Beyond the liver: the broadened tissue tropism of hepatitis E virus.

How hepatitis E virus invades hepatocytes: the mystery of viral entry.

The World Health Organization (WHO) estimates that every year hepa -titis A virus (HAV) infection causes nearly 1.4 million new cases worldwide and the hepatitis E virus (HEV) is re-sponsible for 20 million new infections and over 3 million acute cases. Although in most cases HAV and HEV infections are self-limiting, HAV is estimated to kill 100 000 people each year [1] and HEV nearly 60000 people annually [2]. Pregnant women are at risk of more severe disease, obstetric complications and increased mortality if infected in the third trimester of pregnancy. The faecal–oral route is a well-established mode of transmission for both HAV and HEV and in the case of HEV infec-tion person-to-person transmission is also an important factor in sporadic cases [3]. Outbreaks of HAV and HEV are therefore manifestations of the poor sanitation practices and lack of clean water supplies often found in developing countries. Humanitarian crises with large refugee populations can also be fertile ground for hepatitis outbreaks alongside cholera or other waterborne outbreaks, as was recently seen among Syrian refugees in Iraq [4]. Understanding the importance of the risk of hepatitis outbreaks in displaced populations can help in identifying out-breaks quickly and responding to them in a timely manner to reduce mortality and morbidity. HAV has 7 genotypes, with little variation in their clinical expression. HEV has 4 genotypes with quite differ-ent clinical expressions, responsible for different disease manifestations across developing and developed countries. In developing countries genotype 1 is largely responsible for outbreaks and sporadic cases, via contamination of water and the fecal–oral route. Excep-tions include Mexico in South America and countries in Africa, where genotype 2 is more common [5]. Neither HAV nor HEV have chronic states, although HEV is reported in immunocompro-mised people [6]. Acute HAV infection is often clini-cally indistinguishable from other caus-es of acute viral hepatitis, and laboratory confirmation is necessary. Diagnosis of HEV has its own challenges, which may result in an underestimation of the disease burden [6]. HAV has a very effective vaccine available, and the first vaccine for HEV was approved in China in December 2011, although it is not yet used in any other countries. Whereas HAV seroprevalence increases with age and comes close to 100% in highly endemic countries by the age of 5 years, HEV seroprevalence tends to stay be-tween 5%–60% [7]. In 2010, the WHO World Health Assembly adopted resolution WHA63.18, which called for the pre-vention and control of viral hepatitis, with a focus on HBV and HCV [8]. This resolution came after Member States and WHO understood the gravity of spread of viral hepatitis. In 2012, on the occasion of World Hepatitis Day, Dr Ala Alwan, WHO Regional Director for the Eastern Mediterranean, urged all stakeholders to combine their efforts to confront and combat this silent epi-demic of hepatitis. Many countries of the Eastern Mediterranean Region (EMR) are cur-rently going through major social and political upheavals. With active or proxy wars engulfing many countries in the Region, the public health structures are under stress, creating ideal conditions for the spread of all infectious diseases, especially those spread by contaminated water and lack of sanitation services. Except for a few published articles and outbreak reports, however, very limited data are available about the prevalence of HAV and HEV from these countries or from the Region in general [9–12]. One major reason for this knowledge gap is that we are not looking for the evidence. In most developing countries, including most of those of the EMR, HEV is not routinely considered when a physician asks for investigations into a suspected case of viral hepatitis. Added to the diagnostic challenges is the fact that a majority of hepatitis-infected per-sons do not develop an acute condition that requires major health care interven-tion. HAV is in the same class, with few symptomatic cases reaching health-care settings, and as it is self-limiting in most cases, it is neither investigated nor re-ported to surveillance systems as HAV. Physicians also do not consider HAV or HEV to be serious illnesses, even though fulminant hepatitis, hepatic failure and death can occur from both infections.More than 40% of the population of the EMR lives in just 2 countries, Egypt and Pakistan, both of which are consid-ered endemic for HBC and HCV [8]. In Pakistan, the Field Epidemiology Labo-ratory Training Programme is collecting information about acute viral hepatitis cases via 5 sentinel sites throughout the country [13]. According to reports from the Pakistan viral hepatitis surveillance system HAV—responsible for more

Hepatitis E virus (HEV) is one of the most frequent causes of acute viral hepatitis of enteric transmission worldwide. In South America the overall epidemiology has been little studied, and the burden of the disease remains largely unknown. A research of all scientific articles about HEV circulation in South America until November 2017 was carried out. Human seroprevalences of HEV varied according to the studied population: blood donors presented prevalence rates ranging from 1.8% to 9.8%, while reports from HIV-infected individuals, transplant recipients and patients on hemodialysis showed higher prevalence rates. Only 2 cases of chronic hepatitis in solid-organ transplant patients from Argentina and Brazil have been described. Detection of HEV in the swine population is widely prevalent in the region. Anti-HEV antibodies have also been recently documented in wild boars from Uruguay. Although scarce, studies focused on environmental and food HEV detection have shown viral presence in these kind of samples, highlighting possible transmission sources of HEV in the continent. HEV genotype 3 was the most frequently detected in the region, with HEV genotype 1 detected only in Venezuela and Uruguay. HEV is widely distributed throughout South America, producing sporadic cases of acute hepatitis, but as a possible agent of chronic hepatitis. Finding the virus in humans, animals, environmental samples and food, show that it can be transmitted through many sources, alerting local governments and health systems to improve diagnosis and for the implementation of preventive measures.

Chronic hepatitis E in the immunosuppressed: A new source of trouble?

Novel strains of hepatitis E virus identified from humans and other animal species: is hepatitis E a zoonosis?

Hepatitis E virus (HEV) is an expanding zoonotic viral disease threat. Although HEV causes acute viral hepatitis, it is increasingly being recognized as a systemic pathogen with detection and damage in extrahepatic tissues. The presence of HEV RNA in the semen of chronically infected human patients in the absence of viremia and fecal shedding and presence of HEV in the sperm head underscores the need to understand the interaction of HEV within the male reproduction system. Male accessory glands secrete biofluids necessary for sperm nourishment and to neutralize the acidity of the vagina. The role of male accessory glands in the dissemination and persistence of HEV infection have not been studied. Using an immunosuppressed pig model for chronic HEV infection, we demonstrate infectious HEV in mature sperm cells altering the sperm motility and morphology. HEV isolated from sperm cells remained infectious in human hepatoma cells. Spermatic fluid contained lower virus titers than the sperm cells from chronically infected pigs highlighting that the sperm cells themselves can associate with the virus. Evaluation of the male accessory glands demonstrated viral replication, infiltration of CD45 leukocytes, and apoptosis associated with HEV infection. A decrease in serum testosterone levels was evident in the HEV infected pigs. Even though a lower viral RNA titer was seen in serum and feces of chronically infected, immunosuppressed and ribavirin treated pigs, high viral RNA and infectious particles in sperm is a concern. Our findings necessitate further studies defining the mechanism of sperm cell invasion by HEV, length of HEV survival in sperm cells during chronic HEV infection, and risk of sexual transmission of HEV during both acute and chronic phases of infection.Author SummaryHepatitis E virus, a leading cause of acute viral hepatitis, causes both acute and chronic infection in humans. Recent advances within the HEV field have demonstrated extrahepatic diseases associated with HEV. More recent findings have revealed infectious HEV in the vagina, Sertoli cells, and ejaculate of humans, and sperm cells of pigs. We demonstrate that the male accessory sex glands may have a role in the persistence of HEV infection during chronic infections. We utilized an established immunosuppressed pig model and treated pigs with ribavirin to study the presence of virus in the sperm cells. We demonstrated high viral RNA loads and infectious particles associated with sperm cells. Our study further highlights the importance of the testis, as an immune privileged site, in the maintenance of chronic HEV infection. New studies to evaluate the mechanisms by which HEV associates with sperm cells, the length of HEV survival in sperm cell fractions, and consideration of the testes as a potential HEV reservoir are necessary.

Hepatitis E virus (HEV) is a zoonotic agent today considered a major Public Health issue in industrialized countries. HEV strains belonging to zoonotic genotype 3 are widely present in swine, being today considered important reservoirs for human disease. Unlike in swine, only scarce data are available on the circulation of HEV in wild boar. This study describes the detection and molecular characterization of HEV in livers from sylvatic wild boar hunted in Portugal and destined for consumption. Additionally, the detection of HEV in stools of a confined wild boar population also destined for consumption is also described. A total of 80 liver samples collected during the hunting season of 2011/2012 and 40 stools collected in February 2012 from a wild boar breeding farm in Portugal were tested by a nested broad-spectrum RT-PCR assay targeting open reading frame (ORF) 1. Twenty livers (25.0%) and 4 stools (10%) were positive for HEV. Phylogenetic analysis showed that all strains clustered with sequences classified as HEV genotype 3 subgenotype e. To our knowledge, this is the first report documenting the occurrence and molecular analysis of HEV in sylvatic and captive wild boar destined for human consumption in Portugal. This report demonstrates for the first time the circulation of HEV in wildlife reservoirs of Portugal adding knowledge to the epidemiology of HEV in wild boar populations.

Hepatitis E virus in lettuce and water samples: A method-comparison study

Molecular epidemiology of Hepatitis E virus among humans in the Niger Republic, 2017-2023.

A Novel Immunocompetent Rat Model of HCV Infection and Hepatitis

Paving the way for T cell-based immunotherapies in chronic hepatitis E

Transfusion-associated Hepatitis E, France

In vivo models for studying Hepatitis E virus infection; Updates and applications

To the Editors: Liver enzyme elevations (LEEs) in HIV-positive patients are mostly due to either viral hepatic coinfections or antiretroviral drug–induced hepatotoxicity, leaving a small number of patients with unexplained LEEs.1 However, it was recently shown that a part of the unexplained LEEs in the general population is attributable to locally acquired hepatitis E virus (HEV) genotype 3 infections—probably as the result of zoonotic transmission.2,3 Recent reports have shown that HIV-infected patients are at risk for persistent HEV carriage due to their immunocompromised state.4–6 Since then, the prevalence of HEV in several cohorts of HIV-infected patients has been described.7–12 Renou et al13 observed a geographical gradient in HEV seroprevalence, being lower in HIV-infected patients in northern France compared with the south. However, data from northern Europe are scarce.14,15 Considering the possibility of zoonotic transmission16,17 and the high HEV seroprevalence among pigs and boars in these northern European countries,18,19 investigating the HEV epidemiology in susceptible populations in these countries is important. Therefore, we conducted a study in the Netherlands evaluating the prevalence of locally acquired HEV infection in a cohort of HIV-infected patients with and without unexplained LEEs under follow-up in our clinic. The laboratory results of all HIV-infected patients under follow-up from January 2007 through February 2011 in the University Medical Center Utrecht (n = 1117) were reviewed for the occurrence of elevations in serum alanine aminotransferase (ALT) above the upper limit of normal (45 U/L in our hospital). Because we focused on locally acquired infections with HEV genotype 3, immigrants from regions where non–genotype 3 HEV is highly prevalent were excluded. A number of selected serum samples of unexplained LEEs from the period—negative for hepatitis A, B, or C infection—were obtained for HEV serological testing. As controls, another group of white patients—matched for age and gender—without LEEs was selected and a random sample obtained. All samples were tested—together, in 3 batches—for the presence of both anti-HEV IgM and IgG antibodies with a commercial enzyme-linked immunosorbent assay kit (recomWell HEV; Mikrogen GmbH, Neuried, Germany); positive and borderline results were tested by line blot assay (recomLine HEV IgG/IgM; Mikrogen GmbH). All tests were performed according to the manufacturer's instructions. Patient characteristics at the time the serum sample was obtained were registered. Continuous data were expressed as median value (with range) and analyzed using the Mann–Whitney U test, whereas categorical data are given as a percentage (with number) and analyzed using the Fisher exact test. Results are 2-sided, and P ≤ 0.05 was considered to be statistically significant. A total of 56 HIV-infected patients with unexplained LEEs were selected and matched for age and gender to a total of 50 HIV-infected patients without LEEs. Except for serum ALT values, there were no significant differences in patient characteristics between both groups (Table 1). In the total population (N = 106), 4 patients (3.8%) tested positive for anti-HEV IgG antibodies. Only 1 patient without LEEs tested HEV IgG positive. Within the group of patients with LEEs, 2 patients were HEV IgG positive, whereas 1 patient tested positive for both anti-HEV IgM and IgG antibodies indicating an acute HEV infection. However, a subsequently performed HEV polymerase chain reaction (PCR) on this serum sample was negative. No PCR was performed in the other patients who tested positive for IgG only because this indicated past exposure to HEV. No IgM was detected in any of the collected samples from the group without LEEs. Regarding the 4 HEV-positive patients with detectable IgG antibodies, 3 (75%) were men, with 2 of them being men who have sex with men. Mean age was 32 years, and the mean baseline CD4+ count was 528 cells per cubic millimeter. None of these characteristics differed significantly from the IgG-negative population. Again, only the baseline mean ALT value was significantly higher in the anti-HEV IgG antibody–positive patients (288 vs. 76 IU/L, P < 0.01).TABLE 1: Patient CharacteristicsThis study shows that locally acquired HEV infection occurs among HIV-positive patients in the Netherlands—probably as the result of zoonotic transmission.20 Furthermore, we observed an anti-HEV IgG seroprevalence rate of 3.8% in our whole cohort of HIV-infected patients, with a higher prevalence in those with LEEs (5.4% vs. 2%, P = 0.62). In the patients without LEEs, we did not detect any acute HEV infections. This observed seroprevalence is similar to the one found in the general Dutch population.2 In contrast, 2 recent studies14,15 reported a slightly higher anti-HEV IgG seroprevalence of 4.9% (German cohort) and 9.4% (British cohort) in a predominantly northern European population. No acute infections were detected in these cohorts either. Differences in patient characteristics and different available assays21,22 may have contributed to the observed differences in epidemiology between the study populations. Because recent observations have suggested a delayed anti-HEV seroconversion in HIV patients with very low CD4 counts,4 questions have been raised about the suitability of these tests for the detection of HEV infections,7 and together with the varying diagnostic value between the available assays, they support the argument that serological screening alone may be insufficient to diagnose HEV and should be complemented with detection of HEV by PCR. The recent developments regarding the epidemiology of HEV and the altered natural course in immunocompromised patients led to the discussion of whether HIV-positive patients with unexplained LEEs should be tested for HEV infection.23 With only 1 immunoblot-confirmed case of locally acquired acute HEV infection in the Dutch HIV-positive population, this might not be a cost-effective strategy. In conclusion, although HEV infection should be considered as a causative agent of unexplained LEEs in HIV-infected patients in northern European countries, the number of acute infections is low.

Hepatitis E Virus (HEV) is one of the leading causes of acute viral hepatitis, with ~20 million HEV infections worldwide per annum, and mortality rates up to 25% in pregnant women. However, many aspects of the biology of the virus are poorly understood. HEV has a positive-sense single-stranded RNA genome. ORF1 encodes the non-structural polyprotein required for viral RNA replication. This polyprotein (sometimes termed pORF1) is predicted to contain seven domains based on sequence homology to related viruses and it is hypothesised that this polyprotein must undergo proteolysis to generate functional protein units. However, it is unknown if the pORF1 polyprotein undergoes full proteolysis, the potential locations of any cleavage boundaries and whether a viral or host cell protease is responsible. We have adapted our in vitro-based proteolysis assays to investigate cleavage of HEV pORF1. In comparison to related RNA viruses, our data suggest that pORF1 has no auto-catalytic activity. Previous studies have shown that the liver-produced protease, thrombin, is essential for replication. In the presence of thrombin, we have shown that the ORF1 polyprotein undergoes specific proteolysis to produce eight distinct protein products. Combining bioinformatics with pORF1 truncations/mutagenesis, we have located the position of the pORF1 thrombin cleavage sites. Interestingly, these cleavage sites correspond to the junctions between the predicted pORF1 protein domains. Our data suggests that thrombin is an important cellular protease for controlling pORF1 proteolysis. Work is ongoing to understand the importance of each thrombin cleavage site in viral replication.