AI-Based Comparative Transcriptomics and Gene Network Profiling of Staphylococcus aureus in Astronaut-Associated Missions

Introduction: Spaceflight-associated microgravity alters microbial physiology, raising concerns about the adaptability and pathogenicity of opportunistic bacteria such as Staphylococcus aureus. Understanding transcriptomic responses in this context is essential for astronaut health and mission safety. Methods: Publicly available transcriptomic datasets were retrieved from the NASA GeneLab repository and analyzed in two stages. First, three experimental scenarios were compared: (1) BRIC-23 mission (in vitro Petri dish cultures aboard the ISS; 9 space vs. 9 ground), (2) SpaceX Inspiration4 mission microbiota (40 samples from 10 body sites of 4 astronauts across flight phases), and (3) Dragon capsule surface cultures (30 samples across 10 capsule zones and 3 time points). Differential expression (DESeq2), functional enrichment, heatmaps, co-expression network analysis (WGCNA), and bootstrapping were applied to identify conserved transcriptomic signatures. In the second stage, five candidate genes were selected from 45 consistently altered genes across all conditions. These were validated through statistical significance (BRIC-23, p < 0.05, significant log2 fold change) and consistent presence within co-expression modules. Candidate genes were further integrated with literature-curated virulence and biofilm-associated genes to construct a functional metabolic network using pathway analysis and K-means clustering. Results: Across independent datasets, S. aureus displayed convergence in transcriptomic profiles, particularly involving genes linked to virulence, adhesion, biofilm formation, and metabolic adaptation. Network-level integration revealed metabolic reprogramming and transcriptional plasticity as conserved responses, suggesting tightly regulated adaptation rather than random changes. Discussion: The consistent identification of virulence and biofilm-associated genes across heterogeneous datasets indicates that microgravity imposes selective pressure favoring traits that enhance persistence in extreme environments. These findings support the hypothesis that S. aureus utilizes adaptive regulatory circuits to balance growth, survival, and pathogenic potential in the spaceflight niche. Conclusions: Our integrative bioinformatics approach reveals conserved adaptive strategies in S. aureus under microgravity, characterized by transcriptomic convergence and metabolic reprogramming. These insights underscore the necessity of experimental validation and phenotypic assays to assess microbial risks for astronaut health and to design countermeasures for future long-duration missions.