Tanpiboonsak Suda, Paemanee Atchara, Bunyarataphan Sasinee, Tungpradabkul Sumalee
Department of Biochemistry, Faculty of Science, Mahidol University, RamaVI Road, Bangkok 10400, Thailand.
Mol Cell Probes. 2004 Apr;18(2):97-101. doi: 10.1016/j.mcp.2003.09.010.
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.
鼻疽伯克霍尔德菌和类鼻疽伯克霍尔德菌在临床综合征（鼻疽和类鼻疽）方面表现出高度相似性。其表型和基因型特征也高度相似。为了区分这两种微生物，采用了分子方法。本研究旨在鉴定鼻疽伯克霍尔德菌与类鼻疽伯克霍尔德菌相比不同的DNA片段。鼻疽伯克霍尔德菌和类鼻疽伯克霍尔德菌经Sau3AI酶切后的基因组DNA图谱具有明显差异，尤其是大小在0.9 - 1.5 Kb之间的DNA片段。克隆并测序了鼻疽伯克霍尔德菌一个900 bp的特异性DNA片段。以该特异性DNA片段为探针，进行Southern印迹杂交以区分这两个菌种。杂交图谱结果有效地阐明了这两种伯克霍尔德菌种之间的遗传差异。此外，开发了用Sau3AI酶切的聚合酶链反应 - 限制性片段长度多态性（PCR - RFLP）方法，以便更可靠、快速地鉴定这两个菌种。检测到鼻疽伯克霍尔德菌的一个650 bp的PCR - RFLP产物，而类鼻疽伯克霍尔德菌的PCR - RFLP产物则显示为250 bp和400 bp的两个片段。结果表明，本研究中的特异性DNA片段作为确保鉴定这两个菌种的遗传标记应具有相当大的用途。
