Developing a culturomic approach to build a collection of grapevine foliar microorganisms with potential applications in disease biocontrol

Isolation and conservation of plant-associated microorganisms in pure cultures is important to be able to understand their influence on plant health. However, capturing the whole range of microbial taxa detected by culture-independent approaches remains labour-intensive and challenging since a large proportion fail to grow under laboratory conditions. Culturomic approaches that involve using a variety of culture conditions coupled with high-throughput identification methods are a recommended strategy to effectively capture plant-associated microbial diversity. A number of studies have demonstrated that using plant-based culture media rather than conventional laboratory media is an efficient strategy for increasing the cultivability of plant-associated microorganisms. This study aimed to develop a culturomic approach to build a collection of grapevine foliar microorganisms with potential applications for the biocontrol of foliar diseases. We developed and tested a custom grapevine leaf-based culture medium (GLA) to better mimic the leaf habitat and consequently try to increase the cultivability of foliar microorganisms. Of the 90 genera that were isolated, 17 were isolated exclusively on GLA including 4 bacteria, 1 yeast, and 12 filamentous fungi. Overall, using the GLA allowed for the isolation of a number of taxa that are of interest for their biocontrol potential against plant pathogens. Interestingly, it was also effective for isolating and speeding up the growth of slow-growing filamentous fungi associated with grapevine trunk diseases. Using a combination of laboratory media and the custom GLA resulted in a collection that was representative of 12.5 % of bacterial genera and 17.6 % of fungal genera detected on grapevine leaves. This collection will serve as a repository of pure cultures for conducting experiments. Candidates will be screened and tested both individually and in consortia for potential applications in microbial biocontrol against grapevine diseases, particularly Plasmopara viticola (grapevine downy mildew), with the ultimate goal of developing effective biocontrol products and moving towards more sustainable viticulture.