The Effects of Simvastatin or Interferon-α on Infectivity of Human Norovirus Using a Gnotobiotic Pig Model for the Study of Antivirals

Jung, Kwonil; Wang, Qiuhong; Kim, Yun-Jeong; Scheuer, Kelly; Zhang, Zhenwen; Shen, Quan; Chang, Kyeong‐Ok; Saif, Linda J.

doi:10.1371/journal.pone.0041619

Cited by 68 publications

(104 citation statements)

References 53 publications

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“…Swine have HBGA phenotypes similar to those of humans, making gnotobiotic piglets an excellent model with which to study human NoVs (23-25, 35, 36). Previous studies have shown that gnotobiotic piglets are susceptible to oral infection with several human NoV GII.4 strains, and this animal model has been used for evaluation of the efficacy of vaccine candidates and antiviral therapies against human NoVs (23)(24)(25)37). We found that gnotobiotic piglets inoculated with human NoV GII.4 strain 765 exhibited mild diarrhea, viral RNA shedding in feces, histologic lesions in the intestines, and viral replication in the proximal small intestine.…”

Section: Discussionmentioning

confidence: 73%

“…Second, we determined the infectivity of human NoV following HPP treatment in a gnotobiotic pig model. The gnotobiotic pig is an excellent model for human enteric viruses because pigs share many similarities with humans in gastrointestinal structure, physiology, immunology, and, more importantly, the HBGA phenotypes (types A and H) on enterocytes (23)(24)(25)(26)(27). We found that HPP treatment at 350 MPa for 2 min at an initial temperature of 0°C reduced the amount of human NoV in oysters by approximately 3.7 log 10 units, as determined by a combination of a receptor binding assay and real-time reverse transcription-PCR (RT-PCR).…”

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confidence: 99%

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A Gnotobiotic Pig Model for Determining Human Norovirus Inactivation by High-Pressure Processing

Lou

et al. 2015

Appl Environ Microbiol

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e Human norovirus (NoV) is responsible for over 90% of outbreaks of acute nonbacterial gastroenteritis worldwide and accounts for 60% of cases of foodborne illness in the United States. Currently, the infectivity of human NoVs is poorly understood due to the lack of a cell culture system. In this study, we determined the survival of a human NoV genogroup II, genotype 4 (GII.4) strain in seeded oyster homogenates after high-pressure processing (HPP) using a novel receptor binding assay and a gnotobiotic pig model. Pressure conditions of 350 MPa at 0°C for 2 min led to a 3.7-log 10 reduction in the number of viral RNA copies in oysters, as measured by the porcine gastric mucin-conjugated magnetic bead (PGM-MB) binding assay and real-time RT-PCR, whereas pressure conditions of 350 MPa at 35°C for 2 min achieved only a 1-log 10 reduction in the number of RNA copies. Newborn gnotobiotic piglets orally fed oyster homogenate treated at 350 MPa and 0°C for 2 min did not have viral RNA shedding in feces, histologic lesions, or viral replication in the small intestine. In contrast, gnotobiotic piglets fed oysters treated at 350 MPa and 35°C for 2 min had high levels of viral shedding in feces and exhibited significant histologic lesions and viral replication in the small intestine. Collectively, these data demonstrate that (i) human NoV survival estimated by an in vitro PGM-MB virus binding assay is consistent with the infectivity determined by an in vivo gnotobiotic piglet model and (ii) HPP is capable of inactivating a human NoV GII.4 strain at commercially acceptable pressure levels. Human norovirus (NoV), a member of the Caliciviridae family, is responsible for over 90% of the outbreaks of acute nonbacterial gastroenteritis worldwide and accounts for more than 60% of the cases of foodborne illness in the United States (1, 2). It is estimated that 48 million individuals, or about 17% of the U.S. population, are sickened each year, leading to approximately 128,000 hospitalizations and 3,000 fatalities in the United States (2). Human NoV is transmitted primarily through the fecal-oral route either by direct person-to-person contact or by fecally contaminated food or water. Human NoV is highly contagious and stable, and only a few virus particles are thought to be sufficient to cause an infection (3, 4). Outbreaks frequently occur in restaurants, hotels, day care centers, schools, nursing homes, cruise ships, swimming pools, hospitals, and military installations. Despite the significant economic impact and high morbidity caused by human NoV, no vaccines or antiviral drugs with activity against this virus are currently available (5, 6). This is due in large part to the lack of a cell culture system and a small-animal model for human NoV (6, 7). As a consequence, the survival of human NoV is poorly understood.A major high-risk food for human NoV contamination is seafood, particularly bivalves, such as oysters, mussels, and clams (8-11). These animals are filter feeders and can readily bioaccumulate human NoV in their tissue...

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Section: Discussionmentioning

confidence: 73%

mentioning

confidence: 99%

A Gnotobiotic Pig Model for Determining Human Norovirus Inactivation by High-Pressure Processing

Lou

et al. 2015

Appl Environ Microbiol

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“…The study of human noroviruses (HuNV) has been impeded by the absence of a viable cell culture system for this pathogen and the fact that baby gnotobiotic pigs are the only available smallanimal model (4,5). As a result, other caliciviruses have often been used as surrogate models for the investigation of HuNV biology (6).…”

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confidence: 99%

Structures of the Compact Helical Core Domains of Feline Calicivirus and Murine Norovirus VPg Proteins

Leen

Kwok

Birtley

et al. 2013

J Virol

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fWe report the solution structures of the VPg proteins from feline calicivirus (FCV) and murine norovirus (MNV), which have been determined by nuclear magnetic resonance spectroscopy. In both cases, the core of the protein adopts a compact helical structure flanked by flexible N and C termini. Remarkably, while the core of FCV VPg contains a well-defined three-helix bundle, the MNV VPg core has just the first two of these secondary structure elements. In both cases, the VPg cores are stabilized by networks of hydrophobic and salt bridge interactions. The Tyr residue in VPg that is nucleotidylated by the viral NS7 polymerase (Y24 in FCV, Y26 in MNV) occurs in a conserved position within the first helix of the core. Intriguingly, given its structure, VPg would appear to be unable to bind to the viral polymerase so as to place this Tyr in the active site without a major conformation change to VPg or the polymerase. However, mutations that destabilized the VPg core either had no effect on or reduced both the ability of the protein to be nucleotidylated and virus infectivity and did not reveal a clear structure-activity relationship. The precise role of the calicivirus VPg core in virus replication remains to be determined, but knowledge of its structure will facilitate future investigations.

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“…The recombinant baculovirus (rBac) carrying the VP1 gene (ORF2) of the Hu/NoV/GII.4/HS194/2009/US strain (GenBank accession number GU325839) was produced using the BaculoDirect baculovirus expression system (Invitrogen, Carlsbad, CA). The Gateway entry clone containing the capsid protein (VP1) of HS194 (pENTR/SD/D-TOPO-HS194) was generated and used with BaculoDirect linear DNA to perform an LR (attL [entry clone] ϫ attR [destination vector]) recombination reaction to generate recombinant baculovirus DNA carrying the VP1 gene of HS194 (25). Sf9 cells were cultured at a density of 3 ϫ 10 7 cells/flask (162 cm 2 ) and inoculated with rBac GII.4 at a multiplicity of infection (MOI) of 10, after which the medium was harvested at 10 days postinoculation.…”

Section: Methodsmentioning

confidence: 99%

Recognition of Histo-Blood Group Antigen-Like Carbohydrates in Lettuce by Human GII.4 Norovirus

Gao

Esseili

et al. 2016

Appl Environ Microbiol

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Human norovirus (HuNoV) genogroup II genotype 4 (GII.4) strains account for about 80% of the gastroenteritis outbreaks in the United States. Contaminated food is a major transmission vehicle for this virus. In humans, pigs, and oysters, histo-blood group antigens (HBGAs) act as attachment factors for HuNoVs. In lettuce, although the virus-like particles (VLPs) of a GII.4 HuNoV were found to bind to cell wall carbohydrates, the exact binding site has not been investigated. Here, we show the presence of HBGA-like carbohydrates in the cell wall of lettuce. The digestion of lettuce leaves with cell wall-degrading enzymes exposed more binding sites and significantly increased the level of binding of GII.4 HuNoV VLPs. Competition assays showed that both the HBGA monoclonal antibody, recognizing the H type, and plant lectins, recognizing ␣-L-fucose in the H type, effectively inhibited VLP binding to lettuce tissues. Lettuce cell wall components were isolated and their NoV VLP binding characteristics were tested by enzyme-linked immunosorbent assays. The binding was inhibited by pretreatment of the lettuce cell wall materials with ␣-1,2-fucosidase. Collectively, our results indicate that H-type HBGA-like carbohydrates exist in lettuce tissues and that GII.4 HuNoV VLPs can bind the exposed fucose moiety, possibly in the hemicellulose component of the cell wall. IMPORTANCE Salad crops and fruits are increasingly recognized as vehicles for human norovirus (HuNoV) transmission. A recent studyshowed that HuNoVs specifically bind to the carbohydrates of the lettuce cell wall. Histo-blood group antigens (HBGAs) are carbohydrates and are known as the attachment factors for HuNoV infection in humans. In this study, we show the presence of HBGA-like carbohydrates in lettuce, to which HuNoVs specifically bind. These results suggest that specifically bound HuNoVs cannot be removed by simple washing, which may allow viral transmission to consumers. Our findings provide new information needed for developing potential inhibitors to block binding and prevent contamination. Foodborne illnesses constitute a recurring public health burden, with an estimated 9.4 million illnesses and 1,351 deaths annually in the United States (1). Currently, most illnesses (58%) are caused by human noroviruses (HuNoVs), and others are caused by human bacterial pathogens, such as nontyphoidal Salmonella spp. (11%) and Clostridium perfringens (10%) (1). Noroviruses (NoVs) are nonenveloped single-stranded RNA viruses belonging to the Norovirus genus of the family Caliciviridae. NoVs have been classified into five genogroups (GI-GV), and GI, GII, and GIV viruses cause gastroenteritis in humans (2). Each NoV genogroup is further subdivided into several genotypes based on genetic similarities. As a single genogroup, GII currently accounts for about 86% of HuNoV cases (3, 4). HuNoVs have a very low infectious dose, and the 50% infective dose (ID 50 ) was estimated as about 18 viruses and 2,800 genomic equivalents (GEs) (5, 6). Moreover, NoVs are extremely...

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The Effects of Simvastatin or Interferon-α on Infectivity of Human Norovirus Using a Gnotobiotic Pig Model for the Study of Antivirals

Cited by 68 publications

References 53 publications

A Gnotobiotic Pig Model for Determining Human Norovirus Inactivation by High-Pressure Processing

A Gnotobiotic Pig Model for Determining Human Norovirus Inactivation by High-Pressure Processing

Structures of the Compact Helical Core Domains of Feline Calicivirus and Murine Norovirus VPg Proteins

Recognition of Histo-Blood Group Antigen-Like Carbohydrates in Lettuce by Human GII.4 Norovirus

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