settingsOrder Article Reprints Open AccessArticle The Tripartite Lichen Ricasolia virens: Involvement of Cyanobacteria and Bacteria in Its Morphogenesis by Francisco J. García-Breijo 1ORCID,Arantzazu Molins 2,3,José Reig-Armiñana 2ORCID andEva Barreno 2,*ORCID 1 Departamento de Ecosistemas Agroforestales, ETSIAMN, Universitat Politècnica de València, Camino de Vera s/n, 46022 València, Spain 2 Instituto Cavanilles de Biodiversidad y Biología Evolutiva (ICBiBE), Botánica, Universitat de València, C/Dr. Moliner, 50, 46100 Burjassot, Spain 3 Instituto de Investigaciones Agroambientales y de Economía del Agua (INAGEA), Departamento de Biología, Universitat de les Illes Balears (UIB), Ctra. Valldemossa Km.7., 07122 Palma de Malllorca, Spain * Author to whom correspondence should be addressed. Microorganisms 2023, 11(6), 1517; https://doi.org/10.3390/microorganisms11061517 Received: 1 May 2023 / Revised: 31 May 2023 / Accepted: 5 June 2023 / Published: 7 June 2023 (This article belongs to the Special Issue 10th Anniversary of Microorganisms: Past, Present and Future) Download Browse Figures Review Reports Versions Notes Abstract Ricasolia virens is an epiphytic lichen-forming fungus mainly distributed in Western Europe and Macaronesia in well-structured forests with ecological continuity that lack eutrophication. It is considered to be threatened or extinct in many territories in Europe (IUCN). Despite its biological and ecological relevance, studies on this taxon are scarce. The thalli are tripartite, and the mycobiont has a simultaneous symbiotic relationship with cyanobacteria and green microalgae, which represent interesting models to analyse the strategies and adaptations resulting from the interactions of lichen symbionts. The present study was designed to contribute to a better understanding of this taxon, which has shown a clear decline over the last century. The symbionts were identified by molecular analysis. The phycobiont is Symbiochloris reticulata, and the cyanobionts (Nostoc) are embedded in internal cephalodia. Light, transmission electron and low-temperature scanning microscopy techniques were used to investigate the thallus anatomy, ultrastructure of microalgae and ontogeny of pycnidia and cephalodia. The thalli are very similar to its closest relative, Ricasolia quercizans. The cellular ultrastructure of S. reticulata by TEM is provided. Non-photosynthetic bacteria located outside the upper cortex are introduced through migratory channels into the subcortical zone by the splitting of fungal hyphae. Cephalodia were very abundant, but never as external photosymbiodemes. Keywords: cephalodia; endangered lichens; non-photosynthetical bacteria; Nostoc; Symbiochloris; TEM; tripartite lichen.