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It is necessary to know the antigenic determinants as well as the pathogenicity mechanisms of this amoeba to implement strategies that allow for better antiamoebic therapeutic and diagnostic targets that directly impact the health sector. Therefore, the aim of this work was to analyze some virulence factors as part of extracellular vesicle (EV) cargo secreted by <jats:italic>N. fowleri<\/jats:italic>. The EV secretion to the extracellular medium was evaluated in trophozoites fixed and incubated with anti-<jats:italic>N. fowleri<\/jats:italic> antibody while molecular identification of EV cargo was performed by SDS-PAGE, Western blot, and RT-PCR. Our results showed that <jats:italic>N. fowleri<\/jats:italic> secretes a wide variety of vesicle sizes ranging from 0.2 to\u2009&gt;\u20092\u00a0\u03bcm, and these EVs were recognized by antibodies anti-Naegleropore B, anti-19\u00a0kDa polypeptide band, anti-membrane protein Mp2CL5, anti-protease cathepsin B, and anti-actin. Furthermore, these vesicles were localized in the trophozoites cytoplasm or secreted into the extracellular medium. Specifically in relation to small vesicles, our purified exosomes were recognized by CD63 and Hsp70 markers, along with the previously mentioned proteins. RT-PCR analysis was made through the isolation of EVs from <jats:italic>N. fowleri<\/jats:italic> trophozoite culture by concentration, filtration, and ultracentrifugation. Interestingly, we obtained PCR products for Nfa1, NPB, Mp2CL5, and CatB genes as part of exosomes cargo. This suggests that the molecules identified in this work could play an important role in communication as well as in infectious processes caused by this amoeba. Therefore, the study and characterization of the pathogenicity mechanisms, as well as the virulence factors released by <jats:italic>N. fowleri<\/jats:italic> remains a key point to provide valuable information for the development of therapeutic treatments, vaccine design, or biomarkers for a timely diagnosis against infections caused by protozoa.<\/jats:p>","DOI":"10.1007\/s00436-024-08378-9","type":"journal-article","created":{"date-parts":[[2024,10,21]],"date-time":"2024-10-21T04:01:52Z","timestamp":1729483312000},"update-policy":"https:\/\/doi.org\/10.1007\/springer_crossmark_policy","source":"Crossref","is-referenced-by-count":5,"title":["Analysis of virulence factors in extracellular vesicles secreted by Naegleria fowleri"],"prefix":"10.1007","volume":"123","author":[{"given":"Itzel Berenice","family":"Rodr\u00edguez-Mera","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Sa\u00fal","family":"Rojas-Hern\u00e1ndez","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Karla Alejandra","family":"Barr\u00f3n-Graciano","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Mar\u00eda Maricela","family":"Carrasco-Y\u00e9pez","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"297","published-online":{"date-parts":[[2024,10,21]]},"reference":[{"key":"8378_CR1","doi-asserted-by":"publisher","unstructured":"Bautista-L\u00f3pez, NL, Ndao M, Camargo FV, Nara T, Annoura T, Hardie DB, . . . 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