Stool Dynamics and the Developing Gut Microbiome During Infancy

The infant gut microbiome is a dynamic ecosystem, and it is key to early development, immune maturation, and overall health. Recent insights reveal that the gut microbiota undergoes changes across the 24-h day, raising the possibility that it may act as a "zeitgeber," supporting the host's sleep-wake organization. Despite its importance, timing factors influencing microbiome composition are poorly understood, limiting its use as a health indicator. This study investigates the relationship between stool dynamics (defecation interval, time of sampling), sleep pressure (interval since last sleep), meal timing, and gut microbial composition. Stool samples from 198 healthy infants, aged 3 to 31 months, were analyzed to assess microbial diversity, richness evenness, and abundance. Our findings reveal that longer intervals between bowel movements are associated with increased microbial diversity, evenness, and richness. Stool timing is associated with shifts in microbial composition, especially in younger infants, indicating diurnal microbial fluctuations to become more stable as infants mature. Longer periods of wakefulness were associated with increased microbial diversity in early infancy, although this effect appeared to diminish with age. Feeding schedules had limited effects on the gut microbiome. Longer fasting before sampling showed no significant associations with most microbial parameters, except for a positive association with microbial richness. At the phylum level, results indicate that infant gut microbial composition is influenced by behavior and physiology. Longer intervals between bowel movements were associated with shifts in bacterial abundance, with Proteobacteria decreasing and Actinobacteria increasing. In addition, later stool sampling times revealed higher Actinobacteria levels, and longer fasting was associated with reduced Bacteroidetes. Sleep pressure showed a trend effect with Firmicutes displaying a slight decrease in infants who had been awake longer. Our findings underscore the importance of time-based factors on infant gut microbiome composition.