Virus inactivation by salt (NaCl) and phosphate supplemented salt in a 3D collagen matrix model for natural sausage casings Tinka Wieringa-Jelsma a , Joris J. Wijnker b , Esther M. Zijlstra-Willems a , Aldo Dekker a , Norbert Stockhofe-Zurwieden a , Riks Maas a , Henk J. Wisselink a, ⁎ a Central Veterinary Institute of Wageningen UR, P.O. Box 65, NL-8200 AB Lelystad, the Netherlands b Van Hessen BV, P.O. Box 220 NL-2910 AE, Nieuwerkerk a/d IJssel, the Netherlands abstract article info Article history: Received 4 March 2011 Received in revised form 29 April 2011 Accepted 15 May 2011 Available online 18 May 2011 Keywords: Natural casings Foot-and-mouth disease virus Classical swine fever virus Swine vesicular disease virus African swine fever virus Collagen 3D matrix model Due to possible presence and spread of contagious animal viruses via natural sausage casings the international trade in these food products is subject to veterinary and public health requirements. In order to manage these restrictions we determined the effect of casing preservation on four highly contagious viruses for livestock: foot-and-mouth-disease virus (FMDV), classical swine fever virus (CSFV), swine vesicular disease virus (SVDV) and African swine fever virus (ASFV). We used an in vitro 3D collagen matrix model in which cells, infected with the four different viruses were embedded in a bovine collagen type I gel matrix and treated with either saturated salt (NaCl) or phosphate supplemented saturated salt at four different temperatures (4, 12, 20 and 25 °C) during a period of 30 days. The results showed that all viruses were faster inactivated at higher temperatures, but that stability of the various viruses at 4 °C differed. Inactivation of FMDV in the 3D collagen matrix model showed a clear temperature and treatment effect on the reduction of FMDV titres. At 4 and 12 °C phosphate supplemented salt showed a very strong FMDV inactivation during the first hour of incubation. Salt (NaCl) only had a minor effect on FMDV inactivation. Phosphate supplemented salt treatment increased the effect temperature had on inactivation of CSFV. In contrast, the salt (NaCl) treatment only increased CSFV inactivation at the higher temperatures (20 °C and 25 °C). Also SVDV inactivation was increased by phosphate supplemented salt, but salt (NaCl) treatment only resulted in a significant decrease of SVDV titre at a few time points. The ASFV results showed that both salt (NaCl) and phosphate supplemented salt were capable to inactivate ASFV within 48 h. In contrast to the other viruses (FMDV, CSFV and SVDV), ASFV was the most stable virus even at higher temperatures. The results obtained in this in vitro model underline the efficacy of a combined treatment using phosphate supplemented salt and storage at 20 °C or higher for a period of 30 days. This treatment may therefore be useful in reducing the animal health risks posed by spread of contagious animal viruses by international trade of natural sausage casings. © 2011 Elsevier B.V. All rights reserved. 1. Introduction Various animal viral diseases are notifiable to the OIE, the World Organisation for Animal Health, because of their impact on animal welfare and economic damage. Due to possible presence and spread of infectious viruses via products of animal origin, international shipments are subject to veterinary and public health requirements when importing these goods into a specific region or country. These requirements are often defined in a product- specific Import Risk Assessment by individual countries, using the recommendations of the OIE's Terrestrial Animal Health Code (Office International des Epizooties, 2010) or available scientific literature as a point of reference. Natural sausage casings are sourced, processed and used for sausage production in many different countries and therefore a truly global market exists for this commodity (Wijnker et al., 2008b). Therefore, specific veterinary and public health requirements are applicable to its international trade. The natural casing consists primarily of the submucosal layer of the intestinal tract (Koolmees et al., 2004; Wijnker et al., 2008b), of which collagen is the most important component (Hiltner et al., 1985; Nishiumi and Sakata, 1999). After cleaning, the natural casings are preserved in salt (NaCl) allowing prolonged temperature-independent storage without bacterial spoilage or quality loss (International Scientific Working Group (ISWG), 2009). In addition, previous studies identified a specific mixture of phosphates as preservative added to salt which improves the usability of natural casings for sausage production (Bakker et al., 1999; Houben et al., 2005; Nakae et al., 2008). In the past, available data on virus survival in natural casings were limited to a small number of studies, with little information on International Journal of Food Microbiology 148 (2011) 128–134 ⁎ Corresponding author. Tel.: + 31 320 238403; fax: + 31 320 238961. E-mail address: henk.wisselink@wur.nl (H.J. Wisselink). 0168-1605/$ – see front matter © 2011 Elsevier B.V. All rights reserved. doi:10.1016/j.ijfoodmicro.2011.05.010 Contents lists available at ScienceDirect International Journal of Food Microbiology journal homepage: www.elsevier.com/locate/ijfoodmicro