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The Evolution of the Satratoxin and Atranone Gene Clusters of Stachybotrys chartarum.

ABSTRACT: Stachybotrys chartarum is frequently isolated from damp building materials or improperly stored animal forage. Human and animal exposure to the secondary metabolites of this mold is linked to severe health effects. The mutually exclusive production of either satratoxins or atranones defines the chemotypes A and S. Based upon the genes (satratoxin cluster, SC1-3, sat or atranone cluster, AC1, atr) that are supposed to be essential for satratoxin and atranone production, S. chartarum can furthermore be divided into three genotypes: the S-type possessing all sat- but no atr-genes, the A-type lacking the sat- but harboring all atr-genes, and the H-type having only certain sat- and all atr-genes. We analyzed the above-mentioned gene clusters and their flanking regions to shed light on the evolutionary relationship. Furthermore, we performed a deep re-sequencing and LC-MS/MS (Liquid chromatography-mass spectrometry) analysis. We propose a first model for the evolution of the S. chartarum genotypes. We assume that genotype H represents the most ancient form. A loss of the AC1 and the concomitant acquisition of the SC2 led to the emergence of the genotype S. According to our model, the genotype H also developed towards genotype A, a process that was accompanied by a loss of SC1 and SC3.

SUBMITTER: Ulrich S

PROVIDER: S-EPMC9027890 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Truncated satratoxin gene clusters in selected isolates of the atranone chemotype of Stachybotrys chartarum (Ehrenb.) S. Hughes.

Project description:The fungus Stachybotrys (S.) chartarum was isolated from culinary herbs, damp building materials, and improperly stored animal forage. Two distinct chemotypes of the fungus were described that produced either high-cytotoxic macrocyclic trichothecenes (S type) or low-cytotoxic atranones (A type). Recently, two distinct gene clusters were described that were found to be necessary for the biosynthesis of either macrocyclic trichothecenes (21 SAT (Satratoxin) genes) or atranones (14 ATR (Atranone) genes). In the current study, PCR primers were designed to detect SAT and ATR genes in 19 S. chartarum chemotype S and eight S. chartarum chemotype A strains. Our analysis revealed the existence of three different genotypes: satratoxin-producing strains that harbored all SAT genes but lacked the ATR gene cluster (genotype S), non-satratoxin-producing strains that possessed the ATR genes but lacked SAT genes (genotype A), and a hitherto undescribed hybrid genotype among non-satratoxin-producing strains that harbored all ATR genes and an incomplete set of SAT genes (genotype H). In order to improve the discrimination of genotypes, a triplex PCR assay was developed and applied for the analysis of S. chartarum and S. chlorohalonata cultures. The results show that genes for macrocyclic trichothecenes and atranones are not mutually exclusive in S. chartarum. Correlation of the new genotype-based concept with mycotoxin production data shows also that macrocyclic trichothecenes are exclusively produced by S. chartarum genotype S strains.

| S-EPMC6971138 | biostudies-literature

Correction to: truncated satratoxin gene clusters in selected isolates of the atranone chemotype of Stachybotrys chartarum (Ehrenb.) S. Hughes.

Project description:The original version of this paper was published with error. The supplementary materials originally provided were not captured during article production.

| S-EPMC7645536 | biostudies-literature

Production of Satratoxin G and H Is Tightly Linked to Sporulation in <i>Stachybotrys chartarum</i>.

Project description:Stachybotrys chartarum is a toxigenic fungus that is frequently isolated from damp building materials or improperly stored forage. Macrocyclic trichothecenes and in particular satratoxins are the most potent mycotoxins known to be produced by this fungus. Exposure of humans or animals to these secondary metabolites can be associated with severe health problems. To assess the pathogenic potential of S. chartarum isolates, it is essential to cultivate them under conditions that reliably promote toxin production. Potato dextrose agar (PDA) was reported to be the optimal nutrition medium for satratoxin production. In this study, the growth of S. chartarum genotype S strains on PDA from two manufacturers led to divergent results, namely, well-grown and sporulating cultures with high satratoxin concentrations (20.8 ± 0.4 µg/cm2) versus cultures with sparse sporulation and low satratoxin production (0.3 ± 0.1 µg/cm2). This finding is important for any attempt to identify toxigenic S. chartarum isolates. Further experiments performed with the two media provided strong evidence for a link between satratoxin production and sporulation. A comparison of three-point and one-point cultures grown on the two types of PDA, furthermore, demonstrated an inter-colony communication that influences both sporulation and mycotoxin production of S. chartarum genotype S strains.

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Stachybotrys chartarum

Project description:Stachybotrys chartarum overview

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Stachybotrys chartarum-induced hypersensitivity pneumonitis is TLR9 dependent.

Project description:Hypersensitivity pneumonitis (HP), an inflammatory lung disease, develops after repeated exposure to inhaled particulate antigen and is characterized by a vigorous T helper type 1-mediated immune response, resulting in the release of IL-12 and interferon (IFN)-?. These T helper type 1 cytokines may participate in the pathogenesis of HP. Stachybotrys chartarum (SC) is a dimorphic fungus implicated in a number of respiratory illnesses, including HP. Here, we have developed a murine model of SC-induced HP that reproduces pathology observed in human HP and hypothesized that toll receptor-like 9 (TLR9)-mediated dendritic cell responses are required for the generation of granulomatous inflammation induced by inhaled SC. Mice sensitized and challenged with 10(6) SC spores develop granulomatous inflammation with multinucleate giant cells, accompanied by increased accumulation of neutrophils and CD4(+) and CD8(+) T cells. SC sensitization and challenge resulted in robust pulmonary expression of tumor necrosis factor-?, IL-12, and IFN-?. SC-mediated granulomatous inflammation required IFN-? and was TLR9 dependent, because TLR9(-/-) mice displayed reduced peribronchial inflammation, decreased accumulation and/or activation of polymorphonuclear (PMN) and CD4(+) and CD8(+) T cells, and reduced lung expression of type 1 cytokines and chemokines. T-cell production of IFN-? was IL-12 dependent. Our studies suggest that TLR9 is critical for dendritic cell-mediated development of a type 1 granulomatous inflammation in the lung in response to SC.

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Strain differences influence murine pulmonary responses to Stachybotrys chartarum.

Project description:When the fungus Stachybotrys chartarum is inhaled, its mycotoxins may cause lung injury and inflammation. The severity of human responses to S. chartarum in both occupational and home settings varies widely. To explore these differences, we intratracheally instilled C3H/HeJ, BALB/c, and C57BL/6J mice with S. chartarum spores suspended in saline. One day later, the mice were humanely killed, bronchoalveolar lavage (BAL) was performed, and biochemical and cellular indicators of lung injury and inflammation were measured. BALB/c mice showed the highest myeloperoxidase activity, albumin and hemoglobin levels, and neutrophil numbers in their BAL among the three strains. BALB/c was the only strain to show significant increases in keratinocyte-derived cytokine (KC), monocyte chemotactic protein (MCP)-1, MCP-3, macrophage inflammatory protein (MIP)-1alpha, MIP-1beta, MIP-1gamma, MIP-2, RANTES, IL-1alpha, IL-1beta, IL-3, IL-6, IL-18, leukemia inhibitory factor, macrophage colony-stimulating factor, and TNF-alpha. A model of allergen-induced airway inflammation was examined to assess whether underlying allergic inflammation might contribute to increased susceptibility to S. chartarum-induced pulmonary inflammation and injury. Surprisingly, in BALB/c mice, ovalbumin-induced airway inflammation produced a protective effect against some S. chartarum-induced pulmonary responses. This is the first report of mammalian strain differences affecting responses to S. chartarum. These responses differ from those reported for LPS and other fungi. Analogous underlying genetic differences may contribute to the wide range of sensitivity to Stachybotrys among humans.

| S-EPMC2643262 | biostudies-literature

Biomechanics of conidial dispersal in the toxic mold Stachybotrys chartarum.

Project description:Conidial dispersal in Stachybotrys chartarum in response to low-velocity airflow was studied using a microflow apparatus. The maximum rate of spore release occurred during the first 5 min of airflow, followed by a dramatic reduction in dispersal that left more than 99% of the conidia attached to their conidiophores. Micromanipulation of undisturbed colonies showed that micronewton (microN) forces were needed to dislodge spore clusters from their supporting conidiophores. Calculations show that airspeeds that normally prevail in the indoor environment disturb colonies with forces that are 1000-fold lower, in the nanonewton (nN) range. Low-velocity airflow does not, therefore, cause sufficient disturbance to disperse a large proportion of the conidia of S. chartarum.

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Inflammatory cytokine gene expression in THP-1 cells exposed to Stachybotrys chartarum and Aspergillus versicolor.

Project description:Very little is known about the mechanisms that occur in human cells upon exposure to fungi as well as their mycotoxins. A better understanding of toxin-regulated gene expression would be helpful to identify safe levels of exposure and could eventually be the basis for establishing guidelines for remediation scenarios following a water intrusion event. In this research, cytokine mRNA expression patterns were investigated in the human monocytic THP-1 cell line exposed to fungal extracts of various fragment sizes obtained from Stachybotrys chartarum RTI 5802 and/or Aspergillus versicolor RTI 3843, two common and well-studied mycotoxin producing fungi. Cytokine mRNA expression was generally upregulated 2-10 times following a 24 h exposure to fungal extracts. Expression of the proinflammatory interleukin-1?, interleukin-8, and tumor necrosis factor-? genes increased while the anti-inflammatory gene interleukin-10 also increased albeit at very low level, suggesting that negative feedback regulation mechanism of production of proinflammatory cytokines initiated upon 24 h of incubation. In addition, submicron size extracts of A. versicolor caused significant death of THP-1 cells, whereas extracts of S. chartarum caused no cell death while the mixture of the two fungi had an intermediate effect. There was no general correlation between gene expression and fragment sizes, which suggests that all submicron fragments may contribute to inflammatory response.

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Inhalation of <i>Stachybotrys chartarum</i> Fragments Induces Pulmonary Arterial Remodeling.

Project description:Stachybotrys chartarum is a fungal contaminant within the built environment and a respiratory health concern in the United States. The objective of this study was to characterize the mechanisms influencing pulmonary immune responses to repeatedly inhaled S. chartarum. Groups of B6C3F1/N mice repeatedly inhaled viable trichothecene-producing S. chartarum conidia (strain A or strain B), heat-inactivated conidia, or high-efficiency particulate absolute-filtered air twice per week for 4 and 13 weeks. Strain A was found to produce higher amounts of respirable fragments than strain B. Lung tissue, serum, and BAL fluid were collected at 24 and 48 hours after final exposure and processed for histology, flow cytometry, and RNA and proteomic analyses. At 4 weeks after exposure, a T-helper cell type 2-mediated response was observed. After 13 weeks, a mixed T-cell response was observed after exposure to strain A compared with a T-helper cell type 2-mediated response after strain B exposure. After exposure, both strains induced pulmonary arterial remodeling at 13 weeks; however, strain A-exposed mice progressed more quickly than strain B-exposed mice. BAL fluid was composed primarily of eosinophils, neutrophils, and macrophages. Both the immune response and the observed pulmonary arterial remodeling were supported by specific cellular, molecular, and proteomic profiles. The immunopathological responses occurred earlier in mice exposed to high fragment-producing strain A. The rather striking induction of pulmonary remodeling by S. chartarum appears to be related to the presence of fungal fragments during exposure.

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Toxin Production by Stachybotrys chartarum Genotype S on Different Culture Media.

Project description:Stachybotrys (S.) chartarum had been linked to severe health problems in humans and animals, which occur after exposure to the toxic secondary metabolites of this mold. S. chartarum had been isolated from different environmental sources, ranging from culinary herbs and improperly stored fodder to damp building materials. To access the pathogenic potential of isolates, it is essential to analyze them under defined conditions that allow for the production of their toxic metabolites. All Stachybotrys species are assumed to produce the immunosuppressive phenylspirodrimanes, but the highly cytotoxic macrocyclic trichothecenes are exclusively generated by the genotype S of S. chartarum. In this study, we have analyzed four genotype S strains initially isolated from three different habitats. We grew them on five commonly used media (malt-extract-agar, glucose-yeast-peptone-agar, potato-dextrose-agar, cellulose-agar, Sabouraud-dextrose-agar) to identify conditions that promote mycotoxin production. Using LC-MS/MS, we have quantified stachybotrylactam and all S-type specific macrocyclic trichothecenes (satratoxin G, H, F, roridin E, L-2, verrucarin J). All five media supported a comparable fungal growth and sporulation at 25 °C in the dark. The highest concentrations of macrocyclic trichothecenes were detected on potato-dextrose-agar or cellulose-agar. Malt-extract-agar let to an intermediate and glucose-yeast-peptone-agar and Sabouraud-dextrose-agar to a poor mycotoxin production. These data demonstrate that the mycotoxin production clearly depends on the composition of the respective medium. Our findings provide a starting point for further studies in order to identify individual components that either support or repress the production of mycotoxins in S. chartarum.

| S-EPMC7559122 | biostudies-literature

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