Lissette Retana-Moreira, Elizabeth Abrahams-Sandí, Marco Ruiz-Campos, Johan Alvarado-Ocampo, Julián Castro, Jacob Lorenzo-Morales, Giovanni Sáenz-Arce, Antonio Osuna
{"title":"棘阿米巴T5排泄/分泌产物免疫原性蛋白成分的多克隆抗体检测","authors":"Lissette Retana-Moreira, Elizabeth Abrahams-Sandí, Marco Ruiz-Campos, Johan Alvarado-Ocampo, Julián Castro, Jacob Lorenzo-Morales, Giovanni Sáenz-Arce, Antonio Osuna","doi":"10.1590/0074-02760240190","DOIUrl":null,"url":null,"abstract":"<p><strong>Background: </strong>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.</p><p><strong>Objectives: </strong>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.</p><p><strong>Methods: </strong>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.</p><p><strong>Findings: </strong>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.</p><p><strong>Main conclusion: </strong>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.</p>","PeriodicalId":18469,"journal":{"name":"Memorias do Instituto Oswaldo Cruz","volume":"120 ","pages":"e240190"},"PeriodicalIF":2.5000,"publicationDate":"2025-06-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12158434/pdf/","citationCount":"0","resultStr":"{\"title\":\"Detection of immunogenic protein components in excretion/secretion products of Acanthamoeba T5 using polyclonal antibodies.\",\"authors\":\"Lissette Retana-Moreira, Elizabeth Abrahams-Sandí, Marco Ruiz-Campos, Johan Alvarado-Ocampo, Julián Castro, Jacob Lorenzo-Morales, Giovanni Sáenz-Arce, Antonio Osuna\",\"doi\":\"10.1590/0074-02760240190\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<p><strong>Background: </strong>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.</p><p><strong>Objectives: </strong>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.</p><p><strong>Methods: </strong>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.</p><p><strong>Findings: </strong>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.</p><p><strong>Main conclusion: </strong>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.</p>\",\"PeriodicalId\":18469,\"journal\":{\"name\":\"Memorias do Instituto Oswaldo Cruz\",\"volume\":\"120 \",\"pages\":\"e240190\"},\"PeriodicalIF\":2.5000,\"publicationDate\":\"2025-06-09\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12158434/pdf/\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Memorias do Instituto Oswaldo Cruz\",\"FirstCategoryId\":\"3\",\"ListUrlMain\":\"https://doi.org/10.1590/0074-02760240190\",\"RegionNum\":4,\"RegionCategory\":\"医学\",\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"2025/1/1 0:00:00\",\"PubModel\":\"eCollection\",\"JCR\":\"Q2\",\"JCRName\":\"PARASITOLOGY\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Memorias do Instituto Oswaldo Cruz","FirstCategoryId":"3","ListUrlMain":"https://doi.org/10.1590/0074-02760240190","RegionNum":4,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"2025/1/1 0:00:00","PubModel":"eCollection","JCR":"Q2","JCRName":"PARASITOLOGY","Score":null,"Total":0}
Detection of immunogenic protein components in excretion/secretion products of Acanthamoeba T5 using polyclonal antibodies.
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.
期刊介绍：
Memórias do Instituto Oswaldo Cruz is a journal specialized in microbes & their vectors causing human infections. This means that we accept manuscripts covering multidisciplinary approaches and findings in the basic aspects of infectious diseases, e.g. basic in research in prokariotes, eukaryotes, and/or virus. Articles must clearly show what is the main question to be answered, the hypothesis raised, and the contribution given by the study.
Priority is given to manuscripts reporting novel mechanisms and general findings concerning the biology of human infectious prokariotes, eukariotes or virus. Papers reporting innovative methods for diagnostics or that advance the basic research with these infectious agents are also welcome.
It is important to mention what we do not publish: veterinary infectious agents research, taxonomic analysis and re-description of species, epidemiological studies or surveys or case reports and data re-analysis. Manuscripts that fall in these cases or that are considered of low priority by the journal editorial board, will be returned to the author(s) for submission to another journal.
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