Development and characterization of recombinant tick-borne encephalitis virus expressing mCherry reporter protein: A new tool for high-throughput screening of antiviral compounds, and neutralizing antibody assays

Antiviral Res. 2021 Jan:185:104968. doi: 10.1016/j.antiviral.2020.104968. Epub 2020 Nov 4.


The flavivirus, tick-borne encephalitis virus (TBEV) is transmitted by Ixodes spp. ticks and may cause severe and potentially lethal neurological tick-borne encephalitis (TBE) in humans. Studying TBEV requires the use of secondary methodologies to detect the virus in infected cells. To overcome this problem, we rationally designed and constructed a recombinant reporter TBEV that stably expressed the mCherry reporter protein. The resulting TBEV reporter virus (named mCherry-TBEV) and wild-type parental TBEV exhibited similar growth kinetics in cultured cells; however, the mCherry-TBEV virus produced smaller plaques. The magnitude of mCherry expression correlated well with progeny virus production but remained stable over <4 passages in cell culture. Using well-characterized antiviral compounds known to inhibit TBEV, 2'-C-methyladenosine and 2'-deoxy-2'-β-hydroxy-4'-azidocytidine (RO-9187), we demonstrated that mCherry-TBEV is suitable for high-throughput screening of antiviral drugs. Serum samples from a TBEV-vaccinated human and a TBEV-infected dog were used to evaluate the mCherry-based neutralization test. Collectively, recombinant mCherry-TBEV reporter virus described here provides a powerful tool to facilitate the identification of potential antiviral agents, and to measure levels of neutralizing antibodies in human and animal sera.

Keywords: Antivirals; Neutralization test; Reporter virus; Tick-borne encephalitis virus.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Antibodies, Neutralizing / blood*
  • Antibodies, Viral / blood
  • Antiviral Agents / isolation & purification*
  • Cell Line
  • Cricetinae
  • Encephalitis Viruses, Tick-Borne / genetics*
  • Encephalitis Viruses, Tick-Borne / growth & development
  • Encephalitis, Tick-Borne / immunology
  • Encephalitis, Tick-Borne / virology
  • High-Throughput Screening Assays / methods*
  • Humans
  • Kidney / cytology
  • Luminescent Proteins / genetics*
  • Neutralization Tests*


  • Antibodies, Neutralizing
  • Antibodies, Viral
  • Antiviral Agents
  • Luminescent Proteins
  • red fluorescent protein