Production of a Chaetomium globosum enolase monoclonal antibody.

Chaetomium globosum is a hydrophilic fungal species and a contaminant of water-damaged building materials in North America. Methods to detect Chaetomium species include subjective identification of ascospores, viable culture, or molecular-based detection methods. In this study, we describe the production and initial characterization of a monoclonal antibody (MAb) for C. globosum enolase. MAb 1C7, a murine IgG1 isotype MAb, was produced and reacted with recombinant C. globosum enolase (rCgEno) in an enzyme-linked immunosorbent assay and with a putative C. globosum enolase in a Western blot. Epitope mapping showed MAb 1C7 specific reactivity to an enolase decapeptide, LTYEELANLY, that is highly conserved within the fungal class Sordariomycetes. Cross-reactivity studies showed MAb 1C7 reactivity to C. atrobrunneum but not C. indicum. MAb 1C7 did not react with enolase from Aspergillus fumigatus, which is divergent in only two amino acids within this epitope. The results of this study suggest potential utility of MAb 1C7 in Western blot applications for the detection of Chaetomium and other Sordariomycetes species.

Quantification of C. globosum spores in house dust samples.

Chaetomium globosum is one of the most common fungi that grows in damp buildings and occurs in agricultural and forestry workplaces. Using sera from atopic patients, we characterized and purified an extracellular chitosanase (Chg47) from C. globosum that is antigenic to humans. The study reports the production of monoclonal antibodies to the protein. Three capture ELISAs were developed for Chg47 for detection of spores and spore and mycelial fragments in dust samples using different mono- and polyclonal antibody combinations. One method is based on an enhanced biotinylated polyclonal antibody as the secondary antibody and coating anti-IgM to capture one of two clones of IgM monoclonal antibodies as the capture antibody. The other method makes use of an enhanced rabbit polyclonal antibody as both the primary and capture antibody. The detection limit of the double PAb method for the Chg47 antigen was 7.6 pg/ml. When the anti-IgM+10B3 clone was used, the detection limit was 61 pg/ml and for anti-IgM+5F12, 122 pg/ml. The detection limit of double PAb method is comparable to methods for the allergen and spores of Aspergillus versicolor in house dust and is more sensitive than other immunoassays for allergens in house including for Stachybotrys chartarum, Aspergillus fumigatus and Alternaria alternata. All three methods had limited cross-reactivity to fungi common in house dust representing a diverse array of taxa.

Characterization of an antigenic chitosanase from the cellulolytic fungus Chaetomium globosum.

We are interested in identifying human fungal allergens and antigens from species common on water-damaged or damp building materials for use as marker proteins and diagnostic tests. The cellulolytic fungus Chaetomium globosum is common on damp materials in the building environment worldwide. ELISA and immunoblotting tests identified two related proteins of molecular weights 45 and 47 kDa which were identified as fungal antigens found on spore surfaces and in culture filtrate. The sequences were determined by liquid chromatography tandem mass spectrometry (LC-MS/MS), which indicated that the two proteins were chitosanases, confirmed by enzyme assay. The 47 kDa protein was not glycosylated and had an acidic pI of 4.5. These proteins have not been reported from other fungi and similar antigens were not seen in other fungi common in buildings. The production of polyclonal antibodies in rabbits showed the antigenicity of the target proteins and confirmed they were not artifacts of the isolation process. The proteins isolated are useful biomarkers for the detection of C. globosum in the building environment.

Chaetoglobosin F, a small molecule compound, possesses immunomodulatory properties on bone marrow-derived dendritic cells via TLR9 signaling pathway.

Chaetoglobosin F (Cha F), a cytochalasan-based alkaloid, was obtained from the EtOAc extract of a solid culture of Chaetomium globosum IFB-E019. Dendritic cells (DCs), the most potent antigen presenting cells, are considered as the major target in the modulation of excessive immune responses. Recognition of CpG-DNA by Toll-like receptor 9 (TLR9) on DCs is an important step in the pathogenesis of autoimmune diseases. However, the effect of Cha F on the maturation and immunostimulatory function of CpG-stimulated DCs remains unclear. This study investigated the effects of Cha F on bone marrow (BM)-derived DCs. We found that Cha F inhibits the CpG-induced DCs maturation and function by suppressing the expression of surface molecules (CD40, CD80, CD86 and MHC-II), reducing the production of cytokines and chemokines (IL-12 and CXCL-10), inhibiting the CpG-induced DCs-elicited allogeneic T-cell proliferation, and impairing the migration ability to chemokines. The Cha F-treated DCs were highly efficient at Ag capture, via mannose receptor-mediated endocytosis. Additionally, Cha F was also demonstrated to inhibit CpG-induced activation of MAPKs (p38 and JNK, but not ERK) and the nuclear translocation of NF-κB and STAT1. Furthermore, we confirmed that Cha F was able to suppress TLR9 expression of CpG-induced DCs. Collectively, these findings provide novel insight into the immunopharmacological functions of Cha F, especially with regard to their impact on CpG-induced DCs. These immunosuppressive properties of Cha F may prove useful in controlling DCs-associated autoimmune and/or inflammatory diseases.

Infection of barley roots by Chaetomium globosum: evidence for a protective role of the exodermis.

Plant pathogenesis by fungi is known to be dependent on the host genotype, the virulence of the pathogen and certain environmental conditions influencing fungal establishment. Previously, it has been shown that Chaetomium globosum, a fungus well-characterized for its biocontrol potential, causes necrosis on barley roots grown in Murashige and Skoog (MS)-agar. Using MS-agar and aeroponic culture as axenic plant growth systems, C. globosum pathogenesis was analyzed with serological and histological methods. Irrespective of the growth system, C. globosum infected the root epidermis. Roots grown in MS-agar were extensively colonized intercellularly and intracellularly up to the inner cortex and the tissue underwent necrosis. In contrast, roots grown in aeroponic culture were not colonized beyond the epidermis and the roots appeared healthy. Histochemical analyses revealed that hypodermal suberization stopped fungal invasion. In root tips known to lack suberization, epidermal papilla formation reduced overall infection frequency. The results indicate that specific environmental conditions are important for infection and disease expression in barley roots. Infection is restricted by two spatial and temporal distinct defence mechanisms: (1) papillae in root tips retarding fungal invasion; and (2) suberization of hypodermal cells blocking fungal radial growth.

A study of the aeromycoflora of Cádiz: relationship to anthropogenic activity.

We describe a quantitative and qualitative study of the fungal spores found in the air of Cádiz during 1989 using a Cour-type trap. The results of this study can be extrapolated to other coastal cities of southern Europe with a Mediterranean climate. The spores identified have been classified into 25 taxonomic categories. The most abundant were Cladosporium, Chaetomium and Ustilago, and the most frequent, in addition to those mentioned, were Alternaria, Ascophyta and Venturia. The great abundance of Cladosporium is in accordance with the coastal situation of the city. Cladosporium, Alternaria, Curvularia and Stemphylium reached maximum concentrations jointly in October, 1989. They showed mutual cross-reactions. Ustilago and Nigrospora appeared during the period of cereal harvesting and storage.