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Flavescence dorée is an economically important vector-borne disease of grapevine in Europe caused by phytoplasmas belonging to the 16SrV ribosomal group. Expression profiles of 11 genes of the Flavescence dorée phytoplasma (FDp) were analysed over time following infection of natural (Vitis viniferaand the leafhopper vector Scaphoideus titanus) and experimental (Vicia fabaand Euscelidius variegatus) hosts. Infected and symptomatic grapevine plants (Chardonnay) were sampled under natural field conditions in a productive vineyard in north-western Italy. Broad bean samples were assayed after the experimental inoculation with infectious E. variegatus. Adults of both vector species were analyzed following FDp acquisition from infected broad bean plants. The selected FDp genes were grouped according to their putative functions within different categories, namely ‘Membrane proteins’ (imp), ‘Regulative elements’ (spoVG, rpoD), ‘Protein metabolism, transport and secretion’ (tldD, ysdC, ftsY), ‘Stress response’ (comp83, osmC), ‘ABC transporters’ (CoABC) and ‘Unknown’ function (contig12, comp115). All analyzed genes were expressed in the four different host species suggesting their crucial role during the FDp infection cycle. Moreover, some of them (contig12, CoABC, comp83, and imp) might be considered essential for phytoplasma survival irrespective of the host, while comp115 seems to be required for insect infection. We showed that FDp is metabolically more active in insects than in plants, at least according to the pool of selected genes for this study, indicating that FDp behaves differently in the two hosts compared to other phytoplasma species/strains.

The leafhopper Euscelidius variegatus Kirschbaum (Hemiptera: Cicadellidae) is a natural vector of the ‘Candidatus Phytoplasma asteris’ and a laboratory vector of the phytoplasma associated with grapevine Flavescence dorée. Previous studies indicated that RNA interference (RNAi) by dsRNA injection efficiently works in E. variegatus and that silencing of ATP synthase β increases mortality and impairs phytoplasma multiplication. Here, a nearly complete female sterility was observed when ATP synthase β was silenced. The sterility was associated with the absence of mature eggs in the ovaries. On the contrary, male genitalia morphology and sperm motility were similar to those observed in E. variegatus control specimens, treated with dsRNAs targeting green fluorescent protein (dsGFP). An over-expression of hexamerin (amino acid storage protein) and cathepsin L (lysosome proteinase) was observed at transcript and protein level in dsATP-injected females in comparison with dsGFP-injected insects, in both whole body and dissected ovary samples. Conversely, the expression of other genes known to be involved in oocyte development, namely vitellogenin, perilipin and digestive cystein protein, was not altered in dsATP-injected E. variegatus insects in comparison with dsGFP ones. Possible roles of ATP synthase β, hexamerin and cathepsin L in oocyte and egg development are discussed. Insecticide treatments against vectors are the main strategies to counteract phytoplasma diseases, with negative impact on environment and public health. RNAi is a promising sustainable approach against insect vectors and ATP synthase β could be a valid target gene, as its silencing, besides increasing mortality and reducing phytoplasma multiplication, induces female sterility.

cDNA arrays were used to explore mechanisms controlling fruiting body development in the truffle Tuber borchii. Differences in gene expression were higher between reproductive and vegetative stage than between two stages of fruiting body maturation. We suggest hypotheses about the importance of various physiological processes during the development of fruiting bodies.

ABSTRACTPhytoplasmas are plant-pathogenic bacteria transmitted by hemipteran insects. The leafhopper Euscelidius variegatusis a natural vector of chrysanthemum yellows phytoplasma (CYp) and a laboratory vector of flavescence dorée phytoplasma (FDp). The two phytoplasmas induce different effects on this species: CYp slightly improves whereas FDp negatively affects insect fitness. To investigate the molecular bases of these different responses, transcriptome sequencing (RNA-seq) analysis of E. variegatusinfected with either CYp or FDp was performed. The sequencing provided the first de novotranscriptome assembly for a phytoplasma vector and a starting point for further analyses on differentially regulated genes, mainly related to immune system and energy metabolism. Insect phenoloxidase activity, immunocompetence, and body pigmentation were measured to investigate the immune response, while respiration and movement rates were quantified to confirm the effects on energy metabolism. The activation of the insect immune response upon infection with FDp, which is not naturally transmitted by E. variegatus, confirmed that this bacterium is mostly perceived as a potential pathogen. Conversely, the acquisition of CYp, which is naturally transmitted by E. variegatus, seems to increase the insect fitness by inducing a prompt response to stress. This long-term relationship is likely to improve survival and dispersal of the infected insect, thus enhancing the opportunity of phytoplasma transmission.