Silicon Dioxide Impedes Antiviral Response and Causes Genotoxic Insult During Calicivirus Replication.

Noroviruses (NoV) are the leading cause of nonbacterial gastroenteritis in humans, and replicate extensively in the human gastrointestinal (GI) tract. Silica (also known as silicon dioxide, SiO2) nanoparticles (NPs) used in processed foods, dairy products, and beverages also accumulate in the GI tract. We investigated the effect of silica NPs on NoV replication and host cell response during virus infection, using murine norovirus (MNV-1) infection of RAW 264.7 murine macrophages. Pretreatment with 10 µg/ml silica significantly reduced the viability of macrophages, but no cumulative effects on viability of macrophages were observed with MNV-1 infection. No difference was observed between exposure to control or silica NPs on either the quantity of viral genome copies or the production of infectious virus in macrophages infected with MNV-1. Silica NPs reduced the ability of macrophages to upregulate genes encoding bone morphogenic proteins (BMPs), chemokine ligands and cytokines for which expression levels were otherwise found to be upregulated in response to MNV-1 infection. Furthermore, silica NPs reduced the levels of proinflammatory cytokines secreted by macrophages in response to MNV infection. Finally, silica NPs with MNV-1 infection produced a genotoxic insult to macrophages. Strikingly, this genotoxic insult was also found to occur as a synergistic effect of silica NPs and feline calicivirus infection in feline kidney epithelial cells. Taken together, our study suggests important safety considerations related to reducing exposure to silica NPs affecting the GI tract in individuals infected with NoVs and possibly other foodborne viruses.

Titanium Dioxide Nanoparticles Evoke Proinflammatory Response during Murine Norovirus Infection Despite Having Minimal Effects on Virus Replication.

Noroviruses (NoV) have enhanced tropism for the gastrointestinal (GI) tract and are the major cause of nonbacterial gastroenteritis in humans. Titanium dioxide (TiO2) nanoparticles (NPs) used as food additives, dietary supplements, and cosmetics accumulate in the GI tract. We investigated the effect anatase TiO2 NPs on NoV replication and host response during virus infection, using murine norovirus (MNV-1) infection of RAW 264.7 macrophages. Pretreatment with 20 µg/ml anatase NPs significantly reduced the viability of macrophages alone or during virus infection, but did not alter virus replication. In contrast, pre-incubation with 2 µg/ml anatase NPs reduced virus replication fivefold at 48 h. The presence of anatase NPs during MNV-1 infection evoked a pro-inflammatory response, as measured by a significant increase in expression of cytokines, including IL-6, IFN-γ, TNFα and the TGFß1. No genotoxic insults due to anatase TiO2 NPs alone or to their presence during MNV-1 infection were detected. This study highlights important safety considerations related to NP exposure of the GI tract in individuals infected with noroviruses or other foodborne viruses.

Infection of Murine Macrophages by Salmonella enterica Serovar Heidelberg Blocks Murine Norovirus Infectivity and Virus-induced Apoptosis.

Gastroenteritis caused by bacterial and viral pathogens constitutes a major public health threat in the United States accounting for 35% of hospitalizations. In particular, Salmonella enterica and noroviruses cause the majority of gastroenteritis infections, with emergence of sporadic outbreaks and incidence of increased infections. Although mechanisms underlying infections by these pathogens have been individually studied, little is known about the mechanisms regulating co-infection by these pathogens. In this study, we utilized RAW 264.7 murine macrophage cells to investigate the mechanisms governing co-infection with S. enterica serovar Heidelberg and murine norovirus (MNV). We demonstrate that infection of RAW 264.7 cells with S. enterica reduces the replication of MNV, in part by blocking virus entry early in the virus life cycle, and inducing antiviral cytokines later in the infection cycle. In particular, bacterial infection prior to, or during MNV infection affected virus entry, whereas MNV entry remained unaltered when the virus infection preceded bacterial invasion. This block in virus entry resulted in reduced virus replication, with the highest impact on replication observed during conditions of co-infection. In contrast, bacterial replication showed a threefold increase in MNV-infected cells, despite the presence of antibiotic in the medium. Most importantly, we present evidence that the infection of MNV-infected macrophages by S. enterica blocked MNV-induced apoptosis, despite allowing efficient virus replication. This apoptosis blockade was evidenced by reduction in DNA fragmentation and absence of poly-ADP ribose polymerase (PARP), caspase 3 and caspase 9 cleavage events. Our study suggests a novel mechanism of pathogenesis whereby initial co-infection with these pathogens could result in prolonged infection by either of these pathogens or both together.

Human Respiratory Coronaviruses Detected In Patients with Influenza-Like Illness in Arkansas, USA.

Acute respiratory viruses often result in significant morbidity and mortality. The potential impact of human respiratory coronavirus (CoV) infections was underestimated until the severe acute respiratory syndrome (SARS-CoV) outbreak in 2003, which showed that new, highly pathogenic coronaviruses could be introduced to humans, highlighting the importance of monitoring the circulating coronaviruses. The use of sensitive molecular methods has contributed to the differential diagnosis of viruses circulating in humans. Our study aim was to investigate the molecular epidemiology of human CoV strains circulating in Arkansas, their genetic variability and their association with reported influenza-like symptoms. We analyzed 200 nasal swab samples, collected by the Arkansas Department of Health in 2010, for influenza diagnosis. All samples were from patients showing acute respiratory symptoms while testing negative for influenza. Samples were pre-screened, using a quantitative reverse transcriptase polymerase chain reaction (qRT-PCR) multiprobe for coronavirus, and subjected to confirmatory pancoronavirus and/or strain-specific reverse transcriptase (RT)-PCR followed by sequence analysis. Seventy-nine samples (39.5%) were positive by qRT-PCR and 35 samples (17.5%) were confirmed by conventional RT-PCR. Twenty-three of the confirmed samples (59%) were sequenced. The most frequent strain detected was HCoV-OC43-like followed by NL63-like; only one sample was positive for HCoV-229E and one for HCoV-HKU1. Feline-like CoV strains were detected in three samples, representing possible evidence of interspecies transmission or a new human strain. Seventeen percent of the coronavirus positive samples were also positive for other respiratory viruses, such as Respiratory Syncytial Virus (RSV), Parainfluenza 2 and 3, and Rhinovirus. Thus, HCoV-OC43, NL63, HKU1 and new feline-like strains were circulating in Arkansas in 2010. HCoV was the sole respiratory virus detected in 16% of the patients who showed acute respiratory symptoms with negative diagnoses for influenza virus.

Effects of dietary vitamin A content on antibody responses of feedlot calves inoculated intramuscularly with an inactivated bovine coronavirus vaccine.

OBJECTIVE: To investigate effects of low dietary vitamin A content on antibody responses in feedlot calves inoculated with an inactivated bovine coronavirus (BCoV) vaccine. ANIMALS: 40 feedlot calves. PROCEDURES: Calves were fed diets containing high (3,300 U/kg) or low (1,100 U/kg) amounts of vitamin A beginning on the day of arrival at a feedlot (day 0) and continuing daily until the end of the study (day 140). Serum retinol concentrations were evaluated in blood samples obtained throughout the study. Calves were inoculated IM with an inactivated BCoV vaccine on days 112 and 126. Blood samples obtained on days 112 and 140 were used for assessment of BCoV-specific serum IgG1, IgG2, IgM, and IgA titers via an ELISA. RESULTS: The low vitamin A diet reduced serum retinol concentrations between days 112 and 140. After the BCoV inoculation and booster injections, predominantly serum IgG1 antibodies were induced in calves fed the high vitamin A diet; however, IgG1 titers were compromised at day 140 in calves fed the low vitamin A diet. Other isotype antibodies specific for BCoV were not affected by the low vitamin A diet. CONCLUSIONS AND CLINICAL RELEVANCE: Dietary vitamin A restriction increases marbling in feedlot cattle; however, its effect on antibody responses to vaccines is unknown. A low vitamin A diet compromised the serum IgG1 responses against inactivated BCoV vaccine, which suggested suppressed T-helper 2-associated antibody (IgG1) responses. Thus, low vitamin A diets may compromise the effectiveness of viral vaccines and render calves more susceptible to infectious disease.

Human rotavirus virus-like particle vaccines evaluated in a neonatal gnotobiotic pig model of human rotavirus disease.

The authors discuss here rotavirus-like particle vaccines as an alternative approach to oral live-attenuated rotavirus vaccine and their efficacy in a gnotobiotic pig model. Rotavirus virus-like particles (VLPs) were evaluated in different doses, and routes of administration, and combined with live-attenuated virus and adjuvants. A VLP vaccine composed of rotavirus VP2 and VP6 was immunogenic in gnotobiotic pigs when inoculated intranasally; however, this vaccine failed to confer protection. A combination of oral attenuated human rotavirus and intranasal 2/6VLP vaccines conferred immunogenicity, partial protection against a human rotavirus challenge and induced IFN-γ-producing T cells in the ileum of pigs with similar frequencies to human rotavirus infection. Vaccination through a combination of mucosal inductive sites and live-attenuated vaccine combined with VLP vaccines was the most effective regimen, compared with the use of a single route or a single vaccine alone. However, if formulated with neutralizing antigens, VLP vaccines may constitute a better approach in a high maternal antibody scenario.