The performance and portability of a multiplex real-time PCR assay to detect KPC, NDM, OXA-48-like and VIM carbapenemase gene families from bacterial isolates was assessed using Rotor-Gene Q and ABI 7500 instruments. Gram-negative bacterial isolates (n = 502) were comprised of 100 isolates each with KPC, NDM, VIM or OXA-48-like carbapenemases (including 17 with OXA-181) and 2 isolates with NDM + OXA-48-like enzymes (including 1 with OXA-181) as well as 100 assay-negative isolates comprised of 24 IMP-producers, 24 carbapenem-resistant isolates with no known carbapenemase gene and 52 extended-spectrum β-lactamase-producing carbapenem-susceptible isolates. A multicentre evaluation was carried out in five laboratories using a subset of 100 isolates comprised of 22 isolates each with KPC, NDM, OXA-48-like or VIM alleles and 12 isolates that were negative for the assay targets. Initial validation of the assay on both the Rotor-Gene Q and ABI 7500 instruments demonstrated 100% sensitivity amongst the 402 isolates that were positive for KPC, NDM, OXA-48-like (including OXA-181) and VIM carbapenemase genes, whilst the 100 assay-negative samples were correctly identified indicating 100% specificity. During the multicentre evaluation the same 100% level of sensitivity and specificity was observed in each of the five centres that participated. Neither invalid nor false-positive results were observed. In conclusion, the assay offers a portable and reliable option for the detection of bacteria carrying clinically significant carbapenemases encoded by KPC, NDM, VIM and OXA-48-like carbapenemase genes using either of the two most common real-time PCR instrument platforms.
Bibliographical noteFunding Information:
Funding: This work was funded by the Public Health England (PHE) Strategic Development Fund .
Competing interests: MJE has received financial support from Bruker Daltonik GmbH for attending a conference. PHE's Antimicrobial Resistance and Healthcare Associated Infections (AMRHAI) Reference Unit has received financial support for conference attendance, lectures, research projects or contracted evaluations from numerous sources, including Achaogen Inc., Allecra Antiinfectives GmbH, Amplex, AstraZeneca UK Ltd., Becton Dickinson Diagnostics, The British Society for Antimicrobial Chemotherapy (BSAC), Cepheid, Check-Points B.V., Cubist Pharmaceuticals, Department of Health, Food Standards Agency, GlaxoSmithKline Services Ltd., Henry Stewart Talks, IHMA Ltd., Merck Sharp & Dohme Corp., Meiji Seika Kiasha Ltd., Momentum Biosciences Ltd., Nordic Pharma Ltd., Norgine Pharmaceuticals, Rempex Pharmaceuticals Ltd., Rokitan Ltd., Smith & Nephew UK Ltd., Trius Therapeutics, VenatoRx and Wockhardt Ltd. PH has received lecture/consultancy fees from Eumedica, Merck and Pfizer, and research funding from Novartis and Pfizer in the last 3 years. All other authors declare no competing interests.
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