Immunopeptidomics of Salmonella enterica Serovar Typhimurium-Infected Pig Macrophages Genotyped for Class II Molecules

Salmonellosis is a zoonotic infection that has a major impact on human health; consuming contaminated pork products is the main source of such infection. Vaccination responses to classic vaccines have been unsatisfactory; that is why peptide subunit-based vaccines represent an excellent alternative....

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Autores:
Agudelo, William
Ibarrola, Nieves
Degano, Rosa
Díaz, Jaime
Manzano Román, Raúl
Patarroyo, Manuel A.
Celis Giraldo, Carmen Teresa
Patarroyo, Manuel-Alfonso
Tipo de recurso:
Article of investigation
Fecha de publicación:
2024
Institución:
Universidad de Ciencias Aplicadas y Ambientales U.D.C.A
Repositorio:
Repositorio Institucional UDCA
Idioma:
eng
OAI Identifier:
oai:repository.udca.edu.co:11158/6675
Acceso en línea:
https://repository.udca.edu.co/handle/11158/6675
https://doi.org/10.3390/biology13100832
https://repository.udca.edu.co/
Palabra clave:
630 - Agricultura y tecnologías relacionadas::636 - Producción animal
Cerdos
Péptidos
Salmonella typhimurium
Vacunas
Salmonella typhimurium
Immunopeptidomics
Peptide
Vaccine
Pig
Rights
openAccess
License
https://creativecommons.org/licenses/by-nc-sa/4.0/legalcode.es
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repository_id_str
dc.title.eng.fl_str_mv Immunopeptidomics of Salmonella enterica Serovar Typhimurium-Infected Pig Macrophages Genotyped for Class II Molecules
title Immunopeptidomics of Salmonella enterica Serovar Typhimurium-Infected Pig Macrophages Genotyped for Class II Molecules
spellingShingle Immunopeptidomics of Salmonella enterica Serovar Typhimurium-Infected Pig Macrophages Genotyped for Class II Molecules
630 - Agricultura y tecnologías relacionadas::636 - Producción animal
Cerdos
Péptidos
Salmonella typhimurium
Vacunas
Salmonella typhimurium
Immunopeptidomics
Peptide
Vaccine
Pig
title_short Immunopeptidomics of Salmonella enterica Serovar Typhimurium-Infected Pig Macrophages Genotyped for Class II Molecules
title_full Immunopeptidomics of Salmonella enterica Serovar Typhimurium-Infected Pig Macrophages Genotyped for Class II Molecules
title_fullStr Immunopeptidomics of Salmonella enterica Serovar Typhimurium-Infected Pig Macrophages Genotyped for Class II Molecules
title_full_unstemmed Immunopeptidomics of Salmonella enterica Serovar Typhimurium-Infected Pig Macrophages Genotyped for Class II Molecules
title_sort Immunopeptidomics of Salmonella enterica Serovar Typhimurium-Infected Pig Macrophages Genotyped for Class II Molecules
dc.creator.fl_str_mv Agudelo, William
Ibarrola, Nieves
Degano, Rosa
Díaz, Jaime
Manzano Román, Raúl
Patarroyo, Manuel A.
Celis Giraldo, Carmen Teresa
Patarroyo, Manuel-Alfonso
dc.contributor.author.none.fl_str_mv Agudelo, William
Ibarrola, Nieves
Degano, Rosa
Díaz, Jaime
Manzano Román, Raúl
Patarroyo, Manuel A.
Celis Giraldo, Carmen Teresa
Patarroyo, Manuel-Alfonso
dc.subject.ddc.none.fl_str_mv 630 - Agricultura y tecnologías relacionadas::636 - Producción animal
topic 630 - Agricultura y tecnologías relacionadas::636 - Producción animal
Cerdos
Péptidos
Salmonella typhimurium
Vacunas
Salmonella typhimurium
Immunopeptidomics
Peptide
Vaccine
Pig
dc.subject.lemb.none.fl_str_mv Cerdos
dc.subject.armarc.none.fl_str_mv Péptidos
dc.subject.decs.none.fl_str_mv Salmonella typhimurium
Vacunas
dc.subject.proposal.eng.fl_str_mv Salmonella typhimurium
Immunopeptidomics
Peptide
Vaccine
Pig
description Salmonellosis is a zoonotic infection that has a major impact on human health; consuming contaminated pork products is the main source of such infection. Vaccination responses to classic vaccines have been unsatisfactory; that is why peptide subunit-based vaccines represent an excellent alternative. Immunopeptidomics was used in this study as a novel approach for identifying antigens coupled to major histocompatibility complex class II molecules. Three homozygous individuals having three different haplotypes (Lr-0.23, Lr-0.12, and Lr-0.21) were thus selected as donors; peripheral blood macrophages were then obtained and stimulated with Salmonella typhimurium (MOI 1:40). Although similarities were observed regarding peptide length distribution, elution patterns varied between individuals; in total, 1990 unique peptides were identified as follows: 372 for Pig 1 (Lr-0.23), 438 for Pig 2 (Lr.0.12) and 1180 for Pig 3 (Lr.0.21). Thirty-one S. typhimurium unique peptides were identified; most of the identified peptides belonged to outer membrane protein A and chaperonin GroEL. Notably, 87% of the identified bacterial peptides were predicted in silico to be elution ligands. These results encourage further in vivo studies to assess the immunogenicity of the identified peptides, as well as their usefulness as possible protective vaccine candidates.
publishDate 2024
dc.date.issued.none.fl_str_mv 2024
dc.date.accessioned.none.fl_str_mv 2025-09-30T16:51:30Z
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dc.type.none.fl_str_mv Artículo de revista
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dc.identifier.citation.none.fl_str_mv Celis-Giraldo, C., Suárez, C. F., Agudelo, W., Ibarrola, N., Degano, R., Díaz, J., Manzano-Román, R., & Patarroyo, M. A. (2024). Immunopeptidomics of Salmonella enterica Serovar Typhimurium-Infected Pig Macrophages Genotyped for Class II Molecules. Biology, 13(10), 832. https://doi.org/10.3390/biology13100832
dc.identifier.uri.none.fl_str_mv https://repository.udca.edu.co/handle/11158/6675
dc.identifier.doi.none.fl_str_mv https://doi.org/10.3390/biology13100832
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dc.identifier.instname.none.fl_str_mv Universidad de Ciencias Aplicadas y Ambientales
dc.identifier.reponame.none.fl_str_mv UDCA
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identifier_str_mv Celis-Giraldo, C., Suárez, C. F., Agudelo, W., Ibarrola, N., Degano, R., Díaz, J., Manzano-Román, R., & Patarroyo, M. A. (2024). Immunopeptidomics of Salmonella enterica Serovar Typhimurium-Infected Pig Macrophages Genotyped for Class II Molecules. Biology, 13(10), 832. https://doi.org/10.3390/biology13100832
20797737
Universidad de Ciencias Aplicadas y Ambientales
UDCA
url https://repository.udca.edu.co/handle/11158/6675
https://doi.org/10.3390/biology13100832
https://repository.udca.edu.co/
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language eng
dc.relation.indexed.none.fl_str_mv LaReferencia
dc.relation.citationedition.none.fl_str_mv (Oct., 2024) Artículo número 832
dc.relation.citationendpage.none.fl_str_mv 22
dc.relation.citationissue.none.fl_str_mv 10
dc.relation.citationstartpage.none.fl_str_mv 1
dc.relation.citationvolume.none.fl_str_mv 13
dc.relation.ispartofjournal.none.fl_str_mv Biology
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spelling Agudelo, WilliamIbarrola, NievesDegano, RosaDíaz, JaimeManzano Román, RaúlPatarroyo, Manuel A.Celis Giraldo, Carmen Teresavirtual::912-1Patarroyo, Manuel-Alfonsovirtual::921-12025-09-30T16:51:30Z2025-09-30T16:51:30Z2024Celis-Giraldo, C., Suárez, C. F., Agudelo, W., Ibarrola, N., Degano, R., Díaz, J., Manzano-Román, R., & Patarroyo, M. A. (2024). Immunopeptidomics of Salmonella enterica Serovar Typhimurium-Infected Pig Macrophages Genotyped for Class II Molecules. Biology, 13(10), 832. https://doi.org/10.3390/biology13100832https://repository.udca.edu.co/handle/11158/6675https://doi.org/10.3390/biology1310083220797737Universidad de Ciencias Aplicadas y AmbientalesUDCAhttps://repository.udca.edu.co/Salmonellosis is a zoonotic infection that has a major impact on human health; consuming contaminated pork products is the main source of such infection. Vaccination responses to classic vaccines have been unsatisfactory; that is why peptide subunit-based vaccines represent an excellent alternative. Immunopeptidomics was used in this study as a novel approach for identifying antigens coupled to major histocompatibility complex class II molecules. Three homozygous individuals having three different haplotypes (Lr-0.23, Lr-0.12, and Lr-0.21) were thus selected as donors; peripheral blood macrophages were then obtained and stimulated with Salmonella typhimurium (MOI 1:40). Although similarities were observed regarding peptide length distribution, elution patterns varied between individuals; in total, 1990 unique peptides were identified as follows: 372 for Pig 1 (Lr-0.23), 438 for Pig 2 (Lr.0.12) and 1180 for Pig 3 (Lr.0.21). Thirty-one S. typhimurium unique peptides were identified; most of the identified peptides belonged to outer membrane protein A and chaperonin GroEL. Notably, 87% of the identified bacterial peptides were predicted in silico to be elution ligands. These results encourage further in vivo studies to assess the immunogenicity of the identified peptides, as well as their usefulness as possible protective vaccine candidates.Incluye referencias bibliográficasapplication/pdfenghttps://creativecommons.org/licenses/by-nc-sa/4.0/legalcode.eshttps://creativecommons.org/licenses/by-nc-sa/4.0/info:eu-repo/semantics/openAccessAtribución-NoComercial-CompartirIgual 4.0 Internacional (CC BY-NC-SA 4.0)http://purl.org/coar/access_right/c_abf2https://www.mdpi.com/2079-7737/13/10/832630 - Agricultura y tecnologías relacionadas::636 - Producción animalCerdosPéptidosSalmonella typhimuriumVacunasSalmonella typhimuriumImmunopeptidomicsPeptideVaccinePigImmunopeptidomics of Salmonella enterica Serovar Typhimurium-Infected Pig Macrophages Genotyped for Class II MoleculesArtículo de 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jecuta públicamente en forma digital la Obra o cualquier Obra Derivada u Obra Colectiva, Usted debe mantener intacta toda la información de derecho de autor de la Obra y proporcionar, de forma razonable según el medio o manera que Usted esté utilizando: (i) el nombre del Autor Original si está provisto (o seudónimo, si fuere aplicable), y/o (ii) el nombre de la parte o las partes que el Autor Original y/o el Licenciante hubieren designado para la atribución (v.g., un instituto patrocinador, editorial, publicación) en la información de los derechos de autor del Licenciante, términos de servicios o de otras formas razonables; el título de la Obra si está provisto; en la medida de lo razonablemente factible y, si está provisto, el Identificador Uniforme de Recursos (Uniform Resource Identifier) que el Licenciante especifica para ser asociado con la Obra, salvo que tal URI no se refiera a la nota sobre los derechos de autor o a la información sobre el licenciamiento de la Obra; y en el caso de una Obra Derivada, atribuir el crédito identificando el uso de la Obra en la Obra Derivada (v.g., "Traducción Francesa de la Obra del Autor Original," o "Guión Cinematográfico basado en la Obra original del Autor Original"). Tal crédito puede ser implementado de cualquier forma razonable; en el caso, sin embargo, de Obras Derivadas u Obras Colectivas, tal crédito aparecerá, como mínimo, donde aparece el crédito de cualquier otro autor comparable y de una manera, al menos, tan destacada como el crédito de otro autor comparable.</li>
      <li>
        Para evitar toda confusión, el Licenciante aclara que, cuando la obra es una composición musical:
        <ol type="i">
          <li>Regalías por interpretación y ejecución bajo licencias generales. El Licenciante se reserva el derecho exclusivo de autorizar la ejecución pública o la ejecución pública digital de la obra y de recolectar, sea individualmente o a través de una sociedad de gestión colectiva de derechos de autor y derechos conexos (por ejemplo, SAYCO), las regalías por la ejecución pública o por la ejecución pública digital de la obra (por ejemplo Webcast) licenciada bajo licencias generales, si la interpretación o ejecución de la obra está primordialmente orientada por o dirigida a la obtención de una ventaja comercial o una compensación monetaria privada.</li>
          <li>Regalías por Fonogramas. El Licenciante se reserva el derecho exclusivo de recolectar, individualmente o a través de una sociedad de gestión colectiva de derechos de autor y derechos conexos (por ejemplo, los consagrados por la SAYCO), una agencia de derechos musicales o algún agente designado, las regalías por cualquier fonograma que Usted cree a partir de la obra (“versión cover”) y distribuya, en los términos del régimen de derechos de autor, si la creación o distribución de esa versión cover está primordialmente destinada o dirigida a obtener una ventaja comercial o una compensación monetaria privada.</li>
        </ol>
      </li>
      <li>Gestión de Derechos de Autor sobre Interpretaciones y Ejecuciones Digitales (WebCasting). Para evitar toda confusión, el Licenciante aclara que, cuando la obra sea un fonograma, el Licenciante se reserva el derecho exclusivo de autorizar la ejecución pública digital de la obra (por ejemplo, webcast) y de recolectar, individualmente o a través de una sociedad de gestión colectiva de derechos de autor y derechos conexos (por ejemplo, ACINPRO), las regalías por la ejecución pública digital de la obra (por ejemplo, webcast), sujeta a las disposiciones aplicables del régimen de Derecho de Autor, si esta ejecución pública digital está primordialmente dirigida a obtener una ventaja comercial o una compensación monetaria privada.</li>
    </ol>
  </li>
  <br/>
  <li>
    Representaciones, Garantías y Limitaciones de Responsabilidad.
    <p>A MENOS QUE LAS PARTES LO ACORDARAN DE OTRA FORMA POR ESCRITO, EL LICENCIANTE OFRECE LA OBRA (EN EL ESTADO EN EL QUE SE ENCUENTRA) “TAL CUAL”, SIN BRINDAR GARANTÍAS DE CLASE ALGUNA RESPECTO DE LA OBRA, YA SEA EXPRESA, IMPLÍCITA, LEGAL O CUALQUIERA OTRA, INCLUYENDO, SIN LIMITARSE A ELLAS, GARANTÍAS DE TITULARIDAD, COMERCIABILIDAD, ADAPTABILIDAD O ADECUACIÓN A PROPÓSITO DETERMINADO, AUSENCIA DE INFRACCIÓN, DE AUSENCIA DE DEFECTOS LATENTES O DE OTRO TIPO, O LA PRESENCIA O AUSENCIA DE ERRORES, SEAN O NO DESCUBRIBLES (PUEDAN O NO SER ESTOS DESCUBIERTOS). ALGUNAS JURISDICCIONES NO PERMITEN LA EXCLUSIÓN DE GARANTÍAS IMPLÍCITAS, EN CUYO CASO ESTA EXCLUSIÓN PUEDE NO APLICARSE A USTED.</p>
  </li>
  <br/>
  <li>
    Limitación de responsabilidad.
    <p>A MENOS QUE LO EXIJA EXPRESAMENTE LA LEY APLICABLE, EL LICENCIANTE NO SERÁ RESPONSABLE ANTE USTED POR DAÑO ALGUNO, SEA POR RESPONSABILIDAD EXTRACONTRACTUAL, PRECONTRACTUAL O CONTRACTUAL, OBJETIVA O SUBJETIVA, SE TRATE DE DAÑOS MORALES O PATRIMONIALES, DIRECTOS O INDIRECTOS, PREVISTOS O IMPREVISTOS PRODUCIDOS POR EL USO DE ESTA LICENCIA O DE LA OBRA, AUN CUANDO EL LICENCIANTE HAYA SIDO ADVERTIDO DE LA POSIBILIDAD DE DICHOS DAÑOS. ALGUNAS LEYES NO PERMITEN LA EXCLUSIÓN DE CIERTA RESPONSABILIDAD, EN CUYO CASO ESTA EXCLUSIÓN PUEDE NO APLICARSE A USTED.</p>
  </li>
  <br/>
  <li>
    Término.
    <ol type="a">
      <li>Esta Licencia y los derechos otorgados en virtud de ella terminarán automáticamente si Usted infringe alguna condición establecida en ella. Sin embargo, los individuos o entidades que han recibido Obras Derivadas o Colectivas de Usted de conformidad con esta Licencia, no verán terminadas sus licencias, siempre que estos individuos o entidades sigan cumpliendo íntegramente las condiciones de estas licencias. Las Secciones 1, 2, 5, 6, 7, y 8 subsistirán a cualquier terminación de esta Licencia.</li>
      <li>Sujeta a las condiciones y términos anteriores, la licencia otorgada aquí es perpetua (durante el período de vigencia de los derechos de autor de la obra). No obstante lo anterior, el Licenciante se reserva el derecho a publicar y/o estrenar la Obra bajo condiciones de licencia diferentes o a dejar de distribuirla en los términos de esta Licencia en cualquier momento; en el entendido, sin embargo, que esa elección no servirá para revocar esta licencia o que deba ser otorgada , bajo los términos de esta licencia), y esta licencia continuará en pleno vigor y efecto a menos que sea terminada como se expresa atrás. La Licencia revocada continuará siendo plenamente vigente y efectiva si no se le da término en las condiciones indicadas anteriormente.</li>
    </ol>
  </li>
  <br/>
  <li>
    Varios.
    <ol type="a">
      <li>Cada vez que Usted distribuya o ponga a disposición pública la Obra o una Obra Colectiva, el Licenciante ofrecerá al destinatario una licencia en los mismos términos y condiciones que la licencia otorgada a Usted bajo esta Licencia.</li>
      <li>Si alguna disposición de esta Licencia resulta invalidada o no exigible, según la legislación vigente, esto no afectará ni la validez ni la aplicabilidad del resto de condiciones de esta Licencia y, sin acción adicional por parte de los sujetos de este acuerdo, aquélla se entenderá reformada lo mínimo necesario para hacer que dicha disposición sea válida y exigible.</li>
      <li>Ningún término o disposición de esta Licencia se estimará renunciada y ninguna violación de ella será consentida a menos que esa renuncia o consentimiento sea otorgado por escrito y firmado por la parte que renuncie o consienta.</li>
      <li>Esta Licencia refleja el acuerdo pleno entre las partes respecto a la Obra aquí licenciada. No hay arreglos, acuerdos o declaraciones respecto a la Obra que no estén especificados en este documento. El Licenciante no se verá limitado por ninguna disposición adicional que pueda surgir en alguna comunicación emanada de Usted. Esta Licencia no puede ser modificada sin el consentimiento mutuo por escrito del Licenciante y Usted.</li>
    </ol>
  </li>
  <br/>
</ol>
