Kumar, J.Siraree, Archana2019-04-272019-04-272004-07http://krishikosh.egranth.ac.in/handle/1/5810101203Black rot disease (c.o. Xanthomonas campestris pv. campestris; Xcc) has become a potential threat to cabbage cultivation in many parts of Uttaranchal hills. The pathogen Xanthomonas campestris pv. campestris is both seed and soil borne but infected seed is an important source of inoculum. The use of pathogen free seeds and use of resistant varieties has been recommended for managing the disease. Present investigation was carried out to standardize procedure (s) for an effective isolation and detection of Xcc from cabbage seed lots, standardize a RAPD-PCR protocol for the characterization of Xcc and assessment of genetic diversity in the Xcc population and test efficacy of Pseudomonas fluorescence as biocontrol agents against Xcc. One hundred and fifty bacterial isolates (presumptive Xcc) were generated from 24 cabbage cultivars/ hybrids by direct plating method on nutrient agar medium, and were subjected to various tests/ assays for confirmation. In gram staining test, bacterial isolates were confirmed as gram negative rods, characteristic of genus Xanthomonas. Of the 150 isolates, 87 were positive for starch hydrolysis on NAM. Eighty three isolates as they showed yellow colonies surrounded by a clear zone of starch hydrolysis on NSCA medium were positive for Xcc. While on BSCA medium 56 isolates showed raised light greenish colonies with a clear zone of starch hydrolysis were considered positive for Xcc. Washing assay revealed that seed washings at room temperature for 1.5 hour was adequate for successful recovery of Xcc. Black rot symptoms appeared on cotyledons of cabbage showing the usefulness of growing on test for detecting Xcc. Chlorotic and necrotic ‘V’ shaped lesions appeared on cabbage leaves of four test varieties 4-7 days after syringe inoculation when five Xcc isolates were tested for pathogenicity. Xcc isolates plated in antibiotic (Streptomycin 640 ug/ml, Kanamycin 60 ug/ml, Gentamycin 60 ug/ml) amended NAM showed optimum growth suggesting that antibiotics at above concentration were not inhibitory to Xcc. Thus, 83 isolates were finally considered to be those of Xcc with their origin from 23 cultivars/ hybrids. Only one hybrid, Hybrid hybrid 23, was free of seed-borne infection. RAPD-PCR protocol was standardized with an amplification cycles of 3.30 hrs. Based on 57 polymorphic DNA fragments generated by three decamer primers, isolates were grouped into two major clusters separated at a linkage distance of 20. Shannon diversity index revealed very high (0.972) diversity among Xcc population. In- vitro evaluation of five Pseudomonas fluorescens strains against Xcc isolates revealed high level of antagonism (42.53%) by the strain Pf551. Results obtained in the present investigation revealed that direct plating for isolation of Xcc pathogen and different detection techniques in combination with pathogenicity test could be used for detection of Xcc from cabbage seed lots. The method could be applied to detect seed borne infection of Xcc in the cabbage seed lots before eradication of seed borne inoculum is attempted. Diverse pathogen population underlines necessity of resistance breeding based on Xcc population structure in the region. As was seen, almost all the cultivars/ hybrids that are still in the pipeline for development and release were susceptible to highly susceptible. Eradication of seed borne inoculum and diseases management through use of Pseudomonas fluorescence (Pf 551) needs to be tested under field condition so as to fit the biocontrol strategy under the framework of organic farming.ennullThesis