Human norovirus (NoV) causes gastroenteritis worldwide and has been associated with a number of produce related outbreaks. The design of effective inactivation and prevention procedures requires an understanding of virus survival in environments applicable to the production and processing of fresh produce. To evaluate the extent of NoV risk from farm to fork, the survival of murine norovirus (MNV), a surrogate for human NoV, was studied on stainless steel disks, soil and in bottled water for 42 days and on lettuce for 15 days in the laboratory. Stability experiments were then conducted on farm during one lettuce planting/harvest cycle, for 4 weeks. MNV stability was tested at room temperature in the laboratory or under ambient conditions on the farm. A one log reduction in virus titre was achieved after 30 days in water, 4 days on lettuce, 15 days on stainless steel disks, 12 days on loamy and sandy soil. For farm testing, infectious virus was recovered from both soil and lettuce on the day of inoculation. Although infectious virus was not recovered at later time points, the viral genomes were detected for up to four weeks. The observed long-term persistence of NoV, under both laboratory and field conditions, provides valuable information for developing risk assessments and control procedures to limit the possibility for NoV transmission in the food supply.

Human norovirus (NoV) causes gastroenteritis worldwide and has been associated with a number of produce related outbreaks. The design of effective inactivation and prevention procedures requires an understanding of virus survival in environments applicable to the production and processing of fresh produce. To evaluate the extent of NoV risk from farm to fork, the survival of murine norovirus (MNV), a surrogate for human NoV, was studied on stainless steel disks, soil and in bottled water for 42 days and on lettuce for 15 days in the laboratory. Stability experiments were then conducted on farm during one lettuce planting/harvest cycle, for 4 weeks. MNV stability was tested at room temperature in the laboratory or under ambient conditions on the farm. A one log reduction in virus titre was achieved after 30 days in water, 4 days on lettuce, 15 days on stainless steel disks, 12 days on loamy and sandy soil. For farm testing, infectious virus was recovered from both soil and lettuce on the day of inoculation. Although infectious virus was not recovered at later time points, the viral genomes were detected for up to four weeks. The observed long-term persistence of NoV, under both laboratory and field conditions, provides valuable information for developing risk assessments and control procedures to limit the possibility for NoV transmission in the food supply.

Human norovirus (NoV) causes gastroenteritis worldwide and has been associated with a number of produce related outbreaks. The design of effective inactivation and prevention procedures requires an understanding of virus survival in environments applicable to the production and processing of fresh produce. To evaluate the extent of NoV risk from farm to fork, the survival of murine norovirus (MNV), a surrogate for human NoV, was studied on stainless steel disks, soil and in bottled water for 42 days and on lettuce for 15 days in the laboratory. Stability experiments were then conducted on farm during one lettuce planting/harvest cycle, for 4 weeks. MNV stability was tested at room temperature in the laboratory or under ambient conditions on the farm. A one log reduction in virus titre was achieved after 30 days in water, 4 days on lettuce, 15 days on stainless steel disks, 12 days on loamy and sandy soil. For farm testing, infectious virus was recovered from both soil and lettuce on the day of inoculation. Although infectious virus was not recovered at later time points, the viral genomes were detected for up to four weeks. The observed long-term persistence of NoV, under both laboratory and field conditions, provides valuable information for developing risk assessments and control procedures to limit the possibility for NoV transmission in the food supply.

Human norovirus (NoV) causes gastroenteritis worldwide and has been associated with a number of produce related outbreaks. The design of effective inactivation and prevention procedures requires an understanding of virus survival in environments applicable to the production and processing of fresh produce. To evaluate the extent of NoV risk from farm to fork, the survival of murine norovirus (MNV), a surrogate for human NoV, was studied on stainless steel disks, soil and in bottled water for 42 days and on lettuce for 15 days in the laboratory. Stability experiments were then conducted on farm during one lettuce planting/harvest cycle, for 4 weeks. MNV stability was tested at room temperature in the laboratory or under ambient conditions on the farm. A one log reduction in virus titre was achieved after 30 days in water, 4 days on lettuce, 15 days on stainless steel disks, 12 days on loamy and sandy soil. For farm testing, infectious virus was recovered from both soil and lettuce on the day of inoculation. Although infectious virus was not recovered at later time points, the viral genomes were detected for up to four weeks. The observed long-term persistence of NoV, under both laboratory and field conditions, provides valuable information for developing risk assessments and control procedures to limit the possibility for NoV transmission in the food supply.

Genome-scale ordered RNA structure (GORS) was first identified in 2004. It refers to the presence of secondary structure throughout the length of the RNA genomes of certain genera of RNA virus families, as predicted by bioinformatic analysis. It was also observed that the viruses containing GORS were able to establish persistent infections in their natural hosts, raising the possibility that the presence of GORS could play a role in viral avoidance of the innate immune system. This thesis describes the first study of GORS and its possible role in persistence. Two GORS viruses have been studied, equine rhinitis A virus (ERAV) and murine norovirus (MNV). A 55% seroprevalence of ERAV has been determined in a cohort of Scottish horses indicating a wide exposure to the virus. Equine faecal samples were screened for ERAV by PCR with the intention of identifying a virus, possibly from a persistently infected animal, which would not have undergone any cell culture adaptations as laboratory strains have. Newly identified viruses would then be sequenced, their secondary structures predicted and further studies carried out. Unfortunately, none of the 50 faecal samples screened were positive and clinical isolates of ERAV provided by the Animal Health Trust were sequenced but were identical to laboratory strains, so the study then focussed on MNV. Prevalence of MNV in laboratory mice was determined by PCR of faecal samples to be 67%. MNV was also discovered in the faeces of a pet shop mouse and a wild wood mouse (Apodemus sylvaticus). The complete genomes of 4 laboratory mouse MNVs, the pet shop mouse and wood mouse MNVs were sequenced. Phylogenetic analysis showed the wood mouse MNV had a p distance of 23% from other MNVs, although the laboratory mice and pet shop mouse were closely related to other MNVs. Structural analysis of the genomes of 6 sequenced MNVs, including the wood mouse virus, showed all were GORS viruses. A laboratory strain of MNV, MNV-3, was serially passaged in RAW 264.7 cells to test the hypothesis that in an animal with an intact immune system, there is a pressure for GORS viruses to maintain their genomic RNA structure as a means of immune avoidance, and that cell culture adaptation would attenuate the degree of secondary structure. The complete genome of passage 33 was sequenced, which revealed 7 base mutations, a mutation rate of 0.1 %, which was not considered significant enough to have affected the degree of secondary structure. In order to assess if structured and unstructured RNA behaved differently in cells, replication deficient RNA transcripts were made from the infectious clones of a panel of GORS and non-GORS viruses. These transcripts were electroporated into cells and their rate of decay measured, but there was no difference between the GORS and non-GORS transcripts. The full length and 4 kilobase transcripts were transfected into NIH3T3 cells and the degree of interferon-β induction measured by quantitative PCR and a luciferase reporter assay. The IFN-β response differed across the panel of viruses, and although none of the GORS viruses induced strongly, the non-GORS viruses were variable in their ability to induce an IFN-β response, some inducing strongly, other not at all. This result indicates that during exposure of viral genomes in the cytoplasm during infection, GORS-virus RNAs are unlikely to induce an interferon response, possibly contributing to their ability to persist. It is unclear why some non-GORS-viruses failed to induce IFN and there are likely to be other contributory factors.

Le norovirus murin (MNV) est un agent pathogène de la souris récemment découvert et représente le contaminant le plus courant dans les animaleries de Recherche. Néanmoins, les effets de l'infection au MNV sur la recherche biomédicale ne sont pas encore clairs. Nous avons testé l'hypothèse que l'infection au MNV pourrait modifier la réponse immunitaire chez les souris atteintes d'une infection pulmonaire aiguë. Nous rapportons ici que la co-infection avec MNV augmente la survie des souris ayant une infection pulmonaire aiguë à Pseudomonas aeruginosa et diminue la production in vivo et in vitro de cytokines pro-inflammatoires. Nos résultats suggèrent que l'infection au MNV peut profondément modifier les paramètres étudiés dans les modèles classiques d'infection et mener à de fausses conclusions dans ces modèles expérimentaux. / The murine norovirus (MNV) is a recently discovered mouse pathogen, this virus represents the most common contaminant in laboratory mouse colonies. Nevertheless, the effects of MNV infection on biomedical research are still unclear. We tested the hypothesis that MNV infection could alter immune response in mice with acute lung infection. Here we report that co-infection with MNV increases survival of mice with Pseudomonas aeruginosa acute lung injury and decreases in vivo and in vitro production of pro-inflammatory cytokines. Our results suggest that MNV infection can deeply modify the parameters studied in conventional models of in-fection and lead to false conclusions in experimental models.

No description available.

Même si les traitements thermiques ou la désinfection par les oxydants ont démontré leur efficacité virucide, les mécanismes liés à la perte du caractère infectieux ne sont pas connus. Ceci pose un réel problème d’interprétation de la présence de génome viral en matière de risque infectieux dans les aliments. Ce travail de thèse a pour objectif d’étudier l’évolution des propriétés de surface (charge et hydrophobie) de virus modèles, bactériophage MS2 et norovirus murin, au cours de l’inactivation par la chaleur, l’hypochlorite de sodium et l’ozone. Pour nos deux virus, nous démontrons l’existence d’une température critique au-delà de laquelle la particule virale se déstructure en libérant son génome. Un simple traitement à la RNase permettrait alors de ne détecter que des virus infectieux par biologie moléculaire. Le traitement thermique implique aussi une augmentation de l’hydrophobie soulignant des modifications conformationnelles de la capside. L’hypochlorite de sodium ne modifie que peu les propriétés de surface mais des phénomènes d’oxydation ont lieu au niveau de la capside puisque la charge du bactériophage MS2 est légèrement modifiée. Ces modifications diminuent la résistance thermique du virus. Nous démontrons un effet synergique de l’hypochlorite de sodium et la chaleur sur le bactériophage MS2 (inactivation, RNase et hydrophobie). Quant à l’ozone gazeux, nous soulignons son intérêt pour le traitement virucide des aliments fragiles. Ainsi, ce travail précise les mécanismes d’inactivation des virus et ouvre de nouvelles perspectives tant pour discriminer les virus infectieux et non-infectieux que pour proposer l’exploration de nouveaux traitements technologiques / Although heat treatments or disinfections by oxidants have proven their virucidal efficiencies, mechanisms related to the loss of infectivity are not known. This statement could lead to a misinterpretation of the presence of viral genome on infection risk for humans in food matrices. This thesis aimed to study the evolution of surface properties (charge and hydrophobicity) for model viruses, bacteriophage MS2 and murine norovirus, during the heat, sodium hypochlorite and ozone inactivations. For both viruses, the existence of a critical temperature beyond which the viral particle was disrupted and released its genome was demonstrated. Simple treatment with RNase would then only detect infectious virus by molecular biology. The heat treatment also involved a transient increase in the hydrophobicity which highlighted conformational changes of the viral capsid. Sodium hypochlorite slightly modified the surface properties but oxidation phenomena occurred onto capsid since the bacteriophage MS2 charge has changed a little. These changes decreased the thermal resistance of the virus. Synergistic effects of both sodium hypochlorite and heat were observed on the inactivation of MS2 phages, the sensitivity of their genome to RNases and the increase in hydrophobicity of remaining infectious particles. Regarding gaseous ozone, we underlined its interest in the case of virucidal treatment of fragile food matrices. Therefore, this work specified the virus inactivation mechanisms and opened up new perspectives to discriminate infectious from non-infectious viruses but also to propose the exploration of new technological processes

Challenged by a huge and diverse antigenic stimulus, the intestinal mucosa has developed a unique immune system that mainly functions to maintain tolerance to innocuous antigens while retaining the ability to respond swiftly to pathogenic threats. Central to this specialised immune system are the Intraepithelial Lymphocytes (IELs). These cells are uniquely located between Intestinal Epithelial Cells (IECs) ready to respond to exogenous antigens in the intestinal lumen. The intestinal immune system is constantly influenced, not only by the commensal microbiota, but also by the nutritional status of the host and the availability of certain essential micronutrients that are derived from a healthy-balanced diet. Additionally, age has a significant impact on the efficiency of gut immunity in responding to infectious pathogens, as reflected by the increased burden of gastrointestinal infections at the extremes of age. In this thesis, using the Murine Norovirus (MNV) oral infection model, I aimed to characterize intestinal mucosal antiviral-responses with specific focus on the role of IELs, the impact of aging and the influence of certain micronutrients whose effects are mediated through the Aryl Hydrocarbon Receptor (AhR). Employing different knock-out and adoptive transfer experiments, I concluded that, at least in our experimental conditions and in a viral strain-specific manner, the activated IELs are not essential and may play a minor role in the protective response against MNV infection. This work also demonstrated that various MNV virus strains activate IELs differentially and for the first time (to our knowledge) revealed distinct abilities of these different Norovirus variants to infect IECs. Recognising an impaired response in old (2-year) mice, we were also able to identify a specific defect in the IFN-Lambda response of aged IECs. Furthermore, using the model of MNV infection to investigate the role of AhR signalling, the data I generated suggested a direct link between constitutive AhR signalling and innate interferon-mediated responses. These findings have uncovered a potential preventive/therapeutic targets for enhancing anti-viral responses.

Comparing the protein compositions of three fishes, grass carp exhibited lower band intensity at 47.9 KDa, β-tropomyosin (36.5 KDa), and missed the band at 15.9 KDa myosin light chain. Bigmouth buffalo had a darker tropomodulin (38.8 KDa) band and smaller α-tropomyosin (33-37 KDa) than silver and grass carp. The breaking force (611.8 g) and deformation (11.7 mm) of silver carp cooked gel were significantly higher than the other two fish products. The addition of starches at 2, 4, and 6% to the grass carp paste lowered the breaking force of the cooked gel in a dose-responsive manner compared to the control (P<0.05), but no differences were found in bigmouth buffalo. The bioumulated Murine Norovirus-1 (MNV-1) was found to maintain infectivity during storage of live oysters at 5°C for 15 days while the inoculated MNV-1 kept infectious for 20 days in cooked surimi and salmon fillet. Treatments with 4.0 kGy X-ray achieved the reductions of 3.7 log PFU mL-1 in pure culture or 2.7, 2.2, and 2.0 log PFU g-1 in half-shell oyster, salmon sushi and tuna salad, respectively. X-ray significantly reduced the population of internalized MNV-1 in live oysters from 4.3 ± 0.4 log PFU g-1 to 3.6 ± 0.5, 3.2 ± 0.2, 2.8 ± 0.2, and 2.5 ± 0.1 log PFU g-1, by 1.0, 2.0, 3.0, and 4.0 kGy X-ray, respectively. The population of MNV-1 was reduced to less than 2.0 log PFU g-1 at 5.0 kGy X-ray. The survivability of live oysters was not significantly affected by treatment with 5.0 kGy X-ray, in comparison with the control, for up to 10 days, respectively, during storage at 5°C. Fish sauce was fermented from the by-products of silver carp. The total nitrogen content of fish sauce made in April, and November were 9.86±0.9 and 9.71±4.5 g/l, respectively, which was significantly (p<0.05) higher than the sample of February (8.45±0.25 g/l ), reflecting seasonal effect. The total nitrogen, amino acid nitrogen, pH, and sodium chloride of fish sauce made from silver carp by-products met the international fish sauce standard code of CODEX STAN 302-2011.