Replication of RNA of Tick-Borne Encephalitis Virus in New Transferable Cell Lines of the Natural Host Apodemus peninsulae

Abstract

Cell lines derived from natural virus hosts are one of the most promising tools in virological research. The aim of this work is to establish the dynamics of RNA replication of tick-borne encephalitis virus (TBEV) in new transferable cell line of kidney of the Korean field mouse Apodemus peninsulae (ApnK) and compare it with replication in the traditional culture of fetal porcine kidney cells (SPEVs). The transplantable ApnK cell line was obtained in the laboratory of transmissive infections of the Scientific Center for Family Health and Human Reproduction Problems (Irkutsk), the identity of the culture was established by the analysis of nucleotide sequence of mitochondrial cytochrome B gene (mtCytB, GenBank access number KT983422), and the D-loop sequence of the mitochondrial genome (KT983423). Cells were inoculated with TBEV strain of the Siberian subtype 92M and the total RNA was isolated from them 0, 4, 8, 16, and 24 h after infection. In addition, RNA was also isolated 1, 4, 7, and 19 days after infection. The amount of intracellular RNA of the positive polarity (+RNA) was assessed by quantitative real-time PCR. At the initial stage of infection (0 h after infection) in ApnK cells, the intracellular +RNA concentration was significantly lower than in SPEV, 4.5 lg versus 5.5 lg of genome-equivalents/μL (p = 0.01). After 8 h, in ApnK cells there were no significant difference with points 0 and 4 (p = 0.5 and p = 0.1, respectively), whereas in SPEV at this stage the concentration of RNA was significantly higher than at points 0 and 4 (p = 0.001 and 0.01, respectively). In general, during the first day of infection, the concentration of +RNA in ApnK cells was almost ten times less than in SPEV (p = 0.001). By the fourth day after infection, ApnK cells showed maximum concentrations of +RNA, 7.8 lg of genome-equivalents/μL, with no differences with SPEV (p = 0.06). Subsequently, the amount of RNA in ApnK cells remained at the same level for 19 days, and in SPEV in late terms there was a tendency to decrease in the concentration of +RNA. The results obtained suggest that TBEV in A. peninsulae has an intracellular mechanism for controlling viral infection, which acts both at the stage of the entrance of virion into the cell and by regulating the rate of synthesis of genomic and/or replicative forms of viral RNA.