Project description:BACKGROUND: Stachybotrys chartarum is a filamentous mold frequently identified among the mycobiota of water-damaged building materials. Growth of S. chartarum on suitable substrates and under favorable environmental conditions leads to the production of secondary metabolites such as mycotoxins and microbial volatile organic compounds (MVOCs). The aim of this study was to characterize MVOC emission profiles of seven toxigenic strains of S. chartarum, isolated from water-damaged buildings, in order to identify unique MVOCs generated during growth on gypsum wallboard and ceiling tile coupons. Inoculated coupons were incubated and monitored for emissions and growth using a closed glass environmental growth chamber maintained at a constant room temperature. Gas samples were collected from the headspace for three to four weeks using Tenax TA tubes. RESULTS: Most of the MVOCs identified were alcohols, ketones, ethers and esters. The data showed that anisole (methoxybenzene) was emitted from all of the S. chartarum strains tested on both types of substrates. Maximum anisole concentration was detected after seven days of incubation. CONCLUSIONS: MVOCs are suitable markers for fungal identification because they easily diffuse through weak barriers like wallpaper, and could be used for early detection of mold growth in hidden cavities. This study identifies the production of anisole by seven toxigenic strains of Stachybotrys chartarum within a period of one week of growth on gypsum wallboard and ceiling tiles. These data could provide useful information for the future construction of a robust MVOC library for the early detection of this mold.
Project description:After a single or multiple intratracheal instillations of Stachybotrys chartarum (S. chartarum or black mold) spores in BALB/c mice, we characterized cytokine production, metabolites, and inflammatory patterns by analyzing mouse bronchoalveolar lavage (BAL), lung tissue, and plasma. We found marked differences in BAL cell counts, especially large increases in lymphocytes and eosinophils in multiple-dosed mice. Formation of eosinophil-rich granulomas and airway goblet cell metaplasia were prevalent in the lungs of multiple-dosed mice but not in single- or saline-dosed groups. We detected changes in the cytokine expression profiles in both the BAL and plasma. Multiple pulmonary exposures to S. chartarum induced significant metabolic changes in the lungs but not in the plasma. These changes suggest a shift from type 1 inflammation after an acute exposure to type 2 inflammation after multiple exposures to S. chartarum. Eotaxin, vascular endothelial growth factor (VEGF), MIP-1?, MIP-1?, TNF-?, and the IL-8 analogs macrophage inflammatory protein-2 (MIP-2) and keratinocyte chemoattractant (KC), had more dramatic changes in multiple- than in single-dosed mice, and parallel the cytokines that characterize humans with histories of mold exposures versus unexposed control subjects. This repeated exposure model allows us to more realistically characterize responses to mold, such as cytokine, metabolic, and cellular changes.
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.
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.
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.
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.