Quality analysis of critical control points within the whole food chain and their impact on food quality, safety and health (QACCP)

The overall objective of the project was to optimise organic production and processing in order to improve food quality and increase health promoting aspects in consumer products. The approach was a chain analysis approach which addressed the link between farm and fork and backwards from fork to farm. The objectives were to test food authenticity on farm level and food quality and health in processing. The carrot was chosen as the model vegetable since it is common for the involved partners from industry and is processed for baby food; hence the results are relevant for other vegetables and organic food in general as well. - Identify and define critical and essential product quality parameters useful to optimise organic food quality - Compare products from different farming practices (conventional and within organic) - Performance of QACCP (Quality Analysis Critical Control Point, similar to HACCP methodology) - Test the impact of the food chain (focusing on processing techniques) on the product quality and safety - Test the impact of organic food on healt

Differential protection by cell wall components of Lactobacillus amylovorus DSM 16698T against alterations of membrane barrier and NF-kB activation induced by enterotoxigenic F4+ Escherichia coli on intestinal cells

Background: The role of Lactobacillus cell wall components in the protection against pathogen infection in the gut is still largely unexplored. We have previously shown that L. amylovorus DSM 16698(T) is able to reduce the enterotoxigenic F4(+)Escherichia coli (ETEC) adhesion and prevent the pathogen-induced membrane barrier disruption through the regulation of IL-10 and IL-8 expression in intestinal cells. We have also demonstrated that L. amylovorus DSM 16698T protects host cells through the inhibition of NF-kB signaling. In the present study, we investigated the role of L. amylovorus DSM 16698(T) cell wall components in the protection against F4(+)ETEC infection using the intestinal Caco-2 cell line. Methods: Purified cell wall fragments (CWF) from L. amylovorus DSM 16698T were used either as such (uncoated, U-CWF) or coated with S-layer proteins (S-CWF). Differentiated Caco-2/TC7 cells on Transwell filters were infected with F4(+)ETEC, treated with S-CWF or U-CWF, co-treated with S-CWF or U-CWF and F4(+)ETEC for 2.5 h, or pre-treated with S-CWF or U-CWF for 1 h before F4(+)ETEC addition. Tight junction (TJ) and adherens junction (AJ) proteins were analyzed by immunofluorescence and Western blot. Membrane permeability was determined by phenol red passage. Phosphorylated p65-NF-kB was measured by Western blot. Results: We showed that both the pre-treatment with S-CWF and the co- treatment of S-CWF with the pathogen protected the cells from F4(+)ETEC induced TJ and AJ injury, increased membrane permeability and activation of NF-kB expression. Moreover, the U-CWF pre-treatment, but not the co- treatment with F4(+)ETEC, inhibited membrane damage and prevented NF-kB activation. Conclusions: The results indicate that the various components of L. amylovorus DSM 16698(T) cell wall may counteract the damage caused by F4(+)ETEC through different mechanisms. S-layer proteins are essential for maintaining membrane barrier function and for mounting an anti-inflammatory response against F4(+)ETEC infection. U-CWF are not able to defend the cells when they are infected with F4(+)ETEC but may activate protective mechanisms before pathogen infection.Peer reviewe

Alternatives to in-feed antibiotics in pigs: Evaluation of probiotics, zinc or organic acids as protective agents for the intestinal mucosa. A comparison of in vitro and in vivo results

Weaned pigs are susceptible to frequent infectious diseases. Antibiotics have been used over decades to reduce pathogen infections, but many microorganisms are becoming resistant to antibiotics. Thus, there is an urgent need to find alternatives to in-feed antibiotics. Up to now, various compounds have been used as alternatives, giving promising but sometimes contrasting results. In recent years, the in vitro models of cell culture have been proposed for the screening of various compounds and understanding their mechanisms of action. The intestine is a complex system with a continuous cross-talk among epithelial cells, the local immune system and microflora. Alternatives to in-feed antibiotics may interact with any of these constituents. This review presents data on the protective effects of probiotics, zinc salts and organic acids, used as alternatives to in-feed antibiotics, against pathogen induced damage and inflammatory processes in the intestinal cells and the underlying mechanisms, with an attempt to compare in vitro and in vivo results. All the alternatives tested resulted in effective protection against pathogens by a series of interactions with intestinal cells and pathogens, including the inhibition of adhesion and invasion of pathogens, interference in signalling pathways induced by pathogenic bacteria, maintenance of the epithelial cell cytoskeleton and junctional structure and modulation of the host immune response.Alternatives aux antibiotiques additifs alimentaires chez le porc : évaluation des probiotiques, sels de zinc ou acides organiques comme agents protecteurs de la muqueuse intestinale. Comparaison des résultats in vitro avec les observations in vivo. Le porcelet est sensible aux infections gastro-intestinales au moment du sevrage. Les antibiotiques additifs alimentaires ont été utilisés pendant des décennies pour réduire ces infections, mais de nombreux microorganismes sont devenus résistants à ces substances. Aussi, est-il urgent de trouver des alternatives aux antibiotiques additifs alimentaires. Jusqu’à présent, diverses alternatives ont été envisagées, certaines donnant des résultats prometteurs, d’autres des résultats variables. Au cours des dernières années, les modèles in vitro de cultures cellulaires ont été proposés pour cribler ces substances et élucider leurs mécanismes d’action. L’intestin est un système complexe faisant intervenir des communications permanentes entre les cellules épithéliales, le système immunitaire local et la microflore. Les alternatives aux antibiotiques peuvent interagir avec n’importe lequel de ces composants. Le présent article fait la synthèse des effets protecteurs et des mécanismes d’action des probiotiques, des sels de zinc et des acides organiques, utilisés comme alternatives aux antibiotiques dans les processus inflammatoires et les altérations cellulaires épithéliales in vitro, induites par des agents pathogènes. Elle tente de comparer ces résultats avec les observations in vivo. Toutes les alternatives évaluées ont été efficaces contre les agents pathogènes, via des mécanismes d’interaction cellules épithéliales-agents pathogènes. Ces mécanismes incluaient l’inhibition de l’adhésion et de l’invasion cellulaire par ces pathogènes, la modulation des mécanismes de signalisation intracellulaire induits par les bactéries pathogènes, le maintien de la structure du cytosquelette et des jonctions serrées des cellules épithéliales, et enfin la modulation des réponses immunitaires de la cellule hôte

Impact of organic and conventional carrots on intestinal and peripheral immunity

BACKGROUND: Studies on health effects of organic (ORG) products are still limited and often contradictory. We have investigated the impact of ORG and conventional (CV) carrots from two consecutive harvest years on mouse peripheral and intestinal immunity. RESULTS: Danish carrots (Bolero variety) were grown in three ORG (O1, O2 and O3) and one CV cropping system (D-CV). Italian carrots (Maestro and Excelso varieties) were grown in one ORG and one CV field for each variety. Immune phenotypes of blood, spleen and intestinal lymphocytes, and cytokine serum levels were analyzed in mice fed the different carrots for 30 days. Principal component analysis (PCA) was performed in mice fed the Danish carrots. The consumption of the 'more organic' O2 and O3 carrots induced some changes in lymphocyte populations, including an increase in regulatory T cells. In Italian carrots more differences between ORG and CV were observed in the first as compared to the second year. No relevant differences were observed in cytokine secretion. PCA showed a clear separation among mice fed the O1, O2, O3 and D-CV carrots. CONCLUSIONS: Although a great variability was observed between the two years, an immune stimulation was found after the ORG carrot consumption. (C) 2012 Society of Chemical Industr

Altered expression, localization, and phosphorylation of epithelial junctional proteins in celiac disease

We aimed to study the expression and localization of the molecular components of enterocyte junctions in celiac disease together with the level of tyrosine phosphorylation, a phenomenon known to affect their cellular distribution and function, and to explore the influence of proinflammatory cytokines. Duodenal biopsy specimens from patients with celiac disease and control subjects were used for immunoprecipitation, immunoblotting, and immunolocalization by using antioccludin, anti-zonula occludens (ZO)-1, anti-E-cadherin, anti-beta-catenin, and antiphosphotyrosine antibodies. The same procedures were carried out on filter-grown Caco-2 cells incubated in the absence or presence of interferon g and tumor necrosis factor a. In active celiac disease, the absence of a phosphorylated ZO-1 and the extensive phosphorylation of beta-catenin might be responsible for the absence of membranous localization of occludin and E-cadherin, respectively. The in vitro system showed an influence of the cytokines on the assembly of these complexes that proved the opposite to celiac samples as far as tight junctions were concerned because the presence of a phosphorylated ZO-1 enables occludin to localize in the membrane

Lactobacillus amylovorus inhibits the TLR4 inflammatory signaling triggered by enterotoxigenic Escherichia coli via modulation of the negative regulators and involvement of TLR2 in intestinal Caco-2 cells and pig explants.

Inflammation derived from pathogen infection involves the activation of toll-like receptor (TLR) signaling. Despite the established immunomodulatory activities of probiotics, studies relating the ability of such bacteria to inhibit the TLR signaling pathways are limited or controversial. In a previous study we showed that Lactobacillus amylovorus DSM 16698T, a novel lactobacillus isolated from unweaned pigs, protects the intestinal cells from enterotoxigenic Escherichia coli (ETEC) K88 infection through cytokine regulation. In the present study we investigated whether the ability of L. amylovorus to counteract the inflammatory status triggered by ETEC in intestine is elicited through inhibition of the TLR4 signaling pathway. We used the human intestinal Caco-2/TC7 cells and intestinal explants isolated from 5 week-old crossbreed Pietrain/Duroc/Large-White piglets, treated with ETEC, L. amylovorus or L. amylovorus cell free supernatant, either alone or simultaneously with ETEC. Western blot analysis showed that L. amylovorus and its cell free supernatant suppress the activation of the different steps of TLR4 signaling in Caco-2/TC7 cells and pig explants, by inhibiting the ETEC induced increase in the level of TLR4 and MyD88, the phosphorylation of the IKKα, IKKβ, IκBα and NF-κB subunit p65, as well as the over-production of inflammatory cytokines IL-8 and IL-1β. The immunofluorescence analysis confirms the lack of phospho-p65 translocation into the nucleus. These anti-inflammatory effects are achieved through modulation of the negative regulators Tollip and IRAK-M. We also found that L. amylovorus blocks the up-regulation of the extracellular heat shock protein (Hsp)72 and Hsp90, that are critical for TLR4 function. By using anti-TLR2 antibody, we demonstrate that TLR2 is required for the suppression of TLR4 signaling activation. These results may contribute to develop therapeutic interventions using L. amylovorus in intestinal disorders of piglets and humans