Filamentous fungi differentiate along complex developmental programs directed by abiotic and biotic signals. Currently, intrinsic signals that govern fungal development remain largely unknown. Here we show that an endogenously produced and secreted fungal oxylipin, 5,8-diHODE, induces fungal cellular differentiation, including lateral branching in pathogenic Aspergillus fumigatus and Aspergillus flavus, and appressorium formation in the rice blast pathogen Magnaporthe grisea. The Aspergillus branching response is specific to a subset of oxylipins and is signaled through G-protein coupled receptors. RNA-Seq profiling shows differential expression of many transcription factors in response to 5,8-diHODE. Screening of null mutants of 33 of those transcription factors identifies three transcriptional regulators that appear to mediate the Aspergillus branching response; one of the mutants is locked in a hypo-branching phenotype, while the other two mutants display a hyper-branching phenotype. Our work reveals an endogenous signal that triggers crucial developmental processes in filamentous fungi, and opens new avenues for research on the morphogenesis of filamentous fungi.

Significance: Secondary metabolites (SMs) produced by fungi mediate ecological interactions, define fungal niches, and are of profound pharmacological importance to humans. Most work on SMs has focused on a small number of individuals from each species, not fully reflecting the importance of intraspecific diversity. We demonstrate that even in one of the best-studied model fungi, the carcinogen-producing Aspergillus flavus , more than 25% of SM-producing biosynthetic gene clusters (BGCs) are novel and/or show population-specific variants. These results support the finding that the organization of BGC diversity into population-specific patterns may sometimes result from ecologically important interactions and may inform evolutionary and etiological inferences of SM capacities within a species. Importantly, our work also presents a vision of sources of potential pharmaceuticals.

Fungus-growing attine ants are under constant threat from fungal pathogens such as the specialized mycoparasite Escovopsis, which uses combined physical and chemical attack strategies to prey on the fungal gardens of the ants. In defence, some species assemble protective microbiomes on their exoskeletons that contain antimicrobial-producing Actinobacteria. Underlying this network of mutualistic and antagonistic interactions are an array of chemical signals. Escovopsis weberi produces the shearinine terpene-indole alkaloids, which affect ant behaviour, diketopiperazines to combat defensive bacteria, and other small molecules that inhibit the fungal cultivar. Pseudonocardia and Streptomyces mutualist bacteria produce depsipeptide and polyene macrolide antifungals active against Escovopsis spp. The ant nest metabolome is further complicated by competition between defensive bacteria, which produce antibacterials active against even closely related species.

Three novel dimeric xanthones, cryptosporioptides A–C were isolated from Cryptosporiopsis sp. 8999 and their structures elucidated. Methylation of cryptosporioptide A gave a methyl ester with identical NMR data to cryptosporioptide, a compound previously reported to have been isolated from the same fungus. However, HRMS analysis revealed that cryptosporioptide is a symmetrical dimer, not a monomer as previously proposed, and the revised structure was elucidated by extensive NMR analysis. The genome of Cryptosporiopsis sp. 8999 was sequenced and the dimeric xanthone (dmx) biosynthetic gene cluster responsible for the production of the cryptosporioptides was identified. Gene disruption experiments identified a gene (dmxR5) encoding a cytochrome P450 oxygenase as being responsible for the dimerisation step late in the biosynthetic pathway. Disruption of dmxR5 led to the isolation of novel monomeric xanthones. Cryptosporioptide B and C feature an unusual ethylmalonate subunit: a hrPKS and acyl CoA carboxylase are responsible for its formation. Bioinformatic analysis of the genomes of several fungi producing related xanthones, e.g. the widely occurring ergochromes, and related metabolites allows detailed annotation of the biosynthetic genes, and a rational overall biosynthetic scheme for the production of fungal dimeric xanthones to be proposed.

The filamentous fungus Aspergillus nidulans has been a primary workhorse used to understand fungal genetics. Much of this work has focused on elucidating the genetics of biosynthetic gene clusters (BGCs) and the secondary metabolites (SMs) they produce. SMs are both niche defining in fungi and of great economic importance to humans. Despite the focus on A. nidulans, very little is known about the natural diversity in secondary metabolism within this species. We determined the BGC content and looked for evolutionary patterns in BGCs from whole-genome sequences of two clinical isolates and the A4 reference genome of A. nidulans. Differences in BGC content were used to explain SM profiles determined using liquid chromatography–high-resolution mass spectrometry. We found that in addition to genetic variation of BGCs contained by all isolates, nine BGCs varied by presence/absence. We discovered the viridicatumtoxin BGC in A. nidulans and suggest that this BGC has undergone a horizontal gene transfer from the Aspergillus section Nigri lineage into Penicillium sometime after the sections Nigri and Nidulantes diverged. We identified the production of viridicatumtoxin and several other compounds previously not known to be produced by A. nidulans. One isolate showed a lack of sterigmatocystin production even though it contained an apparently intact sterigmatocystin BGC, raising questions about other genes and processes known to regulate this BGC. Altogether, our work uncovers a large degree of intraspecies diversity in BGC and SM production in this genetic model species and offers new avenues to understand the evolution and regulation of secondary metabolism. IMPORTANCE Much of what we know about the genetics underlying secondary metabolite (SM) production and the function of SMs in the model fungus Aspergillus nidulans comes from a single reference genome. A growing body of research indicates the importance of biosynthetic gene cluster (BGC) and SM diversity within a species. However, there is no information about the natural diversity of secondary metabolism in A. nidulans. We discovered six novel clusters that contribute to the considerable variation in both BGC content and SM production within A. nidulans. We characterize a diverse set of mutations and emphasize how findings of single nucleotide polymorphisms (SNPs), deletions, and differences in evolutionary history encompass much of the variation observed in nonmodel systems. Our results emphasize that A. nidulans may also be a strong model to use within-species diversity to elucidate regulatory cross talk, fungal ecology, and drug discovery systems.