Virus biology of Euscelidius variegatus iflavirus 1: towards the production of an infectious viral clone

Euscelidius variegatus Kirschbaum (Hemiptera Cicadellidae) is a well-known leafhopper vector of phytoplasmas. A new member of a new species of the genus Iflavirus was discovered during transcriptomic analyses of a laboratory population of E. variegatus, and named E. variegatus virus 1 (EVV1). EVV1 infection was asymptomatic, and the virus was constantly detected both in phytoplasma exposed and not exposed vectors. Interestingly, virus load was significantly lower in the former category. Two other E. variegatus populations from France and US were found to be virus free (EVV1-), and one virus free population was therefore established in our lab starting from the French one. EVV1 distribution was assessed by PCR on total RNAs extracted from dissected organs and insects at different life stages. The virus was present in all tested organs and life stages, with different loads. Vertical transmission through the ovary was confirmed. EVV1 was also detected in the honeydew of infected individuals, but viral transmission could not be obtained by feeding on artificial medium containing virus-contaminated honeydew. Horizontal transmission through co-feeding of EVV1-infected and healthy insects, resulted in low infection rates. Plants exposed to infected insects could be contaminated on the plant surface, but were generally virusfree, therefore horizontal transmission through the plant was, at best, extremely inefficient. An infectious clone was constructed and its ability to infect and replicate in virus-free insects was demonstrated. The application of the infectious clone for virus-induced gene silencing to interfere with insect ability to transmit the phytoplasma is ongoing.