Simultaneous detection of the Genus Brucella by combinatorial PCR

Department of Veterinary Science, National Institute of Infectious Diseases, Tokyo 162-8640, Japan.
Japanese journal of infectious diseases (Impact Factor: 1.16). 06/2007; 60(2-3):137-9.
Source: PubMed


We have developed a combinatorial polymerase chain reaction (PCR) procedure to identify four major species of the genus Brucella simultaneously. Four pairs of primers targeting the genes encoding a cell surface protein (BCSP31) and outer membrane proteins (omp2b, omp2a and omp31) were prepared. PCR using these primers gave rise to specific patterns of amplification for each Brucella spp. examined in this study. B. abortus could be identified when fragments of BCSP31 and omp2b/2a were amplified by B. abortus-specific primers. B. melitensis could be identified by the amplification of fragments of BCSP31, omp2b/2a and omp31 using pair of primers B4/B5, JRF/JPR-ab and omp31. Identification of B. canis could be achieved when the amplicons of omp2b/2a were detected by B. canis-specific primers, as could the identification of BCSP31 and omp31. If specific amplifications occurred using all pairs of primers, the strain was identified as B. suis. Combinatorial PCR reported here thus appeared to be an ideal method of identifying Brucella spp., the causative pathogen of human brucellosis.

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    • "For the detection of Brucella species, a multiplex realtime PCR protocol was used, targeting the bcsp31, alkB, and BMEI1162 genes of Brucella species, B. abortus, and B. melitensis, respectively, as previously described [17]. Where it was positive for Brucella species but negative for the other two, B. canis was tested using a PCR protocol targeting the omp2B gene as previously described [18]. For the detection of Leishmania species a real-time PCR protocol targeting the SSU rRNA gene was used, as previously described [19]. "
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