Comparative immunomodulatory effects in mice and in human dendritic cells of five bacterial strains selected for biocontrol of leafy green vegetables.

The market for ready-to eat vegetables is increasing, but unfortunately so do the numbers of food-borne illness outbreaks related to these products. A previous study has identified bacterial strains suitable for biocontrol of leafy green vegetables to reduce the exposure to pathogens in these products. As a tentative safety evaluation, five selected strains (Rhodococcus cerastii MR5x, Bacillus coagulans LMG P-32205, Bacillus coagulans LMG P-32206, Pseudomonas cedrina LMG P-32207 and Pseudomonas punonensis LMG P-32204) were individually compared for immunomodulating effects in mice and in human monocyte-derived dendritic cells (MoDCs). Mice receiving the two B. coagulans strains consistently resemble the immunological response of the normal control, and no, or low, cell activation and pro-inflammatory cytokine expression was observed in MoDCs exposed to B. coagulans strains. However, different responses were seen in the two models for the Gram-negative P. cedrina and the Gram-positive R. cerastii. Moreover, P. punonensis and B. coagulans increased the microbiota diversity in mice as seen by the Shannon-Wiener index. In conclusion, the two strains of B. coagulans showed an immunological response that indicate that they lack pathogenic abilities, thus encouraging further safety evaluation and showing great potential to be used as biocontrol agents on leafy green vegetables.

Seed inoculation with antagonistic bacteria limits occurrence of E. coli O157:H7gfp + on baby spinach leaves.

BACKROUND: During the last decades, outbreaks of foodborne illnesses have increasingly been linked to fresh and/or minimally processed fruit and vegetables. Enterohemorrhagic Escherichia coli was the causal agent for major outbreaks in Europe with leafy green vegetables and sprouts. To improve food safety, microbial antagonism has received attention during recent years and could be one of the solution to prevent contamination of food borne pathogens on fresh produce. Here we investigate the antagonistic effect of three bacterial strains (Pseudomonas orientalis, P. flavescens and Rhodococcus sp.) isolated from spinach leaves against E. coli O157:H7gfp + under laboratory and greenhouse conditions. RESULTS: Our results shows that significantly less culturable E.coli O157:H7gfp + were retrieved from the spinach canopy subjected to antagonist seed treatment than canopy inoculation. Seeds inoculated with Rhodococcus sp. significantly reduced growth of E. coli O157:H7gfp + compared with the other antagonists. The result from the in vitro study shows a significant reduction of growth of E. coli O157:H7gfp+, but only after 44 h when E. coli O157:H7gfp + was propagated in a mixture of spent media from all three antagonists. CONCLUSIONS: The antagonistic effect on phyllospheric E.coli O157:H7gfp + observed after seed inoculation with Rhodococcus sp. might be an indication of induced resistance mechanism in the crop. In addition, there was a small reduction of culturable E.coli O157:H7gfp + when propagated in spent media from all three antagonists. Nutritional conditions rather than metabolites formed by the three chosen organisms appear to be critical for controlling E. coli O157:H7gfp+.

Processed meat products with added plant antioxidants affect the microbiota and immune response in C57BL/6JRj mice with cyclically induced chronic inflammation.

Epidemiological studies have found that there is a correlation between red and processed meat consumption and an increased risk of colorectal cancer. There are numerous existing hypotheses on what underlying mechanisms are causative to this correlation, but the results remain unclear. A common hypothesis is that lipid oxidation, which occurs in endogenous lipids and phospholipids in consumed food, are catalyzed by the heme iron in meat. In this study, five pre-selected plant antioxidant preparations (sea buckthorn leaves and sprouts, summer savory leaves, olive polyphenols, onion skin and lyophilized black currant leaves) were added to a meatball type prone to oxidize (pork meat, 20 % fat, 2% salt, deep-fried and after 2 weeks of storage). Pro-inflammatory markers, neutrophil infiltration and microbiota composition were studied after four months in a chronic inflammation model in C57BL6/J female mice. We found that the bacterial diversity index was affected, as well as initial immunological reactions.

Leaf mineral content govern microbial community structure in the phyllosphere of spinach (Spinacia oleracea) and rocket (Diplotaxis tenuifolia).

The plant microbiome is an important factor for plant health and productivity. While the impact of nitrogen (N) availability for plant growth and development is well established, its influence on the microbial phyllosphere community structure is unknown. We hypothesize that nitrogen impacts the growth and abundance of several microorganisms on the leaf surface. The bacterial and fungal communities of baby leaf spinach (Spinacia oleracea), and rocket (Diplotaxis tenuifolia) were investigated in a field trial for two years in a commercial setting. Nitrogen fertilizer was tested in four doses (basic nitrogen, basic + suboptimal, basic + commercial, basic + excess) with six replicates in each. Culture-independent (Illumina sequencing) and culture-dependent (viable count and identification of bacterial isolates) community studies were combined with monitoring of plant physiology and site weather conditions. This study found that alpha diversity of bacterial communities decreased in response to increasing nitrogen fertilizer dose, whereas viable counts showed no differences. Correspondingly, fungal communities of the spinach phyllosphere showed a decreasing pattern, whereas the decreasing diversity of fungal communities of rocket was not significant. Plant species and effects of annual variations on microbiome structure were observed for bacterial and fungal communities on both spinach and rocket. This study provides novel insights on the impact of nitrogen fertilizer regime on a nutrient scarce habitat, the phyllosphere.

The Hurdle Approach-A Holistic Concept for Controlling Food Safety Risks Associated With Pathogenic Bacterial Contamination of Leafy Green Vegetables. A Review.

Consumers appreciate leafy green vegetables such as baby leaves for their convenience and wholesomeness and for adding a variety of tastes and colors to their plate. In Western cuisine, leafy green vegetables are usually eaten fresh and raw, with no step in the long chain from seed to consumption where potentially harmful microorganisms could be completely eliminated, e.g., through heating. A concerning trend in recent years is disease outbreaks caused by various leafy vegetable crops and one of the most important foodborne pathogens in this context is Shiga toxin-producing Escherichia coli (STEC). Other pathogens such as Salmonella, Shigella, Yersinia enterocolitica and Listeria monocytogenes should also be considered in disease risk analysis, as they have been implicated in outbreaks associated with leafy greens. These pathogens may enter the horticultural value network during primary production in field or greenhouse via irrigation, at harvest, during processing and distribution or in the home kitchen/restaurant. The hurdle approach involves combining several mitigating approaches, each of which is insufficient on its own, to control or even eliminate pathogens in food products. Since the food chain system for leafy green vegetables contains no absolute kill step for pathogens, use of hurdles at critical points could enable control of pathogens that pose a human health risk. Hurdles should be combined so as to decrease the risk due to pathogenic microbes and also to improve microbial stability, shelf-life, nutritional properties and sensory quality of leafy vegetables. The hurdle toolbox includes different options, such as physical, physiochemical and microbial hurdles. The goal for leafy green vegetables is multi-target preservation through intelligently applied hurdles. This review describes hurdles that could be used for leafy green vegetables and their biological basis, and identifies prospective hurdles that need attention in future research.

Effects of household washing on bacterial load and removal of Escherichia coli from lettuce and "ready-to-eat" salads.

Customer demands for fresh salads are increasing, but leafy green vegetables have also been linked to food-borne illness due to pathogens such as Escherichia coli O157:H7. As a safety measure, consumers often wash leafy vegetables in water before consumption. In this study, we analyzed the efficiency of household washing to reduce the bacterial content. Romaine lettuce and ready-to-eat mixed salad were washed several times in flowing water at different rates and by immersing the leaves in water. Lettuce was also inoculated with E. coli before washing. Only washing in a high flow rate (8 L/min) resulted in statistically significant reductions (p < .05), "Total aerobic count" was reduced by 80%, and Enterobacteriaceae count was reduced by 68% after the first rinse. The number of contaminating E. coli was not significantly reduced. The dominating part of the culturable microbiota of the washed lettuce was identified by rRNA 16S sequencing of randomly picked colonies. The majority belonged to Pseudomonadaceae, but isolates from Enterobacteriaceae and Staphylococcaceaceae were also frequently found. This study shows the inefficiency of tap water washing methods available for the consumer when it comes to removal of bacteria from lettuce. Even after washing, the lettuce contained high levels of bacteria that in a high dose and under certain circumstances may constitute a health risk.