Researchers have developed a pipeline to generate genomes from single cells of uncultivated fungi. The approach was tested on several uncultivated fungal species representing early diverging fungi, the earliest evolutionary branches in the fungal genealogy that provide a repertoire of important and valuable gene products.
They can be found on forest floors, swamps and in houses, ranging in size from smaller than the period on your smartphone's keyboard to stretching over several city blocks. More than a million species of fungi are estimated to live on this planet, but most of that diversity remains unknown because the fungi have avoided detection and have not been cultured for study in laboratories.
Reported in Nature Microbiology , a team led by researchers has developed a pipeline to generate genomes from single cells of uncultivated fungi The approach was tested on several uncultivated fungal species representing early diverging fungi, the earliest evolutionary branches in the fungal genealogy that provide a repertoire of important and valuable gene products.
"Most of the phylogenetic diversity represents early diverging fungi," said study co-senior author Tim James. "We know from environmental DNA that they are common in many habitats, but they are presumably microscopic so you really have to look for them, since you can not culture is representative of what you see in environmental DNA to look at the given sample and identify what the cells might look like, but we also want to look at the genomes of the organisms and infer what they're like, that's where single-cell genomics comes in. "
A Technique for Exploring Unknown Diversity
Through projects such as JGI's 1000 Fungal Genomes, researchers aim to expand the known fraction of fungal diversity with representative genome sequences for various lineages. Even with such efforts, the majority of available genomes belong to just two major lines, Ascomycota and Basidiomycota. The early-diverging lines that are closer to the base of the Fungal Tree of Life have few representative genomes.
"Conceptually, this is a pilot project," said JGI data scientist and first author Steven Ahrendt, who began working on the project as a postdoctoral fellow. "This is a similar idea to the approach JGI has taken with microbial dark matter – that the species are out there, but they do not show up in plate-based culturing."
"That mycoparasitic lifestyle might be a factor in why these species are unculturable," Ahrendt noted and Zoopagomycota. In addition, six of the seven fungi are mycoparasites or fungi that attack other fungi. As such, they need to be able to infest the hosts without harming themselves. These species were grown in co-culture with their hosts and then the spores of the parasites were isolated for sampling. "That mycoparasitic lifestyle might be a factor in why these species are unculturable," Ahrendt noted.
Beyond Proof of Principle
There are around 2,000 described species of early diverging fungi, and about 120,000 described species of the Ascomycota and Basidiomycota, James said. "We've described maybe 5 percent of the fungal diversity, and we're in an era where we can start at that missing piece of the diversity.
"This work was a proof-of-principle that the single-cell genomics approach can reconstruct near-complete fungal genomes and provide insights into phylogenetic position and metabolic capacities of diverse unculturable species from environmental samples," agreed JGI Fungal Program head and co- senior author Igor Grigoriev.
"Several genomes in this study represent the first references for fungal phyla containing most of the species that have not been cultured, having genome sequences and metabolic reconstructions of a broad diversity of uncultured fungal species enable us to better understand fungal evolution and expand the catalogs of gene, enzymes and pathways for DOE science and applications. "