甘肃定西地区辣椒病毒病鉴定及BWYV遗传变异分析

doi:10.16420/j.issn.0513-353x.2024-0282

摘要/Abstract

摘要：

利用高通量测序技术和RT-PCR相结合的方法，对甘肃定西地区感染病毒病的辣椒叶片进行病原鉴定。结果显示染病叶片受黄瓜花叶病毒（cucumber mosaic virus，CMV）和甜菜西方黄化病毒（beet western yellows virus，BWYV）的复合侵染，首次在甘肃发现BWYV侵染辣椒。借助特异性引物克隆BWYV的外壳蛋白（coat protein，CP）和运动蛋白（movement protein，MP）基因深入分析BWYV的分子进化和遗传变异，通过序列比对发现其CP基因与其他物种BWYV分离物的核苷酸和氨基酸相似性分别为93.27% ~ 99.51%和90.10% ~ 99.01%，其MP基因的相似性分别为89.20% ~ 99.05%和86.86% ~ 98.86%。此外，系统发育树分析显示BWYV不同分离物可分为4个类群，无明显地理分布特征，寄主植物多样性分析显示具有显著的寄主关联性。

关键词: 辣椒, 甜菜西方黄化病毒, RT-PCR检测, 进化分析

Abstract:

Using a method that combines high-throughput sequencing technology with RT-PCR，pathogen identification was conducted on pepper leaves infected with viral diseases in the Dingxi area of Gansu Province. The results indicated that the infected leaves were co-infected with cucumber mosaic virus and sugar beet western yellow virus. Notably，this study marks the first identification of BWYV infecting peppers in Gansu. Specific primers were employed to clone the coat protein and movement protein genes of BWYV，facilitating an in-depth analysis of the molecular evolution and genetic variation of this virus. Sequence comparisons revealed that the nucleotide and amino acid similarities between the coat protein（CP）gene of BWYV and isolates from other species ranged from 93.27% to 99.51% and from 90.10% to 99.01%，respectively. The similarities of the movement protein（MP） genes were found to be between 89.20% and 99.05% for nucleotides and between 86.86% and 98.86% for amino acids. Furthermore，phylogenetic analysis of BWYV demonstrated that different isolates could be categorized into four distinct groups. These groups exhibited no discernible geographical distribution characteristics，while host plant diversity analysis revealed significant correlations with host species.

Key words: pepper, beet western yellows virus, RT-PCR detection, evolutionary analysis

杨文清, 王嘉荟, 徐秉良, 李惠霞, 梁巧兰, 杨顺义, 牛二波. 甘肃定西地区辣椒病毒病鉴定及BWYV遗传变异分析[J]. 园艺学报, 2025, 52(2): 491-502.

YANG Wenqing, WANG Jiahui, XU Bingliang, LI Huixia, LIANG Qiaolan, YANG Shunyi, NIU Erbo. Identification of Pepper Virus Diseases and Analysis of BWYV Genetic Variation in Dingxi Area of Gansu Province[J]. Acta Horticulturae Sinica, 2025, 52(2): 491-502.

图/表 13

参考文献 38

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引物名称 Primer	引物序列（5′-3′） Primer sequence	产物/bp Fragment	退火温度/℃ T_m	参考文献 Reference
CMV-CP	F：GTCGACCATGGACAAAT；R：CCGGGTCAGACTGGTAGCA	660	56	王德富等,2016
CMV-MP-	F：AGGCATGGCTTTCCAAGGTACCAG；R：AGTGCTAAAGACCGTTAACCA	850	55	杜江等,2023
BWYV-CP	F：CCTCAAAATCTCAGCCCACG；R：TGGCATCATCGGTACTCTGG	780	60	本试验This study
BWYV-MP	F：TGAATACGGTCGTGGGTAGA；R：GGCCATTATACGAGTCGCTG	725	56	本试验This study

引物名称 Primer	引物序列（5′-3′） Primer sequence	产物/bp Fragment	退火温度/℃ T_m	参考文献 Reference
CMV-CP	F：GTCGACCATGGACAAAT；R：CCGGGTCAGACTGGTAGCA	660	56	王德富等,2016
CMV-MP-	F：AGGCATGGCTTTCCAAGGTACCAG；R：AGTGCTAAAGACCGTTAACCA	850	55	杜江等,2023
BWYV-CP	F：CCTCAAAATCTCAGCCCACG；R：TGGCATCATCGGTACTCTGG	780	60	本试验This study
BWYV-MP	F：TGAATACGGTCGTGGGTAGA；R：GGCCATTATACGAGTCGCTG	725	56	本试验This study

基因登录号 Gene ID	株系 Strain	寄主植物 Host plant	地理来源 Geographical origin	核苷酸 Nucleotide	氨基酸 Aminoacid
MK307779.1	SDJN16	辣椒Capsicum annuum	中国山东Shandong，China	99.18	98.51
MK307780.1	SDWF16	辣椒Capsicum annuum	中国山东Shandong，China	99.01	99.01
LC428355.1	S14	菠菜Spinacia oleracea	日本Japan	99.51	99.01
MW349136.1	JJ18	辣椒Capsicum annuum	韩国Korea	99.18	98.51
OQ570952.1	YA35	萝卜Raphanus sativus	韩国Korea	99.18	98.02
LC592344.1	ADHS	北玄参Scrophularia buergeriana	韩国Korea	99.01	99.01
LC428357.1	R3a	萝卜Raphanus sativus	日本Japan	98.85	98.02
OM419176.1	—	香菜Coriandrum sativum	塞浦路斯Cyprus	93.92	91.09
ON924237.1	W11	甜菜Beta vulgaris	美国USA	93.76	91.58
KU521325.1	Rouen1	猪笼草Nepenthes mirabilis	法国French	93.60	90.10
ON924244.1	NYC	甜菜Beta vulgaris	美国USA	93.60	91.58
NC004756.1	USA	甜菜Beta vulgaris	美国USA	93.60	92.08
HM804471.1	BJ^A	甜菜Beta vulgaris	中国北京Beijing，China	97.87	98.51
HM804472.1	BJ^B	甜菜Beta vulgaris	中国北京Beijing，China	95.73	96.04
EU636991.1	IM	甜菜Beta vulgaris	中国内蒙古Inner Mongolia，China	99.18	90.10
EU636990.1	GS	甜菜Beta vulgaris	中国甘肃Gansu，China	93.76	93.56
AB903037.1	KY	菠菜Spinacia oleracea	日本Japan	93.27	94.55

基因登录号 Gene ID	株系 Strain	寄主植物 Host plant	地理来源 Geographical origin	核苷酸 Nucleotide	氨基酸 Aminoacid
MK307779.1	SDJN16	辣椒Capsicum annuum	中国山东Shandong，China	99.18	98.51
MK307780.1	SDWF16	辣椒Capsicum annuum	中国山东Shandong，China	99.01	99.01
LC428355.1	S14	菠菜Spinacia oleracea	日本Japan	99.51	99.01
MW349136.1	JJ18	辣椒Capsicum annuum	韩国Korea	99.18	98.51
OQ570952.1	YA35	萝卜Raphanus sativus	韩国Korea	99.18	98.02
LC592344.1	ADHS	北玄参Scrophularia buergeriana	韩国Korea	99.01	99.01
LC428357.1	R3a	萝卜Raphanus sativus	日本Japan	98.85	98.02
OM419176.1	—	香菜Coriandrum sativum	塞浦路斯Cyprus	93.92	91.09
ON924237.1	W11	甜菜Beta vulgaris	美国USA	93.76	91.58
KU521325.1	Rouen1	猪笼草Nepenthes mirabilis	法国French	93.60	90.10
ON924244.1	NYC	甜菜Beta vulgaris	美国USA	93.60	91.58
NC004756.1	USA	甜菜Beta vulgaris	美国USA	93.60	92.08
HM804471.1	BJ^A	甜菜Beta vulgaris	中国北京Beijing，China	97.87	98.51
HM804472.1	BJ^B	甜菜Beta vulgaris	中国北京Beijing，China	95.73	96.04
EU636991.1	IM	甜菜Beta vulgaris	中国内蒙古Inner Mongolia，China	99.18	90.10
EU636990.1	GS	甜菜Beta vulgaris	中国甘肃Gansu，China	93.76	93.56
AB903037.1	KY	菠菜Spinacia oleracea	日本Japan	93.27	94.55

登录号 Gene ID	株系 Strain	寄主植物 Host plant	地理来源 Geographical origin	核苷酸 Nucleotide	氨基酸 Aminoacid
MK307780.1	SDWF16	辣椒Capsicum annuum	中国山东Shandong，China	99.05	98.86
LC428357.1	R3a	萝卜Raphanus sativus	日本Japan	99.05	98.86
OQ625515.1	NJ22	萝卜Raphanus sativus	韩国Korea	99.05	98.86
LC592344.1	ADHS	北玄参Scrophularia buergeriana	韩国Korea	97.92	94.86
ON759519.1	P03	辣椒Capsicum annuum	韩国Korea	98.86	98.29
MK307779.1	SDJN16	辣椒Capsicum annuum	中国山东Shandong，China	98.67	98.29
OQ570952.1	YA35	萝卜Raphanus sativus	韩国Korea	98.48	98.29
KM076647.1	LS	益母草Leonurus sibiricus	韩国Korea	98.48	97.14
HM804471.1	BJ^A	甜菜Beta vulgaris	中国北京Beijing，China	96.21	93.14
HM804472.1	BJ^B	甜菜Beta vulgaris	中国北京Beijing，China	96.02	92.57
ON924175.1	JJ	小麦Triticum aestivum	中国江苏Jiangsu，China	91.29	89.14
EU636990.1	GS	甜菜Beta vulgaris	中国甘肃Gansu，China	89.96	87.43
HQ543079.1	6pCP	豌豆Pisum sativum	澳大利亚Australia	89.39	89.14
EU636991.1	IM	甜菜Beta vulgaris	中国内蒙古Inner Mongolia，China	89.58	86.86
HQ543082.1	9p	豌豆Pisum sativum	澳大利亚Australia	89.39	88.00
HQ543095.1	27p	豌豆Pisum sativum	澳大利亚Australia	89.20	86.86
HQ543084.1	14p	豌豆Pisum sativum	澳大利亚Australia	89.20	87.43