PCR-RFLP based differentiation of Burkholderia mallei and Burkholderia pseudomallei.
Burkholderia mallei and Burkholderia pseudomallei manifest a high similarity with regard to clinical syndromes, glanders and melioidosis. Phenotypic and genotypic characters are also highly similar. In an attempt to differentiate the two organisms, the molecular method was applied. This study aimed to identify the different DNA fragment in B. mallei, as compared with B. pseudomallei. The Sau3AI-digested genomic DNA patterns of B. mallei and B. pseudomallei are distinctive, especially the DNA fragments between 0.9-1.5 Kb in size. A 900-bp specific DNA fragment of B. mallei was cloned and sequenced. Using the specific DNA fragment as a probe, Southern blot hybridization was performed to differentiate the two species. The results of hybridization patterns are effective in to elucidating the genetic dissimilarities among these two Burkholderia species. Furthermore, polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) digested with Sau3AI was developed to allow a more reliable and rapid identification of the two species. A 650-bp PCR-RFLP product of B. mallei was detected, while two fragments of 250 and 400-bp PCR-RFLP products of B. pseudomallei were visualized. The results suggest that the specific DNA fragment in our study should be of considerable use as a genetic marker for ensuring identification of the two species.
Tanpiboonsak
Suda
S
Department of Biochemistry, Faculty of Science, Mahidol University, RamaVI Road, Bangkok 10400, Thailand.
Paemanee
Atchara
A
Bunyarataphan
Sasinee
S
Tungpradabkul
Sumalee
S
D015373:Bacterial Typing Techniques / Q000379:methods*; D015139:Blotting, Southern; D042726:Burkholderia mallei / Q000145:classification* / Q000235:genetics; D016957:Burkholderia pseudomallei / Q000145:classification* / Q000235:genetics; D016133:Polymerase Chain Reaction / Q000379:methods; D012150:Polymorphism, Restriction Fragment Length