Abstract
A transient expression vector based on Citrus tristeza virus (CTV) is unusually stable. Because of its stability it is being considered for use in the field to control Huanglongbing (HLB), which is caused by Candidatus Liberibacter asiaticus (CLas) and vectored by Asian citrus psyllid, Diaphorina citri. In the absence of effective control strategies for CLas, emphasis has been on control of D. citri. Coincident cohabitation in phloem tissue by CLas, D. citri and CTV was exploited to develop a novel method to mitigate HLB through RNA interference (RNAi). Since CTV has three RNA silencing suppressors, it was not known if CTV-based vector could induce RNAi in citrus. Yet, expression of sequences targeting citrus phytoene desaturase gene by CTV-RNAi resulted in photo-bleaching phenotype. CTV-RNAi vector, engineered with truncated abnormal wing disc (Awd) gene of D. citri, induced altered Awd expression when silencing triggers ingested by feeding D. citri nymphs. Decreased Awd in nymphs resulted in malformed-wing phenotype in adults and increased adult mortality. This impaired ability of D. citri to fly would potentially limit the successful vectoring of CLas bacteria between citrus trees in the grove. CTV-RNAi vector would be relevant for fast-track screening of candidate sequences for RNAi-mediated pest control.
Highlights
Virus-based vectors for expressing foreign genes in plants are routine laboratory tools (Porta and Lomonossoff, 2002; Gleba et al, 2007), generally developed for short term laboratory experiments in herbaceous plants or for making specialty products in these plants
To demonstrate the gene silencing capabilities of Citrus tristeza virus (CTV), transgene green fluorescent protein (GFP) of N. benthamiana line 16c was silenced by CTVVIGS vector carrying truncated GFP
N. benthamiana plants were inoculated with a binary plasmid vector carrying CTV-truncated GFP (tGFP) through agro-infiltration of fully expanded true leaves
Summary
Virus-based vectors for expressing foreign genes in plants are routine laboratory tools (Porta and Lomonossoff, 2002; Gleba et al, 2007), generally developed for short term laboratory experiments in herbaceous plants or for making specialty products in these plants. The rapid spread of devastating disease of citrus, citrus greening known as huanglongbing (HLB), which is threatening the survival of the citrus industry has changed the Citrus tristeza virus (CTV) vector from a laboratory tool to a potential management strategy for citrus groves (Hodges and Spreen, 2012; National Research Council, 2010). At this time, the one of the hopes for survival of the Florida citrus industry is the production of resistant or tolerant trees via transgene (http://www.nytimes.com/2013/07/28/science/ a-race-to-save-the-orange-by-altering-its-dna.html?pagewanted =all& r=0). The viral vector can be deployed more quickly and is being considered as an interim approach (National Research Council, 2010)
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