Adaptive laboratory evolution of Lactiplantibacillus plantarum for enhanced lactose metabolism: Phenotypic changes and underlying molecular mechanisms.

Plant-derived Lactiplantibacillus plantarum possesses strong probiotic potential; however, its limited lactose-metabolizing capacity severely restricts its application in dairy fermentation. In this study, we used an L. plantarum strain isolated from traditionally fermented pickles in Inner Mongolia as the parental strain. The strain underwent 600 generations of adaptive laboratory evolution under stepwise increases in lactose concentration. Phenotypic changes in the evolved strain were systematically characterized, and the underlying molecular basis was investigated through whole-genome sequencing and transcriptomic analyses. The evolved strain exhibited significantly improved growth, acidification, and lactose utilization efficiencies, and the adaptive phenotype was stably inherited. In addition, the activities of key enzymes involved in lactose catabolism, including β-galactosidase, galactokinase, glucokinase, and 6-phosphofructokinase, were markedly increased. Stable nonsynonymous single-nucleotide polymorphisms (SNPs) were identified in the core lactose metabolism genes, lacL and lacS. Transcriptomic analysis further revealed global transcriptional reprogramming, characterized by significant upregulation of genes associated with lactose transport and catabolism, glycolysis, and energy metabolism. Collectively, this study provides a safe and practical strategy for developing dairy-adapted probiotic strains and clarifies the molecular basis of lactose-directed adaptive evolution in L. plantarum.