Global Alert and Response (GAR)

Hepatitis E

The hepatitis E virus

- Morphology and physicochemical properties
- Genome and proteins
- Antigenicity
- Stability

HEV causes self-limited acute viral hepatitis in adults aged 15-40.

HEV is a nonenveloped, spherical, positive-stranded RNA virus. Several different strains have been isolated, partially characterized and molecularly cloned (1990-92).20, 41, 44, 49, 53 Although originally classified within the family of caliciviruses, they are now unclassified.17, 39

Replication in cell culture was first reported in 1993; yields of virus are generally very low.18 In natural infections, the virus replicates in hepatocytes.48 In vivo infected macaque hepatocytes support HEV replication after isolation and placement into tissue culture.50

Man is the natural host for HEV, but certain non-human primates, e.g. chimpanzees, cynomolgus monkeys, rhesus monkeys, pigtail monkeys, owl monkeys, tamarins and African green monkeys are reported to be susceptible to natural infection with human strains of HEV.10, 26, 32, 41, 44, 48, 56

Transmission of human strains of HEV to swine has been reported for the US-2 strain of human HEV, but could not be shown for the Sar-55 and Mex-14 strains.32, 35

A swine strain of HEV has been isolated, identified and characterized and subsequently shown to infect rhesus monkeys and chimpanzees, experimental surrogates of humans. This ability to cross species barriers puts humans at risk for infection with swine HEV.32, 33

In endemic areas, naturally acquired anti-HEV has been found in 42 - 67% of cows, sheep and goats.12, 14

Recent studies show evidence of widespread HEV or HEV-like infection in rodents in the US (prevalence rate of 60% in rats), raising the question of transmission, reservoirs, and strains of HEV in developed countries.13, 24, 47

Click here for: Electron Microscopy (EM) picture

Morphology and physicochemical properties
HEV is a small and structurally simple RNA animal virus.

The virion is nonenveloped and, with a diameter of 27-34 nm, is composed entirely of viral protein and RNA. Electron microscopy (EM) analyses show spherical particles of possible icosahedral symmetry, with indefinite surface substructure, resembling the caliciviruses.18, 41

Morphologically, HEV is similar to Norwalk virus, a member of the calicivirus family, although the sequence of HEV most closely resembles the sequence of rubella virus, a togavirus, and beet necrotic yellow vein virus, a plant furovirus.18, 41, 48

Full virions have a buoyant density of 1.29 g/cm3 in potassium tartrate/glycerol gradients and a sedimentation coefficient of 183 S in neutral sucrose gradients, empty capsids of 165 S under the same conditions.9, 10, 41

Genome and proteins
The hepatitis E genome consists of a linear, single-stranded, positive-sense RNA (that is, mRNA) of approximately 7.5 kb containing a 3' poly(A) tail and short 5' and 3' noncoding (NC) regions.10, 18, 19, 41

Three overlapping open reading frames (ORFs) exist, and all three coding frames are used to express different proteins.10, 29, 41

ORF1 (5 kb) is located towards the 5' end of the genome and encodes a polyprotein of about 1690 amino acids that probably undergoes post-translational cleavage into multiple nonstructural proteins required for virus replication, including a methyltransferase, a putative papain-like cystein protease, an RNA helicase and an RNA-dependent RNA polymerase.18, 19, 29, 41

ORF2 does not overlap with ORF1; it is located at the 3'-end of the genome and encodes the principal and probably only structural protein. It is a capsid protein of 660 amino acids (71 kDa).18, 19, 29, 41, 56

ORF3 begins with the last nucleotide of ORF1; it overlaps extensively with ORF2 and is the shortest of the open reading frames, encoding a small immunogenic 123 amino acid phosphoprotein (14.5 kDa) which associates with the cytoskeleton, suggesting a possible role in the assembly of virus particles.18, 29, 41, 65

The genomes of several HEV strains from different parts of the world have been sequenced and compared. Overall, they appear to fall into four major genetic groups:

1 - South-East Asian (Burmese, some Indian strains), North and Central Asian (strains from China, Pakistan, Kyrgyzstan, and a few from India), and North African strains form one somewhat heterogeneous genotype,
2 - the single North American (Mexico) isolate comprises a second,
3 - the US and swine isolates comprise a third and
4 - a subset of isolates from China and most isolates from Taiwan comprise a newly described fourth group.

Genetically heterogeneous isolates from several European countries have been designated new genotypes, but, at this time, probably should be grouped with the US isolates into a large, heterogeneous group.9, 18, 20, 41, 44, 46, 53 Two novel isolates of HEV have recently been described in Argentina. Distinct from all previously described isolates, they represent two diverse subtypes of a new genotype of HEV.47

The genome of swine HEV, an animal strain of HEV, has recently been identified and characterized. The putative capsid gene (ORF2) of swine HEV shares about 80% sequence identity at the nucleotide level and about 92% identity at the amino acid level with that of human HEV strains. The small ORF3 of swine HEV has about 84% nucleotide sequence identity and about 80% amino acid identity with human HEV strains.33

Click here for: Genetic map
Click here for: Comparison of amino acid sequences of different HEV strains
Click here for: Model of HEV replication

All HEV strains studied to date (1998) appear to comprise a single serotype.29, 40, 41

Major epitopes appear to exist near the carboxyl ends of ORF2 and ORF3. Epitopes contained in ORF2 are more conserved (90.5%) than epitopes contained in ORF3 (73.5%) in different strains.41

Western blot data indicate that type-specific (virus-specific) epitopes exist and can be used to differentiate serologically different isolates.10, 19

Serologic tests for anti-HEV based upon expressed ORF2 sequences are more sensitive for detecting IgM and IgG anti-HEV than are tests based upon antigens containing ORF3 sequences. In fact, proteins expressed from ORF2 measure antibodies that correlate with protection against hepatitis E, whereas no such correlation has been shown for antibodies to ORF3.16

Antibodies to swine HEV cross-react with capsid antigens from strains of human HEV.

No serologic or hybridizing cross-reactivity between HEV and other viral hepatitis agents, including hepatitis A virus (HAV), has been observed.

HEV is extremely sensitive to high salt concentrations (CsCl).9, 10

HEV should be stored as cold as possible, although it is rapidly degraded when freeze-thawed. The virus is sensitive to degradation by proteolytic enzymes.9, 10, 52

HEV is excreted from the liver via the common bile duct into the duodenum of the small intestine. Survival in the gastrointestinal tract suggests relative stability to acid and mild alkaline conditions.10, 18, 23, 41

The amount of infectious virions shed in the faeces during infection is low, consistent with the low rates of secondary spread during epidemics.18

Virions remain unaltered after exposure to trifluorotrichloroethane.41

Outbreaks of HEV have been successfully controlled by chlorination of water supplies. Iodinated desinfectants or autoclaving destroys the virus.2

For transportation, specimens containing HEV should be kept frozen in dry ice (solid CO2, -70°C), or preferably in liquid N2 (-120°C).52


Electron Microscopy (EM) picture

From: Centers for Disease Control and Prevention (CDC), Atlanta, USA:11

Electron microscopy picture of human hepatitis E virus.


Genetic map

Organization of the genome of HEV. The positive-sense single-stranded RNA genome of the prototype strain of HEV, recovered in Myanmar, consists of a 5' NC region of 27 nucleotides, followed by an ORF (ORF1) of 5079 bases. A second ORF (ORF2) begins in coding frame two, 38 nucleotides 3' of the termination of ORF1 and consists of 1980 nucleotides. A 65-nucleotide 3' NC region is contiguous with the termination of ORF2 and is terminated by a 3' stretch of 150 to 200 or more adenosine residues. A third ORF, 369 nucleotides in length, overlaps ORF1 by one nucleotide at its 5' end and extends into ORF2 in coding frame three. The HEV genome is capped. The three open reading frames (ORFs) expressed during infection are shown. Putative functions (protein products of each ORF) are labelled beneath.9, 10, 20, 25, 29, 48, 53

Comparison of amino acid sequences of different HEV strains

Click here for: amino acid sequences (pdf.document)

From: Meng X-J et al.. A novel virus in swine is closely related to the human hepatitis E virus. Proceedings of the National Academy of Sciences USA, 1997, 94:9860-9865,33 "Copyright (1997) National Academy of Sciences, U.S.A.", with permission.

Alignment of the amino acid sequences of ORFs2 (A) and 3 (B) of swine HEV with human strains of HEV. The sequence of Sar55 strain is shown on top, and only differences are indicated. Deletions are indicated by a minus. The putative hypervariable region (HVR) in the ORF3 is indicated by asterisks. Sequences used in this alignment are Burma49, Mexico20, NE8L (Myanmar7), Hyderabad (India37), Madras (India, GenBank accession no.X99441), HEV037 (isolate from a case of fulminant hepatitis, GenBank accession no.X98292), Sar55 (Pakistan53), KS2-87 (China64), Hetian (China, GenBank accession no.L08816), and Uigh179 (China8).20, 33

Model of HEV replication

From: Krawczynski K, Mast EE, and Perdy MA. Hepatitis E : an overview. In: Rizzetto M, Purcell RH, Gerin JL, and Verme G, eds. Viral hepatitis and liver disease, Turin, Minerva Medica, 1997:305-312.28 "Copyright (1997) Edizioni Minerva Medica, Italy", with permission.

Model of HEV replication. HEV: hepatitis E virus. pORF1, 2, 3: proteins encoded by open reading frames 1, 2, 3. 3IP: host protein reacting with protein encoded by ORF3.21 The evidence supporting the existence of HEV subgenomic mRNA derives from Yarbough et al.63, and the data suggesting the compartmentalization of glycosylated pORF2 were generated by Jameel et al.21. The model represents an extension and modification of the HEV replication strategy proposed by Reyes et al.43, 28