BackgroundThe fungal genus Aspergillus is of critical importance to humankind. Species include those with industrial applications, important pathogens of humans, animals and crops, a source of potent carcinogenic contaminants of food, and an important genetic model. The genome sequences of eight aspergilli have already been explored to investigate aspects of fungal biology, raising questions about evolution and specialization within this genus.ResultsWe have generated genome sequences for ten novel, highly diverse Aspergillus species and compared these in detail to sister and more distant genera. Comparative studies of key aspects of fungal biology, including primary and secondary metabolism, stress response, biomass degradation, and signal transduction, revealed both conservation and diversity among the species. Observed genomic differences were validated with experimental studies. This revealed several highlights, such as the potential for sex in asexual species, organic acid production genes being a key feature of black aspergilli, alternative approaches for degrading plant biomass, and indications for the genetic basis of stress response. A genome-wide phylogenetic analysis demonstrated in detail the relationship of the newly genome sequenced species with other aspergilli.ConclusionsMany aspects of biological differences between fungal species cannot be explained by current knowledge obtained from genome sequences. The comparative genomics and experimental study, presented here, allows for the first time a genus-wide view of the biological diversity of the aspergilli and in many, but not all, cases linked genome differences to phenotype. Insights gained could be exploited for biotechnological and medical applications of fungi.Electronic supplementary materialThe online version of this article (doi:10.1186/s13059-017-1151-0) contains supplementary material, which is available to authorized users.

Background: Citric acid, a commodity product of industrial biotechnology, is produced by fermentation of the filamentous fungus Aspergillus niger. A requirement for high-yield citric acid production is keeping the concentration of Mn 2+ ions in the medium at or below 5 µg L −1. Understanding manganese metabolism in A. niger is therefore of critical importance to citric acid production. To this end, we investigated transport of Mn 2+ ions in A. niger NRRL2270. Results: we identified an A. niger gene (dmtA; NRRL3_07789), predicted to encode a transmembrane protein, with high sequence identity to the yeast manganese transporters Smf1p and Smf2p. Deletion of dmtA in A. niger eliminated the intake of Mn 2+ at low (5 µg L −1) external Mn 2+ concentration, and reduced the intake of Mn 2+ at high (> 100 µg L −1) external Mn 2+ concentration. Compared to the parent strain, overexpression of dmtA increased Mn 2+ intake at both low and high external Mn 2+ concentrations. Cultivation of the parent strain under Mn 2+ ions limitation conditions (5 µg L −1) reduced germination and led to the formation of stubby, swollen hyphae that formed compact pellets. Deletion of dmtA caused defects in germination and hyphal morphology even in the presence of 100 µg L −1 Mn 2+ , while overexpression of dmtA led to enhanced germination and normal hyphal morphology at limiting Mn 2+ concentration. Growth of both the parent and the deletion strains under citric acid producing conditions resulted in molar yields (Y p/s) of citric acid of > 0.8, although the deletion strain produced ~ 30% less biomass. This yield was reduced only by 20% in the presence of 100 µg L −1 Mn 2+ , whereas production by the parent strain was reduced by 60%. The Y p/s of the overexpressing strain was 17% of that of the parent strain, irrespective of the concentrations of external Mn 2+. Conclusions: Our results demonstrate that dmtA is physiologically important in the transport of Mn 2+ ions in A. niger, and manipulation of its expression modulates citric acid overflow.

Itaconic acid is a five-carbon dicarboxylic acid with an unsaturated alkene bond, frequently used as a building block for the industrial production of a variety of synthetic polymers. It is also one of the major products of fungal "overflow metabolism" which can be produced in submerged fermentations of the filamentous fungus Aspergillus terreus. At the present, molar yields of itaconate are lower than those obtained in citric acid production in Aspergillus niger. Here, we have studied the possibility that the yield may be limited by the oxygen supply during fermentation and hence tested the effect of the dissolved oxygen concentration on the itaconic acid formation rate and yield in lab-scale bioreactors. The data show that a dissolved oxygen concentration of 2% saturation was sufficient for maximal biomass formation. Raising it to 30% saturation had no effect on biomass formation or the growth rate, but the itaconate yield augmented substantially from 0.53 to 0.85 mol itaconate/mol glucose. Furthermore, the volumetric and specific rates of itaconic acid formation ameliorated by as much as 150% concurrent with faster glucose consumption, shortening the fermentation time by 48 h. Further increasing the dissolved oxygen concentration over 30% saturation had no effect. Moreover, we show that this increase in itaconic acid production coincides with an increase in alternative respiration, circumventing the formation of surplus ATP by the cytochrome electron transport chain, as well as with increased levels of alternative oxidase transcript. We conclude that high(er) itaconic acid accumulation requires a dissolved oxygen concentration that is much higher than that needed for maximal biomass formation, and postulate that the induction of alternative respiration allows the necessary NADH reoxidation ratio without surplus ATP production to increase the glucose consumption and the flux through overflow metabolism.

Itaconic acid is used as a bio-based, renewable building block in the polymer industry. It is produced by submerged fermentations of the filamentous fungus Aspergillus terreus from molasses or starch, but research over the efficient utilization of non-food, lignocellulosic plant biomass is soaring. The objective of this study was to test whether the application of two key cultivation parameters for obtaining itaconic acid from D -glucose in high yields – Mn 2+ ion deficiency and high concentration of the carbon source – would also occur on D -xylose, the principal monomer of lignocellulose. To this end, a carbon and energy balance for itaconic acid formation was established, which is 0.83 moles/mole D -xylose. The effect of Mn 2+ ions on itaconic acid formation was indeed similar to that on D -glucose and maximal yields were obtained below 3 μg L –1 Mn 2+ ions, which were, however, only 0.63 moles of itaconic acid per mole D -xylose. In contrast to the case on D -glucose, increasing D -xylose concentration over 50 g L –1 did not change the above yield. By-products such as xylitol and α-ketoglutarate were found, but in total they remained below 2% of the concentration of D -xylose. Mass balance of the fermentation with 110 g L –1 D -xylose revealed that >95% of the carbon from D -xylose was accounted as biomass, itaconic acid, and the carbon dioxide released in the last step of itaconic acid biosynthesis. Our data show that the efficiency of biomass formation is the critical parameter for itaconic acid yield from D -xylose under otherwise optimal conditions.

Seed contamination with polyketide mycotoxins such as sterigmatocystin (ST) produced by Aspergilli is a worldwide issue. The ST biosynthetic pathway is well-characterized in A. nidulans, but regulatory aspects related to the carbon source are still enigmatic. This is particularly true for lactose, inasmuch as some ST production mutant strains still synthesize ST on lactose but not on other carbon substrates. Here, kinetic data revealed that on d-glucose, ST forms only after the sugar is depleted from the medium, while on lactose, ST appears when most of the carbon source is still available. Biomass-specified ST production on lactose was significantly higher than on d-glucose, suggesting that ST formation may either be mediated by a carbon catabolite regulatory mechanism, or induced by low specific growth rates attainable on lactose. These hypotheses were tested by d-glucose limited chemostat-type continuous fermentations. No ST formed at a high growth rate, while a low growth rate led to the formation of 0.4 mg·L−1 ST. Similar results were obtained with a CreA mutant strain. We concluded that low specific growth rates may be the primary cause of mid-growth ST formation on lactose in A. nidulans, and that carbon utilization rates likely play a general regulatory role during biosynthesis.