Exploring the orchid mycorrhizal symbiosis in the context of conservation: identity, environmental availability, and ontogenesis

Abstract

Orchids make up one of the most diverse plant families globally, and have been valued for their medicinal and ornamental properties for millennia. Despite this, they are increasingly threatened by the actions of humans: from collection, to land clearing, to climate change, we are consistently the biggest threat to global orchid biota. We also represent their most important protectors, with techniques like ex situ germination enabling us to recover species from the brink of extinction. For orchids, conservation relies on an understanding of the mycorrhizal fungi that provide carbohydrates during germination and seedling development; this relationship needs to be recreated to grow seeds ex situ.

Despite their importance, until now mycorrhizal fungi have not been identified for the endangered wetland orchid Corybas carsei. In Chapter 2, fungal associates of C. carsei were identified through both culturing and next-generation sequencing (NGS) approaches. In addition, the fungal associates of three co-occurring wetland orchids were identified through an NGS approach. Culturing and NGS identified different fungal associates for C. carsei, with the most abundant taxa identified through sequencing absent in culturing. The most abundant taxa were identified as members of the genus Basidiodendron (Exidiaceae) and in the order Auriculariales, and do not belong to known orchid mycorrhizal groups. To contrast, the other three wetland orchids associated with the known orchid mycorrhizal family, Ceratobasidiaceae. Fungal associates differed between orchid species and may represent a form of niche partitioning.

The availability of symbiotic fungi in soil can drive recruitment and distribution of orchids. Management of C. carsei through controlled burning has been shown to cause favourable shifts in aboveground (plant) communities but had not been assessed for belowground (fungal) communities. In Chapter 3, shifts in soil fungal communities, in both the short-term (1 month) and long-term (2 years), were detected through an NGS approach. Specific fungal taxa increased in abundance in post-burn treatments, and partly drove observed community shifts. With one primer pair, an amplicon sequencing variant (ASV) identified as Basidiodendron sp. was significantly increased in long-term burn treatments and was found to drive over 3% of community dissimilarity. Orchid mycorrhizal families were generally in low abundance in soils (fewer than 2% of reads), while the potentially mycorrhizal families Exidiaceae and Sebacinaceae were detected in less than half of all soil samples. However, Exidiaceae had a significantly higher abundance in long-term burn soils than in control soils. Controlled burning may thus favour both C. carsei and its fungal partners in the genus Basidiodendron.

Understanding orchid mycorrhizal associations enables the exploitation of mycorrhizal associations, by using fungi to inoculate and promote germination of seed ex situ. This can mark the switch from maintaining existing populations, to generating plants to enhance existing populations or establish new populations. In Chapter 4, I made a preliminary assessment of the fungal associates and germination preferences of several New Zealand orchids. Cultured fungi were diverse and likely non-mycorrhizal, except for those from Corybas oblongus. These were all Tulasnella spp., a known mycorrhizal genus. Interestingly, fungi cultured from Thelymitra longifolia protocorms differed from those identified by directly extracting DNA: culturing identified non-mycorrhizal fungi, while direct extraction identified the genus Tulasnella. This allowed, briefly, a discussion on the potential for mycorrhizal ‘bottlenecks’ and how these may impact germination strategies. I also found that germination preferences varied across species: C. oblongus and Pterosytlis cardiostigma developed further in symbiotic conditions while T. longifolia developed further in asymbiotic conditions. Leaf primordia of T. longifolia developed within 3 months on Modified Knudson C media. These results demonstrate the species-specific responses to different conservation approaches and highlight the need for targeted conservation strategies.

The work presented in this thesis can be used to guide ongoing conservation of C. carsei, and New Zealand terrestrial orchids more generally. In particular, techniques focused on supporting orchid recruitment, including ex situ germination, should be implemented. Although orchids are a group faced by many unique challenges, the future of orchid diversityin New Zealand is hopeful.