Browsing by Author "Deepa James"

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Enhancement of resistance to bacterial wilt in tomato by endophytic microbial communities
(Department of Plant Pathology, College of Horticulture, Vellanikkara, 2015) Deepa James; Sally K Mathew
The study on “Enhancement of resistance to bacterial wilt in tomato by endophytic microbial communities” was conducted in the Department of Plant Pathology, Department of Agricultural Microbiology, and Centre for Plant Biotechnology and Molecular Biology, College of Horticulture, Vellanikkara during the period 2011-2014. The endophytes were isolated from root and stem of healthy tomato plants from 16 locations of north, central and south Kerala. Endophytic microbial population varied with the plant samples and the population was more in root than stem. Bacterial population was higher than fungi and actinomycetes in root and stem. Among 154 endophytes isolated, 12 out of 79 bacteria, 16 out of 68 fungi, and four out of seven actinomycetes were antagonistic to R. solanacearum in in vitro. Among them, five bacteria, eight fungi, and two actinomycetes were promising in planta. Mutually compatible endophytic isolates were selected for the development of consortia and these were identified based on cultural, morphological and molecular characters. Of the five consortia tested, the one consisted of Trichoderma viride-1, T. viride-2, T. harzianum-1, Bacillus subtilis, and Streptomyces thermodiastaticus showed effect in reducing wilt incidence. Comparative study of the microbial consortium with individual endophytes showed the higher efficacy of consortium in reducing the wilt incidence. The endophytes were reisolated from soil, root, and stem of tomato plants. In pot culture experiment, the consortium applied as seed treatment + seedling dip + soil application at 45 DAP showed the minimum wilt incidence. Studies on the mechanism of antagonism of endophytic isolates showed, positive reaction towards ammonia and negative for HCN and siderophore production. The volatile and nonvolatile metabolites of the endophytes inhibited the pathogen. The endophytes showed varying levels of IAA and salicylic acid production with the maximum in T. harzianum-1 and B. subtilis respectively. Study on the effect of secondary metabolites of endophytes on the disease indicated that, seedlings dipped first in culture filtrate for 2 h and later dipped in bacterial inoculum for 30 min before planting showed the lowest per cent wilt incidence with the minimum for T. viride-2. Induction of systemic resistance was studied by assay of defense related compounds such as phenols, oxidative enzymes and PR proteins. The plants treated with microbial consortium showed higher activity of the defense related compounds with the maximum in plants inoculated with both consortium and pathogen. Field evaluation of endophytic consortium against bacterial wilt showed reduction in wilt incidence of 40.85 per cent in highly susceptible variety, PKM-1, 46.94 per cent in susceptible F1 hybrid, COTH-3, and 52.81 per cent in moderately resistant variety, Mukthi. Thus, the study revealed that, the application of endophytic consortium can enhance the resistance mechanism in tomato against bacterial wilt pathogen, R. solanacearum.
Molecular characterisation of Ralstonia solanacearum (Smith) Yabuuchi et al causing bacterial wilt in solanaceous vegetables
(Department of Plant Pathology, College of Horticulture, Vellanikkara, 2001) Deepa James; KAU; Girija, D
Bacterial wilt incited by Ralstonia solanacearum is one of the most devastating diseases of solanaceous vegetable crops in Kerala. Crop losses due to the incidence of this disease may go upto 100 per cent. Existence of different strains, races and biovars has been responsible for breaking down of resistance of varieties evolved through breeding programmes. In view of wide variability, a study was undertaken to characterise the isolates of R. solanacearum collected from three different agro climatic zones ofKerala at molecular level. Nine isolates of R. solanacearum collected from three different locations from brinjal, chilli and tomato were used in the study. These were isolated, purified and maintained in sterile distilled water at room temperature. Inoculation techniques were standardised in brinjal, chilli and tomato plants for assessing the virulence and aggressiveness of the isolates. Virulence and aggressiveness of the isolates were studied on respective host plants and found them highly varying. Vellanikkara and Kumarakom isolates could cross inoculate, whereas Ambalavayal isolates did not. The isolates were characterised by various cultural, morphological and biochemical tests and the variability among them was studied. Biovars, III and IlIA and races, 1 and 3 were identified among the isolates. The isolates were resistant to ampicillin and sensitive to chloramphenicol. Plasmid DNA profile of the isolates were studied and no difference was found in the plasmid DNA profile of the nine isolates. Polymorphism among the isolates was studied using RAPD with ten decamer primers. RAPD profiles exhibited great diversity among biovars III and IlIA as well as among race 1 isolates. Race 3 isolates were less polymorphic with certain primers tested. OPF8 yielded a unique band specific to race 3 isolates. Dendrogram obtained from the pooled data of RAPD profiles also showed high genetic similarity between race 3 isolates. Dendrogram obtained from the pooled data of RAPD profiles also showed high genetic similarity between race 3 isolates. Restriction analysis could not characterise the isolates since no banding pattern was obtained with restricted DNA. No hybridization signal was detected after Southern hybridization in RFLP. Curing of plasmid DNA at high temperatures was found unsuccessful. Plasmid profiles of both mucoid and non-mucoid colonies were compared to assess the role of plasmid in EPS production and the plasmid could be observed in both types of colonies. In the latter, a reduction in size of the plasmid was noticed. Thus the study revealed that great diversity existed among strains of R. solanacearum at different locations of Kerala when molecular techniques, especially RAPD was used as a tool.
Pathogenic variability and management of Lasiodiplodia theobromae causing dieback in mango
(Department of Plant Pathology, College of Agriculture, Vellanikkara, 2025-02-22) Hima Mariya Dixon; Deepa James
Different phenomena like mutation, recombination, natural selection, geographical locations, etc would result in the accumulation of variation in plant pathogenic microorganisms. These variants in the pathogen population often lead to loss of resistance of the cultivar, leaving it highly vulnerable to the disease. Lasiodiplodia theobromae, the causal organism of mango dieback is a highly variable pathogen known to occur in different locations and affecting different mango varieties. In this context, variability studies of Lasiodiplodia theobromae were undertaken to know the variation present among Lasiodiplodia population in Kerala. Purposive sampling surveys were carried out in five districts of Kerala viz., Thrissur (AEU 5, AEU 6, AEU 10 and AEU15), Palakkad (AEU 10, AEU 22 and AEU 23), Wayanad (AEU 20 and AEU 21), Kasaragod (AEU 11) and Kottayam (AEU 4 and AEU 9), thus covering 23 locations and 11 different agroecological units. A total of 37 isolates of the pathogen have been collected from 17 different mango varieties. Per cent disease incidence (PDI) of the different isolates ranged between 25-80 per cent whereas per cent disease severity (PDS) ranged from 17.5-62.5 per cent. General symptoms of dieback under natural conditions were necrotic leaves and black discolouration on the stem and certain isolates showed vascular discolouration too. Upon artificial inoculation of different isolates, a patch of black colour was observed and the range of incubation days varied from 10-15 days while that of lesion size from 0.55 1.1 cm. Diversity in different cultural and morphological characters was also observed. Cultural characters included colony colour (white, greyish white, grey, greyish black, blackish grey, blackish white and black), colony colour on the reverse side of the Petri dish (white and black colour), days taken to cover the Petri plate (2-5), days taken for pycnidia formation (13- 40), days taken for sporulation (22-55) and growth rate (3.34-5.2 cm/day). Certain cultural characters like texture, zonation and margin didn’t show any variation. All the isolates had wooly texture, smooth margin and no zonation was observed in any of the isolates. Different morphological characters also showed wide variation. Morphological characters observed were the colour, size and shape of conidia and the colour and size of hyphae. The range of breadth of hyphae of collected isolates was 3.36-5.18 µm, the range of length of conidia was was 20.83-23.85 µm, and the breadth of conidia was 10.68-13.32 µm. All 37 isolates invariably had cinnamon brown coloured ellipsoid conidia and brown coloured hyphae. The 10 most virulent isolates of Lasiodiplodia theobromae selected for vegetative compatibility studies based on incubation days and lesion size, showed 32 compatible and 13 incompatible out of 45 combinations. The isolates PMuNe,, WMnMa and KoKBn were compatible with maximum number of isolates (eight). Five isolates were selected for pathogenic variability studies based on cluster analysis viz., TNaM (I1), PMuNe (I2), WMnMa (I3), KPB (I4), and KoKBn (I5) and upon molecular characterization confirmed as Lasiodiplodia theobromae. Pathogenic variability of the selected five isolates of Lasiodiplodia theobromae was evaluated against two traditional mango varieties viz., Muvandan and Priyur and four commercial varieties of mango viz., Neelam, Bennet Alphonso, Banganapally, and Imam Pasand. Variability was observed in pathogenicity parameters such as PDI (33.33-100%), PDS (13.33-73.33%), lesion size (0.28-0.95 cm), and incubation period (9-17.33 days). Based on Coefficient of Index (CI) value, Neelam, Bennet Alphonso and Banganapally were categorized as susceptible while the other varieties, Priyur, Muvandan, Bennet Alphonso and Imam Pasand were categorized as moderately susceptible to mango dieback disease. All the isolates produced 100 per cent incidence on Banganapally, confirming its susceptibility. Based on virulence index, three isolates viz., WMnMa (I3), KPB (I4) and KoKBn (I5) with high value of virulence index were selected for their sensitivity to different fungicides and biocontrol agents.