000			06569nam a22002057a 4500
999			_c291787 _d291787
082			_a632.3 _bJOS/MO Ph.D
100			_aJosiya Joy
245			_aMolecular diagnosis and management of Papaya ringspot virus causing papaya ringspot diseases
260			_aVellayani _bDepartment of Plant Pathology, College of Agriculture _c2024
300			_a162,xviiip.
502			_aPh.D
520	3		_aThe research work entitled “Molecular diagnosis and management of Papaya ringspot virus causing papaya ringspot disease” was undertaken in the Department of Plant Pathology, College of Agriculture, Vellayani, Thiruvananthapuram, during 2019-24, with the objectives; molecular diagnosis and recombinant coat protein production of Papaya ringspot virus (PRSV), and evaluation of the efficacy of beneficial microorganisms and botanical inthe management of papaya ringspot disease (PRSD). Roving survey was carried out across five Agro-ecological units (AEUs) of Kerala. The disease incidence (DI) ranged from 50.25 per cent (Kayyur-Cheemeni) to 100 per cent (Kalliyoor, Venganoor, Balaramapuram, Pallichal, Kayamkulam, Mavelikkara, Velukkara, Irinjalakuda and Shoranur). Vulnerability index (VI) of the plants to PRSV in the surveyed locations ranged from 33.54 (Badiyadkka) to 98.22 (Kalliyoor). Serological and molecular detection confirmed the presence of PRSV in all the 20 symptomatic samples collected during survey. Phylogenetic tree constructed with the deduced amino acid sequences of CP gene of 11 Kerala PRSV isolates, revealed that Thiruvananthapuram isolates clustered together, indicating their relatedness during evolution. Mechanical inoculation on two months old Red Lady papaya seedlings under insect proof conditions was carried out to identify the most virulent PRSV isolate collected from different AEUs. At three months after inoculation (MAI), Kalliyoor isolate exhibited highest VI (96.63) followed by Alur (95.48) and Venganoor (95.22) isolates. The lowest VI was observed in Kayyur-Cheemeni isolate with 57.95 VI, followed by Cherpulassery (60.58). The recombinant coat protein of PRSV was induced in pLATE 31 expression vector with C terminal histidine tag, within BL21(DE3)pLysS expression host. PRSV recombinant coat protein was purified using Ni-NTA column chromatography. A single band was observed at 35 kDa in SDS-PAGE analysis and western blotting with PRSV antiserum, confirmed the presence of purified recombinant coat protein in the soluble fraction. Pot culture experiment was conducted to evaluate the management of PRSD using Piriformospora indica. The initial colonization of P. indica inside the papaya roots was observed five days after germination of the seeds grown in P. indica massmultiplied medium. Prophylactic colonization of P. indica exhibited lowest VI (23.10) and 68.27 per cent reduction in VI over control diseased plants at five months after PRSV inoculation (MAI). Double antibody sandwich - Enzyme linked immunosorbent assay (DAS-ELISA) at 5 MAI revealed lowest absorbance (0.23) indicating lowest virus titre in P. indica pre-colonized plants upon PRSV inoculation, compared to control diseased plants (1.23). The accumulation of reactive oxygen species (ROS) i.e., H2O2 and superoxides in the leaves were analyzed using DAB (diaminobenzidine) and NBT (nitro blue tetrazolium chloride) staining respectively. P. indica-colonized plants upon PRSV inoculation indicated a higher initial accumulation of ROS at three weeks after inoculation (3 WAI) and further reduction at 6, 9 and 12 WAI of PRSV, compared to control diseased plants. P. indica-colonized plants also exhibited enhanced antioxidant defence enzyme activity viz., catalase, peroxidase, polyphenol oxidase, phenylalanine ammonia lyase, glutamate synthase and superoxide dismutase compared to control diseased plants. Amongst PRSV inoculated treatments, P. indica pre-colonized plants exhibited highest number of leaves (20.86), leaf area (365.14 cm2), plant height (108.29 cm), stem girth (7.26 cm), shoot biomass (423.43 g), root biomass (133.96 g) and chlorophyll content (2.84 mg g-1 of fw) at five months after transplanting (5 MAT). Effect of P. indica, Bougainvillea spectabilis leaf extract (10 %) and Pseudomonas fluorescens (2 %) were evaluated against natural incidence of PRSD under field conditions at Instructional farm, College of Agriculture, Vellayani and Coconut Research Station, Balaramapuram, Thiruvananthapuram. P. indica-colonized plants exhibited lowest VI (37.63), with highest reduction in VI over control (53.85 %) followed by B. spectabilis treated plants (37.49 %) and P. fluorescens treated plants (33.31 %) at 12 months after planting (12 MAP). In DAS-ELISA, lowest virus titre with absorbance of 0.438 was observed in P. indica-colonized plants at 12 MAP, compared to the highest virus titre in control plants (1.267). B. spectabilis treated plants (0.596) also exhibited reduction in virus titre followed by P. fluorescens treated plants (0.625) at 12 MAP. P. indica-colonized plants exhibited enhancement in growth parameters viz., number of leaves (24.00), leaf area (1127 cm2), stem girth (42.74 cm) and plant height (207.39 cm) at 12 MAP. P. indica-colonized plants also enhanced the yield by 44.68 per cent followed by B. spectabilis treated plants (24.13 %) and P. fluorescens treated plants (17.99 %). Moreover, fruits from P. indica-colonized plants expressed significantly superior quality parameters. Thus, findings from the present study could aid in the preliminary detection and management of the virus, thus mitigating the widespread infection caused by PRSV. The recombinant PRSV coat protein produced in this study could be used for the development of PRSV antiserum. Additionally, our research highlights the efficacy of eco-friendly management strategies for papaya ringspot disease, with P. indica-colonization @ 106 cfu g-1 and also with four foliar sprayings as well as soil drenching of B. spectabilis leaf extract (10 %) applied at fortnightly intervals, starting from one month after planting. More field trials are to be conducted to integrate this strategy in integrated disease management (IDM) package for the effective and sustainable management of PRSD in papaya.
650			_aPlant Pathology
650			_aPapaya
650			_aPapaya ringspot virus
650			_aPapaya ringspot diseases
700			_aRadhika, N S(Guide)
856			_uhttps://krishikosh.egranth.ac.in/handle/1/5810219010
942			_2ddc _cTH