Gut microbiota patterns associated with duration of diarrhea in children under five years of age in Ethiopia

Introduction: Diarrhea claims about 500,000 lives annually among children under five years of age and about 90% of those deaths occur in Sub-Saharan Africa and South Asia. Childhood mortality due to acute diarrhea (AD; less than 7 days of duration) is decreasing, but prolonged (ProD; 7-13 days of duration) and persistent (PD; more than 14 days of duration) diarrhea combined (ProPD) are responsible for increased morbidity and mortality in low- and middle- income countries (LMICs). Objectives: This study aimed to characterize the gut microbiota (GM) composition of children with acute diarrhea (AD) and prolonged or persistent diarrhea (ProPD) by comparing GM composition between these two groups and with non-diarrheal controls in a case-control study. Methods: The study included fecal samples from 1321 children 0-59 months old. After preprocessing based on sequencing depth, we analyzed the GM of 1313 children comparing 554 AD cases, 95 ProPD cases, and 663 frequency matched, non-diarrheal controls. All children were enrolled from Jimma, Ethiopia. The GM composition was determined using 16S rRNA gene (V4 region) amplicon sequencing. The observed zero-radius Operational Taxonomic Units (zOTUs) and Shannon diversity index were assessed, and Bray-Curtis dissimilarity metrics were generated and differences between groups evaluated by PERMANOVA. DESeq2 was used to determine taxa being differentially abundant between diarrhea subcategories and non-diarrheal controls. Results: All diarrhea cases (AD and ProPD) combined and each group separately were associated with a decrease in bacterial diversity compared to non-diarrheal controls. Relative to non-diarrheal controls, the GM of children with AD or ProPD was enriched in Escherichia spp., Campylobacter spp., and Streptococcus spp. The GM of children with AD or ProPD was characterized by a decreased relative abundance of Prevotella copri, Faecalibacterium prausnitzii and Dialister succinatiphilus compared to non-diarrheal controls. Similarly, compared to AD cases, ProPD cases were characterized by a decreased relative abundance of important gut commensals such as F. prausnitzii, Akkermansia muciniphila, Eubacterium hadrum and Blautia spp. Conclusion: The GM of children with AD and ProPD was enriched in putative pathogens as well as facultative anaerobes, while obligate anaerobes were reduced. The GM of children with ProPD was characterized by a more pronounced reduction of obligate anaerobic gut commensals compared to AD, suggesting that prolonged duration of diarrhea is accompanied by a depletion of healthy gut commensals. This indicates a potential role for targeted dietary supplements to restore gut eubiosis and/ or the use of next generation probiotics to improve ProPD treatment.