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Aspergillus fumigatus causes a range of human and animal diseases collectively known as aspergillosis. A . fumigatus possesses and expresses a range of genetic determinants of virulence, which facilitate colonisation and disease progression, including the secretion of mycotoxins. Gliotoxin (GT) is the best studied A . fumigatus mycotoxin with a wide range of known toxic effects that impair human immune cell function. GT is also highly toxic to A . fumigatus and this fungus has evolved self-protection mechanisms that include (i) the GT efflux pump GliA, (ii) the GT neutralising enzyme GliT, and (iii) the negative regulation of GT biosynthesis by the bis -thiomethyltransferase GtmA. The transcription factor (TF) RglT is the main regulator of GliT and this GT protection mechanism also occurs in the non-GT producing fungus A . nidulans . However, the A . nidulans genome does not encode GtmA and GliA. This work aimed at analysing the transcriptional response to exogenous GT in A . fumigatus and A . nidulans , two distantly related Aspergillus species, and to identify additional components required for GT protection. RNA-sequencing shows a highly different transcriptional response to exogenous GT with the RglT-dependent regulon also significantly differing between A . fumigatus and A . nidulans . However, we were able to observe homologs whose expression pattern was similar in both species (43 RglT-independent and 11 RglT-dependent). Based on this approach, we identified a novel RglT-dependent methyltranferase, MtrA, involved in GT protection. Taking into consideration the occurrence of RglT-independent modulated genes, we screened an A . fumigatus deletion library of 484 transcription factors (TFs) for sensitivity to GT and identified 15 TFs important for GT self-protection. Of these, the TF KojR, which is essential for kojic acid biosynthesis in Aspergillus oryzae , was also essential for virulence and GT biosynthesis in A . fumigatus , and for GT protection in A . fumigatus , A . nidulans , and A . oryzae . KojR regulates rglT , gliT , gliJ expression and sulfur metabolism in Aspergillus species. Together, this study identified conserved components required for GT protection in Aspergillus species.

ß-tubulin, calmodulin, internal transcribed spacer and partial Isu-rDNA, RNA polymerase 2, DNA replication licensing factor Mcm7 , and pre-rRNA processing protein Tsr1 were amplified and sequenced from numerous isolates belonging to Aspergillus sect, versicolor. The isolates were analyzed phylogenetically using the concordance model to establish species boundaries. Aspergillus austroafricanus, A. creber, A. cvjetkovicii, A. fructus, A. jensenii, A. puulaauensis, A. subversicolor, A. tennesseensis and A. venenatus are described as new species and A. amoenus, A. protuberus, A. sydowii, A. tabacinus and A. versicolor are accepted as distinct species on the basis of molecular and phenotypic differences. PCR primer pairs used to detect A. versicolor in sick building syndrome studies have a positive reaction for all of the newly described species except A. subversicolor.

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.

Heavy metals (HMs) contamination is a major issue produced by industrial and mining processes, among other human activities. The capacity of fungi to eliminate HMs from the environment has drawn attention. However, the main process by which fungi protect the environment against the damaging effects of these HMs, such as cadmium (Cd), is still unknown. In this study, some fungi were isolated from HMs-polluted soil. The minimum inhibitory concentrations (MICs) and the tolerance indices of the tested isolates against Cd were evaluated. Moreover, molecular identification of the most tolerant fungal isolates ( Aspergillus niger and A. terreus ) was done and deposited in the GenBank NCBI database. The results showed that the colony diameter of A. niger and A. terreus was decreased gradually by the increase of Cd concentration. Also, all the tested parameters were influenced by Cd concentration. Lipid peroxidation (MDA content) was progressively increased by 12.95–105.95% ( A. niger ) and 17.27–85.38% ( A. terreus ), respectively, from 50 to 200 mg/L. PPO, APX, and POD enzymes were elevated in the presence of Cd, thus illustrating the appearance of an oxidative stress action. Compared to the non-stressed A. niger , the POD and PPO activities were enhanced by 92.00 and 104.24% at 200 mg/L Cd. Also, APX activity was increased by 58.12% at 200 mg/L. Removal efficiency and microbial accumulation capacities of A. niger and A. terreus have also been assessed. Production of succinic and malic acids by A. niger and A. terreus was increased in response to 200 mg/L Cd, in contrast to their controls (Cd-free), as revealed by HPLC analysis. These findings helped us to suggest A. niger and A. terreus as the potential mycoremediation microbes that alleviate Cd contamination. We can learn more about these fungal isolates’ resistance mechanisms against different HMs through further studies.

Various strains of Aspergillus oryzae , regarded as a domesticated variant of aflatoxigenic Aspergillus flavus , are utilized in the soybean fermentation industry of Korea. This study compared A. oryzae/flavus strains isolated from various environments in Korea including industrial settings, Meju (brick of dried fermented soybeans), and wild conditions with globally reported strains using genomic analysis to determine their taxonomic positions and risk of mycotoxicity. Using population genomics, five distinct groups (A to E) were identified, with all aflatoxigenic Korean strains in Group C and non-aflatoxigenic Korean strains in Groups A, B, and E. Korean strains from Meju and wild conditions are distributed across Groups A and B, and most of the Korean industrial strains form a sub-cluster with Japanese industrial strains in Group A. Comparing secondary metabolite gene cluster mutation pattern, three gene clusters (Aflatoxin, Cyclopiazonic acid and Ditryptophenaline) were revealed as group specific ones. In aflatoxin and cyclopiazonic acid clusters, most of the Group C strains had intact regions compared to strains in other groups. Since most of the Group C strains produce aflatoxin and have intact Aflatoxin and Cyclopiazonic acid gene clusters, we considered that this group represent A. flavus . Profiling using MALDI-TOF MS analysis also distinguished Group C from Groups A, B and E by specific three proteomic peaks. Among the three peaks, those around 12,700 to 12,900 m/z (Da) are expected to correspond to AflF (nor B), an enzyme involved in Aflatoxin metabolism. These results showed taxonomic positions of Korean strains of A. oryzae/flavus from various environments and also showed possibility to differentiate between A. oryzae and A. flavus with genome and MALDI-TOF MS analysis.

The aspergilli comprise a diverse group of filamentous fungi spanning over 200 million years of evolution. Here we report the genome sequence of the model organism Aspergillus nidulans, and a comparative study with Aspergillus fumigatus, a serious human pathogen, and Aspergillus oryzae, used in the production of sake, miso and soy sauce. Our analysis of genome structure provided a quantitative evaluation of forces driving long-term eukaryotic genome evolution. It also led to an experimentally validated model of mating-type locus evolution, suggesting the potential for sexual reproduction in A. fumigatus and A. oryzae. Our analysis of sequence conservation revealed over 5,000 non-coding regions actively conserved across all three species. Within these regions, we identified potential functional elements including a previously uncharacterized TPP riboswitch and motifs suggesting regulation in filamentous fungi by Puf family genes. We further obtained comparative and experimental evidence indicating widespread translational regulation by upstream open reading frames. These results enhance our understanding of these widely studied fungi as well as provide new insight into eukaryotic genome evolution and gene regulation.

Galactofuranose (Galf)-containing glycostructures are important to secure the integrity of the fungal cell wall. Golgi-localized Galf-transferases (Gfs) have been identified in Aspergillus nidulans and Aspergillus fumigatus. BLASTp searches identified three putative Galf-transferases in Aspergillus niger. Phylogenetic analysis showed that they group in three distinct groups. Characterization of the three Galf-transferases in A. niger by constructing single, double, and triple mutants revealed that gfsA is most important for Galf biosynthesis. The growth phenotypes of the ΔgfsA mutant are less severe than that of the ΔgfsAC mutant, indicating that GfsA and GfsC have redundant functions. Deletion of gfsB did not result in any growth defect and combining ΔgfsB with other deletion mutants did not exacerbate the growth phenotype. RT-qPCR experiments showed that induction of the agsA gene was higher in the ΔgfsAC and ΔgfsABC compared to the single mutants, indicating a severe cell wall stress response after multiple gfs gene deletions.

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.

Members of Aspergillus section Nigri (formerly A. niger group) are distributed worldwide and are regarded as common food spoilage fungi. Some of them are widely used and studied for industrial purposes. They are common sources of extracellular enzymes and organic acids to be used in food processing and are also used in the production of traditional foods, especially in the Orient. Products produced by strains of Aspergillus niger hold the GRAS (Generally Recognised As Safe) status from the FDA. However some species in Aspergillus section Nigri can produce ochratoxin A, a nephrotoxic mycotoxin. In spite of their industrial importance, the taxonomy of black aspergilli ( Aspergillus section Nigri ) is not clear and many attempts have been made in order to find suitable taxonomic criteria. The aim of this paper is to provide an overview of the significance of black aspergilli focusing on all the approaches made in the taxonomy of this group of fungi. Some species, such as A. carbonarius and uniseriate species can be easily recognised. In the A. niger aggregate, although speciation at molecular level has been proposed, no morphological differences can be observed and species identification will therefore remain problematic. Phylogenetic analyses of ITS and 5.8S rDNA gene region of representative black Aspergillus species and a simple key to the most common species that can be easily distinguished by morphological criteria are also included.

BackgroundInvasive fungal disease (IFD) frequently occurs in febrile neutropenic haemato-oncology patients (NHP). Until 2012 no biomarkers were available for the diagnosis of IFD in our Trust. The neutropenic fever pathway was modified to include serial serum Aspergillus galactomannan (GM), serum Aspergillus PCR (APCR) from national reference laboratory and Bronchoalveolar lavage (BAL) GM and APCR.We compared 299 NHP who were investigated with the original pathway between October 2009 and April 2012 with 307 NHP investigated with the novel pathway between April 2013 and 2015. Primary end point was non-inferiority of novel pathway in terms of 12 month mortality. Secondary outcomes were 30 day mortality, length of treatment, length of stay and confidence of diagnosis based on EORTC/MSG criteria1 and concordance of the different biomarkers.MethodProspective patients (2013-15 cohort) were identified from ward lists. Retrospective patients (2009-12 cohort) were identified from haemato-oncology patients who had had blood cultures and were co-incidentally found to be neutropenic. Medical notes, drug charts, discharge letters and microbiology results were reviewed.ResultsThe 2 cohorts were well matched in terms of haemato-oncology diagnosis. The 2009/12 cohort included 561 episodes, with 333 CT chests and 62 bronchospies compared to 508 episodes, 288 CT chests and 86 bronchoscopies in 2013/15 group. 12 month mortality was 42% in 2009/12 versus 34% in 2013/15 cohort. 30 day mortality was 11% for both cohorts. There was no significant difference in length of antifungal treatments, although 24% switched to voriconazole following positive biomarkers.Concordance between serum and BAL GM was 14.8% and APCR was 6.3%. Concordance between serum GM and APCR was 7% and between BAL GM and PCR was 41%.Confidence in diagnosis of IFD increased with the novel pathway See Figure 1.ConclusionThe introduction of a novel NF pathway was found to be non-inferior in terms of 12 month and 30 day mortality. Although there was increased confidence in the diagnosis of IFD, this did not translate to reduced antifungal treatment, although it did influence switching to voriconazole and secondary chemoprophylaxis. Negative serum GM and PCR did not rule out the diagnosis of IFD and BAL biomarkers were more sensitive than serum ones.Reference De Pauw B, et al. Clin Infect Dise 2008; 46(12).[Figure]