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Molecular characterization of tospoviruses associated with ringspot disease in bell pepper from different districts of Himachal Pradesh

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Abstract

Bell pepper (Capsicum annuum L.), an important cash crop for the farmers of Himachal Pradesh was found to be affected with tospovirus like disease. An extensive survey was conducted in the bell pepper grown areas in the five districts of Himachal Pradesh to identify and characterize the causative agent. Hence, 60 symptomatic bell pepper plants exhibiting characteristics symptoms were collected from Solan, Sirmaur, Hamirpur, Kangra and Bilaspur districts. Out of 60 samples, 53 samples were found to be positive by DAS-ELISA with tospovirus group specific antiserum. To confirm the presence of tospovirus, DAC-ELISA was performed using GBNV/CaCV polyclonal antiserum and DAS-ELISA with two monoclonal antibodies i.e. TSWV, GRSV. All the 53 samples were found negative for TSWV and GRSV and positive for GBNV/CaCV. Further, eleven infected isolates from both poly-house and open field conditions were selected for characterization at molecular level. RT-PCR was performed with N gene specific primers for TSWV, GBNV and CaCV. The eleven samples selected for molecular identification were further found to be negative for TSWV and positive for CaCV using RT-PCR. One of the samples from district Sirmaur was found to be positive for mixed infection of GBNV and CaCV. N gene phylogenetic analysis of CaCV/GBNV provided important information about the movement and evolution of tospoviruses in Himachal Pradesh.

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Acknowledgments

The authors would like to thank Prof. P. K. Khosla, Hon’ble Vice-Chancellor, Shoolini University of Biotechnology and Management Sciences, Solan and Foundation for Life Sciences and Business Management (FLSBM), Solan for providing financial support and necessary facilities. Authors are also thankful to Dr Bikash Mandal (Senior Scientist), Indian Agricultural Research Institute (IARI) for kindly providing GBNV/CaCV specific antibodies.

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Correspondence to Saurabh Kulshrestha.

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13337_2016_315_MOESM1_ESM.jpg

Agarose gel electrophoresis of RT-PCR products of samples C1, T, H1, J1, G5. I1, I2, I4, R1 and Chb with N gene/protein specific primer of CaCV. Lane 1 Marker; Lane 2,3 C1 and T isolates (Sirmaur); Lane 4 H1 isolate (Hamirpur); Lane 5, 6 J1 and G5 isolates (Bilaspur); Lanes 7, 8, 9 I1, I2 and I3 isolates (Kangra); Lane 10, 11 R1 and Chb isolates (Solan); Lane12 One Kb plus DNA Marker (Fermentas). Supplementary material 1 (JPEG 21 kb)

Supplementary material 2 (DOCX 18 kb)

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Sharma, A., Kulshrestha, S. Molecular characterization of tospoviruses associated with ringspot disease in bell pepper from different districts of Himachal Pradesh. VirusDis. 27, 188–192 (2016). https://doi.org/10.1007/s13337-016-0315-y

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