Gram T, Ahrens P, Nielsen J P
Danish Veterinary Laboratory, Copenhagen, Denmark.
Vet Microbiol. 1996 Jul;51(1-2):95-104. doi: 10.1016/0378-1135(96)00013-2.
A PCR for the detection of Actinobacillus pleuropneumoniae was evaluated. All of 102 field, isolates of A, pleuropneumoniae reacted in the PCR by amplification of a 985 bp product. No PCR amplification product was observed when examining strains of A. ureae, A. capsulatus, A. hominis, A. equuli, A, rossii, A. suis, Escherichia coli, Bordetella bronchiseptica. Streptococcus suis, Pasteurella haemolytica, Pasteurella multocida, Haemophilus parasuis, Haemophilus taxon Minor group, Haemophilus taxon D/E and haemophilus taxon F. Amplification of a 985 bp product was, however, observed when testing strains of A. lignieresii. The lower detection limit of the PCR test was 10(3) A. pleuropneumoniae CFU/PCR test tube and was not affected by addition of 10(6) E. coli CFU/PCR test tube. Mixed bacterial cultures from tonsils of 101 pigs from 9 different herds were tested by culture and by PCR using four different bacteriological media. While 65% reacted positive in the PCR only 23% were positive by culture, thereby suggesting a superior sensitivity of the PCR test to that of culture. The use of selective media, large inoculum and incubation for 48 h gave the highest number of positive PCR reactions from mixed bacterial cultures. Tonsil cultures from 50 pigs from an A. pleuropneumoniae-negative herd did not react in the PCR. The results show that PCR on mixed bacterial cultures from tonsils may be a highly sensitive method for the detection of A. pleuropneumoniae in pig herds.
对用于检测胸膜肺炎放线杆菌的聚合酶链反应（PCR）进行了评估。102株胸膜肺炎放线杆菌的田间分离株在PCR中均通过扩增出一条985 bp的产物而发生反应。在检测脲放线杆菌、荚膜放线杆菌、人放线杆菌、马驹放线杆菌、罗西放线杆菌、猪放线杆菌、大肠杆菌、支气管败血波氏杆菌、猪链球菌、溶血巴斯德菌、多杀巴斯德菌、副猪嗜血杆菌、次要类嗜血杆菌、D/E类嗜血杆菌和F类嗜血杆菌的菌株时，未观察到PCR扩增产物。然而，在检测林氏放线杆菌的菌株时，观察到了985 bp产物的扩增。PCR检测的下限为10³CFU/PCR试管的胸膜肺炎放线杆菌，并且不受每PCR试管中加入10⁶CFU大肠杆菌的影响。使用四种不同的细菌学培养基，通过培养和PCR对来自9个不同猪群的101头猪扁桃体的混合细菌培养物进行了检测。虽然65%的样本在PCR中呈阳性反应，但通过培养只有23%呈阳性，这表明PCR检测的灵敏度优于培养法。使用选择性培养基、大接种量并培养48小时，可使混合细菌培养物中PCR阳性反应的数量最多。来自胸膜肺炎放线杆菌阴性猪群的50头猪的扁桃体培养物在PCR中无反应。结果表明，对猪扁桃体混合细菌培养物进行PCR检测可能是一种检测猪群中胸膜肺炎放线杆菌的高灵敏度方法。
