Abiven P, Blanchard B, Saillard C, Kobisch M, Bove J M
Centre National D'Etudes Véterinaires et Alimentaires, Laboratoire Central de Recherches Avicole et Porcine, Ploufragan, France.
Mol Cell Probes. 1992 Oct;6(5):423-9. doi: 10.1016/0890-8508(92)90037-x.
Mycoplasma hyopneumoniae is the primary agent of swine enzootic pneumonia. Because of fastidious growth requirements and its serological cross-reactions with other porcine mycoplasmas, we developed a specific DNA probe for its detection. A partial genomic library of M. hyopneumoniae was constructed in plasmid pBR 322 using Hind III chromosomal fragments. The recombinant plasmids were screened by differential hybridization with M. flocculare and M. hyorhinis genomic DNA probes. One non-hybridizing recombinant plasmid was selected and its 1.65 kbp insert (designated I141) tested for specificity against genomic DNA from numerous mycoplasmas, other bacteria species and DNA from lung tissue of specific pathogen free (SPF) piglets. The 32P labelled I141 could detect specifically down to 400 pg of M. hyopneumoniae genomic DNA. To test the suitability of the I141 probe for the laboratory diagnosis of M. hyopneumoniae infections, we used clinical tracheobronchial specimens from piglets which were experimentally infected with M. hyopneumoniae. The results with hybridization on each specimen were compared to findings with an immunofluorescence test. Of the clinical specimen tested, there was agreement in the two tests of 63%.
猪肺炎支原体是猪地方流行性肺炎的主要病原体。由于其苛刻的生长要求以及与其他猪支原体的血清学交叉反应，我们开发了一种用于检测它的特异性DNA探针。使用Hind III染色体片段在质粒pBR 322中构建了猪肺炎支原体的部分基因组文库。通过与絮状支原体和猪鼻支原体基因组DNA探针的差异杂交筛选重组质粒。选择了一个非杂交重组质粒，并对其1.65 kbp插入片段（命名为I141）针对来自多种支原体、其他细菌物种的基因组DNA以及无特定病原体（SPF）仔猪肺组织的DNA进行特异性测试。用32P标记的I141能够特异性检测低至400 pg的猪肺炎支原体基因组DNA。为了测试I141探针对猪肺炎支原体感染实验室诊断的适用性，我们使用了来自经猪肺炎支原体实验感染的仔猪的临床气管支气管标本。将每个标本的杂交结果与免疫荧光试验的结果进行比较。在所测试的临床标本中，两种测试结果的一致性为63%。
