Hepatitis Viruses. Agents of Infectious & Serum Hepatitis.

1. THEME: HEPATITIS VIRUSES. CAUSATIVE AGENTS OF THE INFECTIOUS
AND SERUM HEPATITIS. LABORATORY DIAGNOSTICS AND PROPHYLAXIS
DISEASES.
I. STUDENTS’ INDEPENDENT STUDY PROGRAMME
1. General characteristic and classification of the hepatitis viruses.
2. Morphology, antigen structure, and resistance of the hepatitis B virus.
3. Morphology, antigen structure, and resistance of the hepatitis D, C, and G viruses.
4. Epidemiology of hepatitis with parenteral route of transmission.
5. Morphology, antigen structure, and resistance of the hepatitis A and E viruses.
6. Epidemiology of hepatitis with ingestion (fecal-oral) route of transmission.
7. Pathogenesis and laboratory diagnostics of the infectious hepatitis:
a. Rapid diagnostics (IEM, detection of the viral antigens in the feces with ELISA);
b. Serological method (CFT, PHAT)
8. Pathogenesis and laboratory diagnosis of the parenteral hepatitis:
a. Modern methods (ELISA test, immunoblotting)
9. Specific prophylaxis of hepatitis: characteristics of the vaccines.
1. The term “viral hepatitis” refers to a primary infection of the liver by any one of a
heterogeneous group of “hepatitis viruses”. It consists of types A, B, C, D, E, G.
Hepatitis viruses are taxonomically unrelated (DNA and RNA viruses). The features
common to them are: hepatotropism, ability to cause a similar icteric illness
By epidemiological and clinical criteria, two types of viral hepatitis had been
recognised for long:

2. A first type (infective or infectious hepatitis) is occurred sporadically or as epidemics;
affecting mainly children and young adults; transmitted by the fecal-oral route.A second type
(serum hepatitis or transfusion hepatitis) transmitted mainly by parenteral route.
2. Type B hepatitis (HBV)
HBV is assigned to a separate family Hepadnaviridae
Morphology: HBV is a 42 nm, DNA virus with an outer envelope and inner core, enclosing
the viral genome and a DNA polymerase
Under the electron microscope, sera from type B hepatitis patients show 3 types of particles:
spherical (20 nm in diameter), tubular (20 nm in diameter) and these two types of particles
represent Australia antigen.
The third type of particles are double shelled spherical (42 nm) and also called Danes
particles
Antigen Structure:
HBsAg – hepatitis B surface antigen (glycoprotein)
HBcAg – hepatitis B core antigen (nucleocapsid)
HBeAg – hepatitis B core antigen, is derived from HBcAg (contains viral DNA polymerase
enzyme)
Resistance: HBV is a relatively heat stable virus (It survives at room temperature for long
periods). Heat at 600C for 10 hours reduces infectivity by hundred- to thousand fold
It is susceptible to chemical agents: hypochlorite, 2% gluteraldehyde
3. Type D (Delta) hepatitis (HDV)
HDV is a defective RNA virus depending on the helper function of HBV for its replication
and expression. It belongs to genus Deltavirus
Morphology: It is spherical, 36-38 nm diameter; RNA particle surrounded by HBsAg
envelope. The genome is a single stranded small circular molecule of RNA. It encodes
its own nucleoprotein, the delta antigen, but the outer envelope of HDV is encoded by
the genome of HBV coinfecting the same cell
Epidemiology: There are three important modes of transmission of HBV and HDV infection:
parenteral, per natal, sexual

3. The incubation period is long (about 1- 6 months)
Two types of hepatitis B and D infection are recognized:
Coinfection: delta and HBV are transmitted together at the same time. Coinfection clinically
presents as acute hepatitis B, ranging from mild to fulminant disease
Superinfection: delta infection occurs in a person already harbouring HBV. Superinfection
usually leads to more serious and chronic illness
The clinical picture of hepatitis B is similar to that of type A, but it tends to be more severe
and protracted
The pathogenesis of hepatitis appears to be immune mediated. Hepatocytes carry viral
antigens and are subject to antibody-dependent NK cell and cytotoxic T-cell attack
In the absence of adequate immune response HBV infection may not cause hepatitis, but may
lead to carrier state
Laboratory diagnosis of the HBV infection
Laboratory diagnosis of HBV infections can be carried out by detection of hepatitis B
antigens and antibodies (viral markers). These can be detected by sensitive and specific
tests like ELISA and RIA
HBsAg – it is the first marker to appear in blood after infection. Peak levels of HBsAg are
seen in the preicteric phase of the disease. It remains in circulation throughout the
icteric or symptomatic course of the disease
HBeAg – appears in the serum at the same time as HBsAg. HBeAg is an indicator of active
intrahepatic viral replication and the presence in blood of HBV DNA, virions and DNA
polymerase.
HBcAg – is not detectable in the serum but can be demonstrated in liver cells by
immunofluorescence. Anti-HBc antibody is the earliest antibody to appears in the blood.
Viral DNA polymerase – it appears transiently in serum during preicteric phase (the level
DNA can be detected in serum by PCR)
Laboratory diagnosis of the HDV infection
Delta antigen is primarily expressed in liver cell nuclei, where it can be demonstrated by
immunofluorescence
Anti-delta antibodies appear in serum and can be identified by ELISA test
Type C hepatitis (HCV)
Hepatitis C virus belongs to the family Flaviviridae
Morphology: HCV is a 50-60 nm virus with a linear single stranded positive RNA . Enclosed
within a core and surrounded by an envelope, carrying glycoprotein spikes
HCV infection is seen only in humans. The source of infection is the large number of carriers

4. The incubation period is long (15-160 days). The acute illness is usually mild or unicteric
The hepatitis progress to chronic hepatitis, with some developing cirrhosis and hepatocellular
carcinoma
Laboratory diagnosis
It can be established by detection of anti-HCV by ELISA. Antibody detection becomes positive
only month after the infection
Viral genome (HCV RNA) can be detected by PCR and immunofluorescence.
Type G hepatitis (HGV)
In1996, this virus was first isolated from a patient with chronic hepatitis
It has been placed in family Flaviviridae
Morphology of the virus is like to hepatitis C virus.
HGV RNA has been found in patients with acute, chronic and fulminant hepatitis,
haemophillics, patients with multiple transfusions, blood donors and intravenous drug
addicts
The virus is transmitted parenterally, sexually and from mother to child
Infection may occur in patients coinfected with hepatitis C
HGV infection can be detected by reverse transcriptase polymerase chain reaction
Type A hepatitis (HAV)
Morphology. HAV is a spherical RNA virus, 27-30 nm in diameter, non enveloped, lipid is not
an integral component
Belongs to the Picornaviridae family, genus Hepatovirus
Resistance
HAV is resistant to heat at 60oC for one hour, acid at pH 3, boiling for one minute,
autoclaving 1210C for 20 minutes, 1:4000 formaldehyde at 370C for 72 hours
It survives prolonged storage at a temperature of 40C or below
Epidemiology: HAV transmission is by the fecal-oral rout
Pathogenesis: The virus multiplies in the intestinal epithelium and reaches the liver by
hematogenous spread
The clinical disease consists of two stages: the prodromal (or preicteric) and the icteric stage

5. The onset may be acute or insidious, with fever, malaise, anorexia, nausea, vomiting and liver
tenderness.
There is also jaundice, with yellowing of the skin and the whites of the eyes and the dark urine
typical of liver infections
Liver damage is probably caused by immunological reactions
Laboratory diagnosis
Etiological diagnosis of type A hepatitis may be demonstration of the virus or its antibody
IEM – the virus can be visualized in fecal extracts
Serological tests: CFT, immune adherence, reaction of the passive hemagglutination,
radioimmunoassay and ELISA (by detection of antibody: Ig M and Ig G)
Type E hepatitis (HEV)
HEV is a spherical non enveloped virus, 32-34 nm in diameter, with a single stranded RNA
genome, the surface of the virion shows indentation and spikes
Hepatitis E virus belongs to family Caliciviridae
This virus causes enterically transmitted E hepatitis
Hepatitis E has been shown to occur in epidemic, endemic and sporadic forms
It occurs predominantly in young to middle-aged adults
Clinically the disease resembles that of hepatitis A
The disease is generally mild and self limited, with a low case fatality
A unique feature is the clinical severity and high case fatality in pregnant women, especially
in the last trimester of pregnancy
Laboratory diagnosis
Immunoelectron microscopy – feaces is examined by electron microscopy of aggregated
calicivirus-like particles using monoclonal antibodies
ELISA test and western blot assay : these are used for detection of Ig M and Ig G antibodies
Polymerase chain reaction : HEV RNA can be detected in faeces or acute phase sera of
patients
Prophylaxis of the serum hepatitis includes:

6. General preventive measures (these include health education, improvement of personal
hygiene and strict attention to sterility. Prophylaxis of the serum hepatitis is also
included blood or blood products screening. Avoidance of use of unsterile needles,
syringes and other material is another important general prophylactic measure
Immunisation is carried out only for prophylaxis of the hepatitis A, B, and D.
Passive prophylaxis of the hepatitis B and D– use hepatitis B immunoglobulin (HBIG is
prepared from donors with high titres of anti-HBs
Active of the hepatitis B and D . Following vaccine preparation may be useful:
HBsAg from human carriers;
HBsAg produced in cell line from human hepatocellular carcinoma
HBsAg inserted genome in plasmid (genetic engineering)
Vaccine from polypeptide HBsAg
Prophylaxis of the infectious hepatitis:
General prophylaxis consists of improved sanitary practices; prevention of fecal
contamination of food and water
Specific prophylaxis (only hepatitis A)
Active– use a live attenuated or inactivated vaccine (protection beings 4 weeks after injection
and lasts for 10 to 20 years
Passive – use normal human immuniglobulin
II. Students Practical activities:
1. To diagnose hepatitis A with CFT. Estimate titer of the complement-fixing antibody in
the paired sera of the patient with hepatitis. Make the conclusion based on the antibody
titer rise.
2. Estimate results of the ELISA which have been carried out for revealing of HBsAg in the
test sera collected from 10 patients with hepatitis. Write down the stages of the ELISA
for detection specific antigen in the collected samples.
3. Acquaint with preparations for specific prophylaxis and treatment of the viral hepatitis.
Note medicines for treatment of the serum hepatitis/

7. General preventive measures (these include health education, improvement of personal
hygiene and strict attention to sterility. Prophylaxis of the serum hepatitis is also
included blood or blood products screening. Avoidance of use of unsterile needles,
syringes and other material is another important general prophylactic measure
Immunisation is carried out only for prophylaxis of the hepatitis A, B, and D.
Passive prophylaxis of the hepatitis B and D– use hepatitis B immunoglobulin (HBIG is
prepared from donors with high titres of anti-HBs
Active of the hepatitis B and D . Following vaccine preparation may be useful:
HBsAg from human carriers;
HBsAg produced in cell line from human hepatocellular carcinoma
HBsAg inserted genome in plasmid (genetic engineering)
Vaccine from polypeptide HBsAg
Prophylaxis of the infectious hepatitis:
General prophylaxis consists of improved sanitary practices; prevention of fecal
contamination of food and water
Specific prophylaxis (only hepatitis A)
Active– use a live attenuated or inactivated vaccine (protection beings 4 weeks after injection
and lasts for 10 to 20 years
Passive – use normal human immuniglobulin
II. Students Practical activities:
1. To diagnose hepatitis A with CFT. Estimate titer of the complement-fixing antibody in
the paired sera of the patient with hepatitis. Make the conclusion based on the antibody
titer rise.
2. Estimate results of the ELISA which have been carried out for revealing of HBsAg in the
test sera collected from 10 patients with hepatitis. Write down the stages of the ELISA
for detection specific antigen in the collected samples.
3. Acquaint with preparations for specific prophylaxis and treatment of the viral hepatitis.
Note medicines for treatment of the serum hepatitis/