bioRxiv2019
The reduction of synthetic fungicides in agriculture a major challenge in maintaining sustainable production, protecting the environment and consumers health. Downy mildew caused by the oomycete Plasmopora viticola is the major pathogen in viticulture worldwide and responsible for up to 60% of pesticide treatments. Alternatives to reduce fungicides are thus utterly needed to ensure sustainable vineyard-ecosystems, consumer health and public acceptance. Essential oils (EOs) are amongst the most promising natural plant protection alternatives and have shown their antibacterial, antiviral and antifungal properties on several agricultural crops. However, the efficiency of EOs highly depends on timing, application method and the molecular interactions between the host, the pathogen and EO. Despite proven EO efficiency, the underlying processes are still not understood and remain a black box. The objectives of the present study were: a) to evaluate whether a continuous fumigation of a particular EO can control downy mildew in order to circumvent the drawbacks of direct application, b) to decipher molecular mechanisms that could be triggered in the host and the pathogen by EO application and c) to try to differentiate whether essential oils directly repress the oomycete or act as plant resistance primers.\n\nA custom-made climatic chamber was used for a continuous fumigation of potted vines with different EOs during long-term experiments. The grapevine (Vitis vinifera) cv Chasselas was chosen in reason of its high susceptibility to Plasmopara viticola. Grapevine cuttings were infected with P. viticola. and subsequently exposed to continuous fumigation of different EOs at different concentrations, during 2 application time spans (24 hours and 10 days). Experiments were stopped when infection symptoms were clearly observed on the leaves of the control plants. Plant physiology (photosynthesis and growth rate parameters) were recorded and leaves were sampled at different time points for subsequent RNA extraction and transcriptomics analysis. Strikingly, the Oregano vulgare essential oil vapour treatment during 24h post-infection proved to be sufficient to reduce downy mildew development by 95%. Total RNA was extracted from leaves of 24h and 10d treatments and used for whole transcriptome shotgun sequencing (RNA-seq). Sequenced reads were then mapped onto the V. vinifera and P. viticola genomes. Less than 1% of reads could be mapped onto the P. viticola genome from treated samples, whereas up to 30 % reads from the controls mapped onto the P. viticola genome, thereby confirming the visual observation of P. viticola absence in the treated plants. On average, 80 % of reads could be mapped onto the V. vinifera genome for differential expression analysis, which yielded 4800 modulated genes. Transcriptomic data clearly showed that the treatment triggered the plants innate immune system with genes involved in salicylic, jasmonic acid and ethylene synthesis and signaling, activating Pathogenenesis-Related-proteins as well as phytoalexin synthesis.\n\nThese results elucidate EO-host-pathogen interactions for the first time and indicate that the antifungal efficiency of EO is mainly due to the triggering of resistance pathways inside the host plants. This is of major importance for the production and research on biopesticides, plant stimulation products and for resistance-breeding strategies\n\nAuthor SummaryThe reduction of synthetic plant protection products is a major concern of modern agriculture. The oomycete Plasmopora viticola which causes downy mildew in grapevine is amongst the most important grapevine pests and responsible for the dispersion of huge amounts of pesticides in vineyards. Among the evaluated alternatives to reduce or replace synthetic pesticides, plant volatile compounds could represent a sustainable solution. Some plant essential oils (EOs) have already shown antifungal capacities. However, their application is often difficult in terms of the right timing of treatment, degradation, bad rainfastness, mixability and phytotoxicity.\n\nThe aim of the present work was to investigate whether the vapour phase, applied by a continuous fumigation of different EOs, might inhibit the development of downy mildew on grapevine, and in case of proven efficiency, to study the induced transcriptomic changes by RNA-sequencing in an attempt to elucidate the underlaying molecular interactions. Our results showed that the vapour phases of O. vulgare and T. vulgaris were highly efficient against the pathogen. The study of differentially expressed genes indicated that the EO vapour triggered the main mechanisms of the plants innate immune system such as PTI (Pattern-Triggered Immunity) and ETI (Effector Triggered immunity).\n\nFor the first time these results highlight the effects of EOs vapour on plant genes expression, which is very valuable information for the development of new natural plant protection products, as well as for breeding disease resistant cultivars.