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Suteaki Shioya - One of the best experts on this subject based on the ideXlab platform.
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Enhancement of 1,4-dihydroxy-2-naphthoic acid production by Propionibacterium freudenreichii ET-3 fed-batch culture.
Applied and environmental microbiology, 2007Co-Authors: Keisuke Furuichi, Yoshio Katakura, Kazuaki Ninomiya, Suteaki ShioyaAbstract:The production of 1,4-dihydroxy-2-naphthoic acid (DHNA) was investigated using a fed-batch culture of Propionibacterium freudenreichii ET-3. DHNA is a precursor of menaquinone (MK) and is transformed to MK by combination with an isoprenoid unit. We found that ET-3 stopped MK production and increased DHNA production in an anaerobic fed-batch culture by maintaining the lactose concentration at approximately zero. The maximum DHNA concentration observed in the anaerobic fed-batch culture was markedly higher than the maximum DHNA concentration observed in an anaerobic batch culture. Moreover, MK or DHNA production was affected by the lactose feeding rate; this suggests that lactose metabolism participates in the syntheses of these products. On the other hand, accumulation of propionate was found to inhibit DHNA production in the fed-batch culture. Based on the fact that ET-3 increases DHNA production in an aerobic culture by consuming propionate, we carried out a cultivation experiment in which an anaerobic fed-batch culture was switched to an anaerobic batch culture and found that the DHNA production was increased to a greater extent than the DHNA production in an anaerobic fed-batch culture. These results suggest that DHNA production by ET-3 is markedly influenced by carbon source limitation and the oxygen supply.
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optimal aerobic cultivation method for 1 4 dihydroxy 2 naphthoic acid production by Propionibacterium freudenreichii et 3
Journal of Bioscience and Bioengineering, 2006Co-Authors: Keisuke Furuichi, Yoshio Katakura, Kazuaki Ninomiya, Atsuko Amano, Suteaki ShioyaAbstract:To investigate the effects of oxygen supply on Propionibacterium freudenreichii ET-3 metabolism and 1,4-dihydroxy-2-naphthoic acid (DHNA) production in detail, the strain was cultured by switching from anaerobic condition to aerobic condition at 72 h (termed anaerobic-aerobic switching culture hereafter) employing different oxygen transfer rates (OTRs) in the range of 0.08–0.90 mg/( l ·h). It was found that a 0.08 mg/( l ·h) OTR could not change the metabolism or improve the DHNA production of P. freudenreichii ET-3. When the OTR was in the range of 0.23–0.66 mg/( l ·h), propionate, which inhibits DHNA production significantly, was consumed during the aerobic phase. Final DHNA concentration increased to 0.22 mM, irrespective of OTR. When the OTR was 0.90 mg/( l ·h), a sudden increase in dissolved oxygen (DO) concentration during the aerobic phase resulted in a sudden decrease in DHNA concentration. To attenuate the stresses caused by propionate and oxygen exposure, we designed an optimal cultivation in which the anaerobic and aerobic phases were repeated three times alternately. As a result, propionate concentration was maintained below the level that inhibits DHNA production, and no DO concentration was detected throughout the culture. The final DHNA concentration in this culture was 0.33 mM, which is 2.7-fold that in the anaerobic culture and 1.5-fold that in the anaerobic-aerobic switching culture.
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aerobic culture of Propionibacterium freudenreichii et 3 can increase production ratio of 1 4 dihydroxy 2 naphthoic acid to menaquinone
Journal of Bioscience and Bioengineering, 2006Co-Authors: Keisuke Furuichi, Yoshio Katakura, Kazuaki Ninomiya, Kenichi Hojo, Suteaki ShioyaAbstract:This is the first report on the production of both 1,4-dihydroxy-2-naphthoic acid (DHNA) and menaquinone by Propionibacterium freudenreichii ET-3. DHNA can be a stimulator of bifidogenic growth, and menaquinone has important roles in blood coagulation and bone metabolism. During anaerobic culture, DHNA and menaquinone concentrations reached 0.18 mM and 0.12 mM, respectively. The molar ratio between these products was approximately 3:2, which was not affected by culture pH and temperature over the ranges of 6.0-7.0 and 31-35 degrees C, respectively. As for organic acid, propionate and acetate accumulated at concentrations of 0.3 M and 0.15 M, respectively, and the propionate accumulation particularly inhibited further production of DHNA. To improve DHNA production, we switched from anaerobic condition to aerobic condition during the culture when lactose was depleted. DHNA concentration continued to increase even after lactose exhaustion, reaching 0.24 mM. In contrast to DHNA production, menaquinone production stopped after the switch to aerobic condition. The total molar production of DHNA and menaquinone was 0.3 mM irrespective of aerobic culture and anaerobic-aerobic switching culture. Therefore, the anaerobic-aerobic switching culture could increase the production ratio of DHNA to menaquinone. The DHNA concentration obtained from the anaerobic-aerobic switching culture was 1.3-fold higher than that in the anaerobic culture, because P. freudenreichii ET-3 utilized propionate accumulated in the medium via the reversed methylmalonyl CoA pathway under aerobic condition. The culture method proposed in this study could be applicable to industrial-scale fermentation using 1000 l of media, by which 0.23 mM DHNA was produced.
Houem Rabah - One of the best experts on this subject based on the ideXlab platform.
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Intracellular osmoprotectant concentrations determine Propionibacterium freudenreichii survival during drying
Applied Microbiology and Biotechnology, 2020Co-Authors: Floriane Gaucher, Houem Rabah, Koffigan Kponouglo, Sylvie Bonnassié, Philippe Blanc, Romain Jeantet, Pierre Marchand, Sandrine Pottier, Anne Dolivet, Gwénaël Jan¨Abstract:Propionibacterium freudenreichii is a beneficial bacterium widely used in food as a probiotic and as a cheese-ripening starter. In these different applications, it is produced, dried, and stored before being used. Both freeze-drying and spray-drying were considered for this purpose. Freeze-drying is a discontinuous process that is energy-consuming but that allows high cell survival. Spray-drying is a continuous process that is more energy-efficient but that can lead to massive bacterial death related to heat, osmotic, and oxidative stresses. We have shown that P. freudenreichii cultivated in hyperconcentrated rich media can be spray-dried with limited bacterial death. However, the general stress tolerance conferred by this hyperosmotic constraint remained a black box. In this study, we modulated P. freudenreichii growth conditions and monitored both osmoprotectant accumulation and stress tolerance acquisition. Changing the ratio between the carbohydrates provided and non-protein nitrogen during growth under osmotic constraint modulated osmoprotectant accumulation. This, in turn, was correlated with P. freudenreichii tolerance towards different stresses, on the one hand, and towards freeze-drying and spray-drying, on the other. Surprisingly, trehalose accumulation correlated with spray-drying survival and glycine betaine accumulation with freeze-drying. This first report showing the ability to modulate the trehalose/GB ratio in osmoprotectants accumulated by a probiotic bacterium opens new perspectives for the optimization of probiotics production.
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data from a proteomic analysis highlight different osmoadaptations in two strain of Propionibacterium freudenreichii
Data in Brief, 2020Co-Authors: Floriane Gaucher, Houem Rabah, Sylvie Bonnassié, Pierre Marchand, Sandrine Pottier, Julien Jardin, Valerie Briardbion, Pauline Leverrier, Romain JeantetAbstract:Abstract The article presents a proteomic data set generated by a comparative analysis of the proteomes of Propionibacterium freudenreichii, comparing the CIRM-BIA 129 and CIRM-BIA 1025 strains. The two strains were cultivated until the beginning of the stationary phase in a chemical defined medium (MMO), and in this medium in the presence of NaCl, with or without glycine betaine. Whole-cell proteins were extracted, trypsinolyzed and analyzed by nano LC-MS/MS, prior to identification and classification by function using the X!Tandem pipeline software and the proteomic data from NCBI.nlm.nigh.gov. Quantification of proteins was then carried out in order to detect change in their expression depending on the culture medium. This article is related to the research article entitled “Benefits and drawbacks of osmotic adjustment in Propionibacterium freudenreichii”. The comparative proteomic analysis of the two strains reveal strain-dependent and medium-dependent stress proteomes in the probiotic P. freudenreichii.
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Osmotic adjustments may exert benefits or drawbacks on Propionibacterium freudenreichii viability during freeze-drying.
2019Co-Authors: Floriane Gaucher, Houem Rabah, Sylvie Bonnassié, Philippe Blanc, Romain Jeantet, Pierre Marchand, Gwénaël Jan¨Abstract:Propionibacterium freudenreichii is a beneficial bacterium consumed in cheeses and in probiotic food supplements. Indeed, selected strains of P. freudenreichii combine both technological and probiotic abilities. Selected strains were shown to adhere to intestinal cells and to modulate the mucosal immune response as well as the proliferation/apoptosis balance. P. freudenreichii are thus used as Swiss-type cheese starters and as probiotics in the industry. For these purposes, P. freudenreichii should be converted into a powdered form to be stabilized, without significant loss of viability. It suffers hyperosmotic constraints in different key steps of its use. This includes dry starter preparation, cheese making and transit through the human digestive tract. Osmotic adaptation, and the corresponding adjustments, are prerequisites to the use of P. freudenreichii. Moreover, during its large-scale industrial production, stabilisation of dried bacteria under a long-lasting live form is a critical point. This process being stressful, pre-treatments should aim at an enhanced tolerance. Culture media constitute a means to adapt bacteria in the aim to increase their survival during technological and digestive processes. In this study, we explored the impact of osmoadaptation in two P. freudenreichii strains, in different conditions, on survival. Method The maximal salt concentrations allowing growth, and inducing osmoadapatation, was selected, using a chemically defined medium. For all experiments, cells were collected at the beginning of the stationary phase. Accumulated osmoprotectants were identified and quantified by C and H NMR. Whole cell proteins were identified and quantified by LC-MS/MS. The different cultures were then submitted to oxidative, heat, acid, bile salts challenges and to freeze-drying. Results During growth, both P. freudenreichii strains accumulated trehalose, glutamate and glycine betaine. Osmoadaptation, strain-dependent, had different effects on multiple stress tolerance, depending on the presence of osmoprotectants. One of the two strains growth was restored by provided glycine betaine (GB). In this strain, salt stress preadaptation enhanced tolerance towards heat, oxidative, acid and freeze-drying challenges in the absence of GB. However, transport and accumulation of provided GB had deleterious effects on stress tolerance, while restoring optimal growth under hyperosmotic constraint. In the other strain, neither salt, nor GB, enhanced stress tolerance, which was constitutively low. Accordingly, whole cell proteomics revealed differences between strains, as well as different adaptive mechanisms triggered by salt in the presence and in the absence of GB. Discussion Osmotic adjustments may thus have positive or deleterious effects on industrial abilities of P. freudenreichii. Tuning of the culture medium composition allows driving bacterial adaptation and leads to better survival during freeze-drying and/or digestion. This, in turn, should lead to high yield and high-quality production of starters and probiotics.
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la vectorisation de Propionibacterium freudenreichii cirm bia 129 et de ses proteines immunomodulatrices par la matrice fromagere vers le tube digestif
2019Co-Authors: Houem RabahAbstract:Propionibacterium freudenreichii CIRM-BIA 129 (Pf) est une bacterie benefique utilisee comme levain fromager. Elle presente en outre de nombreuses potentialites probiotiques souche-dependantes, dont la modulation de l’inflammation. Cette propriete resulte de la production de divers metabolites. Les proteines de surface S-layer (Slps), dont la proteine majoritaire SlpB, y jouent egalement un role immunomodulateur. Les proprietes « 2-en-1 », c’est-a-dire a la fois fermentaires et probiotiques, font de Pf un bon candidat pour developper des fromages fonctionnels, afin de prevenir les maladies inflammatoires intestinales. L’objectif de cette these etait d’etudier l’impact de la matrice fromagere sur les proprietes immunomodulatrices de Pf, via ses proteines Slps, par comparaison a une culture sur ultrafiltrat de lait (UF). Les etudes conduites in vitro suggerent que les bacteries provenant du fromage auraient une meilleure capacite de tolerance aux stress gastriques et duodenaux, mais elleauraient une moindre capacite de survie dans le colon, par comparaison a des bacteries provenant d’une culture sur UF. De plus, la proteolyse digestive des proteines de surface ameliore la survie de Pf dans le colon. Parallelement, l’etude de digestion in vitro a montre que la proteolyse des proteines de surface a seulement ete reduite par la matrice fromagere. Cette proteolyse conduit a l’abolition des effets anti-inflammatoires des proteines Slps, qui ne sont pas exprimees de novo dans l’environnement colique. Ces resultats obtenus in vitro etaient coherents avec l’etude in vivo qui a mont
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Propionibacterium freudenreichii CIRM-BIA 129 Osmoadaptation Coupled to Acid-Adaptation Increases Its Viability During Freeze-Drying
Frontiers in Microbiology, 2019Co-Authors: Floriane Gaucher, Houem Rabah, Koffigan Kponouglo, Pierre Marchand, Sandrine Pottier, Julien Jardin, Valérie Briard-bion, Sylvie Bonnassie-rouxin, Jordane Ossemond, Philippe BlancAbstract:Propionibacterium freudenreichii is a beneficial bacterium with documented effects on the gut microbiota and on inflammation. Its presence within the animal and human intestinal microbiota was correlated with immunomodulatory effects, mediated by both propionibacterial surface components and by secreted metabolites. It is widely implemented, both in the manufacture of fermented dairy products such as Swiss-type cheeses, and in the production of probiotic food complements, under the form of freeze-dried powders. The bottleneck of this drying process consists in the limited survival of bacteria during drying and storage. Protective pre-treatments have been applied to other bacteria and may, in a strain-dependent manner, confer enhanced resistance. However, very little information was yet published on P. freudenreichii adaptation to freeze-drying. In this report, an immunomodulatory strain of this probiotic bacterium was cultured under hyperosmotic constraint in order to trigger osmoadaptation. This adaptation was then combined with acid or thermal pre-treatment. Such combination led to accumulation of key stress proteins, of intracellular compatible solute glycine betaine, to modulation of the propionibacterial membrane composition, and to enhanced survival upon freeze-drying. This work opens new perspectives for efficient production of live and active probiotic propionibacteria.
Keisuke Furuichi - One of the best experts on this subject based on the ideXlab platform.
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Enhancement of 1,4-dihydroxy-2-naphthoic acid production by Propionibacterium freudenreichii ET-3 fed-batch culture.
Applied and environmental microbiology, 2007Co-Authors: Keisuke Furuichi, Yoshio Katakura, Kazuaki Ninomiya, Suteaki ShioyaAbstract:The production of 1,4-dihydroxy-2-naphthoic acid (DHNA) was investigated using a fed-batch culture of Propionibacterium freudenreichii ET-3. DHNA is a precursor of menaquinone (MK) and is transformed to MK by combination with an isoprenoid unit. We found that ET-3 stopped MK production and increased DHNA production in an anaerobic fed-batch culture by maintaining the lactose concentration at approximately zero. The maximum DHNA concentration observed in the anaerobic fed-batch culture was markedly higher than the maximum DHNA concentration observed in an anaerobic batch culture. Moreover, MK or DHNA production was affected by the lactose feeding rate; this suggests that lactose metabolism participates in the syntheses of these products. On the other hand, accumulation of propionate was found to inhibit DHNA production in the fed-batch culture. Based on the fact that ET-3 increases DHNA production in an aerobic culture by consuming propionate, we carried out a cultivation experiment in which an anaerobic fed-batch culture was switched to an anaerobic batch culture and found that the DHNA production was increased to a greater extent than the DHNA production in an anaerobic fed-batch culture. These results suggest that DHNA production by ET-3 is markedly influenced by carbon source limitation and the oxygen supply.
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optimal aerobic cultivation method for 1 4 dihydroxy 2 naphthoic acid production by Propionibacterium freudenreichii et 3
Journal of Bioscience and Bioengineering, 2006Co-Authors: Keisuke Furuichi, Yoshio Katakura, Kazuaki Ninomiya, Atsuko Amano, Suteaki ShioyaAbstract:To investigate the effects of oxygen supply on Propionibacterium freudenreichii ET-3 metabolism and 1,4-dihydroxy-2-naphthoic acid (DHNA) production in detail, the strain was cultured by switching from anaerobic condition to aerobic condition at 72 h (termed anaerobic-aerobic switching culture hereafter) employing different oxygen transfer rates (OTRs) in the range of 0.08–0.90 mg/( l ·h). It was found that a 0.08 mg/( l ·h) OTR could not change the metabolism or improve the DHNA production of P. freudenreichii ET-3. When the OTR was in the range of 0.23–0.66 mg/( l ·h), propionate, which inhibits DHNA production significantly, was consumed during the aerobic phase. Final DHNA concentration increased to 0.22 mM, irrespective of OTR. When the OTR was 0.90 mg/( l ·h), a sudden increase in dissolved oxygen (DO) concentration during the aerobic phase resulted in a sudden decrease in DHNA concentration. To attenuate the stresses caused by propionate and oxygen exposure, we designed an optimal cultivation in which the anaerobic and aerobic phases were repeated three times alternately. As a result, propionate concentration was maintained below the level that inhibits DHNA production, and no DO concentration was detected throughout the culture. The final DHNA concentration in this culture was 0.33 mM, which is 2.7-fold that in the anaerobic culture and 1.5-fold that in the anaerobic-aerobic switching culture.
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aerobic culture of Propionibacterium freudenreichii et 3 can increase production ratio of 1 4 dihydroxy 2 naphthoic acid to menaquinone
Journal of Bioscience and Bioengineering, 2006Co-Authors: Keisuke Furuichi, Yoshio Katakura, Kazuaki Ninomiya, Kenichi Hojo, Suteaki ShioyaAbstract:This is the first report on the production of both 1,4-dihydroxy-2-naphthoic acid (DHNA) and menaquinone by Propionibacterium freudenreichii ET-3. DHNA can be a stimulator of bifidogenic growth, and menaquinone has important roles in blood coagulation and bone metabolism. During anaerobic culture, DHNA and menaquinone concentrations reached 0.18 mM and 0.12 mM, respectively. The molar ratio between these products was approximately 3:2, which was not affected by culture pH and temperature over the ranges of 6.0-7.0 and 31-35 degrees C, respectively. As for organic acid, propionate and acetate accumulated at concentrations of 0.3 M and 0.15 M, respectively, and the propionate accumulation particularly inhibited further production of DHNA. To improve DHNA production, we switched from anaerobic condition to aerobic condition during the culture when lactose was depleted. DHNA concentration continued to increase even after lactose exhaustion, reaching 0.24 mM. In contrast to DHNA production, menaquinone production stopped after the switch to aerobic condition. The total molar production of DHNA and menaquinone was 0.3 mM irrespective of aerobic culture and anaerobic-aerobic switching culture. Therefore, the anaerobic-aerobic switching culture could increase the production ratio of DHNA to menaquinone. The DHNA concentration obtained from the anaerobic-aerobic switching culture was 1.3-fold higher than that in the anaerobic culture, because P. freudenreichii ET-3 utilized propionate accumulated in the medium via the reversed methylmalonyl CoA pathway under aerobic condition. The culture method proposed in this study could be applicable to industrial-scale fermentation using 1000 l of media, by which 0.23 mM DHNA was produced.
Gwénaël Jan¨ - One of the best experts on this subject based on the ideXlab platform.
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Intracellular osmoprotectant concentrations determine Propionibacterium freudenreichii survival during drying
Applied Microbiology and Biotechnology, 2020Co-Authors: Floriane Gaucher, Houem Rabah, Koffigan Kponouglo, Sylvie Bonnassié, Philippe Blanc, Romain Jeantet, Pierre Marchand, Sandrine Pottier, Anne Dolivet, Gwénaël Jan¨Abstract:Propionibacterium freudenreichii is a beneficial bacterium widely used in food as a probiotic and as a cheese-ripening starter. In these different applications, it is produced, dried, and stored before being used. Both freeze-drying and spray-drying were considered for this purpose. Freeze-drying is a discontinuous process that is energy-consuming but that allows high cell survival. Spray-drying is a continuous process that is more energy-efficient but that can lead to massive bacterial death related to heat, osmotic, and oxidative stresses. We have shown that P. freudenreichii cultivated in hyperconcentrated rich media can be spray-dried with limited bacterial death. However, the general stress tolerance conferred by this hyperosmotic constraint remained a black box. In this study, we modulated P. freudenreichii growth conditions and monitored both osmoprotectant accumulation and stress tolerance acquisition. Changing the ratio between the carbohydrates provided and non-protein nitrogen during growth under osmotic constraint modulated osmoprotectant accumulation. This, in turn, was correlated with P. freudenreichii tolerance towards different stresses, on the one hand, and towards freeze-drying and spray-drying, on the other. Surprisingly, trehalose accumulation correlated with spray-drying survival and glycine betaine accumulation with freeze-drying. This first report showing the ability to modulate the trehalose/GB ratio in osmoprotectants accumulated by a probiotic bacterium opens new perspectives for the optimization of probiotics production.
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Fermented milks, using Lactobacillus casei or Propionibacterium freudenreichii, prevent mucositis, a side effect of chemotherapy, in mice
2019Co-Authors: Gwénaël Jan¨Abstract:Introduction and Objectives Mucositis, a common side effect of cancer chemotherapy, is a clinically important gastrointestinal inflammatory disease. It consists in a painful inflammation and ulceration of the digestive mucosa, which may compromise proper nutrition of the patient, as well as termination of the treatment. It thus increases mortality and morbidity and contributes to rising health care costs. Its treatment is mainly supportive and often fails to relieve symptoms. Probiotic bacteria may maintain homeostasis and reduce side effects of chemotherapy [1]. A major limit to probiotic bacteria anti-inflammatory effect is their susceptibility to digestive stresses and several studies indicate the key protective role of food matrices in increasing probiotics’ effect via the protection of bacteria against digestive constraints. In this study, we investigated the role of fermented milk, using the immunomodulatory Lactobacillus casei BL23 [2] or Propionibacterium freudenreichii CB129 [3, 4] as a probiotic starter, and of its fortification via addition of whey proteins. Materials and methods We evaluated the role of whey protein isolate (WPI), when added to skim milk fermented by L. casei BL23 or by P. freudenreichii 138, as a protective matrix against in vitro stress challenges. In addition, we investigated in vivo the therapeutic effect of these fermented beverages in a murine model of mucositis induced by 5-Fluorouracil (5-FU). The monitored outcomes included weight loss, ileum histopathological score, villus height, crypt depth and number of mucus-producing goblet cells. Results This study demonstrated that milk supplementation with 30% (w/v) of WPI increases the survival rate of both strains when challenged with acid or bile salts, compared to fermented skim milk without the addition of WPI. Moreover, treatment with the probiotic beverages prevented weight loss and intestinal damages in mice receiving 5-FU. All symptoms of mucositis were drastically reduced by the consumption of developed probiotic fermented milks [5]. Conclusion This study evidenced the protective effect of selected strains of both lactic and propionic acid bacteria, in the context of induced mucositis. It confirmed that selected strains might be used both as starter and probiotic. It opens new avenues for the development of fermented functional foods for target populations.
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Osmotic adjustments may exert benefits or drawbacks on Propionibacterium freudenreichii viability during freeze-drying.
2019Co-Authors: Floriane Gaucher, Houem Rabah, Sylvie Bonnassié, Philippe Blanc, Romain Jeantet, Pierre Marchand, Gwénaël Jan¨Abstract:Propionibacterium freudenreichii is a beneficial bacterium consumed in cheeses and in probiotic food supplements. Indeed, selected strains of P. freudenreichii combine both technological and probiotic abilities. Selected strains were shown to adhere to intestinal cells and to modulate the mucosal immune response as well as the proliferation/apoptosis balance. P. freudenreichii are thus used as Swiss-type cheese starters and as probiotics in the industry. For these purposes, P. freudenreichii should be converted into a powdered form to be stabilized, without significant loss of viability. It suffers hyperosmotic constraints in different key steps of its use. This includes dry starter preparation, cheese making and transit through the human digestive tract. Osmotic adaptation, and the corresponding adjustments, are prerequisites to the use of P. freudenreichii. Moreover, during its large-scale industrial production, stabilisation of dried bacteria under a long-lasting live form is a critical point. This process being stressful, pre-treatments should aim at an enhanced tolerance. Culture media constitute a means to adapt bacteria in the aim to increase their survival during technological and digestive processes. In this study, we explored the impact of osmoadaptation in two P. freudenreichii strains, in different conditions, on survival. Method The maximal salt concentrations allowing growth, and inducing osmoadapatation, was selected, using a chemically defined medium. For all experiments, cells were collected at the beginning of the stationary phase. Accumulated osmoprotectants were identified and quantified by C and H NMR. Whole cell proteins were identified and quantified by LC-MS/MS. The different cultures were then submitted to oxidative, heat, acid, bile salts challenges and to freeze-drying. Results During growth, both P. freudenreichii strains accumulated trehalose, glutamate and glycine betaine. Osmoadaptation, strain-dependent, had different effects on multiple stress tolerance, depending on the presence of osmoprotectants. One of the two strains growth was restored by provided glycine betaine (GB). In this strain, salt stress preadaptation enhanced tolerance towards heat, oxidative, acid and freeze-drying challenges in the absence of GB. However, transport and accumulation of provided GB had deleterious effects on stress tolerance, while restoring optimal growth under hyperosmotic constraint. In the other strain, neither salt, nor GB, enhanced stress tolerance, which was constitutively low. Accordingly, whole cell proteomics revealed differences between strains, as well as different adaptive mechanisms triggered by salt in the presence and in the absence of GB. Discussion Osmotic adjustments may thus have positive or deleterious effects on industrial abilities of P. freudenreichii. Tuning of the culture medium composition allows driving bacterial adaptation and leads to better survival during freeze-drying and/or digestion. This, in turn, should lead to high yield and high-quality production of starters and probiotics.
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Propionibacterium freudenreichii Surface Protein SlpB Is Involved in Adhesion to Intestinal HT-29 Cells.
2018Co-Authors: Gwénaël Jan¨, Floriane Gaucher, Houem Rabah, Julien Jardin, Song Huang, Fillipe Luiz Rosa Do Carmo, Martine Deplanche, Stéphanie Dutertre, Yves Le Loir, Vasco AzevedoAbstract:Propionibacterium freudenreichii is a beneficial bacterium traditionally used as a cheese-ripening starter and more recently for its probiotic abilities based on the release of beneficial metabolites. In addition to these metabolites (short-chain fatty acids, vitamins, and bifidogenic factor), P. freudenreichii revealed an immunomodulatory effect confirmed in vivo by the ability to protect mice from induced acute colitis. This effect is, however, highly strain-dependent. Local action of metabolites and of immunomodulatory molecules is favoured by the ability of probiotics to adhere to the host cells. This property depends on key surface compounds, still poorly characterized in propionibacteria. We thus investigated the involvement of a specific propionibacterial surface protein in adhesion of the probiotic Propionibacterium freudenreichii to host cells. We compared several strains of Propionibacterium freudenreichii, with respect to adhesion to cultured human intestinal epithelial cells, using a well-established model. We inactivated genes using a previously developed method, based on the integration of a non-replicative plasmid, harbouring a portion of the target gene. We analysed adhesion of the wild type and the mutant strains using fluorescent-assisted flow cytometry and confocal microscopy. We also sought inhibition of adhesion using antibodies directed against surface layer proteins. In the present study, we showed different adhesion rates to cultured human intestinal cells, among strains of P. freudenreichii. The most adhesive one was P. freudenreichii CIRM-BIA 129, which is known to expose surface-layer proteins. We evidenced here the involvement of these proteins in adhesion to cultured human colon cells. We then aimed at deciphering the mechanisms involved in adhesion. Adhesion was inhibited by antibodies raised against SlpB, one of the surface-layer proteins in P. freudenreichii CIRM-BIA 129. Inactivation of the corresponding gene suppressed adhesion, further evidencing the key role of slpB product in cell adhesion. This work confirms the various functions fulfilled by surface-layer proteins, including probiotic/host interactions. It opens new perspectives for the understanding of probiotic determinants in propionibacteria, and for the selection of the most efficient strains within the P. freudenreichii species.
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Combining selected immunomodulatory Propionibacterium freudenreichii and Lactobacillus delbrueckii strains: development of an anti-inflammatory cheese.
2018Co-Authors: Gwénaël Jan¨Abstract:Scope. Inflammatory bowel disease (IBD) constitute a growing public health concern in western countries. Bacteria with anti-inflammatory properties are lacking in the dysbiosis accompanying IBD. Selected strains of probiotic bacteria with anti-inflammatory properties accordingly alleviate symptoms and enhance treatment of ulcerative colitis in clinical trials. Such properties are also found in selected strains of dairy starters such as Propionibacterium freudenreichii and Lactobacillus delbrueckii. We thus investigated the possibility to develop a fermented dairy product, combining both starter and probiotic abilities of both lactic acid and propionic acid bacteria, designed to extend remissions in IBD patients. Methods and results. We developed a two-strain P. freudenreichii and L. delbrueckii-fermented experimental pressed cheese using strains previously selected for their anti-inflammatory properties. Consumption of this experimental fermented dairy product protected mice against TNBS-induced colitis, alleviating severity of symptoms, modulating local and systemic inflammation, as well as colonic oxidative stress and epithelial cell damages. As a control, the corresponding sterile dairy matrix failed to afford such protection. Conclusion. This work reveals the probiotic potential of this bacterial mixture, in the context of fermented dairy products. It opens new perspectives for the reverse engineering development of anti-inflammatory fermented foods designed for target populations with IBD, and has provided evidences leading to an ongoing pilot clinical study in ulcerative colitis patients.
Gaucher Floriane - One of the best experts on this subject based on the ideXlab platform.
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Data from a proteomic analysis highlight different osmoadaptations in two strain of Propionibacterium freudenreichii
Elsevier, 2020Co-Authors: Gaucher Floriane, Rabah Houem, Jeantet Romain, Bonnassie Sylvie, Leverrier Pauline, Pottier Sandrine, Jardin Julien, Briard-bion Valérie, Marchand Pierre, Blanc PhilippeAbstract:International audienceThe article presents a proteomic data set generated by a comparative analysis of the proteomes of Propionibacterium freudenreichii, comparing the CIRM-BIA 129 and CIRM-BIA 1025 strains. The two strains were cultivated until the beginning of the stationary phase in a chemical defined medium (MMO), and in this medium in the presence of NaCl, with or without glycine betaine. Whole-cell proteins were extracted, trypsinolyzed and analyzed by nano LC-MS/MS, prior to identification and classification by function using the X!Tandem pipeline software and the proteomic data from NCBI.nlm.nigh.gov. Quantification of proteins was then carried out in order to detect change in their expression depending on the culture medium. This article is related to the research article entitled "Benefits and drawbacks of osmotic adjustment in Pro-pionibacterium freudenreichii". The comparative proteomic analysi
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Differential Adaptation of Propionibacterium freudenreichii CIRM-BIA129 to Cow’s Milk Versus Soymilk Environments Modulates Its Stress Tolerance and Proteome
'Frontiers Media SA', 2020Co-Authors: Tarnaud Florian, Gaucher Floriane, Jardin Julien, Briard-bion Valérie, Do Carmo, Fillipe Luiz Rosa, Illikoud Nassima, Guyomarc'h Fanny, Gagnaire Valérie, Jan GwenaelAbstract:International audiencePropionibacterium freudenreichii is a beneficial bacterium that modulates the gut microbiota, motility and inflammation. It is traditionally consumed within various fermented dairy products. Changes to consumer habits in the context of foodtransition are, however, driving the demand for non-dairy fermented foods, resulting in a considerable development of plant-based fermented products that require greater scientific knowledge. Fermented soymilks, in particular, offer an alternative source of live probiotics. While the adaptation of lactic acid bacteria (LAB) to such vegetable substrates is well documented, little is known about that of propionibacteria. We therefore investigated the adaptation of Propionibacterium freudenreichii to soymilk by comparison to cow’s milk. P. freudenreichii grew in co w’s milk but not in soymilk, but it did grow in soymilk when co-cultured with the lactic acid bacterium Lactobacillus plantarum. When grown in soymilk ultrafiltrate (SUF, the aqueous phase of soymilk), P. freudenreichii cells appeared thinner and rectangular-shaped, while they were thicker and more rounded in cow’s milk utltrafiltrate (MUF, the aqueous phase of cow milk). The amount of extractable surface proteins (SlpA, SlpB, SlpD, SlpE) was furthermore reduced in SUF, when compared to MUF. This included the SlpB protein, previously shown to modulate adhesion and immunomodulation in P. freudenreichii. Tolerance toward an acid and toward a bile salts challenge were enhanced in SUF. By contrast, tolerance toward an oxidative and a thermal challenge were enhancedin MUF. A whole-cell proteomic approach further identified differential expression of 35 proteins involved in amino acid transport and metabolism (including amino acid dehydrogenase, amino acid transporter), 32 proteins involved in arbohydrate transport and metabolism (including glycosyltransferase, PTS), indicating metabolic adaptationto the substrate. The culture medium also modulated the amount of stress proteins | involved in stress remediation: GroEL, OpuCA, CysK, DnaJ, GrpE, in line with the modulation of stress tolerance. Changing the fermented substrate may thus significantly affect the fermentative and probiotic properties of dairy propionibacteria. This needs to be considered when developing new fermented functional foods
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Benefits and drawbacks of osmotic adjustment in Propionibacterium freudenreichii
'Elsevier BV', 2019Co-Authors: Gaucher Floriane, Rabah Houem, Jeantet Romain, Bonnassie Sylvie, Leverrier Pauline, Pottier Sandrine, Jardin Julien, Briard-bion Valérie, Marchand Pierre, Blanc PhilippeAbstract:International audiencePropionibacterium freudenreichii is a beneficial bacterium used as a cheese starter and as a probiotic. Indeed, selected strains of P. freudenreichii combine both technological and health-promoting abilities. Moreover, during large-scale industrial production of dried bacteria and during consumption, P. freudenreichii may undergo different stressful processes. Osmotic adaptation was shown to enhance P. freudenreichii tolerance towards stresses, which are encountered during freeze-drying and during digestion. In this report, we compared the osmoadaptation molecular mechanisms of two P. freudenreichii strains. Both osmotolerance and osmoadaptation were strain-dependent and had different effects on multiple stress tolerance, depending on the presence of osmoprotectants. Availability of glycine betaine (GB) restored the growth of one of the two strains. In this strain, osmotic preadaptation enhanced heat, oxidative and acid stresses tolerance, as well as survival upon freezedrying. However, addition of GB in the medium had deleterious effects on stress tolerance, while restoring optimal growth under hyperosmotic constraint. In the other strain, neither salt nor GB enhanced stress tolerance, which was constitutively low. Accordingly, whole cell proteomics revealed that mechanisms triggered by salt in the presence and in the absence of GB are different between strains. Osmotic adjustment may thus have deleterious effects on industrial abilities of P. freudenreichii. Biological significance: Propionibacteria are found in various niches including fodder, silage, rumen, milk and cheeses. This means adaptation towards different ecological environments with different physicochemical parameters. Propionibacterium freudenreichii, in particular, is furthermore used both as dairy starter and as probiotic and is thus submitted to high scale industrial production. Production and subsequent stabilization still need optimization. Drying processes like freeze-drying are stressful. Osmotic adjustments may modulated tolerance towards drying. However, they are strain-dependent, medium-dependent and may either reduce or increase stress tolerance. A case-by-case study, for each strain-medium thus seems necessary. In this work, we identify key proteins involved in osmoadaptation and give new insights into adaptation mechanisms in P. freudenreichii. This opens new perspectives for the selections of strains and for the choice of the growth medium composition
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Les adaptations bactériennes améliorent la survie de Propionibacterium freudenreichii durant le séchage et le stockage
HAL CCSD, 2019Co-Authors: Gaucher FlorianeAbstract:Probiotic and starters are produced in powder form to facilitate storage, delivery and usage.These beneficial bacteria are usually dried by freezedrying, which is expensive with low productivity. From this point of view, spray drying is an alternative and sustainable process to produce beneficial bacteria. Unfortunately, this process imposes osmotic, heat and oxidative stresses: therefore, it can lead to 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 nonprotein nitrogen sources and 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, combined 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 stabilisés sous forme de poudre pour faciliter leur stockage et leur utilisation. Ils sont généralement séchés par lyophilisation, procédé à la fois coûteux énergétiquement et à faible rendement. Le séchage par pulvérisation représente donc une alternative durable. Cependant, il impose des stress osmotique, chaud et oxydatif, et entraine par suite 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 induites. 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 pulvérisation. De plus, l’addition de glucide combinée à ’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 probiotiques avec des applications plus nombreuses
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Les adaptations bactériennes améliorent la survie de Propionibacterium freudenreichii durant le séchage et le stockage
HAL CCSD, 2019Co-Authors: Gaucher FlorianeAbstract: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
