e-Article
Suppression PCR-Based Selective Enrichment Sequencing for Pathogen and Antimicrobial Resistance Detection on Cell-Free DNA in Sepsis-A Targeted, Blood Culture-Independent Approach for Rapid Pathogen and Resistance Diagnostics in Septic Patients.
Document Type
Academic Journal
Author
Sonntag M; Innovation Field In-Vitro Diagnostics, Fraunhofer Institute for Interfacial Engineering and Biotechnology IGB, 70569 Stuttgart, Germany.; Interfaculty Graduate School of Infection Biology and Microbiology (IGIM), Eberhard Karls University Tuebingen, 72076 Tuebingen, Germany.; Elgeti VK; Innovation Field In-Vitro Diagnostics, Fraunhofer Institute for Interfacial Engineering and Biotechnology IGB, 70569 Stuttgart, Germany.; Faculty of Medicine, Greifswald University Medicine, Fleischmannstr. 8, 17475 Greifswald, Germany.; Vainshtein Y; Innovation Field In-Vitro Diagnostics, Fraunhofer Institute for Interfacial Engineering and Biotechnology IGB, 70569 Stuttgart, Germany.; Jenner L; Innovation Field In-Vitro Diagnostics, Fraunhofer Institute for Interfacial Engineering and Biotechnology IGB, 70569 Stuttgart, Germany.; Mueller J; Innovation Field In-Vitro Diagnostics, Fraunhofer Institute for Interfacial Engineering and Biotechnology IGB, 70569 Stuttgart, Germany.; Max Perutz Labs, Vienna Biocenter Campus (VBC), Vienna Biocenter 5, 1030 Vienna, Austria.; Max Perutz Labs, Department of Structural and Computational Biology, University of Vienna, CIBIV Vienna Biocenter 5, 1030 Vienna, Austria.; Vienna Biocenter PhD Program, a Doctoral School of the University of Vienna and the Medical University of Vienna, 1030 Vienna, Austria.; Brenner T; Department of Anesthesiology and Intensive Care Medicine, University Hospital Essen, University Duisburg-Essen, Hufelandstr. 55, 45147 Essen, Germany.; Decker SO; Department of Anesthesiology, Medical Faculty Heidelberg, Heidelberg University, Im Neuenheimer Feld 420, 69120 Heidelberg, Germany.; Sohn K; Innovation Field In-Vitro Diagnostics, Fraunhofer Institute for Interfacial Engineering and Biotechnology IGB, 70569 Stuttgart, Germany.
Source
Publisher: MDPI Country of Publication: Switzerland NLM ID: 101092791 Publication Model: Electronic Cited Medium: Internet ISSN: 1422-0067 (Electronic) Linking ISSN: 14220067 NLM ISO Abbreviation: Int J Mol Sci Subsets: MEDLINE
Subject
Language
English
Abstract
Sepsis is a life-threatening syndrome triggered by infection and accompanied by high mortality, with antimicrobial resistances (AMRs) further escalating clinical challenges. The rapid and reliable detection of causative pathogens and AMRs are key factors for fast and appropriate treatment, in order to improve outcomes in septic patients. However, current sepsis diagnostics based on blood culture is limited by low sensitivity and specificity while current molecular approaches fail to enter clinical routine. Therefore, we developed a suppression PCR-based selective enrichment sequencing approach (SUPSETS), providing a molecular method combining multiplex suppression PCR with Nanopore sequencing to identify most common sepsis-causative pathogens and AMRs using plasma cell-free DNA. Applying only 1 mL of plasma, we targeted eight pathogens across three kingdoms and ten AMRs in a proof-of-concept study. SUPSETS was successfully tested in an experimental research study on the first ten clinical samples and revealed comparable results to clinical metagenomics while clearly outperforming blood culture. Several clinically relevant AMRs could be additionally detected. Furthermore, SUPSETS provided first pathogen and AMR-specific sequencing reads within minutes of starting sequencing, thereby potentially decreasing time-to-results to 11-13 h and suggesting diagnostic potential in sepsis.