Detection of a new variant of Citrus tristeza virus in Greek citrus crops

Abstract

Citrus tristeza virus (CTV), the most destructive virus of citrus, is a quarantine pathogen in Greece. Since 2000, several accidental imports of infected propagation material have been detected in the country, and while eradication measures were applied, a few disease foci still remain. CTV isolates were collected from Chania (Crete) and the “lemonwood” of Poros (Peloponnese), and their genetic variability was studied using single-strand conformation polymorphism (SSCP). One previously characterized isolate from Argolida grafted on a Mexican lime (GR3) and two Italian isolates from Calamondin were also included in the study. ELISA and RT-PCR tests confirmed CTV presence, and SSCP analysis of the virus amplified coat protein (CP) gene was used to separate either distinct virus isolates for cloning the CP gene or variants (haplotypes) for sequencing. Analyses showed that selected variants of four representative isolates clustered into three of the seven defined phylogenetic groups: groups 3b and 5 (severe isolates) and group M (mild isolates). The prevalent haplotypes detected in the CTV from lemonwood of Poros (GR9) were in group 3b, confirming previous results. However, one sequence variant was identified as a recombinant between haplotypes from groups 3b and 5. Variants of these two groups were also detected in the Italian Calamondin isolate. In the grafted Mexican lime isolate (GR3) from Argolida, only one haplotype was found which belonged to group M, while in the field isolate from Chania (GR6) the only haplotype detected was in group 5. This is the first report of variants of group 5 in Greece, suggesting an unknown virus introduction. The prevalence of severe isolates in the area is of particular concern, and implications for the future of the CTV epidemics are discussed.