Samples of liver and bile were collected from 86 wild boars, 62 deer, 20 wild rabbits, and 78 coypu in the Midi-Pyrénées area. The wild boars and deer were hunted from February 2010 through January 2011, rabbits were hunted from October 2013 through February 2014, and coypu were hunted in April 2011.

RNA was extracted from 30 mg of liver by using RNeasy Mini Kits or from 140 μL of bile by using QIAamp Viral RNA Mini Kits as specified by the manufacturer (QIAGEN, Courtaboeuf, France). Real-time PCR based on ORF3 was used to detect and quantify HEV RNA in plasma samples as previously described (9). The limit of detection was 100 copies/mL.

HEV RNA was detected in 5 (5.8%) wild boar livers, 2 (3.2%) deer livers, 1 (5.0%) wild rabbit liver, and no coypu livers (Table). Thus, the overall prevalence of HEV RNA among wildlife, irrespective of species, was 3.3% (8/246) (95% CI 1.1%–5.5%). Because bile samples were available only from animals with negative HEV RNA liver samples, no bile sample was positive for HEV RNA.

The overall prevalence of HEV RNA in game (3.3%) is similar to that among pigs. A recent nationwide study in France reported that HEV RNA was present in the livers of about 4% of farmed pigs of slaughter age (10). These contaminated livers can then enter the food chain and be responsible for foodborne zoonotic HEV infection. Such pork products, especially raw or undercooked liver, are a major source of exposure to the virus (11). HEV has been found in pig liver sausages (5,12,13). Our findings suggest that game meat from wild boar, deer, and rabbit contribute to HEV epidemiology in the Midi-Pyrénées area. However, residents of this area eat less game meat than pork.

Prevalence rates of HEV RNA in the wild boar populations of other European countries like the Netherlands (2.0%) and Germany (5.0%) are similar (2), and rates of HEV RNA are higher among wild boars in Hungary (12.2%) and Italy (25.0%) (2) and among deer in Spain (13.6%), the Netherlands (15.0%), and Hungary (34.4%) (2). However, despite these HEV RNA prevalence figures, no HEV-hyperendemic region in these countries has been described. Thus, the prevalence of HEV RNA among wildlife alone is not enough to explain the high seroprevalence in a specific area. Contact with the animal reservoir and local dietary practices must be taken into account.

Prevalence of HEV RNA among wild rabbits in this study (5%) is lower than that found by a previous (2010) study of wild rabbits from the Haute-Garonne Department in southwestern France (6/12; 50%) (7). The low prevalence rate reported here could be the result of our shorter collection time (5 months in our study vs. 3 years in the previous study). It is also possible that HEV has become less prevalent since the previous study was done. The description of a strain from humans closely related to a strain from rabbits indicates that zoonotic transmission of HEV from rabbits is possible (7).

A recent study has shown that rats are competent hosts for HEV3, suggesting that rodents (e.g., coypu) may be an alternative reservoir for zoonotic strain HEV3 (14). Coypu meat is eaten as pâté and may be an unexpected source of HEV foodborne zoonotic HEV infection. However, none of the 78 coypu livers tested were positive for HEV RNA. Thus, coypu do not seem to carry zoonotic strains of HEV. However, significant nucleotide changes may be present in the HEV strains in coypu. Thus, the primers used may have not been able to amplify HEV RNA.

In conclusion, the prevalence of HEV RNA in wild boars, deer, and rabbits is similar to that previously reported for pigs. Consumption of the meat of these wild animals, and of pig liver sausage, all contribute to the HEV epidemiology in the Midi-Pyrénées area because of specific local eating habits. Game meat from this part of France should be cooked thoroughly to minimize the risk for HEV infection (15).