Suppression PCR-based selective enrichment sequencing for pathogen and antimicrobial resistance detection on cell-free DNA in sepsis

Sepsis is a life-threatening syndrome triggered by infection and accompanied by high mortality, with antimicrobial resistances (AMRs) further escalating clinical challenges. Rapid and reliable detection of causative pathogens and AMRs is a key factor for fast and appropriate treatment, which increases survival chances for septic patients. However, current diagnostics based on blood culture is limited by low sensitivity and specificity while current molecular approaches fail to enter clinical routine. We therefore developed a Suppression PCR-based selective enrichment sequencing of target sequences approach (SUPSETS), providing a molecular method combining multiplex suppression PCR with Nanopore sequencing to identify most common sepsis pathogens and AMRs using cell-free DNA. Applying only 1 mL of plasma, we target eight pathogens across three kingdoms and ten AMRs in a proof-of-concept study. SUPSETS was successfully tested on first clinical samples and revealed comparable results to clinical metagenomics while clearly outperforming blood culture. Several clinically relevant AMRs could be detected additionally. Furthermore, SUPSETS provided first pathogen and AMR-specific sequencing reads within only few minutes after sequencing start, thereby potentially decreasing time to results to 11 - 13 hours suggesting diagnostic potential in sepsis.