{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,6]],"date-time":"2026-10-06T14:54:53Z","timestamp":1791298493524,"version":"4.1.0"},"reference-count":67,"publisher":"American Society for Microbiology","license":[{"start":{"date-parts":[[2026,10,6]],"date-time":"2026-10-06T00:00:00Z","timestamp":1791244800000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"},{"start":{"date-parts":[[2026,10,6]],"date-time":"2026-10-06T00:00:00Z","timestamp":1791244800000},"content-version":"tdm","delay-in-days":0,"URL":"https:\/\/journals.asm.org\/non-commercial-tdm-license"}],"funder":[{"DOI":"10.13039\/100000060","name":"National Institute of Allergy and Infectious Diseases","doi-asserted-by":"publisher","award":["R01 AI141633"],"award-info":[{"award-number":["R01 AI141633"]}],"id":[{"id":"10.13039\/100000060","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/100000060","name":"National Institute of Allergy and Infectious Diseases","doi-asserted-by":"publisher","award":["T32 AI007539"],"award-info":[{"award-number":["T32 AI007539"]}],"id":[{"id":"10.13039\/100000060","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":["journals.asm.org"],"crossmark-restriction":true},"short-container-title":["J Bacteriol"],"abstract":"<jats:title>ABSTRACT<\/jats:title>\n                  <jats:sec>\n                    <jats:title\/>\n                    <jats:p>\n                      <jats:italic toggle=\"yes\">Francisella tularensis<\/jats:italic>\n                      is a highly virulent, gram-negative bacterial pathogen that causes the zoonotic disease tularemia.\n                      <jats:italic toggle=\"yes\">F. tularensis<\/jats:italic>\n                      infects a variety of host cells and replicates intracellularly while evading and interfering with host immune responses. The molecular mechanisms that facilitate the intracellular replication and virulence of\n                      <jats:italic toggle=\"yes\">F. tularensis<\/jats:italic>\n                      are poorly understood. The\n                      <jats:italic toggle=\"yes\">Francisella<\/jats:italic>\n                      genome contains a set of\n                      <jats:italic toggle=\"yes\">pil<\/jats:italic>\n                      genes that code for the assembly of surface fibers termed type IV pili (T4P). T4P are major bacterial virulence determinants, but the function of the\n                      <jats:italic toggle=\"yes\">pil<\/jats:italic>\n                      system during\n                      <jats:italic toggle=\"yes\">F. tularensis<\/jats:italic>\n                      infection and intracellular growth is unclear. T4P are closely related to the type II secretion pathway, and the\n                      <jats:italic toggle=\"yes\">pil<\/jats:italic>\n                      system of a related\n                      <jats:italic toggle=\"yes\">Francisella<\/jats:italic>\n                      species,\n                      <jats:italic toggle=\"yes\">F. novicida<\/jats:italic>\n                      , was shown to function in protein secretion as well as pilus assembly. To identify proteins secreted by\n                      <jats:italic toggle=\"yes\">F. tularensis<\/jats:italic>\n                      , we analyzed the\n                      <jats:italic toggle=\"yes\">F. tularensis<\/jats:italic>\n                      Live Vaccine Strain (LVS) using bio-orthogonal noncanonical amino acid tagging (BONCAT). Using BONCAT in conjunction with proteomics, we identified candidate proteins secreted by the wild-type LVS, as well as candidate proteins whose extracellular abundance decreased in the absence of the PilF ATPase or the PilE4 pilus subunit. Using epitope tagging of selected candidates, we validated T4P-mediated secretion of the ChiA and ChiD chitinases and the KatG catalase by the LVS. These results further our understanding of the\n                      <jats:italic toggle=\"yes\">pil<\/jats:italic>\n                      system and protein secretion pathways in\n                      <jats:italic toggle=\"yes\">F. tularensis<\/jats:italic>\n                      .\n                    <\/jats:p>\n                    <jats:sec>\n                      <jats:title>IMPORTANCE<\/jats:title>\n                      <jats:p>\n                        <jats:italic toggle=\"yes\">Francisella tularensis<\/jats:italic>\n                        is a highly virulent gram-negative bacterial pathogen and the causative agent of tularemia.\n                        <jats:italic toggle=\"yes\">F. tularensis<\/jats:italic>\n                        lacks secretion systems utilized by other intracellular bacterial pathogens but contains\n                        <jats:italic toggle=\"yes\">pil<\/jats:italic>\n                        genes that encode for type IV pili (T4P) and may also function in protein secretion. T4P are observed on the surface of all\n                        <jats:italic toggle=\"yes\">Francisella<\/jats:italic>\n                        spp., but\n                        <jats:italic toggle=\"yes\">pil<\/jats:italic>\n                        -mediated protein secretion has only been reported for\n                        <jats:italic toggle=\"yes\">F. novicida<\/jats:italic>\n                        , which is not normally pathogenic in humans. In this study, we used bio-orthogonal noncanonical amino acid tagging to identify proteins secreted by\n                        <jats:italic toggle=\"yes\">F. tularensis<\/jats:italic>\n                        , for which there is limited information. We demonstrate that the\n                        <jats:italic toggle=\"yes\">F. tularensis pil<\/jats:italic>\n                        system is capable of protein secretion and validate T4P-mediated secretion of the ChiA and ChiD chitinases and the KatG catalase. These results will facilitate investigation of\n                        <jats:italic toggle=\"yes\">Francisella<\/jats:italic>\n                        virulence mechanisms and may provide targets for therapeutic intervention.\n                      <\/jats:p>\n                    <\/jats:sec>\n                  <\/jats:sec>","DOI":"10.1128\/jb.00253-26","type":"journal-article","created":{"date-parts":[[2026,10,6]],"date-time":"2026-10-06T14:38:36Z","timestamp":1791297516000},"update-policy":"https:\/\/doi.org\/10.1128\/asmj-crossmark-policy-page","source":"Crossref","is-referenced-by-count":0,"title":["Protein secretion by the type IV pilus machinery in\n                    <i>Francisella tularensis<\/i>"],"prefix":"10.1128","author":[{"given":"Angela","family":"DeRosa","sequence":"first","affiliation":[{"id":[{"id":"https:\/\/ror.org\/05qghxh33","id-type":"ROR","asserted-by":"publisher"}],"name":"Department of Microbiology and Immunology, Renaissance School of Medicine, Stony Brook University","place":["Stony Brook, USA"]},{"id":[{"id":"https:\/\/ror.org\/05qghxh33","id-type":"ROR","asserted-by":"publisher"}],"name":"Center for Infectious Diseases, Stony Brook University","place":["Stony Brook, USA"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"P. Todd","family":"Benziger","sequence":"additional","affiliation":[{"id":[{"id":"https:\/\/ror.org\/05qghxh33","id-type":"ROR","asserted-by":"publisher"}],"name":"Department of Microbiology and Immunology, Renaissance School of Medicine, Stony Brook University","place":["Stony Brook, USA"]},{"id":[{"id":"https:\/\/ror.org\/05qghxh33","id-type":"ROR","asserted-by":"publisher"}],"name":"Center for Infectious Diseases, Stony Brook University","place":["Stony Brook, USA"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-5230-9681","authenticated-orcid":true,"given":"Vinaya","family":"Sampath","sequence":"additional","affiliation":[{"id":[{"id":"https:\/\/ror.org\/0324fzh77","id-type":"ROR","asserted-by":"publisher"}],"name":"Department of Diagnostic Health Professions, Long Island University Post","place":["Brookville, USA"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Erik J.","family":"Kopping","sequence":"additional","affiliation":[{"id":[{"id":"https:\/\/ror.org\/05qghxh33","id-type":"ROR","asserted-by":"publisher"}],"name":"Department of Microbiology and Immunology, Renaissance School of Medicine, Stony Brook University","place":["Stony Brook, USA"]},{"id":[{"id":"https:\/\/ror.org\/05qghxh33","id-type":"ROR","asserted-by":"publisher"}],"name":"Center for Infectious Diseases, Stony Brook University","place":["Stony Brook, USA"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Huiyeon","family":"Kim","sequence":"additional","affiliation":[{"id":[{"id":"https:\/\/ror.org\/05qghxh33","id-type":"ROR","asserted-by":"publisher"}],"name":"Department of Microbiology and Immunology, Renaissance School of Medicine, Stony Brook University","place":["Stony Brook, USA"]},{"id":[{"id":"https:\/\/ror.org\/05qghxh33","id-type":"ROR","asserted-by":"publisher"}],"name":"Center for Infectious Diseases, Stony Brook University","place":["Stony Brook, USA"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-9798-9040","authenticated-orcid":true,"given":"David G.","family":"Thanassi","sequence":"additional","affiliation":[{"id":[{"id":"https:\/\/ror.org\/05qghxh33","id-type":"ROR","asserted-by":"publisher"}],"name":"Department of Microbiology and Immunology, Renaissance School of Medicine, Stony Brook University","place":["Stony Brook, USA"]},{"id":[{"id":"https:\/\/ror.org\/05qghxh33","id-type":"ROR","asserted-by":"publisher"}],"name":"Center for Infectious Diseases, Stony Brook University","place":["Stony Brook, USA"]}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"235","published-online":{"date-parts":[[2026,10,6]]},"reference":[{"key":"e_1_3_3_2_2","doi-asserted-by":"publisher","DOI":"10.1128\/CMR.15.4.631-646.2002"},{"key":"e_1_3_3_3_2","doi-asserted-by":"publisher","DOI":"10.1080\/21505594.2023.2274638"},{"key":"e_1_3_3_4_2","doi-asserted-by":"publisher","DOI":"10.3389\/fcimb.2014.00035"},{"key":"e_1_3_3_5_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.celrep.2018.02.096"},{"key":"e_1_3_3_6_2","doi-asserted-by":"publisher","DOI":"10.1101\/cshperspect.a010314"},{"key":"e_1_3_3_7_2","doi-asserted-by":"publisher","DOI":"10.3389\/fmicb.2011.00009"},{"key":"e_1_3_3_8_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.micinf.2005.08.001"},{"key":"e_1_3_3_9_2","doi-asserted-by":"publisher","DOI":"10.1128\/MMBR.05027-11"},{"key":"e_1_3_3_10_2","doi-asserted-by":"publisher","DOI":"10.1128\/IAI.72.6.3204-3217.2004"},{"key":"e_1_3_3_11_2","doi-asserted-by":"publisher","DOI":"10.1189\/jlb.0406287"},{"key":"e_1_3_3_12_2","doi-asserted-by":"publisher","DOI":"10.1111\/j.1365-2958.2009.06947.x"},{"key":"e_1_3_3_13_2","doi-asserted-by":"publisher","DOI":"10.1128\/JB.186.19.6430-6436.2004"},{"key":"e_1_3_3_14_2","doi-asserted-by":"publisher","DOI":"10.3389\/fcimb.2017.00096"},{"key":"e_1_3_3_15_2","doi-asserted-by":"publisher","DOI":"10.3389\/fcimb.2013.00112"},{"key":"e_1_3_3_16_2","doi-asserted-by":"publisher","DOI":"10.1371\/journal.ppat.1000459"},{"key":"e_1_3_3_17_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.cell.2018.01.034"},{"key":"e_1_3_3_18_2","doi-asserted-by":"publisher","DOI":"10.3389\/fcimb.2018.00121"},{"key":"e_1_3_3_19_2","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.0602582103"},{"key":"e_1_3_3_20_2","doi-asserted-by":"publisher","DOI":"10.1128\/JB.02007-12"},{"key":"e_1_3_3_21_2","doi-asserted-by":"publisher","DOI":"10.1021\/pr1009756"},{"key":"e_1_3_3_22_2","doi-asserted-by":"publisher","DOI":"10.3389\/fmicb.2011.00029"},{"key":"e_1_3_3_23_2","doi-asserted-by":"publisher","DOI":"10.1196\/annals.1409.016"},{"key":"e_1_3_3_24_2","doi-asserted-by":"publisher","DOI":"10.1128\/IAI.72.5.3042-3047.2004"},{"key":"e_1_3_3_25_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.sbi.2007.12.009"},{"key":"e_1_3_3_26_2","doi-asserted-by":"publisher","DOI":"10.1111\/j.1365-2958.2008.06197.x"},{"key":"e_1_3_3_27_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.micpath.2011.05.001"},{"key":"e_1_3_3_28_2","doi-asserted-by":"publisher","DOI":"10.1128\/IAI.01726-07"},{"key":"e_1_3_3_29_2","doi-asserted-by":"publisher","DOI":"10.1099\/mic.0.2008\/018077-0"},{"key":"e_1_3_3_30_2","doi-asserted-by":"publisher","DOI":"10.1111\/j.1365-2958.2006.05061.x"},{"key":"e_1_3_3_31_2","doi-asserted-by":"publisher","DOI":"10.1111\/j.1365-2958.2006.05365.x"},{"key":"e_1_3_3_32_2","doi-asserted-by":"publisher","DOI":"10.1093\/femsre\/fuab053"},{"key":"e_1_3_3_33_2","doi-asserted-by":"publisher","DOI":"10.1128\/microbiolspec.psib-0011-2018"},{"key":"e_1_3_3_34_2","doi-asserted-by":"publisher","DOI":"10.1128\/JB.00506-06"},{"key":"e_1_3_3_35_2","doi-asserted-by":"publisher","DOI":"10.1128\/IAI.01006-06"},{"key":"e_1_3_3_36_2","doi-asserted-by":"publisher","DOI":"10.1128\/JB.187.11.3903-3908.2005"},{"key":"e_1_3_3_37_2","doi-asserted-by":"publisher","DOI":"10.1099\/mic.0.028183-0"},{"key":"e_1_3_3_38_2","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.1301740111"},{"key":"e_1_3_3_39_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.chom.2016.10.008"},{"key":"e_1_3_3_40_2","doi-asserted-by":"publisher","DOI":"10.1021\/pr500628k"},{"key":"e_1_3_3_41_2","doi-asserted-by":"publisher","DOI":"10.1128\/IAI.00257-06"},{"key":"e_1_3_3_42_2","doi-asserted-by":"publisher","DOI":"10.3389\/fcimb.2018.00182"},{"key":"e_1_3_3_43_2","doi-asserted-by":"publisher","DOI":"10.1128\/IAI.01717-06"},{"key":"e_1_3_3_44_2","doi-asserted-by":"publisher","DOI":"10.1111\/mmi.13897"},{"key":"e_1_3_3_45_2","doi-asserted-by":"publisher","DOI":"10.1111\/mmi.12509"},{"key":"e_1_3_3_46_2","doi-asserted-by":"publisher","DOI":"10.1371\/journal.pone.0224094"},{"key":"e_1_3_3_47_2","doi-asserted-by":"publisher","DOI":"10.1007\/s00284-017-1217-y"},{"key":"e_1_3_3_48_2","doi-asserted-by":"publisher","DOI":"10.1046\/j.1365-2958.2003.03546.x"},{"key":"e_1_3_3_49_2","doi-asserted-by":"publisher","DOI":"10.1128\/JB.00138-07"},{"key":"e_1_3_3_50_2","doi-asserted-by":"publisher","DOI":"10.1371\/journal.pone.0261938"},{"key":"e_1_3_3_51_2","doi-asserted-by":"publisher","DOI":"10.1128\/JB.00809-07"},{"key":"e_1_3_3_52_2","doi-asserted-by":"publisher","DOI":"10.1111\/cmi.12140"},{"key":"e_1_3_3_53_2","doi-asserted-by":"publisher","DOI":"10.1074\/jbc.M203740200"},{"key":"e_1_3_3_54_2","doi-asserted-by":"publisher","DOI":"10.3389\/fcimb.2022.979693"},{"key":"e_1_3_3_55_2","doi-asserted-by":"publisher","DOI":"10.1128\/AEM.02037-09"},{"key":"e_1_3_3_56_2","doi-asserted-by":"publisher","DOI":"10.1128\/JB.00093-11"},{"key":"e_1_3_3_57_2","doi-asserted-by":"publisher","DOI":"10.3390\/pathogens9121037"},{"key":"e_1_3_3_58_2","doi-asserted-by":"publisher","DOI":"10.1371\/journal.pone.0093119"},{"key":"e_1_3_3_59_2","doi-asserted-by":"publisher","DOI":"10.1128\/iai.00549-22"},{"key":"e_1_3_3_60_2","doi-asserted-by":"publisher","DOI":"10.1038\/nature04249"},{"key":"e_1_3_3_61_2","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.0608279103"},{"key":"e_1_3_3_62_2","doi-asserted-by":"publisher","DOI":"10.1111\/j.1365-2672.2009.04420.x"},{"key":"e_1_3_3_63_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.enzmictec.2022.110118"},{"key":"e_1_3_3_64_2","doi-asserted-by":"publisher","DOI":"10.1371\/journal.ppat.1010167"},{"key":"e_1_3_3_65_2","doi-asserted-by":"publisher","DOI":"10.1371\/journal.pgen.1010561"},{"key":"e_1_3_3_66_2","doi-asserted-by":"publisher","DOI":"10.1099\/mic.0.022491-0"},{"key":"e_1_3_3_67_2","doi-asserted-by":"publisher","DOI":"10.1111\/j.1574-6968.2008.01440.x"},{"key":"e_1_3_3_68_2","doi-asserted-by":"publisher","DOI":"10.1038\/nmeth.1318"}],"container-title":["Journal of Bacteriology"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/journals.asm.org\/doi\/pdf\/10.1128\/jb.00253-26","content-type":"application\/pdf","content-version":"vor","intended-application":"text-mining"},{"URL":"https:\/\/journals.asm.org\/doi\/pdf\/10.1128\/jb.00253-26","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,10,6]],"date-time":"2026-10-06T14:38:37Z","timestamp":1791297517000},"score":1,"resource":{"primary":{"URL":"https:\/\/journals.asm.org\/doi\/10.1128\/jb.00253-26"}},"subtitle":[],"editor":[{"given":"Tao","family":"Dong","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"editor"}]}],"short-title":[],"issued":{"date-parts":[[2026,10,6]]},"references-count":67,"alternative-id":["10.1128\/jb.00253-26"],"URL":"https:\/\/doi.org\/10.1128\/jb.00253-26","relation":{},"ISSN":["0021-9193","1098-5530"],"issn-type":[{"value":"0021-9193","type":"print"},{"value":"1098-5530","type":"electronic"}],"subject":[],"published":{"date-parts":[[2026,10,6]]},"assertion":[{"value":"2026-05-10","order":0,"name":"received","label":"Received","group":{"name":"publication_history","label":"Publication History"}},{"value":"2026-09-12","order":2,"name":"accepted","label":"Accepted","group":{"name":"publication_history","label":"Publication History"}},{"value":"2026-10-06","order":3,"name":"published","label":"Published","group":{"name":"publication_history","label":"Publication History"}}],"article-number":"e00253-26"}}