Korean J Vet Res.  2015 Jun;55(2):105-110. 10.14405/kjvr.2015.55.2.105.

Identification of Brucella melitensis isolates originating from Mongolia and diagnostic real-time PCR evaluation using a specific SNP

• Kang SI
• Kim JY
• Kim SM
• Lee JJ
• Sung SR
• Kim YH
• Jung SC
• Her M

• KMID: 1807798
• DOI: http://doi.org/10.14405/kjvr.2015.55.2.105

Abstract

A real-time PCR assay using hybridization probe (HybProbe) has been developed to detect Brucella (B.) melitensis strains. The primer and HybProbe sets were designed based on the gap gene of chromosome I with a specific single nucleotide polymorphism of B. melitensis. Specificity of the assay was confirmed by comparison to reference Brucella species and other related strains. In the melting curve analysis, B. melitensis generated a peak at 67degrees C unlike those for other Brucella species observed at 61degrees C. Sensitivity of the assay for B. melitensis ranged from 20 ng to 200 fg of genomic DNA. The ability to identify 94 Mongolian B. melitensis isolates using the real-time PCR assay was identical to that of classical biotyping methods and differential multiplex PCR. These data showed that this new molecular technique is a simple and quick method for detecting B. melitensis, which will be important for the control and prevention of brucellosis.

Keyword

Brucella melitensis; HybProbe; melting curve; real-time PCR

MeSH Terms

Brucella
Brucella melitensis*
Brucellosis
DNA
Freezing
Mongolia*
Multiplex Polymerase Chain Reaction
Polymorphism, Single Nucleotide
Real-Time Polymerase Chain Reaction*
Sensitivity and Specificity
DNA