Thèse ou mémoire avec insertion d’articles. / Pour les transformateurs de l'industrie agroalimentaire, garantir la salubrité de leurs produits est une priorité absolue. Chaque étape de la production, de la manipulation des ingrédients aux différents procédés, peut potentiellement engendrer des maladies d'origine alimentaire. Les virus entériques, tels que le norovirus humain et le virus de l'hépatite A, sont des pathogènes pouvant mettre en danger le consommateur. Afin de minimiser ces risques, diverses méthodes de désinfection, tant physiques que chimiques, sont employées. Cependant, les propriétés structurales des différents genres viraux rendent difficile l'obtention d'un traitement virucide à large spectre. Pour cette raison, ce projet porte sur l'utilisation de composés chimiques novateurs à base de peracides pour l'inactivation virale. Pour atteindre cet objectif, quatre désinfectants ont été testés à différentes concentration (50, 80, 250, 500 et 1000 ppm) et temps de contact (0,5 1, 5, et 10 min) en solution liquide, sur de l'acier inoxydable et sur des petits fruits (fraise et bleuet). En solution liquide, l'acide perlevulinique avec du dodécyle sulfate de sodium (SDS) a permis une réduction du norovirus murin (MNV-1) de 3 logs, tandis que l'acide peracétique a provoqué une réduction de 2,24 logs en 0,5 minute. Seul l'acide peracétique a permis une réduction de 3 log en 0,5 minute sur l'acier inoxydable à 80 ppm. L'acide peracétique a d'ailleurs été l'unique désinfectant testé capable d'inactiver 3 logs de MNV-1 à la surface des bleuets à une concentration de 80 ppm pendant 30 secondes. Malheureusement, aucun des désinfectants n'a été en mesure d'inactiver au moins 2 logs de VHA et de VHE, peu importe la concentration et temps de contact. Éventuellement, les résultats de ce projet de maîtrise permettront le développement de nouvelles alternatives efficaces pour permettre l'inactivation de virus entériques dans l'industrie alimentaire par la création de nouveaux composés chimiques efficaces. / For processors in the food industry, ensuring the safety of their products is a top priority. Every step of production, from handling ingredients to different methods, has the potential to cause foodborne illness. Enteric viruses, such as human norovirus and hepatitis A virus, are pathogens that can endanger the consumer. In order to minimize these risks, various methods of prescription, both physical and chemical, are employed. However, the structural properties of the different viral speces make it difficult to achieve broad-spectrum virucidal therapy. For this reason, this project focuses on the use of new chemical compounds based on peracids for viral inactivation. To achieve this objective, four disinfectants were tested at different concentrations (50, 80, 250, 500 and 1000 ppm) and contact time (0.5 1, 5, and 10 min) in liquid solution, on stainless steel and on berries (strawberries and blueberries). In liquid solution, perlevulinic acid with sodium dodecyl sulfate (SDS) provided a 3-log reduction of murine nororivus (MNV-1), while perlevulinic acid caused a 2.24-log reduction in 0.5 min. Only peracetic acid provided a 3 log reduction in 0.5 minutes on stainless steel at 80 ppm. Peracetic acid was the only disinfectant tested to inactivate 3 logs of MNV-1 on the surface of blueberries at a concentration of 80 ppm for 30 seconds. Unfortunately, none of the disinfectants were able to inactivate at least 2 logs of HAV and HEV, regardless of concentration and contact time. Eventually, the results of this master's project will allow the development of new effective alternatives to enable the inactivation of enteric viruses in the food industry through the creation of new effective chemical compounds.

La majorité des gastroentérites non-bactériennes sont causées par les norovirus, qui se transmettent principalement par les surfaces, les aliments et l’eau. La plupart des désinfectants domestiques sont inefficaces contre ces virus à l’exception de l’acide hypochloreux qui, cependant, perd une partie de son activité et génère des sous-produits toxiques en présence de matières organiques. Des méthodes de désinfection alternatives, efficaces et plus sécuritaires sont aujourd’hui nécessaires pour lutter contre la propagation des norovirus. Ce projet de recherche a porté sur deux approches prometteuses, les acides peroxycarboxyliques et les lumières pulsées. Dans un premier temps, quatre acides peroxycarboxyliques ont été évalués en fonction de leur concentration et du temps de contact. Les acides monoperoxycarboxyliques, à savoir les acides peracétique et perpropionique, ont été les plus efficaces en réduisant la charge virale d’environ 4 log10 après un traitement de 50 mg L-1 pendant 5 minutes. Ces molécules ont conservé cette activité contre des norovirus déposés sur de l’acier inoxydable et du PVC, propres ou sales, ainsi qu’immobilisés dans un biofilm artificiel. Dans le cas de l’acide peracétique, les expérimentations ont démontré que cette inactivation résultait principalement de dommages causés sur l’ARN viral, probablement par l’intermédiaire de radicaux libres. À forte concentration, les protéines de la capside étaient aussi altérées. Dans un deuxième temps, les norovirus ont été traités par une lumière pulsée (200 à 1000 nm ; 0,69 J cm-2 par impulsion). Trois impulsions (1,6 seconde) ont diminué la charge virale d’environ 4 log10 dans tous les milieux testés (tampon salin, eaux dures, minérale et usées) à l’exception des eaux turbides. A la turbidité maximum (1000 NTU), la réduction était de 2.4 log10. Cette technologie est aussi efficace pour la désinfection de surfaces propres, même en présence d’un biofilm artificiel (4 log10 après 7 impulsions), mais est affectée par la présence de matières protéiques. Nous avons démontré que les lumières pulsées inactivaient les norovirus en induisant des dommages à la fois sur l’ARN viral et l’intégrité des particules. En conclusion, ces résultats attestent l’efficacité de ces techniques et contribuent à une meilleure compréhension de leur principe d'action. / The majority of non-bacterial gastroenteritis is caused by norovirus, which is transmitted primarily through surfaces, food and water. Most household disinfectants are ineffective against these viruses with the exception of the bleach which, however, loses partially its activity and generates toxic residues in the presence of organic matter. Alternative, effective and safer methods of disinfection are necessary to hamper the spread of norovirus. This research project focused on two promising approaches, namely peroxycarboxylic acids and pulsed light. On the one hand, four peroxycarboxylic acids were evaluated based on their concentration and contact time. Monoperoxycarboxylic acids, namely peracetic and perpropionic acids, were the most effective by reducing the viral load of about 4 log10 after a treatment with 50 mg L-1 for 5 minutes. These molecules maintained their activity against noroviruses attached on stainless steel and PVC, clean or dirty, and entrapped in an artificial biofilm. In the case of peracetic acid, we showed that this inactivation was mainly due to damages on the RNA, probably through free radicals. At high concentrations, the capsid was also altered. On the other hand, noroviruses were treated with pulsed light (200-1000 nm; 0.69 J cm 2 per pulse). Three pulses (1.6 seconds) decreased viral load of approximately 4 log10 in all media tested (buffered saline, hard water, mineral water and sewage treatment effluent) with the exception of turbid water. At the maximum turbidity (1000 NTU), the reduction was 2.4 log10. This technology is also effective to disinfect clean surfaces even in the presence of an artificial biofilm (4 log10 after 7 pulses), but is affected by the presence of proteinaceous material. We demonstrated that the pulsed light inactivated norovirus by inducing damage to both the RNA and the integrity of the particles. In conclusion, these results demonstrate the effectiveness of these approaches and contribute to a better understanding of their mechanism of action.

Microbubbles are small gas-filled bubbles with diameters ranging from 50 to 1 μm, and less than 200 nm are called nanobubbles. Their small sizes and large specific surface area result in a high gas dissolution rate and long lifetime in liquid. Ozone is a strong oxidant that destroys microorganisms and only produces oxygen as the final by-product in fresh water. However, due to the poor stability of aqueous ozone, critical gas waste happens during treatments which leads to a high economic loss. Microbubbles have shown promising enhancement of ozone treatment. In previous studies, ozone microbubbles exhibited excellent efficacy in the removal of organic contaminants and inactivation of microorganisms including bacteria, spores, and fungi, but few articles discuss the virus inactivation of ozone microbubbles treatment. Human noroviruses (NoVs) are the primary cause of foodborne illnesses in the US, and the development of effective inactivation methods is crucial. Because of the absence of suitable in vitro cultivation methods for NoVs and the constraints of the available infectivity models for these viruses, most of the studies about inactivation use surrogate viruses that are similar to NoVs in genetics and structure. Tulane virus is a NoV surrogate that can identify the same putative co-factor. This study focuses on the influence of treatment time, disinfectant air exposure, and the presence of organic contaminants on the inactivation efficacy of ozone microbubbles or millimeter bubbles. The results demonstrate that more than one log10 reduction was produced when the Tulane virus was exposed to ozone millimeter bubbles and ozone microbubbles for a short period of time, even in the presence of high organic load (FBS), and the protective effect of the organic load was shown when the disinfectant induced volume increased. The findings indicate that conducting further research on ozone microbubbles in aqueous applications in food-related applications is useful.

Les virus d'origine alimentaire, comme le virus de l'hépatite A (VHA) et le norovirus (NoV), sont des causes importantes d'infections virales. Chaque année, celles-ci déduisent de grandes sommes monétaires aux systèmes de santé canadiens ainsi qu'aux producteurs alimentaires, sans compter les pertes alimentaires et économiques dues aux rappels d'aliments contaminés. Les stratégies de contrôle conventionnelles ne sont pas suffisamment efficaces et présentent, dans certains cas, des risques pour les travailleurs et/ou pour les consommateurs. Il est donc essentiel d'offrir de nouvelles méthodes d'inactivation naturelles et plus sécuritaires. Les produits issus de bleuet et de canneberge ont le potentiel de devenir ces nouvelles méthodes, notamment par leur concentration forte en polyphénols. De plus, ils sont naturels et sécuritaires pour les travailleurs et les consommateurs. Sans oublier qu'ils sont abondants au Québec, et, de ce fait, à moindres coûts. Le potentiel antiviral de ces produits de bleuet et de canneberge a été étudié et analysé dans ce mémoire envers des virus nus d'origine alimentaire (VHA et NVM-1) en plus de tester un modèle de virus enveloppé, soit le virus de l'herpès simplex de type 1 (HSV-1). NVM-1 et VHS-1 ont été complètement inactivés à l'aide de jus de canneberge et/ou de bleuet (concentrés à 50 %) en 5 minutes de pré-incubation cellulaire ou virale respectivement. VHA a subi une réduction de 3,70 ± 0,54 log ufp/mL à la suite d'une pré-incubation virale de 90 minutes. / Foodborne viruses like the hepatitis A virus (HAV) and the norovirus (NoV) are known as important cause of viral infections. Each year, they cost massive amount to the Canadian health-care systems and to the producers without counting the important food loss due to recalls. The conventional methods are not sufficient and can represent in some case a threat to the worker or to the consumer, which is why it is essential to uncover new inactivation methods that are safer and natural, like berries. The blueberry and cranberry products have the potential to become these new methods considering their interesting composition. Furthermore, they are natural, do not show signs of threat to the worker nor to the consumer and above all they are abundant in Québec and at low cost. The antiviral potential of these products from blueberries and cranberries have been tested and studied in this memoir against the HAV and the MNV-1, naked foodborne viruses as well as against the herpes simplex virus type 1 (HSV-1), an enveloped model. The infectivity of MNV-1 was neutralized completely in the presence of this host cells at 37 °C in as little as 5 min of contact with cranberry or blueberry juices (50 % concentration). HSV-1 was also completely neutralized in viral pre-incubation with the same juice concentration and incubation. HAV infectious titer fell 3.70 ± 0.54 log pfu/mL after 90 minutes of contact with cranberry juice. These results suggest that cranberry and blueberry processing could be useful for reducing the transmission of foodborne viruses.

Les virus d’origine alimentaire, comme le virus de l’hépatite A (VHA) et le norovirus (NoV),sont des causes importantes d’infections virales. Chaque année, celles-ci déduisent de grandes sommes monétaires aux systèmes de santé canadiens ainsi qu’aux producteurs alimentaires, sans compter les pertes alimentaires et économiques dues aux rappels d’aliments contaminés. Les stratégies de contrôle conventionnelles ne sont pas suffisamment efficaces et présentent, dans certains cas, des risques pour les travailleurs et/ou pour les consommateurs. Il est donc essentiel d’offrir de nouvelles méthodes d’inactivation naturelles et plus sécuritaires. Les produits issus de bleuet et de canneberge ont le potentiel de devenir ces nouvelles méthodes, notamment par leur concentration forte en polyphénols. De plus, ils sont naturels et sécuritaires pour les travailleurs et les consommateurs. Sans oublier qu’ils sont abondants au Québec, et, de ce fait, à moindres coûts. Le potentiel antiviral de ces produits de bleuet et de canneberge a été étudié et analysé dans ce mémoire envers des virus nus d’origine alimentaire (VHA et NVM-1) en plus de tester un modèle de virus enveloppé, soit le virus de l’herpès simplex de type 1(HSV-1). NVM-1 et VHS-1 ont été complètement inactivés à l’aide de jus de canneberge et/ou de bleuet (concentrés à 50 %) en 5 minutes de pré-incubation cellulaire ou virale respectivement. VHA a subi une réduction de 3,70 ± 0,54 log ufp/mL à la suite d’une préincubation virale de 90 minutes. / Foodborne viruses like the hepatitis A virus (HAV) and the norovirus (NoV) are known as important cause of viral infections. Each year, they cost massive amount to the Canadian health-care systems and to the producers without counting the important food loss due to recalls. The conventional methods are not sufficient and can represent in some case a threat to the worker or to the consumer, which is why it is essential to uncover new inactivation methods that are safer and natural, like berries. The blueberry and cranberry products have the potential to become these new methods considering their interesting composition. Furthermore, they are natural, do not show signs of threat to the worker nor to the consumer and above all they are abundant in Québec and at low cost. The antiviral potential of these products from blueberries and cranberries have been tested and studied in this memoir against the HAV and the MNV-1, naked foodborne viruses as well as against the herpes simplex virus type 1 (HSV-1), an enveloped model. The infectivity of MNV-1 was neutralized completely in the presence of this host cells at 37 °C in as little as 5 min of contact with cranberry or blueberry juices (50 % concentration). HSV-1 was also completely neutralized in viral pre-incubation with the same juice concentration and incubation. HAV infectious titer fell 3.70 ± 0.54 log pfu/mL after 90 minutes of contact with cranberry juice. These results suggest that cranberry and blueberry processing could be useful for reducing the transmission of foodborne viruses.

In allogeneic bone transplantation, the transmission of viral and non-viral infectious pathogens is the most severe undesirable concomitant phenomenon. The investigations published were examined regarding the inactivating capacity of inactivation procedures that are presently performed in bone banks (peracetic acid/ethanol, gamma irradiation, moist heat) against clinically relevant pathogens (aiming at a virus titer reduction of at least 4 log10 TCID50/ml or titer reduction of non-viral micro-organisms of at least 5 log10 cfu/ml). In the suspension experiments, treatment with peracetic acid/ethanol (peracetic acid 2%, ethanol 96%, aqua ad iniectabilia 2:1:1, 4 hours, 200 mbar, agitation) achieved a titer reduction of > 4 log10 already after 5 minutes for a number of viruses (PSR, PV, BVDV). HIV-2 was also inactivated within 5 minutes below the level of detection ( 4 log10 TCID50/ml was only reached after 4 hours. The results mentioned could be confirmed in the carrier test (contaminated spongiosa cuboids used as ‘worst case’ scenario). In the suspension experiment as well as in the carrier test, the HAV titer was reduced after 4 hours by only 3.7 log10 and 2.87 log10, respectively. The preceding step of defatting the spongiosa tissue by chloroform/ethanol was validated using cell-associated HAV and showed an HAV titer reduction of 7 log10. In the investigations regarding non-viral pathogens, all test organisms were completely inactivated by more than 5 log 10 steps (cfu/ml). Gamma irradiation was the second procedure examined. D10 values (irradiation dose required to reduce 90% of the pathogen titers by one log10 step) that were determined in inactivation kinetics experiments (irradiation conditions: –30°C, 60Co source) corresponded to data published so far. In order to provide for maximal safety, an irradiation dose of 34 kGy was recommended for allogeneic bone transplants using BPV and a diaphysis model from human femurs. The ‘Marburg bone bank system’ was the third procedure examined (thermal disinfection, guaranteed temperature of at least 82.5°C for a minimum of 15 min) using centrally contaminated human femoral heads. All viruses were completely inactivated and their titer reduced by more than 4 log10 steps. Vegetative bacteria and fungi were also completely inactivated (>= 6 log10 in the supernatant). As expected, spores and spore-forming pathogens were not sufficiently inactivated and not inactivated, respectively (titer reduction of less than 2 log10 cfu/ml). However, the latter group can be disregarded, since femoral heads are procured in the operation room under sterile conditions and the following production process rules out a secondary contamination with spores. It could be shown in the investigations presented that all three procedures examined guarantee an inactivation of the viruses investigated according to the recommendations by the senior federal authorities. The three treatment procedures offer additional biosafety by a comprehensive inactivation of non-viral pathogens.