Miller J M, Jenny A L, Ellingson J L
Respiratory and Neurologic Disease Research Unit, National Animal Disease Center, USDA, Agricultural Research Service, PO Box 70, Ames, IA 50010, USA.
J Vet Diagn Invest. 1999 Sep;11(5):436-40. doi: 10.1177/104063879901100508.
A PCR procedure previously developed for identification of Mycobacterium bovis in formalin-fixed tissues was used to identify mycobacteria of the M. avium complex. Tissues were examined from 100 culture-positive cases of M. avium complex infection, including 86 in which the subspecies was not identified and 14 that had been identified as M. avium subsp. paratuberculosis. Each sample was tested with 5 primer sets, 16S ribosomal RNA (rRNA), IS900, IS901, IS1245, and a heat shock protein (hspX), that detect 1 or both M. avium subspecies. The success rate of PCR detection varied with the primers used and the animal species tested. Among the 86 cases with no M. avium subspecies designation, primers for the 16S rRNA gene were clearly the most efficient because they produced amplicons from all samples that reacted with any other primer set. The overall detection rate in this group of samples was 71%: highest in avian tissues (89%) followed by swine (72%) and ruminants (57%) None of the avian or swine tissues reacted with primers for IS900 or hspX, which identify M. a. paratuberculosis. In contrast, 7 of the 12 ruminant samples that were 16S rRNA positive reacted with 1 or both of these primers. All of the 14 cases shown by culture to be M. a. paratuberculosis infections were positive with IS900 primers, whereas only 11 were positive for 16S rRNA. These results indicate that 16S rRNA primers are the most useful for PCR identification of M. avium in formalin-fixed tissues of nonruminant species. However, IS900 primers should also be used when ruminant tissues are examined because these primers provide the greatest sensitivity for detection of M. a. paratuberculosis infections.
一种先前开发用于在福尔马林固定组织中鉴定牛分枝杆菌的聚合酶链反应（PCR）程序，被用于鉴定鸟分枝杆菌复合群的分枝杆菌。对100例鸟分枝杆菌复合群感染的培养阳性病例的组织进行了检查，其中86例未鉴定亚种，14例已鉴定为副结核分枝杆菌鸟亚种。每个样本用5组引物进行检测，即16S核糖体RNA（rRNA）、IS900、IS901、IS1245和一种热休克蛋白（hspX），这些引物可检测一种或两种鸟分枝杆菌亚种。PCR检测的成功率因所用引物和检测的动物种类而异。在86例未指定鸟分枝杆菌亚种的病例中，16S rRNA基因的引物显然是最有效的，因为它们能从所有与任何其他引物组反应的样本中产生扩增子。这组样本的总体检测率为71%：禽类组织中最高（89%），其次是猪（72%）和反刍动物（57%）。没有禽类或猪组织与鉴定副结核分枝杆菌鸟亚种的IS900或hspX引物发生反应。相比之下，16S rRNA呈阳性的12例反刍动物样本中有7例与这些引物中的一种或两种发生反应。培养显示为副结核分枝杆菌鸟亚种感染的14例病例中，所有病例的IS900引物均呈阳性，而16S rRNA仅11例呈阳性。这些结果表明，16S rRNA引物对于在非反刍动物福尔马林固定组织中PCR鉴定鸟分枝杆菌最为有用。然而，在检查反刍动物组织时也应使用IS900引物，因为这些引物对检测副结核分枝杆菌鸟亚种感染具有最高的灵敏度。
