http://orcid.org/0000-0002-0618-5611
Figures & data
a–c Tick cell lines HAE/CTVM8 and HAE/CTVM9 were infected with CCHFV at MOI 0.1 or MOI 1.0. At the indicated time points: a Infectious viral particles released were titrated by the focus forming unit (FFU) assay in Vero cells, error bars = S.D.; b The relative increase in viral RNA in the infected cells was evaluated by qRT-PCR; c expression of the CCHFV-N protein was evaluated by western blot, C = uninfected cells. The experiment was performed three times in duplicate, and sample analyses were performed in duplicate. d HAE/CTVM8 cells were infected with the CCHFV IbAr10200 strain or rCCHFVwt at MOI 0.1. Viral replication was compared by measuring the relative amount of viral RNA in the infected cells at the indicated time points. e–f Human SW13 and tick HAE/CTVM8 cells were infected with rCCHFVwt or rCCHFVmut at MOI 0.1; viral replication was evaluated by: e titration of the virus progeny for both cell lines, and f measuring the relative amount of viral RNA in the infected SW13 cells. *P < 0.05, unpaired t-test. The experiment was performed three times in duplicate, and sample analyses were performed in duplicate. g HuH-7 (JCRB0403) donor cells were transfected with the CCHFV tc-VLP system, using either a wild-type (pC_N) or a mutant (pC_N_D266+269A) N, in the context of a wild-type (L_wt) or a transcriptionally incompetent polymerase (L_D693A). An inactive polymerase (L_∆DD) was used as a control. Minigenome luciferase activity was measured 3 days post transfection. The presence of tc-VLPs in donor cell supernatants was detected by transfer of supernatants onto HuH-7 indicator cells expressing L-wt and N and measurement of luciferase activity 24 h p.i. The results were normalized against the control L_wt+N_wt value set to 100%. The experiment was done in triplicate. h Replication of rCCHFVwt and rCCHFVmut was also evaluated in HAE/CTVM8 cells over 17 days by measuring the relative amount of intracellular viral RNA. *P < 0.002, ANOVA. The experiment was performed three times in duplicate, and sample analyses were performed in duplicate
