Immune responses following Mycobacterium tuberculosis (Mtb) infection or vaccination are frequently assessed by measuring T-cell recognition of crude Mtb antigens, recombinant proteins, or peptide epitopes. We previously showed that not all Mtb-specific T cells recognize Mtb-infected macrophages. Thus, an important question is what proportion of T cells elicited by Mtb infection recognize Mtb-infected macrophages. We address this question by developing a modified elispot assay using viable Mtb-infected macrophages, a low multiplicity of infection and purified T cells. In C57BL/6 mice, CD4 and CD8 T cells were classically MHC restricted. Comparable frequencies of T cells that recognize Mtb-infected macrophages were determined using interferon-γ elispot and intracellular cytokine staining, and lung CD4 T cells more sensitively recognized Mtb-infected macrophages than lung CD8 T cells. Compared to the relatively high frequencies of T cells specific for antigens such as ESAT-6 and TB10.4, low frequencies of total pulmonary T cells elicited by aerosolized Mtb infection recognize Mtb-infected macrophages. Finally, we demonstrate that BCG vaccination elicits T cells that recognize Mtb-infected macrophages. We propose that the frequency of T cells that recognize infected macrophages could correlate with protective immunity and may be an alternative approach to measuring T-cell responses to Mtb antigens.
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We thank members of the Behar lab and Kim West (University of Massachusetts Medical School) for technical assistance and discussion. We thank Dr. Christopher Sassetti, Dr. Kadamaba Papavinasasundaram, Megan Proulx and Dr. Kenneth Rock (University of Massachusetts) for reagents, assistance and discussion. We would like to thank the National Institutes of Health Tetramer Core Facility for providing reagents. Supported by R21 AI136922 and R01 AI106725 (S.M.B.).
The authors declare no competing interests.
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Patankar, Y.R., Sutiwisesak, R., Boyce, S. et al. Limited recognition of Mycobacterium tuberculosis-infected macrophages by polyclonal CD4 and CD8 T cells from the lungs of infected mice. Mucosal Immunol 13, 140–148 (2020). https://doi.org/10.1038/s41385-019-0217-6