Immunoblot analysis of the humoral immune response to Pythium insidiosum in horses with pythiosis.
Abstract: Reactions to Pythium insidiosum by sera from horses with active pythiosis were investigated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and immunoblotting. Five strains of P. insidiosum were grown in nutrient broth and then sonicated. After centrifugation, supernatant antigens were separated by SDS-PAGE. An exoantigen of Conidiobolus coronatus was also tested. Bands with molecular weights between 97,000 and 14,000 were identified by Coomassie blue and silver staining. After being transferred to nitrocellulose, the antigens were reacted against sera from six horses with pythiosis, sera from four horses cured a year earlier by vaccination, and sera from five healthy horses. The sera from horses with pythiosis recognized at least 20 antigens in all strains. Three antigens with molecular weights of 32,000, 30,000, and 28,000 appeared to be immunodominant and specific. Sera from horses cured by immunotherapy showed only five very weak bands, three of them the 32,000-molecular-weight (32K), 30K, and 28K antigens. No bands were observed with sera from healthy horses or sera from horses with a variety of other infections. Sera from horses with pythiosis cross-reacted with the 44K antigen of C. coronatus. The immunodominant antigens described here may be useful for diagnostic purposes and in immunotherapy for this oomycotic infection in horses.
Publication Date: 1992-11-01 PubMed ID: 1452669PubMed Central: PMC270563DOI: 10.1128/jcm.30.11.2980-2983.1992Google Scholar: Lookup
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- Journal Article
- Research Support
- Non-U.S. Gov't
Summary
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The research article explores how the immune responses in horses affected by pythiosis interact with the pathogen Pythium insidiosum. Through analysis, it was found that certain antigens may be key in diagnosing and managing the condition.
Methodology
- The research team conducted an immunoblot analysis, specifically using sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and immunoblotting.
- Five strains of P. insidiosum were cultivated, followed by sonication, a process that utilizes sound waves to disrupt cell structures.
- The supernatant antigens, or the proteins present in the liquid above a sediment or precipitate, were separated using SDS-PAGE.
- The team also tested an exoantigen (an antigen excreted from a pathogenic microorganism) from Conidiobolus coronatus.
- The detection of different protein bands in the antigens occurred through staining methods using Coomassie blue and silver.
Interactions with Sera
- The antigens that underwent separation subsequently reacted against the sera from different horse groups: those suffering from active pythiosis, those cured through vaccination one year prior, and healthy horses.
- The sera from horses with pythiosis identified at least 20 antigens across all strains.
- There were three main antigens with molecular weights of 32,000, 30,000, and 28,000, which appeared to be immunodominant and specific.
- Sera from horses cured through immunotherapy identified far fewer bands and the same three core antigens, although these were faint.
Implications and Findings
- No antigenic bands were detected with the sera from the healthy horses or those with other infections, suggesting the specificity of the identified antigens to pythiosis.
- The sera from horses with pythiosis also cross-reacted with the 44K antigen of C. coronatus, indicating a potential for cross-reactivity in the immune response.
- The conclusion highlights that the discovery of these immunodominant antigens may be beneficial for the diagnosis and immunotherapy of pythiosis, thus serving as key markers of this disease.
Cite This Article
APA
Mendoza L, Nicholson V, Prescott JF.
(1992).
Immunoblot analysis of the humoral immune response to Pythium insidiosum in horses with pythiosis.
J Clin Microbiol, 30(11), 2980-2983.
https://doi.org/10.1128/jcm.30.11.2980-2983.1992 Publication
Researcher Affiliations
- Department of Veterinary Microbiology and Immunology, University of Guelph, Ontario, Canada.
MeSH Terms
- Animals
- Antibodies, Fungal / blood
- Antigens, Fungal / isolation & purification
- Electrophoresis, Polyacrylamide Gel
- Horse Diseases / immunology
- Horses
- Immunoblotting
- Molecular Weight
- Mycoses / immunology
- Mycoses / veterinary
- Pythium / immunology
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This article includes 21 references
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Citations
This article has been cited 11 times.- Yolanda H, Krajaejun T. History and Perspective of Immunotherapy for Pythiosis.. Vaccines (Basel) 2021 Sep 26;9(10).
- Vilela R, Mendoza L. Human Pathogenic Entomophthorales.. Clin Microbiol Rev 2018 Oct;31(4).
- Mendoza L, Vilela R, Voelz K, Ibrahim AS, Voigt K, Lee SC. Human Fungal Pathogens of Mucorales and Entomophthorales.. Cold Spring Harb Perspect Med 2014 Nov 6;5(4).
- Botton SA, Pereira DI, Costa MM, Azevedo MI, Argenta JS, Jesus FP, Alves SH, Santurio JM. Identification of Pythium insidiosum by nested PCR in cutaneous lesions of Brazilian horses and rabbits.. Curr Microbiol 2011 Apr;62(4):1225-9.
- Jindayok T, Piromsontikorn S, Srimuang S, Khupulsup K, Krajaejun T. Hemagglutination test for rapid serodiagnosis of human pythiosis.. Clin Vaccine Immunol 2009 Jul;16(7):1047-51.
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