Evidence for specific interaction between immunodominant membrane protein Imp of Flavescence dorée phytoplasma and insect proteins: preliminary results

Phytoplasmas are phloem-limited plant pathogenic bacteria in the class Mollicutes, family Acheloplasmataceae. Phytoplasma classification is based on 16S rRNA and ribosomal protein (rp) gene sequences and the 'Candidatus' concept is applied for well characterized phytoplasmas. Once acquired by an insect, phytoplasma must cross its midgut membrane, spread in the hemolymph, migrate to the salivary glands and enter the saliva in order to be transmitted to another plant. Previous studies suggested the involvement of different types of membrane proteins of phytoplasmas in promoting phytoplasma internalization in the host cell. In the present study, the involvement of immunodominant membrane protein Imp of Flavescence dorèe type D phytoplasma (FD-Dp) in the interaction with insect proteins was investigated. Six Auchenorrhyncha species known as vectors, potential, and non- vectors of FD-Dp were selected from distantly related species groups. Four species of the infraorder Cicadomorpha, all in the family Cicadellidae. The dnaD-imp-pyrG genomic fragment of FD-D strain was amplified and sequenced from FD-D infected vine leaves. A total of 162 amino acids were predicted for full-length FD-D Imp protein, with residues 44-162 exposed outside the membrane. A His-tagged partial Imp protein and a rabbit polyclonal antibody were synthesized and used for Western and Far-Western dot blots (FWdB) experiments. Total native proteins were extracted from batches of each insect species. FWdB showed interaction of Imp fusion protein with total proteins of St, Ev (known efficient vectors) and Mq (potential vector), while no interaction signal was detected with the other three species, Rs, Zp, and Mp (non-vectors).