TY - JOUR

T1 - Novel infectious cDNA clones of hepatitis C virus genotype 3a (strain S52) and 4a (strain ED43): genetic analyses and in vivo pathogenesis studies

AU - Gottwein, Judith

AU - Scheel, Troels

AU - Callendret, Benoit

AU - Li, Yiping

AU - Eccleston, Heather B

AU - Engle, Ronald E

AU - Govindarajan, Sugantha

AU - Satterfield, William

AU - Purcell, Robert H

AU - Walker, Christopher M

AU - Bukh, Jens

PY - 2010

Y1 - 2010

N2 - Previously, RNA transcripts of cDNA clones of hepatitis C virus (HCV) genotypes 1a (strains H77, HCV-1, and HC-TN), 1b (HC-J4, Con1, and HCV-N), and 2a (HC-J6 and JFH1) were found to be infectious in chimpanzees. However, only JFH1 was infectious in human hepatoma Huh7 cells. We performed genetic analysis of HCV genotype 3a (strain S52) and 4a (strain ED43) prototype strains and generated full-length consensus cDNA clones (pS52 and pED43). Transfection of Huh7.5 cells with RNA transcripts of these clones did not yield cells expressing HCV Core. However, intrahepatic transfection of chimpanzees resulted in robust infection with peak HCV RNA titers of approximately 5.5 log(10) international units (IU)/ml. Genomic consensus sequences recovered from serum at the times of peak viral titers were identical to the sequences of the parental plasmids. Both chimpanzees developed acute hepatitis with elevated liver enzymes and significant necroinflammatory liver changes coinciding with detection of gamma interferon-secreting, intrahepatic T cells. However, the onset and broadness of intrahepatic T-cell responses varied greatly in the two animals, with an early (week 4) multispecific response in the ED43-infected animal (3 weeks before the first evidence of viral control) and a late (week 11) response with limited breadth in the S52-infected animal (without evidence of viral control). Autologous serum neutralizing antibodies were not detected during the acute infection in either animal. Both animals became persistently infected. In conclusion, we generated fully functional infectious cDNA clones of HCV genotypes 3a and 4a. Proof of functionality of all genes might further the development of recombinant cell culture systems for these important genotypes.

AB - Previously, RNA transcripts of cDNA clones of hepatitis C virus (HCV) genotypes 1a (strains H77, HCV-1, and HC-TN), 1b (HC-J4, Con1, and HCV-N), and 2a (HC-J6 and JFH1) were found to be infectious in chimpanzees. However, only JFH1 was infectious in human hepatoma Huh7 cells. We performed genetic analysis of HCV genotype 3a (strain S52) and 4a (strain ED43) prototype strains and generated full-length consensus cDNA clones (pS52 and pED43). Transfection of Huh7.5 cells with RNA transcripts of these clones did not yield cells expressing HCV Core. However, intrahepatic transfection of chimpanzees resulted in robust infection with peak HCV RNA titers of approximately 5.5 log(10) international units (IU)/ml. Genomic consensus sequences recovered from serum at the times of peak viral titers were identical to the sequences of the parental plasmids. Both chimpanzees developed acute hepatitis with elevated liver enzymes and significant necroinflammatory liver changes coinciding with detection of gamma interferon-secreting, intrahepatic T cells. However, the onset and broadness of intrahepatic T-cell responses varied greatly in the two animals, with an early (week 4) multispecific response in the ED43-infected animal (3 weeks before the first evidence of viral control) and a late (week 11) response with limited breadth in the S52-infected animal (without evidence of viral control). Autologous serum neutralizing antibodies were not detected during the acute infection in either animal. Both animals became persistently infected. In conclusion, we generated fully functional infectious cDNA clones of HCV genotypes 3a and 4a. Proof of functionality of all genes might further the development of recombinant cell culture systems for these important genotypes.

KW - Animals

KW - Antibodies, Neutralizing

KW - Cell Line

KW - DNA, Complementary

KW - Genotype

KW - Hepacivirus

KW - Hepatitis C

KW - Hepatitis C Antibodies

KW - Hepatocytes

KW - Humans

KW - Liver

KW - Liver Function Tests

KW - Monkey Diseases

KW - Pan troglodytes

KW - RNA, Viral

KW - T-Lymphocytes

U2 - 10.1128/JVI.02667-09

DO - 10.1128/JVI.02667-09

M3 - Journal article

C2 - 20200247

VL - 84

SP - 5277

EP - 5293

JO - Journal of Virology

JF - Journal of Virology

SN - 0022-538X

IS - 10

ER -