Ability of Euscelidius variegatus to transmit a phytoplasma with a short latency period

Phytoplasma transmission takes place by insect vectors through an Acquisition Access Period (AAP), Latency Period (LP) and Inoculation Access Period (IAP). Phytoplasmas are believed to be transmitted more efficiently by nymphs from the III instar, because they need a long LP to reach the salivary glands becoming infective. The transmission can start from adults as well, but in this case a long LP may exceed the insect lifespan. However, previous evidences indicated that adults can undergo a shorter LP, even though little knowledge is available regarding the phytoplasma temporal dynamics during this period. Here we investigated the minimum time required by the phytoplasma to colonize the vector midgut and salivary glands, and finally to be inoculated into a plant. We used the leafhopper Euscelidius variegatus to investigate the life cycle of flavescence dorée phytoplasma (FDP). Phytoplasma-free E. variegatus adults were left on broad beans (BB) infected by FDP for 7 days AAP. Subsequently, they were individually transferred onto a healthy BB for 7 different IAPs, each one lasting 24 hours from day 8 to 14. Molecular analyses and fluorescence in situ hybridization were performed for FDP detection. FDP was found in the leafhopper midgut since IAP1 with an infection rate reaching 50%, whereas in the salivary glands since IAP2 with and infection rate reaching 30%. FDP was also detected in BBs since IAP4, with infection rates reaching 10%. Our results represent an important step to further deepen the knowledge of phytoplasma transmission and its epidemiology.