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Identification and purification of specific Penicillium marneffei antigens and their recognition by human immune sera
L Jeavons1, A J Hamilton, N Vanittanakom
1Dermatology Laboratory, St. John's Institute of Dermatology, Guy's Hospital, London, United Kingdom. l.jeavons@umds.ac.uk
Abstract:
Disseminated infection with the dimorphic pathogenic fungus Penicillium marneffei is increasingly seen among patients with AIDS in southeast Asian countries. Previous studies have demonstrated the presence of humoral immune responses to this fungus in patient sera; we have confirmed this work using sera from P. marneffei-infected patients (n = 21) to develop Western blots of P. marneffei cytoplasmic yeast antigen (CYA). P. marneffei CYA was then partially purified by liquid isoelectric focusing, and fractions were subjected to sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and Western blotting. Immunoenzyme development of the Western blots with pooled sera from patients with P. marneffei infection and with pooled sera from patients with aspergillosis (n = 20), candidiasis (n = 10), cryptococcosis (n = 9), and histoplasmosis (n = 11) revealed three antigens with relative molecular masses of 61, 54, and 50 kDa. These antigens were specifically recognized by the pooled sera from the P. marneffei-infected patients. The 61- and 54-kDa antigens were subsequently purified to homogeneity by preparative gel electrophoresis, and the 50-kDa antigen was partially purified by the same technique. N-terminal amino acid sequencing revealed that the 61-kDa antigen had a strong homology (87% identity) with the antioxidant enzyme catalase. The three antigens were then subjected to SDS-PAGE and Western blotting and to immunoenzyme development with individual patient sera; sera from 86% of P. marneffei-infected patients recognized the 61-kDa antigen, sera from 71% recognized the 54-kDa antigen, and sera from 48% recognized the 50-kDa antigen. These specifically recognized antigens are the first to be purified from P. marneffei and can be used either singly or in combination to detect antibody responses in a large percentage of individuals infected with P. marneffei.
Insights
Researchers identified three key antigens from the Penicillium marneffei fungus, crucial for detecting immune responses in patients with AIDS. These purified antigens offer a promising diagnostic tool for P. marneffei infections.
Area of Science:
- Mycology
- Immunology
- Infectious Diseases
Background:
- Disseminated Penicillium marneffei infection is a growing concern for AIDS patients in Southeast Asia.
- Previous research indicated humoral immune responses to P. marneffei in infected individuals.
Purpose of the Study:
- To identify and purify specific antigens of P. marneffei recognized by patient antibodies.
- To develop a diagnostic tool for detecting P. marneffei infections through antibody response.
Main Methods:
- Western blotting of P. marneffei cytoplasmic yeast antigen (CYA) using patient sera.
- Antigen purification via liquid isoelectric focusing and preparative gel electrophoresis.
- N-terminal amino acid sequencing and SDS-PAGE for antigen characterization.
Main Results:
- Three specific antigens (61, 54, and 50 kDa) were identified and recognized by sera from P. marneffei-infected patients.
- The 61-kDa antigen showed homology to catalase; 61, 54, and 50 kDa antigens were purified.
- Antibody detection rates were 86% for the 61-kDa, 71% for the 54-kDa, and 48% for the 50-kDa antigen.
Conclusions:
- The purified 61, 54, and 50 kDa antigens are the first to be isolated from P. marneffei.
- These antigens can be used individually or combined for sensitive detection of antibody responses in P. marneffei infections.