猪圆环病毒3型荧光定量PCR检测方法的建立及其在猪体中的组织分布研究

Establishment of a Fluorescent Quantitative PCR Method for Detection of Porcine Circovirus Type 3 and Study on Its Tissue Distribution in Pigs

关键词:  猪圆环病毒3型  荧光定量PCR  组织分布

Key words:  porcine circovirus type 3  fluorescent quantitative PCR  tissue distribution

摘要:

为了解猪圆环病毒3型(PCV3)在猪体中的组织分布情况，针对PCV3 Cap蛋白基因设计筛选出一对特异性引物和探针，通过对荧光PCR引物、探针浓度进行优化，建立了基于TaqMan探针实时荧光定量PCR技术的PCV3检测方法，并应用该方法对人工感染PCV3猪的多种组织器官进行病毒含量检测。结果显示，建立的PCV3实时荧光定量PCR检测方法特异性较好且灵敏度高，可检测低至1 copy/μL的PCV3病毒核酸量，而对其他几种常见猪病毒病原的检测结果均为阴性。PCV3主要存在于肺脏、淋巴结，在扁桃体和脾脏中有少量存在。试验表明，所建立的荧光定量PCR检测方法可用于PCV3的快速定量检测，对PCV3在猪体中的组织分布情况的研究为进一步揭示PCV3的组织嗜性和致病机理提供理论依据。

Abstract:

To explore the distribution of porcine circovirus type 3 (PCV3) in pigs, a pair of specific primers and probe were designed for PCV3 Cap gene. After optimizing the concentrations of PCR primers and probe, a TaqMan probe fluorescence quantitative real-time PCR technology was established, and the viral load of PCV3 in various tissues and organs of pigs artificially infected with PCV3 was studied by this method. The results showed that this method was specific and sensitive for the detection of PCV3, and could detect the amount of virus load as low as l copy/μL, while other common porcine virus pathogens were negative. The results of the detection of the main immune organs and some visceral tissues of artificially infected pigs showed that PCV3 mainly existed in the lungs and lymph nodes, and a small amount of PCV3 existed in tonsils and spleen. This study results indicated that the TaqMan real-time PCR technology could be used for rapid quantitative detection of PCV3, and the study on the tissue distribution of PCV3 in pigs could provide theoretical basis for further revealing the tissue tropism and pathogenic mechanism of PCV3.