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Peer-reviewedCC BYFrontiers in microbiology

Targeted next-generation sequencing: a promising approach for pathogen and antimicrobial resistance gene detection in urinary tract infections

  • Wenzhi Gao1
  • Wencong Han1
  • Jiachang Chen2
  • Zihui Gao3
  • Jingjing Gao4
  • Rui Zhen4
  • Ya Yuan4
  • Yaming Gu3
  • Chen Huang5
  • Chunlin Liu6
  • Gang Wang1
  • Xiaochun Zhang1
8 more authors
  • Zheng Zhao1
  • Gengyan Xiong1
  • Xuesong Li1
  • Pengyuan Zhu2
  • Jun Liu2
  • Chaohui Hu2
  • Yanqing Gong1
  • Zheng Zhang1
6 affiliations
  1. 1Department of Urology, Peking University First Hospital , ,
  2. 2Guangzhou KingCreate Biotechnology Co., Ltd. , ,
  3. 3Department of Urology, Peking University First Hospital-Miyun Hospital , ,
  4. 4Nursing Department, Peking University First Hospital , ,
  5. 5Department of Urology, Jian Gong Hospital , ,
  6. 6Department of Urology, FuXing Hospital Affiliated to Capital Medical University , ,
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Abstract

Objective: Urinary tract infections (UTIs) are common and frequently recurrent. As traditional diagnostic methods fail to meet clinical needs, this study aimed to construct and evaluate the performance of targeted next-generation sequencing (tNGS) for detecting pathogens and antimicrobial resistance genes (ARGs) in urine specimens.

Methods: A total of 446 urine samples from patients with suspected UTIs were tested by tNGS, routine culture, and antimicrobial susceptibility testing (AST), with discordant results resolved by metagenomic NGS (mNGS) and whole-genome sequencing (WGS). The performance of tNGS was evaluated against culture and a composite reference standard (CRS) for pathogen detection, and against AST and WGS for resistance prediction.

Results: tNGS achieved limit of detections of 500–1,000 CFU/mL, with high precision. Compared to culture, tNGS achieved a higher positive rate (62.78%), along with an overall sensitivity of 90.05% and specificity of 56.28%, whereas against the CRS the sensitivity and specificity reached 99.54% and 97.49%, respectively. For the seven common UTI bacteria, both metrics exceeded 90% regardless of the reference standard. Additionally, tNGS outperformed culture in detecting hard-to-culture microbes and polymicrobial infections. Among 132 samples with available AST results, ARG-based prediction showed strong concordance with resistance phenotypes for carbapenems (100.00%) and third generation cephalosporins + β-lactamase inhibitors (91.67%). Factors including sex and urine leukocyte count > 10 cells/μL were found associated with positive tNGS results.

Conclusion: tNGS enables rapid, sensitive and comprehensive detection of UTI pathogens and ARGs, covering hard-to-culture pathogens and mixed infections, thereby showing promise in improving diagnosis. However, it complements rather than replaces phenotypic AST, particularly for penicillin-class antibiotics.

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Cite

BibTeXRIS
BibTeX
@article{gao2026targeted,
  title = {{Targeted next-generation sequencing: a promising approach for pathogen and antimicrobial resistance gene detection in urinary tract infections}},
  author = {Wenzhi Gao and Wencong Han and Jiachang Chen and Zihui Gao and Jingjing Gao and Rui Zhen and Ya Yuan and Yaming Gu and Chen Huang and Chunlin Liu and Gang Wang and Xiaochun Zhang and Zheng Zhao and Gengyan Xiong and Xuesong Li and Pengyuan Zhu and Jun Liu and Chaohui Hu and Yanqing Gong and Zheng Zhang},
  journal = {Frontiers in microbiology},
  year = {2026},
  month = sep,
  doi = {10.3389/fmicb.2026.1945070},
  url = {https://europepmc.org/article/PMC/PMC13661925},
  note = {CC BY},
}
RIS
TY  - JOUR
TI  - Targeted next-generation sequencing: a promising approach for pathogen and antimicrobial resistance gene detection in urinary tract infections
AU  - Wenzhi Gao
AU  - Wencong Han
AU  - Jiachang Chen
AU  - Zihui Gao
AU  - Jingjing Gao
AU  - Rui Zhen
AU  - Ya Yuan
AU  - Yaming Gu
AU  - Chen Huang
AU  - Chunlin Liu
AU  - Gang Wang
AU  - Xiaochun Zhang
AU  - Zheng Zhao
AU  - Gengyan Xiong
AU  - Xuesong Li
AU  - Pengyuan Zhu
AU  - Jun Liu
AU  - Chaohui Hu
AU  - Yanqing Gong
AU  - Zheng Zhang
PY  - 2026
DA  - 2026/09/30
JO  - Frontiers in microbiology
DO  - 10.3389/fmicb.2026.1945070
UR  - https://europepmc.org/article/PMC/PMC13661925
AB  - Objective: Urinary tract infections (UTIs) are common and frequently recurrent. As traditional diagnostic methods fail to meet clinical needs, this study aimed to construct and evaluate the performance of targeted next-generation sequencing (tNGS) for detecting pathogens and antimicrobial resistance genes (ARGs) in urine specimens.  Methods: A total of 446 urine samples from patients with suspected UTIs were tested by tNGS, routine culture, and antimicrobial susceptibility testing (AST), with discordant results resolved by metagenomic NGS (mNGS) and whole-genome sequencing (WGS). The performance of tNGS was evaluated against culture and a composite reference standard (CRS) for pathogen detection, and against AST and WGS for resistance prediction.  Results: tNGS achieved limit of detections of 500–1,000 CFU/mL, with high precision. Compared to culture, tNGS achieved a higher positive rate (62.78%), along with an overall sensitivity of 90.05% and specificity of 56.28%, whereas against the CRS the sensitivity and specificity reached 99.54% and 97.49%, respectively. For the seven common UTI bacteria, both metrics exceeded 90% regardless of the reference standard. Additionally, tNGS outperformed culture in detecting hard-to-culture microbes and polymicrobial infections. Among 132 samples with available AST results, ARG-based prediction showed strong concordance with resistance phenotypes for carbapenems (100.00%) and third generation cephalosporins + β-lactamase inhibitors (91.67%). Factors including sex and urine leukocyte count > 10 cells/μL were found associated with positive tNGS results.  Conclusion: tNGS enables rapid, sensitive and comprehensive detection of UTI pathogens and ARGs, covering hard-to-culture pathogens and mixed infections, thereby showing promise in improving diagnosis. However, it complements rather than replaces phenotypic AST, particularly for penicillin-class antibiotics.
SN  - 1664-302X
ER  -