Hepatitis E virus (HEV) is an important human pathogen and, in developed countries, most human infections are due to a zoonotic cycle, mainly maintained by domestic and wild suids. In European countries several genotype 3 strains have been found to circulate in human population and animal reservoirs, with human infections being related mostly to pork or wild boar meat consumption. In this study, we surveyed HEV circulation in wild boar in Umbria (Italy) during the 20212022 hunting seasons, using a stratified sampling. Liver samples were tested for HEV presence by real-time RT-qPCR. Positive samples were characterized by nested RT-PCR followed by sequencing of partial region of the capsid gene. Overall, 78 out of 179 wild boar liver samples tested positive to HEV (43.6), with viral load ranging between 1.47 and 7.35 log genome copies/g (median 3.20 log). Variations, although not statistically significant, were observed considering geographical, age, weight, and gender factors, in terms of either prevalence or viral load. In particular, in animals younger than 1 year of age, the viral load was 2 log higher (median viral load of 5.50 log genome copies/g of liver tissue) than in older age groups. Sequence analysis characterized the 41 obtained sequences into genotype 3 subtype 3c (n=4), 3f (n=11), and 3e (n=1), while 13 sequences clustered with two genotype 3 genomes (GenBank MF959764 and MK390971) still unassigned to subtypes. Additional, 12 sequences did not cluster with any known subtype or unassigned genome and on phylogenetic analysis segregated into two distinct groups of eight and four sequences, respectively. Interestingly, some wild boar sequences of subtype 3f were intermingled with sequences of HEV strains previously identified in human patients in Central Italy. Sharing of molecular data for HEV in animals is pivotal to decipher the intricate ecology of HEV.

The use of natural substances with antiviral properties might reduce foodborne viral diseases. In this study, we evaluated the virucidal effect of Citrus limon and Thymus serpyllum essential oils (EOs) and of Citrus Limon, Thymus serpyllum and Thymus vulgaris hydrolates on murine norovirus (MNV), a human norovirus surrogate. To assess the virucidal effect of these natural substances, the reduction in viral infectivity was estimated by comparing the TCID₅₀/mL of untreated viral suspension and the viral suspension treated with hydrolates and EOs at different concentrations. The results showed a natural loss of infectivity of the untreated virus after 24 h of approx. 1 log. The EO (1%) of T. serpyllum, and hydrolates (1% and 2%) of T. serpyllum and T. vulgaris immediately caused a reduction in MNV infectivity of about 2 log but did not provide a further significant decrease after 24 h. Instead, the EO (1%) and hydrolate (1% and 2%) of C. limon exerted an immediate reduction in the viral infectivity of about 1.3 log and 1 log, respectively, followed by a further reduction in infectivity of 1 log after 24 h for the hydrolate. These results will allow for the implementation of a depuration treatment based on the use of these natural compounds.

Food-borne transmission is a recognized route for many viruses associated with gastrointestinal, hepatic, or neurological diseases. Therefore, it is essential to identify new bioactive compounds with broad-spectrum antiviral activity to exploit innovative solutions against these hazards. Recently, antimicrobial peptides (AMPs) have been recognized as promising antiviral agents. Indeed, while the antibacterial and antifungal effects of these molecules have been widely reported, their use as potential antiviral agents has not yet been fully investigated. Herein, the antiviral activity of previously identified or newly designed AMPs was evaluated against the non-enveloped RNA viruses, hepatitis A virus (HAV) and murine norovirus (MNV), a surrogate for human norovirus. Moreover, specific assays were performed to recognize at which stage of the viral infection cycle the peptides could function. The results showed that almost all peptides displayed virucidal effects, with about 90% of infectivity reduction in HAV or MNV. However, the decapeptide RiLK1 demonstrated, together with its antibacterial and antifungal properties, a notable reduction in viral infection for both HAV and MNV, possibly through direct interaction with viral particles causing their damage or hindering the recognition of cellular receptors. Hence, RiLK1 could represent a versatile antimicrobial agent effective against various foodborne pathogens including viruses, bacteria, and fungi.

Globally, waterborne viral infections significantly threaten public health. While current European Union regulations stipulate that drinking water must be devoid of harmful pathogens, they do not specifically address the presence of enteric viruses in water used for irrigation or food production. Traditional virus detection methods rely on molecular biology assays, requiring specialized personnel and laboratory facilities. Here, we describe an electrochemical sandwich enzyme-linked immunomagnetic assay (ELIME) for the detection of the hepatitis A virus (HAV) in water matrices. This method employed screen-printed electrodes as the sensing platform and utilized commercially available pre-activated magnetic beads to provide a robust foundation for the immunological reaction. The ELIME assay demonstrated exceptional analytical performance in only 185 min achieving a detection limit of 0.5 genomic copies per milliliter (g.c./mL) and exhibiting good reproducibility with a relative standard deviation (RSD) of 7% in HAV-spiked drinking and processing water samples. Compared with the real-time RT-qPCR method described in ISO 15216-1, the ELIME assay demonstrated higher sensitivity, although the overall linearity of the method was moderate. These analytical attributes highlight the potential of the ELIME assay as a rapid and viable alternative for HAV detection in water used for agriculture and food processing.

Food-borne transmission is a recognized route for many viruses associated with gastrointestinal, hepatic or neurological diseases. Therefore, it is essential to identify new bioactive compounds with broad-spectrum antiviral activity is needed to exploit innovative solutions against these hazards. Lately, antimicrobial peptides (AMPs) have been recognized as promising antiviral agents. Indeed, while the antibacterial and antifungal effects of these molecules have been widely reported, their use as potential antiviral agents has not yet been fully investigated. Herein, the antiviral activity of previously identified or newly designed AMPs, was evaluated against the non-enveloped RNA viruses, Hepatitis A virus (HAV), and Murine Norovirus (MNV), a surrogate for human Norovirus. Moreover, specific assays were performed to recognize at which stage of the viral infection cycle the peptides could function. Results showed that almost all peptides displayed virucidal effects with about 90% of infectivity reduction on HAV or MNV. However, the decapeptide RiLK1 demonstrated, together with its antibacterial and antifungal properties, a notable reduction in viral infection for both HAV and MNV, possibly through direct interaction with viral particles or inhibition of the viral bonding to the cell receptors. Hence, RiLK1 could represent a versatile ‘antimicrobial agent effective against various foodborne pathogens including viruses, bacteria, and fungi.