Determination of hepatitis E virus inactivation during manufacturing of spreadable pork liver sausage and salami-like raw pork sausage.

Abstract

The zoonotic hepatitis E virus (HEV) can cause acute and chronic hepatitis in humans. Meat from domestic pigs, which represent a major animal reservoir of HEV, plays a key role in HEV transmission. Although pork meat products can contain HEV-RNA, it is unknown whether infectious HEV is still present after their manufacturing process. Here, we used a newly developed method for virus extraction from sausages and a quantitative method for detecting HEV infectivity in artificially contaminated sausages to investigate the HEV inactivation during production of spreadable pork liver sausage and salami-like raw pork sausage. The cell culture-adapted HEV genotype 3c strain 47832c was used to contaminate meat preparations intended for production of sausages, which were manufactured based on recipes commonly used in Germany. According to these recipes, spreadable liver sausages of a certain diameter are to be held in a water bath at 70 °C for 30 min. Therefore, the HEV inoculated liver sausage preparations were filled into conical tubes and heated in a 70 °C water bath. After 21 min, the sausages reached a core temperature of 70 °C and samples were taken after further incubation for up to 18 min. For the raw sausages, the HEV inoculated meat preparation was filled into natural casings and sausages were cured at 18 °C and 80 % relative humidity. Samples were taken for up to 21 days. HEV was extracted from all samples, which were quantitatively analyzed for infectious virus and viral RNA using cell culture and RT-qPCR, respectively. During liver sausage production, infectious HEV decreased by four log₁₀ immediately after reaching the core temperature of 70 °C and was completely inactivated (>4.3 log₁₀ decrease) 2 min later (23 min heat treatment). In contrast, the HEV-RNA amount decreased only marginally (<0.6 log₁₀) throughout the whole incubation time. During raw sausage manufacturing, infectious HEV decreased only slightly (<1.3 log₁₀) over three weeks of curing, while the HEV-RNA amount remained unchanged. It can be concluded that the intended heating regime during production of spreadable liver sausages leads to an inactivation of HEV, indicating a low risk of HEV infection for consumers if these sausages are manufactured properly. In contrast, HEV was only slightly inactivated during production of salami-like raw pork sausage. Therefore, raw sausage can contain infectious HEV if starting material with a high HEV amount was used for production. Viral RNA testing cannot be used to predict infectivity of HEV in meat products.