Intestinal release of biofilm-like microcolonies encased in calcium-pectinate beads increases probiotic properties of Lacticaseibacillus paracasei

Abstract

In this study, we show that calcium pectinate beads (CPB) allow the formation of 20 µm spherical microcolonies of the probiotic bacteria Lacticaseibacillus paracasei (formerly designated as Lactobacillus paracasei) ATCC334 with a high cell density, reaching more than 10 log (CFU/g). The bacteria within these microcolonies are well structured and adhere to a three-dimensional network made of calcium-pectinate through the synthesis of extracellular polymeric substances (EPS) and thus display a biofilm-like phenotype, an attractive property for their use as probiotics. During bacterial development in the CPB, a coalescence phenomenon arises between neighboring microcolonies accompanied by their peripheral spatialization within the bead. Moreover, the cells of L. paracasei ATCC334 encased in these pectinate beads exhibit increased resistance to acidic stress (pH 1.5), osmotic stress (4.5 M NaCl), the freeze-drying process and combined stresses, simulating the harsh conditions encountered in the gastrointestinal (GI) tract. In vivo, the oral administration of CPB-formulated L. paracasei ATCC334 in mice demonstrated that biofilm-like microcolonies are successfully released from the CPB matrix in the colonic environment. In addition, these CPB-formulated probiotic bacteria display the ability to reduce the severity of a DSS-induced colitis mouse model, with a decrease in colonic mucosal injuries, less inflammation, and reduced weight loss compared to DSS control mice. To conclude, this work paves the way for a new form of probiotic administration in the form of biofilm-like microcolonies with enhanced functionalities.

Encapsulation of Lactobacillus in Low-Methoxyl Pectin-Based Microcapsules Stimulates Biofilm Formation: Enhanced Resistances to Heat Shock and Simulated Gastrointestinal Digestion

“…As a result, the cell distribution after biofilm formation became more concentrated on the surface and periphery of the MCs. 30 This is also in line with the result of SEM observation on the surface of BMCs in our study.…”

“…The cell release behavior of pectinate beads could likely be ascribed to combined actions of swelling of beads under intestinal environment and enzymatic degradation of beads in the colon. 30 In summary, our study highlights the application of two lowmethoxyl CMPs as ideal coating materials for bacterial encapsulation. Lactobacilli encapsulated in pectin-based MCs showed similar fermentation ability but improved bacterial resistances to the heat shock and simulated gastrointestinal digestion in comparison with the planktonic lactobacilli.…”

“…A few similar studies that worked with alginate-based or pectinbased encapsulation for probiotic Lactobacillus also demonstrated that biofilm-formation encapsulation can provide excellent protection for the encapsulated bacteria. 12,22,30 In this study, although only single-layer pectin-based encapsulations were applied, they were proven to be a feasible and universal way to provide exceptional protections for three Lactobacillus strains. Except for the protective effects, this technology can also work efficiently to develop a high-density probiotic carrier applied as biocatalysts or functional biomaterials.…”

Pili and other surface proteins influence the structure and the nanomechanical properties of Lactococcus lactis biofilms

“…Altogether, this work showing that pili and surface proteins can shape the biofilms of L. lactis may open new doors in probiotic and biotechnological applications in which this bacteria is used. In probiotic issues, for example, it has been shown in lactobacilli that the biofilm mode of life boosts the biological activity mediated by lactobacilli 59 , 60 . Also, it can be hypothesized that more stable biofilms of L. lactis might increase the persistence and retention time of the bacteria in the gastrointestinal tract and thus increase the beneficial effects mediated by this bacteria.…”

Bacterial Materials: Applications of Natural and Modified Biofilms

“…[ 103 ] Similarly, biofilm loaded calcium pectinate beads produced by Heumann et al. [ 104 ] lead to sturdier probiotics; from those biofilm‐spheres, the bacteria were released to the colon as clusters which provided a better anti‐inflammatory effect and protection against GIT disorders than other probiotic forms of this bacterial strain. A better release of biofilm bacteria was also achieved by Vega‐Sagardía et al., [ 105 ] who enriched their formulation with vegetal oil to increase the residence time of the probiotic biofilm in the stomach so that a Helicobacter pylori infection could be efficiently dealt with.…”