The main objective of the project is the development and application of management strategies using rhizospheric microorganisms and useful entomofauna to design sustainable, multi-productive black truffle (Tuber melanosporum) agrosystems. The implementation of these techniques (inoculations of microorganisms, establishment of refuge plants for useful insects, and mixed production with aromatic plants) will be based on the knowledge of the ecology of truffle-producing systems, including their useful fauna (pollinators and natural enemies of pests), the interactions of the fungus with the companion plants, and the dynamics of the associated microbial communities. As a general hypothesis, we propose that sustainable multiproductive systems, based on the know-how of the interactions between macro and microorganisms, will provide an improvement of the ecosystem services related to truffle farming. The TUBERSYSTEMS project is structured in two complementary sub-projects with three general objectives: i) the study of the diversity of macro and micro-organisms and their interactions in truffle producing systems, ii) the selection and management of these organisms for the establishment of multiproductive truffle plantations with aromatic plants, and iii) the analysis of the ecosystem services associated with different truffle producing systems. A network of plantations and wild production forests has been established within the distribution area of the black truffle in Spain to evaluate: i) the diversity of plants forming arbuscular mycorrhizae (AM) located in the burnt areas around the trees (brûlés), ii) the taxonomic and functional diversity of the rhizospheric microbiome (fungi and bacteria), iii) the dynamics of extraradical black truffle mycelium, and iv) the presence of useful insects (pollinators and natural enemies of pests). Initially, the taxonomic and functional diversity of the microorganisms present in the brûlés, and their interaction with T. melanosporum, will be analyzed. Isolations of potential plant growth promoting bacteria (PGPR) will be carried out from truffle production areas, and their effect on the development of Tuber mycorrhizal plants and the persistence of the fungal extraradical mycelium in the soil will be evaluated under controlled mesocosm conditions, and in plantations. In parallel, isolations of the AM fungi associated with plants growing in the brûlés will be obtained and inocula formulated to produce mycorrhizal aromatic plants (lavender, thyme and sage). Mixed plantations using aromatic plants will be established in selected truffle-producing plots and their effect on the extraradical mycelium biomass of T. melanosporum, as well as the presence of this fungus as an endophyte of alternative hosts, will be evaluated. The incidence of Kermes spp. (Hemiptera) and phytoplasmas, and its relation with the trunk tumors syndrome causing serious damage in truffle plantations, will be studied in both monospecific and mixed truffle plantations. Finally, the ecosystem services (productivity, agrosystem regulation and sustainability, landscape, cultural and sociological) provided by the monospecific and multiproductive truffle production systems will be jointly analyzed to generate knowledge-based management guidelines to be applied in sustainable truffle culture.