Evaluation and Diagnosis of Species and Biovars of Brucella among cattles by Multiplex Polymerase Chain Reaction

Alizadehmofrad, Farzad; Parvini, Maryam

year 11, Issue 5 (November - December 2017) Iran J Med Microbiol 2017, 11(5): 107-114 | Back to browse issues page

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Alizadehmofrad F, Parvini M. Evaluation and Diagnosis of Species and Biovars of Brucella among cattles by Multiplex Polymerase Chain Reaction . Iran J Med Microbiol 2017; 11 (5) :107-114
URL: http://ijmm.ir/article-1-651-en.html

Evaluation and Diagnosis of Species and Biovars of Brucella among cattles by Multiplex Polymerase Chain Reaction

Farzad Alizadehmofrad¹

, Maryam Parvini

1- Department of Biology, Urmia Branch, Islamic Azad University, Urmia ,Iran
2- Department of Biology, Urmia Branch, Islamic Azad University, Urmia ,Iran , m.pour4500@gmail.com

Abstract: (7778 Views)

Background and Aims: Brucellosis is an infectious zoonotic disease responsible for reproductive disorders among animals and significant economic losses to the meat and milk supplychain. The purpose of this study was therefore to evaluate and diagnose the species and strains of Brucella isolates among cattles via multiplex polymerase chain reaction.
Materials and Methods: In this study, blood samples were taken from 22 cows diagnosed as having brucellosis by PCR test. Blood samples were cultured in BACTEC vials and incubated at 37 degrees Celsius for 5 days following which each sample was cultured on Brucella agar and kept for 3 days under Co₂ condition.The 16SrRNA gene was used as the target for the detection of Brucella genus and the bcsp31 gene was used for the detection of Brucella biovars. The PCR products were electrophoresed on %1 agarose and compared with standard strains.
Results: The PCR results showed a total of 22 positive samples for Brucella among cattles. Out of the 22 cattle positive samples, 19 were identified as B.abortus (biovars 1, 2, 3, 4, 5) and 3 as B. melitensis (biovars 1, 2, 3).
Conclusions: Brucella abortus biovars 1 and 3 and B .melitensis biovar 1 are the most prevalent biovars among cattles in Lorestan province, Iran. Moreover, multiplex PCR method could be used to improve the qualitative level and the Brucella diagnosis speed in laboratories.

Keywords: Brucellosis, PCR, Brucella species

Full-Text [PDF 748 kb] (1910 Downloads)

Type of Study: Original Research Article | Subject: Medical Bacteriology
Received: 2016/12/25 | Accepted: 2017/09/19 | ePublished: 2017/11/20

References

1. Molina-Flores B. Field experience with the control of Brucella melitensis from selected countries.In: Brucella melitensis in Eurasia and The Middle East. 2010;(10):2-9.

2. Gupta VK, Nayakwadi S, Kumar A, Gururaj K, Kumar A & Pawaiya RS. Markers for the molecular diagnosis of brucellosis in animals [Approaches in diagnosis and management of diseases of livestock and poultry]. Adv Anim. 2014;2(3):31-9. [DOI]

3. Büyük F, Şahin M. Invesigation of Brucella species from various samples of aborted cattle in Turkey by cultural and molecular methods and epidemiological analysis of cases. vetdergi. kafkas.edu 2011;17(5): 809–816. [DOI] [PubMed]

4. Gülhan T, Aksakal A, Ekin İH, Boynukara B. Retrospective Evaluation of Examined Materials for Diagnosis in University Yuzuncu Yil Faculty of Veterinary Medicine Microbiology Department Laboratory. vfdergi.yyu 2011;22(2):127–132. [DOI]

5. Pacheco WA, Genovez ME, Pozzi CR, Silva L.M.P, Azevedo SS, Did C. et al. Excretion of Brucella abortus vaccine B19 strain during a reproductive cycle in dairy cows.Braz. J. Microbiol 2012;43:594-601.

6. Silva J, Carina M, Cleyzer L, Gustavo A, Lara B, Paulo C.M. et al. Brucella abortus detected in cheese from the Amazon region:differentiation of a vaccine strain (B19) from the field strain in the states of Pará, Amapá and Rondônia, Brazil. Pesq. Vet. Bras 2016;36(8):705-710.

7. Silva C.L, Sales G.A, Santos Neto J.G, Silva J.S, De Lara A.P.S.S, Lima S.C.G. et al. Detecção de fraude em amostras comerciais de queijo bubalino por adição de leite bovino por meio da técnica de Reação em Cadeia da Polimerase (PCR) multiplex [Fraud detection in commercial samples of buffalo cheese by addition of bovine milk using multiplex polymerase chain reaction(PCR)]. Revta Inst 2015;74:21-29.

8. Shahbazi Y, Afshari Safavi E, Shavisi N. The epidemiological survey of animal brucellosis in Kermanshah province. IJVCS 2015;10(1):73-97.[In Persian].

9. Mirnejad R, Hosseini Doust R, Kachuei R, Mojtaba Mortazavi S, Khoobdel M, Ahamadi A. Simultaneous detection and differentiates of Brucella abortus and Brucella melitensis by combinatorial PCR. Elsevier 2012;5(1): 24-28.[In Persian].

10. Haghighi M, Ahadi A.M, Madani M, Mahzoonieh M.R. Design and optimization of single reaction Multiplex Nested PCR for enrichment of PCR product in detection of Brucella. IJVCS 2012;7(1): 33-39.[In Persian].

11. Pakzad I, Bahmani S, Ghafouryan S, Hosainzadegan H. Diagnosis of human brucellosis by PCR using l7/l12 and 16srRNA genes compared with common serological tests. AMUJ 2012; 14(3):31-39. [In Persian].

12. Peeri Dogaheh H, Valinejad Z, Pourfarzi F. Evaluation of three DNA extraction methods for detection of brucella DNA in human serum samples. AMUJ 2012;14(3): 40-48.

13. Matrone M, Keid L.B, Rocha V.C.M, Vejarano M.P. et al; Evaluation of DNA extrac tion protocols for Brucella abortus PCR detection in aborted fetuses or calves born from cows experimentally infected with strain 2308;sciel 2009;14(40): 480-489.

14. Aras Z, Taspinar M, Aydin I, Novel A. Polymerase Chain Reaction to Detect Br ucella canis in Dogs. vetdergikafkas 2015;21(2):169-172.

15. Saberi M, Hamali H, Jafari Joozani R , Nofouzi K, Noorsaadat GH. A serological and molecular (PCR) survey on abortions caused by Brucella melitensis Rev-1 vaccine strain in sheep herds of Tabriz-Iran. Jnasci 2013;2:(9)340-343. [In Persian].

16. Mohamed G, Khoudair M, Ramadan, Abdel Mon,em H, Amos PCR as a Rapid Screening Method for Differentiation of Infected and Vaccinated Cattle and Sheep with Brucellosis. Global Veterinaria 10.2013; (6): 748-756.

17. Arasoğlu T, Güllüce M, Özkan H, Adigüzel A, Şahin F. PCR detection of Brucella abortus in cow milk samples collected from Erzurum, Turkey. Turkish J. Med. Sci 2013;43:501-508. [DOI]

18. Marei A, Boghdadi G, Abdel-Hamed N, Hessin R, Abdoel T, Smits H, et al. Laboratory diagnosis of human brucellosis in Egypt and persistence of the pathogen following treatment. J Infect Dev Ctries 2011;5(11): 86-91. [DOI]

19. Wang T, Wang Z, Zhang Y, Bai L, Zhao Y, Liu, Ma A , Yu H. Polymerase chain reaction-based assays for thediagnosis of human brucellosisann-clinmicrob.biomedcentra 2014;13(11)13-31.

20. Mohamed G, Khoudair M, Ramadan, Abdel Mon,em H, Amos PCR as a Rapid Screening Method for Differentiation of Infected and Vaccinated Cattle and Sheep with Brucellosis. Global Veterinaria 10 2013;(6):748-756.

21. Zowghi E, Ebadi A, Yarahmadi M. Isolation and identification of Brucella organisms in Iran. SBMU 2008;3(4):185-188.

22. Doosti A, Ghasemidehkordi P. Application of real-time PCR for identification and differenti ation of Brucella abortus and Brucella melitensisin cattle. tru.uni-sz 2011;4:29-35.

23. Esmaeili H. Brucellosis in Islamic republic of Iran. ISMB 2014;3(3):47-57.

24. Betsy J, Darla R, Steven C, Olsen, Allen E. Evaluation of the Brucella abortus species–specific polymerase chain reaction assay, aneimproved version of the Brucella AMOS polymerase chain reaction assay for cattle, J Vet Diagn Invest 2003;15:374-378. [DOI] [PubMed]