The efficacy of Citrus tristeza virus as a vector for virus induced gene silencing in Huanglongbing-affected citrus

Abstract

Huanglongbing (HLB) is the most damaging disease in citrus. It is caused by the putative bacterium, “Candidatus Liberibacter asiaticus” and is transmitted by the Asian citrus psyllid, Diaphorina citri. Unfortunately, no cure has been found or effective control strategy established for the combat of this disease. Citrus tristeza virus (CTV) was employed successfully for virus induced gene silencing system (VIGS). Previous studies reported the successful use of citrus mediated CTV-based VIGS against D. citri; however, the system was not tested in “Ca. L. asiaticus”-infected citrus. In addition, very little is known about the interaction between CTV and “Ca. L. asiaticus”. Herein, this relationship was explored by infiltrating citrus plants with CTV carrying a truncated phytoene desaturase gene (pds) as a reporter gene. Repressing pds in citrus causes a photo-bleached leaf phenotype. These plants were then graft-inoculated with budwood from “Ca. L. asiaticus”-infected plants. A strong photo-bleaching phenotype indicates that the CTV is present and actively replicating, whereas a weak leaf phenotype indicates reduced or absent virus titer. After infection with “Ca. L. asiaticus”, the leaf-bleaching phenotype gradually became reduced, and the HLB symptoms increased. After three years, the photo-bleaching phenotype was completely absent, while HLB symptoms were fully pronounced. Subsequently, CTV titer was dramatically reduced while “Ca. L. asiaticus” titer increased. In addition, the gene expression of pds was increased indicating a deficit in the RNA interference. On the other hand, enhancing the nutrition of CTV-tPDS plants magnified the photo-bleaching phenotype and diminished the symptoms in HLB-infected plants. Taken together, combining enhanced nutritional programs with the use of CTV as a VIGS vector to target D. citri by citrus mediated RNAi may prolong its efficacy in the fight against HLB. The long-term success of CTV-based VIGS technology is contingent on the balance of power favoring the CTV, rather than the HLB bacterial pathogen, “Ca. L. asiaticus”.