Development of a novel SNP based assay to speciate Brucella isolates
Abstract number: p531
Stubberfield M.R., Scott J.C., Whatmore A.M.
Brucellosis is one of the most important and widespread zoonotic diseases. Therefore, rapid and unambiguous assays for the detection of Brucella species are essential. Here we describe a novel SNP based assay that can speciate Brucella isolates into their six classically recognised species. Previously, molecular approaches to speciation have been hampered by the homogeneity of the genus that has made identification of species-specific markers difficult. The SNP assay described here was developed to overcome the problems presented by this genetic homogeneity as well as mitigating the need for classical culture based biotyping which is both laborious and time consuming.
Several housekeeping genes were sequenced from a large number of Brucella isolates. This sequence data was analysed and a number of stable single nucleotide polymorphisms (SNPs) were identified that appeared to unambiguously define the six classically recognised members of the genus Brucella. Using the primer extension approach to SNP detection a single-tube multiplex assay was developed which incorporated six SNP interrogation primers.
To date more than 400 culturally confirmed isolates have been correctly identified using this assay. These included 73 isolates from human blood cultures that encompassed the pathogenic species B. melitensis, B. suis and B. abortus. The SNP multiplex assay can be used to rapidly and clearly differentiate between the six Brucella species.
In comparison to current molecular based assays this approach is all encompassing and will identify members of all currently recognised biovars within Brucella species. Future objectives involve the inclusion of additional SNPs to facilitate higher resolution beyond the species level.
|Session name:||XXIst ISTH Congress|
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