Introduction: Hepatitis E virus (HEV) is an RNA virus causing an acute generally self-limited disease in humans. Over the last 10 years, in Europe an increasing number of autochthonous cases linked to foodborne transmission of HEV genotype 3 have been reported. Pigs and wild boar are the main reservoirs. Human cases have been frequently linked to the consumption of raw or undercooked pork products and wild boar meat. In this study, the presence of HEV-RNA was investigated in 92 livers and 116 paired liver and muscles from wild boar (Sus scrofa) collected during two hunting seasons 2016 -2017 and 2017-2018, respectively, in Central Italy. Materials and Methods: Fifty mg of liver and muscle samples were used for RNA extraction (RNeasy Mini Kit, Qiagen). The HEV genome was detected and quantified by quantitative Real-Time RT-PCR (RTqPCR) (QuantiFast Pathogen +IC Kits). The RNA of HEV positive liver and muscle samples was analysed by nested RT-PCR amplifying a 348 bp fragment in the ORF2. The 800 bp fragment within the Methyltransferase region (ORF1) was also amplified from paired liver and muscle samples by nested RTPCR (ORF1). The phylogenetic analysis was performed using MEGA7 software. Results: HEV RNA was detected in 62 livers with prevalence ranging from 30% to 65.7% depending on the hunting area. All but one group of hunted animals was negative for HEV. Five animals (8%, 5/62) were positive for HEV in both muscle and liver while none of the animals was positive in muscle only. The ORF2 and ORF1 partial viral sequences were obtained from all five paired livers and muscles. The sequenced strains obtained from paired livers and muscles belonging to the same animal were identical (100% nt.id.). Phylogenetic analysis showed that strains clustered within the different subtypes HEV-3c, HEV-3f and other clusters not assignable to any subtypes described so far. HEV is an emerging public health issue. In mammals, 4 main genotypes have been identified: HEV-1 and -2 infect exclusively humans, HEV-3 and -4 both animals and humans. HEV-3 is frequently associated with autochthonous cases of HEV in industrialized countries and it is widespread worldwide. Swine are considered the most important reservoir of HEV-3 and -4 and the virus has been described in farms worldwide with prevalence up to 50% in animals of 3-5 months of age. The aim of this study is to evaluate the dynamics of HEV infection in 10 pigs housed in the same farm from age of 2 months to slaughter. Individual fecal samples were collected every 2 weeks for 6 months. All samples were tested for the presence of HEV RNA by qRT-PCR. Data collected, showed a progressive reduction of the quantitative of HEV to the point of negative at 28 week. Our study aims to prove how long the virus is shed by animals, if re-infection can occur and if HEV shedder animals are infected at slaughterhouse.

MONITORING OF HEV IN NATURALLY INFECTED PIGS UP TO SLAUGHTERING

De Lucia A.;Ostanello F.;
2019

Abstract

Introduction: Hepatitis E virus (HEV) is an RNA virus causing an acute generally self-limited disease in humans. Over the last 10 years, in Europe an increasing number of autochthonous cases linked to foodborne transmission of HEV genotype 3 have been reported. Pigs and wild boar are the main reservoirs. Human cases have been frequently linked to the consumption of raw or undercooked pork products and wild boar meat. In this study, the presence of HEV-RNA was investigated in 92 livers and 116 paired liver and muscles from wild boar (Sus scrofa) collected during two hunting seasons 2016 -2017 and 2017-2018, respectively, in Central Italy. Materials and Methods: Fifty mg of liver and muscle samples were used for RNA extraction (RNeasy Mini Kit, Qiagen). The HEV genome was detected and quantified by quantitative Real-Time RT-PCR (RTqPCR) (QuantiFast Pathogen +IC Kits). The RNA of HEV positive liver and muscle samples was analysed by nested RT-PCR amplifying a 348 bp fragment in the ORF2. The 800 bp fragment within the Methyltransferase region (ORF1) was also amplified from paired liver and muscle samples by nested RTPCR (ORF1). The phylogenetic analysis was performed using MEGA7 software. Results: HEV RNA was detected in 62 livers with prevalence ranging from 30% to 65.7% depending on the hunting area. All but one group of hunted animals was negative for HEV. Five animals (8%, 5/62) were positive for HEV in both muscle and liver while none of the animals was positive in muscle only. The ORF2 and ORF1 partial viral sequences were obtained from all five paired livers and muscles. The sequenced strains obtained from paired livers and muscles belonging to the same animal were identical (100% nt.id.). Phylogenetic analysis showed that strains clustered within the different subtypes HEV-3c, HEV-3f and other clusters not assignable to any subtypes described so far. HEV is an emerging public health issue. In mammals, 4 main genotypes have been identified: HEV-1 and -2 infect exclusively humans, HEV-3 and -4 both animals and humans. HEV-3 is frequently associated with autochthonous cases of HEV in industrialized countries and it is widespread worldwide. Swine are considered the most important reservoir of HEV-3 and -4 and the virus has been described in farms worldwide with prevalence up to 50% in animals of 3-5 months of age. The aim of this study is to evaluate the dynamics of HEV infection in 10 pigs housed in the same farm from age of 2 months to slaughter. Individual fecal samples were collected every 2 weeks for 6 months. All samples were tested for the presence of HEV RNA by qRT-PCR. Data collected, showed a progressive reduction of the quantitative of HEV to the point of negative at 28 week. Our study aims to prove how long the virus is shed by animals, if re-infection can occur and if HEV shedder animals are infected at slaughterhouse.
Atti XIX Congresso Nazionale S.I.Di.L.V.
89
89
Franco V., Chelli E., De Lucia A., Cerini N., Ostanello F., Di Bartolo I.
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Utilizza questo identificativo per citare o creare un link a questo documento: http://hdl.handle.net/11585/719187
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