Researchers Clone Infectious Hepatitis C Virus
Investigators supported by the National Institute of Allergy and Infectious Diseases (NIAID) have for the first time isolated an infectious clone of the hepatitis C virus (HCV), proving that the virus alone is sufficient to cause disease and broadly expanding the research horizon for HCV investigators.
"This is a major development that no doubt will profoundly influence the field of hepatitis C research," says NIAID Director Anthony S. Fauci, M.D. "Having an infectious HCV clone available will greatly facilitate studies on virus evolution, pathogenesis and host immune response. It will enable scientists to better understand the factors and mechanisms that determine whether virus is cleared from the body or produces a chronic infection."
When the hepatitis C virus was first discovered in 1989, scientists believed they had found the cause of the many cases of hepatitis, often acquired through blood transfusions, that could not be attributed to either the hepatitis A or B viruses. Since then, researchers have detected HCV genetic sequences and antibodies in patients with classic non-A, non-B hepatitis, and screening the blood supply for HCV has lowered significantly the incidence of this disease.
Nevertheless, scientists have been unable to grow the virus efficiently in tissue culture or purify it using other laboratory procedures, necessary steps in the process of verifying that a suspected infectious agent is able to cause disease. Therefore, questions have remained about whether the HCV genetic sequences found in patients truly represent the infectious agent, or are defective or non-infectious versions of the virus.
Charles M. Rice, Ph.D., and colleagues at Washington University in St. Louis, Mo., and at the Food and Drug Administration (FDA) in Bethesda, Md., have now answered these questions. Their findings appear in the July 25, 1997 issue of the journal Science.
The researchers extracted HCV RNA, the virus’s genetic material, from the serum of an individual infected with the virus. They then made thousands of DNA "clones" of the HCV RNA which, in turn, were transcribed back into RNA and injected into the livers of chimpanzees to see if they could cause infection. Chimpanzees are the only known non-human host for HCV.
After 34 of the most promising RNA transcripts failed to cause infection in the chimpanzees, Dr. Rice and his colleagues analyzed the sequences of numerous HCV clones to determine a "consensus sequence" of highly conserved regions within the HCV genome. The researchers then constructed a full-length "consensus clone," reflecting the consensus sequence. RNA transcripts of the consensus clone, which had also been tagged with a unique molecular identification marker, were then injected directly into the livers of two healthy chimpanzees.
In the weeks following injection, the researchers detected signs of infection in the chimpanzees, including enzymes produced in response to liver damage, antibodies to HCV and the presence of HCV RNA. Liver biopsies of each animal also showed tissue damage typical of that seen in HCV-infected chimpanzees. Identification markers on the HCV RNA revealed that the consensus clone had caused infection.
"The RNA levels in the chimps increased steadily in the weeks following inoculation," says Dr. Rice. "If the RNA had come from the original inoculum, instead of from new virus particles, its concentration would have decreased. The RNA also was resistant to RNase, an enzyme that breaks down naked RNA. The enzyme does not degrade RNA that is inside viral particles."
The new finding should be a boon to HCV research. "Viral clones can now be used to establish infections and study the genetics of HCV replication in laboratory animals," says John La Montagne, Ph.D., director of NIAID’s Division of Microbiology and Infectious Diseases.
"Importantly," he adds, "large quantities of infectious HCV RNAs can be produced and used to find ways to grow the virus in cell culture."
Dr. Rice’s research is supported in part through one of four Hepatitis C Cooperative Research Center grants awarded by NIAID in 1996. These Centers are multidisciplinary, collaborative research units designed to generate the knowledge needed to devise preventive and therapeutic strategies for hepatitis C infection.
HCV causes approximately 150,000 cases of acute viral hepatitis each year in the United States. Recovery from infection is rare and between 70 and 90 percent of infected persons become chronic carriers of the virus. According to the Centers for Disease Control and Prevention, chronic liver disease due to HCV causes between 8,000 and 10,000 deaths and leads to about 1,000 liver transplants each year in the United States.
NIAID, a component of the National Institutes of Health (NIH), supports research on AIDS, tuberculosis and other infectious diseases, as well as allergies and immunology. NIH and FDA are agencies of the U.S. Department of Health and Human Services.
NIAID press releases, fact sheets and other materials are available on the Internet via the NIAID home page at http://www.niaid.nih.gov.
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