Major outbreaks of HEV infection in underdeveloped or developing countries have been associated with contamination of drinking water supplies with sewage and poor sanitary conditions. In contrast, in industrialised countries of the West, HEV infection occurs sporadically through autochthonous transmission from an animal reservoir, such as pigs, chickens, rodents, wild boars etc, possibly through consumption of improperly cooked meat.
The virus was first described in 1983 by Balayan and colleagues, some 30 years after the first documented epidemic in New Delhi, following ingestion by one of the team of a faecal extract prepared from stools collected during an outbreak in Afghanistan. Experimental infection of macaques with stool extract led to the isolation and cloning of the virus in 1990, using samples of bile obtained from such animals.
An immunoassay was developed the following year for the detection of antibodies to HEV (anti-HEV). Phylogenetic analysis of sequences of isolates obtained from epidemic and sporadic cases, as well as animals such as pigs revealed the existence of four genotypes of the virus, with genotypes 1 and 2 found entirely in humans in epidemic outbreaks, whilst genotypes 3 and 4 in both humans and pigs in sporadic cases. The latter also explains the zoonotic transmission of the virus.
The virus and genome organisation
The virus is now classified in the Orthohepevirus genus within the Hepeviridae family. The HEV particle is non-enveloped, has a diameter of 27-34nm and contains the 7.2 kb in length single-stranded RNA genome of the virus. The latter has three open reading frames, namely ORF1, 2 and 3, which encode for the non-structural proteins, the capsid protein and another protein of unknown function respectively. The capsid protein carries a number of antigenic epitopes the role of which is pivotal in the detection of antibodies and in vaccine development as discussed later. It is thought that the protein has both linear and conformational epitopes.
As mentioned above, nucleotide sequencing of a number of HEV isolates from different regions of the world followed by phylogenetic tree analyses have identified four different genotypes, designated 1-4, each of which is further subdivided into a number of subtypes. Genotype 1 has five subtypes (designated a-e), genotype 2 has two (a,b), genotype 3 ten (a-j) and genotype 4 seven (a-g). The human genotypes 1 and 2 have been isolated from endemic and epidemic cases in Asia (India, Pakistan, Myanmar, Kyrgyzstan, and China), whilst genotype 2 is more common in Central America and Africa (Mexico, Chad, Morocco, Nigeria, Namibia and Egypt). Genotype 3 is predominantly the cause of sporadic cases occurring worldwide (Asia, Europe, Oceania, North and South America) and genetically it groups together with swine isolates. Genotype 4 stains originate from the Far East and have been isolated from both swine and human samples.
Epidemic outbreaks of HEV infection have involved in some instances many thousands of people and have usually been associated with heavy rains resulting in sewage contamination of drinking water from flooding, and poor hygiene conditions as may happen in refugee camps.
In India, sporadic case of HEV infection occur throughout the year and account for 50-70% of all cases of sporadic viral hepatitis, whilst in industrialised countries of the West sporadic HEV infection was in the past associated with recent travel to endemic areas. Nowadays of course such cases can also be attributed to indigenous transmission through contact with swine (and other animals) as may be the case with pig farmers and sewage workers. Consumption of inadequately cooked meat is another source of infection.
The majority of HEV infections may have a clinically silent course. During outbreaks, the percentage of those affected may vary between 1-15%, being more frequent in adults (3-30%) than children (0.25-10%). Men are more commonly affected than women and person to person spread is not common unless within an affected household.
Deaths as a result of infection range from 0.2-4%, with a much higher fatality rate (20%) due to fulminant hepatitis in pregnant women in the 3rd trimester of pregnancy. Fatalities were observed in 22.2 % of women in the 3rd trimester who developed fulminant disease and 2.8% of men. The more severe outcome in pregnancy appears to be related to diminished cellular immunity and hormonal factors.
Acute HEV infection is normally self-limiting leading to full recovery. However, in recent years protracted infection has been described in liver and kidney transplant patients, as well as patients with haematological malignancies, whereby the infection may culminate in the development of cirrhosis. In all such cases HEV genotype 3 has been implicated. Chronic HEV infection in solid organ transplant patients has been associated with the use of tacrolimus for immunosuppression, low platelet counts and impaired HEV-specific T-cell responses. Chronic HEV infection in Human immunodeficiency virus (HIV)-infected individuals in view of their likely immunosuppression, does not seem to be a problem.
The incubation period may be variable ranging between 2-10 weeks. In icteric cases symptoms may include a short flu-like prodrome with fever, mild chills, abdominal pain, anorexia, nausea, vomiting, clay-coloured stools, dark or tea-coloured urine, diarrhoea, arthralgia, asthenia and a transient macular skin rash. On physical examination the patient may have jaundice, mildly enlarged and tender liver, whilst splenomegaly may also be present. Liver transaminases, alkaline phosphatase and bilirubin in serum are elevated, returning to normal on resolution up to 6 weeks after the appearance of symptoms. Histological examination of liver biopsies may show the presence of ballooning hepatocytes, acidophilic body formation and focal or confluent hepatocyte necrosis. In nearly half of the cases there is evidence of cholestatic hepatitis, whilst the lobules and portal tracts have inflammatory infiltrates. In the case of fulminant disease there is submassive or massive necrosis and collapse of the parenchyma.
Serological and other tests
The presence of IgM anti-HEV in serum is indicative of acute infection, and gives way to IgG anti-HEV in the convalescent phase or in cases (Fig 1). The latter may remain detectable for many years after convalescence in up to half of subjects. Diagnostic tests are based on the detection of anti-HEV using assay formats that employ recombinantly expressed viral capsid proteins, or synthetic peptides representing immunogenic epitopes. However, the sensitivity and specificity of these tests is variable, which may explain some of the discrepancies that are apparent in certain seroprevalence studies.
Detection of HEV-RNA using reverse transcription polymerase chain reaction with appropriate primers is a marker of ongoing infection. HEV-RNA is detectable in patient faeces during the initial few weeks of infection and prior to the rise in transaminase levels (Fig 1). HEV-RNA has been detected in serum for a short period during the acute phase. In view of this, IgM anti-HEV detection still remains the main diagnostic marker of acute infection.
Antibody from convalescent human sera and sera from experimentally infected primates have been shown to exhibit cross-reactivity with diverse strains, to neutralise HEV in vitro and to protect non-human primates against infection following challenge with live HEV. Since the virus was shown to have only one serotype of the virus which elicited neutralising antibodies directed against the capsid protein (ORF 2), then these findings bode well for the development of an effective vaccine. Two vaccines have been produced, both of which contain truncated capsid proteins that self-assemble into virus like particles.
The first recombinant HEV vaccine (a 56kDa protein encompassing aminoacids 112-607) was tested in anti-HEV negative healthy adults, mostly males from the Nepalese army. After 3 doses of the vaccine, 100% of the recipients developed anti-HEV one month post-third dose, and vaccine efficacy was estimated at 95.5%. HEV 239 is the second recombinant vaccine (Hecolin, Xiamen Innovax Biotech, China) expressed in bacteria and encompasses aminoacid positions 368-606 of ORF 2 (Chinese genotype 1 strain). This vaccine was trialled in the Jiangsu province near Shanghai involving over 112000 individuals. After three doses of the vaccine antibodies were detectable in 100% of the immunised individuals after the third dose and complete protection from HEV infection during the 13 months of follow-up was observed. There were 15/48663 (0.0003%) cases of HEV infection in the placebo group, all but one being due to genotype 4.
HEV being a self-limiting disease leading to complete recovery does not warrant antiviral treatment. However, in the case of protracted /chronic HEV infection ribavirin treatment (600-1000mg/day) for at least 3 months has been shown to lead to a virologic response in the majority of treated patients, even though the number treated remains quite small. In patients with organ transplants, reduction in immunosuppression should be considered first, whilst interferon treatment should be weighed against the possibility of organ rejection.