VIRUS HEPATITIS

Viral causes of human hepatitis

Hepatis viruses Other viruses

 Coxsackievirus
 HAV
 Cytomegalovirus
 HBV  Dengue virus
 HCV  Epstein-Barr virus
 Herpes simplex virus
 HDV  Lassa virus
 HEV  Measles virus
 Parainfluenza viruses
 G BV-C  Reovirus
 Rubella virus
 Varicella-zoster virus
 Yellow fever virus
Hepatitis Virus A (HVA)
 Nonenveloped
 Positive-sense, single-
stranded RNA
 Family Picornaviridae
 Hepatovirus genus
 Subclassified into 12
genera based on genotypic
and serologic
Hepatitis A virus (HAV)

 HAV is one kind of picornavirus and used to be classified as

enterovirus type72, but recently, it is considered to be classified as
heparnavirus
 Hepatitis A virion is a naked spherical particle, diameter 27nm
 Consists of a genome of linear, single-stranded RNA, 7.5kb. The
genome may be divided into 3 coding region: P1 region (encoding
structural protein), P2 and P3 regions (encoding non-structure protein)
 During acute stage of infection, HAV can be found in blood and feces
of infected human and primates
 Marmoset and chimpanzee are susceptible animals
Hepatitis A virus (HAV)

 HAV can not cause cytopathy, replicate within cytoplasma of

hepatocytes and via bill are discharged with feces
 7 genetypes, 1, 2, 3, 7 types from humanbody
 Only one antigen-antibody system. Anti-HAV IgM is diagnostic
evidence of recent infection, IgG is protective antibody.
 Resistance of HAV: 56°C, 30 min, usually temperature 1 week, dry
feces at 25°C 30 days, fresh water, sea water ,shellfish or soil for
several months. 70% alcohol at 25°C , 3 min, 100°C, 5 min and
ultraviolet, 1 min
HAV differs from other picornaviruses

 Resistant to high temperatures, low pH, and drugs that

inactivate many picornaviruses
 Replicates very slowly and generally without cytopathic
effect in cell culture
 Nucleotide and amino acid sequences, as well as
predicted sizes of several HAV proteins are dissimilar
 Only one serotype of HAV has been identified, and one
antigenic neutralization site is immunodominant.
Epidemiology
HAV Infection
 Humans and nonhuman primates
 Spread fecal – oral
 Person to person
Hepatitis A virus (HAV) genome organization
Hepatitis A virus (HAV) infectious life cycle
Pathogenesis and Host Immune Responses
Laboratory Diagnosis
Hepatitis C Virus
 HCV is a member of flavivirus family.
 HCV genome is a single stranded positive-sense RNA and
contains 9.4kb
 The genome contains 5’-non coding region, C region, E
region and NS region
 HCV genome may be divided into many types and
subtypes.
 Resistance
 Antigen-antibody system
 The concentration of HCV in blood is low, HCV Ag has not be
detected, anti-HCV is the indicator of infection and the marker of
infectivity
 HCV-RNA
HCV-RNA may be detected from blood or liver tissue, it’s the
direct evidence of infectivity
Hepatitis B virus (HBV)
 HBV is a kind of hepadnavirus
 Three particles in serum:
spherical particles and tubular particles with a diameter of
20 nm, composed of HBsAg
large particles with a diameter of 42 nm, named Dane
particle. It consists of an outer protein shell (envelope, contain
HBsAg) and an inner body ( core, contain HBcAg, HBeAg,
HBV-DNA and DNAP )
Hepatitis B virus (HBV)
 Hepatitis B virion genome is a small circular, partially
double stranded DNA with 3200 nucleotides long.
HBV DNA is asymmetry in length of two strands:
minus strand (long strand, L) has full length. Four
open reading frames (ORF) coded on the minus
strand: C, S, X, and P region
Hepatitis B virus (HBV)

 Four open reading frames (ORF)

S region: include pre-s1, pre-s2 and S gene,
encoded pre-s1 protein, pre-s2 protein and HBsAg.
Pre-s1 protein + pre-s2 protein + HBsAg—large protein
Pre-s2 protein + HBsAg—middle protein
HBsAg—major protein
C region included pre-c and C gene, encode HBeAg and
HBcAg
X region encoded HBxAg
P region encoded DNA polymerase
 Three antigen-antibody system
HBsAg-- anti-HBs system:

 Include HBsAg, anti-HBs, pre-s1,s2 antigen and anti-pres1, s2

 HBsAg appears 1-2 weeks (late to 11-12 weeks) after exposure, persists for
1-6 weeks( even 5 months) in acute hepatitis B.
In chronic patients or carrier, HBsAg persist many years
 HBsAg antigencity but no infectivity
 HBsAg is the marker of infectivity
 HBsAg can be found in humors and secretions: salive, urine, semina, tears,
sweat and breast milk
 10 subtype of HBsAg, 4 major subtypes: adr, adw, ayr, ayw.
 Anti-HBs appear after HBsAg disappear several weeks (or months) anti-HBs
is protective antibody, can persist for many years
 pre-s1 and pre-s2 antigens appear following HBsAg . They are the marker of
infectivity. Anti –pre s2 has the action of clearing virus
 HBcAg—anti-HBc system
 HBcAg can be found in the nuclei of liver cells, no free HBcAg
in serum
 HBcAg is the marker of replication of HBV
 The stage called window phase
 Anti-HBc IgM is a marker of acute infection and acute attack
of chronic infection of HBV. Anti-HBc IgG is the marker of past
infection, high titer means low level replication of HBV
 HBeAg—anti-HBe system
 HBeAg is a soliable antigen
 HBeAg is a reliable indicator of active replication of
HBV
 Anti-HBe is a marker of reduced infectivity. If exist
long may be a marker of integration of HBV into liver
cell
 The marker of molecular biology of HBV
 HBV-DNA
The direct indicator of HBV infection
Can integrate into the genome of hepatocytes
 HBV DNA polymerase
Possesses the ability of reverse transcriptase
and the indicator of the ability of replication of
HBV
Etiology
 HBxAg
Related to chronicity, activity of hepatitis B or liver
cancer
 Resistance
Resistant to heating and common disinfections.
Chimpanzee is susceptible to HBV
Laboratory Tests for HBV
 Serology:
 Many tests available – most common tests are Enzyme
Immunoassays (EIAs, MEIAs)
 First tests available in 1972
 For every rule, there is an exception/caveat
 No single test tells you everything
 Molecular:
 HBV DNA (quantitative)
 HBV genotyping
 HBV resistance testing
 Hepatitis B – Laboratory Tests
Serologic markers:
1) HBsAg (Hepatitis B surface antigen):
• if positive, person is infectious
• Sensitivity = 0.15 ng/ml
• Specificity = 99.5%
2) Anti-HBs (Antibody to HBV surface antigen):
• indicates immunity to HBV and protection from
disease
• Protective level is >10 IU/ml
 Hepatitis B – Laboratory Tests
Serologic markers:

3) Anti - HBc (Antibody to HBV core antigen):

• Total - indicates past or active infection;
present whether person is immune or chronic
carrier
• Specificity = 99.8% to 99.9%
• IgM - early indicator of acute infection
• No antigen test
 Hepatitis B – Laboratory Tests

Serologic markers:
4) HBeAg (Hepatitis Be antigen):
• indicates person is highly infectious
• Selecting patients for therapy
5) Anti-HBe (Antibody to HBVe antigen):
• prognostic for resolution of infection;
less infectious; spontaneous
seroconversion in 10 to 20% of healthy
adults per year
Acute Hepatitis B Virus Infection with Recovery
Typical Serologic Course

Symptoms
HBeAg anti-HBe

Total anti-HBc
Titer

HBsAg IgM anti-HBc anti-HBs

0 4 8 12 16 20 24 28 32 36 52 100
Weeks after Exposure
 Progression to Chronic Hepatitis B Virus
Typical Serologic Course
Acute Chronic
(6 months) (Years)
HBeAg anti-HBe
HBsAg
Total anti-
HBc
Titer

IgM anti-
HBc

0 4 8 12 16 20 24 28 32 36 52 Years
Weeks after Exposure
Virological and Biochemical Course
of Chronic Hepatitis B
 Disease Phases in Chronic HBV
Infection
Phase HBsAg HBeAg Anti- ALT HBV DNA
HBe range
Immune + + - Normal >8 log IU/mL
Tolerant

Immune + + - Normal or 3-8 log IU/mL

Clearance elevated

Inactive + - + Normal <3 log IU/mL

Disease

HBeAg- + - + Normal or 3-8 log IU/mL

negative elevated
Chronic
HBV
Interpretation of Serologic Tests in
Hepatitis B
Hepatitis B – Laboratory Tests

Serologic markers – caveats:

 Persistent HBsAg for >6 mos = chronic infection
 HBsAg and anti-HBs may co-exist in up to 24% of
chronically infected individuals; likely due to mutations
in the “a” determinant of the S gene
 Surface antigen escape mutants described in infants infected
with HBV after HBIG + vaccination and in Liver transplants
after prolonged HBIG
 Anti-HBc IgM may persist for up to 2 years in 20%;
chronically infected individuals may have low titres
which rise during acute flares
 Hepatitis B – Laboratory Tests

Serologic markers – caveats:

 Precore or HBeAg negative mutants:
 Due to mutation in precore (abolishes HBeAg production) or
core promoter region (down-regulates HBeAg production)
 No effect on viral replication (may be enhanced)
 More difficult to treat; greater risk of cirrhosis
 Co-infection with HCV may suppress both HBeAg and
HBsAg
 HBV Viral Genome Organization

HBcAg
Hepatocyte
receptor
bindng site

HBeAg

Protein that HBsAg

transactivates
transcriptional 3200 Base Pair Genome
promotors
HBV DNA Polymerase
 Hepatitis B – Laboratory Tests

Serologic markers – caveats:

 Isolated HBcAb may be due to:
 Remote infection (immune or chronic carrier)
 “Window” period between HBsAg and HBsAb
 Co-infection with HCV
 False positive test result – HBcAb is marker most prone to false
positives
 HBV DNA may help sort this out
 Laboratory Tests for HCV
Serology:
 Detection of anti-HCV antibodies
 Serologic test available since 1990
Molecular:
 HCV RNA detection
 Determination of HCV genotype
 Viral load determination
 Laboratory Tests for HCV
Serology:
 Screening:
 3rd generation EIAs measure antibodies directed against
recombinant peptides NS4, core, NS3, and NS5 proteins
 Sensitivity = 97%
 Detects antibodies within 6 to 8 weeks
 No HCV IgM test available
 Confirmatory/supplementary:
 RIBA, LiPA, Second EIA, HCV RNA
Serologic Pattern of Acute HCV Infection with
Progression to Chronic Infection
anti-
HCV
Symptoms +/-

HCV RNA
Titer

ALT

Normal
0 1 2 3 4 5 6 1 2 3 4
Months Years
Time after Exposure
 Rational Use of HCV Diagnostic Tests

TREATMENT

Diagnosis Prognosis Decision Treatment Response

to treat duration and resistance
Serological Liver assessment
assays histology ALT Genotyping
Qual HCV RNA Liver histology Viral load Qual HCV RNA
Qual HCV RNA Viral load
Deltaviridae, envelope of HDV particles contains the Hepatitis B surface
antigen (HBsAg)
HDV genotypes: I, II, and III;
HDV consists of a single stranded, negative sense, circular RNA virus, with an
envelope made up of HBAg.
Virions are 35-43 nm and are roughly spherical, with no distinct nucleocapsid
structure.
The nucleocapsid is made up of 60 large and small delta antigens. These are the
only proteins encoded by HDV.
HDV relies on host cell machinery for replication, and the viral genome (and
antigenome) serves as ribozymes for self-ligation and cleavage.
Viral replication occurs in the nucleus of primary hepatocytes using a double-
rolling circle mechanism.
New virions can be assembled only in the presence of hepatitis B virus. The
stages of the viral life cycle, including replication, assembly, and transport, depend
on the ratio of small to large delta antigen.
 Hepatitis D Virus - Diagnosis

• Anti-HDV Total (IgG & IgM) available

• Incubation time – similar to Hepatitis B
• High titres of HDV antibodies indicate ongoing
chronic infection
• Available only at National Microbiology Lab in
Winnipeg
 Hepatitis E virus (HEV) is a small, non-enveloped virus
with a single-stranded, positive-sense RNA genome that is
approximately 7.2 kb in size.
 HEV is transmitted mainly via a fecal–oral route
 Hepatitis E Virus - Diagnosis

• Both IgG and IgM antibody tests are available

• Incubation period – 7 to 28 days
• No domestically acquired cases in Canada
• Available only at the National Microbiology Lab
in Winnipeg