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Phyllosphere microorganisms are common contaminants of fruit or vegetable containing foods. The aim of this study was to identify and characterize lactic acid bacteria isolated from fruits and vegetables from Reunion Island, regarding possible application in food. Among 77 isolates, a large diversity of species was observed, with isolates belonging to Lactobacillus plantarum (3 isolates), other species of Lactobacillus (3), Lactococcus lactis (13), Leuconostoc pseudomesenteroides (25), Leuconostoc lactis (1), Leuconostoc mesenteroides (7), Leuconostoc citreum (14), Weissella cibaria (4), Weissella confusa (4), other species of Weissella (2) and Fructobacillus tropaeoli (1). Several of these species, although belonging to lactic acid bacteria, are poorly characterized, because of their low occurrence in dairy products. Lactobacillus, Lactococcus, Leuconostoc and Weissella isolates were classified by (GTG) 5 fingerprinting in 3, 6, 21 and 10 genetic groups, respectively, suggesting a large intra-species diversity. Several Weissella and Lactobacillus isolates were particularly tolerant to acid and osmotic stress, whereas Lc. pseudomesenteroides 60 was highly tolerant to oxidative stress. Isolates of Weissella 30, 64 and 58, Leuconostoc 60 and 12b, Lactobacillus 75 and Fructobacillus 77 present relevant characteristics for their use as starters or as preservative cultures for fruits and vegetables., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Nutritional recommendations emphasize fruit and vegetable consumption. This is related to health-benefits associated with bioactive nutritive molecules (nutrients, vitamins, minerals, fibers, …) as well as non-nutritive phytochemicals (phenolic compounds, flavonoids, bioactive peptides, …) content of these types of foods. Fermented fruit or vegetable products are part of various diets worldwide. Lactic acid bacteria (LAB) fermentation is common among the various fermentation processes used. It produces changes in both profile and types of bioactive compounds. Molecules such as bioactive peptides, short chain fatty acids or polysaccharides are generated while sugar content or anti-nutritional compounds are decreased and phenolic compounds are converted to molecules with added biological value. These transformations, associated with prebiotic and/or probiotic potential supply as well as improvement of food components bioaccessibility and bioavailability, result in modifications of health-related properties. Lastly, interactions between ingested fermented food, intestinal microbiota and their correlations to metabolomics profiles and health represent an important perspective deserving to be further explored., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Spoilage of low-acid canned food during prolonged storage at high temperatures is caused by heat resistant thermophilic spores of strict or facultative bacteria. Here, we performed a bacterial survey over two consecutive years on the processing line of a French company manufacturing canned mixed green peas and carrots. In total, 341 samples were collected, including raw vegetables, green peas and carrots at different steps of processing, cover brine, and process environment samples. Thermophilic and highly-heat-resistant thermophilic spores growing anaerobically were counted. During vegetable preparation, anaerobic spore counts were significantly decreased, and tended to remain unchanged further downstream in the process. Large variation of spore levels in products immediately before the sterilization process could be explained by occasionally high spore levels on surfaces and in debris of vegetable combined with long residence times in conditions suitable for growth and sporulation. Vegetable processing was also associated with an increase in the prevalence of highly-heat-resistant species, probably due to cross-contamination of peas via blanching water. Geobacillus stearothermophilus M13-PCR genotypic profiling on 112 isolates determined 23 profile-types and confirmed process-driven cross-contamination. Taken together, these findings clarify the scheme of contamination pathway by thermophilic spore-forming bacteria in a vegetable cannery., (Copyright © 2015 Elsevier B.V. All rights reserved.)

Geobacillus stearothermophilus is the main thermophilic spore former involved in flat sour spoilage of canned foods. Three typing methods were tested and applied to differentiate strains at intra-species level: panC sequence analysis, REP-PCR and M13-PCR. panC gene was highly conserved within the studied strains, suggesting a low intra-specific diversity. This was supported by REP-PCR primary assays and M13-PCR results. M13-PCR profile analysis succeeded in differentiating six closely related groups (at 79% threshold similarity) among 127 strains from a range of spoiled canned food products and from different canneries. Phenotypic traits were investigated among 20 selected strains representing groups and origins. Ranges of growth under different temperatures (from 40 °C to 70 °C), pH (from 5.0 to 6.5), NaCl concentrations (from 1 to 5%) and sporulation conditions poorly differed between strains, but wet heat resistance of spores showed a 20-fold variation between strains. Furthermore, in this study, strains that belonged to the same M13-PCR genetic group did not share phenotypic characteristics or common origin. The work emphasizes a low diversity within the G. stearothermophilus species but data from this study may contribute to a better control of G. stearothermophilus spoilage in canned food., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

Thermal processing of Low Acid Canned Foods (LACF), which are safe and shelf-stable at ambient temperature for several years, results in heat inactivation of all vegetative microorganisms and the partial or total inactivation of spores. Good Manufacturing Hygienic Practices include stability tests for managing the pathogen risk related to surviving mesophilic bacterial spores. LACF are also often submitted to additional incubation conditions, typically 55 °C for 7 days, to monitor spoilage by thermophiles. In this study we identified the bacterial species responsible for non-stability after prolonged at 55 °C of incubation of LACF from 455 samples collected from 122 French canneries over 10 years. Bacteria were identified by microsequencing or a recent developed tool for group-specific PCR detection (SporeTraQ™). A single species was identified for 93% of examined samples. Three genera were responsible for more than 80% of all non-stability cases: mostly Moorella (36%) and Geobacillus (35%), and less frequently Thermoanaerobacterium (10%). The other most frequent bacterial genera identified were Bacillus, Thermoanaerobacter, Caldanaerobius, Anoxybacillus, Paenibacillus and Clostridium. Species frequency was dependent on food category, i.e. vegetables, ready-made meals containing meat, seafood or other recipes, products containing fatty duck, and related to the intensity of the thermal treatment applied in these food categories. The spore heat resistance parameters (D or δ and z values) from 36 strains isolated in this study were determined. Taken together, our results single out the species most suitable for use as indicators for thermal process settings. This extensively-documented survey of the species that cause non-stability at 55 °C in LACF will help canneries to improve the management of microbial contamination., (Copyright © 2013 Elsevier B.V. All rights reserved.)

Brettanomyces is the major microbial cause for wine spoilage worldwide and causes significant economic losses. Like Saccharomyces cerevisiae, it is well adapted to winemaking, but molecular pathways involved in this acclimatization are still unknown. In this work, we report a time-scale comparison between the two yeasts coping with alcoholic fermentation. Orthologs of some well-characterized stress genes of S. cerevisiae were searched by sequence alignment in the Dekkera/Brettanomyces partial genome; nine genes were finally selected on the basis on their similarity and involvement in adaptation to wine. Transcript analysis during a model grape juice fermentation indicates that a subset of genes (i.e., ATP1, ERG6, VPS34) shows peculiar expression patterns in Brettanomyces bruxellensis but also that some common regulations of stress response exist between the two yeasts, although with different timing (i.e., for MSN4, SNF1, HSP82, NTH1). This suggests that B. bruxellensis efficient survival in such challenging conditions is due to mechanisms unique to it, together with a conserved yeast stress response. This study, although limited by the poor genetic data available on B. bruxellensis, provides first insights into its gene expression remodeling in winemaking and opens new frames for further investigations.

Extracellular proteins from Oenococcus oeni, a wine-making bacterium, were isolated during growth on media differing by their nitrogen content. Analysis by two-dimensional electrophoresis revealed a low number of protein signals. Among the main spots, one signal corresponded to a single protein, which contained a lysine repeat domain characteristic of cell-wall hydrolases. We demonstrated that this major protein, named EprA, was able to hydrolyse several proteins. The heterologous production of this protein in Escherichia coli confirmed the protease activity of EprA. With a MW of 21.3 kDa and a pI of 5.3, EprA presents optimal activity at pH 7.0 and 45 degrees C. This O. oeni protease differs from all lactic acid bacteria proteases so far identified, and thus this bacterium possesses at least three proteases for wine protein hydrolysis.

Aims: The characterization of global aerobic metabolism of Lactobacillus plantarum LP652 under different aeration levels, in order to optimize acetate production kinetics and to suppress H2O2 toxicity., Methods and Results: Cultures of L. plantarum were grown on different aeration conditions. After sugar exhaustion and in the presence of oxygen, lactate was converted to acetate, H2O2 and carbon dioxide with concomitant ATP production. Physiological assays were performed at selected intervals in order to assess enzyme activity and vitality of the strain during lactic acid conversion. The maximal aerated condition led to fast lactate-to-acetate conversion kinetics between 8 and 12 h, but H2O2 immediately accumulated, thus affecting cell metabolism. Pyruvate oxidase activity was highly enhanced by oxygen tension and was responsible for H2O2 production after 12 h of culture, whereas lactate oxidase and NADH-dependent lactate dehydrogenase activities were not correlated to metabolite production. Limited NADH oxidase (NOX) and NADH peroxidase (NPR) activities were probably responsible for toxic H2O2 levels in over-aerated cultures., Conclusion: Modulating initial airflow led to the maximal specific activity of NOX and NPR observed after 24 h of culture, thus promoting H2O2 destruction and strain vitality at the end of the process., Significance and Impact of the Study: Optimal aeration conditions were determined to minimize H2O2 concentration level during growth on lactate.

Aims: The aim of the study was to characterize the effect of various nitrogen sources on Oenococcus oeni growth, carbon source utilization, extracellular protease activity and extracellular proteins. More generally, the goal is to understand how nitrogen-based additives might act to enhance malolactic fermentation in wine., Methods and Results: Five yeast extracts were used. As the amino acid and nitrogen analyses revealed, they were similar in global amino acid composition, except for arginine level. Nevertheless the ratio of amino acids between free/bound, and low/high molecular weight fractions were highly different. One of the yeast extracts led to a significant protease activity in the supernatant and to a poor final biomass of the IOB84.13 strain compared to the other ones. For the IOB84.13 strain specifically, arginine addition to the arginine poor yeast extract did not restore growth. 35S-methionine-labelled extracellular proteins were separated by SDS-PAGE. Signals were detected in all media early in the growth phase and were maintained during 48 h of culture., Conclusions: A significant protease activity was detected for O. oeni supernatants during growth under nitrogen limitation but only for certain nitrogen sources. Moreover, the activity was strain dependent. Peptides (0.5-10 kDa) seemed to be more favourable for growth of wine bacteria than <0.5 kDa nitrogen sources. The extracellular protein signal patterns differed more greatly between the bacterial strains tested than between the nitrogen molecules in the medium., Significance and Impact of the Study: This is the first study extensively considering the role of the nitrogen source composition and level upon O. oeni growth and metabolism.