Visualization of Mycorrhizal Fungi in Orchid Protocorms

The symbiotic relationship between orchids and mycorrhizal fungi is valuable for studying plant–fungal interactions and symbiosis mechanisms. Various staining methods, including trypan blue and acidified blue ink, have been used and compared for their effectiveness and safety. Among these methods, acidified blue ink staining is proposed as a safer alternative to trypan blue, demonstrating its efficacy in visualizing fungal structures within orchid protocorms. The methodological details for conducting the acidified blue ink staining and considerations for sample preparation and microscopy examination are provided. This staining technique offers a simple, efficient, and cost-effective means of detecting fungal infection in orchid protocorms. We also include a concise protocol using the glycol methacrylate (Technovit 7100) embedding method for detailed examinations of the infection process.