基于小RNA深度测序的浙江省丝瓜病毒种类鉴定

doi:10.3969/j.issn.1004-1524.20250501

浙江农业学报 ›› 2026, Vol. 38 ›› Issue (8): 1620-1627.DOI: 10.3969/j.issn.1004-1524.20250501

基于小RNA深度测序的浙江省丝瓜病毒种类鉴定

石玉洁¹^,²(), 柴亚倩¹^,², 高璐瑶¹, 董文其¹, 张鹏¹, 周胜军¹, 王欣¹, 朱育强¹, 孙玉燕¹^,^*()

¹ 浙江省农业科学院蔬菜研究所, 浙江杭州 310021
² 石河子大学农学院, 新疆石河子 832003

收稿日期:2025-05-23 出版日期:2026-08-25 发布日期:2026-09-14
作者简介:石玉洁,研究方向为蔬菜育种。E-mail:shiyujie20001124@163.com
通讯作者: *孙玉燕,E-mail:sunyy@zaas.ac.cn
基金资助:
浙江省重点研发计划(2024C02003-5);浙江省农业(蔬菜)新品种选育重大科技专项子课题(2021C02065-2-4);浙江省瓜菜种质资源表型精准鉴定(2022JZJD003)

Identification of virus species infecting sponge gourd in Zhejiang Province of China based on small RNA deep sequencing

SHI Yujie¹^,²(), CHAI Yaqian¹^,², GAO Luyao¹, DONG Wenqi¹, ZHANG Peng¹, ZHOU Shengjun¹, WANG Xin¹, ZHU Yuqiang¹, SUN Yuyan¹^,^*()

¹ Institute of Vegetables, Zhejiang Academy of Agricultural Sciences, Hangzhou 310021, China
² College of Agriculture, Shihezi University, Shihezi 832003, Xinjiang, China

Received:2025-05-23 Published:2026-08-25 Online:2026-09-14

摘要/Abstract

摘要：

为明确侵染浙江省丝瓜的病毒种类及其分布,本研究采用小RNA深度测序技术对采自浙江嘉兴和绍兴的2份具有典型病毒病症状的丝瓜病叶样本进行鉴定。结果显示,嘉兴样本中鉴定出新德里番茄曲叶病毒(Tomato leaf curl New Delhi virus, ToLCNDV),绍兴样本中鉴定出ToLCNDV、南瓜黄色矮化失调病毒(Cucurbit yellow stunting disorder virus, CYSDV)和瓜类褪绿黄化病毒(Cucurbit chlorotic yellows virus, CCYV)。两地均检测到ToLCNDV,且其重叠群(contig)数量与覆盖度较高。对鉴定到的ToLCNDV DNA-A序列进行系统进化分析,结果表明,当前致病的ToLCNDV属于亚洲株系,其遗传进化关系与地理区域密切相关,且跨宿主传播在各区域普遍发生。进一步利用PCR技术对来自浙江、广西和江西的37份样本进行检测,ToLCNDV检出率分别为87.5%、60%和100%。本研究结果表明,ToLCNDV已成为浙江省和其他省份丝瓜主产区的主要病毒类型,需警惕其快速传播和大规模发生对丝瓜产业造成的潜在严重威胁。

关键词: 丝瓜, 小RNA深度测序, 新德里番茄曲叶病毒, 病毒

Abstract:

To identify the virus species infecting sponge gourd in Zhejiang Province, small RNA deep sequencing technology was performed on two diseased sponge gourd leaves exhibiting typical viral symptoms collected from Jia-xing and Shaoxing, Zhejiang. The results showed that Tomato leaf curl New Delhi virus (ToLCNDV) was detected in the Jiaxing sample, while ToLCNDV, Cucurbit yellow stunting disorder virus (CYSDV) and Cucurbit chlorotic yellows virus(CCYV) were identified in Shaoxing sample. ToLCNDV was detected in both regions with high contig numbers and coverage. Phylogenetic analysis of the identified ToLCNDV DNA-A sequence indicated that the current pathogenic ToLCNDV in this study belongs to the Asian strain, with genetic evolution closely related to geographical regions, and cross-host transmission is widespread across regions. PCR detection of 37 samples from Zhejiang, Guangxi and Jiangxi revealed ToLCNDV detection rates of 87.5%, 60% and 100%, respectively. These results demonstrated that ToLCNDV had become a predominant virus in major sponge gourd production areas of Zhejiang and other provinces, warranting vigilance against the potential severe threat posed by its rapid spread and large-scale outbreaks to the luffa industry.

Key words: sponge gourd, small RNA deep sequencing, Tomato leaf curl New Delhi virus (ToLCNDV), virus

中图分类号:

S436.429

石玉洁, 柴亚倩, 高璐瑶, 董文其, 张鹏, 周胜军, 王欣, 朱育强, 孙玉燕. 基于小RNA深度测序的浙江省丝瓜病毒种类鉴定[J]. 浙江农业学报, 2026, 38(8): 1620-1627.

SHI Yujie, CHAI Yaqian, GAO Luyao, DONG Wenqi, ZHANG Peng, ZHOU Shengjun, WANG Xin, ZHU Yuqiang, SUN Yuyan. Identification of virus species infecting sponge gourd in Zhejiang Province of China based on small RNA deep sequencing[J]. Acta Agriculturae Zhejiangensis, 2026, 38(8): 1620-1627.

图/表 8

表1 本研究中使用的引物

Table 1 Primers used in this study

用途 Usage	引物名称 Primer name	引物序列(5'→3') Primer sequence(5'→3')
DNA-A检测	ToLCNDV-A-61F	CCTATTTAATGCGTGGGGGT
Detection of DNA-A	ToLCNDV-A-1622R	AGCACTGAAGCAGTGGGCAA
验证测序结果准确性	ToLCNDV-A-F	TGCCTGCAGGTCGACTCTAGAACGTCTCCGTCTT
Verification of the accuracy of	ToLCNDV-A-R	TAAGAATTCGAGCTCGGTACCATGGGGTGTTTTCCAGATC
sequencing results	ToLCNDV-B-F	TGCCTGCAGGTCGACTCTAGAACTCAATCGGCGT
	ToLCNDV-B-R	TAAGAATTCGAGCTCGGTACCGAGAGAAACTGCTGCTTCT

表2 丝瓜病样测序基础数据统计分析

Table 2 Statistical analysis of sequencing data for diseased sponge gourd samples

样本 Sample	Read总数 Total reads	有效序列(占比/%) Clean read(proportion/%)	碱基总数 Base sum	GC含量/% GC content/%	N含量/% N content/%	Q30/%
嘉兴 Jiaxing	33 173 336	32 965 570(99.4)	735 041 593	45.86	0	97.39
绍兴 Shaoxing	34 473 140	34 269 880(99.4)	761 658 670	44.03	0.01	97.39

图1 unique sRNA碱基长度分布

Fig.1 Length distribution of unique small RNAs

表3 sRNA测序检测到的病毒种类

Table 3 Virus species detected by small RNA sequencing

样品来源 Sample source	比对序列 Reference sequence	测序总深度 (覆盖度/%) Total sequencing depth (coverage/%)	一致性/% Identity/%	重叠群数量 Contigs number	Contig长度/nt Contig length/nt	病毒 Virus	属 Genus	病毒类型 Virus type
嘉兴	HM007120	2 740 (100)	94.87	27	46~2 740	ToLCNDV	Begomovirus	DNA virus
Jiaxing	FN435312	2 696 (100)	96.08	28	41~1 424	ToLCNDV	Begomovirus	DNA virus
	MH465600	1 770 (65.4)	93.63	17	44~1 424	ToLCNDV	Begomovirus	DNA virus
绍兴	AJ620187	2 740 (100)	96.18	30	45~5 409	ToLCNDV	Begomovirus	DNA virus
Shaoxing	MH157025	2 738 (100)	96.79	31	45~5 409	ToLCNDV	Begomovirus	DNA virus
	FN435312	2 696 (100)	95.62	28	41~2 698	ToLCNDV	Begomovirus	DNA virus
	HG316125	2 577 (94.2)	97.01	30	46~5 409	ToLCNDV	Begomovirus	DNA virus
	MT813029	8 604 (94.4)	99.28	37	42~818	CYSDV	Crinivirus	RNA virus
	MT819949	7 489 (94.6)	99.27	22	45~1 266	CYSDV	Crinivirus	RNA virus
	AB523789	2 570 (32.0)	98.69	42	41~100	CCYV	Crinivirus	RNA virus
	MW629379	1 515 (17.6)	98.88	28	41~99	CCYV	Crinivirus	RNA virus

图2 基于ToLCNDV DNA-A序列的系统进化树

Fig.2 Phylogenetic tree based on DNA-A sequences of ToLCNDV

图3 测序样品的PCR验证 M,DNA marker;1,嘉兴样本的DNA-A验证;2,绍兴样本的DNA-A验证;3,嘉兴样本的DNA-B验证;4,绍兴样本的DNA-B验证。

Fig.3 PCR validation of sequencing samples M, DNA marker; 1, Validation of DNA-A from Jiaxing sample; 2, Validation of DNA-A from Shaoxing sample; 3, Validation of DNA-B from Jiaxing sample; 4, Validation of DNA-B from Shaoxing sample.

图4 浙江省丝瓜样本ToLCNDV的PCR检测结果 M,DNA marker;1~7,嘉兴采集样本;8~13,绍兴采集样本;14和15,嘉善采集样本;16~24,衢州采集样本。

Fig.4 PCR detection results of ToLCNDV in sponge gourd samples from Zhejiang Province M, DNA marker; Samples 1-7 were collected from Jiaxing; Samples 8-13 were collected from Shaoxing; Samples 14 and 15 were collected from Jiashan; Samples 16-24 were collected from Quzhou.

图5 广西和江西丝瓜样本ToLCNDV的PCR检测结果 M,DNA marker;1和2为广西百色市采集的样品,2为健康叶片样品;3~6为广西玉林市采集的样品;7~13为江西赣州采集的样品。

Fig.5 PCR detection results of ToLCNDV in sponge gourd samples from Guangxi Zhuang Autonomous Region and Jiangxi Province M, DNA marker; Samples 1 and 2 were collected from Baise City, Guangxi,with Sample 2 being a healthy leaf; Samples 3-6 were collected from Yulin City, Guangxi; Samples 7-13 were collected from Ganzhou City, Jiangxi.

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基于小RNA深度测序的浙江省丝瓜病毒种类鉴定

Identification of virus species infecting sponge gourd in Zhejiang Province of China based on small RNA deep sequencing

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