Journal Article

Ultra-broad hybrid capture-based targeted next-generation sequencing for sensitive plasma pathogen cfDNA detection in bloodstream infections

Li, Min, Xu, Ke, Gu, Dejian, Mao, Shaowei, Li, Shuangshuang, Wei, Muyun, Zhang, Shengyang, Ai, Xiangzhao

2025

Overview

Background The limited genomic targeting range of current targeted next-generation sequencing (tNGS) workflows results in limited detection of pathogen-derived cell-free DNA (cfDNA), making it challenging to apply this approach to bloodstream infections (BSIs). Here, we developed an ultra-broad hybrid capture-based tNGS method to detect plasma pathogen-derived cfDNA and evaluate its suitability for the diagnosis of BSI. Methods This study introduced an ultra-broad hybrid capture-based tNGS method featuring an ultra-broad pathogen panel (1872 pathogens) and high-density probe coverage. To adequately evaluate its performance, we conducted retrospective tests in 208 suspected BSI patients (139 immunocompromised), comparing tNGS results with mNGS, conventional microbiological testing (CMT), and comprehensive clinical diagnoses. Results In pathogen detection, the concordance between ultra-broad hybrid capture-based tNGS and mNGS results was 93.75%. The diagnostic accuracy of tNGS in BSI was comparable to mNGS (76.44% vs. 75.00%) and significantly higher than CMT (45.67%, p  < 0.0001). In immunocompromised populations, the diagnostic accuracy of tNGS was similar to mNGS (77.70% vs. 76.98%). tNGS detected 92.09% (163/177) of pathogens identified by mNGS. Two of the missed pathogens were not included in the 1872 pathogens panel, and both were from the immunocompromised group. Conclusions Ultra-broad hybrid capture-based tNGS exhibits sensitivity and accuracy comparable to mNGS, effectively covering a relatively wide range of pathogens, and may serve as an economic screening tool for BSI in the future.

• Publisher: BioMed Central,BioMed Central Ltd,Springer Nature B.V,BMC
• Subject: Adult / Aged / Antibiotics / Autoimmune diseases / Biomedical and Life Sciences / Biomedicine / Blood / Blood plasma / Bloodstream infections (BSI) / Cell-Free Nucleic Acids - blood / Cell-Free Nucleic Acids - genetics / CfDNA / Chemotherapy / DNA / DNA sequencing / Female / Genetic aspects / Health aspects / Hematological diseases / High-Throughput Nucleotide Sequencing - methods / Humans / Immunocompetence / Immunocompromised patients / Infections / Male / Medical prognosis / Medical research / Medicine, Experimental / Medicine/Public Health / Metagenomic next-generation sequencing (mNGS) / Middle Aged / Next-generation sequencing / Nucleotide sequencing / Pathogens / Patients / Physiological aspects / Plasma / Retrospective Studies / Sepsis - blood / Sepsis - microbiology / Septicemia / Steroids / Translational Metagenomics / Transplants & implants / Ultra-broad hybrid capture-based targeted next-generation sequencing (tNGS)