In the HeteroDynamic research program we address the evolutionary stability of AM symbiosis, and base our work on the conceptual hypothesis that the symbiosis is stable thanks to a unique genomic organization, a temporally dynamic heterokaryosis, in the fungal partner. This hypothesis is based on the assumptions that 1) heterokaryosis allows for 2) functional variation 3) over time - and my team works from different angles to test these.
Firstly we address heterokaryosis in AM fungi by developing a novel single nucleus genomics method for sequencing and assembly of AM fungal genomes from single nuclei. Reads from individual nuclei can then be mapped back to the assembly to evaluate sequence variance among nuclei from individual strains and even from within single spores. The physical sorting of nuclei using FACS had been established for one species prior to the onset of the HeteoDynamic research program. During the project the method has been developed to accommodate a range of species and a custom-made assembly workflow has been developed to handle the generated data. The method was published in January 2020 (Montoliu-Nerin et al 2020). An added value of this method is that it allows us to assemble genomes from a broad range of AM fungi that could previously not be sequenced due to challenges obtaining sufficient amount and purity of biological material from most AM fungi. Analysis of within strain genetic variation reveales overall low levels of polymorphism and the single nucleotide level. Based on these studies and work on repetitive DNA content in AM fungi the first PhD student defended her thesis in Dec 2020. Low levels of heterokaryosis suggest that the conceptual hypothesis of the HeteroDynamic project should be rejected. However, our comprehensive genome analysis has revealed interesting patterns of rDNA polymorphism, gene copy number variation and possibly variation in ploidy as means of maintaining genetic variation in AM fungal genomes. Through out the project we have continued to analyse these patterns to the extent it is possible with amplified genome DNA . A new PhD student (David) was recruited in December 2019 and has has since focused on verifying genetic polymorphism and examining signals of selection in the existing polymorphism. With the novel workflow for genome sequencing that we have established we have expanded the phylogenetic width of available genome data in Gloeromycota and we have publishing 21 new genomes from 12 genera across seven families.
Secondly, we set out to test for functional heterokaryosis, by simultaneously exposing fungal strains to different hosts. However, the growth experimental part of the HeteroDynamic project has not been as straightforward as the genomic part of the project and in the end the data from these experiments can not be used to test the initial hypothesis. The main reason being lack of fungal colonisation at the time of harvest. Despite this set back, the project has generated major additions of genomic data to the field of AM fungal genomics and ongoing analysis of these resources is building towards a solid understanding of the evolutionary history of these major plant symbionts. Our research is not only important for better understanding evolutionary dynamics of ancient mutualistic interactions between fungi and plants but are potentially important for future agriculture including the use of AM inoculum, optimized rotations and breeding for beneficial AM fungi. While negative feedback between fungi and host plants is important in maintaining high biodiversity in natural ecosystems its role in agriculture has not yet been tested and our studies build the foundation for such studies. To take advantage of growth promoting AM fungal communities in sustainable intensification of agriculture, systems need to be developed where crop rotations are tested for their ability to promote continually beneficial communities.
