The antisense 5' end of the V2 gene confers enhanced resistance against the monopartite begomovirus cotton leaf curl Kokhran virus-Burewala strain

Abstract

Whitefly-transmitted viruses of the genus Begomovirus (the family Geminiviridae) have become a limiting factor for agricultural productivity in many warmer parts of the world. The economies of Pakistan and India have, since the early 1990s, suffered losses due to cotton leaf curl disease (CLCuD). The disease is caused by begomoviruses, the most important of which at this time is cotton leaf curl Kokhran virus strain Burewala (CLCuKoV-Bu), and a disease-specific betasatellite, cotton leaf curl Multan betasatellite (CLCuMuB). Efforts to minimize losses due to CLCuD rely mainly on the use of insecticides to kill the whitefly vector; no resistant cotton varieties are currently commercially available. The study described here has investigated RNA interference technology for its potential to yield resistance against CLCuKoV-Bu and three other begomoviruses; CLCuKoV, tomato leaf curl New Delhi virus (ToLCNDV) and Pedilanthus leaf curl virus (PeLCV). Three fragments of the virion-sense V2 gene of CLCuKoV-Bu were transformed into Nicotiana benthamiana in antisense orientation and transgenic lines expressing virus-specific short RNAs were assessed for their ability to yield resistance. Only CLCuKoV-Bu with the V2 sequence closest to the promoter was resistant. Inoculation of CLCuKoV-Bu with CLCuMuB into transgenic plants did not significantly affect the outcome, although viral DNA was detected in number of plants, suggesting that the betasatellite may impair RNAi resistance. Overall the results indicate that targeting the 5' end of V2 gene using antisense-RNA has the potential to deliver resistance against begomoviruses and that RNAi-based resistance imparts some degree of resistance to heterologous viruses.