基于Nanopore测序技术分析广东地区1株PRRSV-2的基因组特征  

Analysis of the Genomic Characteristics of a PRRSV-2 Strain from Guangdong Region Using Nanopore Sequencing Technology

作　　者：周扬 卢受昇 宫焕宇 陈瑞旭 吴炜姿 曹伟胜[1] ZHOU Yang;LU Shousheng;GONG Huanyu;CHEN Ruixu;WU Weizi;CAO Weisheng(College of Veterinary Medicine,South China Agricultural University,Guangzhou 510642,China;Guangdong Center for Animal Disease Control and Prevention,Guangzhou 510230,China;School of Life Science and Engineering,Foshan University,Foshan 528225,China)

机构地区：[1]华南农业大学兽医学院,广东广州510642 [2]广东省动物疫病预防控制中心,广东广州510230 [3]佛山科学技术学院生命科学院,广东佛山528225

出　　处：《中国兽医杂志》2024年第9期14-22,共9页Chinese Journal of Veterinary Medicine

摘　　要：为促进Nanopore测序技术在美洲型猪繁殖与呼吸综合征病毒(PRRSV-2)全基因组测序中的应用并进一步了解当前广东地区PRRSV-2全基因组特征,本试验从广东地区某屠宰场采集1份PRRSV-2阳性样本,提取核酸进行靶向扩增,对扩增产物进行文库构建和测序,使用Samtools和Medaka等软件对测序数据进行生物信息学分析和拼接,得到样本的全基因组序列。选取5′端1~4 500核苷酸位点和开放阅读框5(ORF5)基因进行一代测序验证,评估本试验的准确性。进一步使用MEGA 11软件对所得的序列进行全基因组遗传进化分析、核苷酸相似性分析和NSP2蛋白氨基酸变异分析;使用RDP4软件和SimPlot软件进行重组分析。结果显示,本试验完成1份PRRSV-2阳性样本的全基因组测序,序列命名为GDYJ0718-7。2Samtools软件分析结果显示,本试验的平均测序深度为1 838×,覆盖PRRSV-2所有编码区。本试验Nanopore测序结果与一代测序结果一致性为100%。全基因组遗传进化分析结果显示,GDYJ0718-7属于QYYZ-like毒株,与参考株QYYZ(JQ308798)的核苷酸相似性仅为86.3%。NSP2蛋白氨基酸变异分析结果显示,GDYJ0718-7呈现1 aa+36 aa+29 aa的独特缺失模式。重组分析发现,GDYJ0718-7可能由QYYZ-like毒株和JXA1-like毒株重组形成。本试验通过Nanopore测序技术发现了广东地区QYYZ-like毒株变异的复杂性和新特点,为PRRSV-2的防控提供了技术和分子信息支持。To promote the application of Nanopore sequencing technology in whole-genome sequencing of the North American type porcine reproductive and respiratory syndrome virus(PRRSV-2) and to further understand the current genomic characteristics of PRRSV-2 in Guangdong,this study collected a PRRSV-2 positive sample from a slaughterhouse in Guangdong.The nucleic acid was extracted for targeted amplification,and the amplified products were used for library construction and sequencing.Bioinformatics analysis and assembly of the sequencing data were performed using Samtools and Medaka software,resulting in the complete genome sequence of the sample.The accuracy of the study was evaluated by Sanger sequencing validation of the 1-4 500 nucleotide positions of the 5′ end and the open reading frame 5(ORF5) gene.Further genetic evolutionary analysis,nucleotide similarity analysis,and NSP2 protein amino acid variation analysis of the obtained sequences were performed using MEGA 11 software;recombination analysis was conducted using RDP4 and SimPlot software.The results showed that the whole genome sequencing of one PRRSV-2 positive sample was completed,and the sequence was named GDYJ0718-7.Samtools software analysis indicated that the average sequencing depth of the study was 1 838×,covering all coding regions of PRRSV-2.The Nanopore sequencing results of this study were 100% consistent with the Sanger sequencing results.Whole-genome genetic evolutionary analysis showed that GDYJ0718-7 belonged to the QYYZ-like strain,with a nucleotide similarity of only 86.3% compared to the reference strain QYYZ(JQ308798).NSP2 protein amino acid variation analysis showed that GDYJ0718-7 exhibited a unique deletion pattern of 1 aa + 36 aa + 29 aa.Recombination analysis revealed that GDYJ0718-7 might have been formed by recombination between QYYZ-like and JXA1-like strains.This study used Nanopore sequencing technology to uncover the complexity and new characteristics of QYYZ-like strain variation in Guangdong,providing technical and molecular

关 键 词：Nanopore测序 猪繁殖与呼吸综合征病毒(PRRSV) 重组 

分 类 号：S852.659.6[农业科学—基础兽医学]