Miles Katie, Churchward Colin P, McAuliffe Laura, Ayling Roger D, Nicholas Robin A J
Department of Statutory and Exotic Bacterial Diseases, Veterinary Laboratories Agency, Addlestone, Surrey, United Kingdom.
J Vet Diagn Invest. 2006 Mar;18(2):168-71. doi: 10.1177/104063870601800205.
Mycoplasma mycoides subspecies mycoides small-colony type (M. m. m. SC) is the cause of the economically important contagious bovine pleuropneumonia. Isolates from Africa and Australia have previously been documented to have a fragment of approximately 8.84 kb, which is absent in European strains. A set of polymerase chain reaction (PCR) primers over this region was designed to identify M. m. m. SC isolates and separate European strains from those of Africa/Australia. Specificity of the PCR assay was achieved through the positioning of an oligonucleotide within the insertion sequence IS1296, upstream of this deletion, which then was paired with a reverse primer, upstream of the deletion, within the 8.84 kb-deleted region or downstream of the deletion, generating fragments of 1.1 kb (all M. m. m. SC strains), 1.4 kb (African/Australian strains only) and 1.3 kb (European strains only), respectively. Identification and differentiation was specific for DNA from M. m. m. SC with no amplification of DNA from other cluster members or closely related species. The PCR products did not require differentiation by use of a restriction endonuclease, and have potential for use in detection of this organism in clinical samples.
丝状支原体丝状亚种小菌落型(M. m. m. SC)是具有重要经济影响的牛传染性胸膜肺炎的病原体。此前有文献记载,来自非洲和澳大利亚的分离株有一段约8.84 kb的片段,而欧洲菌株中没有该片段。针对该区域设计了一组聚合酶链反应(PCR)引物,用于鉴定M. m. m. SC分离株,并将欧洲菌株与非洲/澳大利亚菌株区分开来。PCR检测的特异性是通过在缺失上游的插入序列IS1296内定位一个寡核苷酸来实现的,该寡核苷酸随后与缺失上游的反向引物配对,位于8.84 kb缺失区域内或缺失下游,分别产生1.1 kb(所有M. m. m. SC菌株)、1.4 kb(仅非洲/澳大利亚菌株)和1.3 kb(仅欧洲菌株)的片段。鉴定和区分对M. m. m. SC的DNA具有特异性,不会扩增其他聚类成员或密切相关物种的DNA。PCR产物无需使用限制性内切酶进行区分,具有用于检测临床样本中该病原体的潜力。