F-coliphages, porcine adenovirus and porcine teschovirus as potential indicator viruses of fecal contamination for pork carcass processing

Highlights

• F-coliphages, PAdV and PTV can be traced through the pork slaughter process.

• Significant correlations between viable F-coliphages and PAdV DNA

• Significant correlations between viable F-coliphages and PTV RNA

• Potential food handler contamination at later stages of pork processing

• Potential risk for consumers when consuming undercooked pork

Abstract

There are concerns about the zoonotic transmission of viruses through undercooked pork products. There is a lack of information on suitable indicator viruses for fecal contamination with pathogenic enteric viruses in the meat processing chain. The study compared the incidence and levels of contamination of hog carcasses with F-coliphages, porcine teschovirus (PTV), and porcine adenovirus (PAdV) at different stages of the dressing process to assess their potential as indicator viruses of fecal contamination. One hundred swab samples (200 cm²) were collected from random sites on hog carcasses at 4 different stages of the dressing process and from retail pork over the span of a year from 2 pork processing plants (500/plant). Viable F-coliphages, PAdV DNA and PTV RNA were each detected on ≥ 99% of the incoming carcasses at both plants and were traceable through the pork processing chain. Significant correlations were observed between viable F-coliphages and PAdV DNA and between F-coliphages and PTV RNA but not between PAdV DNA and PTV RNA at the various stages of pork processing. Detection of viable F-coliphages was more sensitive than genomic copies of PAdV and PTV at low levels of contamination, making F-coliphages a preferred indicator in the pork slaughter process as it also provides an indication of infectivity. For plant A, F-RNA coliphages were detected in 25%, 63%, and 21% of carcass swabs after pasteurization, evisceration, and retail pork products, respectively. For plant B, F-coliphages were detected in 33%, 25%, and 13% of carcass swabs after skinning, evisceration, and retail pork samples, respectively. Viable F-RNA coliphages were genotyped. Viable F-RNA GII and GIII were generally not detected at the earlier stages of the slaughter process but they were detected on 13% of carcasses after evisceration and 2% of retail pork samples at plant A, which raises concerns of potential food handler contamination during pork processing. Consumers could be at risk when consuming undercooked meat contaminated with pathogenic enteric viruses.

Introduction

Swine hepatitis E virus (HEV), rotavirus (RV) and porcine enteric calicivirus (PEC) are widespread in herds and some swine strains are genetically related to human strains (Farkas et al., 2005, Huang et al., 2002, Martella et al., 2010, Wang et al., 2005, Ward et al., 2008). The zoonotic transmission of HEV is now well established (Colson et al., 2010, Purcell and Emerson, 2010, Yazaki et al., 2003) and similar concerns about the zoonotic transmission of PEC and RV through undercooked meat products have been raised (Bank-Wolf et al., 2010, Martella et al., 2010, Mattison et al., 2007). As HEV, PEC and RV are found in the intestinal tract and shed in fecal material, it is likely that carcasses become contaminated during slaughter and meat processing operations. In addition, contamination of meat products is also possible as a result of poor personal hygiene from infected food handlers (Mattison et al., 2007, Wilhelm et al., 2015). While enteric viruses do not multiply on meat, they are extremely stable at low temperatures (Brandsma et al., 2012, Jones and Muehlhauser, 2015) and are generally more resistant to environmental stresses and decontaminating treatments than bacteria (Baert et al., 2009). As a result, current strategies to reduce bacterial pathogens in food may not be fully effective against viruses.

Unfortunately, many enteric viruses cannot currently be efficiently cultured and can only be detected by molecular methods. However, the presence of viral nucleic acid does not necessarily represent an infectious virus which has important implications in food safety, particularly for the evaluation of inactivation treatments (Baert et al., 2009, Sobsey et al., 1998). In addition, it is not possible to identify all of the enteric viruses that are pathogenic to humans and the potentially pathogenic viruses may not always be present. Therefore, detection of indicators of fecal contamination may be a better approach to understand the risks of potential zoonotic viruses. There is much information on bacterial indicators for fecal contamination but information on appropriate viral indicators for fecal contamination of carcasses and on contamination and survival of enteric viruses on meat products is extremely limited (Brandsma et al., 2012, Jones and Johns, 2012, Jones et al., 2014b, Jones and Muehlhauser, 2015).

F-coliphages are a normal component of the mammalian gut flora, are abundant in swine and can be readily, rapidly, and economically cultured (Grabow, 2001, Havelaar et al., 1984, Havelaar et al., 1990). F-coliphages are a mixed group of coliphages that infect coliforms via the F-pili, some have a DNA genome and others have an RNA genome (Cole et al., 2003). F-RNA coliphages are considered an attractive candidate as an indicator for enteric viruses in environmental waters and shellfish because they are similar in size and possess similar survival characteristics (Doré et al., 2000, Formiga-Cruz et al., 2003). Moreover, F-RNA coliphages can be differentiated into 4 genogroups, where genogroup (G) I and GIV are isolated mainly from animal feces and GII and GIII are mainly associated with human feces although exceptions have been reported (Cole et al., 2003, Furuse, 1987, Schaper et al., 2002b, Scott et al., 2002, Sundram et al., 2006). Our previous work showed that F-RNA coliphages could potentially be used as an indicator of fecal contamination for enteric viruses as viable F-RNA coliphages were detected in 43% (41/96) of fecal samples from swine presented for slaughter, whereas HEV was detected in only 4 samples (Jones and Johns, 2012). Samples collected from different stages of the dressing process (n = 25/stage) at a research abattoir indicated that numbers of F-RNA coliphages on hog carcasses were reduced to a lesser extent than total aerobic bacteria, coliforms and Escherichia coli (Jones and Johns, 2012). A preliminary study with a small number of samples showed that hog carcasses entering a commercial slaughter facility (n = 20) were heavily contaminated with F-RNA coliphages and HEV but levels and frequency were substantially reduced to almost undetectable levels after carcass pasteurization (n = 10) (Jones and Johns, 2012). The prevalence and levels of viable F-RNA coliphages were lower than total aerobes, coliforms and E. coli on the incoming carcasses, which raised the question if the levels of F-RNA coliphages in swine feces may be too low to enable their tracing through the pork slaughter process (Jones and Johns, 2012).

Porcine adenovirus (PAdV), a double stranded DNA virus, and porcine teschovirus (PTV), a single stranded RNA virus, are abundantly shed in swine feces and have been suggested as possible indicators and markers for source tracking of fecal contamination of swine origin in environmental waters (Jiménez-Clavero et al., 2003, Hundesa et al., 2009). While PTV infections are typically asymptomatic, PTV reportedly caused polioencephalomyelitis, a notifiable disease in Canada, in a small number of pig herds in western Canada (Salles et al., 2011). The potential of using PAdV or PTV as an indicator of fecal contamination during the pork slaughter process has not previously been explored. The objectives of the study are to determine the sources of contamination and the fate of F-coliphages, PAdV and PTV as potential indicator viruses of fecal contamination at various stages of commercial slaughter and pork processing.