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Title: Development and evaluation of an interferon gamma assay for the diagnosis of tuberculosis in red deer experimentally infected with Mycobacterium bovis.
Authors: Risalde, María Ángeles
Thomas, Jobin
Sevilla, Iker
Serrano, Miriam
Ortíz, Jose Antonio
Garrido, Joseba
Domínguez, Mercedes
Domínguez, Lucas
Gortázar, Christian
Ruíz-Fons, Jose Francisco
Keywords: Animal tuberculosis;Cellular immune response;Farmed Cervus elaphus;In vivo test;Mycobacterium tuberculosis complex
metadata.dc.subject.mesh: Animals
Antibodies, Bacterial
Antigens, Bacterial
Enzyme-Linked Immunosorbent Assay
Interferon-gamma Release Tests
Mycobacterium bovis
Issue Date: 16-Nov-2017
Abstract: Red deer (Cervus elaphus) is regarded as an epidemiologically relevant host for Mycobacterium bovis (M. bovis) and closely related members of the Mycobacterium tuberculosis complex that cause animal tuberculosis (TB). The standard antemortem screening test for the detection of TB in deer is the intradermal tuberculin skin test, but the detection of interferon-gamma (IFNγ) produced by white blood cells exposed to M. bovis antigens can be used as an alternative or supplemental assay in most TB eradication/control programs. This study aims to develop an in-house sandwich ELISA for deer IFNγ, based on the cross-reactivity of the antibodies to both cervid and bovine IFNγ, and to evaluate the potential of this assay to detect M. bovis-infected red deer in response to the in vitro stimulation of whole-blood cells with bovine purified protein derivative (bPPD), p22 protein complex derived from bPPD or using the specific tuberculous mycobacterial proteins ESAT-6/CFP-10, Rv3615c and Rv3020c. The positive control stimulant used in this study was pokeweed mitogen, which resulted in a consistent induction of IFNγ in samples from red deer, thus allowing the interpretation of the assay. The percentage of animals correctly classified by this technique as M. bovis non-infected was 100%. The detection of infected animals as positive was high and ranged widely depending upon the antigen and the cut-off value applied, as well as the time after infection. Our findings indicate that this protocol may serve as a reliable assay for the antemortem diagnosis of TB from the initial stage of M. bovis-infection, and may also be adequately sensitive. The suggested optimal antigens and cut-off are bPPD, p22 and the combination of ESAT-6/CFP-10 and Rv3020c with a 0.05 Δ optical density, which yielded a up to 100% correct classification of TB positive and negatve red deer under our experimental conditions. This technique will aid in TB testing of farmed and translocated deer. Future studies should evaluate the ability of this IFNγ assay to detect specific responses under field conditions.
metadata.dc.identifier.doi: 10.1186/s12917-017-1262-6
Appears in Collections:Producción 2020

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