ORIGINAL RESEARCH article
Front. Microbiol.
Sec. Microorganisms in Vertebrate Digestive Systems
Volume 16 - 2025 | doi: 10.3389/fmicb.2025.1633162
This article is part of the Research TopicNew Horizons in Gut Microbiome Research for Enhancing Livestock ProductivityView all 11 articles
Age-Stratified Gut Microbial Changes in Diarrheal Calves: Insights
Provisionally accepted
- 1Inner Mongolia University, Hohhot, China
- 2GuangXi University of Chinese Medicine, Nanning, China
- 3Shanghai Jiao Tong University, Shanghai, China
- 4Fudan University, Shanghai, China
- 5Huashan Hospital Fudan University, Shanghai, China
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Neonatal calf diarrhea (NCD) remains a leading cause of mortality in calves under one month, yet how gut microbial responses vary across developmental stages remains unexplored. This study investigates age-stratified microbiome dynamics during NCD to enable precision interventions.This study investigated 60 female Holstein calves (1, 21, and 30 days old) from a commercial dairy farm, equally divided between healthy and diarrheal groups based on standardized fecal scoring. Fecal samples were collected aseptically, flash-frozen, and processed for 16S rRNA gene sequencing (V3-V4 region) using Illumina NovaSeq. Bioinformatics analyses included DADA2 pipeline for ASV calling, SILVA 138 database for taxonomic annotation, ANCOM-BC2 for differential abundance analysis (FDR<0.05), PICRUSt2 for functional prediction, and SparCC networks (|r|>0.6, p < 0.001) with Gephi visualization. Multivariate statistics, including PERMANOVA and canonical correspondence analysis were performed in QIIME2 and R (phyloseq/vegan packages), with all analyses rarefied to 39,161 sequences/sample.The gut microbiome exhibited age-dependent succession, transitioning from Pseudomonadota dominance (47.2±0.7%) at day 1 to Bacillota/Bacteroidota codominance (85.5±8.2%) by day 30. Age explained significantly more compositional variance than diarrhea status (3.68% vs 1.96%, p < 0.001). Three distinct age-specific diarrheal patterns emerged: (1) Early-stage (1-day-old)showed Bacillota/Pseudomonadota imbalances (84% of differential ASVs) with reduced network complexity (total node count, total edge count, average degree and modularity); (2) Mid-lactation (21-day-old) featured Kurthia as both significantly enriched (log2FC=5.32) and a network hub (degree=14); (3) Mature microbiota (30-day-old) displayed complex multi-phylum dysbiosis involving 10 metabolic pathways. Clostridia_UCG-014 persisted across diarrheal networks, while healthy calves showed age-progressive increases in microbial connectivity (edges: 125 to 1104). Only 2 ASVs demonstrated consistent differential abundance across age groups, confirming the temporal specificity of diarrheal dysbiosis.NCD-associated dysbiosis progresses through distinct developmental phases, from resilient phylum-level shifts in neonates to complex network disruptions in mature microbiota. The identification of stage-specific biomarkers (e.g., day 21 Kurthia) opens new avenues for age-tailored probiotic therapies and early intervention strategies.
Keywords: Neonatal diarrhea, calf, Gut Microbiota, developmental dysbiosis, Microbial networks
Received: 22 May 2025; Accepted: 13 Aug 2025.
Copyright: © 2025 Zhang, Li, Tang, Li, Fu, Hao, Li, Feng and Hu. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) or licensor are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.
* Correspondence:
Jian Li, Inner Mongolia University, Hohhot, China
Xinyu Feng, Shanghai Jiao Tong University, Shanghai, China
Wei Hu, Inner Mongolia University, Hohhot, China
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