Green, variegated, and albino Cremastra variabilis provide insight into mycoheterotrophic evolution associated with wood-decaying fungi.

Abstract

With approximately 31,000 species, orchids begin life as mycoheterotrophs, relying on fungi to meet their carbon demands. Notably, some green orchids retain the ability to acquire carbon through fungal associations (partial mycoheterotrophy) and occasionally produce albino or, more rarely, variegated phenotypes. A linear relationship has been observed between leaf chlorophyll content and dependence on fungal-derived carbon, particularly in orchids associated with ectomycorrhizal (ECM) fungi, but whether such plasticity is similarly robust among orchids associated with non-ECM fungi remains underexplored. Here, we focused on the green, variegated, and albino forms of Cremastra variabilis, which likely lack ECM associations, to investigate (i) whether the degree of mycoheterotrophy, indicated by ¹³C enrichment, correlates with chlorophyll content, and (ii) whether nutritional shifts align with changes in plant structure and mycorrhizal communities. Our results show that rhizoctonia fungi were dominant in green individuals with high chlorophyll levels and lacking coralloid rhizomes, whereas albino and most variegated individuals possessing coralloid rhizomes primarily associate with Psathyrellaceae fungi. Chlorophyll content and carbon stable isotope abundances were negatively correlated, indicating a gradient of increasing mycoheterotrophy from green to albino forms in individuals with coralloid rhizomes. In conclusion, C. variabilis maintains a flexible balance between photosynthesis and mycoheterotrophy, likely shaped by its subterranean morphology and fungal associations, with wood-decaying Psathyrellaceae fungi providing greater support for mycoheterotrophic nutrition than rhizoctonia fungi.