High Throughput-based Surveillance Reveals New Components of Spanish Citrus Virome Related to Tristeza, Impietratura and Yellow Vein Clearing Diseases

Citrus is one of the most important crops cultivated worldwide, representing a strategic source of agricultural income for many countries, including Spain, the main producer within the European Union. The protection of this highly valuable industry against an increasing global movement of pests and pathogens requires effective regulatory measures, including control of plant propagation material, phytosanitary surveillance and risk assessment, which are based not only on knowledge of the established threats but also on potential emerging threats affecting citrus that may circulate unnoticed in the production system. In this work, with the aim of generating knowledge on potential emerging viruses in Spanish citrus orchards, high-throughput sequencing (HTS) analysis has been applied to monitor the sanitary status of several growing areas of one of the main citrus producer regions in Spain, the Valencian Community. The results of this study have revealed a much more complex citrus virome than previously reported, including citrus yellow vein clearing virus (CYVCV), a non-regulated but harmful citrus virus, as well as the T3 genotype of citrus tristeza virus (CTV) and citrus virus A (CiVA), not detected to date in Spain. Moreover, our results indicate the existence of other unknown components of the citrus virome. HTS detection of CYVCV, CTV T3 and CiVA and their presence in Spanish orchards has been confirmed by RT-PCR and Sanger sequencing. These findings have relevant implications in the development of control and regulatory measures against three important viral diseases, tristeza, impietratura and yellow vein clearing diseases, and demonstrate the added value of HTS-based surveillance to discover emerging components of the citrus virome.