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We classified the genes encoding carbohydrate-active enzymes (CAZymes) in 17 sequenced genomes representing 16 evolutionarily diverse Aspergillus species. We performed a phylogenetic analysis of the encoding enzymes, along with experimentally characterized CAZymes, to assign molecular function to the Aspergilli CAZyme families and subfamilies. Genome content analysis revealed that the numbers of CAZy genes per CAZy family related to plant biomass degradation follow closely the taxonomic distance between the species. On the other hand, growth analysis showed almost no correlation between the number of CAZyme genes and the efficiency in polysaccharide utilization. The exception is A. clavatus where a reduced number of pectinolytic enzymes can be correlated with poor growth on pectin. To gain detailed information on the enzymes used by Aspergilli to breakdown complex biomass, we conducted exoproteome analysis by mass spectrometry. These results showed that Aspergilli produce many different enzymes mixtures in the presence of sugar beet pulp and wheat bran. Despite the diverse enzyme mixtures produced, species of section Nigri, A. aculeatus, A. nidulans and A. terreus, produce mixtures of enzymes with activities that are capable of digesting all the major polysaccharides in the available substrates, suggesting that they are capable of degrading all the polysaccharides present simultaneously. For the other Aspergilli, typically the enzymes produced are targeted to a subset of polysaccharides present, suggesting that they can digest only a subset of polysaccharides at a given time.

Background: Laccases can oxidize a broad spectrum of substrates, offering promising applications in various sectors, such as bioremediation, biomass fractionation in future biorefineries, and synthesis of biochemicals and biopolymers. However, laccase discovery and optimization with a desirable pH optimum remains a challenge due to the labor-intensive and time-consuming nature of the traditional laboratory methods., Results: This study presents a machine learning (ML)-integrated approach for predicting pH optima of basidiomycete fungal laccases, utilizing a small, curated dataset against a vast metagenomic data. Comparative computational analyses unveiled the structural and pH-dependent solubility differences between acidic and neutral-alkaline laccases, helping us understand the molecular bases of enzyme pH optimum. The pH profiling of the two ML-predicted alkaline laccase candidates from the basidiomycete fungus Lepista nuda further validated our computational approach, showing the accuracy of this comprehensive method., Conclusions: This study uncovers the efficacy of ML in the prediction of enzyme pH optimum from minimal datasets, marking a significant step towards harnessing computational tools for systematic screening of enzymes for biotechnology applications., (© 2024. The Author(s).)

Aspergillus terreus has attracted interest due to its application in industrial biotechnology, particularly for the production of itaconic acid and bioactive secondary metabolites. As related species also seem to possess a prosperous secondary metabolism, they are of high interest for genome mining and exploitation. Here, we present draft genome sequences for six species from Aspergillus section Terrei and one species from Aspergillus section Nidulantes . Whole-genome phylogeny confirmed that section Terrei is monophyletic. Genome analyses identified between 70 and 108 key secondary metabolism genes in each of the genomes of section Terrei , the highest rate found in the genus Aspergillus so far. The respective enzymes fall into 167 distinct families with most of them corresponding to potentially unique compounds or compound families. Moreover, 53% of the families were only found in a single species, which supports the suitability of species from section Terrei for further genome mining. Intriguingly, this analysis, combined with heterologous gene expression and metabolite identification, suggested that species from section Terrei use a strategy for UV protection different to other species from the genus Aspergillus . Section Terrei contains a complete plant polysaccharide degrading potential and an even higher cellulolytic potential than other Aspergilli, possibly facilitating additional applications for these species in biotechnology.

Background: Colletotrichum fungi infect a wide diversity of monocot and dicot hosts, causing diseases on almost all economically important plants worldwide. Colletotrichum is also a suitable model for studying gene family evolution on a fine scale to uncover events in the genome associated with biological changes., Results: Here we present the genome sequences of 30 Colletotrichum species covering the diversity within the genus. Evolutionary analyses revealed that the Colletotrichum ancestor diverged in the late Cretaceous in parallel with the diversification of flowering plants. We provide evidence of independent host jumps from dicots to monocots during the evolution of Colletotrichum, coinciding with a progressive shrinking of the plant cell wall degradative arsenal and expansions in lineage-specific gene families. Comparative transcriptomics of 4 species adapted to different hosts revealed similarity in gene content but high diversity in the modulation of their transcription profiles on different plant substrates. Combining genomics and transcriptomics, we identified a set of core genes such as specific transcription factors, putatively involved in plant cell wall degradation., Conclusions: These results indicate that the ancestral Colletotrichum were associated with dicot plants and certain branches progressively adapted to different monocot hosts, reshaping the gene content and its regulation., (© The Author(s) 2024. Published by Oxford University Press GigaScience.)

The import of plant-derived small sugars by sugar transporters (STs) has received increasing interest due to its important biological role and great industrial potential. STs are important targets of genetic engineering to improve fungal plant biomass conversion. Comparatively analysis of the genome-wide prevalence and transcriptomics of STs was performed in four filamentous fungi: Aspergillus niger, Aspergillus nidulans, Penicillium subrubescens and Trichoderma reesei. Using phylogenetic analysis and literature mining, their predicted STs were divided into ten subfamilies with putative sugar specificities assigned. In addition, transcriptome analysis revealed complex expression profiles among different STs subfamilies and fungal species, indicating a sophisticated transcriptome regulation and functional diversity of fungal STs. Several STs showed strong co-expression with other genes involved in sugar utilization, encoding CAZymes and sugar catabolic enzymes. This study provides new insights into the diversity of STs at the genomic/transcriptomic level, facilitating their biochemical characterization and metabolic engineering., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Metabolic engineering of filamentous fungi has received increasing attention in recent years, especially in the context of creating better industrial fungal cell factories to produce a wide range of valuable enzymes and metabolites from plant biomass. Recent studies into the pentose catabolic pathway (PCP) in Aspergillus niger have revealed functional redundancy in most of the pathway steps. In this study, a closer examination of the A. niger genome revealed five additional paralogs for the three original pentose reductases (LarA, XyrA, XyrB). Analysis of these genes using phylogeny, in vitro and in vivo functional analysis of the enzymes, and gene expression revealed that all can functionally replace LarA, XyrA, and XyrB. However, they are also active on several other sugars, suggesting a role for them in other pathways. This study therefore reveals the diversity of primary carbon metabolism in fungi, suggesting an intricate evolutionary process that distinguishes different species. In addition, through this study, the metabolic toolkit for synthetic biology and metabolic engineering of A. niger and other fungal cell factories has been expanded.

Many fungi show a strong preference for specific habitats and growth conditions. Investigating the molecular mechanisms of fungal adaptation to varying environmental conditions is of great interest to biodiversity research and is important for many industrial applications. In this study, we compared the transcriptome profiles of two previously genome-sequenced white-rot wood-decay fungi, Trametes pubescens and Phlebia centrifuga, during their growth on two common plant biomass substrates (wheat straw and spruce) at two temperatures (15 °C and 25 °C). The results showed that both fungi partially tailored their molecular responses to different types of carbon sources, differentially expressing genes encoding polysaccharide degrading enzymes, transporters, proteases and monooxygenases. Notably, more lignin modification related AA2 genes and cellulose degradation related AA9 genes were differentially expressed in the tested conditions of T. pubescens than P. centrifuga. In addition, we detected more remarkable transcriptome changes to different growth temperature in P. centrifuga than in T. pubescens, which reflected their different ability to adapt to the temperature fluctuations. In P. centrifuga, differentially expressed genes (DEGs) related to temperature response mainly encode protein kinases, trehalose metabolism, carbon metabolic enzymes and glycoside hydrolases, while the main temperature-related DEGs identified in T. pubescens are only the carbon metabolic enzymes and glycoside hydrolases. Our study revealed both conserved and species-specific transcriptome changes during fungal adaptation to a changing environment, improving our understanding of the molecular mechanisms underlying fungal plant biomass conversion at varying temperatures., Competing Interests: Declaration of Competing Interest The authors declare that they have no conflict of interest., (Copyright © 2023 The Authors. Published by Elsevier GmbH.. All rights reserved.)

l-arabitol is an intermediate of the pentose catabolic pathway in fungi but can also be used as a carbon source by many fungi, suggesting the presence of transporters for this polyol. In this study, an l-arabitol transporter, LatA, was identified in Aspergillus niger . Growth and expression profiles as well as sugar consumption analysis indicated that LatA only imports l-arabitol and is regulated by the arabinanolytic transcriptional activator AraR. Moreover, l-arabitol production from wheat bran was increased in a metabolically engineered A. niger mutant by the deletion of latA , indicating its potential for improving l-arabitol-producing cell factories. Phylogenetic analysis showed that homologs of LatA are widely conserved in fungi.

Fungi play a critical role in the global carbon cycle by degrading plant polysaccharides to small sugars and metabolizing them as carbon and energy sources. We mapped the well-established sugar metabolic network of Aspergillus niger to five taxonomically distant species ( Aspergillus nidulans , Penicillium subrubescens , Trichoderma reesei , Phanerochaete chrysosporium and Dichomitus squalens ) using an orthology-based approach. The diversity of sugar metabolism correlates well with the taxonomic distance of the fungi. The pathways are highly conserved between the three studied Eurotiomycetes ( A. niger , A. nidulans , P. subrubescens ). A higher level of diversity was observed between the T. reesei and A. niger , and even more so for the two Basidiomycetes. These results were confirmed by integrative analysis of transcriptome, proteome and metabolome, as well as growth profiles of the fungi growing on the corresponding sugars. In conclusion, the establishment of sugar pathway models in different fungi revealed the diversity of fungal sugar conversion and provided a valuable resource for the community, which would facilitate rational metabolic engineering of these fungi as microbial cell factories.

Acetyl esterases are an important component of the enzymatic machinery fungi use to degrade plant biomass and are classified in several Carbohydrate Esterase families of the CAZy classification system. Carbohydrate Esterase family 16 (CE16) is one of the more recently discovered CAZy families, but only a small number of its enzyme members have been characterized so far, revealing activity on xylan-derived oligosaccharides, as well as activity related to galactoglucomannan. The number of CE16 genes differs significantly in the genomes of filamentous fungi. In this study, four CE16 members were identified in the genome of Aspergillus niger NRRL3 and it was shown that they belong to three of the four phylogenetic Clades of CE16. Significant differences in expression profiles of the genes and substrate specificity of the enzymes were revealed, demonstrating the diversity within this family of enzymes. Detailed characterization of one of these four A. niger enzymes (HaeA) demonstrated activity on oligosaccharides obtained from acetylated glucuronoxylan, galactoglucomannan and xyloglucan, thus establishing this enzyme as a general hemicellulose acetyl esterase. Their broad substrate specificity makes these enzymes highly interesting for biotechnological applications in which deacetylation of polysaccharides is required., (Copyright © 2022 The Authors. Published by Elsevier B.V. All rights reserved.)

The unique ability of basidiomycete white rot fungi to degrade all components of plant cell walls makes them indispensable organisms in the global carbon cycle. In this study, we analyzed the proteomes of two closely related white rot fungi, Obba rivulosa and Gelatoporia subvermispora , during eight-week cultivation on solid spruce wood. Plant cell wall degrading carbohydrate-active enzymes (CAZymes) represented approximately 5% of the total proteins in both species. A core set of orthologous plant cell wall degrading CAZymes was shared between these species on spruce suggesting a conserved plant biomass degradation approach in this clade of basidiomycete fungi. However, differences in time-dependent production of plant cell wall degrading enzymes may be due to differences among initial growth rates of these species on solid spruce wood. The obtained results provide insight into specific enzymes and enzyme sets that are produced during the degradation of solid spruce wood in these fungi. These findings expand the knowledge on enzyme production in nature-mimicking conditions and may contribute to the exploitation of white rot fungi and their enzymes for biotechnological applications.

Filamentous fungi produce a wide variety of enzymes in order to efficiently degrade plant cell wall polysaccharides. The production of these enzymes is controlled by transcriptional regulators, which also control the catabolic pathways that convert the released monosaccharides. Two transcriptional regulators, GalX and GalR, control d-galactose utilization in the model filamentous fungus Aspergillus nidulans, while the arabinanolytic regulator AraR regulates l-arabinose catabolism. d-Galactose and l-arabinose are commonly found together in polysaccharides, such as arabinogalactan, xylan and rhamnogalacturonan I. Therefore, the catabolic pathways that convert d-galactose and l-arabinose are often also likely to be active simultaneously. In this study, we investigated the interaction between GalX, GalR and AraR in d-galactose and l-arabinose catabolism. For this, we generated single, double and triple mutants of the three regulators, and analysed their growth and enzyme and gene expression profiles. Our results clearly demonstrated that GalX, GalR and AraR co-regulate d-galactose catabolism in A. nidulans. GalX has a prominent role on the regulation of genes of d-galactose oxido-reductive pathway, while AraR can compensate for the absence of GalR and/or GalX., (© 2022 The Authors. Microbial Biotechnology published by Society for Applied Microbiology and John Wiley & Sons Ltd.)

The current impetus towards a sustainable bio-based economy has accelerated research to better understand the mechanisms through which filamentous fungi convert plant biomass, a valuable feedstock for biotechnological applications. Several transcription factors have been reported to control the polysaccharide degradation and metabolism of the resulting sugars in fungi. However, little is known about their individual contributions, interactions and crosstalk. D-galactose is a hexose sugar present mainly in hemicellulose and pectin in plant biomass. Here, we study D-galactose conversion by Aspergillus niger and describe the involvement of the arabinanolytic and xylanolytic activators AraR and XlnR, in addition to the D-galactose-responsive regulator GalX. Our results deepen the understanding of the complexity of the filamentous fungal regulatory network for plant biomass degradation and sugar catabolism, and facilitate the generation of more efficient plant biomass-degrading strains for biotechnological applications., (Copyright © 2022 The Author(s). Published by Elsevier Inc. All rights reserved.)

Xylitol is widely used in the food and pharmaceutical industries as a valuable commodity product. Biotechnological production of xylitol from lignocellulosic biomass by microorganisms is a promising alternative option to chemical synthesis or bioconversion from D-xylose. In this study, four metabolic mutants of Aspergillus niger were constructed and evaluated for xylitol accumulation from D-xylose and lignocellulosic biomass. All mutants had strongly increased xylitol production from pure D-xylose, beechwood xylan, wheat bran and cotton seed hulls compared to the reference strain, but not from several other feed stocks. The triple mutant ΔladAΔxdhAΔsdhA showed the best performance in xylitol production from wheat bran and cotton seed hulls. This study demonstrated the large potential of A. niger for xylitol production directly from lignocellulosic biomass by metabolic engineering., (Copyright © 2021 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

We investigated whether highly available organic residues in Brazil can be used as substrates for the production of the oyster mushroom Pleurotus ostreatus, instead of the conventional cultivation using the eucalyptus sawdust substrate. We assessed the mushroom yield on 13 substrates, of which 12 were formulated with different concentrations of organic residues and one with pure eucalyptus sawdust, and verified whether the raw material used in the substrate formula and the concentration of such alternative residues influenced their biological efficiencies. Substrates containing eucalyptus bark resulted in higher mushroom yield than those containing eucalyptus sawdust, which generally resulted in similar mushroom yield to the remaining formulas. Moreover, the raw material and the concentration of each residue affected the biological efficiency of the substrates. We show that the conventional substrate for P. ostreatus can be replaced by substrates easily accessible to producers without loss in productivity. Furthermore, that the concentration of these mixtures affects the mushroom productivity and should be considered when formulating the growth medium., (© 2021 The Society for Applied Microbiology.)

Xyloglucan is a prominent matrix heteropolysaccharide binding to cellulose microfibrils in primary plant cell walls. Hence, the hydrolysis of xyloglucan facilitates the overall lignocellulosic biomass degradation. Xyloglucanases (XEGs) are key enzymes classified in several glycoside hydrolase (GH) families. So far, family GH44 has been shown to contain bacterial XEGs only. Detailed genome analysis revealed GH44 members in fungal species from the phylum Basidiomycota, but not in other fungi, which we hypothesized to also be XEGs. Two GH44 enzymes from Dichomitus squalens and Pleurotus ostreatus were heterologously produced and characterized. They exhibited XEG activity and displayed a hydrolytic cleavage pattern different from that observed in fungal XEGs from other GH families. Specifically, the fungal GH44 XEGs were not hindered by substitution of neighboring glucosyl units and generated various "XXXG-type," "GXXX(G)-type," and "XXX-type" oligosaccharides. Overall, these fungal GH44 XEGs represent a novel class of enzymes for plant biomass conversion and valorization., Competing Interests: The authors declare no competing interests., (© 2021 The Authors.)

Laccases are multi-copper oxidases that use molecular oxygen as the electron acceptor to oxidize phenolic and indirectly also non-phenolic substrates by mechanisms involving radicals. Due to their eco-friendliness and broad substrate specificity, laccases span a wide range of biotechnological applications. We have heterologously expressed a laccase from the coprophilic basidiomycete Coprinopsis cinerea ( Cc Lcc9) in the methylotrophic yeast Pichia pastoris . The recombinant Cc Lcc9 (r Cc Lcc9) oxidized 2,6-dimethoxyphenol in the neutral pH range, and showed thermostability up to 70°C. The r Cc Lcc9 efficiently oxidized veratryl alcohol to veratraldehyde in the presence of low molecular weight mediators syringyl nitrile, methyl syringate and violuric acid, which are syringyl-type plant phenolics that have shown potential as natural co-oxidants for lignocellulosic materials. In addition, r Cc Lcc9 is able to depolymerize biorefinery hardwood lignin in the presence of methyl syringate and syringyl nitrile as indicated by gel permeation chromatography, and infrared spectral and nucleic magnetic resonance analyses. Furthermore, we showed that several added-value aromatic compounds, such as vanillin, vanillic acid, syringaldehyde, syringic acid and p -hydroxybenzoic acid, were formed during sequential biocatalytic chemical degradation of biorefinery lignin, indicating that r Cc Lcc9 harbors a great potential for sustainable processes of circular economy and modern biorefineries., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Kontro, Lyra, Koponen, Kuuskeri, Kähkönen, Wallenius, Wan, Sipilä, Mäkelä, Nousiainen and Hildén.)

Next to d-glucose, the pentoses l-arabinose and d-xylose are the main monosaccharide components of plant cell wall polysaccharides and are therefore of major importance in biotechnological applications that use plant biomass as a substrate. Pentose catabolism is one of the best-studied pathways of primary metabolism of Aspergillus niger, and an initial outline of this pathway with individual enzymes covering each step of the pathway has been previously established. However, although growth on l-arabinose and/or d-xylose of most pentose catabolic pathway (PCP) single deletion mutants of A. niger has been shown to be negatively affected, it was not abolished, suggesting the involvement of additional enzymes. Detailed analysis of the single deletion mutants of the known A. niger PCP genes led to the identification of additional genes involved in the pathway. These results reveal a high level of complexity and redundancy in this pathway, emphasizing the need for a comprehensive understanding of metabolic pathways before entering metabolic engineering of such pathways for the generation of more efficient fungal cell factories., (© 2021 The Authors. Microbial Biotechnology published by Society for Applied Microbiology and John Wiley & Sons Ltd.)

Dichomitus squalens is an emerging reference species that can be used to investigate white-rot fungal plant biomass degradation, as it has flexible physiology to utilize different types of biomass as sources of carbon and energy. Recent comparative (post-) genomic studies on D. squalens resulted in an increasingly detailed knowledge of the genes and enzymes involved in the lignocellulose breakdown in this fungus and showed a complex transcriptional response in the presence of lignocellulose-derived compounds. To fully utilize this increasing amount of data, efficient and reliable genetic manipulation tools are needed, e.g., to characterize the function of certain proteins in vivo and facilitate the construction of strains with enhanced lignocellulolytic capabilities. However, precise genome alterations are often very difficult in wild-type basidiomycetes partially due to extremely low frequencies of homology directed recombination (HDR) and limited availability of selectable markers. To overcome these obstacles, we assessed various Cas9-single guide RNA (sgRNA) ribonucleoprotein (RNP) -based strategies for selectable homology and non-homologous end joining (NHEJ) -based gene editing in D. squalens . We also showed an induction of HDR-based genetic modifications by using single-stranded oligodeoxynucleotides (ssODNs) in a basidiomycete fungus for the first time. This paper provides directions for the application of targeted CRISPR/Cas9-based genome editing in D. squalens and other wild-type (basidiomycete) fungi.

Fungal laccases are attracting enzymes for sustainable valorization of biorefinery lignins. To improve the lignin oxidation capacity of two previously characterized laccase isoenzymes from the white-rot fungus Obba rivulosa, we mutated their substrate-binding site at T1. As a result, the pH optimum of the recombinantly produced laccase variant rOrLcc2-D206N shifted by three units towards neutral pH. O. rivulosa laccase variants with redox mediators oxidized both the dimeric lignin model compound and biorefinery poplar lignin. Significant structural changes, such as selective benzylic α-oxidation, were detected by nuclear magnetic resonance analysis, although no polymerization of lignin was observed by gel permeation chromatography. This suggests that especially rOrLcc2-D206N is a promising candidate for lignin-related applications., (© 2021 The Authors. Microbial Biotechnology published by Society for Applied Microbiology and John Wiley & Sons Ltd.)

d-xylose reductase is a member of the aldo-keto reductase family, and is involved in d-xylose and l-arabinose conversion through the Pentose Catabolic Pathway (PCP) in fungi. In this study, we biochemically characterized a newly identified second d-xylose reductase (XyrB) from Aspergillus niger . This NADPH-dependent reductase is able to efficiently convert d-xylose and l-arabinose, and it has the highest affinity for these sugars of all currently known fungal pentose reductases. A combination of biochemical data, transcriptomics and phylogenetic analysis further illustrated the role of XyrB in the PCP. Enzymes: D-xylose reductase (EC 1.1.1.307), L-arabinose reductase (EC 1.1.1.21)., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2021 The Authors.)

The filamentous ascomycete Aspergillus niger has received increasing interest as a cell factory, being able to efficiently degrade plant cell wall polysaccharides as well as having an extensive metabolism to convert the released monosaccharides into value added compounds. The pentoses D-xylose and L-arabinose are the most abundant monosaccharides in plant biomass after the hexose D-glucose, being major constituents of xylan, pectin and xyloglucan. In this study, the influence of selected pentose catabolic pathway (PCP) deletion strains on growth on plant biomass and re-routing of sugar catabolism was addressed to gain a better understanding of the flexibility of this fungus in using plant biomass-derived monomers. The transcriptome, metabolome and proteome response of three PCP mutant strains, Δ larA Δ xyrA Δ xyrB , Δ ladA Δ xdhA Δ sdhA and Δ xkiA , grown on wheat bran (WB) and sugar beet pulp (SBP), was evaluated. Our results showed that despite the absolute impact of these PCP mutations on pure pentose sugars, they are not as critical for growth of A. niger on more complex biomass substrates, such as WB and SBP. However, significant phenotypic variation was observed between the two biomass substrates, but also between the different PCP mutants. This shows that the high sugar heterogeneity of these substrates in combination with the high complexity and adaptability of the fungal sugar metabolism allow for activation of alternative strategies to support growth., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Chroumpi, Peng, Markillie, Mitchell, Nicora, Hutchinson, Paurus, Tolic, Clendinen, Orr, Baker, Mäkelä and de Vries.)

Carbon catabolite repression enables fungi to utilize the most favourable carbon source in the environment, and is mediated by a key regulator, CreA, in most fungi. CreA-mediated regulation has mainly been studied at high monosaccharide concentrations, an uncommon situation in most natural biotopes. In nature, many fungi rely on plant biomass as their major carbon source by producing enzymes to degrade plant cell wall polysaccharides into metabolizable sugars. To determine the role of CreA when fungi grow in more natural conditions and in particular with respect to degradation and conversion of plant cell walls, we compared transcriptomes of a creA deletion and reference strain of the ascomycete Aspergillus niger during growth on sugar beet pulp and wheat bran. Transcriptomics, extracellular sugar concentrations and growth profiling of A. niger on a variety of carbon sources, revealed that also under conditions with low concentrations of free monosaccharides, CreA has a major effect on gene expression in a strong time and substrate composition dependent manner. In addition, we compared the CreA regulon from five fungi during their growth on crude plant biomass or cellulose. It showed that CreA commonly regulated genes related to carbon metabolism, sugar transport and plant cell wall degrading enzymes across different species. We therefore conclude that CreA has a crucial role for fungi also in adapting to low sugar concentrations as occurring in their natural biotopes, which is supported by the presence of CreA orthologs in nearly all fungi., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2021 The Author(s).)

Salicylic acid plays an important role in the plant immune response, and its degradation is therefore important for plant-pathogenic fungi. However, many nonpathogenic microorganisms can also degrade salicylic acid. In the filamentous fungus Aspergillus niger , two salicylic acid metabolic pathways have been suggested. The first pathway converts salicylic acid to catechol by a salicylate hydroxylase (ShyA). In the second pathway, salicylic acid is 3-hydroxylated to 2,3-dihydroxybenzoic acid, followed by decarboxylation to catechol by 2,3-dihydroxybenzoate decarboxylase (DhbA). A. niger cleaves the aromatic ring of catechol catalyzed by catechol 1,2-dioxygenase (CrcA) to form cis , cis -muconic acid. However, the identification and role of the genes and characterization of the enzymes involved in these pathways are lacking. In this study, we used transcriptome data of A. niger grown on salicylic acid to identify genes ( shyA and crcA ) involved in salicylic acid metabolism. Heterologous production in Escherichia coli followed by biochemical characterization confirmed the function of ShyA and CrcA. The combination of ShyA and CrcA demonstrated that cis,cis -muconic acid can be produced from salicylic acid. In addition, the in vivo roles of shyA , dhbA , and crcA were studied by creating A. niger deletion mutants which revealed the role of these genes in the fungal metabolism of salicylic acid. IMPORTANCE Nonrenewable petroleum sources are being depleted, and therefore, alternative sources are needed. Plant biomass is one of the most abundant renewable sources on Earth and is efficiently degraded by fungi. In order to utilize plant biomass efficiently, knowledge about the fungal metabolic pathways and the genes and enzymes involved is essential to create efficient strategies for producing valuable compounds such as cis , cis -muconic acid. cis , cis -Muconic acid is an important platform chemical that is used to synthesize nylon, polyethylene terephthalate (PET), polyurethane, resins, and lubricants. Currently, cis , cis -muconic acid is mainly produced through chemical synthesis from petroleum-based chemicals. Here, we show that two enzymes from fungi can be used to produce cis , cis -muconic acid from salicylic acid and contributes in creating alternative methods for the production of platform chemicals., (Copyright © 2021 American Society for Microbiology.)

Lignocellulosic biomass is an abundant and renewable resource, and its utilization has become the focus of research and biotechnology applications as a very promising raw material for the production of value-added compounds. Filamentous fungi play an important role in the production of various lignocellulolytic enzymes, while some of them have also been used for the production of important metabolites. However, wild type strains have limited efficiency in enzyme production or metabolic conversion, and therefore many efforts have been made to engineer improved strains. Examples of this are the manipulation of transcriptional regulators and/or promoters of enzyme-encoding genes to increase gene expression, and protein engineering to improve the biochemical characteristics of specific enzymes. This review provides and overview of the applications of filamentous fungi in lignocellulosic biomass based processes and the development and current status of various molecular engineering strategies to improve these processes., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Utilization of lignin-rich side streams has been a focus of intensive studies recently. Combining biocatalytic methods with chemical treatments is a promising approach for sustainable modification of lignocellulosic waste streams. Laccases are catalysts in lignin biodegradation with proven applicability in industrial scale. Laccases directly oxidize lignin phenolic components, and their functional range can be expanded using low-molecular-weight compounds as mediators to include non-phenolic lignin structures. In this work, we studied in detail recombinant laccases from the selectively lignin-degrading white-rot fungus Obba rivulosa for their properties and evaluated their potential as industrial biocatalysts for the modification of wood lignin and lignin-like compounds. We screened and optimized various laccase mediator systems (LMSs) using lignin model compounds and applied the optimized reaction conditions to biorefinery-sourced technical lignin. In the presence of both N-OH-type and phenolic mediators, the O. rivulosa laccases were shown to selectively oxidize lignin in acidic reaction conditions, where a cosolvent is needed to enhance lignin solubility. In comparison to catalytic iron(III)-(2,2,6,6-tetramethylpiperidin-1-yl)oxyl (TEMPO) oxidation systems, the syringyl-type lignin units were preferred in mediated biocatalytic oxidation systems., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2020 Kontro, Maltari, Mikkilä, Kähkönen, Mäkelä, Hildén, Nousiainen and Sipilä.)

Filamentous fungi are important industrial cell factories used for the production of a wide range of enzymes and metabolites. Their primary metabolism is a significant source of industrially important compounds, as well as of monomeric building blocks for the production of secondary metabolites and extracellular enzymes. Therefore, large efforts have been made towards the development of suitable strains for the industrial scale production of primary metabolites. Over the last decades, metabolic engineering of primary metabolism has become a powerful tool to enhance production of both primary and secondary metabolites. This review summarises the different metabolic engineering methods that have been applied to rationally improve the production of industrially relevant primary metabolites in filamentous fungi, and discusses related challenges and future perspectives., (Copyright © 2020. Published by Elsevier Inc.)

Penicillium subrubescens is able to degrade a broad range of plant biomass and it has an expanded set of Carbohydrate Active enzyme (CAZyme)-encoding genes in comparison to other Penicillium species. Here we used exoproteome and transcriptome analysis to demonstrate the versatile plant biomass degradation mechanism by P. subrubescens during growth on wheat bran and sugar beet pulp. On wheat bran P. subrubescens degraded xylan main chain and side residues from Day 2 of cultivation, whereas it started to degrade side chains of pectin in sugar beet pulp prior to attacking the main chain on Day 3. In addition, on Day 3 the cellulolytic enzymes were highly increased. Our results confirm that P. subrubescens adapts its enzyme production to the available plant biomass and is a promising new fungal cell factory for the production of CAZymes., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

Plant biomass degradation by fungi is a widely studied and applied field of science, due to its relevance for the global carbon cycle and many biotechnological applications. Before the genome era, many of the in-depth studies focused on a relatively small number of species, whereas now, many species can be addressed in detail, revealing the large variety in the approach used by fungi to degrade plant biomass. This variation is found at many levels and includes genomic adaptation to the preferred biomass component, but also different approaches to degrade this component by diverse sets of activities encoded in the genome. Even larger differences have been observed using transcriptome and proteome studies, even between closely related species, suggesting a high level of adaptation in individual species. A better understanding of the drivers of this diversity could be highly valuable in developing more efficient biotechnology approaches for the enzymatic conversion of plant biomass., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

The CRISPR/Cas9 system has been successfully applied for gene editing in filamentous fungi. Previous studies reported that single stranded oligonucleotides can be used as repair templates to induce point mutations in some filamentous fungi belonging to genus Aspergillus. In Aspergillus niger, extensive research has been performed on regulation of plant biomass degradation, addressing transcription factors such as XlnR or GaaR, involved in (hemi-)cellulose and pectin utilization, respectively. Single nucleotide mutations leading to constitutively active forms of XlnR and GaaR have been previously reported. However, the mutations were performed by the introduction of versions obtained through site-directed or UV-mutagenesis into the genome. Here we report a more time- and cost-efficient approach to obtaining constitutively active versions by application of the CRISPR/Cas9 system to generate the desired mutation on-site in the A. niger genome. This was also achieved using only 60-mer single stranded oligonucleotides, shorter than the previously reported 90-mer strands. In this study, we show that CRISPR/Cas9 can also be used to efficiently change functional properties of the proteins encoded by the target gene by on-site genomic mutations in A. niger. The obtained strains with constitutively active XlnR and GaaR versions resulted in increased production of plant biomass degrading enzymes and improved release of d-xylose and l-arabinose from wheat bran, and d-galacturonic acid from sugar beet pulp., Competing Interests: Declaration of Competing Interest The authors declare that they have no competing interests., (Copyright © 2020 Elsevier Inc. All rights reserved.)

White-rot (WR) fungi are pivotal decomposers of dead organic matter in forest ecosystems and typically use a large array of hydrolytic and oxidative enzymes to deconstruct lignocellulose. However, the extent of lignin and cellulose degradation may vary between species and wood type. Here, we combined comparative genomics, transcriptomics and secretome proteomics to identify conserved enzymatic signatures at the onset of wood-decaying activity within the Basidiomycota genus Pycnoporus. We observed a strong conservation in the genome structures and the repertoires of protein-coding genes across the four Pycnoporus species described to date, despite the species having distinct geographic distributions. We further analysed the early response of P. cinnabarinus, P. coccineus and P. sanguineus to diverse (ligno)-cellulosic substrates. We identified a conserved set of enzymes mobilized by the three species for breaking down cellulose, hemicellulose and pectin. The co-occurrence in the exo-proteomes of H2O2-producing enzymes with H2O2-consuming enzymes was a common feature of the three species, although each enzymatic partner displayed independent transcriptional regulation. Finally, cellobiose dehydrogenase-coding genes were systematically co-regulated with at least one AA9 lytic polysaccharide monooxygenase gene, indicative of enzymatic synergy in vivo. This study highlights a conserved core white-rot fungal enzymatic mechanism behind the wood-decaying process., (© The Author(s) 2020. Published by Oxford University Press on behalf of Kazusa DNA Research Institute.)

The kraft lignin's low molecular weight and too high hydroxyl content hinder its application in bio-based carbon fibers. In this study, we were able to polymerize kraft lignin and reduce the amount of hydroxyl groups by incubating it with the white-rot fungus Obba rivulosa . Enzymatic radical oxidation reactions were hypothesized to induce condensation of lignin, which increased the amount of aromatic rings connected by carbon-carbon bonds. This modification is assumed to be beneficial when aiming for graphite materials such as carbon fibers. Furthermore, the ratio of remaining aliphatic hydroxyls to phenolic hydroxyls was increased, making the structure more favorable for carbon fiber production. When the modified lignin was mixed together with cellulose, the mixture could be spun into intact precursor fibers by using dry-jet wet spinning. The modified lignin leaked less to the spin bath compared with the unmodified lignin starting material, making the recycling of spin-bath solvents easier. The stronger incorporation of modified lignin in the precursor fibers was confirmed by composition analysis, thermogravimetry, and mechanical testing. This work shows how white-rot fungal treatment can be used to modify the structure of lignin to be more favorable for the production of bio-based fiber materials., Competing Interests: The authors declare no competing financial interest., (Copyright © 2020 American Chemical Society.)

Saprobic fungi, such as Aspergillus niger, grow as colonies consisting of a network of branching and fusing hyphae that are often considered to be relatively uniform entities in which nutrients can freely move through the hyphae. In nature, different parts of a colony are often exposed to different nutrients. We have investigated, using a multi-omics approach, adaptation of A. niger colonies to spatially separated and compositionally different plant biomass substrates. This demonstrated a high level of intra-colony differentiation, which closely matched the locally available substrate. The part of the colony exposed to pectin-rich sugar beet pulp and to xylan-rich wheat bran showed high pectinolytic and high xylanolytic transcript and protein levels respectively. This study therefore exemplifies the high ability of fungal colonies to differentiate and adapt to local conditions, ensuring efficient use of the available nutrients, rather than maintaining a uniform physiology throughout the colony., (© 2019 The Authors. Environmental Microbiology published by Society for Applied Microbiology and John Wiley & Sons Ltd.)

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.

In fungi, L-rhamnose (Rha) is converted via four enzymatic steps into pyruvate and L-lactaldehyde, which enter central carbon metabolism. In Aspergillus niger, only the genes involved in the first three steps of the Rha catabolic pathway have been identified and characterized, and the inducer of the pathway regulator RhaR remained unknown. In this study, we identified the gene (lkaA) involved in the conversion of L-2-keto-3-deoxyrhamnonate (L-KDR) into pyruvate and L-lactaldehyde, which is the last step of the Rha pathway. Deletion of lkaA resulted in impaired growth on L-rhamnose, and potentially in accumulation of L-KDR. Contrary to ΔlraA, ΔlrlA and ΔlrdA, the expression of the Rha-responsive genes that are under control of RhaR, were at the same levels in ΔlkaA and the reference strain, indicating the role of L-KDR as the inducer of the Rha pathway regulator., (Copyright © 2020 Elsevier GmbH. All rights reserved.)

Heterologous production of fungal ligninolytic cocktails is challenging due to the low yields of catalytically active lignin modifying peroxidases. Production using a natural system, such as a wood-rotting fungus, is a promising alternative if specific or preferential induction of the ligninolytic activities could be achieved. Using transcriptomics, gene expression of the white-rot Dichomitus squalens during growth on mixtures of aromatic compounds, with ring structures representing the two major lignin sub-units, was compared to a wood substrate. Most of the genes encoding lignin modifying enzymes (laccases and peroxidases) categorised as highly or moderately expressed on wood were expressed similarly on aromatic compounds. Higher expression levels of a subset of manganese and versatile peroxidases was observed on di- compared to mono-methoxylated aromatics. The expression of polysaccharide degrading enzymes was lower on aromatic compounds compared to wood, demonstrating that the induction of lignin modifying enzymes became more specific. This study suggests potential for aromatic waste streams, e.g. from lignocellulose pretreatment, to produce a lignin-specific enzyme cocktail from D. squalens or other white-rot fungi., (Copyright © 2019 Elsevier B.V. All rights reserved.)

α-Galactosidases are important industrial enzymes for hemicellulosic biomass degradation or modification. In this study, six novel extracellular α-galactosidases from Penicillium subrubescens were produced in Pichia pastoris and characterized. All α-galactosidases exhibited high affinity to pNPαGal, and only AglE was not active towards galacto-oligomers. Especially AglB and AglD released high amounts of galactose from guar gum, carob galactomannan and locust bean, but combining α-galactosidases with an endomannanase dramatically improved galactose release. Structural comparisons to other α-galactosidases and homology modelling showed high sequence similarities, albeit significant differences in mechanisms of productive binding, including discrimination between various galactosides. To our knowledge, this is the first study of such an extensive repertoire of extracellular fungal α-galactosidases, to demonstrate their potential for degradation of galactomannan-rich biomass. These findings contribute to understanding the differences within glycoside hydrolase families, to facilitate the development of new strategies to generate tailor-made enzymes for new industrial bioprocesses., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

Aromatic compounds derived from lignin are of great interest for renewable biotechnical applications. They can serve in many industries e.g. as biochemical building blocks for bioplastics or biofuels, or as antioxidants, flavor agents or food preservatives. In nature, lignin is degraded by microorganisms, which results in the release of homocyclic aromatic compounds. Homocyclic aromatic compounds can also be linked to polysaccharides, tannins and even found freely in plant biomass. As these compounds are often toxic to microbes already at low concentrations, they need to be degraded or converted to less toxic forms. Prior to ring cleavage, the plant- and lignin-derived aromatic compounds are converted to seven central ring-fission intermediates, i.e. catechol, protocatechuic acid, hydroxyquinol, hydroquinone, gentisic acid, gallic acid and pyrogallol through complex aromatic metabolic pathways and used as energy source in the tricarboxylic acid cycle. Over the decades, bacterial aromatic metabolism has been described in great detail. However, the studies on fungal aromatic pathways are scattered over different pathways and species, complicating a comprehensive view of fungal aromatic metabolism. In this review, we depicted the similarities and differences of the reported aromatic metabolic pathways in fungi and bacteria. Although both microorganisms share the main conversion routes, many alternative pathways are observed in fungi. Understanding the microbial aromatic metabolic pathways could lead to metabolic engineering for strain improvement and promote valorization of lignin and related aromatic compounds., (Copyright © 2019 Elsevier Inc. All rights reserved.)

The extent of carbon catabolite repression (CCR) at a global level is unknown in wood-rotting fungi, which are critical to the carbon cycle and are a source of biotechnological enzymes. CCR occurs in the presence of sufficient concentrations of easily metabolizable carbon sources (e.g., glucose) and involves downregulation of the expression of genes encoding enzymes involved in the breakdown of complex carbon sources. We investigated this phenomenon in the white-rot fungus Dichomitus squalens using transcriptomics and exoproteomics. In D. squalens cultures, approximately 7% of genes were repressed in the presence of glucose compared to Avicel or xylan alone. The glucose-repressed genes included the essential components for utilization of plant biomass- c arbohydrate- a ctive en zyme (CAZyme) and carbon catabolic genes. The majority of polysaccharide-degrading CAZyme genes were repressed and included activities toward all major carbohydrate polymers present in plant cell walls, while repression of ligninolytic genes also occurred. The transcriptome-level repression of the CAZyme genes observed on the Avicel cultures was strongly supported by exoproteomics. Protease-encoding genes were generally not glucose repressed, indicating their likely dominant role in scavenging for nitrogen rather than carbon. The extent of CCR is surprising, given that D. squalens rarely experiences high free sugar concentrations in its woody environment, and it indicates that biotechnological use of D. squalens for modification of plant biomass would benefit from derepressed or constitutively CAZyme-expressing strains. IMPORTANCE White-rot fungi are critical to the carbon cycle because they can mineralize all wood components using enzymes that also have biotechnological potential. The occurrence of carbon catabolite repression (CCR) in white-rot fungi is poorly understood. Previously, CCR in wood-rotting fungi has only been demonstrated for a small number of genes. We demonstrated widespread glucose-mediated CCR of plant biomass utilization in the white-rot fungus Dichomitus squalens This indicates that the CCR mechanism has been largely retained even though wood-rotting fungi rarely experience commonly considered CCR conditions in their woody environment. The general lack of repression of genes encoding proteases along with the reduction in secreted CAZymes during CCR suggested that the retention of CCR may be connected with the need to conserve nitrogen use during growth on nitrogen-scarce wood. The widespread repression indicates that derepressed strains could be beneficial for enzyme production., (Copyright © 2019 American Society for Microbiology.)

Fungal strain engineering is commonly used in many areas of biotechnology, including the production of plant biomass degrading enzymes. Its aim varies from the production of specific enzymes to overall increased enzyme production levels and modification of the composition of the enzyme set that is produced by the fungus. Strain engineering involves a diverse range of methodologies, including classical mutagenesis, genetic engineering and genome editing. In this review, the main approaches for strain engineering of filamentous fungi in the field of plant biomass degradation will be discussed, including recent and not yet implemented methods, such as CRISPR/Cas9 genome editing and adaptive evolution., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Lignocellulosic plant biomass is an important feedstock for bio-based economy. In particular, it is an abundant renewable source of aromatic compounds, which are present as part of lignin, as side-groups of xylan and pectin, and in other forms, such as tannins. As filamentous fungi are the main organisms that modify and degrade lignocellulose, they have developed a versatile metabolism to convert the aromatic compounds that are toxic at relatively low concentrations to less toxic ones. During this process, fungi form metabolites some of which represent high-value platform chemicals or important chemical building blocks, such as benzoic, vanillic, and protocatechuic acid. Especially basidiomycete white-rot fungi with unique ability to degrade the recalcitrant lignin polymer are expected to perform highly efficient enzymatic conversions of aromatic compounds, thus having huge potential for biotechnological exploitation. However, the aromatic metabolism of basidiomycete fungi is poorly studied and knowledge on them is based on the combined results of studies in variety of species, leaving the overall picture in each organism unclear. Dichomitus squalens is an efficiently wood-degrading white-rot basidiomycete that produces a diverse set of extracellular enzymes targeted for lignocellulose degradation, including oxidative enzymes that act on lignin. Our recent study showed that several intra- and extracellular aromatic compounds were produced when D. squalens was cultivated on spruce wood, indicating also versatile aromatic metabolic abilities for this species. In order to provide the first molecular level systematic insight into the conversion of plant biomass derived aromatic compounds by basidiomycete fungi, we analyzed the transcriptomes of D. squalens when grown with 10 different lignocellulose-related aromatic monomers. Significant differences for example with respect to the expression of lignocellulose degradation related genes, but also putative genes encoding transporters and catabolic pathway genes were observed between the cultivations supplemented with the different aromatic compounds. The results demonstrate that the transcriptional response of D. squalens is highly dependent on the specific aromatic compounds present suggesting that instead of a common regulatory system, fine-tuned regulation is needed for aromatic metabolism., (Copyright © 2019 Kowalczyk, Peng, Pawlowski, Lipzen, Ng, Singan, Wang, Grigoriev and Mäkelä.)

Guar gum consists mainly of galactomannan and constitutes the endosperm of guar seeds that acts as a reserve polysaccharide for germination. Due to its molecular structure and physical properties, this biopolymer has been considered as one of the most important and widely used gums in industry. However, for many of these applications this (hemi-)cellulosic structure needs to be modified or (partially) depolymerized in order to customize and improve its physicochemical properties. In this study, transcriptome, exoproteome and enzyme activity analyses were employed to decipher the complete enzymatic arsenal for guar gum depolymerization by Aspergillus niger. This multi-omic analysis revealed a set of 46 genes encoding carbohydrate-active enzymes (CAZymes) responding to the presence of guar gum, including CAZymes not only with preferred activity towards galactomannan, but also towards (arabino-)xylan, cellulose, starch and pectin, likely due to trace components in guar gum. This demonstrates that the purity of substrates has a strong effect on the resulting enzyme mixture produced by A. niger and probably by other fungi as well, which has significant implications for the commercial production of fungal enzyme cocktails., (Crown Copyright © 2019. Published by Elsevier B.V. All rights reserved.)

Here, we report the draft genome sequences of three isolates of the wood-decaying white-rot basidiomycete fungus Dichomitus squalens The genomes of these monokaryons were sequenced to provide more information on the intraspecies genomic diversity of this fungus and were compared to the previously sequenced genome of D. squalens LYAD-421 SS1.

Species in the genus Paecilomyces , a member of the fungal order Eurotiales, are ubiquitous in nature and impact a variety of human endeavors. Here, the biology of one common species, Paecilomyces variotii , was explored using genomics and functional genetics. Sequencing the genome of two isolates revealed key genome and gene features in this species. A striking feature of the genome was the two-part nature, featuring large stretches of DNA with normal GC content separated by AT-rich regions, a hallmark of many plant-pathogenic fungal genomes. These AT-rich regions appeared to have been mutated by repeat-induced point (RIP) mutations. We developed methods for genetic transformation of P. variotii , including forward and reverse genetics as well as crossing techniques. Using transformation and crossing, RIP activity was identified, demonstrating for the first time that RIP is an active process within the order Eurotiales. A consequence of RIP is likely reflected by a reduction in numbers of genes within gene families, such as in cell wall degradation, and reflected by growth limitations on P. variotii on diverse carbon sources. Furthermore, using these transformation tools we characterized a conserved protein containing a domain of unknown function (DUF1212) and discovered it is involved in pigmentation.

Aspergillus section Nigri comprises filamentous fungi relevant to biomedicine, bioenergy, health, and biotechnology. To learn more about what genetically sets these species apart, as well as about potential applications in biotechnology and biomedicine, we sequenced 23 genomes de novo, forming a full genome compendium for the section (26 species), as well as 6 Aspergillus niger isolates. This allowed us to quantify both inter- and intraspecies genomic variation. We further predicted 17,903 carbohydrate-active enzymes and 2,717 secondary metabolite gene clusters, which we condensed into 455 distinct families corresponding to compound classes, 49% of which are only found in single species. We performed metabolomics and genetic engineering to correlate genotypes to phenotypes, as demonstrated for the metabolite aurasperone, and by heterologous transfer of citrate production to Aspergillus nidulans. Experimental and computational analyses showed that both secondary metabolism and regulation are key factors that are significant in the delineation of Aspergillus species.

White-rot fungi, such as Dichomitus squalens, degrade all wood components and inhabit mixed-wood forests containing both soft- and hardwood species. In this study, we evaluated how D. squalens responded to the compositional differences in softwood [guaiacyl (G) lignin and higher mannan content] and hardwood [syringyl/guaiacyl (S/G) lignin and higher xylan content] using semi-natural solid cultures. Spruce (softwood) and birch (hardwood) sticks were degraded by D. squalens as measured by oxidation of the lignins using 2D-NMR. The fungal response as measured by transcriptomics, proteomics and enzyme activities showed a partial tailoring to wood composition. Mannanolytic transcripts and proteins were more abundant in spruce cultures, while a proportionally higher xylanolytic activity was detected in birch cultures. Both wood types induced manganese peroxidases to a much higher level than laccases, but higher transcript and protein levels of the manganese peroxidases were observed on the G-lignin rich spruce. Overall, the molecular responses demonstrated a stronger adaptation to the spruce rather than birch composition, possibly because D. squalens is mainly found degrading softwoods in nature, which supports the ability of the solid wood cultures to reflect the natural environment., (© 2018 The Authors. Environmental Microbiology published by Society for Applied Microbiology and John Wiley & Sons Ltd.)