Viral Hepatitis Author: Nicolene Naidu Bachelor of Biological Science (Cellular Biology), Bachelor of Medical Science (Medical Microbiology) (Honours) The liver is the largest gland in the human body and responsible for a wide range of functions, a few of these include: Filtering harmful substances from the blood Protein synthesis Storage of vitamins (A, D, K and B12), iron and minerals The production of biochemical s like bile, required for digestion Maintenance of blood glucose levels (storage and conversion of glycogen) Manufacturing, breaking down and regulating hormones Fighting infections (by mobilising the macrophages) Regulates cholesterol production Hepatitis is a medical condition that affects the liver. It is characterised by an inflammation of the liver that is caused most frequently by the hepatitis viruses but it can also be caused by other infections, bacteria, parasites, autoimmune diseases, medications, exposure to toxins and even certain plants. The signs and symptoms for the various types are similar because regardless of the cause, all types of hepatitis affect the liver cells. Hepatitis can be infectious (eg. viral) or non-infectious (eg. inflammation caused by alcohol abuse) and while it can be self limiting, when it lasts for a period fewer than six months it is defined as acute hepatitis and chronic hepatitis when the condition persists for longer than 6 months. While all types of viral hepatitis produce clinically similar illnesses, they can be distinguished by their molecular and antigenic properties. Currently there are 5 distinctive types of viral hepatitis: 1. Hepatitis A is caused by injesting food or water contaminated by the Hepatitis A Virus (HAV). Can also be caused by anal-oral contact during sex. Is transmittedalmost exclusively via the faecal-oral route.this form of hepatitis is not chronic and most people who develop Hepatitis A make a full 1
recovery. Parts of the world with poor sanitation are at greatest risk for outbreaks of hepatitis A. HAV is a non-enveloped 27 nm RNA virus that belongs to the Hepatovirus genus of the picornavirus family. The Hepatitis A virion contains 4 capsid polypeptides and has an incubation period of approximately 4 weeks. Up to a 20% variation can occur in the nucleotide sequence of HAV isolates, but despite this all strains belong to one serotype. While the virus is present in the stools, bile, liver and blood, its replication is limited to the liver. Generally once jaundice becomes apparent viremia, faecal shedding of the virus and infectivity rapidly diminish. Anti-HAV antibodies against the HAV antigen can be detected early in the illness (acute phase) and are predominantly of the IgM class, persisting rarely for longer than 6 months to a year. Anti-HAV antibodies of the IgG class become predominant during convalescence. These antibodies remain detectable indefinitely. Diagnosis of hepatitis A is thus made by demonstrating the presence of anti-hav antibodies of the IgM class during acute illness. Patients with serum anti-hav of the IgG class indicate a previous infection and are immune to re-infection after acute illness.(ie. ant- HAV antibodies of the IgG class renders immunity to patients previously infected with the virus.) Figure 1: The Hepatitis A virus (HAV) (http://latinoamerica.abbottdiagnostics.com/ciencia/pdf/4_d.pdf) 2. Hepatitis B - is a sexually transmitted disease caused by the Hepatitis B Virus (HBV). It is spread by contact with infected semen, blood and other body fluids. Hepatitis B infections become chronic in some patients and liver 2
damage could be serious enough to lead to cancer. It is important to note that donated blood is screened for hepatitis B. HBV is made up of 3 particulate forms: a 42 nm double shelled virion, made up of a spherical surface and core; a 27 nm nucleocapsid core and a 22 nm spherical or filamentous form. Is a DNA virus of the Hepadnavirus family. HBV is the ONLY DNA virus among the hepatitis viruses. They possess partially double-stranded and partially single-stranded genomes. HBV has an efficient strategy of encoding proteins from 4 overlapping genes (S,C, P and X), which allows it to achieve its genomic economy (3200 base pair size). While replication occurs in the liver, they exist in other sites. Their replicative strategy is unique among the DNA viruses and is enabled their own endogenous DNA polymerase. The 22 nm spherical/ filamentous forms are the most numerous of the particulate forms. They express an envelope protein on their surface known as hepatitis B surface antigen (HBsAg). HBsAg is detectable in more than 95 % of patients presenting with hepatitis B. HBsAg is usually the first discernable marker of HBV infection. It can be detected during the entire symptomatic phase of infection and precedes clinical symptoms and serum aminotransferase (ALT) activity by 2 to 6 weeks. Within 2 months of the onset of jaundice, HBsAg levels generally become undetectable. It is rare for this antigen to persist longer than a 6 month period. Anti-HBs is produced in response to HBsAg and becomes detectable in serum once HBsAg disappears. It is a protective antibody and persists in the serum indefinitely following infection. The intact virion (42 nm) contains a 27 nm nucleocapsid core. The nucleocapsid core expresses Hepatitis B core antigen (HBcAg) on its surface. Another important HBV antigen to note is Hepatitis B e antigen (HBeAg). It is a soluble, nonpaticulate nucleocapsid protein that is immunologically distinct from the intact HBcAg but a product of the same gene (C gene). HBeAg is produced when translation is initiated in the pre-core region of the C gene and HBcAg is the produced when translation is initiated in the core region of the C gene. Therefore: Translation in precore = HBeAg Translation in core = HBcAg. HBeAg has a signal peptide that leads to its secretion into circulation by binding to the smooth endoplasmic reticulum. HBcAg does not possess a signal peptide and is therefore not secreted into circulation, instead it 3
assembles into nucleocapsid particles. These bind to and incorporate RNA and ultimately the viral DNA. DNA polymerase is also packaged within the nucleocapsid core. HBcAg particles remain the hepatocyte and are only exported when sequestered in an HBsAg envelop. Thus, HBcAg particles do not circulate in the serum. For these reasons, HBcAg is not detectable in the serum of HBVinfected patients. They are however, readily detectable via immuno-histochemical staining of the hepatocyte. In contrast, anti-hbc the antibody produced in response to HBcAg, can be readily detected in serum and presents usually within 2 weeks after the first appearance of HBsAg. Anti-HBc precedes detectable levels of anti-hbs by weeks to months. Immunoglobulin class of anti-hbc can be used to distinguish between recent and older HBV infections. Anti-HBc of the IgM class predominates during the first 6 months of infection (acute infection) whereas, anti-hbc of the IgG class predominates beyond 6 months. Generally, anti-hbs and anti-hbc persist indefinitely in patients that have recovered from an hepatitis B infection. Figure 2: The Hepatitis B virus (HBV) (http://latinoamerica.abbottdiagnostics.com/ciencia/pdf/4_d.pdf) 3. Hepatitis C - is caused by the Hepatitis C Virus (HCV) and is usually spread via direct contact with the blood of an infected person. It is not known to be transmitted enterically (oral-fecal), through breast milk, semen, or saliva. Unlike hepatitis B, risk of liver cancer is only increased in people with cirrhosis and only 20% of hep C patients get cirrhosis. May develop into chronic hepatitis. 4
HCV is an enveloped RNA virus belonging to the Hepacivirus genus (Family : Flaviviridae). Formerly labelled non-a and non-b hepatitis. Virus particles are between 40 60 nm and tend to circulate in relatively low titre (approximately 10 3 10 7 virions/ml), visualisation difficult. HCV infection does not confer lasting immunity against re-infection. The presence of HCV RNA is the most sensitive indicator of HCV infection. Detection of HCV RNA involves molecular amplification (via Polymerase Chain Reaction or Transcription Mediated Amplification). Anti-HCV is produced in response to HCV infection. However, HCV RNA can be detected long before the appearance of anti-hcv and generally persists for the duration of the infection. In patients with chronic HCV infection, HCV RNA may only be detectable intermittently. Figure 3: The Hepatitis C virus (HCV) (http://latinoamerica.abbottdiagnostics.com/ciencia/pdf/4_d.pdf) 4. Hepatitis D transmitted via contact with infected blood, through unprotected sex and contaminated needles. It is caused by the Hepatitis D Virus (HDV). Only a person who is already infected with Hepatitis B can become infected with Hepatitis D. This is because the hepatitis D virus depends on the synthesis of the hepatitis B surface antigen (HBsAg) to be infectious. HDV cannot infect without the HBsAg coating. May develop into chronic hepatitis. HDV is an enveloped hybrid particle with an HDV core and HBsAg coat. 5
It is a defective RNA virus that co-infects with HBV. HDV requires HBV to aide in its replication and expression. It is only slightly smaller than HBV with a virion particle size of between 35-37 nm. The nucleocapsid core expresses delta antigen. HDV can infect a person in one of two ways. Co-infection occurs when HDV infects a person simultaneously with HBV. Super-infection occurs when HDV infects a person already infected with HBV. Because HDV relies solely on HBV, HDV infection can only last for as long as the HBV infection. Anti-HDV is produced in response to HDV infection. While this antibody is produced primarily in the hepatocyte, it is occasionally detectable in serum. With co-infection, anti-hbc of the IgM class predominates with anti- HDV. In super-infection anti-hbc of the IgG class predominates with anti-hdv. 30 40 days may elapse AFTER the onset of symptoms before anti- HDV is detectable. During HDV replication, HDV antigen can be detected in the liver and HDV RNA in the serum and liver. Figure 4: The Hepatitis D virus (HDV) (http://latinoamerica.abbottdiagnostics.com/ciencia/pdf/4_d.pdf) 5. Hepatitis E caused by the Hepatitis E Virus (HEV) and is transmitted enterically via injestion of contaminated food or water. Acute form of hepatitis with no chronic cases reported. HEV is a non-enveloped, 32 to 34 nm,single strand RNA HAV-like virus belonging to the genus Hepevirus (Family: Hepeviridae). 6
While HEV is an HAV-like virus, there is no genomic or antigenic homology between HEV and HAV. Of its 5 genotypes, 4 are detected in humans. Animal reservoirs like swine help perpetuate HEV. Previously called enterically transmitted non-a, non-b hepatitis. Both IgM anti-hev and IgG anti-hev can be detected very early during the course of infection, but both fall rapidly reaching low levels within a year. During late incubation, the virus is excreted in the stool and has been detected in the bile and liver as well. Serologic testing for HEV infection is currently not available routinely. Figure 5: The Hepatitis E virus (HEV) (http://latinoamerica.abbottdiagnostics.com/ciencia/pdf/4_d.pdf) SIGNS AND SYMPTOMS Patients infected with the hepatitis virus usually present with generalized flu like symptoms in the early or acute stage. These include muscle and joint aches, malaise, fever, nausea, vomiting, diarrhea, loss of appetite, aversion to smoking and headaches. Nausea, weight loss and vomiting are usually associated with alterations in taste and olfaction. Symptoms include dark urine and light faeces (clay- coloured), which is usually noticed between 1-5 days before the onset of clinical jaundice (yellowing of skin and eyes). Other symptoms include abdominal discomfort, weight loss, enlarged spleen, tender and enlarged liver. Some may even exhibit enlarged lymph nodes. The enlarged liver is associated with right upper quadrant pain and discomfort. 7
It s important to note that a substantial proportion of patients presenting with viral hepatitis never become icteric. Table1: Showing clinical and epidemiologic features of viral hepatitis. ( Longo D.L, Fauci A.S. Harrison's Gastroenterology and Hepatology. McGraw-Hill. May 2010. Chapter 37: pages 349-372.) Confusion and coma due to hepatic encephalopathy may occur in patients that develop acute liver failure. Chronic hepatitis (caused by hepatitis B, hepatitis B with hepatitis D, hepatitis C) is less common than acute hepatitis and often exhibits no symptoms until the liver is already scarred. At its worst chronic hepatitis can lead to an accumulation of fluid in the abdominal cavity (ascites), cirrhosis, liver failure and deterioration of brain function among others. While the modes of transmission for the various hepatitis infections overlap, clear distinction between then cannot be made solely on the basis of clinical features. The 8
most accurate form of distinguishing between the different types of viral hepatitis involves specific serologic testing. LABORATORY FEATURES Serum ALT and AST show a variable increase during the initial phase of viral hepatitis, which precedes an increase in bilirubin levels. Serum alkaline phosphatise may be normal or only slightly elevated. The level of these enzymes does not however correlate with the degree of liver damage. Peak levels are usually reached once the patient is clinically icteric and progressively diminish doing recovery. When serum bilirubin levels reach > 43 µmol/l, jaundice usually becomes visible in the skin. Transient lymphopenia and neutropenia may occur, proceded by relative lymphocytosis with atypical lymphocytes common. Prolonged Prothrombin Time (PT) may signify extensive hepatocellular necrosis or severe hepatic synthetic defect. About one 1/3 of patients presenting with viral hepatitis present with elevated IgG and IgM serum levels. Low titres of rheumatoid factor may be found. Auto-antibodies in viral hepatisis can be associated with other systemic and viral diseases. However, virus-specific antibodies (during and after infection with hepatitis infection) have diagnostic significance as they can be used as serologic markers. Hepatitis A - Diagnosis based on detection of IgM anti-hav. - Tests for faecal or serum HAV are not routinely available. Hepatitis B - Diagnosis usually made by detection serum HBsAg. - When HBsAg levels are too low for detection, the presence of IgM anti-hbc may be used to establish HBV infection. - Serum HBsAg concentrations correlate inversely with the degree of liver damage. HBV replication can be used as a marker of liver injury. Serum HBV DNA, like HBeAg is an indicator of HBV replication. - At present, HBV DNA testing has shifted from hybridization assays (which are insensitive) to amplification assays like PCR. 9
- HBeAg is another serologic marker in HBV infection. It is used as an indicator of relative infectivity. This form of testing is usually ordered during follow up of chronic infection. - Anti-HBc for IgM can be useful to distinguish between present and past infection. - In the presence of acute viral hepatitis, anti-hbs is rarely detectable and when it is detected, its presence is of no clinical significance. However, anti-hbs is the only serologic marker to appear after immunization with the hepatitis B vaccine (which consists of HBsAg alone). Hepatitis C Figure 6: Showing the diagnostic profile of Hepatitis B. (http://latinoamerica.abbottdiagnostics.com/ciencia/pdf/4_d.pdf) - Diagnosis can be made by demonstrating the presence of serum anti-hcv. - The most sensitive tests for HCV infection, are the assays used to test for HCV RNA and represent the gold standard in establishing Hepatitis C infection. It can be detected before ALT elevation and before the appearance of anti-hcv. - HCV RNA remains detectable indefinitely using amplification techniques. - The two amplification techniques available are branched chain complementary DNA (bdna) and target amplification (via PCR or TMA). - In bdna an enzyme that is colorimetrically detectable is bound to a complementary DNA probe and used as a detection signal which is amplified. - In target amplification, multiple copies of the viral genome are synthesised via reverse transcription of viral RNA to complementary DNA which is then amplified by repeated cycles of DNA synthesis. - While HCV RNA levels are not a reliable marker of the severity of disease, it can be used as a predictor of relative responsiveness to antiviral therapy. 10
Hepatitis D - Intra-hepatic HDV antigen or anti-hdv seroconversion can be used to demonstrate HDV infection. - Early diagnosis of HDV infection is often limited by the 30-40 day delay in the appearance of anti-hdv antigen. - Determining the class of anti-hbc in a patient that presents with serum HBsAg and anti-hdv, can be used to establish the relationship between HBV and HDV. - HDV RNA is a useful indicator of ongoing replication and relative infectivity. Hepatitis E - Commercially available diagnostic assays are available for the determination of Hepatitis E infection. Liver biopsy is rarely required for acute viral hepatitis. Table 2: Showing a simplified diagnostic approach and interpretation in patients presenting with viral hepatitis. (Longo D.L, Fauci A.S. Harrison's Gastroenterology and Hepatology. McGraw-Hill. May 2010. Chapter 37: pages 349-372.) 11
PROGNOSIS In the case of hepatitis A, almost all previously healthy individuals recover completely. In previously healthy individuals with acute hepatitis, a favourable recovery pathway can be anticipated. Older patients and those presenting with serious underlying medical conditions are at greater risk of exhibiting severe hepatitis. When compared to hepatitis B, hepatitis C presents less severely during the acute phase and fatalities in patients infected with hepatitis C, is rare. HDV super-infection in people infected with chronic hepatitis B, increases the mortality rate significantly. Outbreaks of hepatitis E from waterborne sources in India and Asia, have shown an increase in the fatality rate in pregnant women. PROPHYLAXIS Immunization is generally the prophylaxis of choice and differs for each viral type. Hepatitis A Includes active immunization with killed vaccine immunization and passive immunization with IG (immune globulin) which contain enough anti-hav concentrations to confer protection. If IG is administered before infection or early during the incubation period, it is effective in preventing Hepatitis A. Note that IG lots must be free of HCV RNA (determined via PCR) and subjected to viral inactivation steps. Formalin-inactivated vaccines are also effective in preventing hepatitis A. This vaccine is made from strains of HAV attenuated in tissue culture. Hepatitis B Prior to 1982, passive immune-prophylaxis was used, but while it helped reduce frequency of clinical illness, it was not useful in preventing infection. In 1982, active immunization was introduced. It was prepared from the spherical forms (22 nm) of HBsAg derived from the plasma of healthy carriers. In 1987, a genetically engineered vaccine derived from recombinant yeast was introduced. It consists of non-glycosylated HBsAg particles that are indistinguishable from natural HBsAg. 12
Pre-exposure and post-exposure prophylaxis recombinant vaccines exist. Pre-exposure prophylaxis vaccines are given in settings with frequent exposure (eg. laboratory staff, haemodialysis patients, etc). Post-exposure vaccines are given to individuals previously unvaccinated against HBV. Post-exposure prophylaxis involves a combination HBIG( to achieve an increased titre of circulating anti-hbs) and hepatitis B vaccine (to confer longlasting immunity). Note that pregnant women may also be vaccinated. The hepatistis B vaccine may afford immunity for anything between 5-10 years. Hepatitis C IG is not effective in preventing Hepatitis C. HCV s a difficult target for immune-prophylaxis with vaccine because of its ability to neutralise antibodies by its rapid mutation, viral heterogeneity and genotype. Successful measures used to prevent transfusion associated Hepatitis C include, relying on volunteer blood supply instead of commercial blood donors, screening donor bloods with markers that help identify parts of the population at higher risk of blood-borne infections, excluding donors that come from high risk groups for HIV and the use of serologic and virologic tests to screen for HCV infection. Preventative measures for HCV infection include behaviour modification and limiting exposure to infected persons. Hepatitis D Prevention can be achieved by vaccinating persons susceptible with the hepatitis B vaccine. There is no immune-prophylaxis to prevent HDV super-infection in carriers of the HBsAg. Hepatitis E It is not known whether IG prevents HEV infection. At present, a recombinant vaccine is undergoing clinical testing. 13
References 1. http://latinoamerica.abbottdiagnostics.com/ciencia/pdf/4_d.pdf 2. Longo D.L, Fauci A.S. Harrison's Gastroenterology and Hepatology. McGraw- Hill. May 2010. Chapter 37:pages 349-372. 3. http://wikis.lib.ncsu.edu/index.php/group_11_hepatocytes_(liver_cells) 4. http://en.wikipedia.org/wiki/hepatitis. 5. http://www.ehow.com/about_5549588_difference-between-acute-chronichepatitis.html 14