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The primary objective of this study was to characterize lactic acid bacteria (LAB) strains derived from sourdough for possible utilization as functional starters to produce sourdough and various cereal-based fermented foods. A total of 350 autochthonous LAB strains were isolated from 65 Type I sourdough samples and characterized using six random amplified polymorphic DNA (RAPD) primers at intra- and interspecific levels. Species identification of selected strains representing distinct clusters from RAPD analysis was performed based on the 16S rRNA region. The LAB strains were identified as Companilactobacillus crustorum (n = 135), Levilactobacillus brevis (n = 125), Latilactobacillus curvatus (n = 40), Companilactobacillus paralimentarius (n = 32), and Lactiplantibacillus plantarum (n = 18). A total of 66 LAB strains were selected for technological characterization along with two commercial strains. The characterization involved acidity development, EPS production potential, leavening activity, and growth abilities under harsh conditions. Principle component analysis (PCA) identified 2 Lp. plantarum and 14 Lev. brevis strains as the most relevant technologically. Among them, Lp. plantarum L35.1 and Lev. brevis L37.1 were resistant to tetracycline. Evaluation of probiotic characteristics (survival in pH 2.5 and bile presence, auto aggregation capacity, hydrophobic activity, antioxidant activity, antimicrobial activity) by PCA identified four strains with relevance to Lactobacillus rhamnosus GG (LGG), which were further selected for in vitro digestion assays. Lactiplantibacillus plantarum L7.8, Lev. brevis L55.1, and L62.2 demonstrated similar viability indices to LGG, along with increased auto aggregation capacity and antioxidant activity. These strains are promising as candidate starters for producing sourdough and sourdough-related fermented food products. • Sourdough lactic acid bacteria were characterized based on their functionality. • RAPD markers revealed significant intraspecific genetic diversity. • The technological traits of genetically diverse strains were evaluated. • Technologically prominent isolates were further screened for probiotic properties. • Three strains emerged as functional starters. [ABSTRACT FROM AUTHOR]

Abstract In order to encourage Western populations to increase their consumption of vegetables, we suggest turning legumes into novel, healthy foods by applying an old, previously widespread method of food preservation: fermentation. In the present study, a total of 55 strains from different microbial species (isolated from cheese or plants) were investigated for their ability to: (i) grow on a emulsion containing 100% pea proteins and no carbohydrates or on a 50:50 pea:milk protein emulsion containing lactose, (ii) increase aroma quality and reduce sensory off-flavors; and (iii) compete against endogenous micro organisms. The presence of carbohydrates in the mixed pea:milk emulsion markedly influenced the fermentation by strongly reducing the pH through lactic fermentation, whereas the absence of carbohydrates in the pea emulsion promoted alkaline or neutral fermentation. Lactic acid bacteria assigned to Lactobacillus plantarum , Lactobacillus rhamnosus , Lactococcus lactis and Lactobacillus casei species grew well in both the pea and pea:milk emulsions. Most of the fungal strains tested (particularly those belonging to the Mucor and Geotrichum genera) were also able to grow on both emulsions. Although most Actinobacteria and Proteobacteria did not compete with endogenous microbiota (Bacillus), some species such as Hafnia alvei , Acinetobacter johnsonii and Glutamicibacter arilaitensis grew strongly and appeared to restrict the development of the endogenous microbiota when the pea emulsion was inoculated with a combination of three to nine strains. In the mixed emulsions, lactic fermentation inhibited Actinobacteria and Proteobacteria (e.g. Brevibacterium casei , Corynebacterium casei , Staphylococcus lentus) to the greatest extent but also inhibited Bacillus (e.g. Bacillus subtilis and Bacillus licheniformis). Overall, this procedure enabled us to select two microbial consortia able to colonize pea-based products and positively influence the release of volatile compounds by generating a roasted/grilled aroma for the 100% pea emulsion, and a fruity, lactic aroma for the 50:50 pea:milk emulsion. Moreover, the fermentation in the pea-based emulsions reduced the level of hexanal, which otherwise leads to an undesired green pea aroma. Our present results show how the assembly of multiple microbial cultures can help to develop an innovative food product. Highlights • 55 microbial strains from dairy and vegetables were investigated for their ability to ferment pea proteins. • Most of fungi and Firmicutes were suitable to ferment pea proteins emulsions, even without carbohydrates. • Fermentation of pea proteins by relevant microbial combinations generated noteworthy odors. [ABSTRACT FROM AUTHOR]

Abstract This study aimed to evaluate the influence of passion fruit by-product (PFBP) and fructooligosaccharides (FOS) on the viability of Streptococcus thermophilus TH-4 and Lactobacillus rhamnosus LGG in folate bio-enriched fermented soy products and their effect on probiotic survival and folate bio-accessibility under in vitro simulated gastrointestinal conditions during storage of the products at 4 °C for up to 28 days (at days 1, 14, and 28). Kinetic parameters and folate contents before and after fermentation were also evaluated. Four different bio-enriched soy products in which the two microorganisms were used in co-cultures were studied and PFBP and/or FOS were added at 1 g/100 g, except for the control product. No differences (p < 0.05) between the fermented soy products (FSP) were observed for the maximum acidification rate (V max) and the time to reach the V max (T max) or pH 5.5 (T f), indicating that the use of PFBP and/or FOS did not affect the fermentation kinetic parameters. Only Lb. rhamnosus LGG retained the desired viability (>8 log CFU/mL) during storage, whereas St. thermophilus TH-4 populations decreased by day 14 reaching counts between 6.4 and 5.5 log CFU/mL by day 28. The folate content of all FSP increased after fermentation and the simultaneous presence of PFBP and FOS stimulated the co-culture to increase folate production. Folate content in all FSP decreased during storage. Lb. rhamnosus LGG was recovered at the end of the simulated digestion, but PFBP and/or FOS did not affect recovery. The folate content increased during the gastrointestinal assay for all FSP, especially for FSP without supplementation, suggesting an in vitro increase of folate bio-accessibility. Therefore, the bio-enriched probiotic FSP presented a great potential as an innovative functional food by delivering probiotic microorganisms and providing 14% of the recommended daily folate intake. The folate content of the FSP might be increased during gastrointestinal stress conditions, which could contribute to increase the folate bio-accessibility in the gut. Highlights • Lactobacillus rhamnosus LGG was viable during the storage period of FSP. • Folate bio-enriched fermented soy products (FSP) were submitted to in vitro GI stress. • LGG survived the in vitro simulated gastrointestinal conditions. • Folate bio-accessibility increased during the simulated digestion of all FSP. • FSP with passion fruit by-product and FOS may represent 14% of the folate RDI. [ABSTRACT FROM AUTHOR]

In Swiss-type cheeses, characteristic nut-like and sweet flavor develops during the cheese ripening due to the metabolic activities of cheese microbiota. Temperature changes during warm and cold room ripening, and duration of ripening can significantly change the gene expression of the cheese microbiota, which can affect the flavor formation. In this study, a metagenomic and metatranscriptomic analysis of Swiss-type Maasdam cheese was performed on samples obtained during ripening in the warm and cold rooms. We reconstructed four different bacterial genomes ( Lactococcus lactis , Lactobacillus rhamnosus , Lactobacillus helveticus , and Propionibacterium freudenreichii subsp. shermanii strain JS) from the Maasdam cheese to near completeness. Based on the DNA and RNA mean coverage, Lc. lactis strongly dominated (~80–90%) within the cheese microbial community. Genome annotation showed the potential for the presence of several flavor forming pathways in these species, such as production of methanethiol, free fatty acids, acetoin, diacetyl, acetate, ethanol, and propionate. Using the metatranscriptomic data, we showed that, with the exception of Lc. lactis , the central metabolism of the microbiota was downregulated during cold room ripening suggesting that fewer flavor compounds such as acetoin and propionate were produced. In contrast, Lc. lactis genes related to the central metabolism, including the vitamin biosynthesis and homolactic fermentation, were upregulated during cold room ripening. [ABSTRACT FROM AUTHOR]

Diacetyl and the closely related compound acetoin impart desirable buttery flavour and odour to many foods including cheese and are generated through the metabolism of citrate by lactic acid bacteria (LAB). To increase the levels of these compounds, adjunct cultures capable of producing them can be added to cheese fermentations. In this study, we compared the diacetyl and acetoin producing abilities of 13 Lactobacillus rhamnosus strains from cheese sources. Diacetyl and acetoin production was found to be a common feature of Lb. rhamnosus grown in milk, with 12 strains producing these compounds. Whole genome sequencing of four strains revealed that genes encoding the citrate metabolising pathway present in other LAB are conserved in Lb. rhamnosus . One strain was, however, totally defective in diacetyl and acetoin production. This was likely due to an inability to produce the diacetyl/acetoin precursor compound acetolactate resulting from a frameshift mutation in the acetolactate synthase ( als ) gene. Complementation of this defective strain with a complete als gene from a diacetyl producing strain restored production of diacetyl and acetoin to levels equivalent to naturally high producing strains. Introduction of the same als -containing plasmid into the probiotic Lb. rhamnosus strain GG also increased diacetyl and acetoin levels. In model cheesemaking experiments, the als -complemented strain produced very high levels of diacetyl and acetoin over 35 days of ripening. These findings identify the genetic basis for natural variation in production of a key cheese flavour compound in Lb. rhamnosus strains. [ABSTRACT FROM AUTHOR]

Mutandabota is a dairy product consumed as a major source of proteins and micronutrients in Southern Africa. In this study the microbial safety of traditional and a variant of mutandabota fermented with the probiotic Lactobacillus rhamnosus yoba (yoba mutandabota ) was investigated by challenging the products with five important food pathogens: Listeria monocytogenes , Salmonella spp., Campylobacter jejuni , Escherichia coli O157:H7 and Bacillus cereus . Pasteurized full-fat cow's milk was used for producing traditional and yoba mutandabota , and was inoculated with a cocktail of strains of the pathogens at an inoculum level of 5.5 log cfu/mL. Survival of the pathogens was monitored over a potential consumption time of 24 h for traditional mutandabota , and over 24 h of fermentation followed by 24 h of potential consumption time for yoba mutandabota . In traditional mutandabota (pH 3.4 ± 0.1) no viable cells of B. cereus and C. jejuni were detected 3 h after inoculation, while L. monocytogenes , E. coli O157:H7 and Salmonella spp. significantly declined (P < 0.05), but could still be detected (< 3.5 log inactivation) at the end of the potential consumption time. This indicated that consumption of traditional mutandabota exposes consumers to the risk of food-borne microbial infections. In yoba mutandabota , L. rhamnosus yoba grew from 5.5 ± 0.1 log cfu/mL to 9.1 ± 0.4 log cfu/mL in the presence of pathogens. The pH of yoba mutandabota dropped from 4.2 ± 0.1 to 3.3 ± 0.1 after 24 h of fermentation, mainly due to organic acids produced during fermentation. Only Salmonella spp. was able to grow in yoba mutandabota during the first 9 h of fermentation, but then decreased in viable plate count. None of the tested pathogens were detected (> 3.5 log inactivation) after 3 h into potential consumption time of yoba mutandabota . Inactivation of pathogens in mutandabota is of public health significance because food-borne pathogens endanger public health upon consumption of contaminated food, especially in Southern Africa where there are many vulnerable consumers of mutandabota such as children, elderly and immuno-compromised people with HIV/AIDS. The findings of this study demonstrate that mutandabota fermented with L. rhamnosus yoba has antimicrobial properties against the tested pathogens and it is safer compared to the traditional mutandabota . [ABSTRACT FROM AUTHOR]

Thirty-six microorganisms (twenty-one bacteria, twelve yeasts and three fungi) were isolated from surface-ripened cheeses and subjected to in vitro digestive stress. The approach mimicked gastric and/or duodenal digestion. Lactobacillus rhamnosus GG, Escherichia coli Nissle 1917 and Saccharomyces boulardii were used as reference strains. We studied the microorganisms grown separately in culture medium and then included (or not) in a rennet gel. The microorganisms' immunomodulatory abilities were also assessed by profiling cytokine induction in human peripheral blood mononuclear cells (PBMCs). The loss of viability was less than 1 log CFU/mL for yeasts under all conditions. In contrast, Gram-negative bacteria survived gastric and/or duodenal stress well but most of the Gram-positive bacteria were more sensitive (especially to gastric stress). Inclusion of sensitive Gram-positive bacteria in rennet gel dramatically improved gastric survival, when compared with a non-included cultured (with a 4 log CFU/mL change in survival). However, the rennet gel did not protect the bacteria against duodenal stress. The PBMC cytokine assay tests showed that the response to yeasts was usually anti-inflammatory, whereas the response to bacteria varied from one strain to another. [ABSTRACT FROM AUTHOR]

The human-derived potential probiotic strain Lactobacillus rhamnosus CTC1679 was used as a starter culture in reduced fat and sodium low-acid fermented sausages (fuets) to assess its ability to survive through the gastrointestinal tract (GIT) in a human intervention study consisting of 5 healthy volunteers who consumed 25 g fuet a day for 21 days. Faecal samples were analysed during and after consumption. L. rhamnosus CTC1679 produced a transient colonisation of the human GIT and persisted during the ingestion period of fuet containing L. rhamnosus CTC1679 at levels ca. 8 log CFU/g. After 3 days of non-consumption, the strain was still recovered in the faeces of all the volunteers. To evaluate the safety of the nutritionally enhanced manufactured fuets, a challenge test was designed in a separately manufactured batch. L. rhamnosus CTC1679 was able to grow, survive and dominate (levels ca. 108 CFU/g) the endogenous lactic acid bacteria (LAB), prevented the growth of Listeria monocytogenes throughout the whole ripening process of the fuets and eliminated Salmonella. After 35 days of storage at 4 °C, L. monocytogenes was not detected, achieving absence in 25 g of the product. The application of high hydrostatic pressure (HHP) treatment (600 MPa for 5 min) at the end of ripening (day 14) produced an immediate reduction of L. monocytogenes to levels < 1 log CFU/g. After 35 days of storage at 4 °C the pathogen was not detected. Thus, the strain L. rhamnosus CTC1679 is a suitable starter culture for producing safe potentially probiotic fermented sausages. [ABSTRACT FROM AUTHOR]

Abstract: The aim of this work was to study the Lactobacillus spp. intra- and inter- species diversity in a Piedmont hard cheese made of raw milk without thermal treatment and without addition of industrial starter, and to perform a first screening for potential functional properties. A total of 586 isolates were collected during the cheese production and identified by means of molecular methods: three hundred and four were identified as Lactobacillus rhamnosus, two hundred and forty as Lactobacillus helveticus, twenty six as Lactobacillus fermentum, eleven as Lactobacillus delbrueckii, three as Lactobacillus pontis, and two as Lactobacillus gasseri and Lactobacillus reuteri, respectively. A high genetic heterogeneity was detected by using the repetitive bacterial DNA element fingerprinting (rep-PCR) with the use of (GTG)5 primer resulting in eight clusters of L. helveticus and sixteen clusters in the case of L. rhamnosus. Most of isolates showed a high auto-aggregation property, low hydrophobicity values, and a general low survival to simulated digestion process. However, sixteen isolates showed promising functional characteristics. [Copyright &y& Elsevier]

Abstract: The production of viable functional probiotics presupposes stability of strain features in the final product. We evaluated the impact of acquisition of heat-tolerance and subsequent freeze-drying on the adhesion properties of Lactobacillus rhamnosus GG, Lactobacillus casei Shirota, Bifidobacterium lactis Bb-12 and Bifidobacterium animalis IF20/1 and on their ability to inhibit the adhesion of pathogens in a mucus model. Both fresh and freeze-dried cultures were evaluated. Significant differences were observed between fresh, freeze dried, fresh heat-tolerant and freeze dried heat-tolerant strains, especially in the ability of the freeze dried probiotics to exclude, displace or outcompete pathogens. Based on our study characterizing probiotic properties such as adhesion and competitive exclusion, it seems possible to adapt probiotics to processing stresses, such as heat, without significantly changing the probiotic properties of the strains assessed. This may provide new options for future probiotic production technology. However, our results also emphasize that the properties of the stress-adapted strains, as well as the effect of the production processes should always be assessed as these are strain-specific. [Copyright &y& Elsevier]

Abstract: The objective of this study was to develop a new chestnut-based puree, in order to seasonally adjust the offer and use the surplus of undersized production, providing, at the same time, a response to the growing demand for healthy and environmentally friendly products. Broken dried chestnuts have been employed to prepare purees to be fermented with six different strains of Lactobacillus (Lb.) rhamnosus and Lactobacillus casei. The fermented purees were characterized by a technological and sensorial point of view, while the employed strains were tested for their probiotic potential. Conventional in vitro tests have indicated the six lactobacilli strains as promising probiotic candidates; moreover, being the strains able to grow and to survive in chestnut puree at a population level higher than 8 log10 CFU/mL along 40days of storage at 4°C, the bases for the production of a new food, lactose-free and with reduced fat content, have been laid. [Copyright &y& Elsevier]

Abstract: The popularity of functional foods continues to increase as consumers desire flavorful foods that will fulfil their health needs. Among these foods, probiotics may exert positive effects on the composition of gut microbiota and overall health. However, in order to be beneficial, the bacterial cultures have to remain live and active at the time of consumption. The aim of this study was to develop new probiotic food products, such as seasoned cheeses, salami, chocolate and ice-cream with a final probiotic concentration of approximately 109 CFU/daily dose of Lactobacillus rhamnosus IMC 501® and Lactobacillus paracasei IMC 502® mixed 1:1 (SYNBIO®). The survival and viability of probiotics were determined during the foods shelf-life. The values of viable probiotic bacteria of all dairy and non-dairy foods were between 107 and 109 CFU/g of food at the end of the shelf-life and for some of them the values were maintained even after the expiry date. Based on the results of the current study, all the dairy (“Caciotta” cheese, “Pecorino” cheese, “Büscion” Swiss cheese and “Fiordilatte” ice-cream) and non-dairy (“Ciauscolo” salami, Larded salami, Swiss small salami, milk chocolate, dark chocolate, organic jam and chocolate mousse) food products studied would be excellent vehicles to deliver the probiotic health effects because of the high viability of probiotics during the shelf-life of foods and in some cases even after their expiry date. [Copyright &y& Elsevier]

Abstract: This study investigated the effect of microencapsulation on the survival of Lactobacillus rhamnosus GG and Lactobacillus acidophilus NCFM and their acidification in orange juice at 25°C for nine days and at 4°C over thirty five days of storage. Alginate micro beads (10–40μm) containing the probiotics were produced by a novel dual aerosol method of alginate and CaCl2 cross linking solution. Unencapsulated L. rhamnosus GG was found to have excellent survivability in orange juice at both temperatures. However unencapsulated L. acidophilus NCFM showed significant reduction in viability. Encapsulation of these two bacteria did not significantly enhance survivability but did reduce acidification at 25°C and 4°C. In agreement with this, encapsulation of L. rhamnosus GG also reduced acidification in pear and peach fruit-based foods at 25°C, however at 4°C difference in pH was insignificant between free and encapsulated cells. In conclusion, L. rhamnosus GG showed excellent survival in orange juice and microencapsulation has potential in reducing acidification and possible negative sensory effects of probiotics in orange juice and other fruit-based products. [Copyright &y& Elsevier]

Abstract: Parmigiano Reggiano (PR) is an Italian cooked, long-ripened cheese made with unheated cow''s milk and natural whey starter. The microflora is involved in the manufacturing of this cheese, arising from the natural whey starter, the raw milk and the environment. Molecular studies have shown that mesophilic non-starter lactic acid bacteria (NSLAB) are the dominant microflora present during the ripening of PR. In this study, a characterisation of Lactobacillus rhamnosus isolated from a single PR manufacturing and ripening process is reported, using a combination of genotypic fingerprinting techniques (RAPD-PCR and REP-PCR). The intraspecies heterogeneity evidenced for 66 strains is correlated to their abilities to adapt to specific environmental and technological conditions. The detection of biotypes that correlate with specific moments in cheese ripening or differential development throughout this process suggests that these strains may have specific roles closely linked to their peculiar technological properties. [Copyright &y& Elsevier]

Abstract: The immune stimulation induced by Lactobacillus rhamnosus CRL1505 (Lr05) and L. rhamnosus CRL1506 (Lr06) on the resistance to infection with an intestinal pathogen (Salmonella typhimurium) and a respiratory pathogen (Streptococcus pneumoniae) was studied in swiss-albine mice experimental models. The cytokine profiles that induced the innate and specific immune response in both infectious processes were investigated. Both strains were able to improve resistance against the intestinal pathogen. Only Lr05 was able to induce a significant decrease in the number of S. pneumoniae in the lung, prevent its dissemination into the blood and induce a significant increase in Th1 (INF-γ) and Th2 (IL-6, IL-4 and IL-10) cytokine levels in the bronchoalveolar lavages (BAL). The changes in the cytokines profiles in BAL were associated with an increase in the number and activity of phagocytic cells and with the increase in specific antibodies in serum and BAL, which would explain the increased resistance to the challenge. The administration of Lr06 did not induce significant effects at the respiratory mucosal level. The results described in the present paper showed that certain LAB strains can share certain functional properties, although some of them can perform a functional role better than others, so that it is important to perform careful studies on specific strains, according to their therapeutic use. [Copyright &y& Elsevier]

Abstract: We developed a novel selective medium, modified-rhamnose-2,3,5-triphenyltetrazolium chloride-LBS-vancomycin agar (M-RTLV agar), that utilizes the fermentability of l-rhamnose to distinguish Lactobacillus casei and Lactobacillus paracasei from Lactobacillus rhamnosus. Whereas L. casei and L. paracasei formed red colonies on the M-RTLV agar, L. rhamnosus formed either pink-toned colonies or white colonies with a red spot. An intervention study was conducted to confirm the capability of M-RTLV agar to detect ingested L. casei when recovered from human feces. Subjects consumed one bottle daily of a fermented milk product (Yakult® or Yakult® Light, which contains L. casei strain Shirota; LcS) for 7days. Diluents of the fecal samples were cultivated on M-RTLV agar. We were able to enumerate circular medium-sized red colonies, which were morphologically similar to L. casei/L. paracasei but clearly distinguishable from the remaining colonies owing to the color difference. These colonies were then subjected to enzyme-linked immunosorbent assay in order to identify the LcS. The viable counts of LcS were 6.6±0.7log10 CFU/g feces after intake of Yakult® and 6.5±0.6log10 CFU/g feces after intake of Yakult® Light (mean±SD). [Copyright &y& Elsevier]

Abstract: The capability of different fibre preparations to protect the viability and stability of Lactobacillus rhamnosus during freeze-drying, storage in freeze-dried form and after formulation into apple juice and chocolate-coated breakfast cereals was studied. In freeze-drying trials wheat dextrin and polydextrose proved to be promising carriers for the L. rhamnosus strains: both freeze-drying survival and storage stability at 37 °C were comparable to the control carrier (sucrose). Using apple fibre and inulin carriers resulted in powders with fairly good initial freeze-drying survival but with poor storage stability at 37 °C. When fresh L. rhamnosus cells were added into apple juice (pH 3.5) together with oat flour with 20% β-glucan the survival of the cells was much better at 4 °C and at 20 °C than with sucrose, wheat dextrin and polydextrose, whereas with freeze-dried cells no protective effect of oat flour could be seen. The stability of freeze-dried L. rhamnosus cells at 20 °C was higher in chocolate-coated breakfast cereals compared to low pH apple juice. Similar to freeze-drying stability, wheat dextrin and polydextrose proved to be better carriers than oat flour in chocolate-coated breakfast cereals. Regardless of their differing capability to adhere to fibre preparations the two L. rhamnosus strains studied gave parallel results in the stability studies with different carriers. [Copyright &y& Elsevier]

Abstract: This study was undertaken to optimize yeast extract, glucose, and vitamin concentrations; and also culture pH for maximizing the growth of a probiotic bacterium, Lactobacillus rhamnosus, and to assess the effects of these factors by using response surface methodology. A central composite design was used as an experimental design for the allocation of treatment combinations. A polynomial regression model with cubic and quartic terms was used for analysis of the experimental data. It was found that the effects involving yeast extract, glucose, vitamins and pH on the growth of L. rhamnosus were significant, and the strongest effect was given by the yeast extract concentration. Estimated optimum conditions of the factors for the growth of L. rhamnosus are as follows: pH=6.9; vitamin solution=1.28% (v/v); glucose=5.01% (w/v) and yeast extract=6.0% (w/v). [Copyright &y& Elsevier]

To develop a cost-effective fermentation medium, biomass of Debaryomyces hansenii coming from xylitol production and corn steep liquor was evaluated for lactic acid production by Lactobacillus rhamnosus in glucose-containing media. Glucose consumption and lactic acid generation were followed using a variety of media made with different nutrient supplementation. Under selected conditions, media containing glucose, biomass of D. hansenii and corn steep liquor as unique components led to product yields similar to those obtained in a fully supplemented medium. [Copyright &y& Elsevier]

PCR primers derived from Lactobacillus rhamnosus phage Lc-Nu genome were used to screen the presence of phage-related sequences in Lb. rhamnosus strains. Several primer pairs derived from structural and replication gene regions of phage Lc-Nu amplified PCR products of expected sizes from bacterial strains revealing phage-related sequences in 10 of 11 Lb. rhamnosus strains. Strain-specific PCR primers for three probiotic Lb. rhamnosus strains were derived from these phage-related sequences for identification and detection purposes. Specificity of these primers was tested against 11 Lb. rhamnosus strains and over 40 other bacterial strains. [Copyright &y& Elsevier]

Amidst rising demand for non-dairy probiotic foods, and growing interest in coffees with added functionalities, it would be opportune to ferment coffee brews with probiotics. However, challenges exist in maintaining probiotic viability in high-moisture food products. Here, we aimed to enhance the viability of the probiotic bacteria, Lactobacillus rhamnosus GG, in coffee brews by co-culturing with the probiotic yeast, Saccharomyces cerevisiae var. boulardii CNCM-I745. The yeast significantly enhanced the viability of L. rhamnosus GG, as bacterial populations beyond 7 Log CFU/mL were maintained throughout 14 weeks of storage at 4 and 25 °C. In contrast, the single culture of L. rhamnosus GG suffered viability losses below 6 Log CFU/mL within 10 weeks at 4 °C, and 3 weeks at 25 °C. Growth and survival of S. boulardii CNCM-I745 remained unaffected by the presence of L. rhamnosus GG. Volatile profiles of coffee brews were altered by probiotic metabolic activities, but co-culturing led to suppressed generation of diacetyl and ethanol compared to single cultures. Probiotic fermentation did not alter principal coffee bioactive compounds and antioxidant capacities; however, declines in peroxyl radical scavenging capacities were observed after ambient storage. Overall, we illustrate that yeasts are effective in enhancing probiotic bacterial viability in coffee brews, which may be useful in developing shelf stable probiotic food products. • Coffee brews were fermented with probiotic bacteria and yeast. • Survival of L. rhamnosus GG in coffee brews was enhanced by S. boulardii. • Nutrient uptake by S. boulardii limited lactic acid production by the bacteria. • Probiotic metabolites changed headspace flavour compounds of coffee brews. • Major coffee bioactive components and antioxidant capacities were mostly retained. [ABSTRACT FROM AUTHOR]

The present study evaluated impact of moisture content and cell conditions on survival of probiotic strain, Lactobacillus rhamnosus GG, under lethal heat stresses and during long-term storage using freeze-dried cells and oils as matrices. Viable cell counts of freeze-dried L. rhamnosus GG cells suspended in oils had only 1-log-reduction after 5min at 80°C and approximately 3-log-reduction after 20min, while no or very few viable cells were recorded for freeze dried cells suspended in buffer and cultured cells in oils. Surprisingly, freeze-dried cells suspended in oils still contained 4.3 to 6.7logCFU/ml after 5min at 95°C. Long-term storage study indicated that freeze-dried cells suspended in oils kept viable conditions for 4months, and a loss of the viability was only 0.3 to 0.6logCFU/ml. Viable cell counts of cultured cells suspended in oils were not present after 3days to 3months. These results clearly indicate that moisture and cell conditions have a great impact on survival of probiotics under severe heat stress in processing and during long-term storage. Combination of freeze-dried cells and oils as carrier provides beneficial options to preserve viability of probiotics in food processes and storage. [ABSTRACT FROM AUTHOR]

Since cheese is poor in energy for bacterial growth, it is believed that non-starter lactic acid bacteria growth and flavour development are supported by the nutrients from lysis of the starter culture. This study was performed to investigate the dynamics of interaction between starter and non-starter strains from cheese. A starter culture lysate was prepared by enzymatic digestion and tested as a growth substrate for Lactobacillus sp. strains. The two starter culture strains of Lactococcus lactis were also tested on the starter-lysate. All seventeen strains were individually inoculated at the level of 5.0 log 10 cfu mL−1 in M17 broth, with or without 10% starter-lysate, and incubated at 30 °C for 140 h. The optical density 600 nm was modelled with the primary log-transformed Logistic model with delay and lag phase duration, maximum specific growth rate as well as maximum population density obtained. Biphasic growth was mainly observed when the strains were able to utilize the starter-lysate as an energy source. To deal with the lack-of-fit related to the biphasic growth, the observed data points of the curve were divided after graphic evaluation and according to deviation of the residuals from the range ±0.05. Modelling was then performed in two phases by applying the same primary Logistic model in each of the two parts of the growth curve. Values of root-mean-square error and graphic evaluation indicated the good fitting of the data with the suggested approach. The growth of the two Lactococcus lactis strains was not affected by the starter-lysate. However, thirteen of the non-starter strains had their growth rates increased. The increase was greatest for Lactobacillus rhamnosus KU-LbR1, which reached maximum optical densities of 0.23 and 0.58 in the absence and the presence of starter-lysate, respectively. No effect of the starter-lysate was shown for the growth of Lactobacillus curvatus strains. The extend of the growth of non-starter strains on the starter-lysate was shown to be species and strain dependent. • A starter culture lysate was tested on growth of 15 non-starter and 2 starter lactic acid bacteria (SLAB) • Lack-of-fit by applying log-transformed Logistic model was shown due to biphasic growth • A two-phases Logistic modelling approach was proposed and successfully applied • N-glucosamine from SLAB cell wall seem to be the responsible for the increased growth of most tested non-starter LAB • The effect of starter lysate was shown to be species and strain dependent [ABSTRACT FROM AUTHOR]

Abstract: This study evaluated the effect of water activity on the long-term storage stability of the probiotic Lactobacillus rhamnosus GG (LGG) in a dry food matrix. Viability of LGG was further studied in a crushed flaxseed matrix – a new possible product matrix to deliver probiotics – as well as in reference matrices as maltodextrin. Three different water activities (aw =0.11, 0.22 and 0.43) were used, and preparations were stored at room temperature for up to 14months. The viability of LGG was less dependent on the matrix used, but strongly dependent on the water activity. Viability in flaxseed was lost rapidly with aw 0.43: with aw 0.22 the reduction was 2.4 log10 units and with aw 0.11 the reduction of viability was only 0.29 log10 units during the entire storage time. Taken together, regulating water activity to a low value may offer possibilities for extending the shelf life of dry probiotic products. [Copyright &y& Elsevier]

Matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry (MS), a culture based alternative for microbial diversity studies, is an attractive tool to dereplicate large numbers of isolates to a smaller set of representatives for downstream characterization. In the present study, MALDI-TOF MS, combined with a database of reference spectra compiled in previous studies, was applied to identify 88 non-starter lactic acid bacteria (NSLAB) isolated from 18 samples of four different artisanal cheeses produced in the Island of Naxos, Greece, from raw sheep and goat milk without the addition of starters. Eighty-four isolates (95.5%) could be identified directly via MALDI-TOF MS. Lactobacillus brevis and Lactobacillus plantarum were the dominant species, followed by Lactococcus lactis , Leuconostoc mesenteroides Lactobacillus paracasei , Lactobacillus rhamnosus , Pediococcus pentosaceus and Enterococcus faecium. The remaining four isolates represented species present in the database; however, within-species diversity was insufficiently covered. Additionally, pheS sequencing was applied to confirm identification. • Application of MALDI-TOF MS for the identification of lactic acid bacteria • First fingerprinting of the non-starter lactic acid bacteria of artisanal cheeses produced in the Island of Naxos, Greece • PheS sequencing was performed to validate MALDI-TOF MS identication [ABSTRACT FROM AUTHOR]

Fungi are commonly identified as the cause for dairy food spoilage. This can lead to substantial economic losses for the dairy industry as well as consumer dissatisfaction. In this context, biopreservation of fermented dairy products using lactic acid bacteria, propionibacteria and fungi capable of producing a large range of antifungal metabolites is of major interest. In a previous study, extensive screening was performed in vitro and in situ to select 3 dairy fermentates (derived from Acidipropionibacterium jensenii CIRM-BIA1774, Lactobacillus rhamnosus CIRM-BIA1952 and Mucor lanceolatus UBOCC-A-109193, respectively) with antifungal activity. The aim of the present study was to determine the main compounds responsible for this antifungal activity. Fifty-six known antifungal compounds as well as volatiles were targeted using different analytical methods (conventional LC and GC, GC–MS, LC-QToF). The most abundant antifungal compounds in P. jensenii -, L. rhamnosus - and M. lanceolatus -derived fermentates corresponded to propionic and acetic acids, lactic and acetic acids, and butyric acid, respectively. Many other antifungal compounds (organic acids, free fatty acids, volatile compounds) were identified but at lower levels. In addition, an untargeted approach using nano LC-MS/MS identified a 9-amino acid peptide derived from α s2 -casein in the L. rhamnosus -derived fermentate. This peptide inhibited M. racemosus and R. mucilaginosa in vitro. This study provides new insights on the molecules involved in antifungal activities of food-grade microorganism fermentates which could be used as antifungal ingredients in the dairy industry. • The compounds responsible for the antifungal activity of dairy fermentates were investigated. • A large variety of antifungal compounds (organic and fatty acids, volatiles) was evidenced. • A 9-amino acid antifungal peptide was also identified in a Lactobacillus rhamnosus fermentate. • New insights on antifungal molecules of food-grade microorganism fermentates are provided. [ABSTRACT FROM AUTHOR]