{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2025,10,10]],"date-time":"2025-10-10T01:34:40Z","timestamp":1760060080035,"version":"build-2065373602"},"reference-count":24,"publisher":"MDPI AG","issue":"8","license":[{"start":{"date-parts":[[2025,7,31]],"date-time":"2025-07-31T00:00:00Z","timestamp":1753920000000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"CONACHyT-Mexico","award":["926866","59\u20132090\u20131\u2013001-F","003691","2018\u201367015\u201328301"],"award-info":[{"award-number":["926866","59\u20132090\u20131\u2013001-F","003691","2018\u201367015\u201328301"]}]},{"name":"US Agricultural Research Service","award":["926866","59\u20132090\u20131\u2013001-F","003691","2018\u201367015\u201328301"],"award-info":[{"award-number":["926866","59\u20132090\u20131\u2013001-F","003691","2018\u201367015\u201328301"]}]},{"name":"EC-INCO\u2013CT","award":["926866","59\u20132090\u20131\u2013001-F","003691","2018\u201367015\u201328301"],"award-info":[{"award-number":["926866","59\u20132090\u20131\u2013001-F","003691","2018\u201367015\u201328301"]}]},{"name":"USDA-NIFA","award":["926866","59\u20132090\u20131\u2013001-F","003691","2018\u201367015\u201328301"],"award-info":[{"award-number":["926866","59\u20132090\u20131\u2013001-F","003691","2018\u201367015\u201328301"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Vaccines"],"abstract":"<jats:p>Background: Bovine babesiosis, caused by the tick-borne apicomplexan parasite Babesia spp., is an economically significant disease that threatens the cattle industry worldwide. Babesia bovis is the most pathogenic species, leading to high morbidity and mortality in infected animals. One promising approach to vaccination against bovine babesiosis involves the use of multiple protective antigens, offering advantages over traditional live-attenuated vaccines. Tools such as immunobioinformatics and reverse vaccinology have facilitated the identification of novel antigens. Enolase, a \u201cmoonlighting\u201d enzyme of the glycolytic pathway with demonstrated vaccine potential in other pathogens, has not yet been studied in B. bovis. Methods: In this study, the enolase gene from two B. bovis isolates was successfully identified and sequenced. The gene, consisting of 1366 base pairs, encodes a predicted protein of 438 amino acids. Its expression in intraerythrocytic parasites was confirmed by RT-PCR. Two peptides containing predicted B-cell epitopes were synthesized and used to immunize rabbits. Hyperimmune sera were then analyzed by ELISA, confocal microscopy, Western blot, and an in vitro neutralization assay. Results: The hyperimmune sera showed high antibody titers, reaching up to 1:256,000. Specific antibodies recognized intraerythrocytic merozoites by confocal microscopy and bound to a ~47 kDa protein in erythrocytic cultures of B. bovis as detected by Western blot. In the neutralization assay, antibodies raised against peptide 1 had no observable effect, whereas those targeting peptide 2 significantly reduced parasitemia by 71.99%. Conclusions: These results suggest that B. bovis enolase contains B-cell epitopes capable of inducing neutralizing antibodies and may play a role in parasite\u2013host interactions. Enolase is therefore a promising candidate for further exploration as a vaccine antigen. Nonetheless, additional experimental studies are needed to fully elucidate its biological function and validate its vaccine potential.<\/jats:p>","DOI":"10.3390\/vaccines13080818","type":"journal-article","created":{"date-parts":[[2025,8,6]],"date-time":"2025-08-06T15:09:53Z","timestamp":1754492993000},"page":"818","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["Babesia bovis Enolase Is Expressed in Intracellular Merozoites and Contains B-Cell Epitopes That Induce Neutralizing Antibodies In Vitro"],"prefix":"10.3390","volume":"13","author":[{"given":"Alma","family":"C\u00e1rdenas-Flores","sequence":"first","affiliation":[{"name":"Immunology and Vaccines Laboratory, Faculty of Natural Sciences, Autonomous University of Queretaro, Santiago de Queretaro 76230, Queretaro, Mexico"},{"name":"Ph.D. Program in Biological Sciences, College of Natural Sciences, Autonomous University of Queretaro, Santiago de Queretaro 76230, Queretaro, Mexico"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-6354-3200","authenticated-orcid":false,"given":"Minerva","family":"Camacho-Nuez","sequence":"additional","affiliation":[{"name":"Posgrado en Ciencias Genomicas, Universidad Autonoma de la Ciudad de Mexico, Mexico City 03100, Mexico"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-2680-6091","authenticated-orcid":false,"given":"Massaro W.","family":"Ueti","sequence":"additional","affiliation":[{"name":"Animal Diseases Research Unit, USDA-ARS, Pullman, WA 99164, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-5107-194X","authenticated-orcid":false,"given":"Mario","family":"Hidalgo-Ruiz","sequence":"additional","affiliation":[{"name":"College of Veterinary Medicine, Autonomous University of Chiapas, Tuxtla Gutierrez 29000, Chiapas, Mexico"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-5300-6834","authenticated-orcid":false,"given":"Angelina","family":"Rodr\u00edguez-Torres","sequence":"additional","affiliation":[{"name":"Cuerpo Academico de Salud Animal y Microbiolog\u00eda Ambiental, Natural Sciences College, Autonomous University of Queretaro, Santiago de Queretaro 76230, Queretaro, Mexico"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Diego Josimar","family":"Hern\u00e1ndez-Silva","sequence":"additional","affiliation":[{"name":"Immunology and Vaccines Laboratory, Faculty of Natural Sciences, Autonomous University of Queretaro, Santiago de Queretaro 76230, Queretaro, Mexico"},{"name":"Cuerpo Academico de Salud Animal y Microbiolog\u00eda Ambiental, Natural Sciences College, Autonomous University of Queretaro, Santiago de Queretaro 76230, Queretaro, Mexico"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-8837-3578","authenticated-orcid":false,"given":"Jos\u00e9 Guadalupe","family":"G\u00f3mez-Soto","sequence":"additional","affiliation":[{"name":"Cuerpo Acad\u00e9mico de Nutrici\u00f3n y Reproducci\u00f3n Animal, Facultad de Ciencias Naturales, Autonomous University of Queretaro, Santiago de Queretaro 76230, Queretaro, Mexico"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-0042-6856","authenticated-orcid":false,"given":"Masahito","family":"Asada","sequence":"additional","affiliation":[{"name":"National Research Center for Protozoan Diseases, Obihiro University of Agriculture and Veterinary Medicine, Obihiro 080-8555, Hokkaido, Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Shin-ichiro","family":"Kawazu","sequence":"additional","affiliation":[{"name":"National Research Center for Protozoan Diseases, Obihiro University of Agriculture and Veterinary Medicine, Obihiro 080-8555, Hokkaido, Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Alma R.","family":"Tamayo-Sosa","sequence":"additional","affiliation":[{"name":"Instituto de Investigaciones en Ciencias Veterinarias, Universidad Aut\u00f3noma de Baja California, Mexicali 21386, Baja California, Mexico"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Roc\u00edo Alejandra","family":"Ruiz-Manzano","sequence":"additional","affiliation":[{"name":"Departamento de Biolog\u00eda Celular y Fisiolog\u00eda, Instituto de Investigaciones Biom\u00e9dicas, Universidad Nacional Aut\u00f3noma de Mexico, Mexico City 04510, Mexico"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-8892-6390","authenticated-orcid":false,"given":"Juan","family":"Mosqueda","sequence":"additional","affiliation":[{"name":"Immunology and Vaccines Laboratory, Faculty of Natural Sciences, Autonomous University of Queretaro, Santiago de Queretaro 76230, Queretaro, Mexico"},{"name":"Cuerpo Academico de Salud Animal y Microbiolog\u00eda Ambiental, Natural Sciences College, Autonomous University of Queretaro, Santiago de Queretaro 76230, Queretaro, Mexico"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"1968","published-online":{"date-parts":[[2025,7,31]]},"reference":[{"key":"ref_1","unstructured":"Tussaint, M. 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