Variable Membrane Protein A of Flavescence Dorée Phytoplasma Binds the Midgut Perimicrovillar Membrane of Euscelidius variegatus and Promotes Adhesion to Its Epithelial Cells

Abstract

Phytoplasmas are uncultivated plant pathogens and cell wall-less bacteria and are transmitted from plant to plant by hemipteran insects. Phytoplasmas' circulative propagative cycle in insects requires the crossing of the midgut and salivary glands, and primary adhesion to cells is an initial step towards the invasion process. The flavescence dorée phytoplasma possesses a set of variable membrane proteins (Vmps) exposed to its surface, and this pathogen is suspected to interact with insect cells. The results showed that VmpA is expressed by the flavescence dorée phytoplasma present in the midgut and salivary glands. Phytoplasmas cannot be cultivated at present, and no mutant can be produced to investigate the putative role of Vmps in the adhesion of phytoplasma to insect cells. To overcome this difficulty, we engineered the mutant G/6, which lacks the adhesins ScARPs, for VmpA expression and used VmpA-coated fluorescent beads to determine if VmpA acts as an adhesin in adhesion assays and ingestion assays. VmpA specifically interacted with insect cells in culture and promoted the retention of VmpA-coated beads to the midgut of In this latest case, VmpA-coated fluorescent beads were localized and embedded in the perimicrovillar membrane of the insect midgut. Thus, VmpA functions as an adhesin that could be essential in the colonization of the insect by the FD phytoplasmas. Phytoplasmas infect a wide variety of plants, ranging from wild plants to cultivated species, and are transmitted by different leafhoppers, planthoppers and psyllids. The specificity of the phytoplasma-insect vector interaction has a major impact on the phytoplasma plant host range. As entry into insect cells is an obligate process for phytoplasma transmission, the bacterial adhesion to insect cells is a key step. Thus, studying surface-exposed proteins of phytoplasma will help to identify the adhesins implicated in the specific recognition of insect vectors. In this study, it is shown that the membrane protein VmpA of the flavescence dorée phytoplasma acts as an adhesin that is able to interact with cells of , the experimental vector of the FD phytoplasma.

When a Palearctic bacterium meets a Nearctic insect vector: Genetic and ecological insights into the emergence of the grapevine Flavescence dorée epidemics in Europe

“…protein analogous to Vmp1, we could demonstrate that VmpA acts as an adhesin able to bind to insect vector cells in culture and to its midgut perimicrovillar membrane [66,67]. We show here that sequence variations of vmpA and vmpB genes correlate with transmission by insect vectors of different leafhopper subfamilies.…”

“…In the absence of genetic engineering of the uncultivated phytoplasmas, we previously used protein-coated latex beads or recombinant spiroplasmas to mimic phytoplasma surface, and demonstrate that VmpA acts as an adhesin binding to the cells of E. variegatus [66]. Here we found that latex beads had enhanced adhesion to E. variegatus cells in culture and were better retained in E. variegatus and S. titanus midguts, when coated with increasing ratio of FD92 VmpA (cluster II) over PGYA VmpA (cluster I).…”

Evidence suggesting interactions between immunodominant membrane protein Imp of Flavescence dorée phytoplasma and protein extracts from distantly related insect species

“…environmental constraints and biological relationships; Daugherty et al ; Lopes et al ) and affect pathogen‐vector interactions at different spatio‐temporal scales and biological organization levels. Some studies suggested the involvement of specific molecular phytoplasma‐ligand interactions in transmission specificity with insect hosts (Suzuki et al ; Galetto et al ; Arricau‐Bouvery et al ). Different types of phytoplasma membrane proteins (MP) are in direct contact with the host environment and have been recognized to play an important role in promoting phytoplasma internalization in insect cells.…”

Potential role of the alien planthopper Ricania speculum as vector of Flavescence dorée phytoplasma

“…P. asteris' amp protein mediates both pathogen crossing of vector gut epithelium and colonization of salivary glands (Rashidi et al 2015), showing the real involvement of these specific molecular interactions in a compatible phytoplasma/vector combination. A similar specific recognition mechanism was recently demonstrated also for FDp and its vector E. variegatus: the variable membrane protein A (vmpA) expressed on the surface of pathogen cells mediates phytoplasma binding to insect cell culture and pathogen retention at the perimicrovillar membrane of vector midgut epithelium (Arricau-Bouvery et al 2018). Lack of such molecular recognition between R. speculum tissues and FDp membrane proteins could explain the low multiplication rate in this host, lack of salivary gland colonization and the consequent inability of transmission.…”