Metagenomic Cell-free DNA Sequencing for Treatment Monitoring in Sepsis

Sepsis is a life-threatening organ dysfunction caused by a dysregulated response to infection. Early identification of pathogens and accurate assessment of organ injury are critical for improving outcomes, but current methods are often inadequate, especially after initiation of antibiotic treatment. Metagenomic sequencing of cell-free DNA (cfDNA) offers a promising alternative, enabling simultaneous pathogen detection and tissue-of-origin profiling. Contamination, however, can limit its accuracy in low-biomass samples. Here, we apply the Sample-Intrinsic Microbial DNA Found by Tagging and Sequencing (SIFT-seq) assay, which reduces contamination and allows detection of pathogens and organ injury simultaneously. We analyzed 142 plasma specimens: 105 from sepsis patients, 103 collected after initiation of antibiotic treatment, 24 from non-sepsis ICU controls, and 13 from healthy controls. SIFT-seq identified sepsis-causing pathogens in good agreement with pre-antibiotic blood cultures, revealed elevated immune activity and organ injury in sepsis patients, and, when combined with the SOFA score in a multivariate model, improved diagnostic performance (AUC = 0.874). These findings highlight the potential of integrated cfDNA profiling to enhance sepsis diagnosis.