{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,9,25]],"date-time":"2026-09-25T00:16:48Z","timestamp":1790295408395,"version":"4.1.0"},"reference-count":41,"publisher":"Frontiers Media SA","license":[{"start":{"date-parts":[[2026,3,4]],"date-time":"2026-03-04T00:00:00Z","timestamp":1772582400000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100001807","name":"Funda\u00e7\u00e3o de Amparo \u00e0 Pesquisa do Estado de S\u00e3o Paulo","doi-asserted-by":"publisher","award":["(2022\/13917-9, 2023\/02577-5"],"award-info":[{"award-number":["(2022\/13917-9, 2023\/02577-5"]}],"id":[{"id":"10.13039\/501100001807","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100005942","name":"Funda\u00e7\u00e3o Butantan","doi-asserted-by":"publisher","id":[{"id":"10.13039\/501100005942","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":["frontiersin.org"],"crossmark-restriction":true},"short-container-title":["Front. Immunol."],"abstract":"<jats:sec>\n                    <jats:title>Introduction<\/jats:title>\n                    <jats:p>\n                      Tuberculosis (TB) continues to be one of the leading infectious causes of mortality world-wide, while the Bacillus Calmette\u2013Gu\u00e9rin (BCG) vaccine provides variable protection. To address this limitation, our group developed a recombinant BCG strain expressing a detoxified\n                      <jats:italic>Escherichia coli<\/jats:italic>\n                      heat-labile toxin subunit (rBCG-LTAK63), which confers superior protection against\n                      <jats:italic>Mycobacterium tuberculosis<\/jats:italic>\n                      (Mtb). However, the mechanisms underlying this enhanced efficacy remain to be better characterized. Here, we investigated the capacity of rBCG-LTAK63 to enhance inflammasome-associated innate immune responses and its impact on T cell activation and protection against Mtb.\n                    <\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Methods<\/jats:title>\n                    <jats:p>\n                      Bone marrow-derived macrophages (BMDMs) from wild-type (C57Bl\/6),\n                      <jats:italic>Casp-1<\/jats:italic>\n                      -\/-,\n                      <jats:italic>Nlrp3<\/jats:italic>\n                      -\/-,\n                      <jats:italic>Aim-2<\/jats:italic>\n                      -\/- and phenotypically selected AIRmax and AIRmin\n                      <jats:sup>TT<\/jats:sup>\n                      mice were used to evaluate inflammasome-associated responses using IL-1\u03b2 as a primary readout. A co-culture system of inflammasome-activated BMDMs and splenocytes was employed to assess CD4\n                      <jats:sup>+<\/jats:sup>\n                      T cell activation and polarization. Additionally, immunization of AIRmax and AIRmin\n                      <jats:sup>TT<\/jats:sup>\n                      mice with BCG or rBCG-LTAK63 was performed to evaluate protection against Mtb.\n                    <\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Results<\/jats:title>\n                    <jats:p>\n                      rBCG-LTAK63 induced significantly higher IL-1\u03b2 production than parental BCG. IL-1\u03b2 production was largely ASC-associated and predominantly dependent on the NLRP3\/caspase-1 axis; however, residual IL-1\u03b2 production was still detected in\n                      <jats:italic>Casp<\/jats:italic>\n                      -\/- and\n                      <jats:italic>Nlrp3<\/jats:italic>\n                      -\/- BMDMs, indicating the contribution of additional processing pathways. Co-culture of inflammasome-activated BMDMs and splenocytes showed that rBCG-LTAK63-primed macrophages strongly promoted CD4\n                      <jats:sup>+<\/jats:sup>\n                      T cell activation and polarization toward Th1\/Th17 responses.\n                      <jats:italic>In vivo<\/jats:italic>\n                      , BCG only induced protection in AIRmax mice, while rBCG-LTAK63 induced protection in both AIRmax and AIRmin\n                      <jats:sup>TT<\/jats:sup>\n                      genotypes.\n                    <\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Conclusion<\/jats:title>\n                    <jats:p>This demonstrates that protection is achieved in a diminished innate inflammatory response scenario, but its full engagement can further reduce pulmonary bacterial loads. Together, these findings demonstrate that rBCG-LTAK63 enhances protection against TB through a broad innate activation including inflammasome-associated mechanisms.<\/jats:p>\n                  <\/jats:sec>","DOI":"10.3389\/fimmu.2026.1758476","type":"journal-article","created":{"date-parts":[[2026,3,4]],"date-time":"2026-03-04T06:58:16Z","timestamp":1772607496000},"update-policy":"https:\/\/doi.org\/10.3389\/crossmark-policy","source":"Crossref","is-referenced-by-count":2,"title":["Broad innate immune activation enhances the protective efficacy of rBCG-LTAK63 against Mycobacterium tuberculosis"],"prefix":"10.3389","volume":"17","author":[{"given":"Ana Carolina de Oliveira","family":"Carvalho","sequence":"first","affiliation":[{"name":"Laborat\u00f3rio de Desenvolvimento de Vacinas, Instituto Butantan","place":["S\u00e3o Paulo, Brazil"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Monalisa Martins","family":"Trentini","sequence":"additional","affiliation":[{"name":"Laborat\u00f3rio de Desenvolvimento de Vacinas, Instituto Butantan","place":["S\u00e3o Paulo, Brazil"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Dunia","family":"Rodriguez","sequence":"additional","affiliation":[{"name":"Laborat\u00f3rio de Desenvolvimento de Vacinas, Instituto Butantan","place":["S\u00e3o Paulo, Brazil"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"L\u00e1zaro Moreira","family":"Marques-Neto","sequence":"additional","affiliation":[{"name":"Laborat\u00f3rio de Desenvolvimento de Vacinas, Instituto Butantan","place":["S\u00e3o Paulo, Brazil"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Paulo Henrique Santana","family":"Silveira","sequence":"additional","affiliation":[{"name":"Laborat\u00f3rio de Desenvolvimento de Vacinas, Instituto Butantan","place":["S\u00e3o Paulo, Brazil"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Nancy","family":"Starobinas","sequence":"additional","affiliation":[{"name":"Laborat\u00f3rio de Imunogen\u00e9tica, Instituto Butantan","place":["S\u00e3o Paulo, Brazil"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Sergio Costa","family":"Oliveira","sequence":"additional","affiliation":[{"name":"Departamento de Imunologia, Instituto de Ci\u00eancias Biom\u00e9dicas, Universidade de S\u00e3o Paulo","place":["S\u00e3o Paulo, Brazil"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Luciana Cezar de Cerqueira","family":"Leite","sequence":"additional","affiliation":[{"name":"Laborat\u00f3rio de Desenvolvimento de Vacinas, Instituto Butantan","place":["S\u00e3o Paulo, Brazil"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Alex Issamu","family":"Kanno","sequence":"additional","affiliation":[{"name":"Laborat\u00f3rio de Desenvolvimento de Vacinas, Instituto Butantan","place":["S\u00e3o Paulo, Brazil"]}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1965","published-online":{"date-parts":[[2026,3,4]]},"reference":[{"key":"B1","first-page":"1","volume-title":"Global Tuberculosis Report 2024","year":"2024"},{"key":"B2","doi-asserted-by":"publisher","DOI":"10.3389\/fimmu.2017.01134","article-title":"What have we learnt about BCG vaccination in the last 20 years","volume":"8","author":"Dockrell","year":"2017","journal-title":"Front Immunol"},{"key":"B3","doi-asserted-by":"publisher","DOI":"10.3390\/cells12131766","article-title":"Inflammasomes: mechanisms of action and involvement in human diseases","volume":"12","author":"Bult\u00e9","year":"2023","journal-title":"Cells"},{"key":"B4","doi-asserted-by":"publisher","DOI":"10.1038\/s41423-021-00740-6","article-title":"NLRP3 inflammasome activation and cell death","volume":"18","author":"Huang","year":"2021","journal-title":"Cell Mol Immunol"},{"key":"B5","doi-asserted-by":"publisher","DOI":"10.1016\/j.immuni.2019.03.012","article-title":"Interleukin-1 and related cytokines in the regulation of inflammation and immunity","volume":"50","author":"Mantovani","year":"2019","journal-title":"Immunity"},{"key":"B6","doi-asserted-by":"publisher","DOI":"10.1128\/mBio.00679-16","article-title":"Deletion of nuoG from the Vaccine Candidate Mycobacterium bovis BCG \u0394ureC::hly Improves Protection against Tuberculosis","volume":"7","author":"Gengenbacher","year":"2016","journal-title":"mBio"},{"key":"B7","doi-asserted-by":"publisher","DOI":"10.1016\/j.celrep.2017.02.057","article-title":"Recombinant BCG expressing ESX-1 of mycobacterium marinum combines low virulence with cytosolic immune signaling and improved TB protection","volume":"18","author":"Gr\u00f6schel","year":"2017","journal-title":"Cell Rep"},{"key":"B8","doi-asserted-by":"publisher","DOI":"10.1093\/infdis\/jiu675","article-title":"The recombinant BCG \u0394ureC::hly vaccine targets the AIM2 inflammasome to induce autophagy and inflammation","volume":"211","author":"Saiga","year":"2015","journal-title":"J Infect Dis"},{"key":"B9","doi-asserted-by":"publisher","DOI":"10.3389\/fimmu.2024.1436676","article-title":"The role of inflammasomes as central inflammatory hubs in Mycobacterium tuberculosis infection","volume":"15","author":"Theobald","year":"2024","journal-title":"Front Immunol"},{"key":"B10","doi-asserted-by":"publisher","DOI":"10.1155\/2017\/2309478","article-title":"Inflammasomes in mycobacterium tuberculosis-driven immunity","volume":"2017","author":"Wawrocki","year":"2017","journal-title":"Can J Infect Dis Med Microbiol J Can Mal Infect Microbiol M\u00e9dicale"},{"key":"B11","doi-asserted-by":"publisher","DOI":"10.3389\/fimmu.2024.1321657","article-title":"The \u0394fbpA\u0394sapM candidate vaccine derived from Mycobacterium tuberculosis H37Rv is markedly immunogenic in macrophages and induces robust immunity to tuberculosis in mice","volume":"15","author":"Mishra","year":"2024","journal-title":"Front Immunol"},{"key":"B12","doi-asserted-by":"publisher","DOI":"10.1038\/351456a0","article-title":"New use of BCG for recombinant vaccines","volume":"351","author":"Stover","year":"1991","journal-title":"Nature"},{"key":"B13","doi-asserted-by":"publisher","DOI":"10.1080\/14760584.2021.1951243","article-title":"Thirty years of recombinant BCG: new trends for a centenary vaccine","volume":"20","author":"Marques-Neto","year":"2021","journal-title":"Expert Rev Vaccines"},{"key":"B14","doi-asserted-by":"publisher","DOI":"10.3389\/fimmu.2022.867195","article-title":"CRISPR\/cas9 approach to generate an auxotrophic BCG strain for unmarked expression of LTAK63 adjuvant: A tuberculosis vaccine candidate","volume":"13","author":"Moraes","year":"2022","journal-title":"Front Immunol"},{"key":"B15","doi-asserted-by":"publisher","first-page":"2109","DOI":"10.1038\/s41598-017-02003-9","article-title":"Recombinant BCG Expressing LTAK63 Adjuvant induces Superior Protection against Mycobacterium tuberculosis","volume":"7","author":"Nascimento","year":"2017","journal-title":"Sci Rep"},{"key":"B16","doi-asserted-by":"publisher","DOI":"10.1080\/21645515.2019.1669414","article-title":"Recombinant BCG expressing the LTAK63 adjuvant induces increased early and long-term immune responses against Mycobacteria","volume":"16","author":"Carvalho Dos Santos","year":"2019","journal-title":"Hum Vaccines Immunother"},{"key":"B17","doi-asserted-by":"publisher","DOI":"10.3389\/fimmu.2023.1205449","article-title":"Recombinant BCG expressing the LTAK63 adjuvant increased memory T cells and induced long-lasting protection against Mycobacterium tuberculosis challenge in mice","volume":"14","author":"Marques-Neto","year":"2023","journal-title":"Front Immunol"},{"key":"B18","doi-asserted-by":"publisher","DOI":"10.1080\/21645515.2019.1669414","article-title":"Recombinant BCG expressing the LTAK63 adjuvant induces increased early and long-term immune responses against Mycobacteria","volume":"16","author":"Carvalho Dos Santos","year":"2020","journal-title":"Hum Vaccin Immunother"},{"key":"B19","doi-asserted-by":"publisher","DOI":"10.3390\/vaccines10060831","article-title":"Recombinant BCG-LTAK63 vaccine candidate for tuberculosis induces an inflammatory profile in human macrophages","volume":"10","author":"Dos Santos","year":"2022","journal-title":"Vaccines (Basel)"},{"key":"B20","doi-asserted-by":"publisher","first-page":"e0227047","DOI":"10.1371\/journal.pone.0227047","article-title":"LTA1 and dmLT enterotoxin-based proteins activate antigen-presenting cells independent of PKA and despite distinct cell entry mechanisms","volume":"15","author":"Valli","year":"2020","journal-title":"PloS One"},{"key":"B21","doi-asserted-by":"publisher","DOI":"10.4049\/jimmunol.200.Supp.115.10","article-title":"Novel Pycard gene polymorphism impairs Nlpr3 inflammasome-induced IL-1\u03b2 production in mice selected for low inflammatory response","volume":"200","author":"Iba\u00f1ez","year":"2018","journal-title":"J Immunol"},{"key":"B22","doi-asserted-by":"publisher","DOI":"10.1093\/bioinformatics\/btp616","article-title":"edgeR: a Bioconductor package for differential expression analysis of digital gene expression data","volume":"26","author":"Robinson","year":"2010","journal-title":"Bioinformatics"},{"key":"B23","doi-asserted-by":"crossref","DOI":"10.1007\/978-1-0716-2144-8_2","article-title":"In vitro Assays to Study Inflammasome Activation in Primary Macrophages","volume-title":"The Inflammasome. Methods in Molecular Biology","author":"Banerjee","year":"2022"},{"key":"B24","doi-asserted-by":"publisher","DOI":"10.3389\/fimmu.2022.899569","article-title":"Pycard and BC017158 candidate genes of irm1 locus modulate inflammasome activation for IL-1\u03b2 Production","volume":"21","author":"Borrego","year":"2022","journal-title":"Front Immunol"},{"key":"B25","doi-asserted-by":"publisher","DOI":"10.4049\/jimmunol.179.2.1178","article-title":"IL-1 receptor-mediated signal is an essential component of myD88-dependent innate response to mycobacterium tuberculosis infection","volume":"179","author":"Fremond","year":"2007","journal-title":"J Immunol"},{"key":"B26","doi-asserted-by":"publisher","DOI":"10.1111\/j.1348-0421.2001.tb01310.x","article-title":"Role of interleukin (IL)-1 type 1 receptor in mycobacterial infection","volume":"45","author":"Sugawara","year":"2001","journal-title":"Microbiol Immunol"},{"key":"B27","doi-asserted-by":"publisher","DOI":"10.3389\/fphar.2022.1050774","article-title":"AIM2 inflammasome activation benefits the therapeutic effect of BCG in bladder carcinoma","volume":"13","author":"Zhou","year":"2022","journal-title":"Front Pharmacol"},{"key":"B28","doi-asserted-by":"publisher","first-page":"e2308511120","DOI":"10.1073\/pnas.2308511120","article-title":"The total mass, number, and distribution of immune cells in the human body","volume":"120","author":"Sender","year":"2023","journal-title":"Proc Natl Acad Sci U S A"},{"key":"B29","doi-asserted-by":"publisher","DOI":"10.1038\/ni.2222","article-title":"Dectin-1 is an extracellular pathogen sensor for the induction and processing of IL-1\u03b2 via a noncanonical caspase-8 inflammasome","volume":"13","author":"Gringhuis","year":"2012","journal-title":"Nat Immunol"},{"key":"B30","doi-asserted-by":"publisher","first-page":"e40722","DOI":"10.1371\/journal.pone.0040722","article-title":"Mycobacterium tuberculosis Infection of Dendritic Cells Leads to Partially Caspase-1\/11-Independent IL-1\u03b2 and IL-18 Secretion but Not to Pyroptosis","volume":"7","author":"Abdalla","year":"2012","journal-title":"PloS One"},{"key":"B31","doi-asserted-by":"publisher","first-page":"791","DOI":"10.4049\/jimmunol.2200642","article-title":"Bacillus Calmette-Gu\u00e9rin-Trained Macrophages Elicit a Protective Inflammatory Response against the Pathogenic Bacteria Brucella abortus","volume":"211","author":"de Araujo","year":"2023","journal-title":"J Immunol"},{"key":"B32","doi-asserted-by":"publisher","DOI":"10.4049\/jimmunol.0904189","article-title":"Cutting edge: caspase-1 independent IL-1\u03b2 Production is critical for host resistance to mycobacterium tuberculosis and does not require TLR signaling in vivo","volume":"184","author":"Mayer-Barber","year":"2010","journal-title":"J Immunol Baltim Md 1950"},{"key":"B33","doi-asserted-by":"publisher","DOI":"10.1016\/j.molimm.2017.05.001","article-title":"Pattern recognition receptors and coordinated cellular pathways involved in tuberculosis immunopathogenesis: Emerging concepts and perspectives","volume":"87","author":"Mishra","year":"2017","journal-title":"Mol Immunol"},{"key":"B34","doi-asserted-by":"publisher","DOI":"10.3389\/fimmu.2020.592333","article-title":"NOD2\/RIG-I activating inarigivir adjuvant enhances the efficacy of BCG vaccine against tuberculosis in mice","volume":"11","author":"Khan","year":"2020","journal-title":"Front Immunol"},{"key":"B35","doi-asserted-by":"publisher","DOI":"10.1038\/s41423-023-01107-9","article-title":"Integrated NLRP3, AIM2, NLRC4, Pyrin inflammasome activation and assembly drive PANoptosis","volume":"20","author":"Oh","year":"2023","journal-title":"Cell Mol Immunol"},{"key":"B36","doi-asserted-by":"publisher","DOI":"10.1126\/science.aad1210","article-title":"T helper 1 immunity requires complement-driven NLRP3 inflammasome activity in CD4 + T cells","volume":"352","author":"Arbore","year":"2016","journal-title":"Science"},{"key":"B37","doi-asserted-by":"publisher","DOI":"10.3389\/fimmu.2020.01202","article-title":"Characterizing the BCG induced macrophage and neutrophil mechanisms for defense against mycobacterium tuberculosis","volume":"11","author":"Bickett","year":"2020","journal-title":"Front Immunol"},{"key":"B38","doi-asserted-by":"publisher","DOI":"10.1007\/s00011-016-0918-1","article-title":"Distinct gene expression profiles provoked by polyacrylamide beads (Biogel) during chronic and acute inflammation in mice selected for maximal and minimal inflammatory responses","volume":"65","author":"Fernandes","year":"2016","journal-title":"Inflammation Res"},{"key":"B39","doi-asserted-by":"publisher","DOI":"10.3389\/fimmu.2019.01519","article-title":"Recombinant BCG With Bacterial Signaling Molecule Cyclic di-AMP as Endogenous Adjuvant Induces Elevated Immune Responses After Mycobacterium tuberculosis Infection","volume":"10","author":"Ning","year":"2019","journal-title":"Front Immunol"},{"key":"B40","doi-asserted-by":"publisher","DOI":"10.1038\/nm.1928","article-title":"Autophagy enhances the efficacy of BCG vaccine by increasing peptide presentation in mouse dendritic cells","volume":"15","author":"Jagannath","year":"2009","journal-title":"Nat Med"},{"key":"B41","doi-asserted-by":"publisher","DOI":"10.3389\/fimmu.2025.1695560","article-title":"rBCG-LTAK63 enhances protection against tuberculosis by inducing autophagy and circadian gene regulation","volume":"14","author":"Marques-Neto","year":"2025","journal-title":"Front Immunol"}],"container-title":["Frontiers in Immunology"],"original-title":[],"link":[{"URL":"https:\/\/www.frontiersin.org\/articles\/10.3389\/fimmu.2026.1758476\/full","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,3,4]],"date-time":"2026-03-04T06:58:18Z","timestamp":1772607498000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.frontiersin.org\/articles\/10.3389\/fimmu.2026.1758476\/full"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2026,3,4]]},"references-count":41,"alternative-id":["10.3389\/fimmu.2026.1758476"],"URL":"https:\/\/doi.org\/10.3389\/fimmu.2026.1758476","relation":{},"ISSN":["1664-3224"],"issn-type":[{"value":"1664-3224","type":"electronic"}],"subject":[],"published":{"date-parts":[[2026,3,4]]},"article-number":"1758476"}}