Abstract
Watermelon bud necrosis virus (WBNV) and Groundnut bud necrosis virus (GBNV) are two closely related tospovirus species infecting several crops in India. In the present study, specific diagnostic assays for these two most predominant tospoviruses were developed by comparing host reactions and genome sequence information. Bioassay was developed by sap transmission of both the viruses to a set of host plants, cowpea, groundnut and watermelon. WBNV induced only local symptoms including chlorotic spots on cowpea cv. Pusa Komal, typical chlorotic ring spots on cv. C-152, bud necrosis on watermelon and no symptoms on groundnut. Whereas, GBNV induced both local and systemic symptoms including chlorotic/necrotic spots and veinal necrosis on both the cowpea cultivars, systemic yellowing and bud necrosis on groundnut and no symptoms on watermelon. Single and duplex RT-PCR was developed for diagnosis of WBNV and GBNV by designing primers based on differential sequence from Gn/Gc genes located on M RNA and from nucleocapsid protein (NP) gene located on S RNA genome. The duplex RT-PCR using NP gene specific primers was successfully utilised for the diagnosis of natural infection of GBNV in dahlia and muskmelon and mixed infection of GBNV and WBNV in chrysanthemum. The amplified products of duplex RT-PCR were sequenced and phylogenetic analysis confirmed the authenticity of the RT-PCR assay as well as confirmed infection of WBNV and GBNV in the new natural hosts. The bioassay and RT-PCR developed in this study will be useful in detecting single and mixed infection of GBNV and WBNV strains in different crops.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
Abbreviations
- WBNV:
-
Watermelon bud necrosis virus
- GBNV:
-
Groundnut bud necrosis virus
- RT-PCR:
-
Reverse transcription polymerase chain reaction
- NP:
-
Nucleocapsid protein
- Gn/Gc:
-
Glycoprotein precursor
References
Ghanekar AM, Reddy DVR, Iizuka N, Amin PW, Gibbons RW (1979) Bud necrosis of groundnut (Arachis hypogaea) in India caused by tomato spotted wilt virus. Ann Appl Biol 93:173–179
Jain RK, Pappu HR, Pappu SS, Krishana Reddy M, Vani A (1998) Watermelon bud necrosis tospovirus is a distinct virus species belonging to serogroup IV. Arch Virol 143:1637–1644
Jain RK, Pandey AN, Krishnareddy M, Mandal B (2005) Immuno-diagnosis of groundnut and watermelon bud necrosis viruses using polyclonal antiserum to recombinant nucleocapsid protein of Groundnut bud necrosis virus. J Virol Methods 130:162–164
Jain RK, Bag S, Umamaheswaran K, Mandal B (2007) Natural infection by tospoviruses of cucurbitaceous and fabaceous vegetable crops in India. J Phytopathol 155:22–25
Kumar R, Mandal B, Geetanjali AS, Jain RK, Jaiwal PK (2010) Genome organisation and sequence comparison suggest intraspecies incongruence in M RNA of Watermelon bud necrosis virus. Arch Virol 155:1361–1365
Kunkalikar SR, Sudarsana P, Arun BM, Rajagopalan P, Chen TC, Yeh SD, Naidu RA, Zehr UB, Ravi KS (2011) Importance and genetic diversity of vegetable-infecting tospoviruses in India. Phytopathology 101:367–376
Li TJ, Yeh CY, Yeh DS, Joseph A, Raja J, Rajgopalan AP, Liu YL, Chen CT (2011) Complete genome sequence of Watermelon bud necrosis virus. Arch Virol 156:359–362
Mandal B, Jain RK (2010) ELISA kit for tospoviruses detection. ICAR News 16:13
Mandal B, Pappu HR, Culbreath AK (2001) Factors affecting mechanical transmission of Tomato spotted wilt virus to peanut (Arachis hypogaea). Plant Dis 85:1259–1263
Mandal B, Jain RK, Chaudhary V, Varma A (2003) First report of natural infection of Luffa acutangula by Watermelon bud necrosis virus in India. Plant Dis 87:598
Mandal B, Jain RK, Krishnareddy M, Kumar Krishna NK, Ravi KS, Pappu HR (2012) Emerging problems of tospoviruses (Bunyaviridae) and their management in the Indian subcontinent. Plant Dis 96:468–479
Pappu HR, Jones RAC, Jain RK (2009) Global status of tospovirus epidemics in diverse cropping systems: successes achieved and challenges ahead. Virus Res 141:219–236
Rebijit KB, Asokan R, Krishna Kumar NK, Krishna V, Ramamurthy VV (2012) Development of species-specific markers and molecular differences in mtDNA of Thrips palmi Karny and Scirtothrips dorsalis Hood (Thripidae:Thysanoptera), vectors of tospoviruses (Bunyaviridae) in India. Entomol News 122(3):201–213
Reddy M, Reddy DVR, Appa Rao A (1968) A new record of virus disease on peanut. Plant Dis Rep 52:494–495
Renukadevi P, Nagendran K, Nakkeeran S, Karthikeyan G, Jawaharlal M, Alice D, Malathi VG, Pappu HR (2015) First report of Tomato spotted wilt virus infection of chrysanthemum in India. Plant Dis 99:1190
Singh SJ, Krishnareddy M (1996) Watermelon bud necrosis: a new tospovirus disease. Acta Hortic 431:68–77
Takeshita M, Nagai N, Okuda M, Matsuura S, Okuda S, Furuya N, Tsuchiya K (2011) Molecular characterization of Chrysanthemum stem necrosis virus isolates from distinct regions in Japan. Eur J Plant Pathol 131:9–14
Acknowledgments
The doctoral fellowship to the first author by PG School IARI, New Delhi is gratefully acknowledged. Authors are highly thankful to the Director, Directorate of Floriculture Research, IARI, New Delhi for providing chrysanthemum samples.
Funding
This study was funded by Department of Biotechnology (DBT), New Delhi (Grant No. BT/PR7866/AGR/02/379/2006), and NAIP Component-4 (ICAR-World Bank) subproject Novel strategies for molecular diagnosis of plant viruses.
Author information
Authors and Affiliations
Corresponding author
Rights and permissions
About this article
Cite this article
Holkar, S.K., Kumar, R., Yogita, M. et al. Diagnostic assays for two closely related tospovirus species, Watermelon bud necrosis virus and Groundnut bud necrosis virus and identification of new natural hosts. J. Plant Biochem. Biotechnol. 26, 43–51 (2017). https://doi.org/10.1007/s13562-016-0358-6
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s13562-016-0358-6