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Identification and evaluation of a major cytotoxin of A. fumigatus
Abstract:
Aspergillus fumigatus is a highly pathogenic fungus causing a wide spectrum of diseases in immunocompromised as well as immunocompetent hosts. The present work was undertaken to evaluate the cytotoxic nature of fractionated antigens of A. fumigatus against the mammalian cell lines (J774, RAW, CHO and L929). An enriched protein antigenic fraction of A. fumigatus was subjected to con A Sepharose and phenyl Sepharose chromatography. Antigenic fractions, ConAub (conA unbound) and PSC III (fraction III of phenyl Sepharose column) containing low mw antigens showed higher cytotoxicity as compared to other antigenic fractions. PSC III was further purified on HPLC resulting in an 18 kDa homogeneous protein. The purified protein showed high ELISA absorbance values for specific IgG and IgE antibodies in sera of ABPA patients. Monoclonal antibody raised against Asp fl, a major allergen/antigen of A. fumigatus recognised the purified 18 kDa by ELISA and western blot. The 18 kDa allergen/antigen or Asp fl showed similar toxicity towards all the four cell lines (macrophage and fibroblast) with an IC50 of 75 ng/ml or 4.16 nM. Reduction in toxicity of 18 kDa at low temperatures and potentiation in presence of ammonium chloride and monensin indicates mechanism of internalisation of 18 kDa in eukaryotic cells is similar to alpha-sarcin. The present work shows that the 18 kDa allergen/antigen (Asp fl) is a major cytotoxin secreted by A. fumigatus which may play multiple roles in the pathogenesis of Aspergillosis through allergenicity, antigenicity and cytotoxicity.
Insights
A key 18 kDa protein from Aspergillus fumigatus, known as Asp fl, acts as a potent cytotoxin. This fungal antigen contributes to Aspergillosis pathogenesis through its allergenicity, antigenicity, and cytotoxic effects on mammalian cells.
Area of Science:
- Mycology
- Immunology
- Cell Biology
Background:
- Aspergillus fumigatus is a significant pathogen causing diverse diseases in various hosts.
- Understanding its pathogenic mechanisms, particularly the role of its antigens, is crucial for disease management.
Purpose of the Study:
- To investigate the cytotoxic properties of fractionated antigens from Aspergillus fumigatus.
- To identify and characterize a specific cytotoxic antigen responsible for pathogenicity.
Main Methods:
- Fractionation of Aspergillus fumigatus antigens using con A Sepharose and phenyl Sepharose chromatography.
- Purification of a key antigen using High-Performance Liquid Chromatography (HPLC).
- Evaluation of cytotoxicity against mammalian cell lines (J774, RAW, CHO, L929) and characterization using ELISA and Western blot.
Main Results:
- A low molecular weight fraction (PSC III) exhibited significant cytotoxicity.
- An 18 kDa homogeneous protein (Asp fl) was purified, showing high antigenicity in ABPA patient sera.
- The 18 kDa antigen demonstrated potent cytotoxicity (IC50 of 75 ng/ml) against macrophage and fibroblast cell lines, with internalization mechanisms similar to alpha-sarcin.
Conclusions:
- The 18 kDa allergen/antigen (Asp fl) is a major cytotoxin secreted by Aspergillus fumigatus.
- This protein plays a multifaceted role in Aspergillosis pathogenesis via allergenicity, antigenicity, and cytotoxicity.
- Further research into Asp fl could lead to novel therapeutic strategies for Aspergillosis.