Strains Of Propionibacterium Freudenreichii Research Articles

Fat matters: Fermented whole milk potentiates the anti-colitis effect of Propionibacterium freudenreichii

Inflammatory bowel diseases (IBD) constitute a growing concern in western countries. They coincide with gut microbiota dysbiosis, including a loss of immunomodulatory bacteria. Accordingly, probiotic products containing selected immunomodulatory bacterial strains mitigate IBD. Selected strains of Propionibacterium freudenreichii display promising modulatory properties and prevent colitis in animal models. Dairy matrices protect propionibacteria immunomodulatory surface antigens during digestive transit. However, the functional role of the dairy matrix components in such fermented dairy products remains unknown. In the present workthis study, P. freudenreichii CIRM-BIA129, a probiotic strain known for its anti-inflammatory properties, was used to ferment whole milk, skim milk or skim milk ultrafiltrate. The preventive potential of fermented products was tested in DSS-induced mice colitis, in comparison with their unfermented counterparts. P. freudenreichii-fermented milk prevented colitis. Dairy fat in the fermented product potentiated the anti-colitis effects of the probiotic. This work opens new perspectives for developing immunomodulatory functional fermented foods.

Bioprocess Strategies for Vitamin B12 Production by Microbial Fermentation and Its Market Applications.

Vitamin B12 is a widely used compound in the feed and food, healthcare and medical industries that can only be produced by fermentation because of the complexity of its chemical synthesis. For this reason, finding better producer strains and optimizing their bioprocesses have been the main focus of industrial producers over the last few decades. In this review, we initially provide a historical overview of vitamin B12 research and the main biosynthetic characteristics of the two microorganism families typically used for its industrial production: several strains of Propionibacterium freudenreichii and strains related to Pseudomonas denitrificans. Later, a complete summary of the current state of vitamin B12 industrial production as well as the main advances and challenges for improving it is detailed, with a special focus on bioprocess optimization, which aims not only to increase production but also sustainability. In addition, a comprehensive list of the most important and relevant patents for the present industrial strains is provided. Finally, the potential applications of vitamin B12 in different markets are discussed.

Probiotic drug “Propionovy” in the diets of broiler chickens

Currently, probiotics are an alternative to feed antibiotics in poultry farming. Probiotic cultures that are part of these drugs suppress the development of pathogenic and contain conditionally pathogenic bacteria at a safe level, as well as maintain a healthy microflora at a normal level. The mechanism of action of probiotics is studied by specialists from different countries. The purpose of the work was to study the effect of the probiotic drug “Propionovy” on the productive traits and physiological state of broiler chickens. Experimental studies have been carried out under the conditions of LLC “Kuzbassky broiler” in the Kemerovo region. The object of the study was broiler chickens of the cross Hubbard ISA F-15. Comprehensive studies have been conducted on the use of the probiotic drug “Propionovy”, which includes strains of Propionibacterium freudenreichii spp. in the diets of broiler chickens. The influence of various dosages of the studied drug on the productivity and quality of poultry meat, on morphological and biochemical parameters of blood has been studied. The optimal dose of introducing the drug “Propionovy” into the diets of broiler chickens, which is most effective for production from the economic point of view has been determined. It has been found according to the results of the production inspection that the inclusion of the probiotic drug “Propionovy” in the diets of broiler chickens contributed to the increase in the total gain in live weight in the experimental group by 3,76 % and the decrease in the prime-cost of production by 0,45 %, which allowed to obtain an additional profit of 2,83 rubles from 1 head. It has been found that the introduction of the probiotic drug “Propionovy” into the diets of broiler chickens of the cross Hubbard ISA F-15 has the positive effect on their productive traits and physiological state, and also allows us to get additional profit.

Extracellular Vesicles Produced by the Probiotic Propionibacterium freudenreichii CIRM-BIA 129 Mitigate Inflammation by Modulating the NF-κB Pathway.

Extracellular vesicles (EVs) are nanometric spherical structures involved in intercellular communication, whose production is considered to be a widespread phenomenon in living organisms. Bacterial EVs are associated with several processes that include survival, competition, pathogenesis, and immunomodulation. Among probiotic Gram-positive bacteria, some Propionibacterium freudenreichii strains exhibit anti-inflammatory activity, notably via surface proteins such as the surface-layer protein B (SlpB). We have hypothesized that, in addition to surface exposure and secretion of proteins, P. freudenreichii may produce EVs and thus export immunomodulatory proteins to interact with the host. In order to demonstrate their production in this species, EVs were purified from cell-free culture supernatants of the probiotic strain P. freudenreichii CIRM-BIA 129, and their physicochemical characterization, using transmission electron microscopy and nanoparticle tracking analysis (NTA), revealed shapes and sizes typical of EVs. Proteomic characterization showed that EVs contain a broad range of proteins, including immunomodulatory proteins such as SlpB. In silico protein-protein interaction predictions indicated that EV proteins could interact with host proteins, including the immunomodulatory transcription factor NF-κB. This potential interaction has a functional significance because EVs modulate inflammatory responses, as shown by IL-8 release and NF-κB activity, in HT-29 human intestinal epithelial cells. Indeed, EVs displayed an anti-inflammatory effect by modulating the NF-κB pathway; this was dependent on their concentration and on the proinflammatory inducer (LPS-specific). Moreover, while this anti-inflammatory effect partly depended on SlpB, it was not abolished by EV surface proteolysis, suggesting possible intracellular sites of action for EVs. This is the first report on identification of P. freudenreichii-derived EVs, alongside their physicochemical, biochemical and functional characterization. This study has enhanced our understanding of the mechanisms associated with the probiotic activity of P. freudenreichii and identified opportunities to employ bacterial-derived EVs for the development of bioactive products with therapeutic effects.

Beneficial Propionibacteria within a Probiotic Emmental Cheese: Impact on Dextran Sodium Sulphate-Induced Colitis in Mice.

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.

Research on the ability of propionic acid and vitamin B12 biosynthesis by Propionibacterium freudenreichii strain T82

The purpose of this study was to determine the potential for biosynthesis of propionic acid and vitamin B12 by Propionibacterium freudenreichii T82 in a medium containing various sources of carbon (glucose, fructose, and saccharose). These sugars are present in apple pomaces, which are the waste from the production of apple juice. Using statistical analysis design of experiments (DoE), the results allowed us to determine which sugars (carbon sources) exert the most beneficial influence on the biosynthesis of propionic acid and cobalamin. The highest production of propionic acid by the tested bacterial strain was obtained in a medium in which glucose accounted for at least 50% of the available carbon sources. Depending on the culture medium, the concentration of this metabolite ranged from 23 to 40 g/L. P. freudenreichii T82 produced the smallest amount of acid in medium in which the dominant nutrient source was saccharose. The results obtained indicated an inverse relationship between the amount of acid produced by the bacteria and vitamin B12 biosynthesis. Because of the high efficiency of propionic acid biosynthesis by P. freudenreichii T82, the prospect of using this strain to obtain propionate with the simultaneous disposal of waste materials (such as apple pomaces) which contain glucose and/or fructose is very promising.

Exopolysaccharides from co-cultures of Weissella confusa 11GU-1 and Propionibacterium freudenreichii JS15 act synergistically on wheat dough and bread texture

The storage of bread is limited by both physical (staling) and microbial (mainly fungal) spoilage. Exopolysaccharides (EPS) produced by lactic acid bacteria (LAB) and organic acids from propionibacteria (PAB) have been used to enhance texture and extend shelf-life of bakery products. In this study the functionality of EPS of Weissella confusa A3/2-1 (dextran), W. confusa F3/2-2 (dextran and levan), W. confusa 11GU-1 (dextran and ropy capsular polysaccharide) was evaluated in wheat bread. Two strains of Propionibacterium freudenreichii (Pf), shown to produce a heteropolysaccharide (Pf JS15) or a β-glucan (Pf DF30), were tested in single and mixed cultures with W. confusa (Wc). The EPS fermentates were prepared by batch fermentation of cereal- or malt-based medium using sucrose (Wc) or lactic acid (Pf) as carbon source. Incorporation of EPS from single culture fermentates and 1:1 Weissella–Propionibacterium fermentate mixtures revealed strong positive effects of dextran and ropy capsular polysaccharide produced by Wc 11GU-1 on bread staling retardation, with synergistic effects of EPS mixture from Wc 11GU-1 and Pf JS15. A co-fermentation of Wc 11GU-1 and Pf JS15 was developed to produce EPS together with antifungal organic acid mixture (acetate and propionate) in a single step process. The addition of 15% (w/w flour base) co-culture, yielding EPS, acetate and propionate concentrations of 1.5, 0.5 and 1g/kg dough, respectively, resulted in improved bread texture, increased loaf volume and decreased crumb firming during storage for 3days compared with control breads and breads supplemented with equivalent levels of chemical organic acids. Our data showed that EPS could compensate for the negative effects of chemical acetate and propionate in a concentration range exerting antifungal effects. The natural bioingredient produced by Wc 11GU-1 and Pf JS15 has potential for applications as antifungal, texture-building and anti-staling agent in breads, consistent with consumer demands for clean label products.

Mutations and genomic islands can explain the strain dependency of sugar utilization in 21 strains of Propionibacterium freudenreichii.

BackgroundPropionibacterium freudenreichii (PF) is an actinobacterium used in cheese technology and for its probiotic properties. PF is also extremely adaptable to several ecological niches and can grow on a variety of carbon and nitrogen sources. The aim of this work was to discover the genetic basis for strain-dependent traits related to its ability to use specific carbon sources. High-throughput sequencing technologies were ideal for this purpose as they have the potential to decipher genomic diversity at a moderate cost.Results21 strains of PF were sequenced and the genomes were assembled de novo. Scaffolds were ordered by comparison with the complete reference genome CIRM-BIA1, obtained previously using traditional Sanger sequencing. Automatic functional annotation and manual curation were performed. Each gene was attributed to either the core genome or an accessory genome. The ability of the 21 strains to degrade 50 different sugars was evaluated. Thirty-three sugars were degraded by none of the sequenced strains whereas eight sugars were degraded by all of them. The corresponding genes were present in the core genome. Lactose, melibiose and xylitol were only used by some strains. In this case, the presence/absence of genes responsible for carbon uptake and degradation correlated well with the phenotypes, with the exception of xylitol. Furthermore, the simultaneous presence of these genes was in line the metabolic pathways described previously in other species. We also considered the genetic origin (transduction, rearrangement) of the corresponding genomic islands. Ribose and gluconate were degraded to a greater or lesser extent (quantitative phenotype) by some strains. For these sugars, the phenotypes could not be explained by the presence/absence of a gene but correlated with the premature appearance of a stop codon interrupting protein synthesis and preventing the catabolism of corresponding carbon sources.ConclusionThese results illustrate (i) the power of correlation studies to discover the genetic basis of binary strain-dependent traits, and (ii) the plasticity of PF chromosomes, probably resulting from horizontal transfers, duplications, transpositions and an accumulation of mutations. Knowledge of the genetic basis of nitrogen and sugar degradation opens up new strategies for the screening of PF strain collections to enable optimum cheese starter, probiotic and white biotechnology applications.Electronic supplementary materialThe online version of this article (doi:10.1186/s12864-015-1467-7) contains supplementary material, which is available to authorized users.

Biodiversity of dairy Propionibacterium isolated from dairy farms in Minas Gerais, Brazil

Dairy propionibacteria are used as ripening cultures for the production of Swiss-type cheeses, and some strains have potential for use as probiotics. This study investigated the biodiversity of wild dairy Propionibacteria isolates in dairy farms that produce Swiss-type cheeses in Minas Gerais State, Brazil. RAPD and PFGE were used for molecular typing of strains and MLST was applied for phylogenetic analysis of strains of Propionibacterium freudenreichii. The results showed considerable genetic diversity of the wild dairy propionibacteria, since three of the main species were observed to be randomly distributed among the samples collected from different farms in different biotopes (raw milk, sillage, soil and pasture). Isolates from different farms showed distinct genetic profiles, suggesting that each location represented a specific niche. Furthermore, the STs identified for the strains of P. freudenreichii by MLST were not related to any specific origin. The environment of dairy farms and milk production proved to be a reservoir for Propionibacterium strains, which are important for future use as possible starter cultures or probiotics, as well as in the study of prevention of cheese defects.

Great interspecies and intraspecies diversity of dairy propionibacteria in the production of cheese aroma compounds

Flavor is an important sensory property of fermented food products, including cheese, and largely results from the production of aroma compounds by microorganisms. Propionibacterium freudenreichii is the most widely used species of dairy propionibacteria; it has been implicated in the production of a wide variety of aroma compounds through multiple metabolic pathways and is associated with the flavor of Swiss cheese. However, the ability of other dairy propionibacteria to produce aroma compounds has not been characterized. This study sought to elucidate the effect of interspecies and intraspecies diversity of dairy propionibacteria on the production of aroma compounds in a cheese context. A total of 76 strains of Propionibacterium freudenreichii, Propionibacterium jensenii, Propionibacterium thoenii, and Propionibacterium acidipropionici were grown for 15days in pure culture in a rich medium derived from cheese curd. In addition, one strain each of two phylogenetically related non-dairy propionibacteria, Propionibacterium cyclohexanicum and Propionibacterium microaerophilum were included. Aroma compounds were analyzed using headspace trap-gas chromatography-mass spectrometry (GC–MS). An analysis of variance performed on GC–MS data showed that the abundance of 36 out of the 45 aroma compounds detected showed significant differences between the cultures. A principal component analysis (PCA) was performed for these 36 compounds. The first two axes of the PCA, accounting for 60% of the variability between cultures, separated P. freudenreichii strains from P. acidipropionici strains and also differentiated P. freudenreichii strains from each other. P. freudenreichii strains were associated with greater concentrations of a variety of compounds, including free fatty acids from lipolysis, ethyl esters derived from these acids, and branched-chain acids and alcohols from amino acid catabolism. P. acidipropionici strains produced less of these compounds but more sulfur-containing compounds from methionine catabolism. Meanwhile, branched-chain aldehydes and benzaldehyde were positively associated with certain strains of P. jensenii and P. thoenii. Moreover, the production of compounds with a common origin was correlated. Compound abundance varied significantly by strain, with fold changes between strains of the same species as high as in the order of 500 for a single compound. This suggests that the diversity of dairy propionibacteria can be exploited to modulate the flavor of mild cheeses.

Influence of Propionibacteria Supplementation to Rations on Intake, Milk Yield, Composition and Plasma Metabolites of Lactating Buffalos During Early Lactation

Background: A probiotics (direct-fed microbials, DFM) and prebiotics (microbial growth promoters) are specific agents may have provide alternatives to chemical modifiers of manipulate the microbial ecosystem and fermentation characteristics in the rumen and intestinal tracts of livestock animals. The objective of this study was evaluating the effect of two levels propionibacteria strain (P169) on buffalo’s performance. Methods: Six lactating buffaloes in early lactation after four days from calving were divided to three treatments by using two blocks 3×3 complete switch–back design and three successive experimental periods. Each period consisted of 30 days. Animals were fed rations consisting of 70% Concentrate Feed Mixture (CFM) and 30% roughage [20% berseem (B) and 10.0% Rice Straw (RS)] on dry matter basis as control (1). Dietary treatments were: Control ration plus 2 g dairy ProP169® (low dose) (Propionibacterium freudenreichii strain P169) (2) and control ration plus 4 g dairy ProP169 (high dose)/head/day (3). ® Results: Buffaloes fed supplemental Propionibacteria had lower DMI kgG b.wt. (p 0.05) differed between all treatments. Also, blood plasma parameters were not affected by treatments groups. Conclusion: It could be concluded that supplementing rations with dairy Prop P169 might has a little effect on lactating buffalo’s performance.

The physiological state of Lactoccocus lactis and Propionibacterium freudenreichii strains in co-cultures induced by low temperature and osmotic stress

The bacterial physiological state, type of interactions and changes in metabolism ofLactococcus lactis and Propionibacterium freudenreichii strains in co-culture were studied in skimmed milk in response to osmotic [3% (w/v) NaCl] and low temperature (10°C) stress during long-term incubation. Changes in the integrity of cell membrane were examined by LIVE/DEAD® BacLight™ staining, and culture viability and bacterial interactions studies were performed with the use of the plate count technique. The profiles of volatile organic compounds were assessed by static headspace-gas chromatography. During the stationary growth phase, the number of LIVE cells with intact membranes was reduced in the presence of NaCl in comparison with control conditions, and a longer adaptation phase was observed at 10°C. The viability of starter cultures was high at ~108to 109 CFU/ml at the end of the experiment in all tested conditions. Our results point to the possibility of commensalisms interactions between lactic acid and propionic acid bacteria. Prolonged culture incubation contributed to the accumulation of acetoin, and it enhanced acetic acid and propionic acid synthesis during the stationary growth phase of propionic acid bacteria. Key words: Lactococcus lactis, Propionibacterium freudenreichii, viability, volatiles compounds, osmotic stress, low temperature stress.

Multifaceted attributes of dairy propionibacteria: a review

Dairy propionibacteria are Generally Recognized as Safe (GRAS) status microorganisms which have been traditionally used for the manufacture of Swiss type cheeses. In the last two decades various added features and functionalities have been discovered and developed from these bacteria. Propionibacteria are robust organisms with remarkable adaptability to technological and physiological stress conditions. Besides, they also display a multitude of health promoting properties like modulation of gut microbiota, improved gut physiology and immunomodulation suggesting their promising probiotic potential. Propionibacteria produce an interestingly wide range of functional biomolecules like B group vitamins, trehalose, conjugated linoleic acid, propionic acid, bacteriocins, bifidogenic factors etc. These bacteria are thus now being explored for designing novel functional foods as well as for industrial production of nutraceuticals. Growing interest in these bacteria is fueled by the first whole genome sequencing of a Propionibacterium freudenreichii strain providing a platform for better understanding of various pathways and further improvement in related process technologies.

The first dairy product exclusively fermented by Propionibacterium freudenreichii: A new vector to study probiotic potentialities in vivo

Dairy propionibacteria display probiotic properties which require high populations of live and metabolically active propionibacteria in the colon. In this context, the probiotic vector determines probiotic efficiency. Fermented dairy products protect propionibacteria against digestive stresses and generally contain a complex mixture of lactic and propionic acid bacteria. This does not allow the identification of dairy propionibacteria specific beneficial effects. The aim of this study was to develop a dairy product exclusively fermented by dairy propionibacteria. As they grow poorly in milk, we determined their nutritional requirements concerning carbon and nitrogen by supplementing milk ultrafiltrate (UF) with different concentrations of lactate and casein hydrolysate. Milk or UF supplemented with 50 mM lactate and 5 g L−1 casein hydrolysate allowed growth of all dairy propionibacteria studied. In these new fermented dairy products, dairy propionibacteria remained viable and stress-tolerant in vitro during minimum 15 days at 4 °C. The efficiency of milk fermented by the most tolerant Propionibacterium freudenreichii strain was evaluated in piglets. Viability and SCFA content in the colon evidenced survival and metabolic activity of P. freudenreichii. This work results in the design of a new food grade vector, which will allow preclinical and clinical trials.

PROPIONIBACTERIUM FREUDENREICHII STRAINS AS ANTIBACTERIAL AGENTS AT NEUTRAL PH AND THEIR PRODUCTION ON FOOD‐GRADE MEDIA FERMENTED BY SOME LACTOBACILLI

ABSTRACTPropionibacteria are food‐grade microorganisms that can suppress undesirable microbes in food. Profitability of their application depends on their effectiveness against food‐spoiling bacteria and on the accessibility of the protective product. In this study, the ability of Propionibacterium freudenreichii Pr4 to retard the growth of Bacillus subtilis, Bacillus cereus, Pseudomonas putida and Escherichia coli at neutral pH was proven. The efficiency of P. freudenreichii culture broth against all these bacteria, except E. coli, was similar to that of the chemical propionate, taken in the same amounts. Consequent cultivation with certain lactic acid bacteria was found to be a convenient way to achieve active growth of P. freudenreichii on food‐grade media. The pregrowth of Lactobacillus acidophilus 146 in skim milk and milk whey stimulated considerably the growth of P. freudenreichii Pr2 and the production of propionate. The final amount of P. freudenreichii Pr2 and L. acidophilus 146 viable cells in this case was 1.5 and 2 times more than when the bacteria grew separately. Other three Lactobacillus species tested did not provide stimulating effects. Sequential fermentation with Lactobacillus delbrueckii B‐3887 allowed to provide the growth of P. freudenreichii Pr4 in wheat flour medium and to get preparation that contains enough propionate for antirope bread protection.PRACTICAL APPLICATIONSNow we suggest a solid basis for the use of investigated strains of Propionibacterium freudenreichii in food bioprotection; first of all (directly due to our results) in bread manufacturing practice. They can be used as growth inhibitors of Bacillus subtilis, Escherichia coli and some Pseudomonas species not only in acidic but also in pH‐neutral conditions. The two‐stage cultivation of specially selected pairs of lactobacilli and propionibacteria is an approach for obtaining the propionibacteria‐containing preparations in economic and convenient way. The preparations can be applied for antibacterial protection of some other foods: dairy products, surfaces of cheeses, butter, sausages, etc.

Effects of addition of bacterial inoculants to the diets of lactating dairy cows on feed intake, milk yield, and milk composition

Forty-eight lactating Holstein cows (38 multiparous, 10 primiparous) were used in a 10-wk randomized block trial to determine the effect of 2 supplemental bacterial inoculants on DMI, milk yield, and milk composition. Experimental diets were top dressed with ground corn plus one of 2 bacterial inoculant mixtures. Treatments were control (no bacterial inoculant); 2 × 109 cfu/d of Propionibac-terium freudenreichii strain NP24 plus 1 × 109 cfu/d of Lactobacillus acidophilus strain NP51 (B1); and 2 × 109 cfu/d of P. freudenreichii strain NP24 plus 5 × 108 cfu/d each of L. acidophilus strain NP51 and strain NP45 (B2). Cows fed bacterial inoculants had greater yields of milk fat (P = 0.02), FCM (P = 0.007), and energy-corrected milk (P = 0.03) than did controls. Efficiency of milk production (defined as energy-corrected milk yield per unit of DMI) was greater (P = 0.03) for cows fed bacterial inoculants compared with control cows. There was no effect of treatment on DMI, milk fat percentage, or milk protein yield or percentage. There was a tendency (P = 0.08) for increased milk yield with inoculant supplementation compared with the control treatment. Serum urea N concentrations were greater for control cows compared with those receiving inoculants, and there was a tendency for B2 cows to have greater serum urea N concentrations compared with B1 cows. Results of this trial suggest that supplemental bacterial inoculants containing P. freudenreichii and L. acidophilus can improve milk yield and apparent efficiency of nutrient utilization.

In vitro and in vivo gastrointestinal survival, antibiotic susceptibility and genetic identification of Propionibacterium freudenreichii ssp. shermanii JS

The in vitro tolerance of Propionibacterium freudenreichii strains to low pH and high bile concentration, their antibiotic susceptibility and enzyme activity profiles were assessed. The gastrointestinal survival of the P. freudenreichii ssp. shermanii JS strain was also examined when administered to healthy adult volunteers in whey-based fruit juice. For tracking the strain JS in faecal samples, a genetic identification method for the strain was developed. No loss of viability was observed in any of the P. freudenreichii strains during the three-hour treatment at pH 4 or 3, whereas at pH 2 the cell counts of all strains decreased remarkably. Bile salt at 0.5% concentration had no apparent effect on the viability of the strains. No atypical antibiotic resistance was detected. P. freudenreichii ssp. shermanii JS strain was recovered in high numbers in faecal samples during the consumption of whey-based fruit juice enriched with the JS strain, and with few exceptions, it was not detected after the 2-week follow-up period.

Characterization of Growth and Metabolism of Commercial Strains of Propionic Acid Bacteria by Pressure Measurement

AbstractDairy propionibacteria are essential starters for Emmental cheese manufacture. The behavior of three commercial strains of Propionibacterium freudenreichii subsp. shermanii (P.f. 1, P.f. 2 and P.f 3) were studied in a liquid medium under air and N2 atmosphere using an on‐line pressure measurement technique. Growth kinetics and metabolite production were characterized under conditions usually reported as “optimal conditions” (pH 6.5, NaCl 0 %, temperature 30 °C) and also evaluated under “stressful conditions” (pH 5.2, NaCl 2 %, temperature 20 °C) simulating the cheese ripening conditions. In both cases, the effects of oxygen on growth were strain‐dependent. Under “stressful conditions”, two of the three strains were inhibited by oxygen under conditions of air atmosphere, while all three strains grew under conditions of N2 atmosphere. In the latter case, the duration of the lag phase and the maximum rate of pressure variation were significantly different, however, no significant differences were found between the strains with regard to the total fermentation time. Under “optimal conditions” metabolite production was strain‐dependent. In an air atmosphere, all strains produced more acetate and CO2 and less propionate than in a nitrogen atmosphere.

Autolysis of propionibacteria: Detection of autolytic enzymes by renaturing SDS-PAGE and additional buffer studies

Five strains of propionibacteria with 70–90% autolysis in sodium lactate broth (SLB) were studied by renaturing sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). Several lytic bands ranging in size between 25 and 143 kDa were detected by using propionibacteria cells or cell walls as substrate in the gel. Four Propionibacterium freudenreichii strains showed similar autolytic-enzyme profiles, consisting of two autolytic bands, one with molecular mass 162 kDa and one in the range 123–143 kDa. However, the Propionibacterium acidipropionici strain showed a completely different profile, consisting of 8 autolytic bands with molecular masses of 122, 97, 71, 55, 43, 39, 31, and 25 kDa. Lytic enzymes from P. freudenreichii INF-α, P. freudenreichii ISU P-59, P. freudenreichii ISU P-24, and P. freudenreichii ISU P-50 showed lytic activity against cells from all these four strains, but not against P. acidipropionici ATCC 4965. However, P. acidipropionici ATCC 4965 autolysed only its own cells. Effects of pH, temperature, and ions on autolytic activity were tested by renaturing SDS-PAGE and in buffer systems. Results from the SDS-PAGE electrophoresis showed optimal autolytic activity of P. acidipropionici ATCC 4965 at 37 °C and in the pH range 7 to 8.5 and of P. freudenreichii ISU P-59 at 20 °C and in the pH range 5 to 7. The autolytic activity of P. acidipropionici ATCC 4965 was extremely heat stable (100 °C, 2 h), in contrast to the lytic activity of P. freudenreichii ISU P-59, which was heat labile. The autolytic activities of P. acidipropionici ATCC 4965 were inhibited by divalent cations, however, the lytic activities of P. freudenreichii ISU P-59 were activated by Mn 2+, Ca 2+, and Co 2+. In buffer, optimum autolysis of P. acidipropionici ATCC 4965 was observed at pH 8.5 and at 40 °C. P. freudenreichii ISU P-59 showed optimum autolysis in buffer at pH 7.5 and at 30 °C.

Effect of yeast and bacterial adjuncts on the CLA content and flavour of a washed-curd, dry-salted cheese

Three strains of Propionibacterium freudenreichii ssp. shermanii that converted free linoleic acid to conjugated linoleic acid (CLA) in laboratory media were used as adjunct strains, together with strains of Geotrichum candidum and Yarrowia lipolytica, to make a dry-salted, washed-curd cheese. Lactobacillus fermentum was included to produce ethanol (from lactose), a potential substrate for ethyl ester synthesis, while Lactobacillus rhamnosus was used to control the adventitious non-starter lactic acid bacteria population. The total (esterified plus free) level of CLA was similar in the control and experimental cheeses and remained unchanged over 4 months of ripening. Addition of linoleic acid-rich safflower oil to the cheese curd increased the concentration of free linoleic acid generated in the cheese but the CLA content did not change. Free linoleic acid was released by the yeast lipase(s) but there was no conversion to CLA. High concentrations of ethyl esters were produced in the cheeses made with added yeast, giving a fruity flavour.