High-Pressure Inactivation of Human Norovirus Virus-Like Particles Provides Evidence that the Capsid of Human Norovirus Is Highly Pressure Resistant

Department of Food Science and Technology, College of Food, Agricultural and Environmental Sciences, The Ohio State University, Columbus, Ohio, USA.
Applied and Environmental Microbiology (Impact Factor: 3.67). 05/2012; 78(15):5320-7. DOI: 10.1128/AEM.00532-12
Source: PubMed

ABSTRACT Human norovirus (NoV) is the leading cause of nonbacterial acute gastroenteritis epidemics worldwide. High-pressure processing (HPP) has been considered a promising nonthermal processing technology to inactivate food- and waterborne viral pathogens. Due to the lack of an effective cell culture method for human NoV, the effectiveness of HPP in inactivating human NoV remains poorly understood. In this study, we evaluated the effectiveness of HPP in disrupting the capsid of human NoV based on the structural and functional integrity of virus-like particles (VLPs) and histo-blood group antigen (HBGA) receptor binding assays. We found that pressurization at 500 to 600 MPa for 2 min, a pressure level that completely inactivates murine norovirus and feline calicivirus, was not sufficient to disrupt the structure and function of human NoV VLPs, even with a holding time of 60 min. Degradation of VLPs increased commensurate with increasing pressure levels more than increasing time. The times required for complete disruption of human NoV VLPs at 700, 800, and 900 MPa were 45, 15, and 2 min, respectively. Human NoV VLPs were more resistant to HPP in their ability to bind type A than type B and O HBGAs. Additionally, the 23-nm VLPs appeared to be much more stable than the 38-nm VLPs. Taken together, our results demonstrated that the human NoV capsid is highly resistant to HPP. While human NoV VLPs may not be fully representative of viable human NoV, destruction of the VLP capsid is highly suggestive of a typical response for viable human NoV.

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    • "In the absence of a robust in vitro cultivation method, the only source of whole viruses for ligand selection is stool samples from infected individuals. As infectious virus in stool is a difficult sample to obtain and work with, virus-like particles (VLPs) are frequently used instead for many types of studies, from disinfection to immune response characterization (Cheetham et al., 2007; Lou et al., 2012; Nilsson et al., 2009; Souza et al., 2007; Vongpunsawad et al., 2013). VLPs demonstrate similar binding behavior to HBGAs as human NoV particles (Huang et al., 2003; White et al., 1996); however, their production and purification can be costly, time consuming, and variable (Koho et al., 2012). "
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    ABSTRACT: Human noroviruses (NoV) are the leading cause of acute viral gastroenteritis worldwide. Significant antigenic diversity of NoV strains has limited the availability of broadly reactive ligands for design of detection assays. The purpose of this work was to produce and characterize single stranded (ss)DNA aptamers with binding specificity to human NoV using an easily produced NoV target-the P domain protein. Aptamer selection was done using SELEX (Systematic Evolution of Ligands by EXponential enrichment) directed against an Escherichia coli-expressed and purified epidemic NoV GII.4 strain P domain. Two of six unique aptamers (designated M1 and M6-2) were chosen for characterization. Inclusivity testing using an enzyme- linked aptamer sorbent assay (ELASA) against a panel of 14 virus-like particles (VLPs) showed these aptamers had broad reactivity and exhibited strong binding to GI.7, GII.2, two GII.4 strains, and GII.7 VLPs. Aptamer M6-2 exhibited at least low to moderate binding to all VLPs tested. Aptamers significantly (p<0.05) bound virus in partially purified GII.4 New Orleans outbreak stool specimens as demonstrated by ELASA and Aptamer Magnetic Capture (AMC) followed by RT-qPCR. This is the first demonstration of human NoV P domain protein as a functional target for the selection of nucleic acid aptamers that specifically bind and broadly recognize diverse human NoV strains. Copyright © 2015. Published by Elsevier B.V.
    Journal of Biotechnology 06/2015; 37. DOI:10.1016/j.jbiotec.2015.06.389 · 2.87 Impact Factor
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    • "HuNoV virus-like particle (VLP) was also proposed as a potential surrogate for HuNoV because VLPs have similar antigenicity and morphology as HuNoV (Lou et al., 2012). However, since VLPs do not contain viral genetic materials, the reliability of using them as a surrogate for HuNoV is compromised, especially in some treatments targeting the genome of HuNoV for inactivation. "
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    ABSTRACT: Human norovirus (HuNoV) is the leading causative agent for foodborne disease. Currently, studies of HuNoV usually rely on surrogates such as murine norovirus (MNV) due to the lack of a suitable cell culture system and a small animal model for HuNoV. Tulane virus (TV), a monkey calicivirus, is a cultivable enteric calicivirus that not only recognizes the same receptors as HuNoV, but is also genetically closely related to HuNoV. In this study, we determined the pH stability of TV and MNV-1, as well as the effect of high hydrostatic pressure (HHP) on inactivating both viruses in aqueous media, blueberries and oysters. We demonstrated that both TV and MNV-1 were very stable under an acidic environment. They were more resistant to pressure at an acidic environment than at neutral pH. Pressure treatment of 600MPa for 2min at different temperatures (4, 21 and 35°C) barely caused any reduction of TV, as well as MNV-1, on un-wetted (dry) blueberries. However, both TV and MNV-1 on blueberries were successfully inactivated by a pressure of ≤400MPa when blueberries were immersed in phosphate-buffered saline during HHP. Pressure inactivation of both TV and MNV-1 in blueberries and oysters increased as sample temperature decreased in the order of 4>21>35°C. TV was more sensitive to pressure than MNV-1 for the three matrices tested, culture media, blueberries and oysters. This study provides important information on the use of TV as a surrogate for HuNoV study. Results obtained from this study lay a foundation for designing effective HHP treatments for inactivation of HuNoV in high-risk foods such as berries and oysters.
    International journal of food microbiology 01/2013; 162(1):37-42. DOI:10.1016/j.ijfoodmicro.2012.12.016 · 3.08 Impact Factor
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    ABSTRACT: Human norovirus (NoV) is the number one cause of foodborne illness. Despite tremendous research efforts, human NoV is still poorly understood and understudied. There is no effective measure to eliminate this virus from food and the environment. Future research efforts should focus on developing: (1) an efficient cell culture system and a robust animal model, (2) rapid and sensitive detection methods, (3) novel sanitizers and control interventions, and (4) vaccines and antiviral drugs. Furthermore, there is an urgent need to build multidisciplinary and multi-institutional teams to combat this important biodefense agent.
    Infectious disease clinics of North America 09/2013; 27(3):651-74. DOI:10.1016/j.idc.2013.05.009 · 2.73 Impact Factor
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