Exopolysaccharide (EPS)-producing Lacticaseibacillus paracasei EPS DA-BACS was isolated from healthy human feces and its probiotic properties, as well as the structure and prebiotic activity of the EPS from this strain were examined. EPS from L. paracasei EPS DA-BACS had a ropy phenotype, which is known to have potential health benefits and is identified as loosely cell-bounded glucomannan-type EPS with a molecular size of 3.7 × 10⁶ Da. EPS promoted the acid tolerance of L. paracasei EPS DA-BACS and provided cells with tolerance to gastrointestinal stress. The purified EPS showed growth inhibitory activity against Clostridium difficile. L. paracasei EPS DA-BACS cells completely inhibited the growth of Bacillus subtilis, Pseudomonas aeruginosa, and Aspergillus brasiliensis, as well as showed high growth inhibitory activity against Staphylococcus aureus and Escherichia coli. Treatment of lipopolysaccharide-stimulated RAW 264.7 cells with heat-killed L. paracasei EPS DA-BACS cells led to a decrease in the production of nitric oxide, indicating the anti-inflammatory activity of L. paracasei EPS DA-BACS. Purified EPS promoted the growth of Lactobacillus gasseri, Bifidobacterium bifidum, B. animalis, and B. faecale which showed high prebiotic activity. L. paracasei EPS DA-BACS harbors no antibiotic resistance genes or virulence factors. Therefore, L. paracasei EPS DA-BACS exhibits anti-inflammatory and antimicrobial activities with high gut adhesion ability and gastrointestinal tolerance and can be used as a potential probiotic.

This study explored the potential of Lactococcus lactis subsp. lactis CAB701 as a probiotic strain, focusing on its immunostimulatory properties. Despite adverse conditions in the gastrointestinal environment, this strain exhibited remarkable survivability, as evidenced by its tolerance to acid, bile, and pancreatin, coupled with its impressive ability to adhere to Caco-2 cells. It also exhibited significant antioxidant activity, similar to the established probiotic Lacticaseibacillus rhamnosus GG (LGG). Our research elucidates the potent immunostimulatory effects of L. lactis subsp. lactis CAB701. This strain significantly enhanced nitric oxide production in RAW 264.7, far exceeding that obtained with LGG. An in-depth examination revealed elevated expression of key inflammatory mediators, including inducible nitric oxide synthase, tumor necrosis factor-alpha, cyclooxygenase-2, interleukin (IL)-1 beta, and IL-6. L. lactis subsp. lactis CAB701 increases the expression of critical signaling proteins in the mitogen-activated protein kinase pathway. This prompted a substantial increase in the expression of phosphorylated c-Jun N-terminal kinases and extracellular signal-regulated kinases, suggesting their role in modulating these immune-related pathways. Overall, these findings demonstrate the significant immunostimulatory capacity of L. lactis subsp. lactis CAB701, positioning it as a potential candidate for probiotic use, especially in applications that enhance immune responses.

Synbiotics contain health-beneficial bacteria, i.e., probiotics and prebiotics selectively utilized by the probiotics. Herein, three probiotic strains, Leuconostoc lactis CCK940, L. lactis SBC001, and Weissella cibaria YRK005, and the oligosaccharides produced by these strains (CCK, SBC, and YRK, respectively) were used to prepare nine synbiotic combinations. Macrophages (RAW 264.7) were treated with these synbiotic combinations and the corresponding lactic acid bacteria and oligosaccharides alone to evaluate the treatments’ immunostimulatory activities. The level of nitric oxide (NO) production was significantly higher in the macrophages treated with the synbiotics than in those treated with the corresponding probiotic strains and the oligosaccharide alone. The immunostimulatory activities of the synbiotics increased regardless of the probiotic strain and the type of oligosaccharide used. The expressions of tissue necrosis factor-α, interleukin-1β, cyclooxygenase-2, inducible NO synthase genes, and extracellular-signal-regulated and c-Jun N-terminal kinases were significantly higher in the macrophages treated with the three synbiotics than in those treated with the corresponding strains or with the oligosaccharides alone. These results indicate that the synergistic immunostimulatory activities of probiotics and the prebiotics they produced in the studied synbiotic preparations resulted from the activation of the mitogen-activated protein-kinase-signaling pathway. This study suggests the combined use of these probiotics and prebiotics in the development of synbiotic preparations as health supplements.