Pork Liver Sausage Research Articles

Evaluation of hepatitis E virus RNA persistence in experimentally contaminated cured pork liver sausages

Hepatitis E is a disease sustained by RNA viruses, which have four different genotypes, all of which are responsible for acute forms of hepatitis. Genotypes 1 and 2 infect only humans, causing epidemics mainly transmitted by contaminated water, while genotypes 3 and 4 are zoonotic, and the infection is linked to the consumption of raw or undercooked meat or meat products. Hepatitis E virus (HEV) genotypes 3 and 4 have been detected in domestic Suidae, considered the asymptomatic reservoir of HEV, and in wild animals such as wild boar and deer. Despite scientific studies that have highlighted the presence of HEV in cured meat products, such as pork liver sausages, the viral persistence in the different production steps of curing has not been evaluated. Therefore, this study aimed to evaluate the persistence of HEV genotype 3 during the different curing and storage times of experimentally contaminated pork liver sausages using biomolecular methods. The sausages tested positive at all curing and storage times. This study confirms the potential risk attributed to pork liver sausages in HEV transmission. However, to guarantee an efficient risk assessment, future studies will be performed to correlate the presence of HEV RNA with infectious viral particles.

Chronic inflammatory demyelinating polyneuropathy and HEV antibody status: A case-control study from Lazio, Italy

IntroductionFew studies have pointed to the possible role of infectious diseases in triggering Chronic Inflammatory Demyelinating Polyradiculoneuropathy (CIDP). Given the association of Hepatitis E Virus (HEV) with Guillain Barrè syndrome, we conducted a case-control study to determine the possible association of HEV infection with CIDP, analyzing possible risk factors for acquiring HEV infection in both CIDP patients and controls. Materials and methods82 CIDP and 260 from the general population have provided some personal information (demographics, anamnestic data and recognized risk factors for HEV infection) and underwent venipuncture blood sampling for virological assays testing for anti-HEV IgG and IgM with ELISA and RNA-HEV performing RT-PCR. ResultsAnti-HEV IgG seropositivity resulted in 32 CIDP patients (39.0%) and in 45 controls (17.3%), indicating a significant association between anti-HEV IgG positivity and CIDP (OR 3.04; 95% CI 1.70–5.43, p-value <0.001), but in multivariate logistic regression the only significant associations with anti-HEV positivity were eating pork liver sausages (OR 10.443, 95% CI 2.268–60.12, p-value 0.004) and IVIg/SCIg administration (OR 31.32, 95% CI 7.914–171.7, p-value <0.001). DiscussionThe higher prevalence of anti-HEV IgG in CIDP patients than in controls could be justified by chronically administering IVIg/SCIg with a passive acquisition of anti-HEV antibodies. Furthermore, all the 20 CIDP patients who underwent IVIg/SCIg administration reported HEV risk factors, so that they could have acquired the infection. ConclusionsFurther studies in a larger CIDP patient sample in treatment with therapy other than IVIg/SCIg are necessary to rule out the possible confounding effect of IVIg/SCIg.

Detection of hepatitis E RNA in pork products at point of retail in Ireland – Are consumers at risk?

Hepatitis E (HEV), a zoonotic virus, is the leading cause of acute viral hepatitis in Europe. The presence of HEV in domestic pigs can result in infections in humans through consumption of pork products which are undercooked or where processing methods are insufficient to inactivate the virus. In Ireland, pork accounts for 34 % of all meat consumption (CSO, 2022) and the prevalence of HEV in products at point of retail has not previously been characterised.A sampling strategy was designed in which high pork content sausages, fresh pork liver and raw fermented sausages were systematically purchased from three types of retailers between May 2018 and March 2019. In total, 200 pork products were tested using a lysing agent to release the HEV from the product for detection. RT-PCR for HEV was performed on samples with an extraction efficiency >1 % (n = 188/200) (94 %). Low level HEV RNA was detected in 9/188 (4.8 %) pork products tested. The highest incidence of HEV RNA was in pork liver where 6/25 (24 %) samples were positive. The concentration of HEV ranged from 0.02 – to 9.4 genome copies/g of pork.Based on these data an exposure assessment was performed which found that if consumers followed advice from the Food Safety Authority of Ireland to achieve core temperatures of 70 °C or higher when cooking, the risk was likely to be negligible.

Prevalence and risk factors for hepatitis E virus infection in blood donors: a nationwide survey in Italy, 2017 to 2019.

BackgroundIn high-income countries, hepatitis E virus (HEV) infection is mainly a zoonosis. However, it is also transfusion-transmissible and some countries, but not Italy, have introduced HEV screening for blood donations.AimWe assessed HEV infection prevalence and risk factors in a nationwide sample of Italian blood donors.MethodsWe selected 107 blood establishments (BE) distributed in the 20 Italian regions by a stratified two-stage design and invited them to participate in the study. Donors were tested for anti-HEV IgG and IgM and HEV RNA. Sociodemographic data and risk factors were collected through a questionnaire.ResultsOverall, 60 BE from 60 provinces in 19 Italian regions joined the study. We assessed HEV markers in 7,172 blood donors, of whom 6,235 completed the questionnaire. Overall crude and adjusted anti-HEV IgG prevalences were 8.3% and 5.5%, respectively. Overall anti-HEV IgM prevalence was 0.5%, while no blood donor was HEV RNA-positive. Anti-HEV IgG prevalence varied widely among regions (range: 1.3%–27.20%) and hyperendemic prevalences (> 40%) were detected in some provinces in two regions. Older age (AOR = 1.81; 95% CI: 1.36–2.41), foreign nationality (AOR = 2.77; 95% CI: 1.06–7.24), eating raw pork liver sausages (AOR = 2.23; 95% CI: 1.55–3.20) and raw homemade sausages (AOR = 3.63; 95% CI: 2.50–5.24) were independent infection predictors.ConclusionItalian blood donors showed a low to moderate HEV seroprevalence. High levels in some regions and/or provinces were mainly attributable to eating habits. Prevention should include avoiding consumption of raw or undercooked meat and safe production of commercial pork products.

Control of Raw Pork Liver Sausage Production Can Reduce the Prevalence of HEV Infection.

After an acute hepatitis E (HEV) outbreak in Southern Switzerland, in January 2017 the local public health authorities started an active program of food chain control and public education. In this retrospective study, we analysed all laboratory-confirmed acute cases of HEV infection diagnosed between 2014 and 2020. In the period before the public health intervention, the number of cases increased steadily from 2014 (4 of 40 tests, 10%) reaching a peak in the last quarter of 2016 (42 of 285 tests, 14.7 %). Afterwards, the number of positive cases decreased steadily, reaching its lowest value (0.3%) in the second quarter of 2019. There was a statistically significant difference between the frequency of positive cases and period of testing, i.e., before and after the introduction of the public health interventions. Our study shows that active public health measures to control sausages containing raw pork liver can reduce the prevalence of HEV infection.

Hepatitis E virus (HEV) genotype 3 diversity: Identification of a novel HEV subtype in wild boar in Central Italy.

In the last decade in Europe, the number of autochthonous cases of hepatitis E has significantly increased. Most of the cases arise from foodborne infections caused by the zoonotic hepatitis E virus (HEV) genotypes HEV-3 and HEV-4. Several human cases have been linked to consumption of raw or undercooked animal products of both pork (liver sausages) and wild boar meat. In this study, the occurrence of HEV infection was investigated in 611 livers and 88 paired lungs from wild boars collected during the hunting seasons of 2016-2020 in the Umbria-Marche Apennines (Central Italy). Using real-time reverse transcription polymerase chain reaction, 15 liver samples (2.45%) and one lung sample were found to be positive for HEV RNA. The phylogenetic tree built on the partial ORF2 gene revealed that the detected HEV strains belonged to HEV-3f (n=5), HEV-3e (n=1) and HEV-3c (n=1) subtypes. Interestingly, 8 strains were genetically placed in a different cluster, further away from all other subtypes. To corroborate this finding, four complete genomes were obtained by next generation sequencing. The full genome of the HEV strains clustered together with another wild boar strain previously detected in Southern Italy in 2015 but the strains were divergent from all the HEV-3 strains classified in any subtype defined so far. Thus, these strains represent a novel subtype that might have originated in Italy, which we have tentatively named HEV-3n.

Occurrence of HEV-RNA in Italian Regional Pork and Wild Boar Food Products.

Hepatitis E is an emerging threat in industrialized countries. The foodborne transmission linked to consumption of pork and game meat is considered the main source of autochthonous infection. In Europe, small outbreaks have been reported linked to the consumption of pork liver sausages and wild boar meat. Based on previous findings and on increasing evidence of pork and game meat as a vehicle for HEV infections, the present study investigated the occurrence of HEV in 99 pork and 63 wild boar sausages and salami sold in Southern Italy. The HEV genome was detected in four wild boar sausages. Sequencing from 2 wild boar sausages confirmed that the HEV strains detected belonged to HEV-3genotype, not assigned to any defined subtype. Data obtained confirmed the possible occurrence of HEV in pork products and in game. Although the detection rate is low, these products are frequently consumed raw after curing, whose effect on virus viability is still unknown.

Incidence of hepatitis E virus infection among blood donors in a high endemic area of Central Italy.

In Europe, autochthonous hepatitis E virus (HEV) infection is mainly a foodborne zoonosis, but it is also transmitted by blood transfusion. Despite the numerous prevalence surveys, only a few studies have investigated HEV incidence. We aimed to determine HEV incidence and risk factors among blood donors in a hyperendemic area in Central Italy. Of 296 blood donors who had tested HEV negative in two previous seroprevalence surveys in L'Aquila, 198 agreed to undergo at least another blood sampling for estimating HEV incidence nearly 2years after the prevalence surveys. Ten newly acquired infections were detected, yielding an overall incidence of 2.1/100 person-years (95%CI: 1.0-3.9), with an estimated participant's cumulative probability of becoming HEV infected of 6.5% (95%CI: 3.5-12.0) at 4years after enrolment. Seven newly infected blood donors were IgG positive only, two were IgM positive (one also IgG positive) and one was HEV RNA positive only, harbouring subtype 3c. Incident infection was most strongly associated with eating game meat, raw-dried pork liver sausage and raw-dried wild boar sausage. None of these exposures was statistically significant, even if eating raw-dried wild boar sausage approached significance (P=0.06). The HEV incidence we found was considerable compared with other similar studies. The nearly significant association of incident infection with wild boar and other game meat consumption was in agreement with the 3c subtype isolation in the viremic donor. However, beyond eating habits, also other exposure sources are likely important in hyperendemic areas, where incidence and risk exposure studies need to be undertaken for effectively preventing HEV transmission.

Interlaboratory Validation of a Method for Hepatitis E Virus RNA Detection in Meat and Meat Products.

Increasing numbers of hepatitis E cases are currently recognized in many European countries. The zoonotic hepatitis E virus (HEV) genotype 3 mainly circulates in domestic pigs and wild boars, and can be transmitted to humans via consumption of insufficiently heated meat or meat products produced from those animals. Here, a detailed protocol for detection of HEV RNA in meat products is provided, which is based on the method originally described by Szabo et al. (Intl J Food Microbiol 215:149-156, 2015). It consists of a TRI Reagent®/chloroform-based food matrix homogenization, a silica bead-based RNA extraction and a real-time RT-PCR-based RNA detection. The method was further validated in a ring trial with nine independent laboratories using pork liver sausage samples artificially contaminated with different amounts of HEV. The results indicate sufficient sensitivity, specificity, and accuracy of the method for its broad future use in survey studies, routine food control or outbreak investigations.

Acute hepatitis E in French patients and neurological manifestations

Hepatitis E virus (HEV) is a major cause of acute hepatitis worldwide. However, our understanding of the source of contamination is incomplete and the frequency of neurological manifestations in still unknown. 200 eligible cases reported to the French National Reference Center from January 2015 to December 2015 were prospectively included in this case-control study (1 case: 1 control, matched for sex, age and area of living) to investigate the risk of infection. We documented the factors associated with their HEV infection and clinical manifestations. The 200 HEV-infected patients included 137 who were immunocompetent and 63 immunocompromised. The factors associated with an HEV infection were contact with farm animals, eating pork liver sausage and eating unpeeled fruit. The 33 patients (16.5%) who reported neurological symptoms included 14 with neuropathic pain suggesting small fiber neuropathy, 9 with painless sensory disorders, 6 with Parsonage-Turner syndrome, one Guillain-Barre syndrome, one meningitis, one encephalitis and one diplopia. Neurological manifestations were more frequent in immunocompetent patients (22.6% vs 3.2%, p < 0.001). This study highlights the risk of HEV transmission by the environment in industrialized countries. The higher frequency of neurological disorders in immunocompetent patients suggests pathophysiological mechanisms involving the immune system.

천연채소를 활용한 돈간소시지의 품질특성

천연채소를 활용한 돈간소시지의 품질특성

Screening of Ready-to-Eat Meat Products for Hepatitis E Virus in Switzerland

Seroprevalence data for pig herds suggested that there must be a relevant reservoir for hepatitis E virus (HEV) in Switzerland. To know more about the viral presence in ready-to-eat meat products, we screened pork liver sausages and raw meat sausages from the Swiss retail market for the presence of HEV. Testing was performed with a detection method where the virus extraction step was optimized. As for the performance of the improved method, the mean recovery rate for the mengovirus process control was 24.4%, whereas for HEV-inoculated sample matrices between 10.4 and 100% were achieved. The limit of detection was about 1.56 × 103 and 1.56 × 102 genome copies per gram for liver sausages and raw meat sausages, respectively. In the screening programme, HEV-RNA was detected in 10 of total 90 (11.1%) meat products, 7 of 37 (18.9%) liver sausages, and 3 of 53 (5.7%) raw meat sausages. Virus loads of up to 5.54 log10 HEV genome copies per gram were measured. All sequences retrieved from positive samples belonged to HEV genotype 3. The significance of the presented work was a current overview of the HEV prevalence in ready-to-eat meat products on the Swiss retail marked and an improvement of the extraction efficiency of the HEV detection method.

Assessment of the risk of foodborne transmission and burden of hepatitis E in Switzerland

The objective of this study was i) to quantify the risk of hepatitis E for Swiss consumers by specified pork products and ii) to estimate the total burden of human food-borne hepatitis E in Switzerland. A quantitative risk assessment from slaughter to consumption was carried out according to the Codex Alimentarius framework. In the hazard characterization, assumptions were made due to the lack of a dose-response relationship for oral exposure to hepatitis E virus (HEV). The prevalence of HEV in 160 pig livers of 40 different Swiss fattening farms was examined and determined to be 1.3% (CI 0.3%; 4.4%). This result was used as input in the risk assessment model, together with data from other published studies. The annual burden of hepatitis E was estimated in terms of Disability Adjusted Life Years (DALY), using data about hepatitis E cases diagnosed between 2010 and 2015 at two major hospitals located in the canton Ticino. Only the risk of foodborne hepatitis E from products containing pork liver was evaluated, as those containing only pork meat could not be evaluated because of lack of data on HEV load in pork. Assuming that successful oral infection occurs in 1% of servings contaminated with high HEV loads (>105 genome copies), and that acute illness develops in 5% of susceptible consumers, the most likely annual number of foodborne hepatitis E cases in Switzerland was estimated to be 1481 (95% CI 552; 4488) if all products containing pork liver were considered. If only high-risk products, such as plain pork liver and liver sausages (e.g. Saucisse au Foie), were considered, the annual number of cases was estimated to be 176 (95% CI 64; 498). We were unable to calculate the total burden of hepatitis E in Switzerland due to lack of data. Yet, for the canton Ticino, it was shown that a significant increase had occurred from <5 DALY per 100,000 inhabitants before 2012 to >50 DALY per 100,000 inhabitants in 2015. This change could partly be due to an increased reporting and higher awareness among medical practitioners. Extrapolation to other regions could be accomplished if detailed information on food consumption patterns were available. Notification of HEV cases and attempts of cases source attribution would improve the basis for risk assessments.

Detection of Torque Teno Sus Virus in Pork Bile and Liver Sausages.

Torque teno viruses (TTV) are small DNA viruses widespread among humans and pigs. The clinical significance of TTV infections in either humans or pigs is uncertain. In fact, TTV viremia is highly prevalent in patients with different pathologies, but it can also be frequently observed in healthy subjects. Virus infection in pigs is considered a putative cofactor in several diseases; despite being detected frequently in healthy animals, its role still remains unknown. The present study aimed to investigate the presence of Torque teno sus virus (TTSuV) in 62 bile samples collected from pigs at slaughterhouse and in 36 fresh pork liver sausages bought at point of sale. Quantitative Real-Time PCR, confirmed that 19.4 and 58.3% of bile and sausage samples tested positive for TTSuV, respectively. The mean viral load was established as 5.6×104GE/µl for bile and 7.16×103GE/g for sausages. TTSuV nucleotide sequence analysis confirmed a wide heterogeneity among the circulating TTSuV strains, which included both TTSuV1 and TTSuV2.

A nationwide survey of hepatitis E viral infection in French blood donors.

Most cases of hepatitis E viral (HEV) infection in developed countries are autochthonous. Nevertheless, the reported seroprevalence of HEV varies greatly depending on the geographical area and the performance of the immunoassay used. We used validated assays to determine the prevalence of anti-HEV immunoglobulin G (IgG) and IgM among 10,569 French blood donors living in mainland France and three overseas areas. Epidemiological information was collected using a specific questionnaire. We found an overall IgG seroprevalence of 22.4% (8%-86.4%) depending on the geographical area (P < 0.001). The presence of anti-HEV IgG was associated with increasing age (P < 0.001) and eating pork meat (P = 0.03), pork liver sausages (P < 0.001), game meat (P < 0.01), offal (P < 0.001), and oysters (P = 0.02). Conversely, drinking bottled water was associated with a lower rate of anti-HEV IgG (P = 0.02). Overall IgM seroprevalence was 1% (0%-4.6%). The frequency of anti-HEV IgM was higher in donors living in a high anti-HEV IgG seroprevalence area (1.9% versus 0.7%, P < 0.001) and in those eating pork liver sausage (1.4% versus 0.7%, P < 0.01), pâté (1% versus 0.4, P = 0.04), and wild boar (1.3% versus 0.7%, P < 0.01). HEV is endemic in France and hyperendemic in some areas; eating habits alone cannot totally explain the exposure to HEV, and contaminated water could contribute to the epidemiology of HEV infection in France.

Seroprevalence in blood donors reveals widespread, multi-source exposure to hepatitis E virus, southern France, October 2011

The apparent seroprevalence of hepatitis E Virus (HEV)varies greatly among developed countries depending on the geographical area and the sensitivity of immunoassays. We used a validated assay to determine the prevalence of HEV IgG and IgM antibodies among 3,353 blood donors living in southern France,who gave blood during the two first weeks of October 2011 and participated in the study. Demographic and epidemiological information was collected using aspecific questionnaire. We also screened 591 samples for HEV RNA. Overall IgG seroprevalence was 39.1%and varied from 20% to 71.3% depending on the geographical area (p < 0.001) while IgM seroprevalence was 3.31%. Anti-HEV IgG was significantly correlated with increasing age (p < 0.001), eating uncooked pork liver sausages (p < 0.001), offal (p = 0.003), or mussels(p = 0.02). Anti-HEV IgM was associated with being male (p = 0.01) and eating uncooked pork liver sausages(p = 0.02). HEV RNA was detected in one of the 99 anti-HEV IgM-positive samples, but in none of the 492 anti-HEV IgM-negative samples. HEV is hyperendemic in southern France. Dietary and culinary habits alone cannot explain the epidemiology of HEV in this region, indicating that other modes of contamination should be investigated.

Detection of hepatitis E virus in pork liver sausages

Hepatitis E infection is regarded as an emerging public-health concern. The disease is normally self-limiting (mortality rate 1%), but chronic infections have recently been observed in transplanted patients. The etiological agent HEV is a small RNA virus infecting both humans and animals. In humans, the disease may be food-borne and pig is a main reservoir for zoonotic strains. In the present study, we evaluated the presence of HEV and swine fecal cross-contamination in pork liver sausages sold at a grocery store in Italy. HEV genome detection was performed by RT-qPCR, using harmonized protocols that included a process control (murine norovirus) and an internal amplification control. Swine fecal cross-contamination was assessed by determination of the ubiquitous porcine adenovirus.Overall, HEV genome belonging to genotype 3 was detected in both raw (10 out of 45 slices, 250mg each, 22.2%) and dry (1 of 23 slices, 4.3%) liver sausages, but infectivity of the virus was not demonstrated. This pilot study fosters more investigations on HEV presence in pork-derived food, to assess the possible risk for the consumers.

Hepatitis E Virus Genotype 4, Denmark, 2012

Hepatitis E Virus Genotype 4, Denmark, 2012

Hepatitis E virus seroprevalence and risk factors for individuals in working contact with animals

BackgroundIn industrial countries genotypes 3 and 4 of HEV are detected in swine, wild boar, deer and rabbits, and they are associated with autochthonous infections suggesting the existence of zoonotic HEV infections, compatible with the putative involvement of undercooked pork and big game products as a source of infection. ObjectivesTo evaluate the prevalence of anti-HEV antibodies in different population groups in contact with animals and to investigate risk factors for HEV infection. Study designSerum samples were collected from 859 healthy French subjects, including pig farm workers, forestry workers and individuals without working contact with animals (control group). In addition, 58 swine veterinarians were included in the study. Subjects were interviewed using a structured questionnaire, and anti-HEV antibodies were investigated using a sensitive and specific sandwich ELISA. ResultsAnti-Hepatitis E virus (HEV) antibodies were detected in 26% of control population, and in 36% and 44% of forestry and pig farm workers, respectively. In addition, an increase in seroprevalence from the north to the south of France was observed (30.2% versus 40.7%). Consumption of pork liver sausage (AOR 4.4, p<10−4), occupational contact with animals (AOR 1.58, p=0.038 for forestry workers and AOR 2.51, p<10−4 for pig farm workers), and living in southern France (AOR 1.47, p=0.02), were independent risk factors. Wearing working gloves and boots might reduce HEV infection. ConclusionsOccupational exposure to animals and consuming raw or undercooked pork liver sausage or pork liver play a significant role in HEV transmission in industrial countries.

Changes in sensory properties and consumer acceptance of reduced fat pork Lyon-style and liver sausages containing inulin and citrus fiber as fat replacers

The effects of fat reduction in Lyon-style (25% fat) and liver sausages (30% fat) using inulin, citrus fiber and partially rice starch were studied in terms of sensory properties and consumer acceptance. Fat reduced Lyon-style sausages (3 to 17% fat) and liver sausages (3 to 20% fat) were respectively compared to the full-fat controls. Reducing fat in Lyon-style sausages decreased meat flavor, aftertaste meat flavor, greasiness and juiciness, and enhanced color intensity, spiciness, spicy aftertaste, raspy throat, coarseness and firmness scores. But adding inulin and citrus fiber led to sensory characteristics similar to the full-fat reference. Regarding liver sausages, attribute scores in greasiness, creaminess, lumpiness and foamy were decreased with fat reduction and simultaneous addition of fibers. Color intensity, spiciness, firmness and attribute furred tongue were increased. Consumer tests revealed acceptable fat reduced (32 to 90% less than control) and fiber enriched (1.0 to 5.6%) sausages. Drivers of liking were found to relate not only to high-fat but also to low-fat samples.