Abstract

We describe here the application of a novel bovine interleukin-2 (IL-2) enzyme-linked immunosorbent assay (ELISA) for the measurement of antigen-specific IL-2 in cattle naturally infected with Mycobacterium bovis and in cattle vaccinated with Mycobacterium bovis BCG and then experimentally challenged with pathogenic M. bovis. Supernatants from whole-blood cultures stimulated with mycobacterial antigen (bovine purified protein derivative [PPDB] or the peptide cocktail ESAT6-CFP10) were assessed using a sandwich ELISA consisting of a new recombinant monoclonal fragment capture antibody and a commercially available polyclonal anti-bovine-IL-2. The production of IL-2 was compared to the production of gamma interferon (IFN-γ) in the same antigen-stimulated whole-blood supernatants. The data show that cattle infected with M. bovis produced quantifiable levels of antigen-specific IL-2, while IL-2 levels in cattle vaccinated with M. bovis BCG did not. Furthermore, cattle vaccinated with M. bovis BCG and then challenged with pathogenic M. bovis displayed a more rapid induction of IL-2 but ultimately had lower levels of infection-induced IL-2 than did unvaccinated challenge control cattle. These data suggest that IL-2 responses are not detectable post-BCG vaccination and that these responses may require infection with virulent M. bovis to develop. This may be useful to differentiate infected cattle from uninfected or BCG-vaccinated cattle, although the overall sensitivity is relatively low, particularly in single intradermal comparative cervical tuberculin (SICCT)-negative infected animals. Furthermore, the strength of the IL-2 response may correlate with pathology, which poses interesting questions on the immunobiology of bovine tuberculosis in contrast to human tuberculosis, which is discussed.

Highlights

  • We describe here the application of a novel bovine interleukin-2 (IL-2) enzyme-linked immunosorbent assay (ELISA) for the measurement of antigen-specific IL-2 in cattle naturally infected with Mycobacterium bovis and in cattle vaccinated with Mycobacterium bovis BCG and experimentally challenged with pathogenic M. bovis

  • Results are shown for IL-2 and IFN-␥ production in cattle naturally infected with M. bovis, cattle vaccinated with M. bovis BCG and challenged with pathogenic M. bovis, and uninfected controls

  • M. bovis BCG vaccination is known to induce positive IFN-␥ responses to PPDB, which excludes the use of PPDB as a differentiate between infected and vaccinated/protected animals (DIVA) antigen

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Summary

Introduction

We describe here the application of a novel bovine interleukin-2 (IL-2) enzyme-linked immunosorbent assay (ELISA) for the measurement of antigen-specific IL-2 in cattle naturally infected with Mycobacterium bovis and in cattle vaccinated with Mycobacterium bovis BCG and experimentally challenged with pathogenic M. bovis. Cattle vaccinated with M. bovis BCG and challenged with pathogenic M. bovis displayed a more rapid induction of IL-2 but had lower levels of infection-induced IL-2 than did unvaccinated challenge control cattle These data suggest that IL-2 responses are not detectable post-BCG vaccination and that these responses may require infection with virulent M. bovis to develop. The gamma interferon (IFN-␥) test has been used in Great Britain since 2006 as an ancillary test to the single intradermal cervical comparative tuberculin skin test (SICCT) for the diagnosis of preclinical BTB in Great Britain Recent research in both human and bovine TB fields indicates that other cytokines produced in response to TB antigen-specific stimulation may be utilized as potential diagnostic tools. T cells are involved in subunit vaccine boosts of BCG-induced protective immunity both in humans [10] and in mice [11]

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