利用逆转录聚合酶链反应(RT-PCR)和下一代测序技术对伊朗受苹果花叶病影响树木进行病毒组综合分析

Comprehensive Virome Profiling of Apple Mosaic Disease-Affected Trees in Iran Using RT-PCR and Next-Generation Sequencing.

作者信息

Hamedi Anahita, Rakhshandehroo Farshad, Safarnejad Mohammad Reza, Jouzani Gholamreza Salehi, Ben Slimen Amani, Elbeaino Toufic

机构信息

Department of Plant Protection, SR. C., Islamic Azad University, Tehran 14515-775, Iran.

Department of Plant Viruses, Iranian Research Institute of Plant Protection, Agricultural Research, Education and Extension Organization (AREEO), Tehran 19395-1454, Iran.

出版信息

Viruses. 2025 Jul 13;17(7):979. doi: 10.3390/v17070979.

Abstract

Apples (), one of Iran's oldest cultivated fruit crops, hold considerable economic importance. In this study, 170 apple leaf samples representing various commercial cultivars were collected across the country. RT-PCR screening targeted five common apple-infecting viruses and two viroids: apple chlorotic leaf spot virus (ACLSV), apple stem pitting virus (ASPV), apple stem grooving virus (ASGV), apple green crinkle-associated virus (AGCaV), apple mosaic virus (ApMV), apple scar skin viroid (ASSVd), and hop stunt viroid (HSVd). To identify additional or novel agents, 40 RT-PCR-negative samples were pooled into two composite groups and analyzed using next-generation sequencing (NGS). NGS was also performed on individual samples with mixed infections to retrieve full genomes. RT-PCR confirmed the presence of ACLSV, ASPV, ASGV, AGCaV, ApMV, and HSVd. NGS further revealed three additional pathogens: citrus concave gum-associated virus (CCGaV), apple hammerhead viroid (AHVd), and apricot vein clearing-associated virus (AVCaV), which were subsequently detected across the collection by RT-PCR. AGCaV was most prevalent (47.6%), followed by ACLSV (45.8%), HSVd (27.6%), AVCaV (20.5%), ASGV (17%), AHVd (15.2%), ASPV (14.1%), CCGaV (4.7%), and ApMV (3.5%). Mixed infections occurred in 67% of samples. Phylogenetic analysis based on CP genes (ACLSV, ASGV, AGCaV) and full genomes (AVCaV, AHVd) clustered Iranian isolates together, suggesting a common origin. This is the first report in Iran of AGCaV, CCGaV, ApMV, and AVCaV in apple, and notably, the first global report of AVCaV in a non-Prunus host. The findings provide the first comprehensive assessment of the sanitary status of apple trees in Iran.

摘要

苹果是伊朗最古老的栽培水果作物之一,具有相当重要的经济意义。在本研究中,从全国各地收集了代表不同商业品种的170份苹果叶片样本。逆转录聚合酶链反应(RT-PCR)检测针对五种常见的感染苹果的病毒和两种类病毒:苹果褪绿叶斑病毒(ACLSV)、苹果茎痘病毒(ASPV)、苹果茎沟病毒(ASGV)、苹果绿皱相关病毒(AGCaV)、苹果花叶病毒(ApMV)、苹果疤痕果皮类病毒(ASSVd)和啤酒花矮化类病毒(HSVd)。为了鉴定其他或新的病原体,将40份RT-PCR阴性样本合并为两个混合组,并使用下一代测序(NGS)进行分析。对有混合感染的单个样本也进行了NGS,以获取完整基因组。RT-PCR证实了ACLSV、ASPV、ASGV、AGCaV、ApMV和HSVd的存在。NGS进一步揭示了另外三种病原体:柑橘凹皮病相关病毒(CCGaV)、苹果锤头状类病毒(AHVd)和杏叶脉明相关病毒(AVCaV),随后通过RT-PCR在整个样本收集中检测到这些病毒。AGCaV最为普遍(47.6%),其次是ACLSV(45.8%)、HSVd(27.6%)、AVCaV(20.5%)、ASGV(17%)、AHVd(15.2%)、ASPV(14.1%)、CCGaV(4.7%)和ApMV(3.5%)。67%的样本存在混合感染。基于外壳蛋白(CP)基因(ACLSV、ASGV、AGCaV)和完整基因组(AVCaV、AHVd)的系统发育分析将伊朗分离株聚类在一起,表明它们有共同的起源。这是伊朗首次报道苹果中存在AGCaV、CCGaV、ApMV和AVCaV,值得注意的是,这是AVCaV在非李属宿主中的首次全球报道。这些发现首次全面评估了伊朗苹果树的卫生状况。

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