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Abstract

This study presents the most comprehensive spatiotemporal analysis of the ISS microbiome to date. Over a seven-year period, 184 surface samples were collected for live/dead metagenomic profiling, phenotypic characterization of antimicrobial resistance (AMR) and virulence among 102 cultured isolates, and metagenome-assembled genome (MAG) analysis to evaluate evolutionary selection pressures. Despite ecological stability, comparative genomics revealed ongoing microevolution through lateral gene transfer and selection for traits (e.g., radiation resistance and biocide tolerance). Importantly, predictions of AMR and virulence frequently misaligned with experimental outcomes, underscoring the need for functional validation. This dataset highlights a stable core microbiome that persists across years, punctuated by localized adaptation and gene flow. The ISS microbiome exemplifies both ecological resilience and microevolutionary innovation that can inform risk management for long-duration spaceflight.

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72% semantic matchSystems Biology