Hepatitis Viruses

Some studies have found a correlation between tattooing and hepatitis C virus infection in very selected populations, such as prisoners using makeshift tattooing devices.

© Charles Knox/Shutterstock.




“Love is like a virus. It can happen to anybody at any time.”



Maya Angelou, American poet, educator, civil rights activist


This chapter focuses on the group of hepatitis viruses that causes primary hepatitis. The group of viruses is quite diverse, both genetically and taxonomically. The viruses share a liver tissue tropism, causing inflammation and necrosis of the liver.


10.1 The History of Viral Hepatitis


Hepatitis means “inflammation of the liver.” The classic symptom of hepatitis is jaundice, a yellowing of the skin and eyes caused by too much bilirubin in the blood (refer to the clinical features of the virus for further explanation in Section 10.3). Hepatitis is caused by several different viruses and, occasionally, by consumption of alcohol or prescription drugs. Viral hepatitis is one of the most important public health threats worldwide. A notorious case of viral hepatitis in the United States involved the popular American stunt performer Robert Craig “Evel” Knievel, Jr. In 1993, Knievel was diagnosed with a hepatitis C virus infection. He contracted hepatitis C virus from contaminated blood received during one of his 14 reconstructive surgeries used to repair broken bones acquired during a stunt. Knievel received a liver transplant in 1999 and died in 2007 of pulmonary fibrosis at the age of 69. In 2002, the actress from the television series Baywatch and former Playboy model Pamela Anderson claimed she had contracted hepatitis C virus from sharing a tattoo needle with her ex-husband, rock musician Tommy Lee.


Epidemics of jaundice or hepatitis have been recorded throughout history, especially during times of war. For example, outbreaks occurred among Napoleon’s troops during the Egyptian and Russian campaigns that took place in the early 1800s. The crowding and unsanitary conditions of military encampments created an ideal environment for hepatitis A viruses to be transmitted. In 1821, Napoleon Bonaparte died in exile on the island of St. Helena. Theories abound as to the cause of his death, including stomach cancer, arsenic poisoning, inappropriate medical treatment, and hepatitis—an endemic disease of St. Helena.


Approximately 70,000 troops fell ill with hepatitis during the U.S. Civil War. Hepatitis was referred to as “camp jaundice” during the trench warfare battles in the Mediterranean theater in World War I (FIGURE 10-1).


During World War II, at least two significant outbreaks of hepatitis occurred. The first was in 1942, when approximately 330,000 U.S. soldiers were vaccinated against yellow fever virus. Of the vaccinees, 50,000 experienced jaundice and 62 died. It was believed that a virus was present in the serum used to manufacture the yellow fever vaccine. After the lots of yellow fever vaccine administered to the soldiers were destroyed and a human serum–free yellow fever vaccine was manufactured, the cases of jaundice stopped. Years later it was confirmed that the soldiers had serum hepatitis caused by hepatitis B virus.

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FIGURE 10-1 Many soldiers fighting in World War I contracted viral hepatitis during trench warfare, where infectious hepatitis infection was common. Field sanitation during heavy combat was inadequate, and hepatitis A virus was transmitted among the soldiers by a fecal–oral mode of transmission.


The second hepatitis outbreak during the war occurred during the invasion of North Africa and the subsequent invasion of the Italian mainland in the Mediterranean theater of operations (MTO). Even though the antibiotic penicillin was available at this time to treat bacterial infections, and medicine in general had improved, diseases proliferated in combat situations, making treatment of the troops difficult. Malaria and hepatitis epidemics incapacitated 55,000 American assault troops, delaying D-Day. The viral hepatitis outbreak was caused by infectious hepatitis (hepatitis A virus) during the occupation of recaptured land, amid battlefields that were littered with human excrement and corpses. Hepatitis A virus is found in the feces of people infected with hepatitis A virus and is spread by fecal–oral transmission. Unsanitary conditions were common at the frontlines of war. There were no porta-potties (portable restrooms; see VIRUS FILE 10-1) at the battle lines, and troops could not move behind the lines when they needed to go to the bathroom (these were pre-portable-toilet days). Hence, feces got on their boots and clothing and into the bottom of the next shell hole or foxhole they dashed into. Soldiers detected the proximity of the frontlines because of the increasing intensity of odors. Infectious hepatitis was the greatest cause of disabling disease in U.S. and British forces in the MTO.

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FIGURE 10-2 Historical perspective illustrating the separation of hepatitis viruses designated A–E, G that cause primary viral hepatitis. Before they were identified as separate viruses, hepatitis C, E, G, TTV, and SEN-V were designated hepatitis non-A, non-B viruses. Human torque teno virus (TTV), or transfusion transmission virus, was discovered in 1997, and sentinel virus (SEN-V) was identified in 2000. These emerging viruses are associated with hepatitis viruses that are diagnostically separated as non-A–E viruses.


Today, hepatitis C is a major problem among U.S. military veterans. Approximately 8–9% of U.S. Veterans Affairs (VA) Medical Center patients are positive for hepatitis C virus antibodies. Some VA hospitals have had as high as 10–20% of patients with hepatitis C virus antibodies. Hepatitis C virus is transmitted through contact with blood or blood products contaminated with hepatitis C virus. It is not spread through coughing, sneezing, hugging, kissing, breastfeeding (unless bleeding occurs), sharing utensils or glasses, casual contact, or sharing food and water. Multiple factors increased the risk for hepatitis C virus transmission among active duty soldiers during the Vietnam War:1




  • Combat casualties survived multiple blood transfusions in an era prior to hepatitis C virus screening of the blood supply.



  • Some soldiers experimented with injected drugs, such as heroin.



  • Sharing razors or other contaminated instruments resulted in exposure to blood.



  • Many soldiers received tattoos (bleeding may occur during the procedure).



  • Prostitution and promiscuity (more than 15 sexual partners) were common.



  • Healthcare and combat personnel were often exposed to blood/body fluids.



  • Some soldiers received contaminated immunoglobulin for hepatitis A prophylaxis in an era prior to hepatitis C virus screening of the blood supply.


Challenges remain in the treatment of hepatitis C and prevention. Testing and improved treatments will benefit all individuals who suffer from hepatitis C (see Section 10.3).


10.2 Epidemiology of Viral Hepatitis


Hepatitis can be caused by viral infection, trauma, alcohol abuse, or drug-induced toxicity. Viral hepatitis is the most important cause of hepatitis on a global scale. Yellow fever virus, herpes simplex viruses, cytomegalovirus, and Epstein-Barr virus can cause secondary hepatitis, but a range of unrelated human pathogens known as the “hepatitis viruses” cause the vast majority of virally induced primary hepatitis cases. Primary hepatitis occurs immediately after an initial infection by a hepatitis virus. In contrast, secondary hepatitis symptoms follow after another type of infection, disease (e.g., autoimmune disease), treatment (e.g., drug toxicity), or other cause (e.g., alcohol abuse).


Hepatitis Virus Nomenclature


To date, eight hepatitis viruses have been identified. FIGURE 10-2 is a flow diagram used in diagnostics to distinguish the viruses from one another. Originally, each hepatitis virus was named with a letter of the alphabet. Two newly identified members of the group are not named this way: transfusion transmission virus, or torque teno virus (TTV), and sentinel virus (SEN-V). Even though these hepatitis viruses cause similar diseases, they vary widely in their structure and molecular biology (TABLE 10-1).


For years, three main hepatitis viruses were associated with hepatitis: hepatitis A virus (HAV), the cause of infectious hepatitis; hepatitis B virus (HBV), the cause of serum hepatitis; and the hepatitis non-A, non-B viruses. Diagnostic tests were developed to identify HAV and HBV in the 1970s. A new “antigen” found in liver specimens of some patients infected with HBV led to the identification of hepatitis D virus (HDV) in 1977. HDV was only seen in a fraction of patients with HBV, and it was often associated with a more severe course of hepatitis. Hepatitis non-A, non-B viruses were considered the cause of all other unidentified viral causes of hepatitis. In 1989, hepatitis C virus (HCV), the agent that caused non-A, non-B hepatitis, was discovered using modern molecular techniques. Since that time, an additional non-A, non-B hepatitis virus has been discovered and named hepatitis E. Emerging or new viruses causing primary hepatitis are referred to as “non-A–E viruses.” They include hepatitis G virus (HGV), TTV, and SENV. Hepatitis A, B, and C viruses cause over 90% of acute primary viral hepatitis in the United States. TABLE 10-2 compares the various hepatitis viruses by mode of transmission, source or reservoir, whether the course of disease becomes a chronic infection, and how viral infections can be prevented.



1 In 1969, U.S. involvement in the Vietnam conflict was at its peak with more than 500,000 military personnel involved. President Richard Nixon ordered the withdrawal of U.S. forces in 1973.


Table 10-1 Human Hepatitis Viruses: Nomenclature and Characteristics





























































Virus Member Abbreviation Family Enveloped or Naked Genome
Hepatitis A virus HAV Picornaviridae Naked +ssRNA
Hepatitis B virus HBV Hepadnaviridae Enveloped DNA (partially double-stranded)
Hepatitis C virus HCV Flaviviridae Enveloped +ssRNA
Hepatitis D (δ) virus HDV Not assigned* Enveloped –ssRNA
Hepatitis E virus HEV Hepeviridae Naked +ssRNA
Hepatitis G virus HGV Flaviviridae Enveloped +ssRNA
Torque teno virus TTV Circoviridae Naked ssDNA
Sentinel virus SEN-V Circoviridae Naked ssDNA (circular)


*Defective virus, requires HBV for viral assembly.


The geographical distribution of hepatitis viruses A–E is shown in FIGURE 10-3. Hepatitis A commonly occurs in developing parts of the world where sewage disposal and food hygiene are unsatisfactory. HBV is spread through a parenteral mode of transmission (i.e., blood-to-blood transmission), such as through tattooing, body piercing, and injection drug abuse. Transmission can also occur sexually and by fomites (e.g., sharing contaminated razors). High to intermediate prevalence of HBV also occurs in developing countries, Western Europe, Russia, Central America, and the Caribbean. Variants of HBV have been described, and the current hepatitis B vaccine does not protect immunized individuals against the variant HBVs.


Table 10-2 Comparison of Human Hepatitis Virus Infections





























































Virus Source of Virus Mode of Transmission Chronic Infection Prevention
Hepatitis A virus Feces Fecal–oral No Pre-/postexposure immunization
Hepatitis B virus Blood Blood-to-blood contact
Sexual contact
Sweating?*
Yes Pre-/postexposure immunization
Hepatitis C virus Blood Blood-to-blood contact
Sexual contact
Yes Blood donor screening
Risk behavior modification
Hepatitis D virus Blood Blood-to-blood contact Yes Pre-/postexposure modification
Risk behavior modification
Hepatitis E virus Feces Fecal–oral No
Hepatitis G virus Blood Blood-to-blood contact ? Blood donor screening
Torque teno virus Blood Blood-to-blood contact ? ? Blood donor screening
Sentinel virus Blood Posttransfusion hepatitis ? ? Blood donor screening


*New study by Bereket-Yucel, S. 2007. “Risk of hepatitis B infections in Olympic wrestling.” Br J Sports Med. 41:305–310.

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FIGURE 10-3 Geographical prevalence of human hepatitis viruses A–E. (a) Countries or areas at risk for contracting hepatitis A virus. (b) Global prevalence of hepatitis B virus infection among adults. (c) Global prevalence of hepatitis C.

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FIGURE 10-3 (d) Global prevalence of hepatitis D (CDC data). (e) Global prevalence of hepatitis E (CDC data). (Continued)


HCV is spread primarily by direct contact with human blood. The prevalence of hepatitis C is higher among blood donors in the Eastern Mediterranean, Western Pacific, and Southeast Asia compared to some countries in North America and Europe. Data from African countries are limited due to poor reporting.


Hepatitis D virus requires the presence of hepatitis B virus in order to replicate. In general, then, the global pattern of hepatitis D corresponds to the prevalence of hepatitis B virus infection; for example, in countries where chronic hepatitis B cases are low, there is a low prevalence of hepatitis D virus. Hepatitis E virus is spread by the fecal–oral mode of transmission, often through contaminated water. It is endemic and sometimes epidemic in the developing world.


Screening the Blood Supply for Viral Hepatitis Agents


The American Red Cross has been screening for HBsAg since 1971. In 1987, a test for anti-HBcAg was added to the screening protocol. From 1986 to 2003, the U.S. blood supply was also screened for elevated levels of ALT. In 1990, testing began for anti-HCV IgM and IgG, and in 1999 a nucleic acid test for HCV genomic RNA was added to blood donation screening in order to reduce the risk of disease transmission. Blood is not entirely risk-free, but it is very safe. The U.S. blood supply in particular is among the safest in the world.


10.3 Clinical Features of Viruses That Cause Primary Hepatitis


Hepatitis A


Most hepatitis A outbreaks are sporadic and associated with contaminated food or water supplies. Shellfish may become contaminated with sewage. This becomes a problem because they are able to retain and concentrate viruses from the water. The major mode of transmission is fecal– oral. The average incubation period for hepatitis A virus (HAV) is 30 days. Adults experience signs and symptoms more often than children. Pregnant women are at increased risk for serious infection. Symptoms include:




  • Fatigue



  • Abdominal pain



  • Loss of appetite



  • Nausea and vomiting



  • Dark urine



  • Jaundice (in 70–80% of individuals older than 14 years of age; less likely in children)


Signs of jaundice are a yellow color in the skin, mucous membranes, or eyes (FIGURE 10-4). Jaundice occurs when the liver is not functioning properly. The yellow pigment is from bilirubin, a by-product of old red blood cells. You may have noticed that when you have a bruise sometimes the skin goes through a series of colors as it heals. When you saw yellow you were seeing bilirubin.


Hepatitis B


Hepatitis B is relatively rare in developed countries. In endemic areas, a major mode of spread occurs from mother to infant when the mother is a carrier. The blood from the infected mother enters the uninfected fetus. Other high-risk groups for hepatitis B are injection drug users, hemodialysis patients, individuals who have had multiple sex partners, institutionalized patients, and healthcare workers.

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FIGURE 10-4 This person has jaundice of the conjunctiva and facial skin that manifested after HAV infection.


The average incubation period for hepatitis B virus (HBV) is 80 days. About 30% of individuals have no signs or symptoms. When symptoms do occur, they are similar to those of hepatitis A, with the addition of joint pain. Chronic HBV infection occurs in 5–10% of individuals who have been infected by HBV after the age of 5. Chronic HBV infection can lead to liver cirrhosis and hepatocellular (liver) cancer (HCC). Genomic HBV DNA sequences can be detected in the chromosomes of hepatocellular tumors associated with HBV infection. Integration of HBV DNA occurs where there are breaks in the cellular DNA of liver cells. In humans, chronic HBV infection causes an ongoing inflammatory response, resulting in oxidative damage to the DNA, such as double-stranded breaks in the DNA of liver cells. Chronic liver disease is the cause of death in 15–25% of chronically HBV-infected individuals.


Hepatitis C


Prior to 1989, a number of cases of hepatitis after blood transfusions were found not to be caused by hepatitis A or B viruses. Investigators found it difficult to identify the cause of the non-A, non-B hepatitis. Individuals with an acute infection displayed the following symptoms:




  • Fever



  • Fatigue



  • Dark urine



  • Clay-colored stool



  • Abdominal pain



  • Poor appetite



  • Nausea



  • Vomiting



  • Joint pain



  • Jaundice


In 1989, a direct molecular biology approach by Michael Houghton and colleagues provided the tools to identify and characterize the cause of the non-A, non-B hepatitis. Their approach was to clone the non-A, non-B viral genome present in plasma from patients suffering from hepatitis. The new non-A, non-B causative agent of hepatitis was named hepatitis C virus (HCV). Amazingly, HCV was identified without the ability to grow, visualize, or detect the viral agent. Substantial progress has been made in the last decade of research to isolate, grow, and visualize HCV, which is now a major public health problem.


Individuals with chronic HCV infection are asymptomatic, but they may have chronic liver disease that can range from mild to severe, including cirrhosis and liver cancer. HCV infection is a “silent epidemic” because victims show few or no signs of disease for years, or even decades. Early assessments of the total number of infected individuals were underestimated. About 2.7 million people in the United States have chronic HCV infection. HCV infection is most prevalent among those born between 1945 and 1965, the majority of whom were likely infected during the 1970s and 1980s when rates were highest. Before 1992, when blood screening for HCV became available, blood transfusion was a leading means of HCV transmission. Today, CDC experts recommend HCV testing for the following high-risk individuals:

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FIGURE 10-5 HCV prevalence by selected groups in the United States.




  • People born between 1945 and 1965.



  • People who have injected illegal drugs, including those who injected only once many years ago.



  • Recipients of clotting factor prepared before 1987.



  • Recipients of blood transfusions or solid organ transplants before July 1992.



  • Patients who received long-term hemodialysis treatment.



  • Healthcare workers with known HCV exposures after contact with needlesticks involving HCV-positive blood.



  • Recipients of blood or organs from a donor who later tested positive for HCV.



  • People who have tested positive for human immunodeficiency virus (HIV).



  • Patients with signs or symptoms of liver disease (e.g., elevated liver enzymes).



  • Children born to HCV-positive mothers (to avoid detecting material HCV antibodies, the children should not be tested before 18 months of age).


Unlike HBV infection, HCV infection is common in the developed world. About 4 million people in the United States (1.8% of the population) are infected with HCV. HCV is spread almost exclusively through blood contact (FIGURE 10-5). Sexual transmission is rare. Individuals who received blood transfusions or blood products, had long-term kidney dialysis, or had a solid organ transplant before 1992, an era when screening donated blood for HCV was not available, are at risk for having contracted HCV. About 80% of infected individuals have no signs or symptoms. If present, signs and symptoms are similar to other hepatitis virus infections. Between 55% and 85% of infected persons experience a chronic infection, resulting in liver disease. A chronic infection is one of long duration that is characterized by scarring of the liver, preventing it from functioning normally. It is usually insidious, progressing slowly without any signs or symptoms for several decades. Chronic HCV infection can lead to cirrhosis of the liver and HCC in 5–20% of persons. The average incubation period is 6–7 weeks.


Hepatitis D


Hepatitis D virus (HDV) is a defective virus that requires coinfection with HBV to assemble virus particles but not for genomic replication. HDV infections occur during two types of HBV infections: coinfection and superinfection. A coinfection refers to a simultaneous infection in the same host cells by separate pathogens, as by HBV and HDV. A superinfection occurs when a chronic HBV carrier is infected with HDV. Symptoms of hepatitis D are indistinguishable from those of hepatitis B. Less than 5% of cases of acute coinfection with HBD and HDV result in a chronic HDV infection. Sometimes HDV superinfection leads to the clearance of hepatitis b surface antigen (HbsAg), and HDV replication ceases, allowing the acute liver disease to resolve. In people in whom superinfection with HDV and HBV persists, liver disease is severe, and there is an increased risk of fulminant liver failure or progression to cirrhosis. The highest risk factor for HDV infection in the Eastern world is injection drug use: 39–90% of addicts test positive for HDV.


Hepatitis E


Hepatitis E is endemic in the developing countries of Asia and Africa and rare in industrialized nations. Sporadic cases in the developed world almost always occur in travelers returning from endemic areas. The mode of transmission and symptoms of hepatitis E virus (HEV) are similar to HAV (i.e., fecal–oral transmission). Person-to-person transmission is rare. The average incubation period for HEV infection is 40 days. HEV epidemics are often associated with water supplies that are contaminated with fecal matter. Prevention strategies include the avoidance of drinking water and beverages with ice of unknown purity, uncooked shellfish, and uncooked vegetables that have not been peeled or prepared by the traveler.


Infection with HEV causes a more severe illness than HAV. It causes mortality in 1–3% of persons and in 15–25% of HEV-infected pregnant women. Even though hepatitis E is rarely diagnosed in industrialized nations, anti-HEV antibodies have been found in a significant proportion of healthy individuals (up to 28% in some areas). Sporadic cases have been reported in the United States, northern England, Italy, and France. Considerable evidence suggests that HEV may be a zoonotic pathogen (transmitted from animals to people). Human HEV, for which isolates are genetically similar to pig HEV strains, has been isolated from pigs in the United States. It has been shown that pig HEV can cross the species barrier and infect nonhuman primates. Human anti-HEV antibodies cross-react and bind to pig anti-HEV capsid proteins.


Hepatitis “Non-A–E” Viruses


As described earlier, hepatitis A–E viruses were not determined to be the causative agent of all cases of primary hepatitis. The term “hepatitis non-A–E viruses” was used to describe new and emerging viruses that caused primary hepatitis.


Hepatitis G

Hepatitis G virus (HGV) was first cloned from the plasma of a surgeon in 1996. HGV, which is transmitted parenter-ally, is a flavivirus and is a distant relative of HCV. HGV prevalence is global. In the United States, 1.7% of volunteer blood donors test positive for antibodies against HGV. The prevalence of HGV is higher in patients infected with HCV, patients infected with HIV, patients who have had multiple transfusions or organ transplants, and patients undergoing hemodialysis. HGV RNA was detected in patients with non-A–E acute hepatitis, in patients with cirrhosis of unknown origin, in patients with chronic hepatitis, and in patients with HCC (liver cancer). The clinical significance of HGV is not well understood.


Transfusion Transmission Virus, or Torque Teno Virus (TTV)

In 1997, TTV was found using representational difference analysis (RdA) in a Japanese patient who had posttransfusion hepatitis of unknown etiology (hepatitis non-A–E). RDA is a technique used to compare DNA from two different sources, in this case comparing the DNA of patient serum before and after the patient experienced elevated levels of alanine aminotransferase (ALT). Elevated ALT levels are an indicator of liver inflammation that can be the result of viral hepatitis (see Section 10.4). The DNA of each source was amplified using polymerase chain reaction (PCR) and allowed to hybridize. The dsDNA formed was subtracted (removed) from the pool of DNA. The unhybridized DNA was sequenced. The extraneous viral DNA was amplified by PCR and sequenced. It was reported to belong to a new candidate hepatitis virus, named after the initials of the patient, “T. T.” In 2005, the International Committee on Taxonomy of Viruses (ICTV) officially named TTV as torque teno virus (TTV). To date, no conclusive evidence suggests that TTV causes human disease.


Sentinel Virus (SEN-V)

About 10% of transfusion-associated hepatitis and 20% of community-acquired cases of hepatitis are of unknown origin, suggesting the existence of additional causative agents. In 2000, a new virus, called sentinel virus (SEN-V), was identified as a hepatitis non-A–E viral agent. It was thought to be associated with post-transfusion hepatitis. Little is known about the clinical significance of SEN-V.


10.4 Laboratory Diagnosis of Viral Hepatitis Infections


Diagnosis of viral hepatitis is based on symptoms and physical findings as well as blood tests for elevated liver enzymes, antibodies against a virus, and the detection of viral proteins or genomes. Symptoms of acute hepatitis include the following:




  • Fatigue



  • Abdominal pain



  • Loss of appetite



  • Nausea and vomiting



  • Darkening of urine, followed by jaundice or hepatitis


A type of serology testing, enzyme-linked immunosor-bent assay (ElisA), is used to detect antibodies directed against hepatitis viruses or antigens of the hepatitis viruses. Liver enzyme blood tests and molecular techniques to detect viral genomes (e.g., RT-PCR or PCR) also are used to confirm diagnosis of acute viral hepatitis. Blood samples are tested for the presence of elevated levels of two liver enzymes: aspartate aminotransferase (AsT) and alanine aminotransferase (AlT). AST and ALT are normally found in the liver but spill into the bloodstream if the liver is damaged, increasing the enzyme levels in the bloodstream. Nucleic acid tests to detect viral genomes are only available in specialized laboratories. Patients with chronic hepatitis are harder to diagnose because they do not have nausea or jaundice until liver damage is quite advanced.

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FIGURE 10-6 (a) Course of disease and immune response toward HAV infection. Symptoms begin 1–2 weeks after infection. “ALT” represents levels of alanine aminotransferase activity detected in blood. ALT levels peak at 4 weeks postinfection. The IgM antibody response against HAV peaks 4 weeks after infection. HAV is shed in feces through weeks 1–5 after infection. (b) Course of HEV infection is similar to HAV infection except that symptoms are later and continue for a shorter duration than in HAV infections.


HAV infections are reliably diagnosed by the presence of anti-HAV IgM. HEV infections are clinically indistinguishable from HAV infections. Diagnosis of HEV infections is made by serology tests that screen for anti-HEV IgM or by RT-PCR. Tests for HEV are not widely available. HAV and HEV do not persist in the liver, and there is no evidence of progression to chronic liver damage. The courses of HAV and HEV infections are shown in FIGURE 10-6.


HBV infections are diagnosed by the level of IgM antibodies produced against hepatitis B surface antigen (anti-HBsAg) and hepatitis B core-antigen (anti-HBcAg). Antibody levels will vary depending upon whether the patient has an acute or chronic hepatitis infection (TABLE 10-3). FIGURE 10-7A is a graphical representation of the clinical course of acute and chronic hepatitis B over time. A patient is a chronic hepatitis B virus carrier when HBsAg is present for longer than 6 months, but the diagnosis is often suspected much earlier. Hepatitis B e antigen (HBeAg) accumulates during a chronic infection. HBeAg is an HBV protein translated from the same reading frame as HBcAg, but HBeAg is a soluble antigen secreted from infected cells and is found accumulated in serum (see Section 10.7). HBeAg is not a part of the structure of HBV. It serves as an active marker for chronic HBV infection. It is not used in diagnostic testing because it shares homology to HBcAg. Instead, serology testing is used to detect HBcAg and anti-HBcAg. The risk for chronic HBV infection is related to age at the time of infection. About 90% of infants will become chronically infected with HBV as compared to 2–6% of adults. The best way to prevent HBV infection is by getting vaccinated (see Section 10.8).


Table 10-3 Diagnostic Testing for Hepatitis A, B, and C Virus Infections




























Virus Type of Viral Infection Diagnostic Tests
Hepatitis A virus Acute IgM anti-HAV positive
Hepatitis B virus Acute HBcAg positive
Anti-HBcAg positive
IgM anti-HBcAg positive
Anti-HBsAg negative
Hepatitis B virus Chronic HBsAg positive
Anti-HBcAg positive
IgM Anti-HBcAg negative
Anti-HBsAg negative
Hepatitis C virus Acute High serum ALT levels (73 normal limit)
Nucleic acid testing for HCV-RNA
Hepatitis C virus Chronic Anti-HCV positive
HCV RNA ≥ 6 months apart
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FIGURE 10-7 (a) Course of acute and chronic diseases and immune response toward HBV infection. HBeAg is an indicator of a chronic infection. (b) Events in HCV chronic infection. HCV RNA can be detected in acute and chronic HCV infections. ALT levels fluctuate during a chronic HCV infection. (c) Natural history and relationship of acute and chronic HBV and HCV infections. HCV becomes chronic in 75–85% of cases.


Acute HCV infection cannot be reliably diagnosed by serology tests because it can take as long as 3 months for patients to become positive. Acute hepatitis C is diagnosed by blood tests detecting serum ALT levels that are seven times higher than the normal upper limit of liver enzyme levels. Presence of HCV RNA genome in serum samples as determined by nucleic acid testing also is an indicator of HCV infection. The HCV RNA genomes can be detected within the first month of infection and throughout a chronic HCV infection. Early diagnosis is beneficial because it is believed that early treatment can prevent chronic infection with HCV. IgM antibodies disappear after 36 weeks of HCV infection (FIGURE 10-7B).


About 15–25% of individuals infected with HCV can clear the virus from their bodies without treatment and do not develop chronic infection. The reasons for why this occurs are not well known. Chronic HCV infection can occur in 75–85% of untreated patients (FIGURE 10-7C). Diagnosis is based on anti-HCV antibodies and HCV RNA levels (Table 10-3). Individuals who develop chronic hepatitis C mount an immune response toward the virus, but replication of HCV during infection can result in changes whereby the viruses can evade the immune response toward the infection. This may explain how HCV establishes and maintains a chronic infection. Of every 100 individuals infected with HCV, approximately:




  • 75–85 will develop chronic HCV infection.



  • 60–70 will develop chronic liver disease.



  • 5–20 will develop cirrhosis of the liver over a period of 20–30 years.



  • 1 to 5 will die from the consequences of chronic infection (liver cancer or cirrhosis).


Pathogenesis: Chronic Hepatitis


Chronic hepatitis occurs when the inflammation of the liver is active, persists for more than 6 months, and is detectable by increased ALT levels in serum. Hepatitis B, C, D, and G viruses can cause chronic hepatitis infections. A patient who is viremic and has an abnormal ALT level should have a liver biopsy, which is the only way to assess the degree of inflammation of the liver and the stage of liver disease. Chronic liver damage may result in cirrhosis, which is characterized by the formation of fibrous tissues, nodules, and scarring that interfere with liver cell function and blood circulation (FIGURE 10-8). A late complication of cirrhosis is liver cancer (HCC; see Section 10.3), which takes many years to develop.

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FIGURE 10-8 (a) A healthy human liver. (b) A human liver diseased by viral hepatitis and cirrhosis.


10.5 Hepatitis Virus Replication Cycles


The hepatitis viruses belong to five different families, designated by the suffix –viridae (Table 10-1). Families are groups of viruses that share common characteristics. Although there are five families listed in the right-hand column of Table 10-1, one of the viruses is unassigned. Five of the eight hepatitis viruses (hepatitis viruses A–E) discussed in this chapter are well characterized.


Virus Structure


Hepatitis viruses A–E have a spherical shape and icosahedral capsid symmetry 28–50 nm in diameter. Virus particles are either enveloped or nonenveloped. Transmission electron micrographs of HBV and HBC are shown in FIGURE 10-9. Hepatitis B, C, and D viruses are enveloped and relatively sensitive to many physical and chemical agents, whereas HAV is not enveloped and can remain infectious on inanimate surfaces in the environment for a month. Hepatitis A viruses can also retain their infectivity in the hostile environment of the gastrointestinal tract because they are acid and bile resistant.


Virus Replication


Hepatitis viruses enter the bloodstream and are carried to the liver, where they infect the hepatocytes of the liver.

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FIGURE 10-9 Transmission electron micrographs of hepatitis viruses. (a) Hepatitis B virus. (b) Hepatitis C virus.


As the viruses spread and replicate in the liver, hepatocytes may become damaged. Chronic liver damage results in cirrhosis, impairing liver function (FIGURE 10-10). The liver has many functions, such as playing a role in metabolism and the filtering of toxins and waste products in the blood. The liver produces bile, which aids in digestion.


Hepatitis A Virus

HAV is a very stable, naked picornavirus that contains a +ssRNA genome that is 7.5 kilobases (kb) in length. The viral genome contains a small protein, VPg, that is covalently linked to the 5′ end of the genome and a poly(A) tail at the 3′ terminus of the genome. The 5′ terminus also contains a nontranslated region that contains hairpins and pseudoknots. The pseudoknots may function in the recognition of the genome by the viral RNA replicase. The hairpin secondary structures act as an internal ribosome entry site (iREs) that is necessary for cap-independent translation of the viral mRNA (FIGURE 10-11A).


Hepatitis A viruses bind receptors located on the surface of hepatocytes. A candidate cellular receptor that interacts with the virus particle has been identified. It is a mucin-like glycoprotein referred to as the human hepatitis A virus cellular receptor 1 (huHAVcr-1). After entry, the HAV undergoes penetration and uncoating with the release of viral genome into the cytoplasm of the host cell. The +ssRNA viral genome acts directly as an mRNA that is translated by the host cell ribosomal machinery into a large polyprotein. The polyprotein is processed by a viral 3C protease into structural and non-structural proteins of HAV. A viral RNA-dependent RNA polymerase encoded by 3D synthesizes –ssRNA intermediates that are used to create new progeny genomic +ssRNAs that will be packaged into virions. Newly assembled virus particles are transported to the surface of hepatocytes and released (FIGURE 10-11B).


Hepatitis B Virus

American physician and biochemist Baruch (“Barry”) Blumberg (1925–2011) and colleagues began characterizing and comparing serum proteins from hemophiliacs with that from nonhemophiliacs in the 1950s. They collected blood samples from native populations in remote parts of the world with the goal of learning about the genetics of infectious disease susceptibility. They reasoned that hemophiliacs who received multiple blood transfusions would be exposed to serum proteins they had not inherited genetically but instead may have “inherited” from their donors. As a result, the hemophiliacs would produce antibodies against the donor proteins. In 1963, Blumberg’s team discovered an antibody present in the serum of a New York hemophiliac that reacted with an antigen present in the blood of an Australian aborigine infected with hepatitis. Later, they determined that the new unknown Australia antigen was HBsAg. Further experiments by British virologist David M. S. Dane led to the discovery of the complete hepatitis B virus, known as the dane particle, which is now recognized as the agent responsible for serum hepatitis. Blumberg is a corecipient of the 1976 Nobel Prize in Physiology or Medicine (with Daniel Carleton Gajdusek) for this hepatitis B research.

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FIGURE 10-10 Hepatitis A and E viruses are ingested and gain entry into liver cells by way of the bloodstream through the hepatic artery or the portal vein. The portal vein carries blood containing digested food from the small intestine to the liver, thus facilitating the entry of HAV into the liver. Bloodborne hepatitis viruses, such as HCV, enter the liver through the hepatic artery.


Through research and clinical observations, it is known that three types of virus-associated particles are present in the blood of individuals suffering from hepatitis B. The most abundant hepatitis B–like particle present in highly viremic carriers of HBV is a spherical particle approximately 17–25 nm in diameter. The viruses appear as small spheres that contain HBsAg proteins (FIGURE 10-12). Less numerous are filaments approximately 17–20 nm in diameter and up to 200 nm in length (Figure 10-12). The filamentous particles contain small, medium, and large HBsAg proteins (labeled as SHBs, MHBs, and LHBs, respectively, in Figure 10-12), which are not infectious. The large amounts of free HBsAg present in the serum of hepatitis patients correlates with viremia, and in many cases indicate chronic infection.


Dane particles are infectious. The enveloped Dane particles measure 42 nm in diameter and contain small, medium, and large HBsAg proteins. In addition to the HBsAg proteins, the HBV has a DNA-based core that is 27 nm in diameter (Figure 10-12). The core that surrounds the DNA genome contains two proteins: HBcAg and HBeAg. Most of the HBeAg is secreted and is not incorporated into the particle. The viral DNA polymerase (reverse transcriptase), protein kinase C, and heat shock protein 90 are associated with the HBV RNA genome within the core of the particle. Hepatitis B viruses are amazingly stable in the environment. They are resistant to organic solvents, heat, and pH changes. HBV present in blood can withstand drying on a surface for at least a week.


HBV is a hepadnavirus (hepa from “hepatotropic” and dna from its “DNA” genome). The circular genome of HBV is composed of partially double-stranded DNA (dsDNA). The completed full-length strand is approximately 3.2 kb in length; the shorter strand is approximately 1.7 kb in length (FIGURE 10-13). One of the molecular biology hallmarks of hepadnaviruses is that they use a replication strategy common to retroviruses: they replicate through an RNA intermediate, sometimes referred to as a pregenomic RNA, by reverse transcription.

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FIGURE 10-11 (a) Secondary structure of the 5′ noncoding region of hepatitis A virus genome. The pseudoknots are involved in the recognition of the viral RNA-dependent RNA polymerase or replicase, whereas the internal ribosome entry site allows for cap-independent translation of the viral mRNA transcripts into a polyprotein product. The function of the polypyrimidine tract is unknown. (b) Organization of the hepatitis A virus genome. Genes are listed within the genome, and the gene product names are listed above the genes. The genome is translated into a large polyprotein that is processed by the viral 3C protease into three distinct precursors that are processed by proteases to yield smaller precursors and mature viral proteins, etc.

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FIGURE 10-12 Three particles are observed during HBV infection: spheres, filaments, and Dane particles. Only the Dane particles are infectious. Structures are shown for each type of particle. Classic HBsAg, which contains the S domain only, is also called the S protein. Two other proteins share the C-terminal S domain, but differ by length and structure of their N-terminal (pre-S) extensions. The large (L) protein contains the pre-S1, the pre-S2, and the S regions, whereas the medium (M) protein contains the pre-S2 and the S regions only. HBsAg is the most abundant of the S-related antigens. The L and M proteins are expressed at levels of about 5–15% and 1–2%, respectively, compared with S protein.


The host cell receptor used for the attachment and entry of HBV Dane particle into hepatocytes is unknown. A few potential receptors have been shown to interact with HBsAg, such as the transferrin receptor, human liver endonexin, and the asialoglycoprotein receptor molecule, but there is no convincing evidence of their connection to infectivity. Hepatitis B viruses replicate poorly in cell cultures, making it difficult to fully understand the early steps of viral infection. After fusion and entry into the host cell, the viral cores are released into the cytoplasm where the genome is uncoated and enters the nucleus through the nuclear pores.


In the nucleus, host enzymes repair the HBV relaxed circular DNA (rcDNA) by ligating the ends of the HBV genome. HBV DNA replication is completed and gaps in both DNA strands are repaired, resulting in a completed, covalently closed circular DNA (cccDNA) that is a plasmid-like dsDNA molecule. Subsequently, the HBV cccDNA is supercoiled and organized into a chromatin-like structure called an episome, or mini-chromosome. The mini-chromosome replicates independently of the host chromosome in contrast to being integrated into the cellular DNA, as is the case with retroviruses. This is because hepatitis B viruses do not possess integrase activity.


The mini-chromosome acts as a template for two classes of viral mRNAs: viral pregenomic RNAs and genomic RNAs that are transcribed by the host’s RNA polymerase II. The transcription factors identified in the HBV genome recognize at least four promoters, two enhancers, and several binding sites. Noncoding regions are not present on the genome. HBV DNA has a very compact coding organization that codes for four partially overlapping open reading frames (ORFs). The four ORFs are translated into seven known proteins (Figure 10-13). TABLE 10-4 lists the HBV ORFs and their gene products.


Following the replication of full-length HBV pregenomic +ssRNAs, it is complexed with the viral polymerase and protein kinase C protein into core particles. Host-cell heat shock proteins associate with the HBV reverse transcriptase, allowing it to fold into an active conformation. The active viral reverse transcriptase converts this pregenomic RNA into DNA inside of the Dane particles. Unlike retroviruses, the HBV reverse transcriptase activity occurs by protein priming as opposed to RNA priming. Precise replication can only occur in the specialized environment of the intact nucleocapsids.

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FIGURE 10-13 Genomic organization of the hepatitis B virus. The HBV DNA genome is transcribed by the cellular RNA polymerase II. It is under the control of four promoters: pre-S1, pre-S2, pre-C, and X.


Table 10-4 Hepatitis B Virus ORFs and Their Gene Products



















ORF Gene Products
1 (S) Large, middle, and small hepatitis B surface (envelope) antigens (HBsAg)
2 (C) Hepatitis B secreted precore antigen (HBeAg) and hepatitis B core (capsid) antigen (HBcAg)
3 (P) Viral polymerase (DNA-dependent DNA polymerase, reverse transcriptase, with RNase H activity)
4 (X) Transcriptional trans-activating protein kinase

The nucleocapsid cores reach the endoplasmic reticulum (ER), where they associate with the viral surface glyco-proteins and bud into the lumen of the ER and/or the Golgi apparatus out of the hepatocyte host cell (FIGURE 10-14). Empty envelopes containing viral surface proteins embedded in the host cell’s lipid bilayer are continually being shed along with mature, infectious Dane particles.

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FIGURE 10-14 Replication cycle of hepatitis B virus. The HBsAg of the HBV Dane particle attaches to an unknown receptor on the surface of a hepatocyte and enters the cell by endocytosis. The relaxed circular DNA (rcDNA) is transported through the nuclear pores into the nucleus and repaired by cellular enzymes to form covalently closed circular DNA (cccDNA). The HBV cccDNA is supercoiled and organized into a chromatin-like structure called an episome, or mini-chromosome. The host cell’s RNA polymerase II transcribes the HBV mini-chromosome into all of the HBV mRNAs and the pregenomic RNA of HBV. The pregenomic RNA is reverse-transcribed by the HBV reverse transcriptase into new rcDNA inside of the nucleocapsid. HBeAg and HBxAg synthesis are omitted for the sake of simplicity. HBsAg proteins are inserted into the host’s endoplasmic reticulum membrane. The nucleocapsids are enveloped as HBV buds as infectious virions during release.


Coinfection of Hepatitis B Virus and Hepatitis D Virus

As described earlier, HDV is a defective virus that requires the presence of hepatitis B “helper” virus for assembly. HDV is frequently associated with severe acute or chronic hepatitis. HDV infection occurs as a coinfection at the same time as an HBV infection or as a superinfection in individuals who are carriers of HBV. In the latter scenario, a superinfection is defined as an HDV infection following a previous infection by HBV. HDV probably makes use of the same host cellular receptor as HBV in order to attach to and enter hepatocytes.


HDV is a spherical enveloped particle that is approximately 36 nm in diameter. The envelope contains small, medium, and large HBsAgs. The internal nucleocapsid structure of HDV contains the –ssRNA circular genome and about 70 copies of delta antigen. HDV can only form particles in a host cell coinfected with HBV because HBV provides the HBsAg that is required for reinfection into another host cell. Once inside the host cell, HDV can replicate in the absence of HBV.


The genome is 1.7 kb in length, which is similar to plant viral satellites and viroids. It codes for two different types of the same protein (i.e., the delta antigen) that are 195 or 214 amino acids in length. About 70 molecules of delta antigen surround and stabilize the viral RNA genome. The large delta antigen contains four different functional domains: an RNA binding domain, a nuclear localization signal, a virus assembly signal, and a coiled-coil sequence that is responsible for oligomerization of the protein. The delta antigen can form either a homo-or heterodimer. The small form of the delta antigen is required for replication of the RNA genome, whereas the large form is required for the suppression of HDV RNA replication and the assembly and export of virions. The small delta antigen is missing the HDV assembly signal (FIGURE 10-15A).

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FIGURE 10-15 (a) Hepatitis D virus functional domains of the small and large forms of the delta antigen. RBD = RNA-binding domain; CCS = coiled-coil sequence; NLS = nuclear localization sequence; VAS = virus assembly signal. (b) Structure of hepatitis D virus RNAs.


The HDV genome has a high degree of intramolecular base pairing, giving it a double-stranded property similar to that found in plant viroids under natural conditions. When the envelope of the virus is removed inside of an infected cell, the viral genome is targeted to the nucleus. Unlike most RNA viruses, though, it does not encode its own RNA-dependent RNA polymerase for genome replication. Instead, HDV is dependent on the host’s RNA polymerase II and other host factors to replicate its genome. The HDV genomic RNA is a ribozyme that cleaves itself, and ribozyme activity is necessary for HDV replication. HDV genome replication is believed to occur via a rolling circle model similar to that of viroids. FIGURE 10-15B shows the structure of HDV RNAs; a model for the HDV replication cycle is shown in FIGURE 10-16. The HDV RNA –sense (negative-sense) genome acts as a template for synthesis of antigenomic RNA (+sense, or positive sense) and the viral mRNAs. The antigenomic RNA multimers are self-cleaved (step 2 in Figure 10-16) and recircularize, serving as a template for multimers of genomic RNAs, which are self-cleaved and circularized (step 4 in Figure 10-16). The single mRNA transcribed from the viral genome exits the nucleus to produce the small and large forms of the delta antigen (step 7 in Figure 10-16). The ribonucleoprotein complexes (RNPs) assemble and are exported to the ER located in the cytoplasm of the cell, where they associate with the HBsAgs (steps 8 and 9 in Figure 10-16). The HBsAgs are required for the packaging of the hepatitis D virions. Mature hepatitis D viruses are produced and secreted outside of the cell.


Hepatitis C Virus

Hepatitis C virus (HCV) is an icosahedral-shaped, enveloped, +ssRNA flavivirus that causes hepatitis non-A, non-B. Little is known about the ultrastructure of hepatitis C virus because it has been extremely difficult to visualize the virus particles directly from infected serum and tissues. The hypothetical structure of HCV is shown in FIGURE 10-17. It is the only flavivirus that is not transmitted by arthropod vectors (e.g., mosquitoes). Yellow fever virus is the prototype of viruses in the Flaviviridae family. Hepatitis C viruses are also similar to picorna-viruses (e.g., poliovirus), with the main exception that they are enveloped. Since the cloning of HCV nucleic acid in 1989, considerable progress has been made in characterizing the HCV genome and proteins.

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FIGURE 10-16 Proposed model for hepatitis D virus replication cycle.


The HCV genome is about 9.2 kb in length and contains an IRES and one long ORF that encodes a polyprotein precursor. The HCV +ssRNA is translated by host ribosomes by cap-independent translation. The polyprotein precursor is cleaved into structural and non-structural proteins by cellular and viral proteases. Hepatitis C virus NS5b gene encodes its own RNA-dependent RNA polymerase. The HCV genome has substantial sequence variation because NS5b lacks proofreading capability.


A better understanding of the HCV replication cycle, especially the entry process into host cells is needed. HCV entry requires clathrin and more than one receptor. CD81, an integral membrane protein belonging to a family of tetraspanins; scavenger receptor class B type 1 (SR-B1); and claudin 1 were identified as coreceptors based on their interaction with HCV structural E2 or E1 proteins. The lack of patient response to interferon therapy has been correlated with mutations in the E2 gene. The E1 and E2 proteins presumably self-assemble to form the virion (Figure 10-17).

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FIGURE 10-17 Hypothetical structure of the HCV. Nonstructural proteins are labeled “NS” followed by a number. They have diverse functions and form the replication complex. The p7 protein has no known function, and it has not been classified as structural or nonstructural. UTR stands for untranslated region.


Hepatitis E Virus

Hepatitis E virus (HEV) is transmitted by the fecal–oral route, often through contaminated water. Like HAV, it has a +ssRNA genome. It is a member of the Hepeviridae family. Hepatitis E is clinically indistinguishable from hepatitis A, but HEV particles are much less stable than those of HAV. Hepatitis E viruses are approximately 32–34 nm in diameter, nonenveloped, and icosahedral shaped. Its genome is approximately 7.2 kb in length and contains short 5′- and 3′-end noncoding regions. The 3′ end contains a poly(A) tail.


The genome has three ORFs, and all three coding frames are used to express different proteins. ORF1 codes for nonstructural proteins that possess methyltransferase, protease, RNA helicase, and RNA-dependent RNA polymerase (replicase) motifs. ORF2 does not overlap with ORF1 and is located at the 3′ end of the genome and presumably codes for the only capsid (structural) protein. ORF3 begins at the end of ORF1. It overlaps with ORF2 and codes for a small phosphoprotein of undefined function (FIGURE 10-18). Presumably, HEV replicates solely in the cytoplasm of the host cell. Overall, our knowledge of HEV replication is poor. The lack of a suitable cell culture system to propagate HEV is an obstacle in deciphering its molecular biology.

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FIGURE 10-18 Genome organization of HEV. UTR stands for untranslated region. ORF1 codes for a nonstructural, replicative polyprotein; ORF2 codes for a capsid protein; and ORF3 codes for a protein of unknown function.


10.6 Pathophysiology of Chronic Hepatitis Virus Infections


The biggest difference between enveloped and naked hepatitis viruses is that the enveloped viruses (hepatitis B, C, D, and G) cause persistent and chronic infections. These viruses have devised a strategy to escape detection and elimination following natural infection. In other words, the viruses hide from the host’s immune system during a chronic infection. Reports have described reactivation of HBV after liver transplantation with donor livers from HBV-immune individuals. In other words, HBV DNA was recovered from “immune” patients and donors. This suggests that HBV may exist in the hepatocytes in a latent form for a long time. It has been shown that hepatotropic viral clearance is associated with a strong virus-specific cytotoxic TC lymphocyte and TH lymphocyte response. Antibody-mediated responses have been inadequate in clearing the infection.


Evidence is accumulating that the liver damage caused by HBV and HCV during the course of infection may result from an autoimmune reaction directed against hepatocyte membrane antigens that is initiated by HCV or HBV infection. Damage to the liver is mediated by inflammatory cytokines, which are small proteins that communicate between cells and may play a role in the liver. They play an important role in the defense against viral infections. Both HBV and HCV may have the capacity to modulate cytokine gene expression and responses. The chronic infections caused by HBV and HCV may result in an ongoing inflammatory response that activates the process of liver cirrhosis. More studies are needed to develop strategies useful in combating the effects of HBV and HCV infections on the liver.


10.7 Genetic Diversity of Hepatitis Viruses


Viral evolution is the result of genetic variation and selection of a variant from a large viral population. RNA viruses have the highest mutation rates due to the lack of RNA polymerase proofreading activity, followed by ssDNA viruses and lastly dsDNA viruses (for a review of the genome types of the hepatitis viruses, see Table 10.1). Diversity in viral genotype occurs at the time of replication. HAV has only one known serotype, but at least seven different genotypes have been identified worldwide. Genotype refers to the genetic makeup of the virus. Individuals who are infected by HAV in one part of the world are not susceptible to reinfection by a different HAV genotype in another part of the world. Immunoglobulin and vaccines prepared in a variety of countries protect travelers from HAV infection irrespective of their destination.


Both HAV and HEV have +ssRNA genomes. Infections caused by hepatitis A and E viruses are uncommon in the United States and developed countries. The majority of HEV genomic isolates have only been partially sequenced. Based on the genomic sequences that are available, four major genotypes have been identified. Different HEV genomes have been identified in sewage from pig slaughterhouses. This suggests that different HEVs are widespread in the general swine population. There are reports of HEV antibodies present in the sera of farmers working in close contact with pigs, raising the possibility that pig HEV could infect humans. Pigs are considered the major reservoir for hepatitis E viruses. HEV isolates from humans are genetically related to those from pigs in the same area. India has been an exception because genotype 1 HEV is mainly circulating in humans and genotype 4 HEV in swine in this region.


Chronic HBV and HCV share a common theme in that many variants arise during the course of infection. Individuals with chronic hepatitis B or C produce 1010 to 1011 viruses daily, which indicates that the hepatitis viruses are actively replicating. The high replication activity by the viral RNA-dependent RNA polymerases that lack proofreading function explains, in part, the rapid emergence of viral variants. It also may explain the frequent immune-escape phenomena. Mutations in the HBV promoter of the core gene or in the precore region of the core gene are frequently found in patients with chronic hepatitis B.


Like other RNA viruses, HCV has a high mutation rate, resulting in genetic diversity. So far, six main HCV genotypes and more than 50 subtypes have been identified. Genotype 1 is the most common in the United States. Recently, new variants, assigned as types 7–11, have also been recognized. Genotypes are further broken down into subtypes designated by a lowercase letter. For example, the three subtypes of HCV type 1 are designated 1a, 1b, and 1c. The genotypes show general global patterns. Genotypes 1, 2, and 3 are the most common genotypes in North America and Western Europe (FIGURE 10-19). Knowing the genotype helps to predict the likelihood of treatment response (see Section 10.7).


10.8 Management and Prevention of Hepatitis A–E Viruses


Hepatitis A


No specific treatment for hepatitis A infection is available, and management is supportive. Spread of hepatitis A virus infections can be reduced by handwashing and proper sanitary disposal of human feces. Prevention by passive immunization of individuals who likely have been exposed to or in close contact with a person with hepatitis A is achieved by intramuscular injection of human immunoglobulin that contains antibodies against HAV. In general, if administered before exposure or within 2 weeks after exposure, immunoglobulin is 85% effective in preventing hepatitis A.

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FIGURE 10-19 Geographic distribution of the HCV genotypes. Genotype 1 is most prevalent in the United States, in comparison to countries in the Middle East and Africa, where genotypes 4 and 5 are the most dominant.


Hepatitis A vaccines were developed in a manner similar for that of poliovirus vaccines. Formalin-inactivated, cell culture–produced, whole-virus vaccines such as Havrix, Vaqta, and Avaxim are now approved for use in much of the world. HAV is propagated in MRC-5 cells (a cell line of human lung fibroblast cells). The Centers for Disease Control and Prevention (CDC) recommends that all children 12–24 months of age should be vaccinated against HAV. At least two doses that are 6 months apart are needed for lasting protection. In addition to high-risk persons, those who travel to or work in areas with a high prevalence of hepatitis A (e.g., Mexico, Central or South America, Asia, Africa, and Eastern Europe) should be vaccinated. Live, attenuated vaccines similar to the Sabin-poliovirus oral vaccine have been developed and tested in animals and humans but are not yet available, as they have not been able to induce a satisfactory immune response when administered orally.


Hepatitis B


The basic aim of treating hepatitis B patients is to stop HBV replication and prevent end-stage liver disease. Six licensed drug therapies are used against hepatitis B infections: interferon α-2b, lamivudine, adefovir dipivoxil, entecavir, telbivudine, and tenofovir. Interferon is an immune modulator and is administered as an injection. About 46% of patients respond to inter-feron α-2b (intron A) in the first year of treatment. A response is the cessation of viral replication. Interferon α-2b treatment has three drawbacks: it has a large number of side effects, it is expensive, and it is not effective in a high percentage of patients.


Lamivudine (Epivir), adefovir dipivoxil (Hepsera), entecavir (Baraclude), telbivudine (Tyzeka), and tenofovir (Viread) are nucleoside analogs that are potent inhibitors of DNA replication. Telbivudine interferes with +strand synthesis of HBV DNA, thereby inhibiting HBV polymerase. Lamivudine was approved for treatment in 1999 and was first used to treat HIV patients; its mechanism of action is the inhibition of reverse transcriptase. Advantages of lamivudine are that it has few side effects compared to interferon α-2b and has been shown to dramatically inhibit hepatitis B virus DNA replication. The disadvantage is that replication rapidly returns when therapy is stopped. Lamivudine-resistant strains of HBV develop at a rate of 15–20% per year of therapy.


Adefovir dipivoxil has been used since 2002 as a first-line monotherapy, combination therapy, or a therapy in patients who have lamivudine-resistant infections. It can be administered orally, and patients tolerate it well.


Entecavir was approved for use in 2005. It cannot be used as monotherapy in patients with HBV/HIV infection who need treatment for HBV infection. Telbivudine was approved in 2006, but it has not been widely embraced for therapy against HBV. Tenovir was first approved as an HIV reverse transcriptase inhibitor and has similar activity against HBV. It was approved for use in hepatitis B patient treatment in 2008. To date, it has had superior results compared to adefovir and is expected to replace adefovir as a treatment for chronic hepatitis B.


Antibodies against HBsAg protect individuals from acute and chronic hepatitis B infection if used shortly after exposure to hepatitis B virus. Immunoglobulin is used as an adjunct to hepatitis B vaccine in preventing HBV transmission from an infected mother to fetus. If untreated, 70–90% of infants born to HBeAg-positive mothers become infected at birth and develop chronic hepatitis B. Hepatitis B immunoglobulin is also used for postexposure prophylaxis after accidental needlestick or other medical-related injuries in persons who work with infectious body fluids that contain HBsAg.


Early hepatitis B vaccines were prepared by harvesting the 17- to 25-nm particles from the plasma of individuals with chronic HBV infections. The hepatitis B virions were purified and inactivated by heat, formal-dehyde, urea, or pepsin. Plasma-derived vaccines have been available since 1982. Subsequently, vaccine manufacturers have used genetic engineering to express HBsAg in Saccharomyces cerevisiae, a yeast, and mammalian cells. This has led to the development of recombinant DNA vaccines.


Recombinant DNA vaccines were produced by inserting the S gene that codes for the HBsAg into a plasmid DNA. The plasmids containing the S gene were transformed into yeast or transfected into mammalian cells in culture. The HBsAg was expressed and purified by chromatography and filtration (FIGURE 10-20). Under these conditions, purified HBsAg self-assembles into the spherical particles closely resembling the 17- to 25-nm particles found in the serum of people with chronic hepatitis B. The vaccine is not an infectious particle, making this a very safe vaccine.


The CDC recommends that all newborns in the United States receive a birth dose of the recombinant (noninfectious) hepatitis B vaccine before leaving the hospital unless a physician provides a written order to defer the birth dose. A second dose is required at 1–2 months of age and a third dose at 6–18 months of age. HBV can cause liver cancer. An added benefit of the hepatitis B vaccine is that it prevents hepatitis B–related liver cancer. This vaccine was the first to prevent a cancer. From 1990 to 2002, the incidence of hepatitis B decreased 67% in the United States. Hepatitis B vaccination programs have been implemented with considerable success. Unfortunately, the recombinant hepatitis B vaccine is expensive to purchase, which restricts its availability in developing nations. For this reason, oral hepatitis B vaccine candidates are being produced and delivered in plants such as potatoes and soybeans. A number of candidates are suitable for early stage clinical trials.


Hepatitis C


An estimated 2.7–3.9 million people in the United States have chronic hepatitis C and about 275,000 Canadians. Chronic HCV infection affects 170 million people worldwide. HCV infections become chronic in up to 85% of patients. No vaccine is available to prevent HCV infection.

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FIGURE 10-20 Flowchart showing the steps involved in preparing a recombinant DNA hepatitis B vaccine in yeast. The HBV gene that codes for the HBsAg was inserted into a plasmid DNA and transformed into Saccharomyces cerevisiae. The yeast cells expressed the HBsAg. The HBsAg proteins were purified and used as the HBV vaccine. Since the vaccine is using a “part” of a virus, it is a safe vaccine that cannot cause disease.


The HCV genotype determines the duration of treatment and expected efficacy of antiviral activity (see Section 10.9 on genetic diversity and genotypes of HCV). Prior infection with HCV does not protect against later infection with the same or different genotypes of the virus. This is because infected individuals typically have an ineffective immune response toward HCV due to changes in cells caused by HCV during infection. No pre- or postexposure immunoglobulin is available for treatment.


Until 2011, the mainstay for the treatment of acute and chronic HCV was pegylated interferon α-2a or pegylated interferon α-2b and/or ribavirin. Pegylation involves the attachment of a polyethylene glycol molecule to the active interferon molecule. Pegylation improves the half-life of interferon, limiting its degradation in the body, thereby making it long acting. interferon is a glycoprotein produced by the body as part of the innate, or nonspecific, immune system response toward viral infection. Therefore, interferon treatment may prevent HCV replication and the production of new infectious virions in infected individuals. Interferon is administered to the patient as weekly injections. Ribavirin is an oral nucleoside analog that inhibits genome replication of a number of viruses. Response to therapy is measured by determining the levels of serum transaminases and the detection of HCV RNA genomes in serum.


In order to prescribe a treatment plan with the highest probability of success, a person must have his or her particular HCV genotype and subtype identified. Once the genotype is identified, it need not be tested again; genotypes do not change during the course of infection. Knowing the genotype predicts the likelihood of treatment outcome and determines the duration of treatment. The combination of ribavirin/pegylated interferon α-2b treatment over a 6- to 12-month period increased overall cure rates to an average of 40% when treating patients with genotype 1 or 4 and an average of 80% for genotypes 2 and 3. However, the therapy is challenging because of the side effects caused by interferon and ribavirin. Ribavirin is teratogenic. It interferes with normal embryonic development and therefore cannot be taken by patients who are pregnant. The main side effect of ribavirin is anemia. Genotype-5 patients respond similarly to those with genotype 1 toward combination treatment. Patients with genotype 6 have an intermediate response level, between that seen with genotype 1 and genotype 2 or 3. The response to standard treatment is not well established for genotype variants 7–11. Patients needed to be monitored closely for complications or symptoms of adverse reactions by combination therapy. About 50% of patients cannot tolerate the side effects of interferon and ribavirin.


Thus, it was clear that there was a need for more promising orally administered antiviral drugs that interrupted HCV replication and that caused fewer side effects. In 2005, the development of a tissue-culture system to propagate HCV paved the way for researchers to develop new antivirals (VIRUS FILE 10-2).


In 2011, Victrelis (boceprevir) and Incivek (telaprevir) were enthusiastically approved by the FDA as two new treatment options for HCV infection. Boceprevir binds to the active site of the NS3 HCV protease. Telaprevir targets the NS3/4a protease of HCV. Targeting the protease activity cripples HCV replication. The viral proteases are responsible for processing the nonstructural polyprotein located in the N-terminus of the HCV polyprotein. Inhibition of the proteases results in unprocessed HCV polyprotein, which ultimately affects the viral proteins involved in genome replication. Both of the drugs increase the already difficult side effects of standard therapy, such as anemia. However, the drugs do not need to be taken for a full 48-week course of treatment.


Because the drugs target HCV specifically, they are a major improvement over current hepatitis C therapy. Survival rates for people infected with genotype 1 increased from 40% in 2001 to an 80% average in 2011 with the availability of the new drugs that are specific HCV inhibitors. In 2013, two new drugs were approved for chronic hepatitis C: sofosbuvir, an inhibitor of HCV RNA polymerase, and simeprevir, an inhibitor of HCV NS5a replication complex. They were approved for use in combination with or without ribavirin and pegylated interferon. This was a major breakthrough for hepatitis C patients who could not tolerate treatment with interferon due to its toxic effects.


Subsequently, four additional hepatitis C drugs, including the first combination drugs available in pill form that did not require ribavirin or interferon administration, became available during 2014–2015 with the approval of the combination HCV drugs Harvoni and Technivie (TABLE 10-5). The hepatitis C treatment regimens approved in the United States and Canada and by the European Association for the Study of the Liver (EASL) are listed in TABLE 10-6. The interferon-free rows in Table 10-6 are shaded in green. The EASL was formed in 1966 as a small group of 70 hepatologists from 15 European countries to share best medical practices to treat liver disease. Today, the EASL has international influence dedicated to liver disease treatment and research. It has over 4,000 members from all over the world and convenes at the International Liver Congress, where more than 11,000 experts meet and discuss the latest treatment regimens and scientific research. Its mission is educational for physicians and scientists. EASL also facilitates multicenter controlled clinical trials and supports young investigators to ensure that the liver remains at the forefront of research.


The newer drugs that specifically targeted HCV replication resulted in higher than 90% survival rates with less severe side effects because regimens did not require the additional use of interferon and/or ribavirin. This means that HCV is a curable disease. The barrier to treatment is cost. The newer HCV drugs are expensive.


Table 10-5 FDA-Approved Antivirals to Treat Hepatitis C
















































Drug Mechanism of Action Year Approved
Pegylated interferon α-2b Immune system modulator 2001
Ribavirin Nucleoside analog; inhibits viral genome replication 2001
Boceprevir (Victrelis) HCV NS3 protease inhibitor 2011
Telaprevir (Incivek) HCV NS3/4a protease inhibitor 2011
Sofosbuvir (Sovaldi) Inhibits HCV RNA polymerase 2013
Simeprevir (Olysio) HCV NS3/4a protease inhibitor 2013
Ledipasvir (GS-5885) Inhibits HCV NS5a 2014
Harvoni First combo pill of ledipasvir and sofosbuvir 2014
Daclatasvir (Daklinza) Non-nucleoside inhibitor of HCV NS5b polymerase 2015
Technivie Combo pill of ombitasvir (9NS5a inhibitor), paritaprevir (NS3/4A protease inhibitor), and ritonavir (HIV protease inhibitor as a booster) 2015

As of 2014, Harvoni cost $1,125 per pill, or $94,500 for a 12-week course. Sovaldi is a once-daily pill taken with Olysio. The two-drug combination cost about $150,000 for a full course of treatment in 2014. A study by Liverpool University showed that the cost to manufacture a 12-week course of Olysio could be as low as $100–$250. Some insurance companies have decided to limit the availability of the medicine for specific patients (e.g., compensating the “sickest” patients). Access to HCV treatment in low- and middle-income countries is extremely limited because of the high costs of treatment and complexity of patient management. Of the 20 countries with the largest HCV epidemics, 12 are classified as low or lower-middle income. TABLE 10-7 lists income classification by country and most prevalent HCV genotypes.


Table 10-6 Approved Treatment Regimens for Hepatitis C























































































Year Treatment Regimen Duration Genotype
2001 Peg-IFN + RBV 48 weeks 1, 4
2001 Peg-IFN + RBV 24 weeks 2, 3
2011 Peg-IFN + RBV + BOC 24–48 weeks 1
2011 Peg-IFN + RBV + TVR 24–48 weeks 1
2013 Peg-IFN + RBV + SMV 12 weeks 1
2013 Peg-IFN + RBV +SOF 12 weeks 1
2013 Peg-IFN + RBV +SOF 12 weeks 4
2013 RBV + SOF 12 weeks 2
2013 RBV + SOF 24 weeks 3
2014 SOF + LDV 12 weeks 1, 4, 5, 6
2014 SOF + SMV 12 weeks 1,4
2014 HVI 12 weeks 1
2015 SOF + DCV 12 weeks 1–6
2015 RBV + TCV 12 weeks 4
2015 DCV + TCV 12 weeks 1


Abbreviations: Peg-IFN: pegylated interferon; RBV: ribavirin; BOC: boceprevir; TVR: telaprevir; SMV: simeprevir; SOF: sofosbuvir; DCV: daclatasvir; TCV: Technivie; LDV: ledipasvir; HVI: Harvoni.

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Jul 29, 2017 | Posted by in MICROBIOLOGY | Comments Off on Hepatitis Viruses

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