Evaluation of the Amplidiag CarbaR-VRE kit for the accurate detection of carbapenemase-producing bacteria

Introduction Carbapenemase-producing enterobacteriaceae (CPE) and non fermenters (CPNF; pseudomonadaceae and Acinetobacter sp.) have been increasingly reported worldwide. The most clinically relevant carbapenemases belong either to Ambler class A (KPC-type), Ambler class B, i-e metallo-b-lactamases (MBLs such as IMP-, VIM- and NDM-types) or Ambler Class D (OXA-48-like in enterobacteriaceae, Acinetobacter OXA group: OXA-23, OXA-40, OXA-58, OXA-143 and the intrinsic OXA-51-like enzymes, OXA-198 in P. aeruginosa ). Materials and method The Amplidiag ® Carba-R + VRE, a qualitative multiplex nucleic acid-based in-vitro diagnostic test intended for the detection of carbapenemase-producing bacteria and vancomycin resistant enterococci, has been tested on a collection of 100 characterized Gram-negative isolates with reduced susceptibility to carbapenems, and on 200 isolates collected at the National Reference Center for Antibiotic Resistance from 20st January to 10th February 2016. The markers detected by this assay are bla KPC-lik e, bla NDM-like , bla VIM-like , bla IMP-like , bla OXA-48-like , Acinetobacter OXA genes including bla OXA-23 , bla OXA-24/-40 , bla OXA-58 , and bla OXA-51 including upstream promoter ISAba1 , and vanA and vanB . DNA was extracted using QiaAmp DNA extraction kits or by boiling extract. Results The Amplidiag ® CarbaR + VRE was able to detect all KPC, NDM, VIM, IMP, OXA-48 and variants including OXA−162, −181, −204, −232, −244. Similarly, all A. baumannii producing OXA−23, OXA−24/−40, OXA−58 as well as over expression of the chromosomally-encoded OXA−51-like b-lactamase, due to the presence of ISAba1 upstream of the blaOXA-51 gene were detected. The most prevalent carbapenemases encountered in P. aeruginosa have also been detected. However, as claimed by the manufacturer, other carbapenemases such as GES-like carbapenemases (GES-2, GES−5 in P. aeruginosa , GES−14 in A. baumannii ), GIM−1, AIM−1, SPM−1, DIM−1 or OXA−198 in P. aeruginosa , or OXA−143-like in A. baumannii were not detected by the Amplidiag® CarbaR + VRE assay. The PCR worked equally well on purified DNA and on boiling extracted DNA from a colony. The biological performances of the Amplidiag® CarbaR + VRE on the prospective study of 200 CREs isolates collected at the NRC were of 100% and 96% sensitivity and specificity, respectively. Conclusion Amplidiag® Carba-R's performances were high as it was able to detect the five major carbapenemases: NDM, VIM, IMP, KPC, and OXA−48, as well as OXA-type carbapenemases from Acinetobacter sp. It can provide a result directly on colonies growing on selective screening media within two hours. This test needs now to be evaluated on rectal swabs, since it is able to detect the most worrisome resistant traits in Gram negative bacteria.