PG Thesis

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Author: search.filters.author.KAUSubject: search.filters.subject.Plant Pathology

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    Prevalence of sesamum phyllody in Onattukara tract and evaluation of fungal root endophyte Piriformospora indica for its management
    (Department of Plant Pathology, College of Agriculture, Vellayani, 2022) Gifty, K J; KAU; Radhika, N S
    The research work entitled ‘Prevalence of sesamum phyllody in Onattukara tract and evaluation of fungal root endophyte Piriformospora indica for its management’ was conducted during 2019-21 at Department of Plant Pathology, College of Agriculture, Vellayani and Onattukara Regional Agricultural Research Station with the objectives to study the symptomatology, molecular detection and characterization of phytoplasma inciting sesamum phyllody disease in AEU 3 (Onattukara tract); and evaluation of fungal root endophyte P. indica for its management. Phytoplasma infected sesamum samples were collected from D and F blocks of Onattukara Regional Agricultural Research Station and Karthikapally. Karthikapally recorded highest disease incidence (39.44 per cent) and vulnerability index (23.75). Chocolate weed, Melochia corchorifolia, was found to be exhibiting symptoms of shoot proliferation. Hoppers collected from the infected fields were identified as Orosius albicintus, Hishimonas phycitis and Nephotettix sp. Disease symptoms were observed at the stage of flowering of sesamum plants in all the sampled locations. The associated symptoms were reduction in internodal length of stem, axillary bud proliferation, thickening of the floral veins, phyllody and floral proliferation. Microtome sections of infected and healthy leaf, stem of sesamum stained with 4,6-diamidino-2-phenylindole (DAPI) stain, and observed under fluorescence microscope emitted diffuse fluorescence from the infected tissues, which was brighter than the one from the parenchymal cells indicating the presence of phytoplasma in the infected tissues. Studies on variations in the level of gibberellic acid (GA) and indole-3-acetic acid (IAA) in phyllody infected and healthy sesamum was undertaken. GA content was increased by 2.25 times and 10.46 times, and IAA content was decreased by 1.25 times and 1.97 times in leaves and flowers of infected samples compared to the healthy samples. Molecular characterization of sesamum phyllody was performed with leaf samples collected from ORARS lowland, ORARS upland, Vellayani and Karthikapally. Amplicons of 1.4kb was obtained by amplifying with universal primers P1/P6 for detection of phytoplasma. The sequences obtained were subjected to BLAST analysis and the 16S rDNA gene sequence showed that all the isolates shared more than 99 per cent similarity with that of the ‘Candidatus phytoplasma aurantifolia’ strains in GenBank data base. In the phylogenetic tree constructed, the sesamum phyllody phytoplasma under study clustered with the 16SrII group (Candidatus Phytoplasma aurantifolia) phytoplasmas causing sesamum phyllody in various regions. The virtual RFLP pattern generated by iPhyClassifier, derived from 16S rDNA fragment was found to be identical to the reference pattern of 16Sr group II, subgroup D (GenBank accession: Y10097). Based on the results obtained from sequence analysis and virtual RFLP pattern, the phytoplasma associated with sesamum phyllody was identified as ‘'Candidatus Phytoplasma aurantifolia”-related strain belonging to subgroup 16SrII-D. P. indica obtained from Department of Plant Pathology, College of Agriculture, Vellayani was mass multiplied in sterilized coir pith: FYM mixture (1:1) amended with 2 per cent gram flour and sesamum seeds were sown. Colonization was observed seven days after germination. Wedge grafting was standardized in sesamum at 30 days after germination. Pot culture experiment was conducted to evaluate the effect of P. indica against phytoplasma causing sesamum phyllody, by grafting the colonized and non-colonized plants with infected scion. P. indica colonization could significantly reduce the incidence and severity of infection. After 30 and 45 days of grafting, an incidence of 20 and 60 per cent, and severity of 5 and 50 were recorded in the colonized plants grafted with infected scion, whereas an incidence of 60 and 80 per cent and severity of 45 and 75 were recorded in non-colonized plants grafted with infected scion. In colonized plants, enhanced shoot and root length at 30 and 55 days after germination were recorded and also earliness in flowering compared to noncolonized plants. Hence the associated symptoms of phytoplasma infection in sesamum are virescence, thickening of floral veins, phyllody and floral proliferation. The study revealed the association of Candidatus phytoplasma aurantifolia group with sesamum phyllody prevalent in Onattukara tract. The evaluation of beneficial fungal root endophyte P. indica against phytoplasma revealed delayed expression of symptoms in the colonized plants.
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    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.