The Ability of Riboflavin-Overproducing Lactiplantibacillus plantarum Strains to Survive Under Gastrointestinal Conditions

Abstract

Riboflavin, vitamin B2, is essential for humans and has to be obtained from the diet. Some lactic acid bacteria (LAB) produce this vitamin, and they can be used for in-situ fortification of foods. This could be an alternative to supplementation with chemically synthesized vitamin, to palliate riboflavin deficiencies in specific groups of people. Moreover, if the producing LAB could survive in the gastrointestinal stress (GIT) they could be added as probiotics in this environment. In the present study we tested two riboflavin-overproducing Lactiplantibacillus plantarum strains (M5MA1-B2 and M9MG6-B2), spontaneous mutants of LAB isolated from chicha, a traditional Andean beverage. These two LAB, and also their isogenic strains M5MA1-B2[pRCR12] and M9MG6-B2[pRCR12], expressing the mCherry protein from the pRCR12 plasmid, were evaluated in vitro under simulated GIT conditions. Among other, specifically developed protein fluorescence assays were used. The four LAB showed similar levels of adhesion (>6.0%) to Caco-2 cells, higher than that of the probiotic Lacticaseibacillus rhamnosus GG strain (4.51%). Thus, LAB biofilm formation was assessed in the labeled cells by intracellular mCherry fluorescence and in the unlabeled parental strains by crystal violet staining. Both methods detected the formation of consistent biofilms by the L. plantarum strains. The quantification of mCherry fluorescence was also used to analyze LAB auto-aggregation properties. High levels of auto-aggregation were detected for both M5MA1-B2[pRCR12] and M9MG6-B2[pRCR12]. Survival of LAB included in a commercial cereal-based food matrix (Incaparina) under GIT conditions was also evaluated. The four LAB were resistant in vitro to the stomach and intestinal stresses, and proliferated in this environment, indicating a protective and nutritional effect of the Incaparina on the bacteria. Also, M9MG6-B2 survival in the presence or absence of Incaparina was evaluated in vivo in a BALB/c mouse model. The administration of the M9MG6-B2 strain alone or together with Incaparina had no Hernández-Alcántara et al. Riboflavin-Overproducing Lactiplantibacillus Plantarum Gastrointestinal Survival adverse effect on the health, growth and/or well-being of the rodents. In addition, an increment in the villus length/crypt depth ratio was observed. The overall results obtained indicate that the LAB studied have probiotic characteristics of interest for the development of functional foods.

Spontaneous Riboflavin-Overproducing Limosilactobacillus reuteri for Biofortification of Fermented Foods

“…Besides riboflavin production and fermentation capacity, it is important to consider the gastrointestinal adaptation properties of lactobacilli to efficiently implement them for real-life food biofortification or as beneficial supplements ( 16 , 21 ). Here, we showed that L. reuteri AMBV339 survives in simulated gastric juice, suggesting that it is resistant to the harsh conditions of the stomach before, during and after a meal, and can be transported live to the small intestine where riboflavin is absorbed, as we demonstrated via passive transport in the gastrointestinal dialysis model (GIDM-colon) and active transport via human intestinal epithelial cells.…”

“…Moreover, the acidification capacity of these bacteria can inhibit pathogenic growth, typically resulting in longer shelf life ( 20 ). In addition to the ability of the lactobacilli to produce riboflavin in a range of food matrices, it can be beneficial if the selected strains survive the gastrointestinal tract, so that the vitamins can be produced in situ at the target site to further enhance delivery and maximize the chance of reaching required daily intake levels ( 16 , 21 ).…”

Fungal–Lactobacteria Consortia and Enzymatic Catalysis for Polylactic Acid Production

“…Firstly, bacterium–fungus combinations were tested in liquid medium: GPP (Glucose-Proline-Phosphate) minimal medium [ 21 ], Czapek’s Dox [ 29 ], CDM (chemical defined medium) [ 30 ] and plantarum minimal medium (PMM5) [ 27 ], all with glucose as a carbon source. In subsequent experiments, we used MRS broth (Condalab) with glucose as a control medium and MRS broth without dextrose and beef extract (Condalab) with 1% ( w / v ) starch as the carbon source.…”