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Journal of Integrative Agriculture  2021, Vol. 20 Issue (7): 1871-1879    DOI: 10.1016/S2095-3119(20)63275-0
Special Issue: 植物病理合辑Plant Protection—Plant Pathology 植物病毒合辑Plant Virus
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Biological and molecular characterization of tomato brown rugose fruit virus and development of quadruplex RT-PCR detection
YAN Zhi-yong*, ZHAO Mei-sheng*, MA Hua-yu*, LIU Ling-zhi, YANG Guang-ling, GENG Chao, TIAN Yan-ping, LI Xiang-dong
Shandong Provincial Key Laboratory of Agricultural Microbiology, College of Plant Protection, Shandong Agricultural University, Tai’an 271018, P.R.China
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番茄褐色皱果病毒(tomato brown rugose fruit virus,ToBRFV)是2015年首次报道的一种烟草花叶病毒属病毒,是番茄安全生产的严重威胁。该病毒已经传播到美洲、亚洲和欧洲的十个国家。2019年,ToBRFV在中国山东发生。本论文旨在明确ToBRFV山东分离物(ToBRFV-SD)的症状、寄主范围和分子特性,并建立一种有效的检测方法。田间调查ToBRFV-SD在不同品种的症状表现。将ToBRFV-SD接种辣椒、本氏烟、马铃薯、茄子、中烟102和50个番茄品种,鉴定其寄主范围。分段克隆ToBRFV-SD基因组片段,并测定其序列;利用BioEdit version 7.2.6比对ToBRFV所有分离物的全基因组序列,分析序列一致率;利用MEGA version 10.1.5构建系统发育树。根据ToBRFV、烟草花叶病毒(tobacco mosaic virus,TMV)、番茄花叶病毒(tomato mosaic virus,ToMV)和番茄斑萎病毒(tomato spotted wilt virus,TSWV)等四种番茄重要病毒基因组的保守区域设计特异性引物,建立四重RT-PCR检测体系。ToBRFV-SD在番茄叶片引起不同程度的花叶和疱斑,在花萼和花梗上引起坏死,在番茄果实上引起畸形、黄斑和褐色皱缩坏死斑。ToBRFV-SD可侵染番茄、辣椒和本氏烟,隐症侵染马铃薯、茄子和烟草品种中烟102。测试的50个番茄品种均不抗ToBRFV-SD。ToBRFV-SD和以色列分离物ToBRFV-IL基因组的核苷酸和氨基酸一致率最高。在基于全基因组序列的系统进化树中,所有ToBRFV分离物聚集到一个分枝,与烟草花叶病毒分枝距离较近。随后,我们建立了四重RT-PCR检测体系,能够通过一个RT-PCR反应,同时检测并区分ToBRFV、TMV、ToMV和TSWV。本研究明确了ToBRFV-SD的症状、寄主范围和分子特性,建立了能区分ToBRFV、TMV、ToMV和TSWV四重RT-PCR检测体系,对指导ToBRFV的早期检测和防控有积极作用。

Tomato brown rugose fruit virus (ToBRFV) is a novel tobamovirus firstly reported in 2015 and poses a severe threat to the tomato industry.  So far, it has spread to 10 countries in America, Asia, and Europe.  In 2019, ToBRFV was identified in Shandong Province (ToBRFV-SD), China.  In this study, it was shown that ToBRFV-SD induced mild to severe mosaic and blistering on leaves, necrosis on sepals and pedicles, and deformation, yellow spots, and brown rugose necrotic lesions on fruits.  ToBRFV-SD induced distinct symptoms on plants of tomato, Capsicum annumm, and Nicotiana benthamiana, and caused latent infection on plants of Solanum tuberosum, Solanum melongena, and N. tabacum cv. Zhongyan 102.  All the 50 tomato cultivars tested were highly sensitive to ToBRFV-SD.  The complete genomic sequence of ToBRFV-SD shared the highest nucleotide and amino acid identities with isolate IL from Israel.  In the phylogenetic tree constructed with the complete genomic sequence, all the ToBRFV isolates were clustered together and formed a sister branch with tobacco mosaic virus (TMV).  Furthermore, a quadruplex RT-PCR system was developed that could differentiate ToBRFV from other economically important viruses affecting tomatoes, such as TMV, tomato mosaic virus, and tomato spotted wilt virus.  The findings of this study enhance our understanding of the biological and molecular characteristics of ToBRFV and provide an efficient and effective detection method for multiple infections, which is helpful in the management of ToBRFV.
Keywords:  host range        identity        quadruplex RT-PCR detection        phylogenetic tree        symptom        tobamovirus        tomato brown rugose fruit virus  
Received: 28 March 2020   Accepted:
Fund: This study was supported by the grants from the National Natural Science Foundation of China (31720103912 and 31801704) and the ‘Taishan Scholar’ Construction Project, China (TS201712023).
Corresponding Authors:  Correspondence LI Xiang-dong, Tel: +86-538-8242523, E-mail:; GENG Chao, E-mail:    
About author:  YAN Zhi-yong, E-mail:; ZHAO Mei-sheng, E-mail:; MA Hua-yu, E-mail:; * These authors contributed equally to this study.

Cite this article: 

YAN Zhi-yong, ZHAO Mei-sheng, MA Hua-yu, LIU Ling-zhi, YANG Guang-ling, GENG Chao, TIAN Yan-ping, LI Xiang-dong. 2021. Biological and molecular characterization of tomato brown rugose fruit virus and development of quadruplex RT-PCR detection. Journal of Integrative Agriculture, 20(7): 1871-1879.

Adams M J, Adkins S, Bragard C, Gilmer D, Li D, MacFarlane S A, Wong S M, Melcher U, Ratti C, Ryu K H. 2017. ICTV virus taxonomy profile: Virgaviridae. Journal of General Virology, 98, 1999–2000.
Alkowni R, Alabdallah O, Fadda Z. 2019. Molecular identification of tomato brown rugose fruit virus in tomato in Palestine. Journal of Plant Pathology, 101, 719–723.
Broadbent L, Fletcher J T. 1963. The epidemiology of tomato mosaic. Annals of Applied Biology, 52, 233–241.
Brooks C, Nekrasov V, Lipppman Z B, Van Eck J. 2014. Efficient gene editing in tomato in the first generation using the clustered regularly interspaced short palindromic repeats/CRISPR-associated9 system. Plant Physiology, 166, 1292–1297.
Camacho-Beltrán E, Pérez-Villarreal A, Leyva-López N E, Rodríguez-Negrete E A, Ceniceros-Ojeda E A, Méndez-Lozano J. 2019. Occurrence of Tomato brown rugose fruit virus infecting tomato crops in Mexico. Plant Disease, 103, 1440.
FAO (Food and Agriculture Organization). 2020. Online statistical database: Food balance. FAOSTAT. [2020-02-06].*/E
Fidan H, Sarikaya P, Calis O. 2019. First report of Tomato brown rugose fruit virus on tomato in Turkey. New Disease Reports, 39, 18.
Gülser C, Yilmaz N K, Candemir F. 2008. Accumulation of Tobacco mosaic virus (TMV) at different depths clay and loamy sand textural soils due to tobacco waste application. Environmental Monitoring and Assessment, 146, 235–242.
Hall T A. 1999. BioEdit: A user-friendly biological sequence alignment editor and analysis program for Windows 95/98/NT. Nucleic Acids Symposium Series, 41, 95–98.
Hall T J. 1980. Resistance at the TM-2 locus in the tomato to tomato mosaic virus. Euphytica, 29, 189–197.
Hanssen I M, Lapidot M, Thomma B P H J. 2010. Emerging viral diseases of tomato crops. Molecular Plant-Microbe Interactions, 23, 539–548.
Jacobi V, Bachand G D, Hamelin R C, Castello J D. 1998. Development of a multiplex immunocapture RT-PCR assay for detection and differentiation of tomato and tobacco mosaic tobamoviruses. Journal of Virological Methods, 74, 167–178.
Kumar S, Stecher G, Li M, Knyaz C, Tamura K. 2018. MEGA X: Molecular evolutionary genetics analysis across computing platforms. Molecular Biology and Evolution, 35, 1547–1549.
Kumar S, Udaya Shankar A C, Nayaka S C, Lund O S, Prakash H S. 2011. Detection of Tobacco mosaic virus and Tomato mosaic virus in pepper and tomato by multiplex RT-PCR. Letters in Applied Microbiology, 53, 359–363.
Levitzky N, Smith E, Lachman O, Luria N, Mizrahi Y, Bakelman H, Sela N, Laskar O, Milrot E, Dombrovsky A. 2019. The bumblebee Bombus terrestris carries a primary inoculum of Tomato brown rugose fruit virus contributing to disease spread in tomatoes. PLoS ONE, 14, e0210871.
Li R, Gao S, Fei Z, Ling K S. 2013. Complete genome sequence of a new tobamovirus naturally infecting tomatoes in Mexico. Genome Announcements, 1, 247–262.
Ling K S, Tian T, Gurung S, Salati R, Gilliard A. 2019. First report of tomato brown rugose fruit virus infecting greenhouse tomato in the United States. Plant Disease, 103, 1439.
Liu Y, Li F, Li Y Y, Zhang S B, Gao X W, Xie Y, Yan F, Zhang A S, Dai L Y, Cheng Z B, Ding M, Niu Y B, Wang S J, Che H Y, Jiang T, Shi X B, He Z F, Wu Y F, Zhang D Y, Qing L, et al. 2019. Identification, distribution and occurrence of viruses in the main vegetables of China. Scientia Agricultura Sinica, 52, 239–261. (in Chinese)
Luria N, Smith E, Reingold V, Bekelman I, Lapidot M, Levin I, Elad N, Tam Y, Sela N, Abu-Ras A, Ezra N, Haberman A, Yitzhak L, Lachman O, Dombrovsky A. 2017. A new Israeli Tobamovirus isolate infects tomato plants harboring Tm-22 resistance genes. PLoS ONE, 12, e0170429.
Menzel W, Knierim D, Winter S, Hamacher J, Heupel M. 2019. First report of Tomato brown rugose fruit virus infecting tomato in Germany. New Disease Reports, 39, 1.
Meshi T, Motoyoshi F, Maeda T, Yoshiwoka S, Watanabe H, Okada Y. 1989. Mutations in the tobacco mosaic virus 30-kD protein gene overcome Tm-2 resistance in tomato. The Plant Cell, 1, 515.
Panno S, Caruso A G, Davino S. 2019a. First report of tomato brown rugose fruit virus on tomato crops in Italy. Plant Disease, 103, 1443.
Panno S, Ruiz-Ruiz S, Caruso A G, Alfaro-Fernandez A, Ambrosio M I F S, Davino S. 2019b. Real-time reverse transcription polymerase chain reaction development for rapid detection of Tomato brown rugose fruit virus and comparison with other techniques. PeerJ, 7, e7928.
Pelham J. 1966. Resistance in tomato to tobacco mosaic virus. Euphytica, 15, 258–267.
Salem N, Mansour A, Ciuffo M, Falk B W, Turina M. 2016. A new tobamovirus infecting tomato crops in Jordan. Archives of Virology, 161, 503–506.
Skelton A, Buxton-Kirk A, Ward R, Harju V, Frew L, Fowkes A, Long M, Negus A, Forde S, Adams I P, Pufal H, McGreig S, Weekes R, Fox A. 2019. First report of Tomato brown rugose fruit virus in tomato in the United Kingdom. New Disease Reports, 40, 12.
Strasser M, Pfitzner A J P. 2007. The double-resistance-breaking Tomato mosaic virus strain ToMV1-2 contains two independent single resistance-breaking domains. Archives of Virology, 152, 903–914.
Sui X, Zheng Y, Li R, Padmanabhan C, Tian T, Groth-Helms D, Keinath A P, Fei Z, Wu Z, Ling K S. 2016. Molecular and biological characterization of Tomato mottle mosaic virus and development of RT-PCR detection. Plant Disease, 101, 704–711.
Weber H, Schultze S, Pfitzner A J. 1993. Two amino acid substitutions in the tomato mosaic virus 30-kilodalton movement protein confer the ability to overcome the Tm-22 resistance gene in the tomato. Journal of Virology, 67, 6432–6438.
Yan Z Y, Ma H Y, Han S L, Geng C, Tian Y P, Li X D. 2019. First report of tomato brown rugose fruit virus infecting tomato in China. Plant Disease, 103, 2973.
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