Conventional susceptibility testing requires 18-48 h and often delays therapy, whereas existing molecular assays are costly and limited. Real-time PCR's ubiquity in hospitals offers a rapid multiplex screening platform. We established a single-tube multiplex real-time PCR assay coupled with high-resolution melting analysis to detect seven resistance genes (blaKPC, blaNDM, blaOXA-48-like, blaVIM, blaIMP, mcr-1, and mcr-3). Validated on 577 clinical Enterobacterales isolates and a standard strain, the assay exhibited distinct melt peaks for each target, with intra- and inter-run CVs NDM and blaOXA-48-like genes improved the sensitivity from 82.7% to 92.9% when precision melt analysis software was used. The assay demonstrated good quantitative analytical performance, with efficiencies ranging from 91 to 114% (R2 = 0.96-0.99), and a minimum of 102 copies required to confidently detect all targets. In under 4 h, this cost-effective assay leverages existing platforms for comprehensive surveillance of carbapenem and colistin resistance, enabling timely antimicrobial stewardship and infection control.
Luk-in et al. (Tue,) studied this question.