Enteric microbiota influences enteric viral infections, and host response to these pathogens and vaccines. Using gnotobiotic (Gn) pigs transplanted with human gut microbiota (HGM), we studied the effects of HGM on the immune response to oral rotavirus vaccination and rotaviral disease. We also used HGM transplanted Gn pigs to determine the effects of HGM on human norovirus infection. Despite commercially available vaccines, human rotavirus is a leading acute gastroenteritis in children, especially those in developing countries. Human norovirus (HuNoV) is a leading cause of acute gastroenteritis in all age groups worldwide, and no vaccines are commercially available. Further understanding of how enteric microbiota influences these viral diseases may identify therapeutic targets. In our rotavirus study, pigs were colonized with HGM from an infant with low fecal concentrations of enteropathy biomarkers and responded well to their first dose of oral rotavirus vaccine (healthy human gut microbiota "HHGM"); or pigs were colonized with HGM from an infant with high fecal concentrations of enteropathy biomarkers and a poor response to the first dose of oral rotavirus vaccine (unhealthy human gut microbiota "UHGM"). HHGM colonized pigs had stronger cell-mediated and mucosal immune response to oral rotavirus vaccine compared to UHGM pigs based on the number of rotavirus-specific IFN-γ producing T cells in the ileum, spleen, and blood, and trends towards higher rotavirus specific antibody titers in intestinal contents, respectively. Significant correlations between multiple Operational Taxonomic Units (OTUs) of bacteria and frequencies of IFN-γ producing T cells at the time of human rotavirus challenge existed, suggesting that certain members of the microbiota influenced the immune response to the vaccine. After the vaccinated pigs were challenged with human rotavirus, HHGM pigs had less severe and shorter duration of viral shedding and diarrhea compared to UHGM pigs, suggesting that HHGM facilitated development of stronger protective immunity. These results demonstrated that composition of the enteric microbiota influenced host immune response to oral vaccination. In the norovirus study, Gn pigs were colonized with HHGM to determine the effects of microbiota on HuNoV infection. Colonized pigs shed more virus for a longer duration than non-colonized pigs, and also had higher viral titers in the duodenum and distal ileum. Diarrhea was more severe 4-10 days post-infection and lasted longer in colonized compared to non-colonized pigs. Twenty-seven genes related to the immune system were highly upregulated in HuNoV infected, colonized pigs compared to non-colonized controls. These result showed that HHGM influenced infectivity of HuNoV in the Gn pig model and altered host gene expression related to the immune system. These studies showed that HHGM can improve the host immune response and efficacy of rotavirus vaccine, but it can also enhance infection and clinical disease in HuNoV infected Gn pigs. Depending on the virus, gut microbiota may be beneficial or detrimental to the host. Those developing future treatments aimed at altering microbiota to prevent or ameliorate one viral pathogen need to consider the potential for enhancing a different pathogen. These studies demonstrated the usefulness of HGM transplanted Gn pigs for evaluation of microbiota influence on infection and immunity of enteric viral pathogens. / Ph. D. / Gut microbiota influences intestinal viral infections, and host response to these pathogens and vaccines. Using gnotobiotic (Gn) pigs transplanted with human gut microbiota (HGM), we studied the effects of HGM on the immune response to oral rotavirus vaccination and rotaviral disease. We also used HGM transplanted Gn pigs to determine the effect of HGM on human norovirus infection. Despite commercially available vaccines, human rotavirus is a leading acute gastroenteritis in children, especially those in developing countries. Human norovirus (HuNoV) is a leading cause of vomiting and diarrhea in all age groups worldwide, and no vaccines are commercially available. Further understanding of how gut microbiota influences these viral diseases may identify therapeutic targets. In our rotavirus study, pigs were colonized with HGM from an infant without evidence of intestinal disease based on fecal analysis, and who responded well to the first dose of oral rotavirus vaccine (healthy human gut microbiota “HHGM”); or pigs were colonized with HGM from an infant with evidence of potential intestinal dysfunction and a poor response to the first dose of oral rotavirus vaccine (unhealthy human gut microbiota “UHGM”). HHGM colonized pigs had a stronger immune response to the oral rotavirus vaccine compared to UHGM pigs. Significant correlations between multiple Operational Taxonomic Units (OTUs) of bacteria and frequencies of rotavirus-specific immune cells at the time of human rotavirus challenge existed, suggesting that certain members of the microbiota influenced the immune response to the vaccine. After the vaccinated pigs were challenged with human rotavirus, HHGM pigs had less severe and shorter duration of viral shedding and diarrhea compared to UHGM pigs, suggesting that HHGM enhanced vaccine efficacy. These results demonstrated that composition of the gut microbiota influenced host immune response to oral vaccination. In the norovirus study, GN pigs were colonized with HHGM to determine the effects of microbiota on HuNoV infection. Colonized pigs shed more virus for a longer duration than non-colonized pigs, and also had higher viral titers in sections of small intestine. Diarrhea was more severe 4-10 days after infection and lasted longer in colonized compared to non-colonized pigs. Twenty-seven genes related to the immune system were highly upregulated in HuNoV infected, colonized pigs compared to controls. These result showed that HHGM influenced infectivity of HuNoV in the Gn pig model and altered host gene expression related to the immune system. These studies showed how HHGM improved the host immune response and efficacy of rotavirus vaccine, but conversely enhanced infection and clinical disease in HuNoV infected pigs. Depending on the virus, gut microbiota may be beneficial or detrimental to the host. Those developing future treatments aimed at altering microbiota to prevent or ameliorate one viral pathogen need to consider the potential for enhancing a different pathogen. These studies showed the usefulness of HGM transplanted Gn pigs for evaluation of microbiota influence on infection and immunity of intestinal viruses.

Introduction: Acute gastroenteritis is a common illness in humans. It has a high morbidity worldwide and in the United States (U.S); a sizeable mortality is reported in developing countries. Viruses are a common cause of acute gastroenteritis and noroviruses are the leading cause of non-bacterial gastroenteritis (1). Fomites play an important role in the infection cycle of norovirus. This study explored the comparability between a quantitative microbial risk assessment (QMRA) model to predict the probability of illness from norovirus due to environmental contamination with actual observed attack rates, the impact of appropriate cleaning procedures on fomites contamination, and the role of fomites in the transmission of norovirusMethod: Data from three different norovirus illness outbreaks occurring from 2004 to 2005 in Arizona in different venues were used. Questionnaires were used to determine demographics, symptoms associated with gastrointestinal illness, medical visits, and potential exposures such as food items consumed. When available, stool samples were collected from ill participants. Fomites samples were also gathered from the different venues; houseboats, college summer camp living spaces, and a restaurant. These samples were tested for norovirus using reverse transcriptase polymerase chain reaction (RT-PCR). Viral concentration on fomites was calculated using the most probable number methodology.Results: Norovirus was found on fomites from all three outbreaks. Improper cleaning of fomites increased the proportion of norovirus positive fomites from 40 percent to 70 percent in contrast, after cleaning and disinfection with 5000 mg/L free chloride, the proportion of norovirus positive fomites decreased to 33%. The predicted values from the quantitative microbial risk assessment model were consistent with the observed epidemiological attack rates.Conclusion: This study showed unequivocally that fomites play an important role in the transmission cycle of norovirus, and that improper cleaning and disinfection procedures only serve to spread the virus to previously uncontaminated fomites. In the case of outbreaks where people are not always available to answer questionnaires, QMRA should be considered as a worthwhile alternative to a full-scale epidemiological study.

Norovirus is the most common pathogen to cause acute gastroenteritis in the world. Symptoms of acute gastroenteritis include vomiting and/or diarrhea, along with fever, abdominal pain, and malaise. Annually, norovirus causes 685 million cases of acute gastroenteritis and 200,000 deaths, worldwide. Among the 685 million cases occurring every year, 19-21 million occur in the United States. Norovirus can spread through direct or indirect contact (e.g., contaminated food or water). In addition, recent evidence has suggested that norovirus can also be spread via aerosolization. However, no study has determined an indoor generation source for aerosolized norovirus. Therefore, the goals of this study were to optimize sampling and quantification methods for the collection of aerosolized norovirus. Upon optimization, the last was to investigate a potential indoor generation source (i.e., toilet flushing) of aerosolized norovirus. To achieve this goal we devised three studies. In the first study, we optimized a sampling method for the collection of aerosolized norovirus using murine norovirus (MNV) as a surrogate. Optimization of the sampling method was performed using two bioaerosol samplers (SKC BioSampler and the National Institute for Occupational Safety and Health [NIOSH] Bioaerosol Cyclone Sampler 251) and two sampling media (Hanks Balanced Salt Solution [HBSS] and Phosphate Buffered Saline [PBS]). Murine norovirus was aerosolized in a bioaerosol chamber and later collected using the optimized sampler/media combination. After collection, viral RNA was extracted from MNV collected samples and quantified using quantitative polymerase chain reaction (qPCR). Intact capsids of MNV were assessed using propidium monoazide dye in combination with qPCR and confirmed with transmission electron microscopy. There were a total of 10 trials conducted, with each trial lasting for 30 minutes. The SKC BioSampler collected a significantly higher concentration of MNV than the NIOSH-251 sampler did (p-value < 0.0001). However, there were no significant differences in the relative percent of MNV that remained viable between both samplers (p-value = 0.2215). The use of HBSS sampling media yielded a higher concentration of MNV than PBS media (p-value = 0.0125). However, PBS media maintained viability at a significantly higher percentage than HBSS media (p-value < 0.0001). The results support the optimization of a sampling method for the collection of aerosolized MNV and possibly norovirus in different sampling environments. In the second study, we optimized the quantification method for MNV. A relatively new quantification system, droplet digital polymerase chain reaction (ddPCR), was evaluated using the same extracted samples collected in the first study to determine if the same overall outcome could be achieved. In addition, a MNV standard was directly compared between the qPCR and ddPCR. When comparing the same standard, the mean observed concentrations were similar to the nominal concentration. The limit of detection for both instruments was 5 copies per reaction. The coefficient of variation was lower across all ddPCR results than the qPCR results. The range of the R2 was larger for ddPCR compared to qPCR. As for the analysis of bioaerosol samples collected from the first study, the SKC BioSampler collected a significantly higher concentration of MNV compared to the NIOSH-251 sampler (p-value = 0.0002). However, there were no significant differences in the relative percent of MNV that remained viable in both samplers (p-value = 0.6734). The use of HBSS sampling media yielded a higher concentration of MNV than PBS media (p-value = 0.0190). However, PBS media maintained viability at a significantly higher percentage than HBSS media (p-value = 0.0004). The use of ddPCR allows for a simpler workflow and fewer samples and resources. These results support that both PCR systems yield similar results and overall outcomes, thus presenting an optimized quantification method for MNV. In the third study, we used the optimized sampling and quantification methods to conduct a field trial investigation of a potential indoor aerosolization source for norovirus (toilet flushing). To inform bioaerosol sampler placement, two optical particle counters monitored particle size and number distribution of aerosol produced from flushing a toilet across three variables (height, position, and side). The location with the highest mean particle concentration, and where bioaerosol sampling occurred, was behind the toilet and 0.15 m above the toilet bowl rim. A flushometer type toilet was seeded with 105 and 106 PFU/mL of MNV and then flushed. Upon flushing, a SKC BioSampler and Coriolis µ sampler were activated to collect aerosolized MNV. Samples were extracted and then quantified using RT-ddPCR, and viability was quantified using PMA: RT-ddPCR. The concentration of MNV collected after seeding the toilet water ranged from 2.18 x 105 – 9.65 x 106 total copies of MNV. Positive samples of airborne MNV were detected using the Coriolis µ sampler with collected concentrations ranging from 383 – 684 RNA copies/m3 of air. Sample viability for bioaerosol samples were unable to be quantified. The relative percent of MNV virions that remained intact in seeded toilet water was 37-79%. This study provides the first evidence that MNV, a NV surrogate, can be aerosolized when a toilet is flushed.

This work comprises three projects that extend previous models to include features of practical significance for the statistical analysis of infectious disease data. In the first, we find from a simulation study how the degree of heterogeneity in the number contacts that individuals have affects the relationship between estimates of a pathogen's effective population size based on coalescent theory and the true prevalence and incidence of that pathogen. In the second, we find that aggregating data from many small outbreaks allows the parameters of stochastic epidemic models to be consistently estimated with a generalized linear model. Application of this method to a set of 77 small norovirus outbreaks reveals interesting differences in the transmission parameters between hospital and nursing-home outbreaks. In the third project, we gain insight into HIV contact networks in the United States by fitting data from a number of surveys to a simple stochastic model of a dynamic network. / text

Noroviruses are the leading cause of gastroenteritis in the United States, causing an estimated 23 million illnesses, 50,000 hospitalizations and 300 deaths per year. This virus is transmitted via the fecal-oral route and infections can occur from drinking contaminated water or eating contaminated food, contact with contaminated fomites, inhalation of aerosolized vomitus, or person to person spread. Outbreaks of norovirus following exposure to contaminated recreational water have been documented, as swimmers and others engaged in water recreation often consume recreational water, either purposefully or accidentally, during their activities. In Arizona there have been several outbreaks of norovirus gastroenteritis among people using the recreational waters. In the past four years there have been four consecutive outbreaks of norovirus among rafters on the Colorado River, and in 2003 there was an outbreak of norovirus among a large group of houseboaters on Lake Powell.Norovirus is an emerging pathogen in the state of Arizona. In an effort to better understand its epidemiology, and with the aim of providing recommendations for prevention or minimization of future outbreaks, two investigations were undertaken: an investigation of the outbreak of norovirus among houseboaters on Lake Powell, and a survey of water quality and possible sources of norovirus in the Colorado River. Additionally, historical Colorado River water quality data was reviewed and outbreak epidemiology assessed. To better understand the role of viruses in waterborne recreational disease outbreaks, a review of the literature was conducted and an analysis of 48 recreational waterborne disease outbreaks was done.

Shellfish are an important food source however they are known to harbour bacteria, viruses and toxic chemicals that can be detrimental to their human consumers. Oysters have been associated with the gastroenteritis virus Norovirus. New Zealand has some of the highest cases of foodborne illness in the western world. This study investigated a possible link between periods of high rainfall and reported Norovirus outbreaks in four major cities in New Zealand (Auckland, Wellington, Christchurch and Dunedin) as well as national data. Norovirus is a highly infectious foodborne illness. Outbreaks of norovirus have been linked to the consumption of shellfish, and in particular oysters. Norovirus virions can enter the aquatic environment via sewage as a result of human shedding of the virus. This investigation into rainfall and Norovirus outbreaks found no statistically significant relationship, in a monthly or season setting. In addition the relationship between environmental cadmium levels and exposure levels in New Zealand was investigated through meta-analysis. Cadmium is a heavy metal commonly associated with the mining of copper and zinc ores. It is found naturally in the environment, in air land and oceans Increased exposure to cadmium is known to have a number of serious detrimental health effects, in particular this study investigates cadmiums immunosuppressive properties. Concentrations in New Zealand were compared with Canada, Italy and the UK to determine if New Zealand has a relatively high cadmium intake. Interestingly environmental levels (soil and oceanic) in New Zealand were low. Cadmium levels were higher in oysters than in mussels, with New Zealand oysters containing the highest concentration of cadmium presented. New Zealanders also had the highest cadmium burdens in the kidneys and the highest daily intakes. A No Observable Effect Level (NOEL) was calculated from mice data and compared to the daily intakes of the four countries. Both Canada and New Zealand were above this level. Shellfish are a common mechanism for exposure to both Norovirus and cadmium. The levels of cadmium present in the diet of New Zealanders may be sufficiently high to suppress the immune system, making it more vulnerable to infections of enteric diseases such as Norovirus.

GONDIM, R. N. D. G. Diversidade genética de norovírus, co-infecções e estado nutricional de crianças em estudo caso-controle no semiárido brasileiro. 2017. 105 f. Dissertação (Mestrado em Ciências Médicas) - Faculdade de Medicina, Universidade Federal do Ceará, Fortaleza, 2017. / Submitted by Ivone Sousa (ppgcm.ufc@gmail.com) on 2017-08-18T15:44:28Z No. of bitstreams: 1 2017_dis_rndgg.pdf: 3249367 bytes, checksum: 2b719da4016b1f1633735600acd82f2f (MD5) / Approved for entry into archive by Erika Fernandes (erikaleitefernandes@gmail.com) on 2017-08-21T11:27:08Z (GMT) No. of bitstreams: 1 2017_dis_rndgg.pdf: 3249367 bytes, checksum: 2b719da4016b1f1633735600acd82f2f (MD5) / Made available in DSpace on 2017-08-21T11:27:08Z (GMT). No. of bitstreams: 1 2017_dis_rndgg.pdf: 3249367 bytes, checksum: 2b719da4016b1f1633735600acd82f2f (MD5) Previous issue date: 2017-07-26 / Norovirus (NoV) infection is a major global health problem with high incidence morbid-mortality rates. However, the impact of NoV on child development remains poorly described. We wanted to investigate the genetic diversity of circulating NoV strains in children from the semiarid region of Brazil, and its associations with other co-infections and clinical/nutritional status. This case-control study was conducted in six different cities with children aged 2-36 months during the period from April 2010 to March 2011. Cases considered were children who had at least one episode of diarrhea in the last 14 days. Detection and quantification of NoV were performed by RT-qPCR, followed by molecular and phylogenetic analysis. The presence of NoV was 45.2% (75/166), and ranged from 8–77.7% among the cities examined. A lower z-score index was associated with the presence of NoV with weight for age – WAZ (P = 0.034), weight for height – WHZ (P= 0.033), and body mass index for age – BAZ (P = 0.033). In addition, NoV infection was associated with more frequent respiratory symptoms (P= 0.0096). GII.P7 (polymerase genotype) and GII.3 (capsid genotype) were the most frequent NoV genotypes, and analysis of the ORF 1-2 junction identified recombinant NoV strains in 80% (12/15) of the sequenced samples. With regards to co-infections, infections with enteroaggregative Escherichia coli (P = 0.0046) were more frequent in the absence of NoV, whereas in the presence of Shigella a positive correlation was observed with NoV (P = 0.0199). Among the positive cases of NoV, diarrheal episodes were associated with the presence of entero-aggregative E. coli, and simultaneous absence of Cryptosporidium and shiga toxin-producing E. coli (P = 0.0167). The present study highlighted the high genetic variability of NoV in the semiarid region of Brazil, and underlines its role in co-infections and infant nutritional status, thus contributing to better understandings of its impact on infant development and its / A infecção por Norovirus (NoV) é um importante problema de saúde pública mundial devido a altas taxas de morbi-mortalidade. No entanto, o impacto ocasionado pelo NoV no desenvolvimento infantil permanecem pouco descritos. O presente estudo investigou a diversidade genética de cepas de NoV circulantes em crianças no semiárido brasileiro e suas associações como o estado clínico/nutricional e coinfecções. O estudo caso-controle foi realizado em seis diferentes cidades com crianças de 2-36 meses de idade durante o período de abril de 2010 a março de 2011. Foram considerados casos, as crianças que tiveram pelo menos um episódio de diarréia nos últimos 14 dias. A detecção e quantificação de NoV foram realizadas por RT-qPCR, seguidas de análises moleculares e filogenéticas. A taxa de detecção de NoV foi de 45,2% (75/166) e variou de 8% a 77,7% entre as cidades avaliadas. Um menor índice de escores z foi associado a presença de NoV, em peso por idade – WAZ (P = 0,034), peso por altura – WHZ (P= 0,033) e índice de massa corporal por idade – BAZ (P = 0,033). Além disso, a infecção por NoV foi associada à sintomas respiratórios mais freqüentes (P = 0,0096). GII.P7 (genótipo da polimerase) e GII.3 (genótipo do capsídeo) foram os genótipos de NoV mais freqüentes, e a análise da junção de ORF 1-2 identificou cepas recombinantes de NoV em 80% (12/15) das amostras sequenciadas. No que diz respeito às co-infecções, observou-se que as infecções por Escherichia coli enteroagregativa (P = 0,0046) foram mais frequentes na ausência de NoV, enquanto que na presença de Shigella observou-se uma correlação positiva com NoV (P = 0,0199). Entre os casos positivos para NoV, os episódios diarréicos foram associados à presença de E. coli enteroagregativa e ausência simultânea de Cryptosporidium e E. coli produtora de shiga toxina (P = 0,0167). O presente estudo demonstrou alta variabilidade genética de NoV circulantes no semiárido brasileiro e ressalta seu papel em relação a avaliação do estado nutricional infantil e coinfecções, contribuindo assim para uma melhor compreensão do seu impacto no desenvolvimento infantil e da sua epidemiologia.

Hajj is the annual gathering of Islam practitioners in Mecca, Saudi Arabia. During the event, gastrointestinal infections are usually experienced and outbreaks have always been a concern; nevertheless, a deep and integrative study of the etiological agents has never been carried out. Here, I describe for the first time the epidemiology of pathogenic enteric viruses during Hajj 2011, 2012 and 2013. The focus of this study was the common enteric viruses Astrovirus, Norovirus, Rotavirus and Adenovirus. An enzyme Immunoassay established their presence in 14.9%, 15.0% and 6.6% of the reported cases of acute diarrhea for 2011, 2012 and 2013, respectively. For the three years of study, Astrovirus accounted for the majority of the viral infections. To our knowledge, this is the first time an epidemiological study depicts Astrovirus as the main viral agent of gastroenteritis in a mass gathering event. Hajj is rich in strains of Astrovirus, Norovirus and Rotavirus. A first screening by RT-PCR resulted in ten different genotypes. Strains HAstV 2, HAstV 1 and HAstV 5 were identified for Astrovirus. GI.6, GII.3, GII.4 and GII.1 were described for Norovirus and G1P[8], G4P[8] and G3P[8] were found for Rotavirus. The majority of the Astrovirus isolates could not be genotyped suggesting the presence of a new variant(s). Cases like this encourage the use of metagenomics (and nextgeneration sequencing) as a state-of-the-art technology in clinical diagnosis. A sample containing Adenovirus particles is being used to standardize a process for detection directly from stool samples and results will be obtained in the near future. The overall findings of the present study support the concept of Hajj as a unique mass gathering event that potentiates the transmission of infectious diseases. The finding of Norovirus GII.4 Sydney, a variant originated from Australia, suggests that Hajj is a receptor of infectious diseases worldwide. This work is part of the Hajj project, a collaborative effort with the Ministry of Health of the Kingdom of Saudi Arabia in order to describe entirely the epidemiology of gastrointestinal diseases in Hajj. It is expected that the results of this study will serve in the refinement of public health policies.

Les norovirus sont les principaux agents responsables de gastroentérites humaines non bactériennes dans le monde affectant tous les âges. Leur incidence croissante serait liée à leur capacité d’adhésion et de leur persistance dans la chaine alimentaire. Cependant, le mécanisme de cette adhésion demeure inconnu. L’objectif de ce doctorat était d’étudier et de caractériser les norovirus sur les plans structural, moléculaire et fonctionnel afin d’élucider leur mécanisme d’adhésion aux surfaces. Des prototypes «virus-like particle» (VLPs) de norovirus humains des génogroupes GI.1, GII.4 et du calivirus félin (FCV) ont été produits par culture cellulaire, purifiés par ultracentrifugation et caractérisés en fonction du pH, de la force ionique et de la température. Pour cette caractérisation, les charges électriques et les tailles des VLPs ont été analysées en utilisant un Zeta NanoSizer ZS. Les structures secondaires et tertiaires ainsi que leur stabilité ont été analysées par les techniques de dichroïsme circulaire et de fluorescence UV, respectivement. Les énergies de surfaces des VLPs et des surfaces inertes et alimentaires, de même que les énergies libres d’interaction interfaciale entre les VLPs et les surfaces ont été estimées après des mesures d’angle de contact au goniomètre. Enfin, des essais d’adhésion de VLPs de norovirus GII.4 ont permis de faire des corrélations entre les résultats de ces adhésions et les changements structuraux, moléculaires et fonctionnels. Les données ont révélé que l’adhésion des VLPs de GII.4 dépendrait plutôt des changements structuraux moléculaires et fonctionnels des VLPs de GII.4 que des propriétés thermodynamiques des surfaces. Les maxima d’adhésion observés au point isoélectrique avec des diminutions aux pH élevés et les augmentations avec la force ionique ont démontré respectivement, l’importance des interactions acide-bases (hydrophobes) et des forces de van der Waals dans le phénomène d’adhésion. Sur le polyéthylène et la laitue, l’adhésion semble être régie par les interactions hydrophobiques, tandis que sur l’acier inoxydable, elle est apparemment régie par les interactions de van der Waals. À basse température, elle semble reposer sur les résidus hydrophobes à la surface de la capside alors qu’à hautes températures, un déploiement de résidus hydrophobes internes après dénaturation des structures secondaires et tertiaires semble probable dans le phénomène. Le type d’interactions majoritairement impliquées (hydrophobiques) invite à l’utilisation d’agents chaotropiques pour une meilleure élimination des norovirus adhérés et leur contrôle dans le secteur agroalimentaire. / Human noroviruse has been the main cause of acute non-bacterial gastroenteritis in the world through all ages. Their growing incidence has been repeatedly associated with their ability to bind and persist on agrifood surfaces in food processing environments. However, the mechanism of their adhesion phenomenon remains unknown. This study was aimed at identifying structural, molecular and functional elements of human norovirus involved in their adhesion phenomenon. Prototypes as «virus-like particle» (VLPs) have been produced for human noroviruses GI.1 and GII.4 and feline calicivirus (FCV) using cell culture, purified by ultracentrifugation, and characterized with aspect to pH, ionic strength and temperature. In this characterization, their electrical charges and their size were analyzed using a Zeta NanoSizer ZS and their secondary and tertiary structures and stability were analyzed using circular dichroism and intrinsic fluorescence UV techniques, respectively. Their surface energies as well as surface energies of inert surfaces and fresh foods were estimated applying contact angle technic with a goniometer. The total interfacial free energy of interactions between the VLPs and the surfaces were estimated, and GII.4 VLPs adhesion assays perfomed in order to draw correlation between adhesion and structural, molecular and functional changes. Our results revealed that GII.4 VLPs adhesion was rather related to structural, molecular and functional changes than surface thermodynamic proprieties. Maxima observed at isoelectric point followed by decreases with pH and the increasing trend with increasing ionic strength indicate respectively the importance of acid base (hydrophobic) interactions and of van der Waals interactions in adhesion phenomenon. The adhesion on polyethylene and lettuce are seemingly related to hydrophobic interactions, while the adhesion on stainless steel is apparently controlled by van der Waals interactions. At low temperature, adsorption may rely on capsid external hydrophobic residues, while at high temperature, internal hydrophobic residues may plausibly play a part in the process upon denaturation of secondary and tertiary structures. The interactions mainly involved in GII.4 VLPs adsorption are hydrophobic and suggest the use of chaotropic agents to break efficiently adhesion mechanism and limit norovirus outbreaks.

No description available.