Identification of sources of resistance and effective non chemical methods for the management for major viral disease of chilli in Kerala

Abstract

The present study was undertaken in an endeavor to gain some understanding regarding the viral diseases affecting chilli and devising suitable management strategies to contain them. A purposive sampling survey conducted involving 3 districts of Kerala, namely Palakkad, Thrissur and Malappuram covering over 15 locations and 44 samples revealed symptoms descriptive of viral infection like curling, puckering, cupping and yellowing of leaves, dwarfing of plants and low fruit set in affected plants. To identify the etiological agent(s) associated with the symptoms, transmission studies were carried out. Whitefly transmission and graft transmission gave positive results whereas none of the samples that were sap transmitted exhibited symptoms indicating the presence of a geminivirus. Detection methods like Transmission Electron Microscopy (TEM) and Enzyme Linked Immunosorbant Assay (ELISA) were also carried out. Geminate particles were observed but no mosaic causing virus was detected in EM. ELISA performed on samples with mosaic like symptoms for the presence of potyvirus and CMV did not detect the presence of suspected virus in it. Hence, it was concluded that leaf curl disease was the only major disease affecting the crop in the area. Characterization of the virus (s) responsible for the leaf curl disease was attempted. DNA was isolated from seven representative samples after which PCR (Polymerase Chain Reaction) was performed with two sets of primers, degenerate (DENG) primer and a specific primer. The PCR amplified products of four representative isolates were sequenced and analysed for homology with sequences available in NCBI using BLAST software. Three of the isolates showed maximum nucleotide identity to the Pepper leaf curl Bangladesh virus isolate India/Coimbatore/Chilli/2008 segment DNA-A, complete sequence (HM007096.1) and the other isolate showed maximum identity with Chilli leaf curl virus-India isolate Raichur segment DNA-A, complete sequence (MK161454.1). A phylogenetic tree was constructed combining different geminiviruses reported from different crops to study the sequence similarity existing amongst them. The amino acid sequence of the coat protein gene of the isolate PKD-1 was predicted using the ExPASy translation tool available online. With the aim of identifying source (s) of resistance against chilli leaf curl disease in Kerala, two sets of screening experiments were carried out. In the initial field screening of 30 accessions, Violet Mulak, a local accession remained symptomless whereas Arka Lohit (IIHR) and Phule Jyothi (MPKVV) remained highly resistant. The molecular confirmation of the incidence of Chilli leaf curl virus (ChiLCV) in the field was done by performing PCR using primers specific to the CP gene region of the concerned virus. A pot experiment was set up to once again score the disease reaction of 48 accessions including those used in field screening along with a few more accessions. In this experiment, Phule Jyothi remained symptomless and Violet mulak turned out highly resistant whereas Arka Lohit remained HR. Twentysix random infected accessions were confirmed for the presence of ChiLCV in them. The three accessions that were adjudged highly resistant following the conclusions of the abovesaid two screening experiments were subjected to graft transmission. Fifty days old field exposed plants of the resistant sources were used as rootstocks in wedge grafting with scions from known susceptible variety. At 60 days of grafting, rootstocks and scions failed to amplify the primer indicating the absence of virus in them. From all these experiments, what could be inferred is that these accessions possessed true resistance against the virus up to 50 days after transplanting in the field. A management study to evaluate the efficacy of certain plant extracts and biocontrol agents in containing chilli leaf curl disease was undertaken the results of which indicate spraying the leaf extract of Azadirachta indica (10 %) or Mirabilis jalapa (10 %) as the best treatment. The only biocontrol agent with an effect on par with the best treatments was Bacillus subtilis applied as seed treatment (10g kg-1 seed) and foliar spray (20g l-1). All the four plant extracts did have a reducing effect on the disease severity with the least mean recorded in A. indica (10 %).