{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,6,15]],"date-time":"2026-06-15T21:26:22Z","timestamp":1781558782689,"version":"3.54.5"},"reference-count":33,"publisher":"MDPI AG","issue":"9","license":[{"start":{"date-parts":[[2025,9,22]],"date-time":"2025-09-22T00:00:00Z","timestamp":1758499200000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"China Agriculture Research System","award":["CARS36"],"award-info":[{"award-number":["CARS36"]}]},{"name":"China Agriculture Research System","award":["24-1-8-xdny-4-nsh"],"award-info":[{"award-number":["24-1-8-xdny-4-nsh"]}]},{"name":"Qingdao Science and Technology Benefiting the People Demonstration Project","award":["CARS36"],"award-info":[{"award-number":["CARS36"]}]},{"name":"Qingdao Science and Technology Benefiting the People Demonstration Project","award":["24-1-8-xdny-4-nsh"],"award-info":[{"award-number":["24-1-8-xdny-4-nsh"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Microorganisms"],"abstract":"<jats:p>Brucellosis, a zoonotic infection caused by the intracellular pathogen Brucella, leads to chronic multi-organ damage. Currently, rapid, accurate, and sensitive diagnostic technologies are crucial for the prevention and control of brucellosis. This study describes the development of a chemiluminescent immunoassay (Bru-CLIA) for sheep and bovine brucellosis antibody detection, utilizing Brucella abortus strain A19 lipopolysaccharide-coated magnetic particles (LPS-MPs) as the serum antigen and acridinium ester-labeled recombinant streptococcal protein G (AE-SPG) for signal generation. After optimizing the assay\u2019s parameters, the Bru-CLIA demonstrated a sensitivity of approximately 1 IU\/mL and 2 IU\/mL for detecting sheep and bovine brucellosis, respectively. No cross-reactivity was observed with sera from animals immunized with Escherichia coli O157:H7, Mycobacterium tuberculosis, Vibrio cholerae, Legionella, Salmonella, Foot and Mouth Disease virus types O and A, Bovine viral diarrhea virus, Sheep contagious pleuropneumonia, Goat pox virus, or Peste des Petits Ruminants virus, indicating strong specificity. The testing of 81 sheep serum samples and 96 bovine serum samples revealed that Bru-CLIA showed 87.65% and 93.75% concordance with the ID-VET commercial kits for sheep and bovine brucellosis detection, respectively. These results demonstrate that Bru-CLIA offers high specificity, sensitivity, repeatability, and reliability, making it a viable rapid diagnostic tool for the epidemiological surveillance of brucellosis.<\/jats:p>","DOI":"10.3390\/microorganisms13092214","type":"journal-article","created":{"date-parts":[[2025,9,22]],"date-time":"2025-09-22T08:37:39Z","timestamp":1758530259000},"page":"2214","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":2,"title":["Development of a Chemiluminescence Immunoassay for the Serological Diagnosis of Sheep and Bovine Brucellosis"],"prefix":"10.3390","volume":"13","author":[{"given":"Xin","family":"Yan","sequence":"first","affiliation":[{"name":"Laboratory of Zoonoses, China Animal Health and Epidemiology Center, Qingdao 266032, China"},{"name":"Key Laboratory of Animal Biosafety Risk Prevention and Control (South), Ministry of Agriculture and Rural Affairs, Qingdao 266032, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Mingze","family":"Chen","sequence":"additional","affiliation":[{"name":"College of Agriculture, Yanbian University, Yanji 133000, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yuning","family":"Liu","sequence":"additional","affiliation":[{"name":"College of Veterinary Medicine, Qingdao Agricultural University, Qingdao 266109, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Mingjun","family":"Sun","sequence":"additional","affiliation":[{"name":"Laboratory of Zoonoses, China Animal Health and Epidemiology Center, Qingdao 266032, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Mengkun","family":"Huang","sequence":"additional","affiliation":[{"name":"Laboratory of Zoonoses, China Animal Health and Epidemiology Center, Qingdao 266032, China"},{"name":"Key Laboratory of Animal Biosafety Risk Prevention and Control (South), Ministry of Agriculture and Rural Affairs, Qingdao 266032, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jihui","family":"Jin","sequence":"additional","affiliation":[{"name":"Laboratory of Zoonoses, China Animal Health and Epidemiology Center, Qingdao 266032, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jiaqi","family":"Li","sequence":"additional","affiliation":[{"name":"Laboratory of Zoonoses, China Animal Health and Epidemiology Center, Qingdao 266032, China"},{"name":"Key Laboratory of Animal Biosafety Risk Prevention and Control (South), Ministry of Agriculture and Rural Affairs, Qingdao 266032, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Xiangxiang","family":"Sun","sequence":"additional","affiliation":[{"name":"Laboratory of Zoonoses, China Animal Health and Epidemiology Center, Qingdao 266032, China"},{"name":"Key Laboratory of Animal Biosafety Risk Prevention and Control (South), Ministry of Agriculture and Rural Affairs, Qingdao 266032, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-4039-9143","authenticated-orcid":false,"given":"Mengda","family":"Liu","sequence":"additional","affiliation":[{"name":"Laboratory of Zoonoses, China Animal Health and Epidemiology Center, Qingdao 266032, China"},{"name":"Key Laboratory of Ruminant Infectious Disease Prevention and Control (East), Ministry of Agriculture and Rural Affairs, Qingdao 266032, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Haobo","family":"Zhang","sequence":"additional","affiliation":[{"name":"Laboratory of Zoonoses, China Animal Health and Epidemiology Center, Qingdao 266032, China"},{"name":"Key Laboratory of Ruminant Infectious Disease Prevention and Control (East), Ministry of Agriculture and Rural Affairs, Qingdao 266032, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Weixing","family":"Shao","sequence":"additional","affiliation":[{"name":"Laboratory of Zoonoses, China Animal Health and Epidemiology Center, Qingdao 266032, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Shufang","family":"Sun","sequence":"additional","affiliation":[{"name":"Laboratory of Zoonoses, China Animal Health and Epidemiology Center, Qingdao 266032, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Xiaoxu","family":"Fan","sequence":"additional","affiliation":[{"name":"Laboratory of Zoonoses, China Animal Health and Epidemiology Center, Qingdao 266032, China"},{"name":"Key Laboratory of Ruminant Infectious Disease Prevention and Control (East), Ministry of Agriculture and Rural Affairs, Qingdao 266032, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Wenlong","family":"Nan","sequence":"additional","affiliation":[{"name":"Laboratory of Zoonoses, China Animal Health and Epidemiology Center, Qingdao 266032, China"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2025,9,22]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","unstructured":"Moriy\u00f3n, I., Blasco, J.M., Letesson, J.J., De Massis, F., and Moreno, E. (2023). Brucellosis and One Health: Inherited and Future Challenges. Microorganisms, 11.","DOI":"10.3390\/microorganisms11082070"},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"111","DOI":"10.1146\/annurev-animal-021815-111326","article-title":"Brucella spp. Virulence Factors and Immunity","volume":"4","author":"Byndloss","year":"2016","journal-title":"Annu. Rev. Anim. Biosci."},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"1081","DOI":"10.5455\/OVJ.2024.v14.i5.1","article-title":"Brucellosis: Unveiling the complexities of a pervasive zoonotic disease and its global impacts","volume":"14","author":"Khairullah","year":"2024","journal-title":"Open Vet. J."},{"key":"ref_4","doi-asserted-by":"crossref","unstructured":"Nyarku, R., Hassim, A., Jonker, A., and Quan, M. (2020). Development of a Genus-Specific Brucella Real-Time PCR Assay Targeting the 16S-23S rDNA Internal Transcribed Spacer from Different Specimen Types. Vet. Sci., 7.","DOI":"10.3390\/vetsci7040175"},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1080\/01652176.2024.2390944","article-title":"Detection and quantification of Brucella bortus DNA in water buffaloes (Bubalus bubalis) using droplet digital polymerase chain reaction","volume":"44","author":"Fusco","year":"2024","journal-title":"Vet. Q."},{"key":"ref_6","doi-asserted-by":"crossref","unstructured":"Song, S.Z., Li, Z.Y., Liu, Y.Y., Wu, Y.C., Yu, K.Y., and He, Z. (2025). Establishment of a rapid method for the detection of Brucella canis based on recombinase-mediated thermostable nucleic acid amplification technology. Front. Cell. Infect. Microbiol., 14.","DOI":"10.3389\/fcimb.2024.1493492"},{"key":"ref_7","doi-asserted-by":"crossref","unstructured":"Wu, Y., Yan, X., Sun, M., Guo, X., Li, J., Sun, X., Liu, M., Zhang, H., Nan, W., and Shao, W. (2025). Evaluation of twin-arginine translocation substrate proteins as potential antigen candidates for serodiagnosis of brucellosis. Front. Vet. Sci., 12.","DOI":"10.3389\/fvets.2025.1398983"},{"key":"ref_8","doi-asserted-by":"crossref","unstructured":"Loubet, P., Magnan, C., Salipante, F., Pastre, T., Keriel, A., O\u2019Callaghan, D., Sotto, A., and Lavigne, J.P. (2024). Diagnosis of brucellosis: Combining tests to improve performance. PLoS Negl. Trop. Dis., 18.","DOI":"10.1371\/journal.pntd.0012442"},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"2080","DOI":"10.14202\/vetworld.2023.2080-2085","article-title":"Comparison of diagnostic tests for detecting bovine brucellosis in animals vaccinated with S19 and RB51 strain vaccines","volume":"16","author":"Ibarra","year":"2023","journal-title":"Vet. World"},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"2295398","DOI":"10.1080\/07853890.2023.2295398","article-title":"Brucellosis: Epidemiology, pathogenesis, diagnosis and treatment-a comprehensive review","volume":"55","author":"Qureshi","year":"2023","journal-title":"Ann. Med."},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"171","DOI":"10.3329\/bjnm.v18i2.35240","article-title":"Chemiluminescence Immunoassay: Basic Mechanism and Applications","volume":"18","author":"Hasan","year":"2018","journal-title":"Bangladesh J. Nucl. Med."},{"key":"ref_12","doi-asserted-by":"crossref","unstructured":"Matsunaga, H., Makino, A., Kato, Y., Murakami, T., Yamaguchi, Y., Kumanogoh, A., Oba, Y., Fujimi, S., Honda, T., and Tomonaga, K. (2021). Radioligand Assay-Based Detection of Antibodies against SARS-CoV-2 in Hospital Workers Treating Patients with Severe COVID-19 in Japan. Viruses, 13.","DOI":"10.3390\/v13020347"},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"950","DOI":"10.1021\/acs.jafc.7b04923","article-title":"Phage-Mediated Competitive Chemiluminescent Immunoassay for Detecting Cry1Ab Toxin by Using an Anti-Idiotypic Camel Nanobody","volume":"66","author":"Qiu","year":"2018","journal-title":"J. Agric. Food Chem."},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"127080","DOI":"10.1016\/j.talanta.2024.127080","article-title":"Multivalent bifunctional nanobody to enhance the sensitivity of direct competitive chemiluminescence immunoassay for the detection of microcystin LR in lake water","volume":"283","author":"Zhao","year":"2025","journal-title":"Talanta"},{"key":"ref_15","doi-asserted-by":"crossref","unstructured":"Satyaputra, F., Hendry, S., Braddick, M., Sivabalan, P., and Norton, R. (2021). The Laboratory Diagnosis of Syphilis. J. Clin. Microbiol., 59.","DOI":"10.1128\/JCM.00100-21"},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"114269","DOI":"10.1016\/j.jviromet.2021.114269","article-title":"Development of a chemiluminescence immunoassay to accurately detect African swine fever virus antibodies in serum","volume":"298","author":"Yang","year":"2021","journal-title":"J. Virol. Methods"},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"338","DOI":"10.1111\/tbed.12811","article-title":"Development of a chemiluminescence immunoassay using recombinant non-structural epitope-based proteins to accurately differentiate foot-and-mouth disease virus-infected and vaccinated bovines","volume":"65","author":"Liu","year":"2018","journal-title":"Transbound. Emerg. Dis."},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"11","DOI":"10.1002\/bio.3908","article-title":"Chemiluminescence assay for Listeria monocytogenes based on Cu\/Co\/Ni ternary nanocatalyst coupled with penicillin as generic capturing agent","volume":"36","author":"Zhang","year":"2021","journal-title":"Luminescence"},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"13","DOI":"10.1186\/1475-2859-5-13","article-title":"Brucella spp noncanonical LPS: Structure, biosynthesis, and interaction with host immune system","volume":"5","author":"Cardoso","year":"2006","journal-title":"Microb. Cell Fact."},{"key":"ref_20","doi-asserted-by":"crossref","unstructured":"Wang, X., Wang, Y., Ma, L., Zhang, R., De, Y., Yang, X., Wang, C., and Wu, Q. (2015). Development of an improved competitive ELISA based on a monoclonal antibody against lipopolysaccharide for the detection of bovine brucellosis. BMC Vet. Res., 11.","DOI":"10.1186\/s12917-015-0436-3"},{"key":"ref_21","doi-asserted-by":"crossref","unstructured":"Abdelgawad, H.A., Lian, Z., Yin, Y., Fang, T., Tian, M., and Yu, S. (2023). Characterization of Brucella abortus Mutant A19mut2, a Potential DIVA Vaccine Candidate with a Modification on Lipopolysaccharide. Vaccines, 11.","DOI":"10.3390\/vaccines11071273"},{"key":"ref_22","doi-asserted-by":"crossref","unstructured":"Wang, L., Li, D., Zeng, D., Wang, S., Wu, J., Liu, Y., Peng, G., Xu, Z., Jia, H., and Song, C. (2024). Development of a fully automated chemiluminescent immunoassay for the quantitative and qualitative detection of antibodies against African swine fever virus p72. Microbiol. Spectr., 12.","DOI":"10.1128\/spectrum.00809-24"},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"390","DOI":"10.1002\/vms3.165","article-title":"Bovine brucellosis in Pakistan; an analysis of engagement with risk factors in smallholder farmer settings","volume":"5","author":"Arif","year":"2019","journal-title":"Vet. Med. Sci."},{"key":"ref_24","doi-asserted-by":"crossref","unstructured":"Munyua, P., Osoro, E., Hunsperger, E., Ngere, I., Muturi, M., Mwatondo, A., Marwanga, D., Ngere, P., Tiller, R., and Onyango, C.O. (2021). High incidence of human brucellosis in a rural Pastoralist community in Kenya, 2015. PLoS Negl. Trop. Dis., 15.","DOI":"10.1371\/journal.pntd.0009049"},{"key":"ref_25","doi-asserted-by":"crossref","unstructured":"Gong, F., Wei, H.X., Li, Q., Liu, L., and Li, B. (2021). Evaluation and Comparison of Serological Methods for COVID-19 Diagnosis. Front. Mol. Biosci., 8.","DOI":"10.3389\/fmolb.2021.682405"},{"key":"ref_26","doi-asserted-by":"crossref","unstructured":"Zheng, X., Duan, R.H., Gong, F., Wei, X., Dong, Y., Chen, R., Yue Liang, M., Tang, C., and Lu, L. (2022). Accuracy of serological tests for COVID-19: A systematic review and meta-analysis. Front. Public Health, 10.","DOI":"10.3389\/fpubh.2022.923525"},{"key":"ref_27","doi-asserted-by":"crossref","unstructured":"Ji, H., Chang, L., Zhao, J., Zhang, L., Jiang, X., Guo, F., and Wang, L. (2019). Evaluation of ELISA and CLIA for Treponema pallidum specific antibody detection in China: A multicenter study. J. Microbiol. Methods, 166.","DOI":"10.1016\/j.mimet.2019.105742"},{"key":"ref_28","doi-asserted-by":"crossref","unstructured":"Sekacheva, M., Boroda, A., Fatyanova, A., Rozhkov, A., and Bagmet, N. (2023). Clinical validation of the novel CLIA-CA-62 assay efficacy for early-stage breast cancer detection. Front. Oncol., 13.","DOI":"10.3389\/fonc.2023.1009863"},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"689","DOI":"10.3390\/pathogens10060689","article-title":"Evaluation of a Novel CLIA Monotest Assay for the Detection of Anti-Hepatitis E Virus-IgG and IgM: A Retrospective Comparison with a Line Blot and an ELISA","volume":"10","author":"Dichtl","year":"2021","journal-title":"Pathogens"},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"116","DOI":"10.1016\/j.ijid.2021.12.316","article-title":"An automated chemiluminescent immunoassay (CLIA) detects SARS-CoV-2 neutralizing antibody levels in COVID-19 patients and vaccinees","volume":"115","author":"Liu","year":"2022","journal-title":"Int. J. Infect. Dis."},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"3275","DOI":"10.1039\/D0NR08008J","article-title":"Ultrasensitive chemiluminescence immunoassay with enhanced precision for the detection of cTnI amplified by acridinium ester-loaded microspheres and internally calibrated by magnetic fluorescent nanoparticles","volume":"13","author":"Zhao","year":"2021","journal-title":"Nanoscale"},{"key":"ref_32","doi-asserted-by":"crossref","unstructured":"Miao, C., Shao, J., Yang, S., Wen, S., Ma, Y., Gao, S., Chang, H., and Liu, W. (2024). Development of plate-type and tubular chemiluminescence immunoassay against African swine fever virus p72. Appl. Microbiol. Biotechnol., 108.","DOI":"10.1007\/s00253-024-13249-5"},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"204","DOI":"10.1016\/j.ab.2010.07.025","article-title":"Enhanced immunoassay sensitivity using chemiluminescent acridinium esters with increased light output","volume":"406","author":"Natrajan","year":"2010","journal-title":"Anal. Biochem."}],"container-title":["Microorganisms"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/2076-2607\/13\/9\/2214\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,9]],"date-time":"2025-10-09T18:46:49Z","timestamp":1760035609000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/2076-2607\/13\/9\/2214"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2025,9,22]]},"references-count":33,"journal-issue":{"issue":"9","published-online":{"date-parts":[[2025,9]]}},"alternative-id":["microorganisms13092214"],"URL":"https:\/\/doi.org\/10.3390\/microorganisms13092214","relation":{},"ISSN":["2076-2607"],"issn-type":[{"value":"2076-2607","type":"electronic"}],"subject":[],"published":{"date-parts":[[2025,9,22]]}}}