Evaluation of the AID carbapenemase line probe assay for rapid detection and identification of carbapenemase genes in Gram-negative bacilli.

Abstract

This study evaluated the AID carbapenemase line probe assay (LPA) for the detection and identification of carbapenem resistance genes in Enterobacteriaceae and other Gram-negative bacilli (GNB) using bacterial cultures and DNA extracts directly from patient urine samples. The AID carbapenemase LPA detects 13 different carbapenemase genes. Test probe accuracy was verified for using clinical Enterobacteriaceae isolates harbouring bla KPC , bla VIM , bla NDM , bla GIM , bla AIM , bla SPM , bla IMP and bla OXA-48 and a well-characterized set of Escherichia coli DH5α strains transformed with the vector plasmid pUC57- kan harbouring bla BIC , bla SIM , bla DIM , bla IMI-3 , bla IMI-1 and bla NMC-A . Sensitivity and specificity was determined by testing 151 clinical GNB strains previously characterized for the production of carbapenemase activity and carbapenemase genes. Direct detection of carbapenemase genes using the LPA was determined using 299 clinical urine specimens. Analytical sensitivity for detection in urine was determined by testing serial dilutions of bla KPC and bla NDM in clinical Klebsiella pneumoniae strains. All carbapenemase gene probes showed 100% accuracy without cross-reactions. Sensitivity and specificity of the LPA using clinical isolates was 100% for each. Analytical sensitivity for detection of bla KPC and bla NDM in urine was 10 1 -10 2 cfu. The LPA detected carbapenemase genes in 20 urines, which were confirmed in 12 samples by conventional multiplex PCR. Remarkably, 0 of the 20 urines grew carbapenemase-suspicious GNB applying EUCAST recommendations. The AID carbapenemase LPA is an accurate, sensitive and easy-to-use test for the detection and identification of carbapenemase genes, which can readily be implemented in any diagnostic laboratory.