Persistence of caliciviruses on environmental surfaces and their transfer to food

Abstract

The noroviruses (NoV) are a common cause of human gastroenteritis whose transmission by foodborne routes is well documented. Fecally contaminated surfaces are likely to contribute to this foodborne transmission and to the propagation of viral disease outbreaks. The purpose of this study was to (i) investigate the stability of NoV on various food preparation surfaces; and (ii) evaluate the degree of virus transfer from these surfaces to a model-ready-to-eat (RTE) food. For the virus persistence experiments, stainless steel, formica and ceramic coupons were artificially contaminated with Norwalk virus (NV), the prototype genogroup I NoV; NV RNA; or feline calicivirus (FCV) F9 (a NoV surrogate), stored at ambient temperature for up to 7 d, and periodically assayed for detection. In the transfer experiments, stainless steel coupons were inoculated with NV or FCV F9 and allowed to dry for 10, 30 and 60 min, after which lettuce leaves were exposed to the surface of the coupons at various contact pressures (10, 100, and 1000 g/9 cm²). Virus recovery was evaluated by RT-PCR (for NV and NV RNA) or by plaque assay (for FCV F9) using Crandell Reese Feline Kidney (CRFK) cells. NV and FCV were detected on all three surfaces for up to 7 d post-inoculation; for FCV, there was an approximate 6 to 7-log₁₀ drop in virus titer over the 7 d evaluation period. By contrast, when stainless steel was inoculated with purified NV RNA, RT-PCR detection was not possible beyond 24 h. Transfer of both NV and FCV from stainless steel surfaces to lettuce occurred with relative ease. This study confirms lengthy NoV persistence on common food preparation surfaces and their ease of transfer, confirming a potential role for environmental contamination in the propagation of viral gastroenteritis.

Introduction

The noroviruses (NoV) are now recognized as a major cause of foodborne illness worldwide (Mead et al., 1999, Lopman et al., 2002). These viruses are transmitted directly by person-to-person contact or indirectly via contaminated food, water, or fomites. Contamination and subsequent transmission can occur by two routes, i.e., fecal–oral and aerosol formation following projectile vomiting (Patterson et al., 1997). For the so-called ready-to-eat (RTE) foods, transmission of enteric viruses by infected food handlers is well documented; indeed, recent epidemiological surveillance data indicate that poor personal hygiene of infected food handlers is the most commonly cited factor contributing to foodborne outbreaks of NoV-associated gastroenteritis (Bean et al., 1996). Surfaces and fomites contaminated through the deposition of fecal material or serving as a repository for aerosolized vomitus also contribute to foodborne transmission and subsequent disease outbreaks (Levy et al., 1975, Mbithi et al., 1991, Patterson et al., 1997). Indeed, previous studies have suggested prolonged environmental persistence of the NoV, as viral RNA has been detected on environmental surfaces such as sinks, commodes, and carpets for several days after initial contamination (Cheesbrough et al., 1997, Green et al., 1998, Green et al., 1999, Liu et al., 2003). Such environmental stability may be at least partially responsible for recent high-profile cruise ship outbreaks of NoV gastroenteritis (Center for Disease Control and Prevention, 2002).

Early studies with cultivable (including hepatitis A virus and rotavirus) human enteric viruses have confirmed virus persistence on materials commonly found in food preparation environments (Abad et al., 1994), along with the possibility for their mechanical transfer from contaminated objects (Lo et al., 1994, Mbithi et al., 1992). Other studies have focused on factors influencing the environmental stability of these cultivable enteric viruses, including relative humidity (RH), temperature, and the type of surface contaminated (Sattar et al., 1986, Mbithi et al., 1991). In the absence of animal or cell culture models for the propagation of the NoV, investigators have used the cultivable feline caliciviruses (FCV) as a surrogate in environmental persistence and transfer studies (Doultree et al., 1999, Lin et al., 2003, Bidawid et al., 2004).

This study was undertaken in an effort to better characterize the persistence of NoV on environmental surfaces commonly used in food preparation, and the relative efficiency with which these viruses can be transferred from contaminated surfaces to foods. Using a two-pronged experimental approach, the persistence and transfer of NoV in human fecal extracts were evaluated using reverse-transcription PCR (RT-PCR), while the same experiments also were done using the FCV model, with survival monitored by mammalian cell culture infectivity assay. Accordingly, we were able to attain both molecular and enumerative (infectivity) data on the degree of virus persistence and transfer.