A simple method to detect and differentiate Burkholderia pseudomallei and Burkholderia thailandensis using specific flagellin gene primers.
We have previously shown that Burkholderia pseudomallei, the causative pathogen of melioidosis, may be discriminated from the closely related non-pathogenic species Burkholderia thailandensis by the presence of a 15 base pair deletion in the flagellin gene of B. thailandensis. Using specific flagellin gene primers flanking the distinctive region, PCR products of 191 and 176 bp in size were detected for B. pseudomallei and B. thailandensis, respectively. The sensitivity of detection is 20-80 colony forming units/reaction of B. pseudomallei and B. thailandensis cell suspension. To mimic the expected environmental situation, mixed populations of the two species were analyzed. The results showed that the PCR-based method could be use to distinguish the two species in a duplex reaction. In addition, we have developed a simplified method for direct PCR-based detection from soil samples. The result indicated that about 200 colonies of bacteria per reaction could be detected. This method can be applied to epidemiological studies, especially for investigating the ecological relationship between these two species in the environment.
Sonthayanon
Piengchan
P
Institute of Molecular Biology and Genetics, Mahidol University, Thailand. stprj@mahidol.ac.th
Krasao
Piamnukul
P
Wuthiekanun
Vannaporn
V
Panyim
Sakol
S
Tungpradabkul
Sumalee
S
D001210:Asia, Southeastern; D019117:Burkholderia / Q000145:classification* / Q000235:genetics; D019121:Burkholderia Infections / Q000175:diagnosis; D016957:Burkholderia pseudomallei / Q000235:genetics; D017931:DNA Primers / Q000592:standards*; D004269:DNA, Bacterial / Q000032:analysis*; D005408:Flagellin / Q000235:genetics*; D006801:Humans; D016133:Polymerase Chain Reaction; D012680:Sensitivity and Specificity; D012988:Soil Microbiology / Q000592:standards
D017931:DNA Primers; D004269:DNA, Bacterial; D005408:Flagellin