Methods for Detecting and Differentiating Burkholderia pseudomallei and Burkholderia mallei with a Single Probe DNA SNP Assay


Researchers at TGen and Northern Arizona University (NAU) have developed a method for detecting Burkholderia pseudomallei and Burkholderia mallei from a single DNA SNP in a sample using Taqman SNP dual-probe allelic discrimination real-time PCR assay.  This assay provides a single-step, single reaction for identifying and differentiating between B. pseudomallei and B. mallei.  Further, in cases in which circumstances eliminate the possibility of one of the two species, this assay can be used as a single-probe assay for specific identification.


Melioidosis is an infectious disease endemic to Southeast Asia and northern Australia caused by the Gram-negative bacterium B. pseudomallei, with estimated mortality rates in northeast Thailand of 50% and Australia of 19%.  Additionally, B. mallei is closely related to B. pseudomallei and causes additional serious infectious diseases, such as glanders.  These infectious diseases have long incubation periods, and have been poorly diagnosed in the past due to factors including the mimicking of other common bacteria infections and the low concentration of bacteria cells in common types of diagnostic samples.  With these challenges presented by pathogenic Burkholderia species,  the methods of microbiological culture, polysaccharide microarray, ELISA, and real time PCR assays targeting DNA from a Type III Secretion System gene (TTS1) have had difficulty achieving the requisite sensitivity and specificity for a rapid and accurate diagnosis, which delays treatment and increases the risk for patients.


The assay developed by the researchers at TGen and NAU targets a DNA SNP from a subsequent point mutation that occurred after the B. mallei lineage diverged from B. pseudomallei.  Real-time PCR is carried out on both the forward and reverse primers of each probe. The assay may use samples from the environment, a human, or an animal.



Link to US Issued Patent US 9,340,837 B2

Patent Information:
For Information, Contact:
Katie Bray
Intellectual Property Counsel
The Translational Genomics Research Institute
Jolene Bowers
David Engelthaler
Paul Keim