Detection of Extended-Spectrum β-Lactamase and Klebsiella pneumoniae Carbapenemase Genes Directly from Blood Cultures by Use of a Nucleic Acid Microarray

摘要

The growing crisis of multidrug-resistant (MDR) Gram-negative bacteria requires that current technologies permit the rapid detection of extended-spectrum β-lactamase (blaESBL) and Klebsiella pneumoniae carbapenemase (blaKPC) genes. In the present study, we assessed the performance characteristics of a commercially available nucleic acid microarray system for the detection of blaESBL and blaKPC genes directly from positive blood cultures. Using blood cultures (BCs) that contained Gram-negative bacilli identified by Gram staining, we isolated bacterial DNA using spin columns (BC-C) and rapid water lysis (BC-W). Twenty ESBL/KPC-positive and 20 ESBL/KPC-negative blood culture samples, as well as 20 non-lactose-fermenting organisms, were tested. The 20 isolates that were ESBL positive by phenotypic testing were also evaluated on solid medium (SM), and the DNA was extracted by use of a spin column (SM-C). The resulting 140 DNA extractions were assessed for DNA quantity and quality using 260/280-nm absorbance ratios, and DNA microarray analysis was performed in a blinded fashion. Microarray and phenotypic results were concordant for 98.3% of BC-W, 90% of BC-C, and 95% of SM-C samples. Compared to phenotypic testing, the sensitivity and specificity for BC-C samples were 88.9% and 100%, respectively, and for BC-W samples, the sensitivity and specificity were 94.4% and 100%, respectively. BC-W samples yielded the highest concordance with phenotypic results. Nucleic acid microarrays offer promise in the identification of blaESBL and blaKPC genes directly from blood cultures, thereby reducing the time to identification of these important pathogens.