Virome Analysis of Lemon Plants with Vein Clearing Symptoms Reveals Mixed Infection of Citrus Vein Clearing Virus, Iris Domestica Betaflexivirus 1 and Hop Stunt Viroid

Citrus yellow vein clearing virus (CYVCV) is the causative agent of the yellow vein clearing disease (YVCD), a worldwide and highly destructive disease in lemon (<i>Citrus lemon</i>) and sour orange trees (<i>C. aurantium</i>). The typical symptoms of vein clearing are believed to be associated with CYVCV infection in citrus, so virus-specific diagnostic systems are currently used to confirm infection. In the present study, virome analysis based on high-throughput sequencing (HTS) on a lemon plant showing YVCD revealed mixed infection of CYVCV, iris domestica betaflexyviridae 1 (IDBV), and hop stunt viroid (HSVd). This multiple infection was confirmed in other two lemon plants with similar symptoms using virus/viroid specific primers. This is the first report of IDBV in lemon. Through molecular characterization and the reconstruction of phylogenetic relationships, a possible origin of the viruses/viroid identified in lemon has been hypothesized. Such mixed infections raise new questions about their role in the expression of YVCD symptoms observed on lemon.