Visualization of Positive and Negative Sense Viral RNA for Probing the Mechanism of Direct-Acting Antivirals against Hepatitis C Virus

Viruses. 2019 Nov 8;11(11):1039. doi: 10.3390/v11111039.


RNA viruses are highly successful pathogens and are the causative agents for many important diseases. To fully understand the replication of these viruses it is necessary to address the roles of both positive-strand RNA ((+)RNA) and negative-strand RNA ((-)RNA), and their interplay with viral and host proteins. Here we used branched DNA (bDNA) fluorescence in situ hybridization (FISH) to stain both the abundant (+)RNA and the far less abundant (-)RNA in both hepatitis C virus (HCV)- and Zika virus-infected cells, and combined these analyses with visualization of viral proteins through confocal imaging. We were able to phenotypically examine HCV-infected cells in the presence of uninfected cells and revealed the effect of direct-acting antivirals on HCV (+)RNA, (-)RNA, and protein, within hours of commencing treatment. Herein, we demonstrate that bDNA FISH is a powerful tool for the study of RNA viruses that can provide insights into drug efficacy and mechanism of action.

Keywords: RNA FISH; Zika virus; direct acting antivirals; hepatitis C virus; negative-sense; positive-sense.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Antiviral Agents / pharmacology*
  • Cell Line
  • Hepacivirus* / drug effects
  • Hepacivirus* / genetics
  • Hepatitis C / drug therapy
  • Hepatitis C / virology
  • Humans
  • In Situ Hybridization, Fluorescence / methods
  • RNA, Viral* / drug effects
  • RNA, Viral* / metabolism
  • Virus Replication / drug effects
  • Zika Virus / drug effects
  • Zika Virus / genetics
  • Zika Virus Infection / drug therapy
  • Zika Virus Infection / virology


  • Antiviral Agents
  • RNA, Viral