Genomic features of lichen-associated black fungi

Abstract

Lichens are mutualistic associations consisting of a primary fungal host, and one to few primary phototrophic symbiont(s), usually a green alga and/or a cyanobacterium. They form complex thallus structures, which provide unique and stable habitats for many other microorganisms. Frequently isolated from lichens are the so-called black fungi, or black yeasts, which are mainly characterized by melanized cell walls and extremophilic lifestyles. It is presently unclear in which ways these fungi interact with other members of the lichen symbiosis. Genomic resources of lichen-associated black fungi are needed to better understand the physiological potential of these fungi and shed light on the complexity of the lichen consortium. Here, we present high-quality genomes of 14 black fungal lineages, isolated from lichens of the rock-dwelling genus Umbilicaria. Nine of the lineages belong to the Eurotiomycetes (Chaetothyriales), four to the Dothideomycetes, and one to the Arthoniomycetes, representing the first genome of a black fungus in this class. The PacBio-based assemblies are highly contiguous (5-42 contigs per genome, mean coverage of 79-502, N50 of 1.0-7.3 mega-base-pair (Mb), Benchmarking Universal Single-Copy Orthologs (BUSCO) completeness generally ≥95.4%). Most contigs are flanked by a telomere sequence, suggesting we achieved near chromosome-level assemblies. Genome sizes range between 26 and 44 Mb. Transcriptome-based annotations yielded ~11,000-18,000 genes per genome. We analyzed genome content with respect to repetitive elements, biosynthetic genes, and effector genes. Each genome contained a polyketide synthase gene related to the dihydroxynaphthalene-melanin pathway. This research provides insights into genome content and metabolic potential of these relatively unknown, but frequently encountered lichen associates.

Keywords: PacBio; biosynthetic gene clusters; diversity; funannotate; melanin; phylogenetics; symbiosis.

FIGURE 1

Phylogenetic placement of 14 black fungal lineages isolated from lichens. The new lineages are highlighted in red and group in the Eurotiomycetes, Dothideomycetes, and Arthoniomycetes. Presented trees are maximum likelihood phylogenies based on concatenated ITS and nuLSU sequences. Branches in bold indicate ML bootstrap support ≥70%. All data retrieved from GenBank are provided in Table S2. Full phylogenetic trees, without collapsed clades and detailed BS, are available as Figures [Link], [Link].

FIGURE 2

Genome assembly and annotation statistics of 14 de novo genomes of black fungi isolated from lichens. Indicated are genome size, number of contigs, number of exons, percentage of repeats, number of annotated genes and proteins, and composition of secondary metabolite gene clusters. Phylogenetic relationships of the analyzed taxa are indicated (yellow—Eurotiomycetes, orange—Dothideomycetes, blue—Arthoniomycetes). Secondary metabolite categories comprise the following: type I and III PKS, NRPS and NRPS‐like, terpenes, (fungal‐)ribosomally synthesized and post‐translationally modified peptides (RiPPs) and (fungal‐)RiPP‐like, and others. Others include phosphate‐like, betalacetones, indoles, non‐alpha poly‐amino acids, and non‐ribosomal peptide‐metallophores (NRP‐metallophore). Secondary metabolite gene clusters were identified using antiSMASH. A detailed overview on genome assembly and annotation statistics is provided in Table S3.

FIGURE 3

Number of predicted effectors and secreted proteins in genomes of black fungi, supplemented with the percentage of effectors in relation to all annotated proteins. Apoplastic and cytoplasmic effectors are summed up. Effectors were predicted using EffectorP.

FIGURE 4

Phylogenetic relationships of the keto synthase (KS) domain of polyketide synthase genes. This is a maximum likelihood phylogeny of non‐reducing PKSs (NR‐PKSs). ML bootstrap support values ≥70% are shown, and a tree scale bar is indicated at the bottom of the tree. Black fungal isolates of the present study are highlighted in red. We included all Ascomycota NR‐PKSs linked to a secondary metabolite from the MIBiG database. Clade numbers follow previous publications., , Example metabolites or metabolite classes are indicated.