A VEM (Vector Enabled Metagenomic) approach to investigate the presence of potentially invasive geminiviruses in Southern Italy in the frame of GeMed, a research project involving five Mediterranean countries

Geminiviruses are monopartite or bipartite circular single-stranded DNA viruses, infecting major vegetable crops in the Mediterranean area, i.e., tomato and cucurbits. They are considered among the most worrying plant viruses due to their economic impact, the frequent introduction of exotic species, and their high propensity to recombine, continuously generating potentially invasive and resistance-breaking strains. Moreover, geminiviruses can be associated to different satellite molecules, i.e. betasatellites, further increasing their pathogenic potential. To prevent and control the spread of new and invasive geminiviruses (NIGs), and of betasatellites, a vector-enabled metagenomics (VEM) approach has been used to identify the population of NIGs and satellites circulating in the South of Italy. Insects present in the surveyed areas (mainly Bemisia spp., leafhopper spp., aphids spp., and mirids spp.) have been collected in Campania and Sicily regions, considered as two different agroecosystems, from 2020 to 2022. DNA was extracted from 15 different pools of insects, and circular DNA molecules were amplified by Rolling Circle Amplification (RCA). The RCA products were subjected to high-throughput generation sequencing (Illumina) and data were analysed using ad hoc bioinformatic pipelines, focusing on NIGs and betasatellites. To confirm the presence of virus/betasatellite in insects, validations by specific PCR were carried out. Due to its potential to detect new geminiviruses at an early invasion stage, this approach will help to develop prompt prevention and control measures. In the frame of the GeMed PRIMA Project, analogous VEM approaches have been carried out also in Tunisia and Morocco.