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Optimization of Media Composition to Maximize the Yield of Exopolysaccharides Production by Lactobacillus rhamnosus Strains.

ABSTRACT: Growth media composition is a critical factor influencing the yield of bacterial exopolysaccharides (EPSs), which have attracted the interest of researchers around the world due to their diverse physicochemical and biological properties. This work presents the optimization of media for EPS synthesis by three Lactobacillus rhamnosus strains, namely ?OCK 0943, ?OCK 0935, and OM-1. The optimized media led to a more than 13-fold increase in EPS yield for L. rhamnosus ?OCK 0943 (from 85 to 1138.2 mg/L), an almost 9-fold increase for L. rhamnosus ?OCK 0935 (from 103.67 to 900 mg/L), and a more than 7-fold increase for L. rhamnosus OM-1 (from 133.67 to 987.84 mg/L) as compared to cultures in standard MRS medium (de Man, Rogosa, and Sharpe). It has been found that the main medium-related determinant of EPS synthesis by the studied L. rhamnosus strains are the carbon source-in this case, it was fructose and sucrose.

SUBMITTER: Oleksy-Sobczak M

PROVIDER: S-EPMC7306023 | biostudies-literature | 2020 Jun

REPOSITORIES: biostudies-literature

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Optimization of Media Composition to Maximize the Yield of Exopolysaccharides Production by Lactobacillus rhamnosus Strains.

Oleksy-Sobczak Magdalena M Klewicka Elżbieta E

Probiotics and antimicrobial proteins 20200601 2

Growth media composition is a critical factor influencing the yield of bacterial exopolysaccharides (EPSs), which have attracted the interest of researchers around the world due to their diverse physicochemical and biological properties. This work presents the optimization of media for EPS synthesis by three Lactobacillus rhamnosus strains, namely ŁOCK 0943, ŁOCK 0935, and OM-1. The optimized media led to a more than 13-fold increase in EPS yield for L. rhamnosus ŁOCK 0943 (from 85 to 1138.2 mg/ ...[more]

PMID: 31410767

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Project description:The traditional media used for the fermentation of Lactobacilli always contain carbohydrate polymers, which interfere with the analysis of the exopolysaccharide (EPS) produced by the bacteria. In this investigation, a novel medium formulation that could avoid such interference was successfully developed. The beef extract, yeast extract, and peptone used in this formulation were subjected to the removal of polysaccharides before use. The factors affecting the EPS production were optimized by a single factor test, Plackett–Burman design, and Box–Behnken design. The optimum formula was ascertained as: 7.5 g L–1 yeast extract, 12.5 g L–1 beef extract, 10 g L–1 peptone, 21.23 g L–1 maltose, 5.51 g L–1 yeast nitrogen base, 2 g L–1 K2HPO4, 5 g L–1 anhydrous sodium acetate, 2 g L–1 ammonium citrate, 0.58 g L–1 MgSO4⋅7H2O, 0.25 g L–1 MnSO4⋅H2O, and 1 mL L–1 Tween 80. The initial pH of the medium was 6.5. The optimized conditions for fermentation of the strain to produce EPS were as follows: seed size 1%, culture temperature 37°C, and culture time 20 h. Optimum results showed that EPS yield was 496.64 ± 3.15 mg L–1, being 76.70% higher than that of unoptimized conditions (281.07 ± 5.90 mg L–1). The EPS was mainly comprised of glucose and guluronic acid, with a weight average molecular weight of 19.9 kDa; it was also characterized by Fourier transform infrared spectroscopy and UV analysis. EPS was found to significantly enhance the phagocytic capacity, promote the NO, TNF-α, IL-1β, and IL-6 secretion, and improve mRNA expression of cytokines in RAW 264.7 macrophages, indicating its considerable immunomodulatory activity. Western bolt and immunofluorescence results demonstrated that the EPS was able to increase p65 nuclear translocation in the macrophages, indicating that EPS enhanced immunity via the NF-κB signaling pathway. EPS investigated in this work has potential as an attractive functional food supplement candidate for the hypoimmunity population.

Project description:BackgroundSalmonella typhimurium (S.T), as an important foodborne bacterial pathogen, can cause diarrhea and gastroenteritis in humans and animals. Numerous studies have confirmed that exopolysaccharides (EPSs) have various biological functions, but the mechanism through which EPSs improve the immunity of animals against the invasion of pathogenic bacteria is unclear. Here, we explored the protective effect of EPSs of Lactobacillus rhamnosus GG (LGG) on the S.T-infected intestine.MethodsMice received adequate food and drinking water for one week before the start of the experiment. After 7 d of prefeeding, 2×108 CFU/mL S.T solution and an equivalent volume of saline (control group) were given orally for 1 d. On the fourth day, the mice were treated with 0.5 mg/mL EPSs, 1.0 mg/mL EPSs, 2.0 mg/mL EPSs, or 2.0 mg/mL penicillin for 7 d. Finally, the body and relative organ weight, histological staining, and the levels of antioxidant enzyme activity and inflammatory cytokines were determined.ResultsThe S.T-infected mice exhibited symptoms of decreased appetite, somnolence, diarrhea and flagging spirit. Treatment with EPSs and penicillin improved the weight loss of the mice, and the high dose of EPSs showed the best therapeutic effect. EPSs significantly ameliorated S.T-induced ileal injury in mice. High-dose EPSs were more effective than penicillin for alleviating ileal oxidative damage induced by S.T. The mRNA levels of inflammatory cytokines in the ileum of mice showed that the regulatory effects of EPSs on inflammatory cytokines were better than those of penicillin. EPSs could inhibit the expression and activation of key proteins of the TLR4/NF-κB/MAPK pathway and thereby suppress the level of S.T-induced ileal inflammation.ConclusionsEPSs attenuate S.T-induced immune responses by inhibiting the expression of key proteins in the TLR4/NF-κB/MAPK signaling pathway. Moreover, EPSs could promote bacterial aggregation into clusters, which may be a potential strategy for reducing the bacterial invasion of intestinal epithelial cells.

Project description:Lactobacillus rhamnosus GG is one of the most widely marketed and studied probiotic strains. In L. rhamnosus GG, the spaCBA-srtC1 gene cluster encodes pili, which are important for some of the probiotic properties of the strain. A previous study showed that the DNA sequence of the spaCBA-srtC1 gene cluster was not present in some L. rhamnosus GG variants isolated from liquid dairy products. To examine the stability of the L. rhamnosus GG genome in an industrial production process, we sequenced the genome of samples of L. rhamnosus GG (DSM 33156) collected at specific steps of the industrial production process, including the culture collection stock, intermediate fermentations, and final freeze-dried products. We found that the L. rhamnosus GG genome sequence was unchanged throughout the production process. Consequently, the spaCBA-srtC1 gene locus was intact and fully conserved in all 31 samples examined. In addition, different production batches of L. rhamnosus GG exhibited consistent phenotypes, including the presence of pili in final freeze-dried products, and consistent characteristics in in vitro assays of probiotic properties. Our data show that L. rhamnosus GG is highly stable in this industrial production process.IMPORTANCELactobacillus rhamnosus GG is one of the best-studied probiotic strains. One of the well-characterized features of the strain is the pili encoded by the spaCBA-srtC1 gene cluster. These pili are involved in persistence in the gastrointestinal tract and are important for the probiotic properties of L. rhamnosus GG. Previous studies demonstrated that the L. rhamnosus GG genome can be unstable under certain conditions and can lose the spaCBA-srtC1 gene cluster. Since in vitro studies have shown that the loss of the spaCBA-srtC1 gene cluster decreases certain L. rhamnosus GG probiotic properties, we assessed both the genomic stability and phenotypic properties of L. rhamnosus GG throughout an industrial production process. We found that neither genomic nor phenotypic changes occurred in the samples. Therefore, we demonstrate that L. rhamnosus GG retains the spaCBA-srtC1 cluster and exhibits excellent genomic and phenotypic stability in the specific industrial process examined here.

Dataset Information

Optimization of Media Composition to Maximize the Yield of Exopolysaccharides Production by Lactobacillus rhamnosus Strains.

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Optimization of Media Composition to Maximize the Yield of Exopolysaccharides Production by Lactobacillus rhamnosus Strains.

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