Thermal Inactivation of Enteric Viruses and Bioaccumulation of Enteric Foodborne Viruses in Live Oysters (Crassostrea virginica)

Abstract

f Human enteric viruses are among the main causative agents of shellfish-associated outbreaks. In this study, the kinetics of viral bioaccumulation in live oysters and the heat stabilities of the predominant enteric viruses were determined both in tissue culture and in oyster tissues. A human norovirus (HuNoV) GII.4 strain, HuNoV surrogates (murine norovirus [MNV-1], Tulane virus [TV]), hepatitis A virus (HAV), and human rotavirus (RV) bioaccumulated to high titers within oyster tissues, with different patterns of bioaccumulation for the different viruses. We tested the thermal stability of each virus at 62, 72, and 80°C in culture medium. The viruses can be ranked from the most heat resistant to the least stable as follows: HAV, RV, TV, MNV-1. In addition, we found that oyster tissues provided protection to the viruses during heat treatment. To decipher the mechanism underlying viral inactivation by heat, purified TV was treated at 80°C for increasing time intervals. It was found that the integrity of the viral capsid was disrupted, whereas viral genomic RNA remained intact. Interestingly, heat treatment leading to complete loss of TV infectivity was not sufficient to completely disrupt the receptor binding activity of TV, as determined by the porcine gastric mucin-magnetic bead binding assay. Similarly, HuNoV virus-like particles (VLPs) and a HuNoV GII.4 strain retained some receptor binding ability following heat treatment. Although foodborne viruses have variable heat stability, 80°C for >6 min was sufficient to completely inactivate enteric viruses in oysters, with the exception of HAV.A pproximately 7.6 million to 14.5 million illnesses in the United States are attributed to the consumption of contaminated seafood each year, and enteric viruses are responsible for more than 50% of these cases (1). In a review of the available epidemiological evidence, human norovirus (HuNoV) and hepatitis A virus (HAV) were the leading viruses associated with shellfish, accounting for 83.7% and 12.8% of outbreaks, respectively (2). The type of shellfish most frequently associated with viral outbreaks was oysters, which were the vehicle in 58.4% of outbreaks (2). In some regions, human enteric viruses are practically ubiquitous in harvested shellfish. Keller et al. (3) showed that 100% of shellfish samples collected from Vitória Bay, Espírito Santo, Brazil, were positive for rotavirus (RV) and adenovirus. However, only 80% of the growing water samples were positive for these pathogens. Viral titers were 400 times higher in the shellfish samples than in the growing water, indicating high levels of natural bioaccumulation (3). In the Galician Rias area, the largest shellfish production area in the European Union, 55% of mussel, clam, and cockle samples were contaminated by HuNoV genogroup I (GI) and GII and HAV (4). Thus, understanding of the ecology and persistence of enteric viruses in shellfish is needed to help prevent future outbreaks.The consumption of uncooked contaminated bivalve shellfish continues to pose a public ...

Thermal Inactivation Kinetics of Tulane Virus in Cell‐Culture Medium and Spinach

“…However, Tian, Yang, Quigley, Chou, and Jiang () and Arthur and Gibson () reported higher D ‐values at 56 °C of 11.8 and 4.03 min, respectively, whereas at 63 °C, reported D ‐values were 2.6 and 1.18 min, respectively, in 1.5 mL microcentrifuge tubes using the first‐order model. Others reported a D ‐value of 2.38 min at 62 °C in cell‐culture media in capillary tube with the first‐order model (Araud et al., ). In this study, a D ‐value of 1.74 min at 56 °C in 2‐mL glass vials was obtained, which appears to be in the lower range of earlier reported values at 56 °C, though somewhat closer and higher than the values reported for 55 °C by Hirneisen and Kniel ().…”

“…Studies on the heat treatment analysis of the capsid integrity of TV showed that at 80 °C after the shortest time point of 5 s, the appearance of the heated virus changed with the loss of normal round structure, which was accompanied with appearance of rough edges (Araud et al., ). It is known that the protruding domain of the capsid protein is responsible for viral entry into the host cell (Yu et al., ); therefore, the loss of capsid integrity can potentially lead to reduction in host cell binding.…”

“…Cultivable TV was not available when these previous studies were undertaken in our laboratory (Bozkurt, D'Souza, & Davidson, , , ; Bozkurt, Ye, Harte, D'Souza, & Davidson, ). Although studies have been performed to analyze the thermal inactivation of TV in cell‐culture media, only 1.5 mL centrifuge tubes (Arthur & Gibson, ; Tian et al., ) or 0.2 mL PCR tubes (Hirneisen & Kniel, ), or more recently, capillary tubes (Araud et al., ), were used. The heat‐inactivation kinetics of TV in 2‐mL glass vials (using different glass container sizes compared to glass capillary tubes) as well as the z ‐values have not been reported to date.…”

Research Progress in Viral Inactivation Utilizing Human Norovirus Surrogates

“…However, considerably more study of different treatments and additional controls on the internal temperature and its distribution in the food matrix would need to be conducted to confirm this. In another study in oysters, a similar phenomenon of a protective effect of oyster matrices against heat treatment of MNV and TV was observed (Araud et al, 2016), but as with the strawberry puree, the mechanism of the protective effect was beyond the scope of the study. One potential mechanism of a protective effect could be viral binding to carbohydrates present in the strawberry and oyster matrices, as such binding has been shown to protect the virus in the case of binding to bacterial cell surface carbohydrates .…”

The effects of X-ray treatments on bioaccumulated murine norovirus-1 (MNV-1) and survivability, inherent microbiota, color, and firmness of Atlantic oysters (Crassostrea virginica) during storage at 5 °C for 20 days

“…Several post-harvest techniques have been proposed to reduce HuNoV in live oysters including cold and high temperature, UV treatment, and high-hydrostatic pressure. However, most of these treatments have low log reduction on norovirus or they kill oysters (Araud et al, 2016, Ye et al, 2015Venugopal, 2005).…”