{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,7,29]],"date-time":"2026-07-29T22:28:50Z","timestamp":1785364130910,"version":"3.55.0"},"reference-count":33,"publisher":"Springer Science and Business Media LLC","issue":"1","license":[{"start":{"date-parts":[[2024,12,18]],"date-time":"2024-12-18T00:00:00Z","timestamp":1734480000000},"content-version":"tdm","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0"},{"start":{"date-parts":[[2024,12,18]],"date-time":"2024-12-18T00:00:00Z","timestamp":1734480000000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0"}],"funder":[{"name":"The National Key Research and Development Program of China","award":["2021YFD1800402"],"award-info":[{"award-number":["2021YFD1800402"]}]}],"content-domain":{"domain":["link.springer.com"],"crossmark-restriction":false},"short-container-title":["Vet Res"],"abstract":"<jats:title>Abstract<\/jats:title><jats:p><jats:italic>Brucella<\/jats:italic> is an intracellular parasitic pathogen that causes the worldwide zoonotic disease brucellosis. The type IV secretion system (T4SS) is utilized to secrete various effectors to help <jats:italic>Brucella<\/jats:italic> form <jats:italic>Brucella<\/jats:italic>-containing vacuoles within the cell and accomplish intracellular trafficking and replication. <jats:italic>Brucella<\/jats:italic> has fewer recognized effector proteins than other intracellular parasites in the <jats:italic>Proteobacteria<\/jats:italic>, indicating that <jats:italic>Brucella<\/jats:italic> may contain a large number of unidentified effector proteins. In this study, the optimal conditions for inducing protein secretion from <jats:italic>Brucella<\/jats:italic> were screened, and the secreted proteins of 2308 and the T4SS-deficient mutant SV123 under optimal conditions were collected for comparative proteomics analysis. By label-free quantitative proteomics, we identified 15 differential proteins. Through the \u03b2-lactamase TEM1 assay and indirect immunofluorescence assay, we identified RS15060 and RS10635 as novel T4SS effectors. Furthermore, by constructing mutation strains and performing cell\/mouse infection experiments, we found that deletion of the <jats:italic>rs15060<\/jats:italic> gene reduced the capacity of <jats:italic>Brucella<\/jats:italic> to replicate in cells and cause chronic infection in mice. In conclusion, a novel <jats:italic>Brucella<\/jats:italic> T4SS effector protein, RS15060, was identified to be associated with virulence in this study, and the discovery of effector proteins is conducive to a more comprehensive elucidation of T4SS function as well as to uncovering the cryptic strategies of <jats:italic>Brucella<\/jats:italic> survival in cells.<\/jats:p>","DOI":"10.1186\/s13567-024-01417-4","type":"journal-article","created":{"date-parts":[[2024,12,18]],"date-time":"2024-12-18T11:31:11Z","timestamp":1734521471000},"update-policy":"https:\/\/doi.org\/10.1007\/springer_crossmark_policy","source":"Crossref","is-referenced-by-count":9,"title":["Identification of Brucella RS15060 as a novel type IV secretion system effector associated with bacterial virulence"],"prefix":"10.1186","volume":"55","author":[{"given":"Yi","family":"Yin","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Mingxing","family":"Tian","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Guangdong","family":"Zhang","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Hai","family":"Hu","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Chan","family":"Ding","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-1586-4277","authenticated-orcid":false,"given":"Shengqing","family":"Yu","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"297","published-online":{"date-parts":[[2024,12,18]]},"reference":[{"key":"1417_CR1","doi-asserted-by":"publisher","first-page":"1168","DOI":"10.1016\/j.meegid.2009.07.001","volume":"9","author":"AM Whatmore","year":"2009","unstructured":"Whatmore AM (2009) Current understanding of the genetic diversity of Brucella, an expanding genus of zoonotic pathogens. Infect Genet Evol 9:1168\u20131184","journal-title":"Infect Genet Evol"},{"key":"1417_CR2","doi-asserted-by":"publisher","first-page":"523","DOI":"10.1146\/annurev-micro-090110-102905","volume":"65","author":"VL Atluri","year":"2011","unstructured":"Atluri VL, Xavier MN, de Jong MF, den Hartigh AB, Tsolis RM (2011) Interactions of the human pathogenic Brucella species with their hosts. Annu Rev Microbiol 65:523\u2013541","journal-title":"Annu Rev Microbiol"},{"key":"1417_CR3","doi-asserted-by":"publisher","first-page":"209","DOI":"10.1016\/S0378-1135(02)00210-9","volume":"90","author":"E Moreno","year":"2002","unstructured":"Moreno E, Cloeckaert A, Moriyon I (2002) Brucella evolution and taxonomy. Vet Microbiol 90:209\u2013227","journal-title":"Vet Microbiol"},{"key":"1417_CR4","doi-asserted-by":"publisher","first-page":"1","DOI":"10.1016\/j.vetmic.2007.11.023","volume":"129","author":"MN Seleem","year":"2008","unstructured":"Seleem MN, Boyle SM, Sriranganathan N (2008) Brucella: a pathogen without classic virulence genes. Vet Microbiol 129:1\u201314","journal-title":"Vet Microbiol"},{"key":"1417_CR5","doi-asserted-by":"publisher","first-page":"137","DOI":"10.1038\/nrmicro753","volume":"1","author":"E Cascales","year":"2003","unstructured":"Cascales E, Christie PJ (2003) The versatile bacterial type IV secretion systems. Nat Rev Microbiol 1:137\u2013149","journal-title":"Nat Rev Microbiol"},{"key":"1417_CR6","doi-asserted-by":"publisher","first-page":"13637","DOI":"10.3390\/ijms222413637","volume":"22","author":"X Xiong","year":"2021","unstructured":"Xiong X, Li BW, Zhou ZX, Gu GJ, Li MJ, Liu J, Jiao HW (2021) The VirB system plays a crucial role in intracellular infection. Int J Mol Sci 22:13637","journal-title":"Int J Mol Sci"},{"key":"1417_CR7","doi-asserted-by":"publisher","first-page":"72","DOI":"10.3389\/fcimb.2015.00072","volume":"5","author":"Y Ke","year":"2015","unstructured":"Ke Y, Wang Y, Li W, Chen Z (2015) Type IV secretion system of Brucella spp. and its effectors. Front Cell Infect Microbiol 5:72","journal-title":"Front Cell Infect Microbiol"},{"key":"1417_CR8","doi-asserted-by":"publisher","DOI":"10.3389\/fmicb.2020.599205","volume":"11","author":"ZC Ma","year":"2020","unstructured":"Ma ZC, Li RR, Hu RR, Deng XY, Xu YM, Zheng W, Yi JH, Wang Y, Chen CF (2020) Brucella abortus BspJ is a nucleomodulin that inhibits macrophage apoptosis and promotes intracellular survival of Brucella. Front Microbiol 11:599205","journal-title":"Front Microbiol"},{"key":"1417_CR9","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.2105324118","volume":"118","author":"JB Luizet","year":"2021","unstructured":"Luizet JB, Raymond J, Lacerda TLS, Barbieux E, Kambarev S, Bonici M, Lembo F, Willemart K, Borg JP, Celli J, G\u00e9rard FCA, Muraille E, Gorvel JP, Salcedo S (2021) The Brucella effector BspL targets the ER-associated degradation (ERAD) pathway and delays bacterial egress from infected cells. Proc Natl Acad Sci USA 118:e2105324118","journal-title":"Proc Natl Acad Sci USA"},{"key":"1417_CR10","doi-asserted-by":"publisher","first-page":"102","DOI":"10.1038\/s41467-022-35763-8","volume":"14","author":"A Louche","year":"2023","unstructured":"Louche A, Blanco A, Lacerda TLS, Cancade-Veyre L, Lionnet C, Berge C, Rolando M, Lembo F, Borg JP, Buchrieser C, Nagahama M, G\u00e9rard FCA, Gorvel JP, Gueguen-Chaignon V, Terradot L, Salcedo SP (2023) Brucella effectors NyxA and NyxB target SENP3 to modulate the subcellular localisation of nucleolar proteins. Nat Commun 14:102","journal-title":"Nat Commun"},{"key":"1417_CR11","doi-asserted-by":"publisher","first-page":"e0072624","DOI":"10.1128\/mbio.00726-24","volume":"15","author":"A Gimenez","year":"2024","unstructured":"Gimenez A, Del Giudice MG, Lopez PV, Guaimas F, Samano-Sanchez H, Gibson TJ, Chemes LB, Arregui CO, Ugalde JE, Czibener C (2024) Brucella NpeA is a secreted Type IV effector containing an N-WASP-binding short linear motif that promotes niche formation. MBio 15:e0072624","journal-title":"MBio"},{"key":"1417_CR12","doi-asserted-by":"publisher","first-page":"81","DOI":"10.1128\/br.22.2.81-98.1958","volume":"22","author":"P Gerhardt","year":"1958","unstructured":"Gerhardt P (1958) The nutrition of brucellae. Bacteriol Rev 22:81\u201398","journal-title":"Bacteriol Rev"},{"key":"1417_CR13","doi-asserted-by":"publisher","first-page":"3045","DOI":"10.1128\/JB.01663-14","volume":"196","author":"A Zuniga-Ripa","year":"2014","unstructured":"Zuniga-Ripa A, Barbier T, Conde-Alvarez R, Martinez-Gomez E, Palacios-Chaves L, Gil-Ramirez Y, Grillo MJ, Letesson JJ, Iriarte M, Moriyon I (2014) Brucella abortus depends on pyruvate phosphate dikinase and malic enzyme but not on Fbp and GlpX fructose-1,6-bisphosphatases for full virulence in laboratory models. J Bacteriol 196:3045\u20133057","journal-title":"J Bacteriol"},{"key":"1417_CR14","doi-asserted-by":"publisher","first-page":"1044","DOI":"10.1111\/j.1462-5822.2011.01601.x","volume":"13","author":"M de Barsy","year":"2011","unstructured":"de Barsy M, Jamet A, Filopon D, Nicolas C, Laloux G, Rual JF, Muller A, Twizere JC, Nkengfac B, Vandenhaute J, Hill DE, Salcedo SP, Gorvel JP, Letesson JJ, De Bolle X (2011) Identification of a Brucella spp. secreted effector specifically interacting with human small GTPase Rab2. Cell Microbiol 13:1044\u20131058","journal-title":"Cell Microbiol"},{"key":"1417_CR15","doi-asserted-by":"publisher","first-page":"2904","DOI":"10.1128\/JB.00357-11","volume":"193","author":"F Wang","year":"2011","unstructured":"Wang F, Hu S, Gao Y, Qiao Z, Liu W, Bu Z (2011) Complete genome sequences of Brucella melitensis strains M28 and M5\u201390, with different virulence backgrounds. J Bacteriol 193:2904\u20132905","journal-title":"J Bacteriol"},{"key":"1417_CR16","doi-asserted-by":"publisher","first-page":"17","DOI":"10.1186\/s13567-017-0422-9","volume":"48","author":"Y Bao","year":"2017","unstructured":"Bao Y, Tian M, Li P, Liu J, Ding C, Yu S (2017) Characterization of Brucella abortus mutant strain Delta22915, a potential vaccine candidate. Vet Res 48:17","journal-title":"Vet Res"},{"key":"1417_CR17","doi-asserted-by":"publisher","DOI":"10.1128\/spectrum.02074-23","volume":"11","author":"Y Yin","year":"2023","unstructured":"Yin Y, Fang T, Lian Z, Zuo D, Hu H, Zhang G, Ding C, Tian M, Yu S (2023) Erythronate utilization activates VdtR regulating its metabolism to promote Brucella proliferation, inducing abortion in mice. Microbiol Spectr 11:e0207423","journal-title":"Microbiol Spectr"},{"key":"1417_CR18","doi-asserted-by":"publisher","first-page":"153","DOI":"10.1038\/nprot.2006.24","volume":"1","author":"KH Choi","year":"2006","unstructured":"Choi KH, Schweizer HP (2006) mini-Tn7 insertion in bacteria with single attTn7 sites: example Pseudomonas aeruginosa. Nat Protoc 1:153\u2013161","journal-title":"Nat Protoc"},{"key":"1417_CR19","doi-asserted-by":"publisher","DOI":"10.1371\/journal.ppat.1003556","volume":"9","author":"S Myeni","year":"2013","unstructured":"Myeni S, Child R, Ng TW, Kupko JJ 3rd, Wehrly TD, Porcella SF, Knodler LA, Celli J (2013) Brucella modulates secretory trafficking via multiple type IV secretion effector proteins. PLoS Pathog 9:e1003556","journal-title":"PLoS Pathog"},{"key":"1417_CR20","unstructured":"APT-BioCloud. https:\/\/bio-cloud.aptbiotech.com\/."},{"key":"1417_CR21","doi-asserted-by":"publisher","first-page":"1261","DOI":"10.1111\/j.1462-5822.2011.01618.x","volume":"13","author":"MI Marchesini","year":"2011","unstructured":"Marchesini MI, Herrmann CK, Salcedo SP, Gorvel JP, Comerci DJ (2011) In search of Brucella abortus type IV secretion substrates: screening and identification of four proteins translocated into host cells through VirB system. Cell Microbiol 13:1261\u20131274","journal-title":"Cell Microbiol"},{"key":"1417_CR22","doi-asserted-by":"publisher","first-page":"1378","DOI":"10.1111\/j.1365-2958.2008.06487.x","volume":"70","author":"MF de Jong","year":"2008","unstructured":"de Jong MF, Sun YH, den Hartigh AB, van Dijl JM, Tsolis RM (2008) Identification of VceA and VceC, two members of the VjbR regulon that are translocated into macrophages by the Brucella type IV secretion system. Mol Microbiol 70:1378\u20131396","journal-title":"Mol Microbiol"},{"key":"1417_CR23","doi-asserted-by":"publisher","first-page":"4041","DOI":"10.1128\/IAI.67.8.4041-4047.1999","volume":"67","author":"F Porte","year":"1999","unstructured":"Porte F, Liautard JP, Kohler S (1999) Early acidification of phagosomes containing Brucella suis is essential for intracellular survival in murine macrophages. Infect Immun 67:4041\u20134047","journal-title":"Infect Immun"},{"key":"1417_CR24","doi-asserted-by":"publisher","first-page":"1544","DOI":"10.1073\/pnas.032514299","volume":"99","author":"ML Boschiroli","year":"2002","unstructured":"Boschiroli ML, Ouahrani-Bettache S, Foulongne V, Michaux-Charachon S, Bourg G, Allardet-Servent A, Cazevieille C, Liautard JP, Ramuz M, O\u2019Callaghan D (2002) The Brucella suis virB operon is induced intracellularly in macrophages. Proc Natl Acad Sci USA 99:1544\u20131549","journal-title":"Proc Natl Acad Sci USA"},{"key":"1417_CR25","doi-asserted-by":"publisher","first-page":"141","DOI":"10.1111\/j.1365-2958.2009.06751.x","volume":"73","author":"M Llosa","year":"2009","unstructured":"Llosa M, Roy C, Dehio C (2009) Bacterial type IV secretion systems in human disease. Mol Microbiol 73:141\u2013151","journal-title":"Mol Microbiol"},{"key":"1417_CR26","doi-asserted-by":"publisher","DOI":"10.1016\/j.vetmic.2021.109007","volume":"254","author":"Z Cheng","year":"2021","unstructured":"Cheng Z, Li Z, Yin Y, Lian Z, Abdelgawad HA, Hu H, Guan X, Zuo D, Cai Y, Ding C, Wang S, Li T, Qi J, Tian M, Yu S (2021) Characteristics of Brucella abortus vaccine strain A19 reveals its potential mechanism of attenuated virulence. Vet Microbiol 254:109007","journal-title":"Vet Microbiol"},{"key":"1417_CR27","doi-asserted-by":"publisher","first-page":"3766","DOI":"10.1099\/mic.0.2007\/010462-0","volume":"153","author":"KA Aly","year":"2007","unstructured":"Aly KA, Baron C (2007) The VirB5 protein localizes to the T-pilus tips in Agrobacterium tumefaciens. Microbiology (Reading) 153:3766\u20133775","journal-title":"Microbiology (Reading)"},{"key":"1417_CR28","doi-asserted-by":"publisher","first-page":"10","DOI":"10.1136\/jmedgenet-2011-100466","volume":"49","author":"BJ van den Bosch","year":"2012","unstructured":"van den Bosch BJ, Gerards M, Sluiter W, Stegmann AP, Jongen EL, Hellebrekers DM, Oegema R, Lambrichs EH, Prokisch H, Danhauser K, Schoonderwoerd K, de Coo IF, Smeets HJ (2012) Defective NDUFA9 as a novel cause of neonatally fatal complex I disease. J Med Genet 49:10\u201315","journal-title":"J Med Genet"},{"key":"1417_CR29","doi-asserted-by":"publisher","first-page":"111","DOI":"10.1111\/cge.13089","volume":"93","author":"F Baertling","year":"2018","unstructured":"Baertling F, Sanchez-Caballero L, van den Brand MAM, Fung CW, Chan SH, Wong VC, Hellebrekers DME, de Coo IFM, Smeitink JAM, Rodenburg RJT, Nijtmans LGJ (2018) NDUFA9 point mutations cause a variable mitochondrial complex I assembly defect. Clin Genet 93:111\u2013118","journal-title":"Clin Genet"},{"key":"1417_CR30","doi-asserted-by":"publisher","first-page":"41","DOI":"10.1016\/S0306-9877(98)90252-3","volume":"51","author":"R Ramirez-Romero","year":"1998","unstructured":"Ramirez-Romero R (1998) Is Brucella abortus a facultative intracellular pathogen with mitochondria-like activity? Med Hypotheses 51:41\u201345","journal-title":"Med Hypotheses"},{"key":"1417_CR31","doi-asserted-by":"publisher","first-page":"5035","DOI":"10.1128\/IAI.01998-05","volume":"74","author":"Y He","year":"2006","unstructured":"He Y, Reichow S, Ramamoorthy S, Ding X, Lathigra R, Craig JC, Sobral BW, Schurig GG, Sriranganathan N, Boyle SM (2006) Brucella melitensis triggers time-dependent modulation of apoptosis and down-regulation of mitochondrion-associated gene expression in mouse macrophages. Infect Immun 74:5035\u20135046","journal-title":"Infect Immun"},{"key":"1417_CR32","doi-asserted-by":"publisher","first-page":"415","DOI":"10.1006\/bbrc.1998.9786","volume":"253","author":"JL Loeffen","year":"1998","unstructured":"Loeffen JL, Triepels RH, van den Heuvel LP, Schuelke M, Buskens CA, Smeets RJ, Trijbels JM, Smeitink JA (1998) cDNA of eight nuclear encoded subunits of NADH:ubiquinone oxidoreductase: human complex I cDNA characterization completed. Biochem Biophys Res Commun 253:415\u2013422","journal-title":"Biochem Biophys Res Commun"},{"key":"1417_CR33","doi-asserted-by":"publisher","first-page":"172","DOI":"10.3390\/microorganisms7060172","volume":"7","author":"R Islam","year":"2019","unstructured":"Islam R, Brown S, Taheri A, Dumenyo CK (2019) The gene encoding NAD-dependent epimerase\/dehydratase, wcaG, affects cell surface properties, virulence, and extracellular enzyme production in the soft rot phytopathogen. Pectobacterium carotovorum Microorganisms 7:172","journal-title":"Pectobacterium carotovorum Microorganisms"}],"container-title":["Veterinary Research"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/link.springer.com\/content\/pdf\/10.1186\/s13567-024-01417-4.pdf","content-type":"application\/pdf","content-version":"vor","intended-application":"text-mining"},{"URL":"https:\/\/link.springer.com\/article\/10.1186\/s13567-024-01417-4\/fulltext.html","content-type":"text\/html","content-version":"vor","intended-application":"text-mining"},{"URL":"https:\/\/link.springer.com\/content\/pdf\/10.1186\/s13567-024-01417-4.pdf","content-type":"application\/pdf","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2024,12,18]],"date-time":"2024-12-18T12:15:38Z","timestamp":1734524138000},"score":1,"resource":{"primary":{"URL":"https:\/\/veterinaryresearch.biomedcentral.com\/articles\/10.1186\/s13567-024-01417-4"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2024,12,18]]},"references-count":33,"journal-issue":{"issue":"1","published-online":{"date-parts":[[2024,12]]}},"alternative-id":["1417"],"URL":"https:\/\/doi.org\/10.1186\/s13567-024-01417-4","relation":{},"ISSN":["1297-9716"],"issn-type":[{"value":"1297-9716","type":"electronic"}],"subject":[],"published":{"date-parts":[[2024,12,18]]},"assertion":[{"value":"1 June 2024","order":1,"name":"received","label":"Received","group":{"name":"ArticleHistory","label":"Article History"}},{"value":"23 September 2024","order":2,"name":"accepted","label":"Accepted","group":{"name":"ArticleHistory","label":"Article History"}},{"value":"18 December 2024","order":3,"name":"first_online","label":"First Online","group":{"name":"ArticleHistory","label":"Article History"}},{"order":1,"name":"Ethics","group":{"name":"EthicsHeading","label":"Declarations"}},{"value":"The animal experiments were conducted under a protocol approved by the Institutional Animal Care and Use Committee at Shanghai Veterinary Research Institute, the Chinese Academy of Agricultural Sciences, in strict accordance with the guidelines in the Guide for the Care and Use of Laboratory Animals (Permission#: SV-20230319\u201306).","order":2,"name":"Ethics","group":{"name":"EthicsHeading","label":"Ethics approval and consent to participate"}},{"value":"The authors declare that they have no competing interests.","order":3,"name":"Ethics","group":{"name":"EthicsHeading","label":"Competing interests"}}],"article-number":"168"}}