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Detection of immunogenic protein components in excretion/secretion products of Acanthamoeba T5 using polyclonal
Lissette Retana-Moreira1,2, Elizabeth Abrahams-Sandí1,2, Marco Ruiz-Campos1
1Universidad de Costa Rica, Facultad de Microbiología, Departamento de Parasitología, San José, Costa Rica.
Background:
Acanthamoeba is a free-living amoeba widely distributed, responsible for keratitis and granulomatous amoebic encephalitis. The presence of virulence factors in its excretion/secretion products has been demonstrated. Characterisation of these products, including the determination of immunogenic protein components using polyclonal antibodies, could be the basis for the development of new diagnostic tools and help to understand aspects related to its pathogenesis.
Objectives:
To identify immunogenic protein components in Acanthamoeba conditioned medium (ACM) and extracellular vesicles (EVs) using polyclonal anti-Acanthamoeba antibodies produced in the laboratory and to evaluate the effect of these antibodies in adhesion and cytopathic effect.
Methods:
Excretion/secretion products were obtained after the axenic culture of a potentially pathogenic environmental Acanthamoeba T5 isolate. The presence of immunogenic components in lysates of trophozoites, ACM and EVs was determined using polyclonal anti-Acanthamoeba antibodies produced in Wistar rats. Proteomic analyses to identify the immunogenic protein components in ACM and EVs were included. Experiments to evaluate the effect of polyclonal anti-Acanthamoeba antibodies in adhesion and cytopathic effect in vitro were also performed in Vero cells.
Findings:
Protein recognition by anti-Acanthamoeba antibodies in lysates, ACM and EVs was demonstrated, and these components were identified using proteomics. Decreases in adhesion and cytopathic effect after the preincubation of trophozoites with antibodies, prior to the contact with cells, were observed.
Main Conclusion:
The development of polyclonal antibodies, capable of recognising proteins related to pathogenesis in ACM and EVs, and with significant effects in adhesion, provides an important tool for the search for new therapeutic and diagnostic targets in infections caused by Acanthamoeba.
Insights
Researchers developed polyclonal antibodies against Acanthamoeba proteins found in conditioned medium and extracellular vesicles. These antibodies reduced Acanthamoeba adhesion and cytopathic effects, offering potential diagnostic and therapeutic targets for infections.
Area of Science:
- Microbiology
- Immunology
- Parasitology
Background:
- Acanthamoeba is a ubiquitous protozoan parasite.
- It causes severe infections like keratitis and granulomatous amoebic encephalitis.
- Virulence factors in Acanthamoeba secretions are key to pathogenesis.
Purpose of the Study:
- Identify immunogenic proteins in Acanthamoeba conditioned medium (ACM) and extracellular vesicles (EVs).
- Utilize laboratory-produced polyclonal anti-Acanthamoeba antibodies for identification.
- Assess the impact of these antibodies on Acanthamoeba adhesion and cytopathic effects.
Main Methods:
- Cultured Acanthamoeba and collected excretion/secretion products (ACM, EVs).
- Used polyclonal anti-Acanthamoeba antibodies to detect immunogenic proteins in lysates, ACM, and EVs.
- Performed proteomic analysis to identify specific proteins.
- Evaluated antibody effects on Acanthamoeba adhesion and cytopathic effects in vitro.
Main Results:
- Successfully demonstrated protein recognition by anti-Acanthamoeba antibodies in all tested Acanthamoeba fractions.
- Identified specific immunogenic protein components via proteomics.
- Observed significant reduction in Acanthamoeba adhesion and cytopathic effects when pre-treated with antibodies.
Conclusions:
- Developed effective polyclonal antibodies targeting pathogenic Acanthamoeba proteins.
- These antibodies recognize virulence factors in ACM and EVs.
- Antibodies show therapeutic potential by inhibiting Acanthamoeba adhesion and cytopathicity, aiding in the development of new diagnostic and therapeutic strategies.
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