CC BY 4.0Brenner, ThorstenThorstenBrennerDecker, Sebastian OliverSebastian OliverDeckerVainshtein, YevhenYevhenVainshteinGrumaz, SilkeSilkeGrumazManoochehri, MehdiMehdiManoochehriFeißt, ManuelManuelFeißtSeidel-Glätzer, AndreaAndreaSeidel-GlätzerPletz, Mathias Wilhelm RudolfMathias Wilhelm RudolfPletzBracht, HendrikHendrikBrachtBerger, Marc MoritzMarc MoritzBergerFuest, Kristina E.Kristina E.FuestBlobner, ManfredManfredBlobnerBach, FriedhelmFriedhelmBachMöerer, OnnenOnnenMöererBrandenburger, TimoTimoBrandenburgerDimski, ThomasThomasDimskiSuchodolski, KlaudiuszKlaudiuszSuchodolskiJäkel, UlrikeUlrikeJäkelZischkau, JanaJanaZischkauHäberle, Helene AnnaHelene AnnaHäberleRosenberger, PeterPeterRosenbergerSchürholz, TobiasTobiasSchürholzLindau, SimoneSimoneLindauSchaller, Stefan J.Stefan J.SchallerPutensen, Christian P.Christian P.PutensenDusse, FabianFabianDussePetros, SirakSirakPetrosGaasch, MaxMaxGaaschNusshag, ChristianChristianNusshagWeigand, Markus AlexanderMarkus AlexanderWeigandSohn, KaiKaiSohnGlanz, KarolinaKarolinaGlanzBarth, EberhardEberhardBarthWinkler, Martin SebastianMartin SebastianWinklerGillmann, Hans J.Hans J.Gillmann2025-09-042025-09-042025https://publica.fraunhofer.de/handle/publica/494892https://doi.org/10.24406/publica-528910.1016/j.jinf.2025.10656510.24406/publica-52892-s2.0-10501332698440774415Objectives: Despite limited sensitivity and specificity, blood cultures (BCs) still represent the gold standard of diagnostic care in septic patients. We aimed to overcome current diagnostic limitations by unbiased next-generation sequencing (NGS) of circulating microbial cell-free DNA (mcfDNA) in plasma samples. Methods: We performed a prospective, observational, non-interventional, multicenter study (Next GeneSiS-Trial) to compare positivity rates for NGS-based identification of causative pathogens with BCs in patients suffering from sepsis or septic shock. An independent expert panel (n=3) retrospectively evaluated the plausibility of NGS-based findings and the potential for anti-infective treatment adaptations based on NGS results. Results: The positivity rate of NGS-based diagnostics (NGS+) for 491 septic patients was 70.5% compared to positive BCs (BC+) with 19.4% within the first three days after sepsis onset. NGS+ results were evaluated as plausible in 98.6% of cases by the expert panel. Based on the experts´ recommendations, additional knowledge of NGS-based pathogen findings would have resulted in anti-infective treatment adaptations in 32.6% of all patients. Potentially inadequately treated NGS+/blood culture negative (BC-) patients showed worse outcomes. Conclusion: The integration of NGS-based pathogen diagnostics in sepsis has the potential to improve patients´ outcomes as compared to a treatment strategy based on standard-of-care microbiological diagnostics alone.entrueBacteremiaBlood cultureClinical metagenomicsCritical careMicrobial cell-free DNAMolecular diagnosticsNext-generation sequencingNGSOutcomejournal article