pecies in the genus Aspergillus are of broad interest to medical 1 , applied 2,3 , and basic research 4. Members of Aspergillus section Nigri ('black aspergilli') are prolific producers of native and heterologous proteins 5,6 , organic acids (in particular citric acid 2,7,8), and secondary metabolites (including biopharmaceuticals and mycotoxins like ochratoxin A). Furthermore, the section members are generally very efficient producers of extracellular enzymes 9,10 ; they are the production organisms for 49 out of 260 industrial enzymes 11,12. Among the most important of these, in addition to A. niger, are A. tubingensis, A. aculeatus, and A. luchuensis (previously A. acidus, A. kawachii, and A. awamori 13-15 , respectively). Members of Aspergillus section Nigri are also known as destructive degraders of foods and feeds, and some isolates produce the potent mycotoxins ochratoxin A 16 and fumonisins 17-19. In addition, some species in this section have been proposed to be pathogenic to humans and other animals 20. It is thus of interest to further examine section Nigri for industrial exploitation, as well as prevention of food spoilage, toxin production, and pathogenicity caused by these fungi. A combined phylogenetic and phenotypic approach has shown that section Nigri contains at least 27 species 21-25. Recent results have shown that the section contains species with high diversity and may consist of two separate clades: the biseriate species and the uniseriate species 26 , which show differences in sexual states 27 , sclerotium formation 28 , and secondary metabolite production 29. In the section, only six species have had their genome sequenced: A. niger 2,8 , A. luchuensis 15,30 , A. carbonarius 31 , A. aculeatus 31 , A. tubingensis 31 , and A. brasiliensis 31. This section, with its combination of species richness and fungal species with a diverse impact on humanity, is thus particularly interesting for studying the diversification of fungi into species. In this study, we have de novo-sequenced the genomes of 20 species of section Nigri, thus completing a genome compendium of 26 described species in the section. Further, we have genome-sequenced three

Section Flavi encompasses both harmful and beneficial Aspergillus species, such as Aspergillus oryzae, used in food fermentation and enzyme production, and Aspergillus flavus, food spoiler and mycotoxin producer. Here, we sequence 19 genomes spanning section Flavi and compare 31 fungal genomes including 23 Flavi species. We reassess their phylogenetic relationships and show that the closest relative of A. oryzae is not A. flavus, but A. minisclerotigenes or A. aflatoxiformans and identify high genome diversity, especially in sub-telomeric regions. We predict abundant CAZymes (598 per species) and prolific secondary metabolite gene clusters (73 per species) in section Flavi. However, the observed phenotypes (growth characteristics, polysaccharide degradation) do not necessarily correlate with inferences made from the predicted CAZyme content. Our work, including genomic analyses, phenotypic assays, and identification of secondary metabolites, highlights the genetic and metabolic diversity within section Flavi.

The fungal genus of is highly interesting, containing everything from industrial cell factories, model organisms, and human pathogens. In particular, this group has a prolific production of bioactive secondary metabolites (SMs). In this work, four diverse species (, ,, and )have been whole-genome PacBio sequenced to provide genetic references in three sections. and also were sequenced for SM elucidation. Thirteen genomes were analyzed with comparative genomics to determine phylogeny and genetic diversity, showing that each presented genome contains 15-27% genes not found in other sequenced Aspergilli. In particular, was compared with the pathogenic species This suggests that can produce most of the same allergens, virulence, and pathogenicity factors as, suggesting that could be as pathogenic as Furthermore, SMs were linked to gene clusters based on biological and chemical knowledge and analysis, genome sequences, and predictive algorithms. We thus identify putative SM clusters for aflatoxin, chlorflavonin, and ochrindol in ,, and , respectively, and novofumigatonin,-cycloechinulin, and -aszonalenins in Our study delivers six fungal genomes, showing the large diversity found in the genus; highlights the potential for discovery of beneficial or harmful SMs; and supports reports of pathogenicity. It also shows how biological, biochemical, and genomic information can be combined to identify genes involved in the biosynthesis of specific SMs.

The increased interest in secondary metabolites (SMs) has driven a number of genome sequencing projects to elucidate their biosynthetic pathways. As a result, studies revealed that the number of secondary metabolite gene clusters (SMGCs) greatly outnumbers detected compounds, challenging current methods to dereplicate and categorize this amount of gene clusters on a larger scale. Here, we present an automated workflow for the genetic dereplication and analysis of secondary metabolism genes in fungi. Focusing on the secondary metabolite rich genus Aspergillus, we categorize SMGCs across genomes into SMGC families using network analysis. Our method elucidates the diversity and dynamics of secondary metabolism in section Nigri, showing that SMGC diversity within the section has the same magnitude as within the genus. Using our genome analysis we were able to predict the gene cluster responsible for biosynthesis of malformin, a potentiator of anti-cancer drugs, in 18 strains. To proof the general validity of our predictions, we developed genetic engineering tools in Aspergillus brasiliensis and subsequently verified the genes for biosynthesis of malformin.