Autophagy pathway induced by a plant virus facilitates viral spread and transmission by its insect vector
(A) At different days padp, 30 live dsRNAs-treated leafhoppers positive for transcript of RGDV P8 gene were used for assay of viral genome copies, which were calculated as the log of the copy number of P8 gene/μg insect RNA. Means (±SD) from three biological replicates are shown. The statistical significance was related to the dsGFP control. *P < 0.05. (B) At 48 hpi, effects of dsAtg5, dsAtg8 or dsTorc1 on transcript levels of Atg8 gene in dsRNAs-treated leafhoppers as revealed by RT-qPCR assay. ACTB was used as the internal control. Means (±SD) from three biological replicates are shown. *P < 0.05. (C) The accumulation levels of ATG8 and SQSTM1 in dsRNAs-treated leafhoppers were detected by western blot assay using ATG8- and SQSTM1-specifc IgG, respectively. Insect ACTB was detected with ACTB-specific IgG as a control. (D) RGDV infection in the intestine of leafhoppers receiving dsRNAs at 4 days padp, as detected by immunolabelling with P8-rhodamine (red) and the ACTB dye phalloidin-FITC (green). Bars, 200 μm. (E) Effects of autophagy pathway on the transmission of RGDV by R. dorsalis. Leafhoppers treated with dsRNAs were used to infect susceptible rice plants, and viral transmission rate was determined by RT-PCR assay. Means (±SD) from three independent replicates are shown. The statistical significance is related to the dsGFP control. *P < 0.05. fc, filter chamber. mg, midgut. amg, anterior midgut. mmg, middle midgut. pmg, posterior midgut.