巴基斯坦的一种新型禽源戊型肝炎病毒。
A novel avian isolate of hepatitis E virus from Pakistan.
机构信息
Department of Biochemistry and Biotechnology, University of Gujrat, Gujrat, 50700, Pakistan.
Division of Viral Hepatitis, Centers for Disease Control and Prevention (CDC), MS-A33, 1600 Clifton Rd NE, Atlanta, GA, 30329, USA.
出版信息
Virol J. 2019 Nov 21;16(1):142. doi: 10.1186/s12985-019-1247-0.
BACKGROUND
Avian hepatitis E virus (aHEV) has been associated with hepatitis-splenomegaly syndrome (HSS) in chickens along with asymptomatic subclinical infection in many cases. So far, four genotypes have been described, which cause infection in chickens, specifically in broiler breeders and layer chickens. In the present study, we isolated and identified two novel aHEV strains from the bile of layer chickens in Pakistan evincing clinical symptoms related to HSS.
METHODOLOGY
Histology of liver and spleen tissues was carried out to observe histopathological changes in these tissues. Bile fluid and fecal suspensions were used for viral RNA isolation through MegNA pure and Trizol method which was further used for viral genome detection and characterization by cDNA synthesis and amplification of partial open reading frame (ORF) 1, ORF2 and complete ORF3. The bioinformatics tools; Molecular Evolutionary Genetics Analysis version 6.0 (MEGA 6), Mfold and ProtScale were used for phylogenic analysis, RNA secondary structure prediction and protein hydropathy analysis, respectively.
RESULTS
Sequencing and phylogenetic analysis on the basis of partial methyltranferase (MeT), helicase (Hel) domain, ORF2 and complete ORF3 sequence suggests these Pakistani aHEV (Pak aHEV) isolates may belong to a Pakistani specific clade. The overall sequence similarity between the Pak aHEV sequences was 98-100%. The ORF1/ORF3 intergenic region contains a conserved cis-reactive element (CRE) and stem-loop structure (SLS). Analysis of the amino acid sequence of ORF3 indicated two hydrophobic domains (HD) and single conserved proline-rich domain (PRD) PREPSAPP (PXXPXXPP) with a single PSAP motif found in C-terminal. Amino acid changes S15 T, A31T, Q35H and G46D unique to the Pak aHEV sequences were found in the N-terminal region of ORF3.
CONCLUSIONS
Our data suggests that Pak aHEV isolates may represent a novel Pakistani clade and high sequence homology to each other support the supposition they may belong to a monophyletic clade circulating in the region around Pakistan. The data presented in this study provide further information for aHEV genetic diversity, genotype mapping, global distribution and epidemiology.
背景
禽戊型肝炎病毒（aHEV）与鸡的肝炎-脾肿大综合征（HSS）有关，在许多情况下还伴有无症状的亚临床感染。迄今为止，已描述了四种基因型，它们会引起鸡感染，特别是在肉鸡饲养者和蛋鸡中。在本研究中，我们从巴基斯坦的蛋鸡胆汁中分离并鉴定了两株新型 aHEV 株，这些鸡表现出与 HSS 相关的临床症状。
方法
对肝和脾组织进行组织学检查，以观察这些组织的组织病理学变化。使用 MegNA pure 和 Trizol 方法从胆汁和粪便悬液中提取病毒 RNA，进一步通过 cDNA 合成和部分开放阅读框（ORF）1、ORF2 和完整 ORF3 的扩增来检测和表征病毒基因组。使用分子进化遗传学分析版本 6.0（MEGA 6）、Mfold 和 ProtScale 等生物信息学工具进行系统发育分析、RNA 二级结构预测和蛋白质疏水性分析。
结果
基于甲基转移酶（MeT）、解旋酶（Hel）结构域、ORF2 和完整 ORF3 序列的测序和系统发育分析表明，这些巴基斯坦 aHEV（Pak aHEV）分离株可能属于巴基斯坦特有的一个分支。Pak aHEV 序列之间的整体序列相似性为 98-100%。ORF1/ORF3 基因间区含有一个保守的顺式反应元件（CRE）和茎环结构（SLS）。ORF3 氨基酸序列分析表明，存在两个疏水区（HD）和单个保守的脯氨酸丰富区（PRD）PREPSAPP（PXXPXXPP），在 C 末端有一个 PSAP 基序。在 ORF3 的 N 末端发现了 aHEV 序列特有的氨基酸变化 S15T、A31T、Q35H 和 G46D。
结论
我们的数据表明，Pak aHEV 分离株可能代表一个新的巴基斯坦分支，彼此之间的高序列同源性支持它们可能属于巴基斯坦周边地区流行的单系分支的假设。本研究提供的资料进一步说明了 aHEV 的遗传多样性、基因型图谱、全球分布和流行病学情况。
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