Metabolic profiling and growth characteristics of a spaceflight-induced mutant of Lacticaseibacillus rhamnosus: Unveiling enhanced carbohydrate and amino acid metabolism for improved probiotic potential

Introduction: Lacticaseibacillus (L.) rhamnosus Probio-M9 (Probio-M9) is a probiotic strain known for its positive effects on the intestinal microbial community, host immunity, and the well-being of stressed adults. Lacticaseibacillus rhamnosus R7970 (R7970) is a mutant cell line of Probio-M9 obtained through a spaceflight mutagenesis experiment. Objective: This study aimed to investigate the molecular-level differences in growth characteristics and metabolism between R7970 and Probio-M9. Methods: Growth behavior comparisons included colony morphology, viable cell count, pH, and titratable acidity in the growth medium. Transcriptomics analysis was performed on bacterial cell lysates, while non-targeted metabolomics analysis was conducted on both the lysate and cell culture supernatant samples. Targeted metabolomics analysis validated the quantities of selected differential metabolites. Results: R7970 exhibited a stronger acid-producing ability, higher viable cell counts, and larger colony sizes compared to Probio-M9. Integrated metabolomics and transcriptomics analysis revealed more active carbohydrate and amino acid metabolism in R7970 than in Probio-M8. Quantitative analysis confirmed increased levels of lactic acid, citric acid, free amino acids, and tryptophan metabolites in R7970 compared to Probio-M9. Additionally, R7970 exhibited higher levels of beneficial metabolites, including alpha-ketoglutarate, indole-3acetamide, indole-3-lactic acid, nicotinic acid, and picolinic acid. Conclusions: This study provides valuable insights into the molecular mechanisms underlying the distinct growth behavior of the R7970 mutant line, particularly highlighting its differential carbohydrate and amino acid metabolism. The enhanced probiotic properties of R7970 make it a valuable addition to existing probiotic resources, with promising potential for the development of novel functional products.