Han Wen, Luo Yuzi, Zhao Bibo, Sun Yuan, Li Su, Qiu Huaji
State Key Laboratory of Veterinary Biotechnology, Harbin Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Harbin 150001, China.
Wei Sheng Wu Xue Bao. 2013 Feb 4;53(2):197-203.
Extreme varieties of viruses exist in the environment and animals, some of which are unknown. However, many unknown viruses are barely detected by means of conventional virus isolation and PCR assay.
To develop a technology platform for detecting unknown viruses.
We established the technology based on viral metagenomics in combination with novel molecular diagnostics. The technology is consisted of removal of host nucleic acid, random PCR amplification, large-scale sequencing, and bioinformatics.
The technology was applied to detect classical swine fever virus (CSFV)-infected cells and a tissue sample of a pig infected with porcine circovirus type 2 (PCV2). We amplified 13.7% sequences of CSFV genome and 47.2% those of PCV2 genome, respectively. Moreover, we amplified 16.4% sequences of the simian parainfluenza virus type 5 genome from an unknown virus cell culture using the developed method. In addition, using the developed method combined with the high-throughput sequencing, we detected 1.1% virus sequences, including CSFV, PCV2, torque teno sus virus (TTSuV), porcine bocavirus (PBoV) and human adenovirus type 6 (Ad6) from 7 clinical swine samples of unknown causative agents.
The developed metagenomics-based method showed good sensitivity for detection of both DNA and RNA viruses from diverse swine samples, and has potential for universal detection of known and unknown viruses. It might facilitate the diagnosis of emerging viral diseases.
环境和动物中存在着种类繁多的病毒，其中一些尚不为人所知。然而，许多未知病毒通过传统的病毒分离和聚合酶链反应（PCR）检测方法很难被检测到。
开发一种检测未知病毒的技术平台。
我们基于病毒宏基因组学并结合新型分子诊断技术建立了该技术。该技术包括去除宿主核酸、随机PCR扩增、大规模测序和生物信息学。
该技术应用于检测感染经典猪瘟病毒（CSFV）的细胞以及感染猪圆环病毒2型（PCV2）的猪的组织样本。我们分别扩增出了CSFV基因组13.7%的序列和PCV2基因组47.2%的序列。此外，使用所开发的方法，我们从一种未知病毒细胞培养物中扩增出了5型猿猴副流感病毒基因组16.4%的序列。另外，使用所开发的方法结合高通量测序，我们从7份病因不明的临床猪样本中检测到了1.1%的病毒序列，包括CSFV、PCV2、猪细小病毒（TTSuV）、猪博卡病毒（PBoV）和6型人腺病毒（Ad6）。
所开发的基于宏基因组学的方法对检测来自不同猪样本的DNA和RNA病毒均显示出良好的敏感性，并且具有检测已知和未知病毒的潜力。它可能有助于新发病毒性疾病的诊断。
