Enhancement of the Molecular and Serological Assessment of Hepatitis E Virus in Milk Samples (original) (raw)
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Evidence of Hepatitis E Virus in Goat and Sheep Milk
Viruses, 2020
Hepatitis E virus (HEV) is the etiological agent behind hepatitis E infection. Domestic pigs and wild boars are the main animal reservoirs of HEV. Very few papers describe HEV infection in goats and sheep. As the data pertaining to the presence of HEV virus in the milk of small ruminants in Europe are lacking, the aim of this paper was to examine a representative number of milk samples from these animals. The detection of HEV genome (HEV RNA) was performed using reverse transcriptase real-time polymerase chain reaction (RT-qPCR). HEV RNA was found in 2.8% of the examined samples. Positivity ranged from 101 to 103 genome equivalents/mL (GE/mL) with a median of 9.99 × 102 GE/mL. On the basis of these results, the milk of small ruminants could represent a source of HEV infection to consumers.
Hepatitis E Virus in the Food of Animal Origin: A Review
Foodborne Pathogens and Disease
Hepatitis E virus (HEV) is a cosmopolitan foodborne pathogen. The viral agent infects humans through the consumption of contaminated food (uncooked or undercooked). Most cases of infection are asymptomatic and for this reason, this pathology is considered underdiagnosed. Domestic and wild animals are considered natural reservoirs: that is, domestic pig, wild boar, sheep, goat, deer, rabbit, and so on. Therefore, various work categories are at risk: that is, veterinarians, farmers, hunters, slaughterhouse workers, and so on. In these last decades, researchers found a high percentage of positivity to the molecular viral detection in several food matrices included: ready-to-eat products, processed meat products, milk, and shellfish. This review aims to provide an international scenario regarding HEV ribonucleic acid (RNA) detection in several foodstuffs. From this investigative perspective, the study aims to highlight various gaps of the current knowledge about technologies treatments' impact on viral loads. The purpose was also to provide an innovative point of view ''One Health''-based, pointing out the strategic role of environmental safety.
Detection of enteroviruses and hepatitis a virus RNA in cow milk by RT-PCR
Acta Veterinaria-beograd, 2010
Viral RNA was extracted using the rapid acid guanidinium thiocyanatephenol-chloroform method. HAV primers were selected from sequences of the VP2 and VP4 capsid region. Enterovirus primers were selected from the 5' non-translated region, which is the most conserved region in an enterovirus genome. Four samples (8%) raw milk were positive for EV RNA. However, HAV RNA was not determined the milk samples. This is the first report on the detection of HAV and EV RNA in cow milk samples in Turkey. This study highlights the interest for contamination by HAV and EV in milk samples from different areas.
Animals
Identifying Hepatitis E virus (HEV)-positive pig farms is important to implement surveillance programs for this emerging zoonotic agent. The aim of this study was to evaluate the use of serosanguineous fluids obtained as part of castration practice (processing fluids (PFs)) to detect anti-HEV antibodies in newborn piglets. Ninety-five paired serum and PF samples were collected from piglets of 29 different litters and tested with a commercial ELISA kit. A significant positive correlation (Spearman’s rho: 0.600; p < 0.01) was found between anti-HEV antibodies in serum and PF samples. In 26 out of 29 litters (89.7%), there was at least one positive piglet in the serum. Sixteen litters out of 29 (55.2%) were also positive in PFs. To simulate the use of PF as pooled samples, the limit of detection of the ELISA was assessed mixing the PF sample with strong, medium, medium-weak and weak ELISA titres with 3, 4, 5 and 6 negative PF samples. Our results suggest that it is still possible to...
Food Control, 2008
The objective of this study was to detect HAV antigen in raw cow's milk supplied in Mashhad, Iran using ELISA technique. To ascertain the best recovery method for detecting HAV antigen in raw milk, different dilutions of HAV antigen (0.00, 1.0, 10 À3 , 10 À6 and 10 À9 ml/l) were added to UHT milk. Results obtained from the recovery of HAV antigen from treated milk showed that milk with acidic coagulation followed by filtration with paper filters and membrane filters, had the most optical density and considered to be the best recovery method. Then, the detection of HAV in raw milk in two seasons spring and summer was investigated. HAV antigen was detected in 13.3% and 34.48% of milk samples of spring and summer, respectively.
Journal of Clinical Microbiology, 2014
Hepatitis E virus (HEV) is a major cause of acute viral hepatitis in people in many developing countries and is also endemic in many industrialized countries. Mammalian HEV (mHEV) isolates can be divided into at least four recognized major genotypes. Several nucleic acid amplification techniques have been developed for mHEV detection, with great differences in sensitivity. The aim of this study was to compare the performances of two singleplex real-time reverse transcriptase (RT) PCR assays for broad detection of all four mHEV genotypes (assays A and B) and two duplex real-time RT-PCR assays for detection and differentiation of mHEV genotypes 3 and 4 (assays C and D). RNAs extracted from 28 fecal samples from pigs experimentally inoculated with HEV genotype 3 and 186 fecal samples from commercial pigs with unknown HEV exposure were tested by all four assays. In experimental samples, HEV RNA was detected in 96.4% (assay A), 39.2% (assay B), 14.2% (assay C), and 0% (assay D) of the samples. In field samples with unknown HEV exposure, HEV RNA was detected in 67.2% (assay A), 36.4% (assay B), 1.1% (assay C), and 0.5% (assay D) of the samples. The assays showed overall poor agreement ( ؍ 0.19 to 0.03), with differences in detection rates between assays (P < 0.01). Assays A and B, which broadly detect HEV genotypes 1 to 4, had significantly higher detection rates for HEV RNA than the duplex assays C and D, which were both designed to detect and differentiate between HEV genotypes 3 and 4.
Emerging Infectious Diseases, 2020
I nfection with hepatitis E viruses (HEVs) is one of the major causes of acute hepatitis in humans (1). Most HEV strains infecting humans belong to the virus species Orthohepevirus A (HEV-A) (2). HEV-A comprises 8 genotypes; genotypes 1-4 and 7 are found in humans. HEV-A genotypes 1 and 2 seem to be restricted to humans. The other 3 genotypes have also been detected in animals, including pigs (genotypes 3 and 4) and camelids (genotype 7) (1). The most likely source of human zoonotic HEV infection is consumption of contaminated food. Typically, human HEV infections lead to acute and self-limiting disease or asymptomatic seroconversion, but chronic hepatitis E has also been reported, mainly in transplant recipients (3,4). Infection with camel-associated HEV-A genotype 7 was reported in a patient from the United Arab Emirates with chronic hepatitis after liver transplantation (4,5). This infection was likely acquired through consumption of contaminated camel products. Despite the risk for zoonotic transmission, data about shedding and immunity of HEV-A genotype 7 infection in naturally infected dromedaries are scarce. The Study We investigated HEV-A RNA and specific antibody levels in dromedary calves and corresponding dams from 1 farm at monthly intervals over the course of the calves' first year of life. We included serum samples from 11 dam-calf pairs in the United Arab Emirates. The farm contained ≈4,500 camels. The 11 dam-calf pairs investigated in this study were kept in different fenced compartments within 100-150 m of each other but were housed together with other dam-calf pairs in the same paddock throughout lactation (6). All calves were born during June 2014. Serum samples were obtained during the first week and then at monthly intervals until 1 year after birth. We tested samples for HEV RNA by using 2 reverse transcription PCRs (RT-PCRs) and for HEV antibodies by an HEV-A genotype 7 IgG ELISA (Appendix, https://wwwnc.cdc.gov/EID/article/26/9/19-1758-App1.pdf). In the studied cohort, all calves were naturally infected by HEV-A, as confirmed by RNA detection in serum samples, or seroconversion. In 9 of the 11 calves (#1-#9) (Figure 1), HEV-A RNA was detected in >1 serum sample. In 2 calves (#10 and #11) no RNA was detected, but an increase in ELISA ratio, equivalent to seroconversion, confirmed recent HEV-A infection. The average age for infection of calves was 4.6 months (range 1-6 months). All HEV-A-RNA positive calves cleared the virus from their blood and showed accompanying seroconversion (Figure 1). Average viremia was 2.1 months (range 1-4 months). Viral RNA concentrations in serum samples ranged from 6.6 × 10 2 IU/mL to 2.3 × 10 6 IU/mL (mean 4.6 × 10 5 IU/mL).
Hepatitis E virus in lettuce and water samples: A method-comparison study
International journal of food microbiology, 2018
The hepatitis E virus (HEV), which is an increasing cause of acute viral hepatitis in Europe, is a zoonotic virus that is mainly transmitted through contaminated water, consumption of raw or undercooked meat from pigs or wild boar, blood transfusion, and organ transplantation. Although the role of HEV transmission through contaminated produce has not been confirmed, the presence of HEV has been reported in irrigation waters and in vegetables. The present study used a World Health Organization (WHO) international standard and clinical samples to evaluate the performance characteristics of three RT-qPCR assays for detection and quantification of HEV. Two of the evaluated assays provided good analytical sensitivity, as 250 international units (IU) per ml could be detected. Then, experiments focused on evaluating the elution conditions suitable for HEV release from vegetables, with the method proposed by the ISO 15216:2017 selected for evaluation in three types of fresh vegetables. The ...
Journal of Veterinary Diagnostic Investigation, 2006
The aim of this study was to determine at which production stages hepatitis E virus (HEV) is shed by the highest number of pigs and to estimate the relative risk associated with each stage. For this purpose, 146 fecal samples of pigs from 21 farms were studied. In addition, 1 sample from the manure ditch and another sample of drinking water, collected directly from the trough located in the pen, were taken from 16 farms. HEV RNA was detected in fecal samples from 34 pigs (23.29%). The production stages in which most pigs excreted HEV were weaners (41.7%) and pigs in the first month of feeding (60%). The results of the statistical analysis showed that the principal significant risk stage in HEV shedding was the first month of feeding (odds ratio [OR] 19.5, 95% CI 3.59–106.07, P = 0.001) followed by the weaners stage (OR 9.3, 95% CI.78–48.42, P = 0.008). In 8 out of 16 farms tested (50%) HEV RNA was detected in raw manure and in the water trough of only 1. Detection of HEV in manure d...
Detection of hepatitis E virus (HEV) in goats
Virus Research, 2016
Hepatitis E virus (HEV) is a major cause of acute hepatitis worldwide. Genotypes 1 and 2 are restricted to humans, whereas genotypes 3 and 4 also occur in animals and are recognized as zoonotic pathogens. In this study, by screening goat faecal samples collected from six small farms located in the province of Teramo (Abruzzo region, Italy), HEV RNA was found with an overall prevalence of 9.2% (11/119). Upon sequence analysis of a 0.8 kb portion of the ORF2 gene, four strains were grouped with animal and human genotype 3 HEVs, subtype c, with the highest match (94.2-99.4% nt identity) to a wild boar strain, WB/P6-15/ITA, identified in the same geographical area in which the six goat farms were located. Further investigations are needed in order to assess if goat may represent an additional active host for HEV.