Publication: Caracterización imunoproteómica de derivados proteicos de "Mycobacterium bovis, Mycobacterium avium (subespecies avium y paratuberculosis) y Corinebacterium pseudotuberculosis"
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2019-04-26
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Universidad Complutense de Madrid
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Abstract
La tuberculosis animal es una zoonosis bacteriana producida por bacterias del complejo Mycobacterium tuberculosis, especialmente M. bovis y M. caprae que representa un importante problema sanitario en el hombre y los animales (domésticos y salvajes). Por esta razón, en la Unión Europea, los bovinos están sometidos a un programa de erradicación que se basa en el “diagnóstico y sacrificio” de los animales reaccionantes a las pruebas de diagnóstico oficiales, cuyo reactivo básico es la tuberculina bovina o PPDb. Esta es un derivado proteico purificado de M. bovis de composición y titulación compleja. Por todo ello nos planteamos el estudio de este reactivo, para a través del conocimiento de la composición de las tuberculinas poder mejorar el diagnóstico. En primer lugar, llevamos a cabo una caracterización inmunoproteómica utilizando anticuerpos monoclonales (AcM). Obtuvimos 4 AcMo tipo que nos han permitido identificar cinco proteínas inmunodominantes presentes en la PPDb: MPB70, MPB83, HspX y ESAT-6, exclusivas del MTBC, y meromycolate acyl carrier protein presente también en la PPDa (derivado proteico purificado de M. avium subsp. avium). Las cuatro primeras forman parte de un complejo multiproteico, que se obtiene siempre por inmunopurificación, a partir de la PPDb, con el AcM SIM 377-18 el cual reconoce un epítopo común en las proteínas MPB70 y MPB83. A este complejo multiproteico lo denominamos P22, dado que en el análisis por western-blot muestra una proteína mayoritaria con movilidad electroforética a la altura de 22 kDa.
Animal tuberculosis is a zoonotic infection caused by members of the Mycobacterium tuberculosis complex (MTBC), especially M. bovis and M. caprae. It represents an important health problem, in humans, domestic animals and wildlife. For this reason, in the European Union, cattle are subject to an eradication programme, based on " test-and slaughter" strategies, which implies the removal of animals that react positively to diagnostic tests. The official diagnostic tests are based on the cell immune response against the bovine tuberculin or PPDb. This is a purified protein derivative (PPD) prepared from M. bovis which has a complex composition and needs a careful titration. This is why, in this work, by means of studying the composition of this bovine tuberculin, we considered improving the diagnosis of the tuberculosis disease.Firstly, we carried out the immunoproteomic characterization of PPDb using monoclonal antibodies (mAbs). We obtained four types of mAb that allowed us to identify five immunodominant proteins present in PPDb, which are MPB70, MPB83, HspX and ESAT-6 exclusives in MTBC, and meromycolate acyl carrier protein, also present in the PPDa, the purified protein derivative from M. avium subsp avium. The PPDb exclusive identified proteins are included in a multiprotein complex which is always obtained by immunopurification from the PPDb with the mAb SIM 377-18 and we named P22, because in the western blot analysis shows a major protein with electrophoretic mobility at 22 kDa..
Animal tuberculosis is a zoonotic infection caused by members of the Mycobacterium tuberculosis complex (MTBC), especially M. bovis and M. caprae. It represents an important health problem, in humans, domestic animals and wildlife. For this reason, in the European Union, cattle are subject to an eradication programme, based on " test-and slaughter" strategies, which implies the removal of animals that react positively to diagnostic tests. The official diagnostic tests are based on the cell immune response against the bovine tuberculin or PPDb. This is a purified protein derivative (PPD) prepared from M. bovis which has a complex composition and needs a careful titration. This is why, in this work, by means of studying the composition of this bovine tuberculin, we considered improving the diagnosis of the tuberculosis disease.Firstly, we carried out the immunoproteomic characterization of PPDb using monoclonal antibodies (mAbs). We obtained four types of mAb that allowed us to identify five immunodominant proteins present in PPDb, which are MPB70, MPB83, HspX and ESAT-6 exclusives in MTBC, and meromycolate acyl carrier protein, also present in the PPDa, the purified protein derivative from M. avium subsp avium. The PPDb exclusive identified proteins are included in a multiprotein complex which is always obtained by immunopurification from the PPDb with the mAb SIM 377-18 and we named P22, because in the western blot analysis shows a major protein with electrophoretic mobility at 22 kDa..
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Tesis de la Universidad Complutense de Madrid, Facultad de Veterinaria, Departamento de Sanidad Animal, leída el 14-12-2018
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Veterinaria
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3109 Ciencias Veterinarias