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ThesisItem Open Access Molecular characterization, host range and integrated management of bhindi yellow vein mosaic disease(Department of Plant Pathology, College of Horticulture, Vellanikkara, 2019) Chinju, E A; KAU; Anita Cherian, KBhindi (Abelmoschus esculentus (L.) Moench) is one of the most important vegetable crops cultivated across the globe. However, its cultivation is often hindered by biotic stresses like incidence of pests and diseases. Among the diseases, yellow vein mosaic disease is one of the major constraints in bhindi cultivation which leads to 100 per cent yield loss especially when infected at an early stage of the crop. In recent years, the evolution of new viral strains is a serious problem especially among Begomoviruses belonging to the family Geminiviridae which has an adverse effect on the host plant resistance. Considering these facts, the present study was undertaken to carry out the molecular characterization of the virus causing bhindi yellow vein mosaic disease (BYVMD), to study the host range and seed transmission of the virus and to develop a sustainable disease management strategy. The project was initiated with purposive sampling survey conducted in elevan different locations of Thrissur district, Kerala. The disease incidence recorded during the survey ranged from 61.20 to 98.16 per cent while the disease severity ranged from 48 to 90 per cent. The predominant symptoms observed on the leaves of infected plants under natural conditions were vein clearing, vein thickening, reduction in leaf area, bleached appearance and marginal necrosis. A novel type of symptom observed during the survey was general yellowing of leaves with severe puckering along the veins and upward curling of leaves. Linear cholorotic striations were observed on the calyx of the flower buds. The immature fruits produced by the infected plants showed linear chlorotic striations, while the mature fruits were bleached in appearance along with reduction in fruit size. The plants infected during the early vegetative stage were extremely stunted. The major symptoms developed under artificial conditions were vein clearing, vein thickening and puckering of leaves. xxiv Histopathological studies of the infected leaf revealed disruption of parenchymtous cells in the epidermis, disintegration of chloroplast, reduction in number of metaxylem and protoxylem along with abnormality of phloem vessels. The studies on virus transmission confirmed that it is transmitted through grafting and insect vector, Bemisia tabaci. The presence of virus inside the insect body was also confirmed through polymerase chain reaction (PCR) based molecular technique. The studies on seed transmission revealed that BYVMD is not seed-borne. Host range studies revealed that weed species Synedrella nodiflora and Hemidesmus indicus were proved to be hosts of the begomovirus. Molecular detection of the virus causing BYVMD was standardized through PCR, using universal primer specific to the core coat protein gene of Begomovirus which yielded amplicons at expected size of about 550 bp. The amplification was also carried out using primers specific to coat protein gene of bhindi yellow vein mosaic virus (BYVMV) and okra enation leaf curl virus (OELCuV) which yielded amplicons at expected band size of about 770 bp. The molecular characterization of the elevan isolates was carried out through in silico analysis to identify the virus associated with BYVMD and for diversity analysis. The results revealed that all the isolates showed 99-100 per cent nucleotide homology to OELCuV. BLASTp analysis of the isolates also showed 100 per cent identity with coat protein of OELCuV and thus confirming that the virus causing yellow vein mosaic disease in bhindi in Kerala is okra enation leaf curl virus. The identity was further confirmed through DNA barcoding technique and species demarcation analysis. A field experiment was also conducted to develop a disease management package against BYVMD. Among the seven treatments, T5 i.e., integrated management with early seedling protection using insect proof net + yellow sticky trap + seed bio-priming and foliar spray of PGPR mix II + alternate foliar spray of Bougainvillea spectabilis and azadiractin was found to be most effective with xxv lowest disease incidence and severity, least whitefly population and maximum yield. It is concluded that yellow vein mosaic disease affecting bhindi cultivation in Kerala is caused by okra enation leaf curl virus, an evolved strain of BYVMV. This virus is transmitted through grafting and the insect vector Bemisia tabaci and not though seeds. The outcome of the study would also facilitate early detection and elimination of sources of infection so as to reduce the spread of the disease. An integrated disease management package was also developed for the benefit of farming community.ThesisItem Open Access Characterization and evaluation of Pseudomonas spp. for abiotic stress tolerance(Department of Plant Pathology, College of Horticulture, Vellanikkara, 2019) Reshma, K S; KAU; Reshmy, VijayaraghavanPseudomonas spp. are one among the most extensively used biocontrol agent in plant disease management to control foliar, soil borne or seed borne pathogens. However, prevalence of abiotic stresses such as drought, high temperature, salinity and acidity may affect the field performance due to poor survival under adverse conditions. To date, very little effort has been taken to tap microbial diversity from stressed ecosystems of Kerala. Thus, a study was undertaken to isolate native strains of Pseudomonas spp. having inherent stress tolerance. Purposive soil sampling survey was conducted during the period February- April 2018 and a total of 26 representative soil samples were collected from four districts viz., Ernakulam, Palakkad, Thrissur and Alappuzha. It was revealed that various locations of Palakkad and Thrissur district experienced high temperature (40-46oC) and low moisture of 1.1- 3.9 per cent. Samples procured from Alappuzha and Ernakulam districts were extremely acidic (pH 3.4- 4.5) andsaline (EC 4.1-6.68 dSm-1). The results of soil analysis confirmed that the collected soil samples were abiotically stressed and there could be chances of obtaining stress tolerant native isolates of Pseudomonas spp. For isolation of Pseudomonas spp., soil samples weresubjected to serial dilution and plating technique in King’s B agar and Pseudomonas agar base, among which King’s B agar yielded optimum number of Pseudomonas colonies at 105 dilution.Maximum population of Pseudomonas spp. was recorded in samples procured from Moncombu (1.3-7.8x106cfu g-1), whereas, minimum population was observed in soil samples collected from Vyttila (3.14- 6.01 x 104cfu g-1). A total of 15 isolateswere purified and designated with sample codes based on the place of collection. Among the nine fluorescent isolates, M7 and P7 were identified as Pseudomonas aeruginosa, an opportunistic human pathogen based on its growth at 42oC and on King’s A agar medium and thus, were eliminated. Isolates of Pseudomonas spp. were subjected to in vitro screening for abiotic stress tolerance and compared with reference culture of KAU. The isolates P2, M4 and T5 were selected as temperature tolerant (50oC), M4 and V4 as salt tolerant (1.5 M NaCl), P4 and T4 as xxxii drought tolerant (30% PEG) and M4 and M5 as acid tolerant (pH 3.5) strains. Subsequently, a total of seven abioticstress tolerant isolates viz., M4, M5, P2, P4, V4, T4 and T5 were evaluated further for their in vitro antagonistic potential against five major soil borne fungal pathogens using Bangle method.The four isolatesviz., P2, P4, M4 and M5 were selected as the potential antagonistic strains since they outperformed reference culture of KAU and showed higher and consistent biocontrol activity with per cent growth inhibition ranging from 62.21 to 91.00 per cent against Phytophthora capsici, Pythium aphanidermatum, Sclerotium rolfsii, Rhizoctonia solani and Fusarium oxysporum. Further studies revealed M4, M5 as better producers of ACC deaminase enzyme and M4, P4 as better producers of exopolysaccharide. Highest cellulase and β-1, 3 glucanase production was recorded by the isolates P2 and M4 respectively. A preliminary screening of the isolates were carried out for testing the bioefficacy on cowpea seeds against Rhizoctonia solani using roll towel method. The results revealed that the isolate M4 was highly efficient, reference culture as efficient and the other isolates P2, P4 and M5 as moderately efficient.All the four isolates were further evaluated for their in vivo activity through a pot culture experiment using cowpea- R. solani system. Eventhough, M4 outperformed all other isolates with highest seed germination, biometric characters and yield as well as with lowest per cent disease incidence, other three isolates were also found superior or equally efficient as reference culture of KAU. The best performing isolates with promising traits of stress tolerance, antagonism and plant growth promotion viz., P2, P4, M4 and M5 were identified based on cultural, morphological, biochemical and molecular characterization. The isolate P2 was identified as Pseudomonas putida, P4 as P. fluorescens and M4 and M5 as P. aeruginosa. Thus, the current investigation has thrown light on the prevalence of abiotic stress tolerant strains of Pseudomonas spp. in stressed ecosystems which would significantly help the farming community to overcome such drastic climate changes. However, the study should further be complemented with large scale multilocational trials to prove their efficacy under field conditions.ThesisItem Open Access Characterisation of Ralstonia solanacearum (Smith) Yabuuchi et al. infecting solanaceous vegetables in relation to Physico-chemical and Biological properties of soil(Department of Plant Pathology, College of Horticulture, Vellanikkara, 2019) Anjali, V A; KAU; Sainamole Kurian, PRalstonia solanacearum, the causal agent of vascular wilt disease of crop plants has been ranked as the second most important bacterial pathogen in the world next to Pseudomonas syringae. The high diversity exhibited by the pathogen hampers the breeding for resistance, consequently the resistant varieties developed may not express uniform level of resistance in different geographical locations. Being a soil inhabitant, the survival of R. solanacearum is influenced by physico-chemical and biological properties of soil. Considering these facts, present investigation was carried out with the objective of characterisation of R. solanacearum from different agro ecological units (AEUs) of Kerala and to determine the effect of soil physical, chemical and biological properties on the pathogen. Purposive sampling survey was conducted in four AEUs of Kerala viz., North Central laterite (NCL), Marayur hills (MH), Southern laterite (SL) and Palakkad central plains (PCP) from March to September 2018. Two locations from each AEUs were selected for the survey. The per cent incidence in different locations ranged from 20 to 88 per cent. The pathogen was isolated from infected plant samples collected during the survey on triphenyl tetrazolium chloride (TZC) agar and pathogenicity was proved by inoculation on respective hosts. The rhizosphere soil samples of healthy and diseased plants were collected from each location. The population of the pathogen present in the soil was quantified and it ranged from 20.66 x 104 cfu g-1of soil to 137.66 x 104 cfu g-1 of soil. A strong positive correlation was observed between pathogen density in soil and per cent disease incidence (PDI). A total of eight isolates were collected, purified and maintained for all the experiments. The colony characters of different isolates on TZC agar showed considerable variation and the morphology of the bacterial cells was studied using scanning electron microscopy. Typical rod shaped cells with size 0.3-0.5 µm x 1.2-1.7 µm were observed. The molecular characterization of the isolates was done by PCR amplification and sequencing of 16S rRNA. The sequences were subjected to in-silico analysis which confirmed the identity of all isolates as R. solanacearum (Smith) Yabuuchi et al. The phylogenetic analysis revealed that the eight isolates collected from different AEUs clustered on different branches of the tree while those from the same AEUs clustered together. This indicates considerable variation among the isolates in accordance with location which can be attributed to the difference soil parameters in these locations. The isolates of the pathogen were further categorized into races and biovars based on pathogenicity on differential hosts and utilization of disaccharides and hexose-alcohols respectively. The results revealed that two isolates from Marayur hills (MH 1 and MH 2) belong to race 3, biovar II whereas two from Palakkad central plains (PCP 1 and PCP 2) belong to race 1 biovar III A. The other four isolates collected from Northern central laterite (NCL 1 and NCL 2) and Southern laterite (SL 1 and SL 2) were identified as race 1, biovar III. The physico-chemical and biological properties of the soil samples collected during the survey were analysed using standard protocols. The statistical analysis using paired sample t-test revealed significantly higher soil pH, organic carbon, available K, Ca and Fe content and soil microflora in rhizosphere soil of healthy plant compared to diseased. A significant positive correlation was observed between PDI and soil parameters viz., water holding capacity and bulk density whereas soil pH and available Ca content exhibited a negative correlation with PDI. A similar trend was observed in the case of pathogen population also. Further, multiple regression analysis was performed to assess the extent of variation contributed by different soil parameters on PDI and pathogen population. The results indicate that 96.8 per cent variation in the bacterial wilt incidence is explained by soil pH and available Ca content in the rhizosphere soil with negative correlation whereas bulk density and Ca content contributed 92.2 per cent in the build-up of population of Ralstonia solanacearum in soil. The study revealed the influence of soil factors on bacterial wilt disease incidence, population of R. solanacearum and pathogen variability. Hence, it is concluded that, manipulation of soil factors can play a major role in integrated management of the disease. Furthermore, the variability of pathogen according to geographical region, may be considered while planning resistance breeding programmes against bacterial wilt disease.ThesisItem Open Access Physiological and cultural studies on blue oyster mushroom (Hypsizygus ulmarius (Bull.:Fr.) Readhead)(Department of Plant Pathology, College of Agriculture, Vellayani, 2016) Sumi, I; KAU; Geetha, DThe present study entitled “Physiological and cultural studies on blue oyster mushroom (Hypsizygus ulmarius (Bull.:Fr.) Redhead)” was carried out in the mushroom unit, Instructional Farm, College of Agriculture, Vellayani during 2014-2016, with the objective to standardize the technology for cultivation of Hypsizygus ulmarius and to study its morphological and physiological aspects. The initial culture of H. ulmarius was isolated from the mushroom beds maintained in the mushroom unit of instructional farm through tissue culture method and purified by hyphal tip method. Morphological studies of H. ulmarius showed that the sporocarps were medium to large in size having a dark blue colour in the pinhead stage which became creamy white on maturity with an irregularly shaped, convex pileus with gills attached to the stem, but not decurrent and cylindrical, smooth and eccentric stipe. Microscopic studies revealed septate hypahe with clamp connection, oval shaped, hyaline basidiospores and the spore print was white. Studies on developmental morphology showed that H. ulmarius took an average of five days from the day of pinhead formation to complete maturity. The maximum mycelial growth was recorded on potato dextrose agar. A temperature of 25 0C, pH of 8 and dark conditions are found favourable for maximum mycelial growth. Evaluation of different substrates for spawn production revealed that paddy grains was the best medium in which spawn run was completed in fifteen days with thick fluffy growth and recorded less contaminants followed by wheat and sorghum. Evaluation of different substrates for mushroom production revealed that paddy straw was the best material for the cultivation of blue oyster with a total yield of 985 g kg-1 from three harvests followed by rubber sawdust (905 g kg-1). The minimum time for mushroom production was recorded for sugarcane bagasse and the maximum time for rubber sawdust. The average weight of sporocarp was maximum in mushrooms harvested from rubber sawdust and the maximum number of sporocarps was recorded in paddystraw. Beds prepared from sugarcane bagasse were heavily contaminated with Trichoderma sp. When compared with Pleurotus florida, H. ulmarius took more time (18 days) for complete spawn run in paddy grains and the yield was higher on paddy straw (1.096 kg kg-1) than P. florida (976 g kg-1). Infestation of pests viz., phorid flies (Megaselia sp.) and staphylinid beetles were prevalent during spawn run as well as sporocarp formation. The competitor moulds recorded were Trichoderma sp., Aspergillus sp. and Coprinus sp. Analysis for the proximate constituents in H. ulmarius revealed that it contains appreciable amount of carbohydrate (29 %), protein (32 %) and fibre (17.69 %). Sensory evaluation was done on steam cooked mushrooms for attributes like appearance, colour, texture, flavor and taste using five point score card and an overall acceptability score of 3.6 was obtained for H. ulmarius compared to P. florida (3.0). In the preference study conducted for both the mushrooms using Hedonic rating scale, 30 per cent of evaluators extremely liked H. ulmarius than P. florida (10 %). The study on the keeping quality of mushrooms in normal atmospheric condition indicated a shelf life of eight hours for H. ulmarius compared to six hours for P. florida. The study also showed that blue oyster mushrooms stored under refrigeration (4 0C) in perforated polythene covers had better shelf life (5 days) compared to P. florida (3 days). The present study indicated that blue oyster mushroom can be cultivated successfully in tropical areas on locally available materials like paddy straw and rubber saw dust under favourable climatic conditions viz., 26-28 0C temperature, more than 90 per cent relative humidity and good aeration. The variety is superior to the presently growing oyster mushroom (P. florida) in terms of yield, presence of appreciable amount of proximate constituents and keeping quality.ThesisItem Open Access Integrated management of rhizoctonia leaf blight of amaranthus (Amaranthus tricolor L.)(Department of Plant Pathology, College of Agriculture, Vellayani, 2016) Gireesh; KAU; Radhakrishnan, N VThe study entitled “Integrated management of Rhizoctonia leaf blight of Amaranthus (Amaranthus tricolor L.)” was conducted at the College of Agriculture, Vellayani and Coconut Research Station, Balaramapuram during 2014-2016 with the objective to investigate the effect of soil solarization, biocontrol agents, chemical activator, indigenous formulations and new generation fungicides on growth, yield and severity of foliar blight of amaranthus. Samples of the infected leaves showing Rhizoctonia leaf blight in amaranthus were collected from Vellayani, Kalliyoor, Venganoor and Kakkamoola locations. Among the four isolates of the pathogen, the Vellayani isolate gave significantly superior growth rate with minimum of six days for sclerotial formation. Koch‟s postulates were proved for the pathogenicity of different isolates of Rhizoctonia solani. All the four isolates have taken three days for the first symptom development but the progression of lesion size of Vellayani isolate was maximum compared to all other isolates, hence the Vellayani isolate was selected as the most virulent isolate for use in further in vitro studies. Evaluation of biocontrol agents for in vitro suppression of R. solani showed that Trichoderma harzianum completely overgrown the pathogen with maximum inhibition of 49.56 % compared to Pseudomonas fluorescens (28.30 %). Under in vitro evaluation of chemical activator, different concentrations of Acibenzolar-S- Methyl (ASM) against pathogen, 100 ppm concentration recorded the maximum mycelial inhibition of 75.67 % and 5 ppm concentration recorded the minimum mycelial inhibition of 27.70 %. Among indigenous organic formulations, turmeric powder and baking soda combination inhibited the maximum growth of the pathogen by 64.40 %. In the in vitro studies with new generation fungicides,mancozeb in cow dung supernatant (0.4 %) and tebuconazole (0.1 %) recorded the 100 % mycelial inhibition of the pathogen. Field studies on disease suppression and plant growth promotion was carried out as two experiments, one in soil solarized plots and the other in non solarized plots. Soil solarization along with soil application of ASM (75 ppm) and foliar application of ASM (100 ppm) recorded the lowest disease incidence of 30.41 % and 30.42 % respectively, which was superior when compared with foliar application of ASM (100 ppm) and soil application of ASM (75 ppm) with the disease incidence of 37.06 % and 38.84 %. Soil solarization + foliar spray of tebuconazole (0.1 %) recorded the minimum disease index of 37.85 % which was superior compared to foliar spray of tebuconazole (0.1 %) with the disease index of 39.28 %. Among the biocontrol agents soil solarization + foliar spray of Pseudomonas fluorescens (2 %) gave minimum disease index of 45.22 % which was greater compared to foliar spray of P. fluorescens (2 %) with the disease index of 51.66 %. In case of indigenous organic formulations, soil solarization + foliar spray of fish amino acid (5 %) given the maximum control of the disease with the disease index of 49.51 % which was superior to foliar spray of fish amino acid (5 %) with disease index of 63.59 %. The number of days taken for flowering in soil solarized plots ranged from 28.67 to 35 days where as the number of days taken for the flowering of amaranthus in non solarized plots was ranged from 27.27 to 31.67 days. At the time of harvest, soil solarization + mancozeb in cow dung supernatant (0.4 %) recorded maximum plant height of 127.07 cm which was higher compared to foliar spray of azoxystrobin (0.15 %) with plant height of 117.60 cm. Maximum of 78.00 number of leaves were recorded by soil solarization + foliar spray of azoxystrobin (0.15 %) which was greater compared to foliar spray of azoxystrobin (0.15 %) with 67.67 number of leaves.Soil solarization + foliar spray of azoxystrobin (0.15 %) gave the highest yield in terms of fresh weight by 26975.00 kg/ha and dry weight of 4233.33 kg/ha which was superior when compared with foliar spray of tebuconazole (0.1 %) with the fresh weight of 23375.00 kg/ha and dry weight of 3362.50 kg/ha. It is concluded that soil solarization for 31 days with the foliar application of tebuconazole (0.1%) can effectively control the Rhizoctonia leaf blight disease severity with plant growth and yield promotion under field conditions.ThesisItem Open Access Characterization of biointensive management of fungal fruit rots of cucurbits(Department of Plant Pathology, College of Horticulture, Vellanikkara, 2019) Muhammad Suhaib, Ismayil M; KAU; Yamini Varma, C KCucurbits are the most popular and widely cultivated vegetables in Kerala. One of the main constraints in the production of these crops is the occurrence of fungal fruit rots, on which no detailed and systematic studies have been conducted in Kerala. The study was carried out during 2017-2019 at College of Agriculture, Padannakkad with the objective to identify and characterize the fungal pathogens causing fruit rots in cucurbits, occurring in the northern zone of Kerala and to study the management of selected fruit rot diseases under in vitro and most severe and predominant disease in vivo conditions. Purposive sampling surveys were conducted for the occurrence of fungal fruit rots in cucurbits in Kasargod, Kannur and Kozhikkode districts. Diseased plant samples were collected. Results of survey showed prevalence of seven different fungal diseases with a range of 6- 51 per cent disease incidence. Isolations done from the infected fruit samples collected during the survey yielded seven genera of fungal pathogens. Characterization of the selected pathogens were carried out based on the cultural and morphological characters and identified up to generic level. Further identification of species of each genus was done by molecular characterization by sequencing the ITS region of each fungus by in silico analysis and confirmed as Choanephora cucurbitarum, Sclerotium rolfsii, Rhizoctonia solani, Phytophthora nicotianae, Pythium deliense, Corynespora cassiicola and Fusarium equiseti. Symptomatologies of these fungal diseases were studied in detail both under natural and artificial conditions. In vitro evaluation of fungicides, botanicals and biocontrol agents was done against the selected five pathogens. Four fungicides and three botanicals at three concentrations and five biocontrol agents were selected for the studies. In vitro studies showed that, against Choanephora cucurbitarum fruit rot, tebuconazole 5EC showed 100 per cent inhibition at all the three concentrations. Among botanicals, garlic extract at medium and higher concentrations and azadirachtin 0.1% at lower concentration were the best. Among biocontrol agents, Trichoderma viride was the best. Against Sclerotium rolfsii fruit rot, tebuconzole 5EC (Folicur), were significantly superior than all other fungicides at lower concentration, which showed 100 per cent inhibition. The fungicides tebuconzole 5EC and mancozeb 64% + cymoxanil 8% were superior at recommended and higher concentration and both of them showed cent per cent inhibition. Among botanicals, garlic extract and azadirachtin 0.1% were the most effective at the lower and recommended concentrations. At higher concentration garlic extract was the best. Among biocontrol agents, Bacillus subtilis and PGPM were the most effective treatments and they were at par with each other. Against Rhizoctonia solani fruit rot, all the chemical fugicides recorded cent per cent inhibition at all concentrations. Garlic extract was the best at all concentrations. The best biocontrol agets against Rhizoctonia solani was Trichoderma viride. Against Phytophthora nicotianae fruit rot, all the fungicides at all concentrations showed cent per cent inhibition. The data on the botanicals revealed that, garlic extract shows maximum inhibition (66.66%, 77.77%, 100%) at all concentrations. Among biocontrol agents, Trichoderma viridae and PGPM were the best treatments and they showed cent per cent inhibition. Against Corynespora cassiicola fruit rot, mancozeb 64% + cymoxanil 8% were superior (100% inhibition) at the lower concentration. At recommended concentrations, mancozeb 64% + cymoxanil 8% and mancozeb 64% + carbendazim 12% showed cent per cent inhibition. At higher concentrations, tebuconzole 5EC, mancozeb 64% + cymoxanil 8% and mancozeb 63% + carbendazim 12% were showed 100 per cent inhibition and they were significantly not different. Among the botanicals garlic extract was the most effective at all concentrations. The best biocontrol agent against Corynespora cassiicola was PGPM, which showed cent per cent inhibition. In vivo evaluation of management of Choanephora cucurbitarum fruit rot in pumpkin emphasized that tebuconazole 5 EC and mancozeb 64%+cymoxanil 8% are the best treatments. Botanicals viz., azadirachtin 0.1 % and garlic extract came as next best treatments and biocontrol agents, T. viride and B.subtilis were the least effective treatments. These can be recommended as effective antipathogenic agents for field application for the management of the fungal fruit rot disease in pumpkin. The present work resulted in a detailed and systematic study on the fungal fruit rots of cucurbits in northern zone of Kerala. Future line work should be concentrated on the field level studies of all the fruit rots of cucurbits in different districts of Kerala and on the recommendations of local specific management practices.ThesisItem Open Access Ecofriendly Management of Major Fungal Foliar diseases affecting yard long bean in Polyhouse(Department of Plant Pathology, College of Agriculture, Vellayani, 2019) Deepthi Nair, S; KAU; Sajeena, AThe study entitled ‘Ecofriendly management of major fungal foliar diseases affecting yard long bean in polyhouse’ was conducted at College of Agriculture, Vellayani and Integrated Farming System Research Station (IFSRS), Karamana during 2017- 2019 with the objectives to determine the major fungal foliar diseases affecting yard long bean grown in polyhouse; and their management using natural resources and ecofriendly methods. A survey was conducted in 15 polyhouses located at different parts of Thiruvananthapuram district, where yard long bean was cultivated. NS 621 was the most common variety of yard long bean cultivated in polyhouses. Powdery mildew (Disease Incidence (DI) - 100% and Per Cent Disease Index (PDI) - 60.70) and Diaporthe leaf spot (DI - 80% and PDI - 45.77) were the major fungal foliar diseases of the crop observed in these polyhouses. Five varieties of yard long bean viz. Lola, Vellayani Jyothika, Gitika (KAU Varieties), VS 50 (KAU pre release culture) and NS 621 (Namdhari Seeds Private Limited) were screened in the polyhouse at IFSRS, Karamana in completely randomized design (CRD) with four replications, to assess the natural incidence and severity of different fungal foliar diseases. The most susceptible variety to natural incidence of powdery mildew (PDI - 50.89) and Diaporthe leaf spot (PDI - 58.66) diseases was NS 621, whereas VS 50 was tolerant to powdery mildew disease (PDI - 11.11). In vitro evaluation of organic preparations viz, egg-lemon juice extract (10%) and sunflower oil (25 ml) + baking soda (10 g) + bar soap (10 g) + water (5 l), botanicals viz. fermented weed (Setaria barbata) extract (10%) and nimbecidine (0.5%), and non-hazardous compounds viz. potassium silicate (1%) and sodium bicarbonate (0.5%) against Diaporthe tulliensis by poisoned food technique in potato dextrose agar (PDA) medium revealed that fermented extract of S. barbata (10%), sodium bicarbonate (0.5%) and a combination of egg-lemon juice extract (10%) and fermented extract of S. barbata (10%) resulted in cent per cent inhibition of the mycelial growth of the fungus, which were further used for in vivo evaluation. Fermented egg-lemon juice extract (10%), potassium silicate (1%) as well as Psuedomonas fluorescens (KAU isolate) had no inhibitory effect on the mycelial growth of the fungus. A trial was conducted to assess the efficacy of selected treatments for the management of powdery mildew and Diaporthe leaf spot diseases in NS 621 in the polyhouse of IFSRS, Karamana in CRD with eight treatments replicated thrice. The plants sprayed with tebuconazole 50% + trifloxystrobin 25% (WG) (0.04%) (positive control) at 20, 40 and 60 days after planting (DAP) resulted in maximum control of Diaporthe leaf spot (PDI - 4.89) and powdery mildew (PDI - 1.22) diseases. Among the treatments, the combined application of arbuscular mycorrhizal fungi (AMF) (KAU isolate) @ 5g plant-1 at the time of planting along with the foliar application of fermented extract of S. barbata (10%) at 20, 40 and 60 DAP resulted in significant control of Diaporthe leaf spot (PDI - 11.35), whereas the combined application of AMF @ 5g plant-1 at the time of planting along with the foliar application of sodium bicarbonate (0.5%) at 20, 40 and 60 DAP resulted in significant suppression of powdery mildew disease (PDI -11.59). Soil application of AMF alone and foliar application of P. fluorescens (2%) had least effect in reducing the severity of both the diseases. Thus, the present study revealed that powdery mildew and Diaporthe leaf spot were the most important diseases of yard long bean in polyhouses of Thiruvananthapuram district. Soil application of AMF @ 5g plant-1 at the time of planting and foliar spray of fermented extract of S. barbata (10%) or sodium bicarbonate (0.5%) at 20, 40 and 60 DAP were the most effective treatments against Diaporthe leaf spot and powdery mildew diseases respectively, which could be used as a green technology to produce safe-to-eat yard long bean.ThesisItem Open Access Seed Biopriming and Spraying at fruit set for the management of Chilli Anthracnose caused by Colletotrichum capsicii (Sydow) Butler and Bisby(Department of Plant Pathology, College of Agriculture, Vellayani, 2019) Athira, P V; KAU; Radhkrishnan, N VThe study entitled ‘Seed biopriming and spraying at fruit set for the management of chilli anthracnose caused by Colletotrichum capsici (Sydow.) Butler and Bisby’ was conducted at College of Agriculture, Vellayani and Coconut Research Station (CRS), Balaramapuram during 2017- 2019 with the objective to study the seed borne nature of chilli anthracnose/fruit rot caused by Colletotrichum capsici and the effect of seed biopriming and spraying during fruit set for its management. Samples of chilli seeds were collected from five agro-ecological zones of Kerala viz. Northern, High range, Central, Special problem and Southern zones. The lowest percentage of infection was found in seed samples collected from Vellayani (6.62) and the highest in Parassala (43.33). Both externally and internally seed borne fungi and bacteria were isolated from the infected seeds. Fruit rot samples were collected from five locations of Kerala viz. Vellayani, Kumarakom, Thrissur, Wayanad and Kasargod. Five isolates (C1 to C5) of C. capsici were obtained. Vellyani isolate (C1) was the most virulent isolate which recorded a lesion size of 3.81 cm in detached fruit assay at seven days after inoculation (DAI). The isolate C1 was used for further studies. Six biocontrol agents viz. Trichoderma viride (KAU isolate), Pseudomonas fluorescens PN 026 (KAU isolate), Bacillus subtilis VLY 62 Bacillus pumilus VLY 17, Bacillus amyloliquefaciens VLY 24 and Piriformospora indica were screened in vitro against C. capsici using dual culture method. B. amyloliquefaciens recorded the highest percentage suppression of mycelial growth of C. capsici (62.96) followed by B. subtilis (56.30) and T. viride (51.85). P. fluorescens did not show the mycelial growth suppression of C. capsici. In vitro assays on chilli fruits revealed that B. amyloliquefaciens was most effective in controlling fruit rot with a lesion size of 2.71 cm 10 DAI followed by B. subtilis (lesion size - 2.97 cm) and T. viride (lesion size – 3.81 cm). Peroxidase and polyphenol oxidase activities were also higher in B. amyloliquefaceins treated chilli fruits before and after treatment. Standardization of priming techniques revealed that soaking of seeds for 1 h was found effective for T. viride and 4 h for other biocontrol agents. The effective soaking durations were followed for the respective treatments in further in vivo studies. Based on the in vitro assays, the best three treatments viz., B. amyloliquefaciens, B. subtilis and T. viride were taken for in vivo studies. In vivo evaluation of seed biopriming on the control of fruit rot revealed that the lowest disease severity was recorded in B. subtilis (PDI – 37.19) followed by B. amyloliquefaciens and T. viride (PDI in both - 40.00). The severity of anthracnose in leaves was lowest in B. subtilis (PDI – 10.60) which was on par with T. viride (PDI – 11.30) and B. amyloliquefaciens (PDI – 11.90). The number of fruits per plant, 100 seed weight and seed yield per plant were higher in T. viride. In vivo studies on the effect of seed biopriming and spraying during fruit set recorded the lowest disease severity for B. amyloliquefaciens (PDI – 28.13 at 15 DAI) which was on par with carbendazim 0.1 per cent (PDI - 29.38) and was followed by B. subtilis (PDI - 31.25) and T. viride (PDI - 32.5). The severity of anthracnose in leaves was lowest in B. subtilis (PDI - 8.00) which was on par with B. amyloliquefaciens (PDI - 8.10) and T. viride (PDI - 8.50). The number of fruits per plant, 100 seed weight and seed yield per plant were highest in T. viride treated plants. Thus, the present study indicated that seed biopriming and spraying with B. amyloliquefaciens VLY 24 at fruit set was found to be the most effective for the management of fruit rot / anthracnose of chilli which could be used as an ecofriendly measure to produce safe-to-eat crop.ThesisItem Open Access Characterization of different viruses infecting small Cardamom (Elettaria cardamomum Maton) and production of disease free plants(Department of Plant Pathology, College of Agriculture, Vellayani, 2019) Vangala Bhavana,; KAU; Radhika, N SThe study entitled “Characterization of viruses infecting small cardamom (Elettaria cardamomum Maton) and production of disease free plants” was conducted at Department of Plant Pathology, College of Agriculture, Vellayani during 2017-2019, with the objective to study the occurrence and distribution of viruses infecting small cardamom, molecular characterization of the viruses and elimination of viruses through meristem culture for the production of disease free planting material. Survey was conducted in Kattappana, Pampadumpara and Ambalapara panchayats of Kattappana block; and Nedumkandam, Thukkupalam and Chembalam panchayats of Nedumkandam blocks of Idukki district during November 2018 – May 2019. The incidence of katte disease caused by Cardamom mosaic virus (CdMV) was present in all the panchayats surveyed and it ranged from 3.75 to 43.0 per cent in Kattappana block and 5.0 to 31.33 per cent in Nedumkandam block. Disease incidence for chlorotic streak caused by Banana bract mosaic virus (BBrMV) was recorded from Kattappana (41%), Pampadumpara (30%) and Nedumkandam (8.33%) panchayats. The aphids infestation was absent in all the surveyed plots. Colocasia spp. and Alpinia spp. were the major plants observed in and around the cardamom fields and were not having visible symptoms of the viral infections. The virus inoculums were maintained under insect proof net house at Cardamom Research Station, Pampadumpara. Katte disease produced slender chlorotic flecks developing into pale green discontinuous stripes running parallel to veins from midrib to leaf margin of the infected leaves. Mosaic mottling and chlorotic specks were seen on the infected leaves and young pseudostems. In case of severe infection, plants produced stunted tillers. Chlorotic streak disease was characterised by continuous and discontinuous chlorotic streaks along veins and midribs of the infected leaves and green discontinuous spindle streaks on pseudostem. CdMV (a potyvirus) and BBrMV in cardamom was detected using polyclonal antibodies of Potato virus Y (PVY) and BBrMV respectively procured from DSMZ, Germany by direct antigen coating- Enzyme linked immunosorbent assay (DAC-ELISA) and Dot immunobinding assay (DIBA).The highest virus titre of CdMV and BBrMV was obtained in samples collected from Pampadumpara and Kattappana respectively. Molecular detection of the viruses was carried out using reverse transcriptase - polymerase chain reaction (RT-PCR) with specific primers for CdMV and BBrMV; and obtained amplicons of expected size of 879-905 bp for CdMV- and 625-633 bp for BBrMV- infected samples. The sequences of the isolates of CdMV from Kattappana, Pampadumpara and Nedumkandam were subjected to BLAST analysis and found to be similar to Indian cardamom mosaic virus isolates from Thalathamane and Appangala with > 96 per cent similarity. The BBrMV in cardamom from Kattappana, Pampadumpara and Nedumkandam was similar to Banana bract mosaic virus (BBrMV) CdM isolate of Karnataka (91.01%), Coimbatore (90.29%) and Thrissur (95.76%) respectively. Phylogeny tree constructed in MEGA 6.0 software differentiated CdMV and BBrMV into four clades, in which CdMV Kattappana and Nedumkandam isolates were clustered together whereas CdMV Pampadumpara isolate was in separate clade. Similarly, BBrMV isolates of Pampadumpara and Nedumkandam clustered together while BBrMV Kattappana was in separate clade. Meristem of 2 mm size separated from infected plants were grown in Murashige and Skoog medium supplemented with 3 mg benzyl amino purine (BAP), 1.5 mg indole acetic acid (IAA) and 0.8 mg kinetin expressed direct organogenesis but multiple shoots were not produced. The TC plants were subjected to DAC-ELISA with the specific polyclonal antibodies and PCR with specific primers of the viruses and confirmed that the plants produced from meristems were free of both the viruses. Thus, the present study revealed that two viral diseases viz., katte and chlorotic streak affecting small cardamom in Idukki. Serologically and molecularly it was detected that katte disease was caused by Cardamom mosaic virus (CdMV) and chlorotic streak disease was caused by Banana bract mosaic virus (BBrMV), and the viruses could be eliminated from the infected plants through meristem tip culture to produce the diseases free plants.