Evaluation of a diagnostic algorithm for rapid identification of Gram-negative species and detection of extended-spectrum β-lactamase and carbapenemase directly from blood cultures

Objectives: To evaluate a rapid diagnostic algorithm based on MALDI-TOF MS, lateral flow immunoassays (LFIAs) and molecular testing performed directly from positive blood cultures (BCs) for Gram-negative species identification and detection of CTX-M extended-spectrum beta-lactamases and main carbapenemases. Methods: Non-duplicate BCs positive to Gram-negative bacteria at microscope examination were subjected to species identification by direct MALDI-TOF MS following recovery of bacterial pellet by Rapid MBT Sepsityper (R) kit. Subsequently, NG-Test (R) CARBA 5 and NG-Test (R) CTX-M MULTI LFIAs were performed according to identified microbial species. Eazyplex (R) SuperBug CRE molecular assay was performed in cases of NG-Test (R) CARBA 5 negative results in patients with documented carbapenemase-producers carriage. Results of rapid diagnostic workflow were compared with those obtained by conventional diagnostic routine. Results: Overall, the direct MALDI-TOF MS protocol allowed reliable identification to the species level of 92.1% of the 2133 monomicrobial BCs. Rate of matched identification was significantly higher for Enterobacterales (97.3%) in comparison to non-fermenting Gram-negative species (80.2%), obligate anaerobic bacteria (42.1%) and fastidious Gram-negative species (41.5%). The overall sensitivity of NG-Test (R) CARBA 5 and NG-Test (R) CTX-M MULTI was 92.2% and 91.6%, respectively. Integration of Easyplex (R) SuperBug CRE allowed the detection of bla(KPC) mutants associated with ceftazidime/avibactam resistance, reaching 100% sensitivity in carbapenemase detection. Both LFIAs and molecular testing showed no false-positive results. Conclusions: Algorithms based on MALDI-TOF MS, LFIAs and molecular testing may represent a cost-effective tool to timely identify Gram-negative species and detect resistance markers directly from BCs. According to local epidemiology, these results may allow antimicrobial stewardship interventions including prompt use of new approved drugs.

Automated summary

This study evaluated a rapid diagnostic algorithm based on MALDI-TOF MS, lateral flow immunoassays (LFIAs), and molecular testing for the identification of Gram-negative species and detection of beta-lactamase and carbapenemase genes directly from positive blood cultures (BCs). The results showed that this algorithm allowed reliable species identification and timely detection of resistance markers. Integration of molecular testing further improved the detection of specific resistance genes.