Fungi produce a vast number of secondary metabolites that shape their interactions with other organisms and the environment. Characterising the genes underpinning metabolite synthesis is therefore key to understanding fungal evolution and adaptation. Lichenised fungi represent almost one-third of Ascomycota diversity and boast impressive secondary metabolites repertoires. However, most lichen biosynthetic genes have not been linked to their metabolite products. Here we used metagenomic sequencing to survey gene families associated with production of anthraquinones, UV-protectant secondary metabolites present in various fungi, but especially abundant in a diverse order of lichens, the Teloschistales (class Lecanoromycetes, phylum Ascomycota). We successfully assembled 24 new, high-quality lichenised fungal genomes de novo and combined them with publicly available Lecanoromycetes genomes from taxa with diverse secondary chemistry to produce a whole-genome tree. Secondary metabolite biosynthetic gene cluster (BGC) analysis showed that whilst lichen BGCs are numerous and highly dissimilar, core enzyme genes are generally conserved across taxa. This suggests metabolite diversification occurs via re-shuffling existing enzyme genes with novel accessory genes rather than BGC gains/losses or de novo gene evolution. We identified putative anthraquinone BGCs in our lichen dataset that appear homologous to anthraquinone clusters from non-lichenised fungi, suggesting these genes were present in the common ancestor of the subphylum Pezizomycotina. Finally, we identified unique transporter genes in Teloschistales anthraquinone BGCs that may explain why these metabolites are so abundant and ubiquitous in these lichens. Our results support the importance of metagenomics for understanding the secondary metabolism of non-model fungi such as lichens.

Outbreaks of fungal diseases have devastated plants and animals throughout history. Over the past century, the repeated emergence of coffee wilt disease caused by the fungal pathogen Fusarium xylarioides severely impacted coffee production across sub-Saharan Africa. To improve the disease management of such pathogens, it is crucial to understand their genetic structure and evolutionary potential. We compared the genomes of 13 historic strains spanning 6 decades and multiple disease outbreaks to investigate population structure and host specialisation. We found that F. xylarioides comprised at least 4 distinct lineages: 1 host-specific to Coffea arabica, 1 to C. canephora var. robusta, and 2 historic lineages isolated from various Coffea species. The presence/absence of large genomic regions across populations, the higher genetic similarities of these regions between species than expected based on genome-wide divergence and their locations in different loci in genomes across populations showed that horizontal transfers of effector genes from members of the F. oxysporum species complex contributed to host specificity. Multiple transfers into F. xylarioides populations matched different parts of the F. oxysporum mobile pathogenicity chromosome and were enriched in effector genes and transposons. Effector genes in this region and other carbohydrate-active enzymes important in the breakdown of plant cell walls were shown by transcriptomics to be highly expressed during infection of C. arabica by the fungal arabica strains. Widespread sharing of specific transposons between F. xylarioides and F. oxysporum, and the correspondence of a putative horizontally transferred regions to a Starship (large mobile element involved in horizontal gene transfers in fungi), reinforce the inference of horizontal transfers and suggest that mobile elements were involved. Our results support the hypothesis that horizontal gene transfers contributed to the repeated emergence of coffee wilt disease.

Urban areas may contain a wide range of potential habitats and environmental gradients and, given the many benefits to human health and well-being, there is a growing interest in maximizing their biodiversity potential. However, the ecological patterns and processes in urban areas are poorly understood. Using a widely applicable ecological survey method, we sampled epiphytic lichen communities, important bioindicators of atmospheric pollution, on host Quercus trees in urban parks of London, UK, to test if common patterns relating to lichen diversity are mirrored in urban green spaces. We found lichen diversity to be dependent on host species identity, and negatively related to local tree crowding. In addition, we found a strong negative effect of tree size on lichen diversity, leaving large trees as unexploited niches. A novel network analysis revealed the presence of only pioneer communities, showing the lichen communities are being held in successional stasis, likely due to the heritage effects of SO₂ emissions and current nitrogen pollution and particulate emissions. Our study highlights that jointly assessing species richness, community structure and the successional stage can be key to understanding diversity patterns in urban ecosystems. Subsequently, this may help best determine the optimum conditions that will facilitate biodiversity increase within cities.

Lichen-forming fungi are a diverse and ecologically important group of obligate mutualistic symbionts. Due to difficulties with maintaining them in culture and their extremely slow growth, lichenologists are increasingly opting for metagenomic sequencing followed by symbiont genome separation using bioinformatic pipelines. However, without knowing the true genome size of the lichen-forming fungus, we cannot quantify the completeness of the genome assembly and the efficacy of the bioinformatic filtering. To address this issue, we report here the first whole-genome assembly for the lichen-forming fungus Ramalina farinacea (L.) Ach. sequenced with Oxford Nanopore long-read technology alongside direct measurements of its genome size using flow cytometry. The assembly showed high contiguity (N50 = 1.55 Mb) and gene set completeness (BUSCO = 95.8%). The highly robust genome size of 33.61 Mb/1C (coefficients of variation = 2.98) that was obtained showed our assembly covered 97% of the entire genome. Our results demonstrate that accurate genome size measurements can be obtained directly from lichen thalli and used to provide a benchmark for assessing true cytometric completeness of metagenome-derived assemblies.

Summary: High‐throughput molecular studies of museum specimens (museomics) have great potential in biodiversity research, but fungal historical collections have scarcely been examined, leading to no comprehensive methodological assessments. Here we present a whole genome sequencing (WGS) project conducted at the Fungarium of the Royal Botanic Gardens, Kew. DNA was extracted from 2104 specimens collected between 1770 and 2023, and we found that the specimen age had the smallest effect, while DNA purification and taxonomic identity had the greatest effect on DNA yield. We barcoded 771 specimens, and WGS was conducted on a subset of 394 specimens that were empirically selected for in‐depth analysis. We developed an automated assembly pipeline, integrating 16 different approaches. Starting from 220 libraries (excluding lichenised specimens), we produced 3143 assemblies using these approaches and found that there is no universal assembly method that can provide good‐quality genomes for all; rather, different approaches should be used depending on the library size and the specimen's age. Producing high‐quality genomes from specimens over 100 yr old is possible by using customised DNA extraction protocols and applying a multimethod bioinformatic approach. Whole genomes from historical collections will enrich genomics resources, accelerating biodiversity and evolutionary research, amongst others.

Bark and ambrosia beetles are ecologically important and widespread forest organisms. While flying between host plants and tunnelling into bark, they vector a high diversity of plant and insect-associated fungi within and between forests. These fungal communities live under the bark and rarely produce visible spore-bearing structures, making them difficult to sample and identify at scale. By combining passive insect trapping across the United Kingdom with individual beetle metabarcoding, we generated a reference DNA barcode library for fungi associated with over 1000 beetles, representing 25 native weevil species in the subfamily Scolytinae (Curculionidae). Sampling sites span longitudinal and latitudinal gradients, multiple forest types, and both natural and plantation forests. We use state-of-the-art fungal ITS2 OTU clustering and identification, resulting in 5274 identified fungal OTUs. This fungal diversity data can be used alongside extensive site and sample metadata to explore geographic and ecological biodiversity patterns. As these associations can be highly invasive and damaging, this resource provides a baseline to understand current fungal communities and monitor future changes.