Evaluation of loop-mediated isothermal amplification for the rapid identification of bacteria and resistance determinants in positive blood cultures
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The use of molecular assays to rapidly identify pathogens and resistance genes directly from positive blood cultures (BCs) contribute to shortening the time required for the diagnosis of bloodstream infections. In this work, loop-mediated isothermal amplification (LAMP) assays have been examined for their potential use in BC diagnosis. Three different assays were applied. The commercially available eazyplex® MRSA test detects Staphylococcus aureus, S. epidermidis, mecA, and mecC. Two in-house assays [Gram-positive (GP) and Gram-negative (GN)] have been developed for the detection of streptococci, enterococci, vanA, vanB, Pseudomonas spp., Enterobacteriaceae, and the bla CTX-M family. A total of 370 positive BCs were analyzed. LAMP test results were obtained within 30 min, including sample preparation. Amplification was measured by real-time fluorescence detection. The threshold time for fluorescence intensity values ranged from 6.25 to 13.75 min. The specificity and sensitivity of the assays varied depending on the target. Overall, from 87.7% of BCs, true-positive results were obtained, compared to routine standard diagnosis. Twenty-one tests were true-negative because of the lack of an appropriate target (5.7%). The concordance of positive test results for resistance genes with subsequent antibiotic susceptibility testing was 100%. From 15 BC bottles with mixed cultures, eazyplex® assays produced correct results in 73% of the cases. This study shows that LAMP assays are fast and cost-saving tools for rapid BC testing in order to expedite the diagnostic report and improve the antibiotic stewardship for sepsis patients.
KeywordsBlood Culture Antimicrobial Susceptibility Testing Cefpodoxime Lamp Assay Blood Culture Bottle
Compliance with ethical standards
The study protocol for the evaluation of LAMP eazyplex® assays for clinical BCs was reviewed and approved by the ethics committee of the Friedrich Schiller University of Jena (4400-04/15).
This study was supported by internal funding from the Jena University Hospital and by a grant from the German Federal Ministry of Education and Research (BMBF, 13N13890).
Conflict of interest
J. Rödel received financial support from Amplex BioSystems to attend the 67th Annual Meeting of the German Society for Hygiene and Microbiology in 2015. No funding was received from the manufacturer of eazyplex® tests for the purpose of the study.
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