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The current study aimed to investigate molecular identification, cyst histological examination and seroprevalence of <jats:italic>Sarcocystis<\/jats:italic> spp. in water buffaloes slaughtered in the main abattoirs of 6 Egyptian Governorates; Cairo, Giza, Beni-suef, Al-Sharqia, Qalyubia, and El-Beheira. Each buffalo was visually inspected for the presence of <jats:italic>Sarcocystis<\/jats:italic> macrocysts and blood samples were collected. Out of 900 examined water buffaloes (<jats:italic>Bubalus bubalis<\/jats:italic>), 246 (27.3%) were found to be infected based on macroscopic examination. Histological examination of the macrocyst revealed metrocytes close to the cyst's edge and elongated curving basophilic bradyzoites occupying most of the cyst. Depending on age, we categorized naturally infected buffaloes into 3 groups: young (&lt;\u20092 y.), adult (2\u20135 y.) and old (5\u201310 y.). The highest infection rate was observed in older buffaloes aged 5\u201310 years, meanwhile female animals exhibited a higher prevalence of infection compared to males. The esophagus had the highest presence of <jats:italic>Sarcocystis<\/jats:italic> compared to other organs. Using PCR based on <jats:italic>18S<\/jats:italic> rRNA gene and sequencing, we isolated 3 <jats:italic>Sarcocystis<\/jats:italic> species from infected tissues: <jats:italic>S. buffalonis<\/jats:italic>, <jats:italic>S. fusiformis<\/jats:italic> and <jats:italic>S. hirsuta-<\/jats:italic>like. The prepared whole cyst antigen of <jats:italic>S<\/jats:italic>. <jats:italic>fusiformis<\/jats:italic> was characterized by 9 major polypeptides (140, 120, 78, 66, 53, 39, 32, 24 and 19\u00a0kDa) on 10% SDS-PAGE. Using western blotting, we identified 2 common immunogenic reactive bands (66 and 32\u00a0kDa) against naturally infected buffalo sera and hyperimmune rat sera. Additionally, sarcocystosis positive seroprevalence rate was 46.4% (418\/900) with a sensitivity of 97.6% and specificity of 100% using indirect ELISA based on <jats:italic>S. fusiformis<\/jats:italic> whole cyst antigen. In conclusion, this study highlights the molecular identification, cyst histological examination and seroprevalence of <jats:italic>Sarcocystis<\/jats:italic> sp. among buffaloes in Egypt. ELISA using the <jats:italic>S. fusiformis<\/jats:italic> whole cyst antigen could be adapted to detect antibodies to <jats:italic>Sarcocystis<\/jats:italic> sp., in live animals, with an acceptable specificity and sensitivity.<\/jats:p>","DOI":"10.1186\/s12917-025-04933-3","type":"journal-article","created":{"date-parts":[[2025,7,22]],"date-time":"2025-07-22T04:33:30Z","timestamp":1753158810000},"update-policy":"https:\/\/doi.org\/10.1007\/springer_crossmark_policy","source":"Crossref","is-referenced-by-count":2,"title":["Sarcocystis species: molecular identification and seroprevalence in water buffaloes (Bubalus bubalis)"],"prefix":"10.1186","volume":"21","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-9718-5130","authenticated-orcid":false,"given":"Nagwa I.","family":"Toaleb","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-2620-4189","authenticated-orcid":false,"given":"Raafat M.","family":"Shaapan","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-9673-015X","authenticated-orcid":false,"given":"Haitham","family":"Elaadli","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-0723-7276","authenticated-orcid":false,"given":"Kadria N.","family":"Abdel Megeed","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-2581-6503","authenticated-orcid":false,"given":"Dina","family":"Aboelsoued","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"297","published-online":{"date-parts":[[2025,7,22]]},"reference":[{"issue":"1","key":"4933_CR1","doi-asserted-by":"publisher","first-page":"12","DOI":"10.1186\/s13568-024-01817-x","volume":"15","author":"D Aboelsoued","year":"2025","unstructured":"Aboelsoued D, Toaleb NI, Abdel Megeed KN. 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