Microbial dynamics across commercial spaceflights of varying duration

Spaceflight introduces environmental stressors that can alter human microbiomes and immune responses. We analyzed 259 and 17 biospecimens for microbiome and single-cell studies from n = 6 astronauts across two commercial International Space Station (ISS) missions: Axiom 2 (10-day mission) and Axiom 3 (21-day mission). Samples included saliva, stool, urine, and body swabs from 10 anatomical sites, profiled via 16S ribosomal RNA (rRNA) gene amplicon sequencing. Gut and oral microbiomes remained stable, while skin-associated communities exhibited transient diversity shifts post-flight. Taxonomic analysis revealed individual and site-specific patterns, as well as from the spaceflight/ISS environment. Cytokine profiling from single-cell data indicated immune activation, with IL-32 and IL-16 transcripts elevated in Axiom 2 and Axiom 3. Microbiome and interleukin correlation analyses found 14 significant associations in Axiom 2 data. These findings provide an integrated view of microbiome individuality. Our study represents a descriptive comparison of two missions, informing strategies for crew health on future long-duration missions.