Low agreement between serological and molecular tests for the diagnosis of cattle brucellosis

Iranian Journal of Veterinary Science and Technology

دوره 17، شماره 3 - شماره پیاپی 40، آذر 2025، صفحه 61-68   

نوع مقاله: Research Articles

شناسه دیجیتال (DOI): 10.22067/ijvst.2025.89499.1412

نویسندگان

Mehran Ghaemi^{* 1}؛ Mohammad Sadegh Golvajouei²

¹Department of Pathobiology, Faculty of Veterinary Medicine, Ferdowsi University of Mashhad, Iran.

²Department of Pathobiology, School of Veterinary Medicine, Shiraz University, Iran.

چکیده

Bovine brucellosis, caused mainly by Brucella abortus, is an important cows disease that has created a widespread public health problem in humans. Diagnosis primarily  relies on serological testing; however, these assays lack sensitivity and, more importantly, specificity. In this study, we tried to compare the performance of  serological tests routinely applied in Iran with antigen detection tests. Also, we examined Brucella species circulating in cows of Fars province, Iran. In addition, the infection rate of Yersinia entrocolitica O9 strain as a probable interfering agent in Brucella spp. serological tests were evaluated. Supramammary lymph nodes were sampled from 98 Brucella spp. reactor cows of Fars province, Iran, analyzed by bacterial culture and molecular tests, including conventional, multiplex, and real-time PCR. Brucella spp. was isolated from 5.1% of cultured samples, while conventional and real-time PCR detected in 15 (15.3%) and 21 (21.4%) samples, respectively. All positive samples were identified as B. abortus. Notably, 78.6% of seropositive cows tested Brucella spp. negative by both molecular tests and culture at the time of slaughtering, which showed a high false-positive rate of serological testing. As Y. enterocolitica O9 was not detected in any lymph node samples, it could be concluded that immunological cross-reaction with this bacterium was not the reason for the few real-time PCR-positive results among Brucella reactor cows. In conclusion, real-time PCR provides valuable information about the Brucella species circulating in the slaughtered cows of each region.

کلیدواژه‌ها

Brucella abortus؛ Yersinia entrocolitica؛ real-time PCR؛ serological test

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مراجع

1.    Chisi SL, Marageni Y, Naidoo P, Zulu G, Akol GW, Van Heerden H. An evaluation of serological tests in the diagnosis of bovine brucellosis in naturally infected cattle in KwaZulu-Natal province in South Africa. Journal of the South African Veterinary Association. 2017 Feb 28;88. Doi:10.4102/jsava.v88i0.1381.
2.    Alamian S, Esmaelizad M, Zahraei T, Etemadi A, Mohammadi M, Afshar D, et al. A Novel PCR Assay for Detecting Brucella abortus and Brucella melitensis. Osong Public Health and Research Perspectives. 2017 Feb 28;8(1):65–70. Doi:10.24171/j.phrp.2017.8.1.09.
3.    Poester FP, Samartino LE, Santos RL. Pathogenesis and pathobiology of brucellosis in livestock. Revue Scientifique et Technique de l’OIE. 2013 Apr 1;32(1):105–15.Doi:org/10.20506/rst.32.1.2193.
4.    WOAH (World organization for animal health). Brucellosis (Brucella abortus, B. melitensis and B. suis). Manual of Diagnostic Tests and Vaccines for Terrestrial Animals, 13th edition. 2024. Chapter 3.1.4.
5.    Yu WL, Nielsen K. Review of Detection of Brucella sp. by Polymerase Chain Reaction. Croatian Medical Journal. 2010 Aug;51(4):306–13. Doi:10.3325/cmj.2010.51.306.
6.    CSFPH (The center for food safety and public health) Brucellosis: Brucella abortus factsheet, 2018 available from:  http://www.cfsph.iastate.edu/DiseaseInfo/factsheets.htm.
7.    Muñoz P, Marin C, Monreal D, González D, Garin-Bastuji B, Díaz RF, et al. Efficacy of several serological tests and antigens for diagnosis of bovine brucellosis in the presence of false-positive serological results due to Yersinia enterocolitica O:9. Clinical and diagnostic laboratory immunology. 2005 Jan 1;12(1):141–51. Doi:10.1128/CDLI.12.1.141-151.2005.
8.    Nielsen K, Gall D, Bermudez R, Renteria T, Moreno F, Corral A, et al. Field trial of the brucellosis fluorescence polarization assay. Journal of Immunoassay and Immunochemistry. 2002 Aug 29;23(3):307–16. Doi:10.1081/IAS-120013030.
9.    O’Leary S, Sheahan M, Sweeney T. Brucella abortus detection by PCR assay in blood, milk and lymph tissue of serologically positive cows. Research in Veterinary Science. 2006 Oct;81(2):170–6. Doi:10.1016/j.rvsc.2005.12.001.
10.    Tiwari A, Pal V, Afley P, Sharma DK, Bhatnagar CS, Bhardwaj B, et al. Real-time PCR carried out on DNA extracted from serum or blood sample is not a good method for surveillance of bovine brucellosis. Tropical Animal Health and Production. 2014 Sep 4;46(8):1519–22. Doi:10.1007/s11250-014-0664-8.
11.    Carvalho TPD, Silva LAD, Castanheira TLL,Souza TDD, Paixão TAD, Lazaro-Anton L, Tsolis RM, Santos RL. Cell and Tissue Tropism of Brucella spp.. Infection and Immunity. 2023 91:e00062-23.Doi: 10.1128/iai.00062-23.
12.    Leal-Klevezas DS, Martinez-Vazquez IO, Lopez-Merino A, Martinez-Soriano JP. Single-step PCR for detection of Brucella spp. from blood and milk of infected animals. Journal of Clinical Microbiology. 1995; 33(12):3087-3090. 
13.    Hamdy MER, Amin AS. Detection of Brucella Species in the Milk of Infected Cattle, Sheep, Goats and Camels by PCR. The Veterinary Journal. 2002 May;163(3):299–305. Doi:10.1053/tvjl.2001.0681.
14.    Amin AS, Hamdy MER, Ibrahim AK. Detection of Brucella melitensis in semen using the polymerase chain reaction assay. Veterinary Microbiology. 2001 Oct;83(1):37–44. Doi:10.1016/s0378-1135(01)00401-1.
15.    Kaynak-Onurdag F, Okten S, Sen B. Screening Brucella spp. in bovine raw milk by real-time quantitative PCR and conventional methods in a pilot region of vaccination, Edirne, Turkey. Journal of Dairy Science. 2016 May;99(5):3351–7. Doi:10.3168/jds.2015-10637.
16.    Ali S, Ali Q, Melzer F, Khan I, Akhter S, Neubauer H, et al. Isolation and identification of bovine Brucella isolates from Pakistan by biochemical tests and PCR. Tropical Animal Health and Production. 2013 Jul 19;46(1):73–8. Doi:10.1007/s11250-013-0448-6.
17.    Hoffman T, Rock K, Denis Rwabiita Mugizi, Shaman Muradrasoli, Lindahl-Rajala E, Erume J, et al. Molecular detection and characterization of Brucella species in raw informally marketed milk from Uganda. Infection ecology & epidemiology. 2016 Jan 1;6(1):32442–2. Doi:10.3402/iee.v6.32442.
18.    Dadar M, Alamian S, Behrozikhah AM, Yazdani F, Kalantari A, Etemadi A, et al. Molecular identification of Brucella species and biovars associated with animal and human infection in Iran. Veterinary Research Forum. 2019;10(4):315–21. Doi:10.30466/vrf.2018.89680.2171.
19.    Sharifiyazdi H, Haghkhah M, Behroozikhah AM, Nematgorgani E. Bacteriological and molecular investigation of B. melitensis in dairy cows in Iran. Comparative Clinical Pathology. 2010;21:269-273. 
20.    Amiri K, Jahan peyma D, Makenali AS. et al. National control of animal brucellosis manual. Iranian Veterinary Organization Press. 2016. [in Persian]   
21.    García-Yoldi D, Marín CM, Miguel MJ, Muñoz PM, Vizmanos JL, López-Goñi I. Multiplex PCR Assay for the Identification and Differentiation of all Brucella Species and the Vaccine Strains Brucella abortus S19 and RB51 and Brucella melitensis Rev1. Clinical Chemistry. 2006 Apr 1;52(4):779–81. https://doi.org/10.1373/clinchem.2005.062596
22.    Winchell JM, Wolff BJ, Tiller R, Bowen MD, Hoffmaster AR. Rapid Identification and Discrimination of Brucella Isolates by Use of Real-Time PCR and High-Resolution Melt Analysis. 2010 Mar 1;48(3):697–702. Doi:10.1128/JCM.02021-09.
23.    Stenkova AM, Isaeva MP, Rasskazov VA. Development of a multiplex PCR for detection of the Yersinia genus with identification of pathogenic species (Y. pestis, Y. pseudotuberculosis, Y. enterocolitica). Molecular Genetics, Microbiology and Virology. 2008;3:18-23.
24.    Lübeck PS, Mikael Skurnik, Ahrens P, Hoorfar J. A Multiplex PCR-Detection Assay for Yersinia enterocolitica Serotype O:9 and Brucella spp. Based on the Perosamine Synthetase Gene. Advances in experimental medicine and biology. 2004 Jan 1;451–4. Doi:10.1007/0-306-48416-1_89.