建立并应用 PCR 检测方法鉴定棉签中强毒和弱毒鸭瘟病毒 DNA
Establishment and application of a PCR assay for the identification of virulent and attenuated duck plague virus DNA in cotton swabs.
机构信息
Institute of Preventive Veterinary Medicine, Sichuan Agricultural University, Wenjiang, China; Avian Disease Research Center, College of Veterinary Medicine of Sichuan Agricultural University, Wenjiang, China; Key Laboratory of Animal Disease and Human Health of Sichuan Province, Sichuan Agricultural University, Wenjiang, China.
Avian Disease Research Center, College of Veterinary Medicine of Sichuan Agricultural University, Wenjiang, China; Key Laboratory of Animal Disease and Human Health of Sichuan Province, Sichuan Agricultural University, Wenjiang, China.
出版信息
Poult Sci. 2023 May;102(5):102555. doi: 10.1016/j.psj.2023.102555. Epub 2023 Feb 2.
Duck plague is an acute, febrile, and septic infectious disease caused by duck plague virus (DPV), which causes serious harm to the duck industry in China. Ducks latently infected with DPV display a clinically healthy state, which is one of the epidemiological characteristics of duck plague. In the present study, to rapidly distinguish vaccine-immunized ducks from wild virus-infected ducks during production, a PCR assay based on the newly identified LORF5 fragment was developed to effectively and accurately identify viral DNA in cotton swab samples and was used to assess artificial infection models and clinical samples. The results showed that the established PCR method had good specificity and that only the virulent and attenuated DNA of duck plague virus was specifically amplified, as the results for the detection of common duck pathogens (duck hepatitis B virus, duck Tembusu virus, duck hepatitis A virus type 1, novel duck reovirus, Riemerella anatipestifer, Pasteurella multocida, and Salmonella) were negative. The amplified fragments of virulent and attenuated strains were 2,454 bp and 525 bp, and their minimum detection amounts were 0.46 pg and 46 pg, respectively. The detection rate of the virulent and attenuated DPV strains in duck oral and cloacal swabs was lower than that of the gold standard PCR method (GB-PCR, which is unable to distinguish virulent and attenuated strains), and cloacal swabs from clinically healthy ducks were more suitable for detection than oral swabs. In conclusion, the PCR assay established in the present study can be used as a simple and effective method for the clinical screening of ducks that are latently infected with virulent strains of DPV and shedding virus, which can provide technical support for the elimination of duck plague from duck farms.
鸭瘟是由鸭瘟病毒（DPV）引起的一种急性、热性、败血性传染病，对中国养鸭业危害严重。DPV 潜伏感染鸭呈临床健康状态，这是鸭瘟的流行病学特征之一。本研究旨在生产过程中快速区分疫苗免疫鸭和野毒感染鸭，基于新鉴定的 LORF5 片段建立了 PCR 检测方法，能够有效、准确地检测棉拭子样本中的病毒 DNA，并用于人工感染模型和临床样本的评估。结果表明，所建立的 PCR 方法具有良好的特异性，仅特异性扩增鸭瘟病毒的强毒和弱毒 DNA，对常见鸭病病原体（鸭乙型肝炎病毒、鸭坦布苏病毒、鸭甲型肝炎病毒 1 型、新型鸭呼肠孤病毒、鸭疫里默氏杆菌、多杀性巴氏杆菌和沙门氏菌）的检测结果均为阴性。强毒和弱毒株的扩增片段分别为 2454 bp 和 525 bp，最小检测量分别为 0.46 pg 和 46 pg。鸭口腔和泄殖腔棉拭子中强毒和弱毒 DPV 株的检出率均低于金标准 PCR 方法（无法区分强毒和弱毒株），且临床健康鸭的泄殖腔拭子比口腔拭子更适合检测。综上所述，本研究建立的 PCR 检测方法可作为临床筛查鸭潜伏性强毒感染和排毒的简单、有效方法，为鸭场鸭瘟净化提供技术支持。
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