Fungal endophytes, both mycorrhizal and non-mycorrhizal, are involved in the development of the life cycle of orchids, providing potential beneficial relationships. Here, we assess the succession of changes in the diversity of fungal symbionts associated with a terrestrial temperate orchid species, Anacamptis morio subsp. champagneuxii, over three phenological stages: developed leaves but no stem elongation, flowering, and fruiting. Fungi endophyte associated with roots were obtained by culture in sterile conditions. A total of 18 morphotypes-one Mortierellomycota, two Basidiomycota and 15 Ascomycota-were differentiated, and were also characterized using PCR and DNA sequencing techniques. Only three of the 18 OTUs are shared among the three phenological stages examined: Westerdykella sp., a member of Ceratobasidiaceae, and Fusarium oxysporum, representing a relative abundance of between 28% (fruiting) to 41% (flowering). Our research confirmed that fungal symbionts varied among the different phenological stages examined, the peak of endophyte diversity appearing in the flowering stage. The availability of a diverse mycobiota seems to be important for the survival of orchid plants because it may cover particular physiological needs, and knowledge concerning this mycobiota is of special relevance in the establishment of reliable conservation programmes.
Keywords: Anacamptis; ITS; Mycorrhizal fungi; Orchid endophytes; Phenological stages; Symbiosis.