1. KAUTIR (Kerala Agricultural University Theses Information and Retrieval)

The present research work entitled ‘Characterization of virus(es) involved in Chilli leaf curl disease complex in Chilli (Capsicum annuum L.)’ was carried out at Department of Plant Biotechnology and Department of Plant Pathology, College of Agriculture, Vellayani, during the year 2020-2021, with the primary objective to characterize the virus(es) causing Chilli leaf curl disease from five agro-ecological zones of Kerala. Symptomatic chilli plants were collected from different chilli growing agro-ecological zones (AEZs) of the state viz., Nedumangad from Thiruvananthapuram (coastal plains), Kumarakom from Kottayam (Central midlands), Vellanikkara from Thrissur (South Midlands), Thekkepotta from Palakkad (Palakkad plains) and Meenangadi from Wayanad (High Ranges). Highest disease incidence of 80 per cent was observed in Thiruvananthapuram. Symptomatological studies revealed that the infected plants showed extensive curling, crinkling and puckering, apical bunchy appearance, slight yellowing with mosaic patterns on the leaf lamina. Inter-veinal chlorosis accompanied with necrosis and vein-banding was also observed. The association of white- flies (Bemisia tabaci) on the adaxial surfaces of leaves was the best indication for viral infection in chilli plants. Genomic DNA isolated from infected chilli leaf samples using CTAB method showed an absorbance value of 1.8 and genomic RNA extracted using TRIzol method gave an absorbance value of 2.0. Partial characterization of coat protein gene of DNA and RNA viruses were carried out using coat protein specific primers. It produced amplicons of ~550bp specific to AV-AC primers from Thiruvananthapuram, Thrissur and Palakkad and ~500bp amplicons specific to Deng primers from all the AEZs under study. Synthesized cDNA was amplified using the reported coat-protein specific primers viz., CMV and CVMV primers. ~550bp sized amplicons were obtained specific to CMV primers from all the zones except Wayanad where the amplicons obtained were of size ~700bp specific to CVMV primers. Eluted amplicons were ligated to pMD20-T vector, cloned to competent DH5α E.coli cells and screened through Blue- white screening on LB ampicillin/X-Gal plate wherein recombinant transformed cells appeared as white colonies. Those transformed colonies that gave an expected band size in Colony PCR were chosen for sequencing. Sequenced coat protein gene of DNA virus isolated from Thiruvananthapuram, Kottayam, Thrissur, Palakkad and Wayanad resulted in sequence lengths of 558bp, 512bp, 560bp, 515bp and 520bp respectively while that of RNA viruses yielded sequences of length 560bp, 556bp, 558bp, 563bp and 690bp respectively. BLASTn analysis of the DNA viral sequences revealed that Thiruvananthapuram (OM719034), Kottayam (OM719035), Thrissur (OM719036), Palakkad (OM719037) and Wayanad (OM719038) had 98 per cent, 98 per cent, 97 per cent, 98 per cent and 89 per cent similarity with Tomato leaf curl New Delhi Virus. RNA isolates from Thiruvananthapuram (OM719039), Kottayam (OM719040), Thrissur (OM719041) and Palakkad (OM719042) showed an identity of 99.46 per cent, 99.09 per cent, 98.54 per cent and 99.46 per cent with Cucumber mosaic virus isolates and that from Wayanad (OM719043) showed 97 per cent similarity with Chilli veinal mottle virus. Phylogenetic relationship among the DNA isolates showed maximum relatedness between Thiruvananthapuram and Thrissur followed by Kottayam and Palakkad and the Wayanad isolate was categorized as an out group. RNA isolates revealed maximum relatedness between Thiruvananthapuram and Kottayam while that from Palakkad was categorized as out group. To conclude, partial characterization of viruses isolated from chilli plants selected for the study revealed that the chilli plants collected for the study were subjected to mixed infections caused by a complex of viruses. The plants from Thiruvananthapuram, Kottayam, Thrissur and Palakkad were infected by a complex of viruses that belong to genera Begomovirus and Cucumovirus while those plants from Wayanad were infected by a complex comprising of viruses of genera Begomovirus and Potyvirus

1. KAUTIR (Kerala Agricultural University Theses Information and Retrieval)

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