Standardization of Real Time Polymerase Chain Reaction (PCR) Against Multiplex PCR Via FilmArray To Detect Colistin And Carbapenem Resistant Enterobacterales Genes Amongst Septicemic Patients For Developing Countries’,

Objectives: To develop inhouse real time PCR for carbapenam and colistin resistant Genes i.e Imipenem (IMP), Klebsiella pneumoniae carbapenemase (KPC), oxacillinase (OXA-48), new Delhi metallo-β-lactamase (NDM), Verona integron-encoded metallo-β- lactamase (VIM), and mobilized colistin resistance (mcr-1), keeping multiplex PCR via biofire filmarray (BFA) as gold standard, amongst septicemic patients admitted in intensive care units (ICUs).

Study Setting: It was a multi disciplinary study conducted in the Microbiology department of Armed Forces Institute of Pathology (AFIP), Rawalpindi. The sampling was done from acute medical unit (AMU), medical ICU (MICU), surgical ICU (SICU), neonatal ICU (NICU), pediatric ICU (PICU), male and female oncology wards of Combined Military Hospital and Pak Emirates Military Hospital (PEMH), Rawalpindi, Pakistan.

Study Design And Sample Size: Diagnostic Accuracy (DA) Study which involved 116(N)

Methods: The admitted confirmed septicemic patients fulfilling guidelines mentioned in sepsis 3 definition, were enrolled for the study. The samples showing positive signal by best available control technology (BacT) alert system for blood culture proceedings were selected for BFA and real time PCR processing.

Results: The primers for colistin (mcr-1), carbapenam and microbial AMR genes were successfully designed and validated i.e mcr-1, IMP, KPC, NDM, OXA 48. The DA of BFA for microbial Colistin & Carbapenam Resistant Genes detection is 84.5 %, for colistin resistant gene detection is 86.56 % and Carapenam Genes Detection is 61.7 % upon comparison with phenotypic detection via disc diffusion testing.

Conclusion: The primers validation process via real time PCR for IMP, KPC, OXA-48, NDM, and mcr- 1 were successfully completed.