Molecular Detection of Carbapenemases in Enterobacterales: A Comparison of Real-Time Multiplex PCR and Whole-Genome Sequencing.

Carbapenem-resistant Enterobacterales are a growing problem in healthcare systems worldwide. While whole-genome sequencing (WGS) has become a powerful tool for analyzing transmission and possible outbreaks, it remains laborious, and the limitations in diagnostic workflows are not well studied. The aim of this study was to compare the performance of WGS and real-time multiplex PCR (RT-qPCR) for diagnosing carbapenem-resistant Enterobacterales. In this study, we analyzed 92 phenotypically carbapenem-resistant Enterobacterales, sent to the University Hospital Heidelberg in 2019, by the carbapenem inactivation method (CIM) and compared WGS and RT-qPCR as genotypic carbapenemase detection methods. In total, 80.4% of the collected isolates were identified as carbapenemase producers. For six isolates, discordant results were recorded for WGS, PCR and CIM, as the carbapenemase genes were initially not detected by WGS. A reanalysis using raw reads, rather than assembly, highlighted a coverage issue with failure to detect carbapenemases located in contigs with a coverage lower than 10×, which were then discarded. Our study shows that multiplex RT-qPCR and CIM can be a simple alternative to WGS for basic surveillance of carbapenemase-producing Enterobacterales. Using WGS in clinical workflow has some limitations, especially regarding coverage and sensitivity. We demonstrate that antimicrobial resistance gene detection should be performed on the raw reads or non-curated draft genome to increase sensitivity.