Gebhart C J, Ward G E, Murtaugh M P
Department of Veterinary Pathobiology, College of Veterinary Medicine, University of Minnesota, St. Paul 55108.
J Clin Microbiol. 1989 Dec;27(12):2717-23. doi: 10.1128/jcm.27.12.2717-2723.1989.
Conventional microbiological methods for isolating and identifying Campylobacter species are laborious, tedious, and subjective. Because of the increasing importance of Campylobacter species in human and animal diseases and the recent emergence of many new species and atypical strains, we are developing chromosomal DNA probes for rapid and simple identification of Campylobacter species, especially those of veterinary importance. We report the cloning and characterization of chromosomal DNA fragments from Campylobacter hyointestinalis, an organism isolated from pigs with proliferative enteritis. To obtain C. hyointestinalis-specific probes, chromosomal DNA fragments from C. hyointestinalis were cloned into plasmid vector pGEM-3Z. Recombinant plasmids were screened for C. hyointestinalis-specific inserts by DNA hybridization, using chromosomal DNA from either C. hyointestinalis or C. fetus which had been 32P labeled. Recombinants which hybridized to C. hyointestinalis, but not C. fetus, DNA were 32P labeled and screened further for sensitivity and specificity. Three probes were identified that were species specific and capable of detecting 10(4) C. hyointestinalis organisms by bacterial spot blotting in 48 h. We anticipate that these probes will be useful for routine species identification and for epidemiological studies.
用于分离和鉴定弯曲杆菌属菌种的传统微生物学方法既费力、繁琐又主观。由于弯曲杆菌属菌种在人类和动物疾病中的重要性日益增加，以及最近出现了许多新菌种和非典型菌株，我们正在开发染色体DNA探针，用于快速、简单地鉴定弯曲杆菌属菌种，特别是那些具有兽医重要性的菌种。我们报告了从猪增生性肠炎分离出的猪肠弯曲杆菌染色体DNA片段的克隆和特性分析。为了获得猪肠弯曲杆菌特异性探针，将猪肠弯曲杆菌的染色体DNA片段克隆到质粒载体pGEM-3Z中。使用经32P标记的猪肠弯曲杆菌或胎儿弯曲杆菌的染色体DNA，通过DNA杂交筛选重组质粒中猪肠弯曲杆菌特异性插入片段。与猪肠弯曲杆菌DNA杂交但不与胎儿弯曲杆菌DNA杂交的重组体用32P标记，并进一步筛选其敏感性和特异性。鉴定出三种探针，它们具有种特异性，能够在48小时内通过细菌斑点印迹法检测到10(4)个猪肠弯曲杆菌菌体。我们预计这些探针将有助于常规菌种鉴定和流行病学研究。
