Evaluation of Virus Inactivation by Hydrostatic Pressure

APPROACH: Virus and virus contaminated food samples will be prepared and sealed in the ARS laboratory. Samples will be pressure-treated using Dallas Hoover's high pressure unit. After pressurization, samples will be assayed by and within the ARS lab.

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PROGRESS: 2006/10 TO 2007/09<BR>
Progress Report Objectives (from AD-416) The objective is to evaluate the utility of high pressure processing for inactivation of food-borne viruses within shellfish and other foods while retaining uncooked character and flavor. Approach (from AD-416) Virus and virus contaminated food samples will be prepared and sealed in the ARS laboratory. Samples will be pressure-treated using Dallas Hoover's high pressure unit. After pressurization, samples will be assayed by and within the ARS lab. Significant Activities that Support Special Target Populations 1935-42000-059-02N - This report serves to document research conducted under a Non-Funded Cooperative Agreement between ARS and the University of Delaware. A project (continued since the 2006 report) has been completed to evaluate the effects of sugar (sucrose), salt (NaCl), and pH on the inactivation of a human norovirus surrogate, feline calicivirus (FCV), when subjected to high pressure processing (HPP). Results indicate over the range of pH 3¿8, FCV was most resistant to pressure at greater than or equal to pH 5.2. Sucrose and NaCl, applied individually or in combination, retarded the inactivation for FCV by HPP. Increasing the NaCl concentration from 0 to 15%, or sucrose concentration from 0 to 40%, increased this effect, above this plateau of 15% for NaCl and 40% for sucrose, respectively, no additional reduction of pressure inactivation was observed. When both NaCl and sucrose were added to the FCV stock, the two factors had an additive effect, further limiting pressure inactivation of FCV. A second project was initiated to further evaluate temperature effects on inactivation of HPP. Previous work demonstrated that hepatitis A virus (HAV) in tissue culture media was less readily inactivated at colder temperature, the opposite result obtained for other viruses tested. Recent work with HAV-contaminated shellfish has shown that this phenomenon does not appear to hold up for the virus within shellfish tissues, suggesting a specific media-dependent effect of pressure on HAV. Current experiments are seeking to ferret out this issue. Monitoring activities for this project are through conference calls and site visits.
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PROGRESS: 2005/10/01 TO 2006/09/30<BR>

Progress Report 4d Progress report. This report serves to document research conducted under a Non-funded Cooperative Agreement between ARS and the University of Delaware. This study is a sub-component of the parent project 1935-42000-059-00D, Molecular & Biochemical Detection and Intervention Methods for Bacterial and Viral Pathogens within Aquaculture Products. A project has been completed involving an evaluation of high pressure processing time and temperature for the inactivation of hepatitis A virus. A second high pressure project was completed in which feline calicivirus was used to evaluate matrix composition effects for virus inactivation. Results indicate that increasing salt (NaCl) and sucrose composition reduce the efficiency of virus inactivation under pressure. The pH effects were also evaluated. At acidic pH, there was minimal inactivation compared to controls, while at pH 6-8, inactivation was more efficient.
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PROGRESS: 2004/10/01 TO 2005/09/30<BR>

4d Progress report. This report serves to document research conducted under a non-funded Cooperative Agreement between ARS and the University of Delaware. Additional details of this research can be found in the report for the parent CRIS 1935-42000-045-00D, Development of Methods and Strategies to Improve the Microbiological Safety of Aquaculture Products. Projects have been completed involving an evaluation of high pressure processing (HPP) to inactivate hepatitis A virus (HAV) in green onions and strawberry puree and the evaluation of time and temperature effects on HPP treatment parameters for the norovirus surrogate, feline calicivirus. Results indicate that HAV within blended strawberries or HAV-contaminated green onions can be inactivated by 5-min HPP treatments at approximately 3800 atmospheres at room temperature, suggesting that HPP may be a viable commercial intervention strategy for produce. For feline calicivirus, extended treatment time indicated a diminishing rate of inactivation characterized by a nonlinear or tailing effect for longer periods of HPP treatment. Temperatures both above and particularly below room temperature enhanced inactivation. For 5-min, 2000-atmosphere treatments, inactivation differed by as much as 10,000 fold when compared to room temperature. Projects to evaluate the effect of salt, sugar, pH, and soluble protein content on HPP efficiency are ongoing. A second project evaluating the effect of temperature and treatment time on HAV is also ongoing.
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PROGRESS: 2003/10/01 TO 2004/09/30<BR>

4. What were the most significant accomplishments this past year? D. Progress Report: This report serves to document research conducted under a non-funded cooperative agreement between ARS and the University of Delaware. Additional details of this research can be found in the report for the parent CRIS 1935-42000-045-00D titled 'Development of methods and strategies to improve the microbiological safety of Aquaculture products'. Human picornaviruses (i.e. hepatitis A virus, coxsackieviruses, Aichi virus, and parechoviruses) and caliciviruses (i.e. Genogroup I and II norovirus) are replicate in the human intestine, are shed in stool, and subsequently transmitted by oral (food-borne) routes. Since these viruses are quite stable, foods can become contaminated with human waste either by improper food handler hygiene, spraying of non-potable water on vegetables or via bioconcentration by filter feeding shellfish from contaminated growing waters. For uncooked foods such as oysters and produce (i.e. strawberries, green onions, etc.), virus contamination has been linked to numerous virus outbreaks. For shellfish, virus contamination is difficult to detect and there are no good intervention strategies to inactivate purge infectious virus from shellfish beyond adequate cooking. An evaluation of the potential of high hydrostatic pressure, within the Food Science Laboratory at the University of Delaware, indicated that feline calicivirus, a research surrogate for the non-replicatable norovirus is very sensitive to inactivation by high pressure and hepatitis A virus is moderately sensitive to pressure. Since high pressure processing is currently practiced by the oyster industry for automated shell shucking and as a process to inactivate the pathogenic estuarine bacteria Vibrio vulnificus, these results suggest promise for commercial applications. Consequently this specific cooperative agreement was recently initiated. While a formal cooperative agreement was recently initiated, a less well defined research relationship between the Hoover laboratory at the University of Delaware and the USDA, Dover laboratory has existed for the past couple of years. Past accomplishments include the inactivation of HAV and a norovirus surrogate, feline calicvirus, with high pressure. This work has in part led to a collaboration between the USDA-Dover and the FDA-Dauphin Island laboratories which has demonstrated that high pressure can inactivate HAV directly within contaminated oysters. Other picornaviruses such as Aichi-, parecho-, polio- and coxsackie- virus were also evaluated. Results indicate that these other picornavirus have varied sensitivities to inactivation by high pressure. Lambda phage has been identified as a non-pathogenic indicator organism for high pressure processing effectiveness. Evaluation of lambda phage inactivation within different solutions and modeling of inactivation vs. time of pressure application has been performed. Ongoing work is in two areas. First, evaluation of HAV inactivation on the surface of chopped green onion and within blended strawberry puree indicated that HAV can be readily inactivated by HPP in these environments. Second, modeling of inactivation of feline calicivirus is being performed. Inactivation curves of the norovirus surrogate are being generated at constant pressure vs. time (log10 titer reduction vs. time) and constant pressure and time vs. temperature (log10 virus reduction vs. temperature). Current results indicate a diminishing increase of inactivation with time. For temperature, results indicate that colder temperatures (-10 and 0C) actually result in enhanced inactivation of virus as opposed to room temperature.