Separation and interference of strains from a citrus tristeza virus isolate evidenced by biological activity and double‐stranded RNA (dsRNA) analysis

Abstract

Separation of strains of citrus tristeza virus (CTV), differentiated by their double-stranded RNA (dsRNA) profiles, was obtained by graft-inoculating citron plants from a Mexican lime that had been recently aphid-or graft-inoculated with a mild CTV isolate (T-385). Up to 24 sub-isolates with differing dsRNA profiles were obtained from the aphid-inoculated lime. Some of these sub-isolates induced stronger symptoms in several citrus species than the original T-385 isolate. One sub-isolate, T-385-33, was mild in Mexican lime, but induced stem pitting on sweet orange. Inoculation of this isolate on Mexican lime, sour orange and Eureka lemon induced mild or no symptoms when inoculum was taken from citron, but very severe symptoms when the inoculum was from sweet orange. Mexican lime and sweet orange plants co-inoculated with T-385-33 from sweet orange in combination with the other 23 sub-isolates showed mild symptoms. The results obtained suggest that there is natural cross-protection among sub-isolates in the original T-385 isolate. RESULTS dsRNA profilesA mild CTV isolate from the IVIA collection, T-385 (Moreno etal., 1991), was graft-inoculated to two citron plants, which were kept in the greenhouse. When these plants were periodically analysed for dsRNA content, variations in the electrophoretic profiles could be observed (Fig. 1).The original T-385 field isolate was aphidtransmitted to a Mexican lime plant and 2 months later, when the lime first reacted positively io CTV by ELISA, was graft-inoculated to 40 citron plants. When these citron plants were individually analysed 2 years after inoculation, up to 24 different dsRNA profiles could be identified (Fig, Plant Pathology 42,(48)(49)(50)(51)(52)(53)(54)(55)(56)(57)

Regeneration of transgenic citrus plants under non selective conditions results in high-frequency recovery of plants with silenced transgenes

“…The full-length p25 coat protein ORF from the strain T-305 of CTV (Moreno et al 1993) was reverse transcribed and amplified by PCR (RT-PCR) with the primers 5¢-CGCTAGAGGATCCA-CCATGGACTAGTAAACAAAG-3¢ and 5¢-CCGACGGGATC-CTCAACGTGTGTTGAATTTCC-3¢, both of which carry BamHI sites (indicated in italics) at their 5¢ ends, and, after digestion with BamHI, the cDNA was cloned into pMOG 180 (Mogen International). The p25 sequence was modified by introducing two consecutive stop codons 3 nt downstream of the start codon (underlined in the primer sequence) to make its transcripts untranslatable.…”

Estudio de la interacción entre naranjo amargo y el virus de la tristeza de los cítricos

“…El proceso de decaimiento puede durar varios años, sin embargo, en ocasiones se observa un síndrome de colapso que acaba con la vida del árbol en pocas semanas (Ben Ze'ev et al, 1989). La mayoría de los aislados de tristeza causan decaimiento de las combinaciones sobre NA, si bien se han descrito algunos que son asintomáticos sobre esta combinación (Bové et al, 1988;Albertini et al, 1988;Moreno et al, 1991, Albiach-Martí, et al, 2000b.…”

Molecular Virology and Pathogenicity of Citrus tristeza virus

“…These could vary in the pathogenicity induced depending on the citrus host and in transmission efficiency by aphids Moreno et al, 2008). The strains composing a field population are unequally distributed within a CTV infected tree (d'Urso et al, 2000) and could be somehow separated from the former mixture by aphid or graft transmission or by host passage creating a new mixture of strains, thus a new isolate, which could generate a completely distinct symptomatology in citrus plants (Albiach et al, 2000a;Ayllon et al, 1999b;Moreno et al, 1993;Roy & Brlansky, 2009;Velazquez-Monreal et al, 2009;Weng et al, 2007). Additionally, aphid or graft transmission and host passage could modify the composition of the D-RNAs population in the CTV isolates (Albiach-Martí et al, 2000a;Mawassi et al, 1995b) The sequencing of the genomes of nineteen CTV isolates from distant places in the planet, which represented a subset of its local CTV population, helped the understanding of the evolution of CTV and the complex structure of the actual CTV isolates.…”

“…Wider application of natural mild strain cross-protection has been limited by difficulty in finding mild isolates of CTV that effectively protect against SP and QD pathotypes (Bar-Joseph et al, 1989). Another problem is that natural mild CTV isolates may contain minor severe stem pitting variants that, upon aphid transmission, could become prevalent (Moreno et al, 1993;Velazquez-Monreal et al, 2009). Since only isolates within a closely related sequence group will cross-protect , naturally occurring mild T30-like isolates (Albiach-Martí et al, 2000b), would not protect against disease inducing isolates from other genotypes.…”