Studies on the genome heterogeneity and its host passage effect of apple stem grooving capillovirus

利用逆转录-聚合酶链式反应扩增苹果、日本梨和欧洲梨的苹果茎沟病毒(ASGV)和柑橘碎叶病毒(CTLV)的基因组3‘端序列，并对扩增产物进行序列分析。结果表明，苹果、日本梨和欧洲梨的ASGV分离物都含有至少2~4个核苷酸序列差异较大的‘序列变异’。对已报道的ASGV和CTLV分离株或变异株的ORF1和ORF2蛋白氨基酸序列的比较表明，ASGV(P-209)ORF1蛋白1868~2105位氨基酸序列高度保守，15个分离物或变异株ORF2蛋白的氨基酸序列相似性在92.8~100%之间，21个分离物或变异株的CP相似性在93.2~100%之间。相反，在15个分离物和序列变异体中，编码ORF2蛋白的A284AA区(在本文中称为V区)相当于ORF1蛋白在另一帧中的1584-1868位氨基酸序列的相似性高度不同(53.5-99.3%)。15例中V区氨基酸保守率仅为20.4%。根据CP、ORF2或V区蛋白的AA序列构建的系统发育树表明，无论来源寄主植物是苹果、日本梨、欧洲梨、柑橘和百合，分离物或序列变异体都被分成几个簇。在由V区蛋白构建的树中，两个CTLV分离物被分成两个不同的簇，每个簇包含来自苹果和日本梨的分离物或序列变体。这些结果表明，柑橘CTLV不能与玫瑰科植物的ASGV在分子水平上区分开来。

The 3'-terminal regions of the genomes of apple stem grooving virus (ASGV) isolated from apple, Japanese pear and European pear plants and citrus tatter leaf virus (CTLV) from citrus plants were amplified by the reverse transcription-polymerase chain reaction and then the DNA products were sequenced. The results indicated that all ASGV isolates from apple, Japanese pear and European pear comprise at least two to four 'sequence variants' which significantly differ in nucleotide sequence from each other. Comparison of the amino acid (aa) sequences of the ORF1 and ORF2 proteins among isolates or sequence variants including isolates of ASGV and CTLV reported previously showed that the aa sequences of the ORF2 protein (320 aa) and the putative coat protein (CP) (237aa) equivalent to aa positions 1868 to 2105 of the ORF1 protein of ASGV (P-209) were highly conserved among isolates or sequence variants.Similarities ranged from 92.8 to 100% for the ORF2 protein among 15 isolate or sequence variants and 93.2 to 100% for the CP among 21 isolates or sequence variants. On the contrary, the aa sequence similarities of a284 aa region (named 'V-region' in this paper) equivalent to aa positions 1584 to 1868 of the ORF1 protein which encodes ORF2 protein in another frame were highly variable (53.5 to 99.3%) among 15 isolates and sequence variants. The percentage of conserved V-region aa in all 15 was only 20.4%. Phylogenetic trees constructed from the aa sequences of CP,ORF2 or V-region proteins showed that isolates or sequence variants were separated into several clusters, regardless of the source host plant species (apple, Japanese pear, European pear, citrus and lily). In a tree constructed by V-region proteins, two CTLV isolates were grouped into two different clusters each of which contained isolates or sequence variants from apple and Japanese pear. These results demonstrate that CTLV from citrus cannot be distinguished from ASGV from Rosaceous plants at the molecular level.