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This review provides a comprehensive overview of the current state of probiotic research, covering a wide range of topics, including strain identification, functional characterization, preclinical and clinical evaluations, mechanisms of action, therapeutic applications, manufacturing considerations, and future directions. The screening process for potential probiotics involves phenotypic and genomic analysis to identify strains with health-promoting properties while excluding those with any factor that could be harmful to the host. In vitro assays for evaluating probiotic traits such as acid tolerance, bile metabolism, adhesion properties, and antimicrobial effects are described. The review highlights promising findings from in vivo studies on probiotic mitigation of inflammatory bowel diseases, chemotherapy-induced mucositis, dysbiosis, obesity, diabetes, and bone health, primarily through immunomodulation and modulation of the local microbiota in human and animal models. Clinical studies demonstrating beneficial modulation of metabolic diseases and human central nervous system function are also presented. Manufacturing processes significantly impact the growth, viability, and properties of probiotics, and the composition of the product matrix and supplementation with prebiotics or other strains can modify their effects. The lack of regulatory oversight raises concerns about the quality, safety, and labeling accuracy of commercial probiotics, particularly for vulnerable populations. Advancements in multi-omics approaches, especially probiogenomics, will provide a deeper understanding of the mechanisms behind probiotic functionality, allowing for personalized and targeted probiotic therapies. However, it is crucial to simultaneously focus on improving manufacturing practices, implementing quality control standards, and establishing regulatory oversight to ensure the safety and efficacy of probiotic products in the face of increasing therapeutic applications. [ABSTRACT FROM AUTHOR]

The haemoglobin content in meat is consistently associated with an increased risk of colorectal cancer, whereas calcium may play a role as a chemopreventive agent. Using rodent models, calcium salts have been shown to prevent the promotion of haem-induced and red meat-induced colorectal carcinogenesis by limiting the bioavailability of the gut luminal haem iron. Therefore, this study aimed to compare impacts of dietary calcium provided as calcium salts or dairy matrix on gut homoeostasis perturbations by high haeminic or non-haeminic iron intakes. A 3-week intervention study was conducted using Fischer 344 rats. Compared to the ferric citrate-enriched diet, the haemoglobin-enriched diet led to increased faecal, mucosal, and urinary lipoperoxidation-related biomarkers, resulting from higher gut luminal haem iron bioavailability. This redox imbalance was associated to a dysbiosis of faecal microbiota. The addition of calcium to haemoglobin-enriched diets limited haem iron bioavailability and counteracted redox imbalance, with improved preventive efficacy when calcium was provided in dairy matrix. Data integration revealed correlations between haem-induced lipoperoxidation products and bacterial communities belonging to Peptococcaceae, Eubacterium coprostanoligenes group, and Bifidobacteriaceae. This integrated approach provides evidence of the benefits of dairy matrix as a dietary calcium vehicle to counteract the deleterious side-effects of meat consumption. [ABSTRACT FROM AUTHOR]

The growing incidence of human diseases involving inflammation and increased gut permeability makes the quest for protective functional foods more crucial than ever. Propionibacterium freudenreichii (P. freudenreichii) is a beneficial bacterium used in the dairy and probiotic industries. Selected strains exert anti-inflammatory effects, and the present work addresses whether the P. freudenreichii CIRM-BIA129, consumed daily in a preventive way, could protect mice from acute colitis induced by dextran sodium sulfate (DSS), and more precisely, whether it could protect from intestinal epithelial breakdown induced by inflammation. P. freudenreichii CIRM-BIA129 mitigated colitis severity and inhibited DSS-induced permeability. It limited crypt length reduction and promoted the expression of zonula occludens-1 (ZO-1), without reducing interleukin-1b mRNA (il-1b ) expression. In vitro, P. freudenreichii CIRM-BIA129 prevented the disruption of a Caco-2 monolayer induced by proinflammatory cytokines. It increased transepithelial electrical resistance (TEER) and inhibited permeability induced by inflammation, along with an increased ZO-1 expression. Extracellular vesicles (EVs) from P. freudenreichii CIRM-BIA129, carrying the surface layer protein (SlpB), reproduced the protective effect of P. freudenreichii CIRM-BIA129. A mutant strain deleted for slpB (DslpB), or EVs from this mutant strain, had lost their protective effects and worsened both DSS-induced colitis and inflammation in vivo. These results shown that P. freudenreichii CIRM-BIA129 daily consumption has the potential to greatly alleviate colitis symptoms and, particularly, to counter intestinal epithelial permeability induced by inflammation by restoring ZO-1 expression through mechanisms involving S-layer protein B. They open new avenues for the use of probiotic dairy propionibacteria and/or postbiotic fractions thereof, in the context of gut permeability. NEW & NOTEWORTHY Propionibacterium freudenreichii reduces dextran sodium sulfate (DSS)-induced intestinal permeability in vivo. P. freudenreichii does not inhibit inflammation but damages linked to inflammation. P. freudenreichii inhibits intestinal epithelial breakdown through S-layer protein B. The protective effects of P. freudenreichii depend on S-layer protein B. Extracellular vesicles from P. freudenreichii CB 129 mimic the protective effect of the probiotic. [ABSTRACT FROM AUTHOR]

Introduction: The mechanisms underlying innate immune memory (trained immunity) comprise epigenetic reprogramming of transcriptional pathways associated with alterations of intracellular metabolism. While the mechanisms of innate immune memory carried out by immune cells are well characterized, such processes in non-immune cells, are poorly understood. The opportunistic pathogen, Staphylococcus aureus, is responsible for a multitude of human diseases, including pneumonia, endocarditis and osteomyelitis, as well as animal infections, including chronic cattle mastitis that are extremely difficult to treat. An induction of innate immune memory may be considered as a therapeutic alternative to fight S. aureus infection. Methods: In the current work, we demonstrated the development of innate immune memory in non-immune cells during S. aureus infection employing a combination of techniques including Enzyme-linked immunosorbent assay (ELISA), microscopic analysis, and cytometry. Results: We observed that training of human osteoblast-like MG-63 cells and lung epithelial A549 cells with β-glucan increased IL-6 and IL-8 production upon a stimulation with S. aureus, concomitant with histones modifications. IL-6 and IL-8 production was positively correlated with an acetylation of histone 3 at lysine 27 (H3K27), thus suggesting epigenetic reprogramming in these cells. An addition of the ROS scavenger N-Acetylcysteine, NAC, prior to β-glucan pretreatment followed by an exposure to S. aureus, resulted in decreased IL-6 and IL-8 production, thereby supporting the involvement of ROS in the induction of innate immune memory. Exposure of cells to Lactococcus lactis resulted in increased IL-6 and IL-8 production by MG-63 and A549 cells upon a stimulation with S. aureus that was correlated with H3K27 acetylation, suggesting the ability of this beneficial bacterium to induce innate immune memory. Discussion: This work improves our understanding of innate immune memory in non-immune cells in the context of S. aureus infection. In addition to known inducers, probiotics may represent good candidates for the induction of innate immune memory. Our findings may help the development of alternative therapeutic approaches for the prevention of S. aureus infection. [ABSTRACT FROM AUTHOR]

Bacterial extracellular vesicles (EVs) are natural lipidic nanoparticles implicated in intercellular communication. Although EV research focused mainly on pathogens, the interest in probiotic-derived EVs is now rising. One example is Propionibacterium freudenreichii , which produces EVs with anti-inflammatory effects on human epithelial cells. Our previous study with P. freudenreichii showed that EVs purified by size exclusion chromatography (SEC) displayed variations in protein content according to bacterial growth conditions. Considering these content variations, we hypothesized that a comparative proteomic analysis of EVs recovered in different conditions would elucidate whether a representative vesicular proteome existed, possibly providing a robust proteome dataset for further analysis. Therefore, P. freudenreichii was grown in two culture media, and EVs were purified by sucrose density gradient ultracentrifugation (UC). Microscopic and size characterization confirmed EV purification, while shotgun proteomics unveiled that they carried a diverse set of proteins. A comparative analysis of the protein content of UC- and SEC-derived EVs, isolated from cultures either in UF (cow milk ultrafiltrate medium) or YEL (laboratory yeast extract lactate medium), showed that EVs from all these conditions shared 308 proteins. This EV core proteome was notably enriched in proteins related to immunomodulation. Moreover, it showed distinctive features, including highly interacting proteins, compositional biases for some specific amino acids, and other biochemical parameters. Overall, this work broadens the toolset for the purification of P. freudenreichii -derived EVs, identifies a representative vesicular proteome, and enumerates conserved features in vesicular proteins. These results hold the potential for providing candidate biomarkers of purification quality, and insights into the mechanisms of EV biogenesis and cargo sorting. [ABSTRACT FROM AUTHOR]

Inflammatory bowel diseases (IBD), a heterogeneous group of inflammatory conditions that encompass both ulcerative colitis and Crohn’s disease, represent a major public health concern. The etiology of IBD is not yet fully understood and no cure is available, with current treatments only showing long-term effectiveness in a minority of patients. A need to increase our knowledge on IBD pathophysiology is growing, to define preventive measures, to improve disease outcome, and to develop new effective and lasting treatments. IBD pathogenesis is sustained by aberrant immune responses, associated with alterations of the intestinal epithelial barrier (IEB), modifications of the enteric nervous system, and changes in microbiota composition. Currently, most of the treatments target the inflammation and the immune system, but holistic approaches targeting lifestyle and diet improvements are emerging. As dysbiosis is involved in IBD pathogenesis, pre-, pro-, syn-, and postbiotics are used/tested to reduce the inflammation or strengthen the IEB. The present review will resume these works, pointing out the stage of life, the duration, and the environmental conditions that should go along with microbiota or microbiota-derived treatments. [ABSTRACT FROM AUTHOR]

Microbial agents have promise for the bioremediation of Pb(II)-polluted environments and wastewater, the biodecontamination of foods, and the alleviation of toxicity in living organisms. The dairy bacterium Propionibacterium freudenreichii is poorly able to remove Pb(II) from aqueous solution at 25 ppm, ranging from 0 to 10% of initial concentration. Here, we report on an original strong enhancement of this activity (ranging from 75% to 93%, p < 0.01) following the addition of a polysorbate detergent (Tween® 80) during or either shortly after the growth of a P. freudenreichii culture. We evaluated the optimal Tween® 80 concentration for pretreatment conditions, documented the role of other detergents, and explored the possible mechanisms involved. Our results reveal a novel, environmentally friendly, low-cost pretreatment procedure for enhancing the selective removal of lead from water by probiotic-documented bacteria. [ABSTRACT FROM AUTHOR]

Bacteria used in the production of fermented food products have been investigated for their potential role as modulators of inflammation in gastrointestinal tract disorders such as inflammatory bowel diseases (IBD) that cause irreversible changes in the structure and function of gut tissues. Ulcerative colitis (UC) is the most prevalent IBD in the population of Western countries, and it is marked by symptoms such as weight loss, rectal bleeding, diarrhea, shortening of the colon, and destruction of the epithelial layer. The strain Propionibacterium freudenreichii CIRM-BIA 129 recently revealed promising immunomodulatory properties that greatly rely on surface-layer proteins (Slp), notably SlpB. We, thus, cloned the sequence encoding the SlpB protein into the pXIES-SEC expression and secretion vector, and expressed the propionibacterial protein in the lactic acid bacterium Lactococcus lactis NCDO 2118. The probiotic potential of L. lactis NCDO 2118 harboring pXIES-SEC: slpB (L. lactis -SlpB) was evaluated in a UC-mice model induced by Dextran Sulfate Sodium (DSS). During colitis induction, mice receiving L. lactis -SlpB exhibited reduced severity of colitis, with lower weight loss, lower disease activity index, limited shortening of the colon length, and reduced histopathological score, with significant differences, compared with the DSS group and the group treated with L. lactis NCDO 2118 wild-type strain. Moreover, L. lactis -SlpB administration increased the expression of genes encoding tight junction proteins zo-1 , cln -1, cln-5 , ocln , and muc-2 in the colon, increased IL-10 and TGF-β, and decreased IL-17, TNF-α, and IL-12 cytokines in the colon. Therefore, this work demonstrates that SlpB recombinant protein is able to increase the probiotic potential of the L. lactis strain to alleviate DSS-induced colitis in mice. This opens perspectives for the development of new approaches to enhance the probiotic potential of strains by the addition of SlpB protein. [ABSTRACT FROM AUTHOR]

Propionibacterium freudenreichii is a beneficial food-grade actinobacterium, widely implemented, and thus consumed, in various food products. As the main application, P. freudenreichii is used as a cheese-ripening starter, mostly in hard type cheeses. Indeed, during manufacture of "Swiss-type" cheeses (or opened-body cheeses), the technological process favors propionibacteria growth, as well as the corresponding propionic fermentation. This leads to the characteristic flavor of these cheeses, through the release of short chain fatty acids and through lipolysis, as well as to their specific texture. To fulfil this ripening, massive amounts of propionibacteria are industrially produced, dried and stored, prior to cheese making. Furthermore, P. freudenreichii is commercialized in various probiotic food supplements aiming at preserving intestinal health and comfort, in line with its ability to produce beneficial metabolites (short chain fatty acids, vitamins), as well as immunomodulatory compounds. Other industrial applications of P. freudenreichii include the production of food-grade vitamins of the B group, of trehalose, of conjugated linoleic acid, and of biopreservatives. For these different applications, maintaining survival and activity of propionibacteria during production, drying, storage and finally implementation, is crucial. More widely, maintaining live and active probiotic bacteria represents a challenge as the market for probiotic products increases. Probiotic bacteria are, for a bulk majority, freeze-dried, but spray drying is also more and more considered. Indeed, this process is both continuous and more cost-efficient, as it utilizes less energy compared to freeze-drying; on the other hand, it exposes bacteria to higher heat and oxidative stresses. Apart from process optimization and strain selection, it is possible to enhance the resistance of bacteria by taking advantage of their adaptation capacity. Indeed, P. freudenreichii stress tolerance can be boosted by different pretreatments applied before the drying step, thus considerably increasing its final survival. In particular, adaptation to hyperosmotic conditions improves stress tolerance, while the presence of osmoprotectants may mitigate this improvement. Thermal adaptation also modulates tolerance towards these technological challenges. The composition of the growth medium, including the ratio between the carbohydrates provided and the non-protein nitrogen, plays a key role in driving the accumulation of osmoprotectants. This, in turn, determines P. freudenreichii tolerance towards different stresses, and overall towards both freeze-drying and spray-drying. As an example, the accumulation of trehalose enhances its spray-drying survival, while the accumulation of glycine betaine enhances its freeze-drying survival. Growth of propionibacteria in hyperconcentrated whey was used to trigger multiple stress tolerance acquisition, underpinned by overexpression of key stress protein, accumulation of cytoplasmic storage compounds, and leading to enhanced spray-drying survival. A simplified process, from cultivation to atomization, was developed by using whey as a 2-in-1 medium in which propionibacteria were grown, protected and dried with minimal cell death. This innovative process was then subjected to scaling up at the industrial level. In this aim, a gentle multi-stage drying process offering mild drying conditions by coupling spray drying with belt drying, led to final probiotic survival close to 100% when stress tolerance acquisition was previously implemented. Such innovation opens new avenues for the efficient, cost-effective and sustainable development of new probiotic production technologies, as well as probiotic application in the context of food and feed. Key points: • Propionibacteria acquire multi-stress tolerance when grown in hyper-concentrated whey. • Spray drying of osmo-adapted probiotic bacteria is possible with limited cell death. • A two-in-one drying method is developed to grow and dry probiotic bacteria in the same matrix. [ABSTRACT FROM AUTHOR]

Inflammatory bowel diseases (IBDs) constitute disturbances of gastrointestinal tract that cause irreversible changes in the structure and function of tissues. Ulcerative colitis (UC), the most frequent IBD in the population, is characterized by prominent inflammation of the human colon. Functional foods containing probiotic bacteria have been studied as adjuvants to the treatment or prevention of IBDs. The selected probiotic strain Lactococcus lactis NCDO 2118 (L. lactis NCDO 2118) exhibits immunomodulatory effects, with promising results in UC mouse model induced by dextran sodium sulfate (DSS). Additionally, cheese is a dairy food that presents high nutritional value, besides being a good delivery system that can be used to improve survival and enhance the therapeutic effects of probiotic bacteria in the host. Therefore, this work investigated the probiotic therapeutic effects of an experimental Minas Frescal cheese containing L. lactis NCDO 2118 in DSS-induced colitis in mice. During colitis induction, mice that consumed the probiotic cheese exhibited reduced in the severity of colitis, with attenuated weight loss, lower disease activity index, limited shortening of the colon length, and reduced histopathological score. Moreover, probiotic cheese administration increased gene expression of tight junctions' proteins zo-1 , zo-2 , ocln , and cln-1 in the colon and increase IL-10 release in the spleen and lymph nodes. In this way, this work demonstrates that consumption of probiotic Minas Frescal cheese, containing L. lactis NCDO 2118, prevents the inflammatory process during DSS-induced colitis in mice, opening perspectives for the development of new probiotic functional foods for personalized nutrition in the context of IBD. [ABSTRACT FROM AUTHOR]

Propionibacterium 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 food transition 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 cow'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 enhanced in 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 carbohydrate transport and metabolism (including glycosyltransferase, PTS), indicating metabolic adaptation to 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. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

Propionibacterium freudenreichii is a beneficial Gram-positive bacterium, traditionally used as a cheese-ripening starter, and currently considered as an emerging probiotic. As an example, the P. freudenreichii CIRM-BIA 129 strain recently revealed promising immunomodulatory properties. Its consumption accordingly exerts healing effects in different animal models of colitis, suggesting a potent role in the context of inflammatory bowel diseases. This anti-inflammatory effect depends on surface layer proteins (SLPs). SLPs may be involved in key functions in probiotics, such as persistence within the gut, adhesion to host cells and mucus, or immunomodulation. Several SLPs coexist in P. freudenreichii CIRM-BIA 129 and mediate immunomodulation and adhesion. A mutant P. freudenreichii CIRM-BIA 129Δ slpB (CB129Δ slpB) strain was shown to exhibit decreased adhesion to intestinal epithelial cells. In the present study, we thoroughly analyzed the impact of this mutation on cellular properties. Firstly, we investigated alterations of surface properties in CB129Δ slpB. Surface extractable proteins, surface charges (ζ-potential) and surface hydrophobicity were affected by the mutation. Whole-cell proteomics, using high definition mass spectrometry, identified 1,288 quantifiable proteins in the wild-type strain, i.e., 53% of the theoretical proteome predicted according to P. freudenreichii CIRM-BIA 129 genome sequence. In the mutant strain, we detected 1,252 proteins, including 1,227 proteins in common with the wild-type strain. Comparative quantitative analysis revealed 97 proteins with significant differences between wild-type and mutant strains. These proteins are involved in various cellular process like signaling, metabolism, and DNA repair and replication. Finally, in silico analysis predicted that slpB gene is not part of an operon, thus not affecting the downstream genes after gene knockout. This study, in accordance with the various roles attributed in the literature to SLPs, revealed a pleiotropic effect of a single slpB mutation, in the probiotic P. freudenreichii. This suggests that SlpB may be at a central node of cellular processes and confirms that both nature and amount of SLPs, which are highly variable within the P. freudenreichii species, determine the probiotic abilities of strains. [ABSTRACT FROM AUTHOR]

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 favored by the ability of probiotics to adhere to the host cells. This property depends on key surface compounds, still poorly characterized in propionibacteria. 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. [ABSTRACT FROM AUTHOR]

Dairy propionibacteria are used as cheese ripening starters, as biopreservative and as beneficial additives, in the food industry. The main species, Propionibacterium freudenreichii, is known as GRAS (Generally Recognized As Safe, USA, FDA). In addition to another dairy species, Propionibacterium acidipropionici, they are included in QPS (Qualified Presumption of Safety) list. Additional to their well-known technological application, dairy propionibacteria increasingly attract attention for their promising probiotic properties. The purpose of this review is to summarize the probiotic characteristics of dairy propionibacteria reported by the updated literature. Indeed, they meet the selection criteria for probiotic bacteria, such as the ability to endure digestive stressing conditions and to adhere to intestinal epithelial cells. This is a prerequisite to bacterial persistence within the gut. The reported beneficial effects are ranked according to property's type: microbiota modulation, immunomodulation, and cancer modulation. The proposed molecular mechanisms are discussed. Dairy propionibacteria are described as producers of nutraceuticals and beneficial metabolites that are responsible for their versatile probiotic attributes include short chain fatty acids (SCFAs), conjugated fatty acids, surface proteins, and 1,4-dihydroxy-2-naphtoic acid (DHNA). These metabolites possess beneficial properties and their production depends on the strain and on the growth medium. The choice of the fermented food matrix may thus determine the probiotic properties of the ingested product. This review approaches dairy propionibacteria, with an interest in both technological abilities and probiotic attributes. [ABSTRACT FROM AUTHOR]

Dairy propionibacteria are actinomycetales found in various fermented food products. The main species, Propionibacterium freudenreichii, is generally recognized as safe and used both as probiotic and as cheese starter. Its probiotic efficacy tightly depends on its tolerance towards digestive stresses, which can be largely modulated by the ingested delivery vehicle. Indeed, tolerance of this bacterium is enhanced when it is consumed within a fermented dairy product, compared to a dried probiotic preparation. We investigated both stress tolerance and protein neosynthesis upon growth in i) chemically defined or ii) aqueous phase of Emmental cheeses. Although the same final population level was reached in both media, a slower growth and an enhanced survival of CIRM BIA 1 strain of P. freudenreichii subsp. shermanii was observed in Emmental juice, compared to chemically defined medium. This was accompanied by differences in substrates used and products released as well as overexpression of various early stress adaptation proteins in Emmental juice, compared to chemically defined medium, implied in protein folding, in aspartate catabolism, in biosynthesis of valine, leucine and isoleucine, in pyruvate metabolism in citrate cycle, in the propionate metabolism, as well as in oxidoreductases. All these changes led to a higher digestive stress tolerance after growth in Emmental juice. Mechanisms of stress adaptation were induced in this environment, in accordance with enhanced survival. This opens perspectives for the use of hard and semi-hard cheeses as delivery vehicle for probiotics with enhanced efficacy. [ABSTRACT FROM AUTHOR]

Background: Gastric cancer is one of the most common cancers in the world. The "economically developed countries" life style, including diet, constitutes a risk factor favoring this cancer. Diet modulation may lower digestive cancer incidence. Among promising food components, dairy propionibacteria were shown to trigger apoptosis of human colon cancer cells, via the release of short-chain fatty acids acetate and propionate. Methodology/Principal Findings: A fermented milk, exclusively fermented by P. freudenreichii, was recently designed. In this work, the pro-apoptotic potential of this new fermented milk was demonstrated on HGT-1 human gastric cancer cells. Fermented milk supernatant induced typical features of apoptosis including chromatin condensation, formation of apoptotic bodies, DNA laddering, cell cycle arrest and emergence of a subG1 population, phosphatidylserine exposure at the plasma membrane outer leaflet, reactive oxygen species accumulation, mitochondrial transmembrane potential disruption, caspase activation and cytochrome c release. Remarkably, this new fermented milk containing P. freudenreichii enhanced the cytotoxicity of camptothecin, a drug used in gastric cancer chemotherapy. Conclusions/Significance: Such new probiotic fermented milk may thus be useful as part of a preventive diet designed to prevent gastric cancer and/or as a food supplement to potentiate cancer therapeutic treatments. [ABSTRACT FROM AUTHOR]

Propionibacterium freudenreichii is used as a ripening culture in Swiss cheese manufacture. It grows when cheeses are ripened in a warm room (about 24°C). Cheeses with an acceptable eye formation level are transferred to a cold room (about 4°C), inducing a marked slowdown of propionic fermentation, but P. freudenreichii remains active in the cold. To investigate the P. freudenreichii strategies of adaptation and survival in the cold, we performed the first global gene expression profile for this species. The time-course transcriptomic response of P. freudenreichii CIRM-BIA1T strain was analyzed at five times of incubation, during growth at 30°C then for 9 days at 4°C, under conditions preventing nutrient starvation. Gene expression was also confirmed by RT-qPCR for 28 genes. In addition, proteomic experiments were carried out and the main metabolites were quantified. Microarray analysis revealed that 565 genes (25% of the protein-coding sequences of P. freudenreichii genome) were differentially expressed during transition from 30°C to 4°C (P<0.05 and ∣fold change∣>1). At 4°C, a general slowing down was observed for genes implicated in the cell machinery. On the contrary, P. freudenreichii CIRM-BIA1T strain over-expressed genes involved in lactate, alanine and serine conversion to pyruvate, in gluconeogenesis, and in glycogen synthesis. Interestingly, the expression of different genes involved in the formation of important cheese flavor compounds, remained unchanged at 4°C. This could explain the contribution of P. freudenreichii to cheese ripening even in the cold. In conclusion, P. freudenreichii remains metabolically active at 4°C and induces pathways to maintain its long-term survival. [ABSTRACT FROM AUTHOR]

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Copyright of Dairy Science & Technology (EDP Sciences) is the property of EDP Sciences and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Propionibacterium freudenreichii, a food-grade bacterium able to kill colon cancer cell lines in vitro by apoptosis, may exert an anticarcinogenic effect in vivo. To assess this hypothesis, we administered daily 2 × 1010 colony-forming units (CFU) of P. freudenreichii TL133 to human microbiota-associated (HMA) rats for 18 d. Either saline or 1,2-dimethylhydrazine (DMH) was also administered on days 13 and 17 and rats were killed on day 19. The levels of apoptosis and proliferation in the mid and distal colon were assessed by terminal deoxynucleotide transferase-mediated deoxyuridine triphosphate nick end labelling (TUNEL) and proliferating cell nuclear antigen (PCNA) immunolabelling, respectively. The administration of P. freudenreichii TL133 significantly increased the number of apoptotic cells in DMH-treated rats compared to those given DMH only (P < 0·01). Furthermore, propionibacteria were able to decrease the proliferation index in the distal colon after treatment with DMH (P < 0·01). Conversely, propionibacteria alone did not exert such an effect on healthy colonic mucosa. P. freudenreichii TL133 thus facilitated the elimination of damaged cells by apoptosis in the rat colon after genotoxic insult and may play a protective role against colon cancer. [ABSTRACT FROM PUBLISHER]

Proteomic analysis was performed to determine and differentiate the composition of the secretomes of Phanerochaete chrysosporium CIRM-BRFM41, a peroxidase hypersecretory strain grown under ligninolytic conditions and on softwood chips under biopulping conditions. Extracellular proteins from both cultures were analyzed by bidimensional gel electrophoresis and matrix-assisted laser desorption/ionization time-of-flight tandem mass spectrometry. A total of 37 spots were identified. The secretome in liquid synthetic medium comprised mainly peroxidases, while several wood-degrading enzymes and enzymes involved in fungal metabolism were detected in biopulping cultures on softwood. This prompted an analysis of the impact of secretome modulation in the presence of softwood chips. Biotreated wood was submitted to kraft cooking and chemical bleaching using chlorine dioxide. The fungal pre-treatment led to a significant increase in pulp yield and a better bleachability of the pulp. This bleachability improvement could be explained by the production of specific lignocellulose-degrading enzymes. [ABSTRACT FROM AUTHOR]

In addition to their use in cheese technology, dairy propionibacteria have been identified as potential probiotics. However, to have a probiotic effect, propionibacteria have to survive and to remain metabolically active in the digestive tract. The aim of the present study was to investigate the survival and metabolic activity of Propionibacterium freudenreichii within the gastrointestinal tract of human microbiota-associated rats, and its influence on intestinal microbiota composition and metabolism. Twenty-five dairy Propionibacterium strains were screened for their tolerance towards digestive stresses and their ability to produce propionate in a medium mimicking the content of the human colon. Three strains were selected and a daily dose of 2 × 1010 colony-forming units was fed to groups of human microbiota-associated rats for 20 d before microbiological, biochemical and molecular investigations being carried out. These strains all reached 8-log values per g faeces, showing their ability to survive in the gastrointestinal tract. Transcriptional activity within the intestine was demonstrated by the presence of P. freudenreichii-specific transcarboxylase mRNA. The probiotic efficacy of propionibacteria was yet species- and strain-dependent. Indeed, two of the strains, namely TL133 and TL1348, altered the faecal microbiota composition, TL133 also increasing the caecal concentration of acetate, propionate and butyrate, while the third strain, TL3, did not have similar effects. Such alterations may have an impact on gut health and will thus be taken into consideration for further in vivo investigations on probiotic potentialities of P. freudenreichii. [ABSTRACT FROM PUBLISHER]

The human probiotic Propionibacterium freudenreichii kills colorectal adenocarcinoma cells through apoptosis in vitro via its metabolites, the short chain fatty acids (SCFA), acetate and propionate. However, the precise mechanisms, the kinetics of cellular events and the impact of environmental factors such as pH remained to be specified. For the first time, this study demonstrates a major impact of a shift in extracellular pH on the mode of propionibacterial SCFA-induced cell death of HT-29 cells, in the pH range 5.5 to 7.5 prevailing within the colon. Propionibacterial SCFA triggered apoptosis in the pH range 6.0 to 7.5, a lethal process lasting more than 96 h. Indeed at pH 7.5, SCFA induced cell cycle arrest in the G2/M phase, followed by a sequence of cellular events characteristic of apoptosis. By contrast, at pH 5.5, the same SCFA triggered a more rapid and drastic lethal process in less than 24 h. This was characterised by sudden mitochondrial depolarisation, inner membrane permeabilisation, drastic depletion in ATP levels and ROS accumulation, suggesting death by necrosis. Thus, in digestive cancer prophylaxis, the observed pH-mediated switch between apoptosis and necrosis has to be taken into account in strategies involving SCFA production by propionibacteria to kill colon cancer cells. [ABSTRACT FROM AUTHOR]

Two membrane proteins from the avian pathogen Mycoplasma gallisepticum have been previously purified using a simple, efficient and non-denaturing method: a lipoprotein P67 (pMGA) and P52. In the current study, the lipid part of P67 was chemically analysed. The molecular structure of the lipoprotein-lipid component was determined to be S-glyceryl cysteine with two O-ester-linked acyl chains. Fatty acid analysis of the purified P67 indicated a heterogeneous composition: palmitic acid (16:0)>stearic acid (18:0)>oleic acid (18:1c)>myristic acid (14:0), with 16:0 as the major component. These findings, along with previously published results, support the conclusion that P67 is pMGA1.2, a true membrane-associated lipoprotein although not N-acylated. In contrast to P67, P52 is not a lipoprotein. Topological experiments using in situ treatment with proteases and growth inhibition in the presence of anti-P52 serum provided evidence of the surface exposition of the polypeptide. The N-terminal sequence of P52 was found to be similar to the dihydrolipoamide acetyltransferase from several mollicutes; this enzyme is a membrane-associated component of the pyruvate dehydrogenase complex. Immunoblotting techniques revealed that the surface antigens P52 and P67 were specific to the species M. gallisepticum and the closely related species M. imitans. No antigenic difference was revealed within these species with the anti-P52 serum, while anti-P67 serum confirmed the antigenic variability of P67. The potential of P52 and P67 as antigens in serological diagnosis tests or as candidates for anti-mycoplasma subunit vaccines is discussed. [ABSTRACT FROM AUTHOR]

Prohormone convertase 2 (PC2) is a subtilisin-like protease involved in the intracellular processing of prohormones and proneuropeptides. Like its substrates, it is synthesised as a prepropeptide which undergoes proteolysis during transit through the regulated secretory pathway. Previous studies have shown that aggregation and membrane association of proPC2 occurs in a calcium-dependent and pH-dependent manner and that the pro-region of PC2 may be involved in this process. These events may be involved in the sorting of proteins to the regulated secretory pathway. To investigate this further, we made a chimeric protein containing both the signal peptide and pro-region of PC2 and the N-terminal part of α1-antitrypsin, called pro2α1. PC2, α1-antitrypsin and pro2α1 were compared with regard to their membrane association and aggregation properties using, respectively, sucrose gradient centrifugation after expression in Xenopus oocytes, and an in vitro aggregation assay. The chimeric protein, pro2α1, underwent low-pH-dependent aggregation and membrane association similar to wild-type PC2. Membrane association occurred at pH 5.5 in the absence of calcium and at pH 6.0 in the presence of 10 mM calcium but not at pH 6.5 or 7.0. α1-antitrypsin, as expected of a constitutively secreted protein, did not aggregate at low pH, nor associate with membranes. Pro2α1 thus exhibits the membrane association and aggregation properties of PC2, confirming the role of the pro-region in these processes. A series of deletions were performed within the 84-residue propeptide in order to define the sequences involved. Deletion of amino acids 52-77 reduced aggregation but large deletions in the pro-region had only a minimal effect on membrane association. These data suggest that several regions within the pro-peptide are important in these events. [ABSTRACT FROM AUTHOR]

Toxic metals (such as lead, cadmium, and, to a lesser extent, aluminum) are detrimental to health when ingested in food or water or when inhaled. By interacting with heavy metals, gut and food-derived microbes can actively and/or passively modulate (by adsorption and/or sequestration) the bioavailability of these toxins inside the gut. This "intestinal bioremediation" involves the selection of safe microbes specifically able to immobilize metals. We used inductively coupled plasma mass spectrometry to investigate the in vitro ability of 225 bacteria to remove the potentially harmful trace elements lead, cadmium, and aluminum. Interspecies and intraspecies comparisons were performed among the Firmicutes (mostly lactic acid bacteria, including Lactobacillus spp., with some Lactococcus, Pediococcus, and Carnobacterium representatives), Actinobacteria, and Proteobacteria. The removal of a mixture of lead and cadmium was also investigated. Although the objective of the study was not to elucidate the mechanisms of heavy metal removal for each strain and each metal, we nevertheless identified promising candidate bacteria as probiotics for the intestinal bioremediation of Pb(II) and Cd(II). [ABSTRACT FROM AUTHOR]

Backgrounds and Aims. Inflammatory Bowel Diseases (IBD), including Ulcerative Colitis (UC), coincide with alterations in the gut microbiota. Consumption of immunomodulatory strains of probiotic bacteria may induce or prolong remission in UC patients. Fermented foods, including cheeses, constitute major vectors for bacteria consumption. New evidences revealed anti-inflammatory effects in selected strains of Propionibacterium freudenreichii. We thus hypothesized that consumption of a functional cheese, fermented by such a strain, may exert a positive effect on IBD. Methods. We investigated the impact of cheese fermented by P. freudenreichii on gut inflammation. We developed an experimental single-strain cheese solely fermented by a selected immunomodulatory strain of P. freudenreichii, CIRM-BIA 129. We moreover produced, in industrial conditions, an Emmental cheese using the same strain, in combination with Lactobacillus delbrueckii CNRZ327 and Streptococcus thermophilus LMD-9, as starters. Consumption of both cheeses was investigated with respect to prevention of Dextran Sodium Sulphate (DSS)-induced colitis in mice. Results. Consumption of the single-strain experimental cheese, or of the industrial Emmental, both fermented by P. freudenreichii CIRM-BIA 129, reduced severity of subsequent DSS-induced colitis, weight loss, disease activity index and histological score. Both treatments, in a preventive way, reduced small bowel Immunoglobulin A (IgA) secretion, restored occludin gene expression and prevented induction of Tumor Necrosis Factor α (TNFα), Interferon γ (IFNγ) and Interleukin-17 (IL-17). Conclusions. A combination of immunomodulatory strains of starter bacteria can be used to manufacture an anti-inflammatory cheese, as revealed in an animal model of colitis. This opens new perspectives for personalised nutrition in the context of IBD. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

Propionibacterium freudenreichii, a dairy starter, can reach a population of almost 109 propionibacteria per gram in Swiss-type cheese at the time of consumption. Also consumed as a probiotic, it displays strain-dependent anti-inflammatory properties mediated by surface proteins that induce IL-10 in leukocytes. We selected 23 strains with varied anti-inflammatory potentials in order to identify the protein(s) involved. After comparative genomic analysis, 12 of these strains were further analysed by surface proteomics, eight of them being further submitted to transcriptomics. The omics data were then correlated to the anti-inflammatory potential evaluated by IL-10 induction. This comparative omics strategy highlighted candidate genes that were further subjected to gene-inactivation validation. This validation confirmed the contribution of surface proteins, including SlpB and SlpE, two proteins with SLH domains known to mediate non-covalent anchorage to the cell-wall. Interestingly, HsdM3, predicted as cytoplasmic and involved in DNA modification, was shown to contribute to anti-inflammatory activity. Finally, we demonstrated that a single protein cannot explain the anti-inflammatory properties of a strain. These properties therefore result from different combinations of surface and cytoplasmic proteins, depending on the strain. Our enhanced understanding of the molecular bases for immunomodulation will enable the relevant screening for bacterial resources with anti-inflammatory properties. [ABSTRACT FROM AUTHOR]