Despite widespread use of the bacille Calmette–Guérin (BCG) vaccine, tuberculosis (TB) remains a leading cause of global mortality from a single infectious agent (Mycobacterium tuberculosis or Mtb). Here, over two independent Mtb challenge studies, we demonstrate that subcutaneous vaccination of rhesus macaques (RMs) with rhesus cytomegalovirus vectors encoding Mtb antigen inserts (hereafter referred to as RhCMV/TB)—which elicit and maintain highly effector-differentiated, circulating and tissue-resident Mtb-specific CD4+ and CD8+ memory T cell responses—can reduce the overall (pulmonary and extrapulmonary) extent of Mtb infection and disease by 68%, as compared to that in unvaccinated controls, after intrabronchial challenge with the Erdman strain of Mtb at ∼1 year after the first vaccination. Fourteen of 34 RhCMV/TB-vaccinated RMs (41%) across both studies showed no TB disease by computed tomography scans or at necropsy after challenge (as compared to 0 of 17 unvaccinated controls), and ten of these RMs were Mtb-culture-negative for all tissues, an exceptional long-term vaccine effect in the RM challenge model with the Erdman strain of Mtb. These results suggest that complete vaccine-mediated immune control of highly pathogenic Mtb is possible if immune effector responses can intercept Mtb infection at its earliest stages.
Subscribe to Journal
Get full journal access for 1 year
only $4.92 per issue
All prices are NET prices.
VAT will be added later in the checkout.
Tax calculation will be finalised during checkout.
Rent or Buy article
Get time limited or full article access on ReadCube.
All prices are NET prices.
Cambier, C.J., Falkow, S. & Ramakrishnan, L. Host evasion and exploitation schemes of Mycobacterium tuberculosis. Cell 159, 1497–1509 (2014).
Orme, I.M., Robinson, R.T. & Cooper, A.M. The balance between protective and pathogenic immune responses in the TB-infected lung. Nat. Immunol. 16, 57–63 (2015).
Shaler, C.R., Horvath, C.N., Jeyanathan, M. & Xing, Z. Within the enemy's camp: contribution of the granuloma to the dissemination, persistence and transmission of Mycobacterium tuberculosis. Front. Immunol. 4, 30 (2013).
Pai, M. et al. Tuberculosis. Nat. Rev. Dis. Primers 2, 16076 (2016).
Comas, I. et al. Human T cell epitopes of Mycobacteriumtuberculosis are evolutionarily hyperconserved. Nat. Genet. 42, 498–503 (2010).
Comas, I. et al. Out-of-Africa migration and Neolithic coexpansion of Mycobacteriumtuberculosis with modern humans. Nat. Genet. 45, 1176–1182 (2013).
Dorhoi, A. & Kaufmann, S.H. Pathology and immune reactivity: understanding multidimensionality in pulmonary tuberculosis. Semin. Immunopathol. 38, 153–166 (2016).
Hawn, T.R. et al. Tuberculosis vaccines and prevention of infection. Microbiol. Mol. Biol. Rev. 78, 650–671 (2014).
Kaufmann, S.H. Future vaccination strategies against tuberculosis: thinking outside the box. Immunity 33, 567–577 (2010).
Griffiths, K.L. et al. Targeting dendritic cells to accelerate T cell activation overcomes a bottleneck in tuberculosis vaccine efficacy. Nat. Commun. 7, 13894 (2016).
Barclay, W.R. et al. Protection of monkeys against airborne tuberculosis by aerosol vaccination with bacillus Calmette–Guerin. Am. Rev. Respir. Dis. 107, 351–358 (1973).
Kaushal, D. et al. Mucosal vaccination with attenuated Mycobacteriumtuberculosis induces strong central memory responses and protects against tuberculosis. Nat. Commun. 6, 8533 (2015).
Verreck, F.A.W. et al. Variable BCG efficacy in rhesus populations: pulmonary BCG provides protection where standard intradermal vaccination fails. Tuberculosis (Edinb.) 104, 46–57 (2017).
Jeyanathan, M. et al. AdHu5Ag85A respiratory mucosal boost immunization enhances protection against pulmonary tuberculosis in BCG-primed nonhuman primates. PLoS One 10, e0135009 (2015).
Darrah, P.A. et al. Aerosol vaccination with AERAS-402 elicits robust cellular immune responses in the lungs of rhesus macaques but fails to protect against high-dose Mycobacteriumtuberculosis challenge. J. Immunol. 193, 1799–1811 (2014).
Verreck, F.A. et al. MVA.85A boosting of BCG and an attenuated, phoP-deficient M.tuberculosis vaccine both show protective efficacy against tuberculosis in rhesus macaques. PLoS One 4, e5264 (2009).
Tameris, M.D. et al. Safety and efficacy of MVA85A, a new tuberculosis vaccine, in infants previously vaccinated with BCG: a randomized, placebo-controlled phase 2b trial. Lancet 381, 1021–1028 (2013).
Tameris, M. et al. The candidate TB vaccine, MVA85A, induces highly durable TH1 responses. PLoS One 9, e87340 (2014).
Jarvis, M.A., Hansen, S.G., Nelson, J.A., Picker, L.J. & Früh, K. in Cytomegaloviruses: From Molecular Pathogenesis to Intervention Vol. 2 (ed. Reddehase, M.J.) 450–462 (Caister Academic Press, 2013).
Cicin-Sain, L. et al. Cytomegalovirus-specific T cell immunity is maintained in immunosenescent rhesus macaques. J. Immunol. 187, 1722–1732 (2011).
Sylwester, A.W. et al. Broadly targeted human-cytomegalovirus-specific CD4+ and CD8+ T cells dominate the memory compartments of exposed subjects. J. Exp. Med. 202, 673–685 (2005).
Hansen, S.G. et al. Effector memory T cell responses are associated with protection of rhesus monkeys from mucosal simian immunodeficiency virus challenge. Nat. Med. 15, 293–299 (2009).
Hansen, S.G. et al. Profound early control of highly pathogenic SIV by an effector memory T cell vaccine. Nature 473, 523–527 (2011).
Hansen, S.G. et al. Immune clearance of highly pathogenic SIV infection. Nature 502, 100–104 (2013).
Hansen, S.G. et al. Broadly targeted CD8+ T cell responses restricted by major histocompatibility complex E. Science 351, 714–720 (2016).
Scanga, C.A. & Flynn, J.L. Modeling tuberculosis in nonhuman primates. Cold Spring Harb. Perspect. Med. 4, a018564 (2014).
Sharpe, S. et al. Ultra-low-dose aerosol challenge with Mycobacteriumtuberculosis leads to divergent outcomes in rhesus and cynomolgus macaques. Tuberculosis 96, 1–12 (2016).
Gormus, B.J., Blanchard, J.L., Alvarez, X.H. & Didier, P.J. Evidence for a rhesus monkey model of asymptomatic tuberculosis. J. Med. Primatol. 33, 134–145 (2004).
Sibley, L. et al. Route of delivery to the airway influences the distribution of pulmonary disease but not the outcome of Mycobacteriumtuberculosis infection in rhesus macaques. Tuberculosis 96, 141–149 (2016).
Mothé, B.R. et al. The TB-specific CD4+ T cell immune repertoire in both cynomolgus and rhesus macaques largely overlap with humans. Tuberculosis (Edinb.) 95, 722–735 (2015).
Langermans, J.A. et al. Divergent effect of bacillus Calmette–Guérin (BCG) vaccination on Mycobacteriumtuberculosis infection in highly related macaque species: implications for primate models in tuberculosis vaccine research. Proc. Natl. Acad. Sci. USA 98, 11497–11502 (2001).
Hsu, T. et al. The primary mechanism of attenuation of bacillus Calmette–Guerin is a loss of secreted lytic function required for invasion of lung interstitial tissue. Proc. Natl. Acad. Sci. USA 100, 12420–12425 (2003).
Hansen, S.G. et al. Cytomegalovirus vectors violate CD8+ T cell epitope recognition paradigms. Science 340, 1237874 (2013).
Zak, D.E. et al. A blood RNA signature for tuberculosis disease risk: a prospective cohort study. Lancet 387, 2312–2322 (2016).
Cliff, J.M., Kaufmann, S.H., McShane, H., van Helden, P. & O'Garra, A. The human immune response to tuberculosis and its treatment: a view from the blood. Immunol. Rev. 264, 88–102 (2015).
Berry, M.P. et al. An interferon-inducible neutrophil-driven blood transcriptional signature in human tuberculosis. Nature 466, 973–977 (2010).
Kaforou, M. et al. Detection of tuberculosis in HIV-infected and uninfected African adults using whole-blood RNA expression signatures: a case–control study. PLoS Med. 10, e1001538 (2013).
De Libero, G., Singhal, A., Lepore, M. & Mori, L. Nonclassical T cells and their antigens in tuberculosis. Cold Spring Harb. Perspect. Med. 4, a018473 (2014).
Rayner, E.L. et al. Early lesions following aerosol infection of rhesus macaques (Macacamulatta) with Mycobacteriumtuberculosis strain H37RV. J. Comp. Pathol. 149, 475–485 (2013).
Steinert, E.M. et al. Quantifying memory CD8 T cells reveals regionalization of immunosurveillance. Cell 161, 737–749 (2015).
Thome, J.J. & Farber, D.L. Emerging concepts in tissue-resident T cells: lessons from humans. Trends Immunol. 36, 428–435 (2015).
Dallenga, T. & Schaible, U.E. Neutrophils in tuberculosis—first line of defense or booster of disease and targets for host-directed therapy? Pathog. Dis. 74, ftw012 (2016).
Mishra, B.B. et al. Nitric oxide prevents a pathogen-permissive granulocytic inflammation during tuberculosis. Nat. Microbiol. 2, 17072 (2017).
Ong, C.W. et al. Neutrophil-derived MMP-8 drives AMPK-dependent matrix destruction in human pulmonary tuberculosis. PLoS Pathog. 11, e1004917 (2015).
Mattila, J.T., Maiello, P., Sun, T., Via, L.E. & Flynn, J.L. Granzyme B–expressing neutrophils correlate with bacterial load in granulomas from Mycobacterium tuberculosis–infected cynomolgus macaques. Cell. Microbiol. 17, 1085–1097 (2015).
Lyadova, I.V. Neutrophils in tuberculosis: heterogeneity shapes the way? Mediators Inflamm. 2017, 8619307 (2017).
Martineau, A.R. et al. Neutrophil-mediated innate immune resistance to mycobacteria. J. Clin. Invest. 117, 1988–1994 (2007).
Warren, E., Teskey, G. & Venketaraman, V. Effector mechanisms of neutrophils within the innate immune system in response to Mycobacteriumtuberculosis infection. J. Clin. Med. 6, E16 (2017).
Seiler, P. et al. Early granuloma formation after aerosol Mycobacteriumtuberculosis infection is regulated by neutrophils via CXCR3-signaling chemokines. Eur. J. Immunol. 33, 2676–2686 (2003).
Jeyanathan, M. et al. Differentially imprinted innate immunity by mucosal boost vaccination determines antituberculosis immune protective outcomes, independent of T cell immunity. Mucosal Immunol. 6, 612–625 (2013).
Beverley, P.C. et al. A novel murine cytomegalovirus vaccine vector protects against Mycobacteriumtuberculosis. J. Immunol. 193, 2306–2316 (2014).
Knight, G.M. et al. Impact and cost-effectiveness of new tuberculosis vaccines in low- and middle-income countries. Proc. Natl. Acad. Sci. USA 111, 15520–15525 (2014).
Obermoser, G. et al. Systems-scale interactive exploration reveals quantitative and qualitative differences in response to influenza and pneumococcal vaccines. Immunity 38, 831–844 (2013).
Fabregat, A. et al. The Reactome pathway Knowledgebase. Nucleic Acids Res. 44 (D1), D481–D487 (2016).
Clemmensen, S.N. et al. Olfactomedin 4 defines a subset of human neutrophils. J. Leukoc. Biol. 91, 495–500 (2012).
Ambrose, L.R., Morel, A.S. & Warrens, A.N. Neutrophils express CD52 and exhibit complement-mediated lysis in the presence of alemtuzumab. Blood 114, 3052–3055 (2009).
National Research Council. Guide for the Care and Use of Laboratory Animals 8th edn. (The National Academies Press, 2011).
Capuano, S.V. III et al. Experimental Mycobacteriumtuberculosis infection of cynomolgus macaques closely resembles the various manifestations of human M. tuberculosis infection. Infect. Immun. 71, 5831–5844 (2003).
Rumboldt, Z., Huda, W. & All, J.W. Review of portable CT with assessment of a dedicated head CT scanner. AJNR Am. J. Neuroradiol. 30, 1630–1636 (2009).
Rubin, G.D. Lung nodule and cancer detection in computed tomography screening. J. Thorac. Imaging 30, 130–138 (2015).
Luciw, P.A. et al. Stereological analysis of bacterial load and lung lesions in nonhuman primates (rhesus macaques) experimentally infected with Mycobacterium tuberculosis. Am. J. Physiol. Lung Cell. Mol. Physiol. 301, L731–L738 (2011).
Zvi, A., Ariel, N., Fulkerson, J., Sadoff, J.C. & Shafferman, A. Whole-genome identification of Mycobacterium tuberculosis vaccine candidates by comprehensive data mining and bioinformatic analyses. BMC Med. Genomics 1, 18 (2008).
Zeileis, A. Object-oriented computation of sandwich estimators. J. Stat. Softw. 16, 1–16 (2006).
Zeileis, A. Econometric computing with HC and HAC covariance matrix estimators. J. Stat. Softw. 11, 1–17 (2004).
R Core Team. R: A Language and Environment for Statistical Computing (R Foundation for Statistical Computing, 2015).
Holm, S. A simple sequentially rejective multiple-test procedure. Scand. J. Stat. 6, 65–70 (1979).
Agresti, A. & Coull, B.A. Approximate is better than “exact” for interval estimation of binomial proportions. Am. Stat. 52, 119–126 (1998).
Dobin, A. et al. STAR: ultrafast universal RNA-seq aligner. Bioinformatics 29, 15–21 (2013).
Zimin, A.V. et al. A new rhesus macaque assembly and annotation for next-generation sequencing analyses. Biol. Direct 9, 20 (2014).
Anders, S., Pyl, P.T. & Huber, W. HTSeq—a Python framework to work with high-throughput sequencing data. Bioinformatics 31, 166–169 (2015).
Robinson, M.D. & Oshlack, A. A scaling normalization method for differential expression analysis of RNA-seq data. Genome Biol. 11, R25 (2010).
Robinson, M.D., McCarthy, D.J. & Smyth, G.K. edgeR: a Bioconductor package for differential expression analysis of digital gene expression data. Bioinformatics 26, 139–140 (2010).
Ritchie, M.E. et al. limma powers differential expression analyses for RNA sequencing and microarray studies. Nucleic Acids Res. 43, e47 (2015).
Zeileis, A. & Hothorn, T. Diagnostic checking in regression relationships. R News 2, 7–10 (2002).
We thank C. Kahl, S. Hagen, J. Bae, I. Pelletier, Y. Guo, E.M. Borst, L.S. Uebelhoer and J. Womack for technical assistance, C. Scanga and J. Flynn for guidance on the RM model of TB (including sharing of NHP protocols and provision of Mtb challenge stocks), P. Barry (University of California, Davis) and T. Shenk (Princeton University) for the 68-1 and 68-1.2 BACs, respectively, J. Flynn (University of Pittsburgh) for Mtb Erdman, W. Hanekom, L. Stuart and D. Barber for helpful discussions, D. Casimiro for manuscript review, J. Strussenberg for management of the BSL3 facility, L. Boshears for administrative assistance and A. Townsend for figure preparation. This work was supported by AERAS, the Bill and Melinda Gates Foundation (grant no. OPP1087783; A.A. and D.E.Z.) and the US National Institutes of Health (NIH; grant no. U19 AI106761 (A.A.), P51 OD011092 (ONPRC); U42 OD010426 (ONPRC)).
OHSU, L.J.P., S.G.H., D.M. and K.F. have a significant financial interest in Vir Biotechnology, Inc., a company that may have a commercial interest in the results of this research and technology. The potential individual and institutional conflicts of interest have been reviewed and managed by OHSU. T.G.E. has served as a clinical consultant to Vir Biotechnology and also has a significant financial interest in that company.
Supplementary Figures 1–10 (PDF 1296 kb)
Life Sciences Reporting Summary (PDF 164 kb)
1482 Genes comparably regulated in TB patients and unvaccinated RM from Study 1 and Study 2 after Mtb challenge (XLSX 254 kb)
Pathway enrichments for genes comparably regulated in TB patients and unvaccinated RM from Study 1 and Study 2 after Mtb challenge (XLSX 15 kb)
214 genes with post-challenge expression patterns in vaccinated and unvaccinated RM that are highly significantly associated with scaled combined outcome measure (XLSX 56 kb)
Pathway enrichments for genes with post-challenge expression patterns that are highly significantly associated with scaled combined outcome measure (XLSX 11 kb)
258 genes with pre-challenge expression patterns in RhCMV-vaccinated RM that are significantly associated with scaled combined outcome measure (XLSX 77 kb)
Lists of genes exhibiting plausible associations with specific leukocyte populations for RhCMV/TB-vaccinated RM on the day of challenge (XLSX 262 kb)
Cell population enrichments for genes with pre-challenge expression patterns in RhCMV-vaccinated RM that are significantly associated with scaled combined outcome measure (XLSX 10 kb)
Pathway enrichments for genes with pre-challenge expression patterns in RhCMV-vaccinated RM that are significantly associated with scaled combined outcome measure (XLSX 9 kb)
Wilcoxon rank sum test statistics comparing expression levels of protection signature genes in the RhCMV/TB and BCG+RhCMV/TB groups from Study 1 (XLSX 12 kb)
Source data to Fig. 1
Source data to Fig. 2
Source data to Fig. 3
Source data to Fig. 4
Source data to Fig. 5
Source data to Fig. 6
About this article
Cite this article
Hansen, S., Zak, D., Xu, G. et al. Prevention of tuberculosis in rhesus macaques by a cytomegalovirus-based vaccine. Nat Med 24, 130–143 (2018). https://doi.org/10.1038/nm.4473
CD4 T Cell Help Prevents CD8 T Cell Exhaustion and Promotes Control of <i>Mycobacterium tuberculosis</i> Infection
SSRN Electronic Journal (2021)
Ultra-low Dose Aerosol Infection of Mice with Mycobacterium tuberculosis More Closely Models Human Tuberculosis
Cell Host & Microbe (2021)
Cell Host & Microbe (2021)
Prophylactic and therapeutic HBV vaccination by an HBs‐expressing cytomegalovirus vector lacking an interferon antagonist in mice
European Journal of Immunology (2021)
Guinea Pig Cytomegalovirus Protective T Cell Antigen GP83 Is a Functional pp65 Homolog for Innate Immune Evasion and Pentamer-Dependent Virus Tropism
Journal of Virology (2021)