Integrated Transcriptomic and Regulatory Network Analysis of Childhood Infectious Diarrhea Reveals Broad Whole-Blood Host Remodeling with Pathway-Level Validation in an Independent Cohort

Background: Childhood infectious diarrhea elicits systemic host responses, but computational findings from public transcriptomic data require independent validation. Methods: We reanalyzed the pediatric whole-blood RNA-sequencing dataset GSE69529 using differential-expression, functional-enrichment, protein-association, miRNA-network, promoter-sequence, motif, and transcription-factor activity analyses. Pathway-level external validation used the independent whole-blood cohort GSE276395, comprising 278 samples from multiple diarrhea groups and healthy controls. Results: We identified 7,050 differentially expressed genes among 15,740 analyzed genes at adjusted P < 0.05. Enrichment implicated immune signaling, RNA metabolism, cell-cycle control, intracellular organization, apoptosis, membrane trafficking, and metabolism. Protein-association and experimentally supported miRNA-gene networks were broadly connected. Promoter composition was similar between regulatory directions; AME identified no differentially enriched HOCOMOCO motifs, STREME candidates failed the prespecified E-value threshold, and no CollecTRI-ULM regulator remained significant after false-discovery-rate correction. The independent cohort reproduced innate-immune, neutrophil-centered, and immunometabolic themes, but its 12-gene signature did not overlap the 12 discovery candidates or validate CFTR- or SLC26A3-linked hypotheses. Conclusions: Broad whole-blood host remodeling is supported across cohorts, whereas gene-, motif-, and regulator-level findings remain exploratory and require experimental validation.