Mycorrhizal divergence in Lepanthes

Mycorrhizal fungi form essential mutualisms with the majority of land plants, yet their role on plant community composition and diversification remains poorly understood. Orchids provide a model system for studying these interactions because the orchid life cycle obligatorily depends on mycorrhizae. Historically, studies have emphasized the role of niche partitioning and competition avoidance resulting in distinct mycobiome compositions among coexisting orchids. Since closely related orchid species have been found to associate with similar groups of fungi, it has been speculated that different fungal species are needed for coexistence but not for speciation. However, fungi have often been examined at lower resolution levels (i.e., class, order, family, or genus). Using third-generation long-read sequencing (PacBio) of the full ITS region, we characterized fungal communities at fine scale with high accuracy. We evaluated alpha- and beta-diversity of fungal communities in four closely related, narrowly endemic epiphytic orchid species from the rapidly diversifying genus Lepanthes in one of the world’s richest biodiversity hotspots. Our analyses reveal that orchid species have distinct mycobiont assemblages, with differences in composition unevenly distributed across species. These results suggest that shifts in fungal partners may contribute to speciation and rapid diversification in Lepanthes. This study highlights the potential evolutionary role of mycorrhizal fungi in orchid diversification and demonstrates the value of high-resolution sequencing in uncovering cryptic fungal diversity.