Detection and Phylogenetic Analysis of the Hepatitis E Virus in a Canadian Swine Production Network

Viral contamination along the production chain is a significant concern in both food safety and livestock health. Pigs have been reported to act as a reservoir for zoonotic viruses, sometimes emerging ones, and epidemiological studies have shown direct links between the consumption of uncooked pork offal and cases of hepatitis caused by the hepatitis E virus (HEV) genotype 3 in humans. The presence of HEV in swine herds has been reported, but its dissemination in pork production environments is still unknown. To investigate viral contamination sources in the swine industry, 452 environment and fecal samples, including samples from livestock transportation vehicles, were collected over a period of 11 months from ten farms and one slaughterhouse that together represent a single production network. Hepatitis E virus RNA was detected by nested RT-PCR in 32 samples from both inside and outside farm buildings, on trucks, and, mostly, from fomites collected in the slaughterhouse yard, such as on a utility vehicle. Phylogenetic analysis showed a wide diversity of HEV genotype 3 strains, similar to human and swine strains previously found. According to the results of this study, the movements of trucks and utility vehicles might play an important role in HEV dissemination on a slaughterhouse site and throughout an entire network.     

Prevalence and Molecular Characterization of the Hepatitis E Virus in Retail Pork Products Marketed in Canada

Infection with the hepatitis E virus (HEV) is very common worldwide. HEV causes acute viral hepatitis with approximately 20 million cases per year. While HEV genotypes 1 and 2 cause large waterborne and foodborne outbreaks with a significant mortality in developing countries, genotypes 3 and 4 are more prevalent in developed countries with transmission being mostly zoonotic. In North America and Europe, HEV has been increasingly detected in swine, and exposure to pigs and pork products is considered to be the primary source of infection. Therefore we set out to investigate the prevalence of HEV in retail pork products available in Canada, by screening meal-size portions of pork pâtés, raw pork sausages, and raw pork livers. The presence of the HEV genomes was determined by RT-PCR and viral RNA was quantified by digital droplet PCR. Overall, HEV was detected in 47% of the sampled pork pâtés and 10.5% of the sampled raw pork livers, but not in the sampled pork sausages, and sequencing confirmed that all HEV strains belonged to genotype 3. Further phylogenetic analysis revealed that except for one isolate that clusters with subtype 3d, all isolates belong to subtype 3a. Amino acid variations between the isolates were also observed in the sequenced capsid region. In conclusion, the prevalence of HEV in pâtés and raw pork livers observed in this study is in agreement with the current HEV distribution in pork products reported in other developed countries.     

First Serological Study of Hepatitis E Virus Infection in Backyard Pigs from Serbia

Hepatitis E virus (HEV) genotype 3 infections in pigs and humans have been lately reported in Europe. In the present study, the prevalence of anti-HEV IgG antibodies in swine was investigated, for the first time, in Serbia by means of an ELISA based on a recombinant open reading frame 2 protein of HEV genotype 3. A total of 315 serum samples from 3 to 4 months-old healthy backyard pigs, collected in 63 herds from 28 towns and villages of 4 different districts of the Vojvodina province, in the northern part of Serbia, were tested. A 34.6% (109/315) of the sera tested were positive. The prevalence of anti-HEV antibodies varied widely between municipalities (range 16.7–75.0%) and herds (range 0–100%). These data indicate that HEV infection is widespread in Serbian backyard pigs.     

Hepatitis E Virus (Genotype 3) in Slurry Samples from Swine Farming Activities in Italy

Hepatitis E virus (HEV) is an emergent causative agent of acute hepatitis, transmitted by fecal–oral route. Infection with HEV is a global cause for morbidity and mortality throughout the world: it mainly causes large outbreaks in endemic areas and sporadic autochthonous cases in industrialized countries where HEV infections seem to be an emergent zoonotic disease. Infection of porcine livestock and its relationship with the human cases have been demonstrated. The present study describes an investigation on the prevalence and diversity of HEV in pig slurry in Italy. Slurry samples (24) were collected from ten farms located in North Italy during 2015 and analyzed for HEV, using four broad-range nested PCR assays targeting ORF1 (MTase), ORF2 (capsid) genes, and ORF2/3 regions. Overall, 18 samples (75%) were positive for HEV RNA, and characterized as genotype 3. Nine samples could be subtyped by ORF2 sequencing: Eight belonged to subtype 3f, while one sequence could not be characterized by blast analysis and phylogenetic analysis and may actually represent a new subtype. Furthermore, similarity of 99% was found between 3f Italian HEV sequences of human and swine origins. Real-Time PCR assay was also performed, in order to obtain quantitative data on positive samples. Two swine slurry samples were positive, containing 600 and 1000 UI per mL of sewage. The results of this study show that HEV strains belonging to zoonotic genotype 3 are widely present in swine excreta, and have high degree of identity with strains detected in autochthonous HEV cases. Improving swine farming operations safety and increasing operators’ awareness of the zoonotic potential connected with the handling of swine effluents turn out to be key points in order to reduce the environmental and sanitary problem represented by the possible dissemination of HEV to water bodies.     

Potential Approaches to Assess the Infectivity of Hepatitis E Virus in Pork Products: A Review

The zoonotic transmission of hepatitis E, caused by the hepatitis E virus (HEV), is an emerging issue. HEV appears common in pigs (although infected pigs do not show clinical signs), and evidence suggests that a number of hepatitis E cases have been associated with the consumption of undercooked pork meat and products. Little information is available on whether cooking can eliminate HEV, since there is currently no robust method for measuring its infectivity. HEV infectivity can be clearly demonstrated by monitoring for signs of infection (e.g., shedding of virus) in an animal model. However, this approach has several disadvantages, such as lack of reproducibility and unsuitability for performing large numbers of tests, high costs, and not least ethical considerations. Growth in cell culture can unambiguously show that a virus is infectious and has the potential for replication, without the disadvantages of using animals. Large numbers of tests can also be performed, which can make the results more amenable to statistical interpretation. However, no HEV cell culture system has been shown to be applicable to all HEV strains, none has been standardized, and few studies have demonstrated their use for measurement of HEV infectivity in food samples. Nonetheless, cell culture remains the most promising approach, and the main recommendation of this review is that there should be an extensive research effort to develop and validate a cell culture-based method for assessing HEV infectivity in pork products. Systems comprising promising cell lines and HEV strains which can grow well in cell culture should be tested to select an assay for effective and reliable measurement of HEV infectivity over a wide range of virus concentrations. The assay should then be harnessed to a procedure which can extract HEV from pork products, to produce a method suitable for further use. The method can then be used to determine the effect of heat or other elimination processes on HEV in pork meat and products, or to assess whether HEV detected in any surveyed foodstuffs is infectious and therefore poses a risk to public health.     

Prevalence of Hepatitis E Virus (HEV) Antibodies in Mexican Pigs

The hepatitis E virus (HEV) is the causative agent of Hepatitis E, an enterically transmitted disease. HEV infections in pigs and humans have been reported worldwide, but data from Mexico are scarce. In the present study, the prevalence of anti-HEV IgG antibodies was investigated in a quite large number of swine from Mexico by means of an ELISA based on a recombinant open reading frame 2 protein of HEV genotype 3. Serum samples from 683 healthy pigs (1–48 months old), collected during 2010–2013 in 109 herds from 48 municipalities located in 9 states in the centre of the country were assayed. A 30.75 % (210/683) of the sera tested were positive, and they were distributed along all the states included in the study. The prevalence of anti-HEV antibodies varied widely between municipalities and herds, and it was higher in pigs 4–6 months of age. No relationships were detected between seroprevalences and farm characteristics. Forty individual faecal samples were analysed by RT-PCR and all resulted negative. These data indicate that HEV infection is widespread in Mexican pigs; thus, representing a potential zoonotic risk for humans.     

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.     

Hiding in Plain Sight? It’s Time to Investigate Other Possible Transmission Routes for Hepatitis E Virus (HEV) in Developed Countries

Historically in developed countries, reported hepatitis E cases were typically travellers returning from countries where hepatitis E virus (HEV) is endemic, but now there are increasing numbers of non-travel-related (“autochthonous”) cases being reported. Data for HEV in New Zealand remain limited and the transmission routes unproven. We critically reviewed the scientific evidence supporting HEV transmission routes in other developed countries to inform how people in New Zealand may be exposed to this virus. A substantial body of indirect evidence shows domesticated pigs are a source of zoonotic human HEV infection, but there is an information bias towards this established reservoir. The increasing range of animals in which HEV has been detected makes it important to consider other possible animal reservoirs of HEV genotypes that can or could infect humans. Foodborne transmission of HEV from swine and deer products has been proven, and a large body of indirect evidence (e.g. food surveys, epidemiological studies and phylogenetic analyses) support pig products as vehicles of HEV infection. Scarce data from other foods suggest we are neglecting other potential sources of foodborne HEV infection. Moreover, other transmission routes are scarcely investigated in developed countries; the role of infected food handlers, person-to-person transmission via the faecal–oral route, and waterborne transmission from recreational contact or drinking untreated or inadequately treated water. People have become symptomatic after receiving transfusions of HEV-contaminated blood, but it is unclear how important this is in the overall hepatitis E disease burden. There is need for broader research efforts to support establishing risk-based controls.     

菲对湘江沉积物吸附镉的影响

制备了镉单独污染和镉-菲复合污染的模拟水样,研究了菲对湘江沉积物吸附模拟水样中镉的动力学和热力学特征的影响,并进一步研究了在不同水样pH和ρ(沉积物)下,菲存在与否及ρ(菲)对沉积物吸附镉的影响.结果表明,无论菲存在与否,湘江沉积物吸附镉的热力学数据均能够用Langmuir方程拟合,相关系数达到0.99以上,属单分子层吸附;动力学数据均可用拉格朗日二级动力学方程拟合,相关系数接近1;以化学吸附过程为主,且在湘江沉积物上的吸附均以快速吸附为主;但是菲的存在削弱了沉积物对镉的吸附能力,提高了镉的解吸率.在试验范围内随着水样pH逐渐增大,菲对沉积物吸附镉的影响越小;ρ(菲)越大,沉积物对镉的吸附量越小;菲对沉积物吸附镉的影响主要是由于菲与镉竞争沉积物颗粒矿物之间的孔隙以及甲醇化的菲分子与颗粒的结合掩蔽了颗粒物表面的部分极性基团两方面原因造成的.      

Expression and Immunoreactivities of Hepatitis E Virus Genotype 3 Open Reading Frame-2 (ORF-2) Recombinant Proteins Expressed in Insect Cells

Hepatitis E virus (HEV) is a fecal-orally transmitted virus that is endemic in many geographical areas with poor sanitary conditions and inadequate water supplies. In Europe, a low-endemic area, an increased number of autochthonous sporadic human cases of patients infected with HEV strains of genotype 3, have been reported lately. The relatively high prevalence of HEV genotype 3 infections in European pigs has raised concerns about a potential zoonotic transmission to humans. Determination of HEV seroprevalence in pigs would help to clarify its incidence and possible zoonotic implications. To this purpose, we have expressed and partially characterized swine genotype 3 HEV open reading frame-2 proteins upon infection of Sf21 insect cells with recombinant baculoviruses. The use of the expressed proteins as diagnostic antigens for the detection of antibodies to HEV has been further assayed with human and swine sera.     

北京城区水体中PPCPs的分布特征及潜在风险

采集北京城区河流34个表层水样和23个沉积物样品,采集后海、前海和西海这3个城市湖泊10个表层水样和5个沉积物样品,应用HPLC-MS/MS技术对表层水样及沉积物样品中10种药物及个人护理品(PPCPs)进行检测分析,发现河流表层水样和城市湖泊表层水样中PPCPs的浓度范围分别为N.D.~655 ng·L~(-1)和N.D.~252 ng·L~(-1).在河流表层水样中,10种PPCPs的检出率范围为0~100%,其中咖啡因的检出率为100%,磺胺甲唑、地尔硫卓和泰乐菌素均未检出;在湖泊表层水样中,10种PPCPs的检出率范围为0~100%,其中对乙酰氨基酚、林可霉素、咖啡因和阿奇霉素的检出率为100%,磺胺甲唑、地尔硫卓、泰乐菌素及卡马西平都未检出;河流沉积物和城市湖泊沉积物中PPCPs的含量范围分别为N.D.~1 709ng·g~(-1)和N.D.~35.9 ng·g~(-1),在河流表层沉积物中,10种PPCPs的检出率范围为4%~96%,其中甲氧苄啶的检出率最高,泰乐菌素的检出率最低;在湖泊表层沉积物中,10种PPCPs的检出率范围为0~100%,其中地尔硫卓的检出率最高,咖啡因、泰乐菌素和卡马西平均未检出.从各河流看,永定引水渠、凉水河、通惠河及坝河表层水体及沉积物中PPCPs的含量较高,永定河、昆玉河表层水体中PPCPs的含量较低.应用风险商值(RQ)模型评价北京城区各河流及湖泊表层水体与沉积物中PPCPs残留对生态环境的影响,发现北京城区河流及湖泊表层水体中的10种PPCPs的RQ值均低于0.1,对河流生态环境具有低风险.城区河流及湖泊沉积物中,对乙酰氨基酚对永定引水渠、通惠河及坝河底栖生态环境具有中等风险;林可霉素对永定引水渠、温榆河、通惠河、护城河、坝河、凉水河及后海底栖生态环境具有中等风险;甲氧苄啶对永定引水渠、清河、温榆河、通惠河、护城河、坝河、亮马河、凉水河及后海底栖生态环境具有中等风险;阿奇霉素对亮马河及凉水河底栖生态环境具有中等风险,对永定引水渠、清河、温榆河、通惠河、坝河及后海底栖生态环境具有高风险.     

Preliminary studies on occurrence of monensin antibiotic in Bosque River Watershed

Water quality impact due to excessive nutrients has been extensively studied. In recent years, however, micro-pollutants such as pharmaceuticals and hormonal products used in animal agriculture have added an additional impact to overall water quality. Pharmaceuticals used in the poultry, swine, beef, and dairy industries have been detected in various environmental matrices such as, soil, groundwater and surface water. In this study, 26 surface water samples were collected throughout the Bosque River Watershed (BRW) with samples representing a range of land use conditions and locations of major dairy operations. Samples were analyzed using commercially available Enzyme-Linked Immunosorbent Assay test. Of the 26 samples, three samples consistently tested positive for monensin antibiotic with concentration ranging from 0.30 to 3.41 μg/L. These three samples were collected from sites that received varying amount of agriculture wastes (11.7% to 31.3%) and located downstream from sites associated with moderate levels of animal agriculture. The preliminary results suggest that there is a potential for monensin occurrence in the BRW, although initial findings indicate only very low levels.     

Prevalence and Genetic Characterization of Aichivirus in Environmental Waters in Thailand

Aichivirus 1 (AiV-1) is an enteric virus that has been documented to be the causative agent of diarrhea in humans. It is transmitted by fecal–oral route, through person-to-person contact, consumption of contaminated food or water, or recreation of contaminated water. AiV-1 is highly prevalent in water samples and has been proposed as a potential indicator of fecal contamination in water reservoirs. This study aimed to investigate the prevalence and genetic diversity of AiV-1 in environmental water samples in Thailand. A total of 126 samples were collected monthly from November 2016 to July 2018 from various sources of environmental water including irrigation water, reservoir, river, and wastewater. The presence of AiV-1 was detected by RT-nested PCR of the 3CD region and further analyzed by phylogenetic analysis. The AiV-1 was detected in 28 out of 126 (22.2%) of tested samples. A high frequency of AiV-1 detection was in wastewater (52.4%). All 28 AiV-1 strains detected in this study belonged to the genotype B and were closely related to AiV strains detected previously in environmental waters and in humans worldwide. This study demonstrated, for the first time, the contamination of AiV-1 in various sources of water samples in Thailand and provided a better insight into the prevalence of AiV-1 in environmental waters and its potential risk of human health.

     

Survival of Porcine Reproductive and Respiratory Syndrome Virus in Pork Products

There is a risk of virus transmission through contaminated pork, and many viruses are considered potential hazards for both humans and livestock. The risk of transmission may be elevated with importation/exportation of meat between countries globally. Survival of porcine reproductive and respiratory syndrome virus (PRRSV) in different pork products has not been studied. The present study evaluated PRRSV survival in four different products: fresh sausage, ham, bacon, and acidified sausage prepared with experimentally contaminated pork. These products were prepared according to standard methods used by the manufacturers of pork products, and then stored at room temperature, 4 °C and ?20 °C. PRRSV was detected only in fresh sausage for up to 15 days at 4 °C and for 30 days at ?20 °C. No PRRSV was detected at any temperature in any of the other three products. These preliminary data provide valuable information for the pork processing industry, as well as in planning for import/export of these products among different countries.     

松花江流域渔业生态环境汞污染现状评价

2007～2009年对松花江干流、嫩江下游江段、第二松花江干流水质、底质和鱼类汞含量状况进行了调查研究,并参照渔业水质标准、无公害水产品产地环境要求、无公害水产品安全要求和松花江流域汞的背景值对流域生态环境汞污染进行评价.结果表明:江水中总汞含量0.000010～0.000172 mg/L,符合渔业水质标准要求;第二松...     

Hepatitis E Seroprevalence and Genotyping in a Cohort of Wild Boars in Southern Germany and Eastern Alsace

In the last few years it has been realized that the hepatitis E virus (HEV) is endemic in most industrialized countries and that it is a zoonotic disease. Potential reservoirs for HEV have been identified to be wild boars and deers, but HEV has also been found in domestic pigs and other animals. Due to the probable spread of the virus via contaminated food or contact to infected animals, HEV antibodies are present in more than 16% of the German adult population and rates are increasing with age. We collected blood from 104 wild boars in southern Germany and the border region of Alsace. We found an anti-HEV seroprevalence of 11.5% in our cohort, using ELISA. Furthermore, we observed active infection in 3.85% of the animals by positive HEV PCR in the sera of the boars. In our cohort, no regional differences of seroprevalence or active infection were seen. Sequencing revealed rather close homology of some detected HEV sequences to genotypes isolated from patients in Germany. Hence wild boars are a potential source of HEV infection in Middle Europe and the rate of infectious animals is quite high.     

低溶解氧状态下河网区不同类型沉积物的氮释放规律

采集浙江温州温瑞塘河河网中4个受不同人类行为干扰的沉积物样品,分别代表受河道上游来水、制革厂排水、建筑垃圾和生活污水排放的影响,研究了低溶解氧(DO)状态下不同类型沉积物各形态氮(N)的释放规律.结果表明,低DO状态下,不同上覆水条件能明显影响沉积物-水系统中N迁移转化.上覆水污染越重越不利于沉积物中N释放,水质改善后易促进沉积物中N的释放.屿田河高营养盐水平的上覆水中的N仍能在扩散作用下进入沉积物,并导致已严重污染的沉积物中总氮(TN)和硝酸盐氮(NO3--N)含量增加,控制河流外源N持续输入利于沉积物污染负荷的降低.低DO状态下各类沉积物中氨氮(NH4+-N)均会释放进入上覆水,而NO3--N则迁至沉积物中,并在反硝化作用下脱离系统.NH4+-N的释放量取决于其在沉积物中的含量.低DO状态下NO3--N释放的风险小于NH4+-N.