PDF(Pubmed)
Abstract：
PCR is the most effective method for detecting difficult-to-cultivate pathogens and pathogens that are part of mixed infections in animals, such as Ornithobacterium rhinotracheale, which causes bird ornithobacteriosis, or Avibacterium paragallinarum, which causes infectious coryza. In this work, we developed and validated two efficient and sensitive diagnostic assays for the rapid and accurate detection of A. paragallinarum and O. rhinotracheale DNA in bacterial isolates and clinical samples using real-time PCR with TaqMan-like probes. When designing the PCR assays, we performed in silico analysis, optimized DNA isolation methods and PCR conditions, and assessed the analytical and diagnostic performance of PCR. We designed primers and probes that have no mismatches with published whole-genome sequences of bacteria. The optimization of conditions showed that the PCR assays are sufficiently robust to changes in temperature and oligonucleotide concentration. The validation showed that the developed assays have high analytical and diagnostic sensitivity and specificity. These assays are expected to improve the differential diagnosis of respiratory diseases in chickens and turkeys.
摘要：
PCR是检测难以培养的病原体和作为动物混合感染一部分的病原体的最有效方法，比如鼻气管鸟状杆菌，会导致鸟类鸟粪杆菌病，或者副细菌，会导致传染性鼻炎.在这项工作中,我们开发并验证了两种高效和灵敏的诊断方法,用于使用TaqMan样探针进行实时PCR,快速和准确地检测细菌分离株和临床样品中的副鸡根瘤菌和鼻气管O.DNA.在设计PCR检测时，我们进行了硅分析，优化的DNA分离方法和PCR条件，并评估了PCR的分析和诊断性能。我们设计了与已发表的细菌全基因组序列没有错配的引物和探针。条件的优化显示PCR测定对于温度和寡核苷酸浓度的变化是足够稳健的。验证表明所开发的测定具有高的分析和诊断灵敏度和特异性。这些测定有望改善鸡和火鸡呼吸道疾病的鉴别诊断。
