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  4. 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
 
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May 17, 2024
Journal Article
Title

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

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.
Author(s)
Sonntag, Mirko  
Fraunhofer-Institut für Grenzflächen- und Bioverfahrenstechnik IGB  
Elgeti, Vanessa Kimberly
Fraunhofer-Institut für Grenzflächen- und Bioverfahrenstechnik IGB  
Vainshtein, Yevhen  orcid-logo
Fraunhofer-Institut für Grenzflächen- und Bioverfahrenstechnik IGB  
Jenner, Lucca
Fraunhofer-Institut für Grenzflächen- und Bioverfahrenstechnik IGB  
Müller, Jan
Fraunhofer-Institut für Grenzflächen- und Bioverfahrenstechnik IGB  
Brenner, Thorsten
Decker, Sebastian
Sohn, Kai  
Fraunhofer-Institut für Grenzflächen- und Bioverfahrenstechnik IGB  
Journal
International journal of molecular sciences  
Open Access
File(s)
Download (2.62 MB)
Rights
CC BY 4.0: Creative Commons Attribution
DOI
10.3390/ijms25105463
10.24406/publica-3129
Additional full text version
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Language
English
Fraunhofer-Institut für Grenzflächen- und Bioverfahrenstechnik IGB  
Keyword(s)
  • antimicrobial resistances

  • cell-free DNA

  • precision diagnostics

  • next-generation sequencing

  • suppression PCR

  • real-time diagnostics

  • Nanopore sequencing

  • sepsis

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