Effects of prolonged confined space operations on human gut microbiota and serum metabolome

Psychophysiological stress caused by long-term closed environments can disturb the homeostasis of the intestinal flora, which can aggravate bodily stress through metabolic dysfunction, forming a vicious circle. The purpose of this study was to analyze the mechanisms of the intestinal flora and serum metabolism of individuals working in closed environments, reveal the characteristics of the flora and metabolomes of such workers, and provide the basis for health protection. Fecal and serum samples of nine healthy volunteers were collected before entering the cabin, and on days 14 and 28, the intestinal microflora was analyzed via 16S rRNA sequencing. Serum metabolomics analysis was performed via LC-MS. Serum inflammatory factors were analyzed using the targeted proteome Olink technique. A total of 6,373 OTUs were identified. At the gate level, 5/9 of the subjects exhibited significantly increased levels of Proteobacteria on day 14, and 6/9 of the subjects exhibited increased levels of Firmicutes on Day 28. Bacteroides levels continued to decrease. Linear discriminant analysis effect size showed a decrease in Prevotellamassilia timonensis and Alloprevotella and an increase in Clostridiales spp. by day 28. LC-MS revealed that 93 metabolites were upregulated and 178 metabolites were downregulated compared with the baseline. KEGG enrichment analysis showed that amino acid biosynthesis was significantly activated on day 14 (P < 0.05), while steroid hormone and fatty acid biosynthesis dominated on day 28. Olink analysis revealed significantly increased expression of IL-2 and CCL23 on day 28 (P < 0.05). The closed environment can induce structural adjustment of the microbial community (enrichment of Firmicutes and reduction of Bacteroides), which can lead to dynamic reprogramming of serum metabolites and gradual activation of inflammation. These findings suggest the temporal adaptability of microbial metabolic functions and host inflammatory response. • .Phased Metabolic Adaptation: Distinct metabolic shifts occur over time: Early phase (14 days) activates amino acid biosynthesis for energy/microbiota needs, while the Late phase (28 days) shifts to steroid hormone and fatty acid synthesis for chronic stress adaptation. • Novel Biomarker Discovery: Identified two crucial biomarkers: Ergothioneine (EGT) shows a sustained increase (unlike spaceflight), suggesting dietary intervention value, and Alloprevotella exhibits a significant, first-reported decline, serving as an early-warning indicator for health risks due to its role in producing gut-protective SCFAs. • .Microbiota Dynamics Trump Diversity: "Key microbiota regulation" is a more sensitive health indicator than overall diversity. Critical taxa like Alloprevotella and Clostridium show significant fluctuations despite stable diversity metrics. • .Targeted Health Interventions: Proposes developing precision monitoring systems integrating EGT, Alloprevotella, and steroid pathways, and implementing targeted interventions like EGT-rich diets/prebiotics for antioxidants and Alloprevotella probiotics to restore gut-barrier function.