Genetically-engineered citrus tristeza virus: establishing and examining its interaction with wild type CTV

Authors: Yokomi, Raymond - Ray; SELVARAJ, VIJAYANANDRAJ - Foreign Agricultural Service (FAS, USDA); MAHESHWARI, YOGITA - Foreign Agricultural Service (FAS, USDA); HAJERI, SUBHAS - Central California Tristeza Eradication Agency

Submitted to: Citrograph
Publication Type: Trade Journal
Publication Acceptance Date: 6/10/2020
Publication Date: 9/30/2020
Citation: Yokomi, R.K., Selvaraj, V., Maheshwari, Y., Hajeri, S. 2020. Genetically-engineered citrus tristeza virus: establishing and examining its interaction with wild type CTV. Citrograph. 10(4):66-69.

Interpretive Summary: A genetically-engineered Citrus tristeza virus (GE-CTV) from Florida with a T36 genotype (GE-T36) was developed in Florida to express foreign genes in citrus phloem to combat Huanglongbing (HLB), a devastating citrus disease. GE-T36 was introduced in a quarantine greenhouse in Parlier to study its biology and interaction when co-infected with wild-type California CTV strains. Vector transmission of GE-T36 by the cotton aphid occurred at a low level (0.86%). Reciprocal graft challenge experiments with GE-T36 with California CTV strains of T30, T36, RB, S1, and VT showed two important results. 1) GE-T36 successfully co-infected citrus with T30, S1 and VT genotypes but not T36. This was expected due to superinfection exclusion phenomenon of cross-protection that occurs between CTV isolates of the same strain but not between different strains. 2) Co-infection and expression of the GE-T36 was severely limited by the RB strain. These data suggests efficacy of GE-T36 to express genes to mitigate HLB will be limited by preexisting T36 and RB strains in California. Fortunately, T36 and RB genotypes are not widespread in central California, where majority of the citrus is grown. However, other GE-CTV genotypes or expression vectors should be developed to overcome cross protection if this proves to be a significant constraint on GE-T36 efficacy.

Technical Abstract: A prototype genetically-engineered Citrus tristeza virus (GE-CTV) from Florida with a T36 genotype (GE-T36) was introduced in a quarantine greenhouse in Parlier to develop protocols to study the biology of GE-CTV being developed in California. Four GE-T36 clones (500, 525, 527, 588), obtained from the Dawson Laboratory, Lake Alfred, FL, were successfully established in Citrus macrophylla (Alemow) and Madam Vinous sweet orange (C. sinensis). Vector transmission of the GE-CTV clones by the cotton aphid was shown to occur at a low level (0.86%). Since field trees are usually infected with two or more strains of CTV, to study the efficacy of GE-CTV under co-infection situation, Alemow seedlings were preinfected with California strains of T30, T36, RB, S1, and VT followed by challenge-inoculation with the GE-T36 clone 527. Two important outcomes were observed. 1) GE-T36-527 successfully co-infected with T30, S1 and VT genotypes but not T36. This was expected and can be explained by the superinfection exclusion phenomenon of cross-protection, which occurs between CTV isolates of the same strain but not between different strains. 2) Surprisingly, a significant limitation of co-infection and expression of the GE-T36 by the RB strain was observed which suggested that efficacy of a GE-CTV based on T36 genotype carrying genes to mitigate HLB will be affected by the preexisting T36 and RB infection. The good news is that the T36 and RB genotypes are relatively less widespread in central California groves, where majority of the citrus is grown. However, it would be a wise strategy to develop multiple California GE-CTVs to overcome cross protection and grove-specific application of GE-CTV based on the previous history of CTV in that grove.