Plasma Cell-Free DNA Sequencing Now Detects Antimicrobial Resistance Markers Directly from Blood
This validation study demonstrates that plasma microbial cell-free DNA sequencing can directly detect seven key antimicrobial resistance markers (including mecA, vanA, and blaCTX-M) across 18 target pathogens without culture. Clinical validation showed 95% positive and 95% negative percent agreement for MRSA detection, and 100% agreement for vancomycin-resistant enterococci. The addition of AMR detection to an existing mcfDNA platform gives clinicians a culture-independent tool for optimizing antimicrobial therapy, particularly in critically ill patients.
The original study
Analytical and clinical validation of direct detection of antimicrobial resistance markers by plasma microbial cell-free DNA sequencing.
- Authors
- Christians FC, Akhund-Zade J, Jarman K, Venkatasubrahmanyam S, Noll N, Blauwkamp TA, et al.
- Journal
- Journal of clinical microbiology
- Type
- Journal Article, Validation Study
- PMID
- 39194269
Original abstract
UNLABELLED: Sequencing of plasma microbial cell-free DNA (mcfDNA) has gained increased acceptance as a valuable adjunct to standard-of-care testing for diagnosis of infections throughout the body. Here, we report the analytical and clinical validation of a novel application of mcfDNA sequencing, the non-invasive detection of seven common antimicrobial resistance (AMR) genetic markers in 18 important pathogens. The AMR markers include SCCmec, mecA, mecC, vanA, vanB, blaCTX-M, and blaKPC. The AMR markers were computationally linked to the pathogens detected. Analytical validation showed high reproducibility (100%), inclusivity (54 to 100%), and exclusivity (100%). Clinical accuracy was assessed with 114 unique plasma samples from patients at seven study sites with concordant culture results for target bacteria from a variety of specimen types and correlated with available phenotypic antimicrobial susceptibility test results and genotypic results. The positive percent agreement (PPA), negative percent agreement (NPA), and diagnostic yield (DY) were estimated for each AMR marker. DY was defined as the percentage of tests that yielded an actionable result of either detected or not detected. The results for the combination of SCCmec and mecA for staphylococci were PPA 19/20 (95.0%), NPA 21/22 (95.4%), DY 42/60 (70.0%); vanA for enterococci were PPA 3/3 (100%), NPA 2/2 (100%), DY 5/6 (83.3%); blaCTX-M for gram-negative bacilli were PPA 5/6 (83.3%), NPA 29/29 (100%), DY 35/49 (71.4%); and blaKPC for gram-negative bacilli were PPA 0/2 (0%), NPA: 23/23 (100%), DY 25/44 (56.8%). The addition of AMR capability to plasma mcfDNA sequencing should provide clinicians with an effective new culture-independent tool for optimization of therapy. IMPORTANCE: This manuscript is ideally suited for the Innovative Diagnostic Methods sections as it reports the analytical and clinical validation of a novel application of plasma microbial cell-free DNA sequencing for direct detection of seven selected antimicrobial resistance markers in 18 target pathogens. Clearly, it has potential clinical utility in optimizing therapy and was incorporated into the Karius test workflow in September 2023. In addition, the workflow could readily be adapted to expand the number of target bacteria and antimicrobial resistance markers as needed.