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Peer-reviewedCC BY 4.0mSystems

Comparative gut microbiome in diarrheal and non-diarrheal children: an individually matched case-control study

  • Jialiang Xu1
  • Yufeng Fan2
  • Geruo Qu1,3
  • Jiaqi Li1,3
  • Yao Peng3
  • Mengyu Wang3,4
  • Jingyun Zhang3
  • Yuqing Feng5
  • Xiao Liu3
  • Yongfei Hu5
  • Biao Kan3,4
  • Zhenpeng Li3
2 more authors
  • Mei Zeng6
  • Xin Lu3
6 affiliations
  1. 1School of Light Industry Science and Engineering, Beijing Technology and Business University, Beijing, China
  2. 2Department of Clinical Laboratory, Shanxi Bethune Hospital, Shanxi Academy of Medical Sciences, Third Hospital of Shanxi Medical University, Tongji Shanxi Hospital, Taiyuan, China
  3. 3National Key Laboratory of Intelligent Tracking and Forecasting for Infectious Diseases, National Institute for Communicable Disease Control and Prevention, Chinese Center for Disease Control and Prevention and Chinese Academy of Preventive Medicine, Beijing, China
  4. 4School of Public Health, Cheeloo College of Medicine, Shandong University, Jinan, Shandong, China
  5. 5State Key Laboratory of Animal Nutrition and Feeding, College of Animal Science and Technology, China Agricultural University, Beijing, China
  6. 6Department of Infectious Disease, Children’s Hospital of Fudan University, National Children’s Medical Center, Shanghai, China
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Abstract

Diarrhea, a leading cause of under-five mortality in developing countries, drives therapeutic challenges amid global antibiotic resistance. This study aimed to compare gut microbiota, antibiotic resistance genes (ARGs), mobile genetic elements (MGEs), and virulence factors (VFs) in diarrheal and non-diarrheal children, explore reasons for asymptomatic pathogen carriage, and develop a microbiome-based model to predict the potential etiology of diarrhea. This individually matched case-control study analyzed 42 paired fecal samples (selected from 716 diarrhea/non-diarrhea cases) to compare gut microbiome profiles, ARGs, MGEs, and VFs using metagenomic sequencing and pathogen-specific PCR. Diarrheal children showed reduced alpha diversity, increased Proteobacteria, and elevated VFs and MGEs. Surprisingly, ARGs were more abundant in non-diarrheal children, suggesting ARG colonization in healthy hosts. Actinomyces was enriched in non-diarrheal pathogen carriers and may be associated with asymptomatic bacterial pathogen carriage. A random forest (RF) model incorporating non-pathogenic bacteria achieved high accuracy in predicting diarrheal status and pathogen carriage. This study reveals distinct microbial ecologies between diarrheal and healthy children. The enrichment of specific bacterial taxa in asymptomatic carriers may be associated with pathogen tolerance. The unexpected ARG abundance in healthy children highlights a hidden antimicrobial resistance reservoir. These findings inform microbiome-based diagnostics and antibiotic stewardship in pediatric diarrhea.

IMPORTANCE: This study is significant because it provides robust, individually matched case-control evidence linking childhood diarrhea to gut microbiome dysbiosis and the distribution of pathogenic and resistance-related genetic elements. By integrating metagenomic sequencing with pathogen-specific polymerase chain reaction (PCR), it offers a comprehensive comparison of microbial composition, virulence factors (VFs), mobile genetic elements (MHEs), and antibiotic resistance genes (ARGs) between diarrheal and non-diarrheal children. The findings demonstrate reduced microbial diversity and enrichment of Proteobacteria, virulence factors, and mobile genetic elements in diarrheal cases, highlighting microbiome instability during infection. Importantly, the unexpectedly higher abundance of antibiotic resistance genes in non-diarrheal children underscores the underestimated role of healthy populations as reservoirs of resistance. Furthermore, the establishment of predictive models for diarrhea status and pathogen carriage enhances the translational value of the study. Overall, this work advances the understanding of pediatric diarrheal disease and informs prevention, surveillance, and treatment strategies in the context of global antimicrobial resistance.

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Cite

BibTeXRIS
BibTeX
@article{xu2026comparative,
  title = {{Comparative gut microbiome in diarrheal and non-diarrheal children: an individually matched case-control study}},
  author = {Jialiang Xu and Yufeng Fan and Geruo Qu and Jiaqi Li and Yao Peng and Mengyu Wang and Jingyun Zhang and Yuqing Feng and Xiao Liu and Yongfei Hu and Biao Kan and Zhenpeng Li and Mei Zeng and Xin Lu},
  journal = {mSystems},
  year = {2026},
  month = sep,
  doi = {10.1128/msystems.00198-26},
  url = {https://doi.org/10.1128/msystems.00198-26},
  note = {CC BY 4.0},
}
RIS
TY  - JOUR
TI  - Comparative gut microbiome in diarrheal and non-diarrheal children: an individually matched case-control study
AU  - Jialiang Xu
AU  - Yufeng Fan
AU  - Geruo Qu
AU  - Jiaqi Li
AU  - Yao Peng
AU  - Mengyu Wang
AU  - Jingyun Zhang
AU  - Yuqing Feng
AU  - Xiao Liu
AU  - Yongfei Hu
AU  - Biao Kan
AU  - Zhenpeng Li
AU  - Mei Zeng
AU  - Xin Lu
PY  - 2026
DA  - 2026/09/30
JO  - mSystems
DO  - 10.1128/msystems.00198-26
UR  - https://doi.org/10.1128/msystems.00198-26
AB  - Diarrhea, a leading cause of under-five mortality in developing countries, drives therapeutic challenges amid global antibiotic resistance. This study aimed to compare gut microbiota, antibiotic resistance genes (ARGs), mobile genetic elements (MGEs), and virulence factors (VFs) in diarrheal and non-diarrheal children, explore reasons for asymptomatic pathogen carriage, and develop a microbiome-based model to predict the potential etiology of diarrhea. This individually matched case-control study analyzed 42 paired fecal samples (selected from 716 diarrhea/non-diarrhea cases) to compare gut microbiome profiles, ARGs, MGEs, and VFs using metagenomic sequencing and pathogen-specific PCR. Diarrheal children showed reduced alpha diversity, increased Proteobacteria, and elevated VFs and MGEs. Surprisingly, ARGs were more abundant in non-diarrheal children, suggesting ARG colonization in healthy hosts. Actinomyces was enriched in non-diarrheal pathogen carriers and may be associated with asymptomatic bacterial pathogen carriage. A random forest (RF) model incorporating non-pathogenic bacteria achieved high accuracy in predicting diarrheal status and pathogen carriage. This study reveals distinct microbial ecologies between diarrheal and healthy children. The enrichment of specific bacterial taxa in asymptomatic carriers may be associated with pathogen tolerance. The unexpected ARG abundance in healthy children highlights a hidden antimicrobial resistance reservoir. These findings inform microbiome-based diagnostics and antibiotic stewardship in pediatric diarrhea.  IMPORTANCE: This study is significant because it provides robust, individually matched case-control evidence linking childhood diarrhea to gut microbiome dysbiosis and the distribution of pathogenic and resistance-related genetic elements. By integrating metagenomic sequencing with pathogen-specific polymerase chain reaction (PCR), it offers a comprehensive comparison of microbial composition, virulence factors (VFs), mobile genetic elements (MHEs), and antibiotic resistance genes (ARGs) between diarrheal and non-diarrheal children. The findings demonstrate reduced microbial diversity and enrichment of Proteobacteria, virulence factors, and mobile genetic elements in diarrheal cases, highlighting microbiome instability during infection. Importantly, the unexpectedly higher abundance of antibiotic resistance genes in non-diarrheal children underscores the underestimated role of healthy populations as reservoirs of resistance. Furthermore, the establishment of predictive models for diarrhea status and pathogen carriage enhances the translational value of the study. Overall, this work advances the understanding of pediatric diarrheal disease and informs prevention, surveillance, and treatment strategies in the context of global antimicrobial resistance.
SN  - 2379-5077
ER  -