Isolation and Immunocharacterization of Lactobacillus salivarius from the Intestine of Wakame-Fed Pigs to Develop Novel “Immunosynbiotics”

Abstract

Emerging threats of antimicrobial resistance necessitate the exploration of effective alternatives for healthy livestock growth strategies. ‘Immunosynbiotics’, a combination of immunoregulatory probiotics and prebiotics with synergistic effects when used together in feed, would be one of the most promising candidates. Lactobacilli are normal residents of the gastrointestinal tract of pigs, and many of them are able to exert beneficial immunoregulatory properties. On the other hand, wakame (Undaria pinnafida), an edible seaweed, has the potential to be used as an immunoregulatory prebiotic when added to livestock feed. Therefore, in order to develop a novel immunosynbiotic, we isolated and characterized immunoregulatory lactobacilli with the ability to utilize wakame. Following a month-long in vivo wakame feeding trial in 8-week-old Landrace pigs (n = 6), sections of intestinal mucous membrane were processed for bacteriological culture and followed by identification of pure colonies by 16S rRNA sequence. Each isolate was characterized in vitro in terms of their ability to assimilate to the wakame and to differentially modulate the expression of interleukin-6 (IL-6) and interferon beta (IFN-β) in the porcine intestinal epithelial (PIE) cells triggered by Toll-like receptor (TLR)-4 and TLR-3 activation, respectively. We demonstrated that feeding wakame to pigs significantly increased the lactobacilli population in the small intestine. We established a wakame-component adjusted culture media that allowed the isolation and characterization of a total of 128 Lactobacilli salivarius colonies from the gut of wakame-fed pigs. Interestingly, several L. salivarius isolates showed both high wakame assimilation ability and immunomodulatory capacities. Among the wakame assimilating isolates, L. salivarius FFIG71 showed a significantly higher capacity to upregulate the IL-6 expression, and L. salivarius FFIG131 showed significantly higher capacity to upregulate the IFN-β expression; these could be used as immunobiotic strains in combination with wakame for the development of novel immunologically active feeds for pigs.

Efficient Selection of New Immunobiotic Strains With Antiviral Effects in Local and Distal Mucosal Sites by Using Porcine Intestinal Epitheliocytes

“…Of note, the biotechnological properties of the two strains are different. Whereas L. rhamnosus CRL1505 has been used mainly in the development of dairy functional products (2,44), the MPL16 strain has shown a remarkable ability to growth and ferment wakame (Undaria pinnatifida) that is the most popular and economically important edible brown algae in Asian countries (24,45). Then, the different biotechnological properties of L. plantarum MPL16 could potentiate the development of non-dairy functional foods or feeds with the ability to improve antiviral immunity in the intestine and the respiratory tract.…”

“…From that study, we were able to select a set of potential biomarkers that would allow us to efficiently select new immunobiotic strains with antiviral capabilities including IFNα, IFNβ, RIG1, TLR3, OAS1, RNASEL, MX2, A20, CXCL5, CCL4, IL-15, SELL, SELE, EPCAM, PTGS2, PLA2G4A, PTEGES, and PTGER4. Then, in order to validate this assumption, PIE cells were stimulated with different lactobacilli including immunomodulatory (L. rhamnosus CRL1505, L. plantarum CRL1506, L. rhamnosus IBL07, and L. plantarum MPL16) (8,16,24,25) and non-immunomodulatory (L. rhamnosus CRL576 and L. plantarum CRL681) strains and then challenged with poly(I:C). The expression of the biomarkers was then evaluated.…”

Draft Genome Sequence of Ligilactobacillus salivarius FFIG58, Isolated from the Intestinal Tract of Wakame-Fed Pig

“… Ligilactobacillus salivarius (basonym: Lactobacillus salivarius ) ( 1 ) is considered an important member of the swine intestinal microbiota ( 2 , 3 ) because of its capacity to protect against intestinal infections through inhibition of the growth of pathogens, competition for mucosal binding sites, or modulation of the immune system ( 4 – 7 ). Our previous work demonstrated that feeding pigs wakame ( Undaria pinnatifida ), economically important edible algae in Asian countries ( 8 ), was able to modify the gastrointestinal microbiota, inducing a significant increase in the abundance of lactobacilli ( 9 ). A library of L. salivarius strains isolated from the intestine of pigs fed wakame was constructed ( 9 ), and we investigated the capacities of those strains to modulate the innate antiviral immune response in porcine intestinal epithelial (PIE) cells.…”

“…Our previous work demonstrated that feeding pigs wakame ( Undaria pinnatifida ), economically important edible algae in Asian countries ( 8 ), was able to modify the gastrointestinal microbiota, inducing a significant increase in the abundance of lactobacilli ( 9 ). A library of L. salivarius strains isolated from the intestine of pigs fed wakame was constructed ( 9 ), and we investigated the capacities of those strains to modulate the innate antiviral immune response in porcine intestinal epithelial (PIE) cells. Among the strains evaluated, L. salivarius FFIG58 had a remarkable capacity to improve interferon beta levels in PIE cells after Toll-like receptor 3 activation and to protect against rotavirus infection ( 9 ).…”

“…A library of L. salivarius strains isolated from the intestine of pigs fed wakame was constructed ( 9 ), and we investigated the capacities of those strains to modulate the innate antiviral immune response in porcine intestinal epithelial (PIE) cells. Among the strains evaluated, L. salivarius FFIG58 had a remarkable capacity to improve interferon beta levels in PIE cells after Toll-like receptor 3 activation and to protect against rotavirus infection ( 9 ).…”