利用宏基因组学鉴定藏猪腹泻粪便病毒种群

为进一步了解藏猪腹泻粪便中病毒种群，从青藏高原地区16个藏猪场采集腹泻粪便样本146份，将样本混合成一个pool样，提取RNA反转录成cDNA，建库后利用TruSeq Illumina测序，对测序的序列进行整理分析及相关病毒的分子特征进行研究。数据分析结果表明：藏猪腹泻粪便样本病毒种群包括11个病毒科的19种病毒，主要以线性和环状的小DNA病毒为主，如猪粪相关环状病毒7型、猪腺病毒和猪博卡病毒（PBoV）等；与腹泻相关的病原有猪流行性腹泻病毒（PEDV）、猪圆环病毒2型（PCV-2）和牛病毒性腹泻病毒1型（BVDV-1）3种；与腹泻相关的新发病原有porcine bufavirus、rabovirus和pasivirus 3种。利用SOAP软件组装出猪细小病毒6型（PPV-6）、PCV-2、PBoV-2完整或接近全长的基因组和戊型肝炎病毒（HEV）完整的ORF2基因序列，系统发育结果显示PPV-6和PCV-2与参考毒株有较近的遗传进化关系，PBoV-2与参考毒株有较远的遗传进化关系，单独聚为一簇，可能是一个全新的基因型。为进一步调查PCV-2在藏猪腹泻粪便中的检出率，对采集146份样本进行检测，检出率为10.96%（16/146，95% CI：6.4%~17.2%），且与四川地区PCV-2流行株有较近的亲缘关系。本研究发现藏猪腹泻粪便中病毒种类复杂多样，且可能存在与其他动物和人交叉感染情况，具有重要的公共卫生学意义，也为藏猪腹泻病防控提供一定的理论依据。

Identification of Viral Community in Diarrheal Feces of Tibetan Pig by Metagenomics

This study was conducted to further understand the viral community in diarrheal feces of Tibetan pig. The 146 diarrheal fecal samples were collected from 16 Tibetan pig farms in Qinghai-Tibet Plateau. The samples were mixed into a pool sample, then RNAs were purified and synthesized into cDNAs. TruSeq Illumina sequencing was used to analyze the sequences and molecular characteristics of the diarrhea-associated viruses. The results showed that the virus species in the fecal samples of diarrheal Tibetan pigs included 19 viruses from 11 families. They were mainly linear and cyclic small DNA viruses, such as porcine stool-associated circular virus type 7, porcine adenovirus and porcine bocavirus(PBoV). There are three diarrheal viral pathogens, such as porcine epidemic diarrhea virus (PEDV), porcine circovirus type 2 (PCV-2) and bovine viral diarrhea virus type 1 (BVDV-1), as well as three new pathogens, such as porcine bufavirus, rabovirus and pasivirus. The complete or nearly full-length genome of porcine parvovirus type 6(PPV-6), PCV-2, PBoV-2 and complete ORF2 gene sequence of hepatitis E virus (HEV) were assembled by SOAP software. The phylogenetic results showed that PPV-6 and PCV-2 had a relatively close genetic and evolutionary relationship with the reference strain, while PBoV-2 had a relatively distant genetic and evolutionary relationship with the reference strain, which were gathered into a single cluster, and may be a completely new genotype. In order to further investigate the detection rate of PCV-2 in diarrheal feces of Tibetan pigs, 146 samples were collected and the detection rate was 10.96% (16/146, 95% CI:6.4%-17.2%), and it was closely related to the epidemic strain of PCV-2 in Sichuan region. In this study, it was found that the viruses in diarrheal feces of Tibetan pigs are complex and varied, and may be cross-infected with other animals and humans, which has important public health significance, and also provides theoretical basis for prevention and control of diarrhea in Tibetan pigs.