Nagpur, Maharashtra, India a Tertiary Care Hospital involving non-duplicate Gram negative carbapenem resistant isolates. NG-Test CARBA 5 was employed to detect carbapenemase production in different bacterial isolates. The study was conducted in duration of six months from August 2023 to February 2024. All the ertapenem resistant Gram negative bacterial isolates obtained from clinical samples during the study period were included in the study. Types of carbapeneamase produced by multidrug resistant Gram negative bacterial isolates obtained from clinical samples by of NG-Test CARBA 5, by immunochromatographic assay during the study period were documented. For 30 randomly selected isolates and a negative reference strain, the results obtained from NG-Test CARBA 5 were confirmed by TaqMan based multiplex RT-qPCR, for detection of carbapeneamase genes, NDM, KPC, IMP, VIM, OXA. The test was performed as per the kit protocol. PCR results were considered as the gold standard for detection of genes. Sensitivity, specificity, Positive Predictive Value (PPV) and Negitive Predictive Value (NPV) for NG-Test CARBA 5 was calculated using Microsoft Excel 2016. Results: Out of 277 Gram negative isolates, 75 (27.07%) isolates which were resistant to carbapenam were evaluated for presence of carbapeneamase genes by NG-Test CARBA assay. The results revealed that 54 (72%) isolates were positive for one or more carbapenemase genes, whereas 21 (28%) isolates were confirmed to be negative. The negative control (E. coli ATCC 25922) showed appropriate results. To validate and evaluate the feasibility of the NG-Test CARBA 5 assay, RT-qPCR test was performed for 30 (20 isolates NG-Test CARBA 5 Positive and 10 isolates NG-Test CARBA 5 negative) randomly selected isolates, and a negative reference strain; all the isolates tested by RT-qPCR, showed presence of one or more genes and the reference negative strain was confirmed as a negative. Conclusion: As per the present study findings, it is 64.5% sensitive and 100% specific, The PPV and NPV of NG-Test CARBA 5 is 100% and 8.4%. NG-Test CARBA 5 showed promise in detecting carbapenemase producing isolates although it needs further evaluation with large number of samples under Indian environmental conditions.