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

ABSTRACT

The growing crisis of multidrug-resistant (MDR) Gram-negative bacteria requires that current technologies permit the rapid detection of extended-spectrum β-lactamase (bla _ESBL) and Klebsiella pneumoniae carbapenemase (bla _KPC) genes. In the present study, we assessed the performance characteristics of a commercially available nucleic acid microarray system for the detection of bla _ESBL and bla _KPC 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 bla _ESBL and bla _KPC genes directly from blood cultures, thereby reducing the time to identification of these important pathogens.