Accurate Differentiation of Trichosporon Asahii and Trichosporon Asteroides from other Human Pathogenic Yeasts and Moulds by using Highly Specific Monoclonal Antibodies

Abstract

Members of the genus Trichosporon are emerging opportunistic pathogens of humans, causing the invasive fungal disease trichosporonosis in immunocompromised patients, and summer-type hypersensitivity pneumonitis (SHP) in immunocompetent individuals through inhalation of arthroconidia. Trichosporonosis is frequently misdiagnosed as candidiasis or cryptococcosis due to a lack of awareness and the inaccuracy of immunodiagnostic tests for these yeast pathogens. Delays in identification and differentiation of Trichosporon spp. from other yeasts and timely administration of appropriate antifungal drug treatments add to the poor prognosis and high mortality rate associated with this trichosporonosis. This thesis describes the use of hybridoma technology to produce two highly specific murine monoclonal antibodies (MAbs), CA7 and TH1, for detection and differentiation of Trichosporon from other yeast pathogens. The MAbs react with extracellular antigens from T. asahii and T. asteroides, the two most common pathogenic agents of trichosporonosis. CA7 and TH1 do not recognise related Trichosporon spp., or unrelated pathogenic yeasts and moulds including Candida spp., Cryptococcus spp., species of Aspergillus, Fusarium, Scedosporium, and etiologic agents of mucormycosis. Immunofluorescence and western blotting studies show that MAb CA7, an immunoglobulin G1 (IgG1), binds to a major ~60kDa glycoprotein antigen produced on the surface of hyphae, while TH1, an immunoglobulin M (IgM), binds to an antigen produced on the surface of conidia. I show how the MAbs can be used with standard mycological growth medium (Sabouraud Dextrose Agar) and an enzyme-linked immunosorbent assay (ELISA) to accurately differentiate T. asahii from Candida albicans and Cryptococcus neoformans in single and mixed species cultures.