The Toll-Dorsal Pathway Is Required for Resistance to Viral Oral Infection in Drosophila
Figure 6
Toll Pathway mutant flies are less resistant to other RNA viruses oral infection.
(A) Survival of pll−/− and w1118 iso flies after CrPV oral infection (1.76×1010 TCID50/ml) or buffer. pll−/− flies were significantly more sensitive to CrPV than w1118 iso (Cox proportional hazard mixed effect model, p<0.001). (B) CrPV RNA levels in pll−/− and w1118 iso flies upon oral infection (1.76×1010 TCID50/ml). CrPV loads are significantly different between pll−/− and w1118 iso line (Wilcoxon test, p<0.005). (C) pll−/− and w1118 iso flies were systemically infected with CrPV at three different concentrations (106, 107, 108 TCID50/ml). pll−/− mutant flies were not more susceptible to CrPV systemic infection than w1118 iso control flies (Cox Proportional Hazards Model, p = 0.966, p = 1.000 and p = 0.974, respectively). (D) Survival of pll−/− and w1118 iso flies upon Nora oral infection or buffer. pll−/− flies were not more sensitive than w1118 iso (Cox proportional hazard mixed effect model, p = 0.887). (E) Nora RNA levels upon oral infection. Nora loads are significantly different between pll−/− and w1118 iso line (Wilcoxon test, p<0.005). (F) Survival of pll−/− and w1118 iso flies upon FHV oral infection (1010 TCID50/ml) or buffer. pll−/− flies were significantly more sensitive than w1118 iso (Cox proportional hazard mixed effect model, p<0.001). (G) FHV RNA levels upon oral infection (1010 TCID50/ml). FHV loads are significantly different between pll−/− and w1118 iso line (Wilcoxon test, p<0.005) (in the other independent replicate the difference in medians is 20-fold and p = 0.05). (H) pll−/− and w1118 iso flies were systemically infected with FHV at three different concentrations (106, 107, 108 TCID50/ml). pll−/− mutant flies were not more susceptible to FHV systemic infection than w1118 iso control flies (Cox Proportional Hazards Model, p = 0.819, p = 0.709 and p = 0.225, respectively). For survival experiments (A, C, D, F and H) sixty 3–6 days old males of each line per treatment were used and survival was scored daily. Survival experiments for oral infections were performed thrice, yielding similar results. Survival data of all replicates was analysed together using the Cox proportional hazard mixed effect model. For viral loads experiments (B, E, G) 3–6 days old males of each line were orally infected with the virus of interest and collected 5–6 dpi for RNA extraction and RT-qPCR. Relative amount of virus was calculated using host Rpl32 mRNA as a reference and values are relative to the median of the w1118 iso samples. Each point represents the relative virus amount of a single fly and lines are medians of these values. All viral loads experiments were performed twice yielding similar results.