Characterisation of non-transmissible mutants of TYLCSV in the whitefly vector Bemisia tabaci

The monopartite Begomovirus (Geminiviridae) Tomato yellow leaf curl Sardinia virus (TYLCSV) has a capsid protein (CP) indispensable for plant infection and vector transmission. Four amino acids (aa) are essential for transmissibility: Q129, N130, Q134, and D152. In this study, three non-transmissible (NT) mutants were characterized: a single mutant N130D (named QDQD), a double mutant Q129P and Q134H (PNHD), and a triple mutant with a further D152E change (PNHE), in comparison with the wild-type virus (wt or QNQD). All these mutants formed virions in infected plants.To understand the reasons for their non-transmissibility, a detailed study on their relationship with whiteflies was undertaken. Using quantitative dot-blot hybridization and real-time PCR, the kinetics of these mutants were studied in the vector B. tabaci and the non-vector Trialurodes vaporariorum exposed to agroinoculated infected plants. Mutant QDQD was not detected by dot-blot in the insects, even at the end of the acquisition. PNHD was acquired and circulated in both B. tabaci and T. vaporariorum for at least 7 days, while the PNHE mutant circulated in B. tabaci only. We studied whether the pre-acquisition of NT mutants interfered with the transmission of the wt virus. Transmission of the wt virus was inhibited when B. tabaci were pre-exposed to plants infected by the PNHE mutant, but not by other mutants. An immunolocalisation protocol was set up to detect the viral CP in B. tabaci previously exposed to infected plants. Data will be presented on the localization of the CP in the vector by immunoelectron microscopy.