Transcriptional profiling of probiotic Escherichia coli Nissle 1917 under multiple environmental stresses: Simulated microgravity, microaerobic, and co-culture conditions

Environmental factors can substantially alter the physiology of probiotics, yet their transcriptomic responses under multiple simultaneous stresses remain poorly understood. Among these factors, a previous study has shown that the low-shear environment induced by simulated microgravity elicits mild stress in the well-known probiotic Escherichia coli Nissle 1917 (EcN). In this study, we investigated the impact of additional environmental stressors on the transcriptomic profile of EcN by introducing microaerobic conditions and co-culture with a pathogenic Pseudomonas aeruginosa PAO1 (PAO1), in combination with simulated microgravity. Under the combined condition of simulated microgravity and microaerobic stress, EcN maintained stable growth while upregulating genes involved in anaerobic respiration and biofilm formation, indicating effective adaptation to oxygen-limited environments. However, when co-cultured with PAO1 under the same conditions, EcN exhibited pronounced growth inhibition. Transcriptomic analysis revealed downregulation of genes involved in energy metabolism and biofilm formation, while genes related to metal ion uptake remained active. Further comparison between simulated microgravity and normal gravity under these compound stress conditions revealed that microgravity further exacerbated growth suppression in the context of overlapping environmental stresses, particularly via altered amino acid metabolism. These findings suggest that EcN's stress responses and viability are impaired in complex environmental conditions, emphasizing the importance of developing more resilient probiotic strains for therapeutic applications in extreme environments such as spaceflight.