Review: Adaptation of Beneficial Propionibacteria, Lactobacilli, and Bifidobacteria Improves Tolerance Toward Technological and Digestive Stresses

This review deals with beneficial bacteria, with a focus on lactobacilli, propionibacteria, and bifidobacteria. As being recognized as beneficial bacteria, they are consumed as probiotics in various food products. Some may also be used as starters in food fermentation. In either case, these bacteria may be exposed to various environmental stresses during industrial production steps, including drying and storage, and during the digestion process. In accordance with their adaptation to harsh environmental conditions, they possess adaptation mechanisms, which can be induced by pretreatments. Adaptive mechanisms include accumulation of compatible solutes and of energy storage compounds, which can be largely modulated by the culture conditions. They also include the regulation of energy production pathways, as well as the modulation of the cell envelop, i.e., membrane, cell wall, surface layers, and exopolysaccharides. They finally lead to the overexpression of molecular chaperones and of stress-responsive proteases. Triggering these adaptive mechanisms can improve the resistance of beneficial bacteria toward technological and digestive stresses. This opens new perspectives for the improvement of industrial processes efficiency with regard to the survival of beneficial bacteria. However, this bibliographical survey evidenced that adaptive responses are strain-dependent, so that growth and adaptation should be optimized case-by-case

Growth in Hyper-Concentrated Sweet Whey Triggers Multi Stress Tolerance and Spray Drying Survival in Lactobacillus casei BL23: From the Molecular Basis to New Perspectives for Sustainable Probiotic Production

Lactobacillus casei BL23 has a recognized probiotic potential, which includes immune modulation, protection toward induced colitis, toward induced colon cancer and toward dissemination of pathogens. In L. casei, as well as in other probiotics, both probiotic and technological abilities are highly dependent (1) on the substrate used to grow bacteria and (2) on the process used to dry and store this biomass. Production and storage of probiotics, at a reasonable financial and environmental cost, becomes a crucial challenge. Food-grade media must be used, and minimal process is preferred. In this context, we have developed a “2-in-1” medium used both to grow and to dry L. casei BL23, considered a fragile probiotic strain. This medium consists in hyper-concentrated sweet whey (HCSW). L. casei BL23 grows in HCSW up to 30% dry matter, which is 6 times-concentrated sweet whey. Compared to isotonic sweet whey (5% dry matter), these growth conditions enhanced tolerance of L. casei BL23 toward heat, acid and bile salts stress. HCSW also triggered intracellular accumulation of polyphosphate, of glycogen and of trehalose. A gel-free global proteomic differential analysis further evidenced overexpression of proteins involved in pathways known to participate in stress adaptation, including environmental signal transduction, oxidative and metal defense, DNA repair, protein turnover and repair, carbohydrate, phosphate and amino acid metabolism, and in osmoadaptation. Accordingly, HCSW cultures of L. casei BL23 exhibited enhanced survival upon spray drying, a process known to drastically affect bacterial viability. This work opens new perspectives for sustainable production of dried probiotic lactobacilli, using food industry by-products and lowering energy costs

Collaboration entre probiotiques lactiques et propioniques : nouveau procédé de production durable

il s'agit d'un type de produit dont les métadonnées ne correspondent pas aux métadonnées attendues dans les autres types de produit : DISSERTATIONDairy propionibacterium are present in many fermented products. Propionibacterium freudenreichii is considered as a GRAS bacteria (Generally Recognize As Safe), and is used as probiotic and as cheese starters. The strain P. freudenreichii CIRM-BIA 129 has surface layers proteins with anti-inflammatory immunomodulatory properties. Production and stabilization of dairy bacteria can be optimized. In addition, during the production of lactic bacteria, lactic acid is formed, and co-products must be treated before elimination. The collaboration between lactic bacteria and dairy bacteria is experimented. The use of P. freudenreichii for the treatment of coproducts, and the stabilization of these bacteria by spray drying are also studied.Experiments show that P. freudenreichii CIRM-BIA 129 can grow in these coproducts and uses lactic acid. After the strain development, the lactic acid is eliminated, the coproduct is therefore treated. The strain still has its immunomodulatory potential after development in the new culture medium. The addition of excipient such as sweet whey and maltodextrin can increase the dry extract of the culture and causes a better dryability. P.freudenreichii resists to the culture spray drying if the culture contains 15% of excipient. However P.freudenreichii, does not survive during storage of powders in used drying conditionsLes bactéries propioniques laitières sont présentes dans de nombreux produits fermentés. Propionibacterium freudenreichii est reconnue comme bactérie GRAS (considérée sûre), et utilisée comme probiotique et ferment fromager. La souche P. freudenreichii CIRM-BIA 129 possède des protéines de surface ayant des capacités immunomodulatrice anti-inflammatoires. La production et la stabilisation des bactéries propioniques peuvent être optimisées. De plus, lors de la production de bactéries lactiques, de l’acide lactique est formé, et les coproduits doivent donc être traités avant d’être éliminés. Une collaboration entre bactéries lactiques et propioniques est expérimentée. L’utilisation de P. freudenreichii pour le traitement de l’acide lactique et la stabilisation de ces bactéries par séchage par atomisation sont également étudiées.Les expériences montrent que P. freudenreichii CIRM-BIA 129 peut se développer dans ces coproduits et qu’elle utilise l’acide lactique. Après le développement de la souche, l’acide lactique est éliminé, le coproduit est donc traité. La souche comporte toujours son potentiel immunomodulateur après développement dans le nouveau milieu de culture. L’ajout d’excipient comme le lactosérum doux et la maltodextrine permet d’augmenter l’extrait sec des cultures, et entraine une meilleure séchabilité. P. freudenreichii résiste au séchage par atomisation des cultures comportant 15% d’excipient. Cependant P. freudenreichii ne survit pas au cours du stockage des poudres avec les conditions de séchage utilisées

Taking Advantage of Bacterial Adaptation in Order to Optimize Industrial Production of Dry Propionibacterium freudenreichii

Propionibacterium freudenreichii is a beneficial bacterium, used both as a probiotic and as a cheese starter. Large-scale production of P. freudenreichii is required to meet growing consumers’ demand. Production, drying and storage must be optimized, in order to guarantee high P. freudenreichii viability within powders. Compared to freeze-drying, spray drying constitutes the most productive and efficient, yet the most stressful process, imposing severe oxidative and thermal constraints. The aim of our study was to provide the tools in order to optimize the industrial production of dry P. freudenreichii. Bacterial adaptation is a well-known protective mechanism and may be used to improve bacterial tolerance towards technological stresses. However, the choice of bacterial adaptation type must consider industrial constraints. In this study, we combined (i) modulation of the growth medium composition, (ii) heat-adaptation, and (iii) osmoadaptation, in order to increase P. freudenreichii tolerance towards technological stresses, including thermal and oxidative constraints, using an experimental design. We further investigated optimal growth and adaptation conditions, by monitoring intracellular compatible solutes accumulation. Glucose addition, coupled to heat-adaptation, triggered accumulation of trehalose and of glycine betaine, which further provided high tolerance towards spray drying and storage. This work opens new perspectives for high quality and fast production of live propionibacteria at the industrial scale

Improving the survival of beneficial bacteria during drying and storage by the exploitation of bacteria adaptation

Les probiotiques et les ferments sont produits sous forme de poudre pour faciliter leur stockage et leur utilisation. Ils sont généralement stabilisés par lyophilisation, procédé à la fois cher et à faible rendement. Le séchage par atomisation représente donc une alternative durable. Cependant, le séchage par atomisation impose des stress osmotique, chaud et oxydatif, et entraine alors la mort d’une part importante des bactéries.Dans ce travail, les adaptations de Propionibacterium freudenreichii durant différents traitements ont été étudiées pour mieux comprendre les protections-croisées apportées. Les mécanismes d’adaptation comprennent l’accumulation de solutés compatibles, la surproduction des protéines générales de stress et la modulation de la composition en acides gras membranaires. Pendant l’osmoadaptation, l’accumulation des solutés compatibles a été modulée par la quantité d’azote non-protéique et de glucides du milieu de culture.De grandes quantités de glycine bétaïne accumulées ont permis d’obtenir un fort taux de survie pendant la lyophilisation. En revanche, l’accumulation conjointe de glycine bétaïne et de tréhalose a permis d’obtenir un fort taux de survie pendant le séchage par atomisation. De plus, l’addition de glucide combinée à l’osmoadaptation ou à une thermo-adaptation provoque l’accumulation de trehalose et de glycine bétaïne.L’adaptation peut donc être optimisée pour obtenir des bactéries plus résistantes aux procédés de séchage, de stockage et de la digestion. Ces travaux offrent de nouvelles solutions pour la production de levains et de prProbiotic and starters are produced in powder form to facilitate storage, and usage. These beneficial bacteria are usually dried by freeze-drying, which is expensive with low productivity. Therefore, spray drying is an alternative and sustainable process to produce beneficial bacteria. Unfortunately, spray drying imposes osmotic, heat and oxidative stresses, this process can thus lead to the death of an important bacterial death.In this work, Propionibacterium freudenreichii adaptation to different treatments were studied to better understand their impact on cross-protections. Adaptation mechanisms included compatible solutes accumulation, general stress protein over-production and modulation of membrane fatty acids composition. During osmoadaptation, compatible solutes accumulation was fine-tuned by the medium composition and especially by the amount of non-protein nitrogen sources and of carbohydrates.Accumulation of high amounts of glycine betaine during osmoadaptation, led to high survival during freeze-drying. By contrast, accumulation of high amounts of glycine betaine and trehalose, led to high viability during spray drying. Different adaptations can trigger the same adaptation mechanisms. Accumulation of high amounts of trehalose and of glycine betaine was triggered by the addition of saccharides, combine to osmoadaptation or to heat-adaptation. Adaptation can be optimized to confer bacteria enhanced resistance during drying processes, storage and digestion. This work opens new avenues for the production of starters and probiotics with enhanced robustness

Les adaptations bactériennes améliorent la survie de Propionibacterium freudenreichii durant le séchage et le stockage

Probiotic and starters are produced in powder form to facilitate storage, and usage. These beneficial bacteria are usually dried by freeze-drying, which is expensive with low productivity. Therefore, spray drying is an alternative and sustainable process to produce beneficial bacteria. Unfortunately, spray drying imposes osmotic, heat and oxidative stresses, this process can thus lead to the death of an important bacterial death.In this work, Propionibacterium freudenreichii adaptation to different treatments were studied to better understand their impact on cross-protections. Adaptation mechanisms included compatible solutes accumulation, general stress protein over-production and modulation of membrane fatty acids composition. During osmoadaptation, compatible solutes accumulation was fine-tuned by the medium composition and especially by the amount of non-protein nitrogen sources and of carbohydrates.Accumulation of high amounts of glycine betaine during osmoadaptation, led to high survival during freeze-drying. By contrast, accumulation of high amounts of glycine betaine and trehalose, led to high viability during spray drying. Different adaptations can trigger the same adaptation mechanisms. Accumulation of high amounts of trehalose and of glycine betaine was triggered by the addition of saccharides, combine to osmoadaptation or to heat-adaptation. Adaptation can be optimized to confer bacteria enhanced resistance during drying processes, storage and digestion. This work opens new avenues for the production of starters and probiotics with enhanced robustness.Les probiotiques et les ferments sont produits sous forme de poudre pour faciliter leur stockage et leur utilisation. Ils sont généralement stabilisés par lyophilisation, procédé à la fois cher et à faible rendement. Le séchage par atomisation représente donc une alternative durable. Cependant, le séchage par atomisation impose des stress osmotique, chaud et oxydatif, et entraine alors la mort d’une part importante des bactéries.Dans ce travail, les adaptations de Propionibacterium freudenreichii durant différents traitements ont été étudiées pour mieux comprendre les protections-croisées apportées. Les mécanismes d’adaptation comprennent l’accumulation de solutés compatibles, la surproduction des protéines générales de stress et la modulation de la composition en acides gras membranaires. Pendant l’osmoadaptation, l’accumulation des solutés compatibles a été modulée par la quantité d’azote non-protéique et de glucides du milieu de culture.De grandes quantités de glycine bétaïne accumulées ont permis d’obtenir un fort taux de survie pendant la lyophilisation. En revanche, l’accumulation conjointe de glycine bétaïne et de tréhalose a permis d’obtenir un fort taux de survie pendant le séchage par atomisation. De plus, l’addition de glucide combinée à l’osmoadaptation ou à une thermo-adaptation provoque l’accumulation de trehalose et de glycine bétaïne.L’adaptation peut donc être optimisée pour obtenir des bactéries plus résistantes aux procédés de séchage, de stockage et de la digestion. Ces travaux offrent de nouvelles solutions pour la production de levains et de p

Driving adaptation of probiotic and starter bacteria in order to improve industrial production process

Driving adaptation of probiotic and starter bacteria in order to improve industrial production process. STLOpenday

The Cheese Matrix Modulates the Immunomodulatory Properties of Propionibacterium freudenreichii CIRM-BIA 129 in Healthy Piglets

International audiencePropionibacterium freudenreichii is a beneficial bacterium, used as a cheese starter, which presents versatile probiotic properties. These properties are strain-dependent. We hypothesized they may also be delivery vehicle-dependent. In this study, we thus explored in healthy piglets how the cheese matrix affects the immunomodulatory properties of P. freudenreichii. During 2 weeks, three groups of weaned piglets consumed, respectively, P. freudenreichii as a liquid culture (PF-culture), P. freudenreichii under the form of a cheese (PF-cheese), or a control sterile cheese matrix (Cheesematrix). The in vivo metabolic activity of P. freudenreichii was assessed by determining short chain fatty acids (SCFA) concentration and bifidobacteria population in feces. Whatever the delivery vehicle, P. freudenreichii was metabolically active in piglets’ colon and enhanced both bifidobacteria and SCFA in feces. P. freudenreichii consumption decreased the secretion of TNFa and of IL-10 by peripheral blood mononuclear cells (PBMC). It did not alter IL-10, IFNg, IL-17, and TNFa secretion in mesenteric lymph node immune cells (MLNC). PF-cheese enhanced significantly Treg phenotype, while PF-culture decreased significantly Th17 phenotype in PBMC and MLNC. Remarkably, only PF-cheese induced an increase of Th2 phenotype in PBMC and MLNC. Ex vivo stimulation of PBMC and MLNC by Lipopolysaccharides and Concanavalin A emphasized the difference in the immunomodulatory responses between PF-culture and PF-cheese group, as well as between PBMC and MLNC. This study shows the importance to consider the delivery vehicle for probiotic administration. It confirms the anti-inflammatory potential of P. freudenreichii. It opens new perspectives for the use propionibacteria-fermented products as preventive agents for inflammatory bowel diseases and intestinal infectious diseases

L’adaptation de Propionibacterium freudenreichii permet d’augmenter sa survie au séchage

IntroductionP. freudenreichii est utilisé comme ferment pour les fromages de type emmental et également comme probiotique. La stabilisation de P. freudenreichii sous forme de poudre est essentielle pour garantir une bonne stabilité. La lyophilisation et le séchage par atomisation peuvent être utilisés pour produire des ferments et des probiotiques en poudre. Le séchage par atomisation demande moins d’énergie que la lyophilisation. Cependant le séchage par atomisation expose les bactéries à de très hautes températures. Ce séchage est donc un procédé stressant, provocant des stress thermique, oxydatif et osmotique. En vue de résister à ce procédé, les bactéries doivent donc être adaptées. Les milieux de culture sont un moyen d’adapter les bactéries dans le but d’augmenter leur survie au séchage par atomisation.ObjectifDans cette étude, nous avons exploré l’influence de la composition du milieu de culture sur 1) l’accumulation d’osmoprotecteurs et 2) la survie des bactéries lors du séchage par atomisation.Matériels et méthodesPour le milieux riche YEL (Yeast Exctract Lactate) et les milieux laitiers, les concentrations de sel les plus élevées permettant la croissance de P. freudenreichii ont été choisies. Les osmoprotecteurs accumulés par P. freudenreichii ont été identifiés et quantifiés par RMN du proton et du carbone. Les différentes cultures ont ensuite subi les challenges oxydatifs, thermique et le séchage par atomisation.RésultatsPendant la croissance, P. feudenreichii CIRM-BIA 129 accumulent du tréhalose, du glutamate et de la glycine betaine. En condition hyper osmotique, l’accumulation de ces osmoprotecteurs est plus élevés. Le lactose naturellement présent dans le lactosérum, ou ajouté dans le YEL, provoque une adaptation au stress acide, mais augmente également l’accumulation intracellulaire du tréhalose. Le ratio tréhalose/glycine betaine accumulés est déterminé par le ratio azote/carbone du milieu de culture. Avec un ratio tréhalose/glycine betaine optimisé, P. freudenreihcii a une meilleure survie au challenge thermique et oxydatif, comme au séchage par atomisation.ConclusionPiloter la composition du milieu de culture de P. freudenreichii permet de diriger l’adaptation et conduit à une meilleure survie durant le séchage par atomisation. Ceci permettra par la suite la production de qualité des ferments et probiotiques avec de hauts rendement