Development of a real-time PCR assay for the detection of Brucella ovis DNA in clinical samples-Reference-Cited by-同舟云学术

Development of a real-time PCR assay for the detection of Brucella ovis DNA in clinical samples

Published:2021-06-24 Issue:1-2 Volume:7 Page:17-20
ISSN:2411-0388
Container-title:Journal for Veterinary Medicine, Biotechnology and Biosafety
language:en
Short-container-title:J. Vet. Med. Biotechnol. Biosafety

Author:

Marchenko N. V.¹^ORCID,Lymanska O. Yu.¹^ORCID,Gerilovych A. P.¹^ORCID,Bolotin V. I.¹^ORCID

Affiliation:

1. National Scientific Center ‘Institute of Experimental and Clinical Veterinary Medicine’

Abstract

The etiological agent of infectious ovine epididymitis is Brucella ovis and for its direct indication in clinical samples several PCR protocols are proposed. This study describes a design and selection of the oligonucleotides for real-time PCR targeting conservative BOV_A0504 gene. The specificity of a real-time PCR was validated using 25 B. ovis field isolates and 14 microorganisms of closely related species. The detection limit of B. ovis in bacterial culture was determined as 3.5×101 CFU/mL with Ct value of 37.8. There are no detectable fluorescence signals in the clinical samples from intact animals, whereas bacteriologically confirmed material such as urine and testicle tissue samples were positive. It confirms that the assay is highly specific for detection of B. ovis DNA. Thus, the proposed real-time PCR assay enables fast detection and quantification of B. ovis in clinical material, which can be used as additional test for estimation of the health status of a sheep herd

Publisher

Kharkiv Entomological Society

Subject

Pharmacology (medical),Complementary and alternative medicine,Pharmaceutical Science

Reference14 articles.

1. Alton, G. G., Jones, L. M., Angus, R. D. and Verger, J. M. (1988) Techniques for the Brucellosis Laboratory. Paris: Institut National de la Recherche Agronomique. ISBN 2738000428.

2. Altschul, S. F., Gish, W., Miller, W., Myers, E. W. and Lipman, D. J. (1990) ‘Basic local alignment search tool’, Journal of Molecular Biology, 215(3), pp. 403–410. doi: https://doi.org/10.1016/S0022-2836(05)80360-2.

3. Bricker, B. J. (2002) ‘PCR as a diagnostic tool for Brucellosis’, Veterinary Microbiology, 90(1–4), pp. 435–446. doi: https://doi.org/10.1016/S0378-1135(02)00228-6.

4. Costa, L. F., Nozaki, C. N., Lira, N. S. C., Antunes, J. M. A. P., Xavier, M. N., Costa, E. A., Paixão, T. A., Santos, R. L. and Megid, J. (2013) ‘Species-specific nested PCR as a diagnostic tool for Brucella ovis infection in rams’, Arquivo Brasileiro de Medicina Veterinária e Zootecnia, 65(1), pp. 55–60. doi: https://doi.org/10.1590/S0102-09352013000100009.

5. Hinić, V., Brodard, I., Thomann, A., Cvetnić, Ž., Makaya, P. V., Frey, J. and Abril, C. (2008) ‘Novel identification and differentiation of Brucella melitensis, B. abortus, B. suis, B. ovis, B. canis, and B. neotomae suitable for both conventional and real-time PCR systems’, Journal of Microbiological Methods, 75(2), pp. 375–378. doi: https://doi.org/10.1016/j.mimet.2008.07.002.