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首页> 外文期刊>Journal of Veterinary Research >Real-time quantitative PCR for detection and identification of Actinobacillus pleuropneumoniae serotype 2
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Real-time quantitative PCR for detection and identification of Actinobacillus pleuropneumoniae serotype 2

机译：实时定量PCR检测和鉴定胸膜肺炎放线杆菌血清型2

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Introduction: Porcine pleuropneumonia inflicts important economic losses on most commercial herds. Detection of subclinical or chronic infection in animals still remains a challenge, as isolation and identification of A. pleuropneumoniae serotypes is difficult and quantification of the bacteria on agar plates is often almost impossible. The aim of the study was to develop and evaluate a serotype-specific quantitative TaqMan probe-based PCR for detection of serotype 2 in pig lungs, tonsils, and nasal swabs. Material and Methods: The primers were designed from the capsular polysaccharide biosynthesis genes of A. pleuropneumoniae serotype 2. PCR specificity and sensitivity were evaluated using reference strains and several other bacterial species commonly isolated from pigs. Results: The real-time qPCR for detection of A. pleuropneumoniae serotype 2 was highly specific and gave no false positives with other serotypes or different bacterial species of pig origin. The detection limit for pure culture was 1.2 x 10(4) CFU/mL, for lung tissue and nasal swabs it was 1.2 x 10(5) CFU/mL, and for tonsils - 1.2 x 10(5) CFU/mL. Conclusion: The method can be used to serotype A. pleuropneumoniae isolates obtained during cultivation and to detect and identify A. pleuropneumoniae serotype 2 directly in nasal swabs and tonsil scrapings obtained from live pigs or lung tissue and tonsils collected post-mortem.
机译：简介：猪胸膜肺炎对大多数商业畜群造成重大经济损失。动物的亚临床或慢性感染检测仍然是一个挑战，因为分离和鉴定胸膜肺炎链球菌血清型是困难的，并且在琼脂平板上定量细菌通常几乎是不可能的。该研究的目的是开发和评估基于血清型的定量TaqMan探针定量PCR，以检测猪肺，扁桃体和鼻拭子中的血清型2。材料和方法：引物是从胸膜肺炎链球菌血清型2的荚膜多糖生物合成基因设计的。PCR的特异性和敏感性使用参考菌株和其他几种通常从猪中分离出的细菌进行了评估。结果：实时定量PCR检测胸膜肺炎链球菌血清型2具有高度特异性，并且与其他血清型或猪不同细菌种均无假阳性反应。纯培养物的检出限为1.2 x 10（4）CFU / mL，肺组织和鼻拭子的检出限为1.2 x 10（5）CFU / mL，扁桃体为1.2 x 10（5）CFU / mL。结论：该方法可用于在培养过程中获得血清型肺炎链球菌分离株的血清型，并直接在从活猪或肺组织中采集的鼻拭子和扁桃体刮片和死后采集的扁桃体中直接检测和鉴定血清胸膜肺炎链球菌血清型2。

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