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  • ThesisItemOpen Access
    DEVELOPMENT AND EVALUATION OF MANALI TAMARIND (Pithecellobium dulce) BASED FUNCTIONAL FOODS
    (PROFFESSOR JAYASHANKAR TELANGANA STATE AGRICULTURAL UNIVERSITY, 2023-01-04) Rachana Sree, S; Supraja, T.
    Underutilized horticultural crops were those that are not sold widely or farmed on a significant scale for commercial purposes. Local people grow, trade and consume these products. These crops are easier to grow, even in unfavourable climatic and soil circumstances. Therefore, taking advantage of underutilized horticulture crops can help to address the socioeconomic issues of poor health, nutrition, unemployment, and poverty. By satisfying the dietary needs of vulnerable individuals, the consumption of underused fruit crops provides nutrition to the poor and destitute tribal people. They are a rich source of carbohydrates, lipids, proteins, energy, vitamins A, B1, B2, B3, B6, B9, B12, C, folic acid, minerals, Ca, P, Fe and dietary fibre. Manali tamarind was procured from different places like Nalgonda, Jagityal, Hyderabad and Bhadrachalam. Raw material was cleaned thoroughly, tray-dried at 60°C for 5 ½ hours and stored in plastic containers for further use. The fresh and dried aril was analysed and the dried aril (DA) proximate, mineral content was increased due to the increased concentrations of the nutrients when subjected to the drying process compared to FA. The vitamin C and aw content of fresh aril (FA) was high due to their heat and light-sensitive nature compared to DA. The intensity of the colour increased for DA due to drying temperature and time. Functional parameters like WAI, WSI, ORC and FC were good for DA powder due to its high solubility because of the uniform particle size and shape. Ready-to-use products (chutney powder, beverage powder, pickle 10% to 70% incorporation) and ready-to-eat products like (curd, yoghurt and ice cream 5% to 35% incorporation) were developed. The best formulation from each product was selected by using sensory evaluation as follows chutney powder with 40% (SCP4), beverage powder with 30% (SBP3), ice cream with 10% dried aril powder (SIC2), pickle with 30% (SP3), curd with 5% (SC1) and yoghurt with 10% incorporation of fresh aril paste (SYH2) was further subjected to proximate, physical and functional analysis. All the selected products were low in CHO and energy compared to the control. The total sugars, reducing sugars and non-reducing sugars were higher for samples than that of controls due to the incorporation of aril powder and paste. The physical parameters like TSS, TA and TI of the samples increased due to the aril powder present in them. The most accepted product during the sensory evaluation was pickle followed by chutney powder. The functional properties of sample pickles were high compared to the control due to the aril incorporation where WAI was 1.82±0.04ml/g, WSI was 28.73±8.86%, ORC was 3.51±0.07ml/g and FC was 1.79±0.00%. The moisture, protein, total sugars, reducing sugars and non-reducing sugars content of the sample pickle was high compared to the control which was 35.9±81.05%, 3.82±0.01%, 15.99±0.06%, 4.10±0.06% and 11.89±0.03%. The shelf life of the selected products based on TMC and TBC revealed that chutney powder (40%), beverage powder (30%) and pickle (30%) performed well during the entire study period of 90 days. Whereas, for sample ice cream (10%) it was 45 days, curd (5%) and yoghurt (10%) performed well for 30 days. All the storage parameters like pH, TSS, TA and moisture content showed variation during the storage period. All the products were subjected to sensory evaluation during the 15-day interval period and all the products were high to moderately liked by the panellists. Consumer evaluation was conducted with 100 respondents of the age group 15- 65 years, all the selected products scored well and the acceptability of yoghurt was high compared to the other products. The respondents were ready to use them in their day-to day lives and can afford up to 30/- to 50/- for the 100g based on the variability of products.
  • ThesisItemOpen Access
    VARIABILITY AND FUNGICIDE MANAGEMENT OF CHILLI ANTHRACNOSE AND FRUIT ROT DISEASE CAUSED BY Colletotrichum spp.
    (PROFFESSOR JAYASHANKAR TELANGANA STATE AGRICULTURAL UNIVERSITY, 2022-07-02) Mounika, M.; Vidya Sagar, B.
    Chilli (Capsicum annuum L.) is one of the most economically important vegetable crops in the world. It is affected by various biotic and abiotic constraints. Among different biotic constraints, anthracnose disease caused by various species of Colletotrichum is the major limiting factor affecting yield and production of chilli crop. Symptoms of the disease include fruit rot, leaf spots, dieback on stem, seedling blight, or damping off. Severity of the disease varies with the strain of the pathogen responsible for the disease. Hence, accurate identification of pathogen up to species level is needed for choosing the appropriate management strategy for controlling this disease. Keeping these points in view, the present investigation on “variability and fungicide management of chilli anthracnose and fruit rot disease caused by Colletotrichum spp.” was carried out with the objectives to survey for the occurrence of anthracnose and fruit rot disease incidence in major chilli growing districts of Telangana, to study the variability among the isolates of Colletotrichum spp. with respect to cultural, morphological and molecular characteristics and to study the efficacy of fungicides against Colletotrichum spp. in vitro and glass house conditions. A roving survey was conducted in major chilli growing districts of Telangana i.e., Khammam, Warangal and Mahabubabad during Rabi 2020 and ten isolates of Colletotrichum spp. were collected. Highest per cent incidence of anthracnose and fruit rot disease of 34.20 was recorded in Vasnari village of Khammam district. While, lower per cent incidence of 09.85 was recorded in Manchu Thanda village of Mahabubabad. Commonly grown varieties were ‘Teja’ and its derivatives which were severely affected by anthracnose and fruit rot disease. Pathogenicity test was conducted for all the ten isolates of Colletotrichum spp. on chilli fruits by pin prick method. All the isolates produced small brown water-soaked lesion which gradually enlarged and covered the fruit surface within 48hrs with isolate C3 being the most virulent producing early and severe symptoms and was selected for further studies of management. The variability among the isolates of Colletotrichum spp. was studied with respect to their morphological, cultural and molecular characters among the isolates. The isolates showed variation in colour when tested on different media and this change was observed within the media and also among the isolates. The colour varied from white, gray to black. The isolates showed a varied colony texture from fluffy to scanty mycelium on different media tested. There was no significant difference among the isolates tested for radial growth within the media and the significant difference was observed between the media when tested for radial growth. Among five media tested, PDA supported the maximum growth While, minimum growth was recorded in Corn Meal Agar medium. Most of the isolates produced distinct concentric zonation’s except isolate C2 and C8 in which zonation’s were absent. Sporulation was more on Host Extract Agar medium. Least sporulation was found in Corn Mean Agar medium supported. Acervuli were disc shaped, waxy, sub epidermal, typically with dark needle like septate setae. The average number of acervuli for 5 mm disc was 25. All the isolates produced dark brown setae which were smooth, septate, rigid, swollen at the base and tapered towards the acute apex. Length of the setae of test isolates ranged from 57.30 × 05.07 µm to 61.00 × 5.83 µm. Influence of temperature on the radial growth of Colletotrichum spp. was studied at different temperatures. Highest growth of all the isolates was recorded at 30oC. While, no growth was observed at 10oC. Influence of pH on radial growth, wet and dry mycelial weight of Colletotrichum spp. was studied and it was found that the maximum radial growth, wet and dry mycelial weight of all the isolates was recorded at pH 7.0. While the lowest was recorded at pH 8.0. Molecular sequencing of all the isolates of Colletotrichum spp. revealed that out of ten isolates, seven isolates were Colletotrichum truncatum and three isolates were identified as Colletotrichum capsici. Further variability using RAPD markers revealed that there was no relationship between the geographical origin and molecular clustering among the isolates. Poisoned technique was conducted using thirteen fungicides with Captan and Carbendazim as standard fungicide control. The results showed all the fungicides were effective in inhibiting radial growth of Colletotrichum truncatum. Among the six systemic fungicides tested, Propiconazole was found to be most effective with maximum inhibition at all the concentrations tested which was on par with Carbendazim followed by Difenoconazole. Least inhibition was shown by Tebuconazole. Among the five combination fungicides tested, Pyraclostrobin + Metiram was found to be effective followed by Azoxystrobin + Tebuconazole. Least inhibition was shown by Picoxystrobin + Tricyclazole. Among the four non-systemic fungicides tested, Captan was found to be most effective followed by Copper Hydroxide. Lowest inhibition was shown by Copper Oxychloride. The fungicides which were effective under in vitro were further evaluated for their efficacy under glass house conditions. Seeds of the susceptible cultivar ‘sonal’ were grown under glass house conditions. Out of all the treatments tested, Propiconazole was found to be effective in reducing the per cent reduction of the disease over control followed by Difenoconazole. While the least reduction was shown by Copper Hydroxide.
  • ThesisItemOpen Access
    STUDIES ON VARIABILITY AND INTEGRATED DISEASE MANAGEMENT OF WILT OF SAFFLOWER (Carthamus tinctorius L.) CAUSED BY Fusarium oxysporum f. sp. carthami
    (PROFFESSOR JAYASHANKAR TELANGANA STATE AGRICULTURAL UNIVERSITY, 2022-07-02) Rajendra prasad, M.; Vidya Sagar, B.
    Safflower (Carthamus tinctorius L.) is one of the major Rabi oilseed crop cultivated in the Deccan Plateau region of India. Wilt of safflower caused by Fusarium oxysporum f. sp. carthami Klisiewicz and Houston (Foc) results in yield loss up to 93 per cent in susceptible varieties. Wilt samples of safflower were collected from different locations of Telangana and Andhra Pradesh and fifteen isolates of Fusarium oxysporum f. sp. carthami were isolated from the collected diseased samples and they designated as FOC 1 to FOC 15. Morphological variations among collected isolates were studied. Based on the observed length of microconidia the isolates could be divided into 5 distinct groups i.e. Group I to V, based on macroconidia length the isolates could be divided into 3 distinct groups i.e. Group I to III. Based on chlamydospore diameter, the isolates could be divided into four distinct groups (Group I to IV), based on the cultural characters they were categorized into eight distinct groups (Group I to VIII) and based on colony diameter the isolates grouped into seven distinct groups (Group I to Group VII). The variation was present among the isolates at molecular level and the isolates were grouped into different clusters. The results of molecular variability revealed that There was a relationship exists between clustering of isolates based on molecular variability with the location. Pathogenic variability of the fifteen isolates of Fusarium oxysporum f. sp. carthami was tested using susceptible safflower cultivar PBNS-12 under high inoculum pressure in glasshouse conditions. Wilt incidence was recorded in the range of 53.3 % (FOC-11) to 93.3 % (FOC-I). Forty one germplasm lines were collected from ARS, Tandur and were screened against Fusarium oxysporum f. sp. carthami. Based on the disease scale (0-9), they were categorized from highly resistant to highly susceptible. Seed mycoflora detected from the seeds of 41 germplasm lines by using standard blotter method and agar plate methods. In both the methods, eight fungal genera viz., Alternaria spp., Aspergillus flavus, Aspergillus niger, Chaetomium spp., Rhizopus spp., Curvularia spp., Macrophomina phaseolina and Fusarium spp. were recorded. Among the observed fungi A. niger was found predominant (40.28 % and 42.52 %), while M. phaseolina was least (2.39 % and 2.02 %). Effect of fungicides and biocontrol agents on seed germination, seedling vigour index and seed mycoflora was evaluated on four popular verities of safflower viz., Nira, ISF-764, PBNS-12 and Manjira by standard blotter method and paper towel method. Seed treatment with captan (0.25 %) was found superior followed by seed treatment with Trichoderma harzianum (1 %), in all the four popular varieties tested in the testes parameters. Susceptible variety PBNS-12 was used to find the effect of seed treatment with bioagents, organic amendments and carbendazim and their combinations on per cent seedling emergence, pre emergence and post emergence mortality in glass house conditions. Among all the treatments tested, T11-combination treatment of (T1 + T2 + T3 + T4 (seed inoculation with test pathogen followed by seed treatment with Trichoderma harzianum (10 g kg-1 seed) + T2 (seed treatment with Pseudomonas fluorescens (10 g kg 1 seed) + T3 (seed treatment with carbendazim (1 g kg-1 seed) + T4 (soil application with neem cake (10 g kg-1 soil) was found superior in all the parameters discussed. When tested for growth parameters, shoot length was recorded highest in T11 (59.2 cm), root length was recorded highest in T6 (10 cm) and Total length was recorded highest in T11 (68.2 cm). Highest fresh weight (272.10 g) and dry weight (105.35 g) was also recorded from the same combination treatment. In integrated disease management experiment, when the cost benefit ratio is compared, T16 (Seed treatment with carbendazim) was found effective with C:B ratio of 1:67 followed by T14 Seed treatment with Trichoderma harzianum + Seed treatment with Pseudomonas fluorescens (1:52) and T3 Seed treatment with Trichoderma harzianum (1:50). Among the above treatments, T16 recorded highest yield (1681 kg ha 1 ) highest germination (88.42 %) and lowest wilt incidence (15.12 %) followed by T14 (Seed treatment with Trichoderma harzianum + Seed treatment with Pseudomonas fluorescens) recorded yield of 1580 kg ha-1 , germination of 82.33 per cent and wilt incidence of 22.10 per cent respectively.
  • ThesisItemOpen Access
    ISOLATION AND CHARACTERIZATION OF RHIZOSPHERE MICROFLORA AND DEVELOPMENT OF MICROBIAL CONSORTIUM FOR MANAGEMENT OF STEM ROT DISEASE IN GROUNDNU
    (PROFFESSOR JAYASHANKAR TELANGANA STATE AGRICULTURAL UNIVERSITY, 2022-07-02) Akash, A. U.; Ramya, V.
    Groundnut [Arachis hypogaea L.], is an important edible oilseed crop used extensively for oil extraction, cooking, and domestic purposes. In Telangana state, groundnut is predominantly cultivated as a Rabi crop. Among the various diseases affecting groundnut, stem rot caused by Sclerotium rolfsii Sacc. is a major threat to successful groundnut cultivation and can cause heavy yield losses. The present study was aimed at developing a native microbial consortium using microbes from groundnut rhizosphere soil for the management of stem rot disease in groundnut. Groundnut rhizosphere soil samples and infected plant samples were collected from twelve different fields in ten villages covering six mandals of Nagarkurnool district of Telangana state during Rabi 2019-20. Rhizosphere soil microflora and test pathogen S. rolfsii were isolated. Seven isolates of test pathogen S. rolfsii were studied for variability in colony morphology and sclerotial morphology. All the isolates recorded cent per cent disease incidence in pathogenicity tests conducted using groundnut variety Kadiri-6. Isolate GNS1 was found to be the most virulent among seven tested isolates and hence was used for further studies. A total of 190 rhizosphere isolates (111 from King’s B medium, nutrient agar medium and actinomycetes isolation agar medium; 79 from potato dextrose agar medium and Martin’s Rose Bengal agar medium) isolated from rhizosphere soil samples were evaluated for their cultural, morphological and biochemical characteristics. Gram-positive organisms dominated the groundnut rhizosphere soil. Among the fungal isolates, Aspergillus spp. (majorly A. niger and A. flavus) dominated the groundnut rhizosphere soil. Preliminary screening of these isolates for antagonistic activity against S. rolfsii, identified 33 potential rhizobacterial isolates and 9 fungal isolates. Further screening of these isolates using dual culture assay revealed eight potential biocontrol isolates including seven bacterial isolates viz. S1NA7, S2NA6, S3KB6, S4KB5, S8KB2, S9KB4 xvi and S10KB2 and 1 fungal isolate viz. Tricho5, all of which were selected for further studies. Molecular characterization of test pathogen isolate GNS1 identified it as Athelia rolfsii. Bacterial biocontrol isolate S9KB4 was identified as Bacillus amyloliquefaciens and the remaining six bacterial biocontrol isolates were identified as Bacillus subtilis. Fungal biocontrol isolate Tricho5 was identified as Trichoderma asperellum. Compatibility among these eight biocontrol isolates was tested and isolates were screened for plant growth-promoting biochemical traits. Among the bacterial biocontrol isolates, six isolates were compatible with each other except B. subtilis isolate S1NA7 which was compatible with fungal biocontrol isolate Tricho5. Three microbial consortia were developed by combining compatible isolates considering antagonism against S. rolfsii as primary factor viz., RMC1 (B. subtilis isolate S2NA6 + B. subtilis isolate S3KB6 + B. subtilis isolate S4KB5), RMC2 (B. subtilis isolate S8KB2 + B. amyloliquefaciens isolate S9KB4) and RMC3 (B. subtilis isolate S1NA7 + T. asperellum isolate Tricho5). Further, these consortia were tested for plant growth promotion and efficacy in managing stem rot disease of groundnut under glasshouse conditions. Seed treatment with microbial consortium RMC2 (B. subtilis isolate S8KB2 + B. amyloliquefaciens isolate S9KB4) recorded the highest disease control in glasshouse studies while seed treatment with RMC3 (B. subtilis isolate S1NA7 + T. asperellum isolate Tricho5) resulted in highest plant growth promotion. The study demonstrated the efficiency of the consortia in both plant growth promotion and managing stem rot disease in groundnut compared to seed treatment with individual biocontrol isolates and chemical control. Future research on the efficiency of these native microbial consortia under field conditions can help in developing an efficient and sustainable management for stem rot disease in groundnut.
  • ThesisItemOpen Access
    GREEN SYNTHESIS OF NANOPARTICLES FOR THE MANAGEMENT OF SEED BORNE FUNGAL PATHOGENS OF SOYBEAN (Glycine max L.)
    (PROFFESSOR JAYASHANKAR TELANGANA STATE AGRICULTURAL UNIVERSITY, 2022-07-02) Shaik Reshma; Bharathi, V.
    Soybean (Glycine max L.), popularly known as the “Wonder Crop” and “Golden bean” is one of the globally important cultivated legumes. It is widely cultivated for food, feed and industrial purposes throughout the world. In India, soybean is the second largest cultivated oilseed crop after groundnut. As soybean cultivation has spread globally, diseases have become more prevalent and severe. The majority of important soybean fungal diseases are seed-borne and have emerged as one of the most critical yield-limiting factors in global soybean production. Hence, maintaining seed quality by lowering the rate of seed deterioration is a top priority. The current research focuses on isolating and identifying seed-borne fungal pathogens associated with soybean, as well as conducting in vitro studies to assess the effectiveness of green and chemically synthesized ZnO and MgO nanoparticles in limiting the growth of major seed-borne fungal pathogens associated with soybean. Further, the effect of seed treatment with nanoparticles was evaluated on seed health and quality parameters of soybean against major seed-borne fungal pathogens by following standard ISTA procedures. In the present study, a total of 62 soybean seed samples were collected from different locations of Adilabad, Nizamabad, Kamareddy and Jagtial districts of Telangana state during Kharif 2020. The soybean seed samples collected were subjected to standard blotter and agar plate methods for detection of seed borne mycoflora, while seed quality parameters were evaluated using rolled paper towel method. A total of eight fungal species belonging to six genera viz., Fusarium oxysporum, Fusarium semitectum, Macrophomina phaseolina, Colletotrichum truncatum, Curvularia spp., Aspergillus niger, Aspergillus flavus and Penicillium spp. were assessed by using both the methods of isolation, and the pathogenicity for the major seed borne fungal pathogens was also proved under glass house conditions. The antifungal activity of neem-based green synthesized ZnO and MgO nanoparticles synthesized using leaf extracts of neem (Azadirachta indica) and chemically synthesized ZnO and MgO nanoparticles at different concentrations was evaluated for mycelial growth inhibition against four major seed borne fungal pathogens of soybean (F. oxysporum, F. semitectum, M. phaseolina and C. truncatum), for their mycelial growth inhibition by employing both agar well and disc diffusion methods. Green synthesized ZnO and MgO nanoparticles have shown highly significant inhibition when compared to chemically synthesized ZnO and MgO nanoparticles in a dose dependent manner. Among the methods tested for antifungal assay (agar well and disc diffusion), the pathogens were more sensitive to agar well method when compared to disc diffusion method under in vitro. It was observed from the results that, F. oxysporum and F.semitectum were more sensitive to neem-MgO nanoparticle at 2000 mg/kg with highest mycelial growth inhibition of 70.10 and 76.47 per cent, respectively, while the fungi M. phaseolina and C. truncatum were more sensitive to neem-ZnO nanoparticle at 2000 mg/kg with highest mycelial growth inhibition of 76.47 and 81.58 per cent, respectively, under in vitro conditions. The antifungal activity of nanoparticles was also evaluated for spore germination against F. oxysporum and highest inhibition in spore germination was resulted with neem-MgO nanoparticle followed by neem-ZnO, MgO and ZnO nanoparticles. Further the antifungal effect of nanoparticles was confirmed by SEM analysis of F. oxysporum treated with 2000 mg/kg of neem-ZnO and neem-MgO nanoparticles. The SEM images demonstrated that the nanoparticles clearly damaged the hyphae of F. oxysporum, exhibiting deformed hyphal walls in treated hyphae when compared to untreated control. Effectiveness of seed treatment (dry and wet seed treatment) with neem-based green and chemically synthesized ZnO and MgO nanoparticles of different concentrations was evaluated against three major seed borne fungal pathogens (F. oxysporum, M. phaseolina and C. truncatum) to determine their effect on seed health and seed quality parameters of soybean using blotter test and rolled paper towel method, respectively. In blotter test, all the tested nanoparticles used for seed treatment reduced the fungal infection significantly in a concentration dependent manner. Neem-MgO NP against F. oxysporum and neem-ZnO NP against M. phaseolina and C. truncatum, showed maximum inhibition at the highest concentration (2000 mg/kg) when compared to chemically synthesized nanoparticles at the same concentrations and over control. From rolled paper towel method, it was observed that, all the tested metal oxide nanoparticles had influential effect on seed quality parameters of soybean against all the tested pathogens and wet seed treatment was found to be more effective as compared to dry seed treatment. Among all the treatments with respect to seed quality parameters, the effect of seed treatment with neem-ZnO nanoparticle was more pronounced followed by neem-MgO nanoparticle when compared to chemically synthesized nanoparticles. All the seed quality parameters significantly increased from lower to higher concentrations with maximum effect at 1500 mg/kg but thereafter decline in seed quality parameters was observed at increased concentration (2000 mg/kg) due to phytotoxicity. Further, the uptake of nanoparticles by soybean seed after seed treatment was confirmed by SEM analysis of ten day old seedlings treated with neem-MgO and neem-ZnO NPs. The findings of this study provide a significant step towards enhancing soybean protection measures, involving green synthesised nanoparticles as one of the most eco friendly components, exhibiting antifungal activity for the effective management of seed borne fungal diseases
  • ThesisItemOpen Access
    ASSESSMENT OF NATIVE ISOLATES OF BIOCONTROL AGENTS FROM THE RHIZOSPHERE OF PIGEON PEA (Cajanus cajan (L.) Millsp.) AGAINST Fusarium udum
    (PROFFESSOR JAYASHANKAR TELANGANA STATE AGRICULTURAL UNIVERSITY, 2022-09-27) Pravalika, Y.; Padmaja, G.
    Pigeonpea (Cajanus cajan (L.) Millsp.) is one of the most important pulse crops of semiarid tropic and sub tropic regions viz., Asia and Africa (Maesen, 1990). It is the second most important crop in the world and Fusarium wilt is the major disease causing yield loss. Among the major biotic stresses the Fusarium wilt, caused by Fusarium udum is the most important soil borne disease of pigeonpea and cause 30-100 per cent loss in grain yield and depends upon the stage at which the plant is affected and can approach over 50 per cent and even up to 100 per cent when wilt occurs at the pre pod stage (Kannaiyan and Nene, 1981; Okiror, 2002). Use of chemicals is one of the widely practiced methods to manage the disease. However, it offers many disadvantages such as emergence of fungicide-resistant populations, disruption of local ecosystems, cost, large labor requirements and threats to human health. Therefore, it is important to assess the potential biocontrol agents for secondary metabolites and their antifungal activity against Fusarium udum to develop effective and environmental friendly methods to control the disease. Total of fourty two rhizosphere soil samples were collected from different pigeon pea growing areas of Telangana state to isolate bio control agents. Data was collected on GPS coordinates, soil type etc. From the rhizosphere, 124 bacterial isolates and 81 fungal isolates were isolated. Pure cultures were maintained by periodical sub – culturing for further use. Diversity of bacterial microflora among all the bacterial isolates were characterized for their cultural and biochemical characteristics. Biochemical characteristics like Methyl red and Voges Proskauer test, Citrate utilization, Oxidase test, Catalase test, Indole test, KOH and Grams staining was performed with the 124 rhizosphere isolates. From the 124 isolates, 49 isolates tested positive for Methyl red test, Author : Y. PRAVALIKA Title of the thesis : “ASSESSMENT OF NATIVE ISOLATES OF BIOCONTROL AGENTS FROM THE RHIZOSPHERE OF PIGEON PEA (Cajanus cajan (L.) Millsp.) AGAINST Fusarium udum Degree : MASTER OF SCIENCE IN AGRICULTURE Faculty : AGRICULTURE Major field : PLANT PATHOLOGY Chairperson : Dr. G. PADMAJA University : PROFESSOR JAYASHANKAR TELANGANA STATE AGRICULTURAL UNIVERSITY Year of submission : 2022 32 isolates tested positive for Voges Proskauer test, 103 isolates tested as positive for Citrate utilization, 86 isolates tested as positive for Oxidase test, 98 tested positive for the Catalase test only 23 isolates tested positive for the Indole test. Of the total 124 isolates tested, 53 isolates were KOH test positive and had negative Gram reaction while 71 tested negativefor the KOH test and gave a positive Gram reaction. All the fungal and bacterial isolates were screened primarily for antagonism against F. udum. Among the ten Trichoderma isolates tested in dual culture, S36RFI-42 recorded the maximum inhibition of 87.15 per cent over control followed by isolate S18RFI-23 i.e., 86.13 per cent. The isolates S24RFI-30, S39RFI-47, S2RFI-2, have shown 84.65, 84.27 and 84.19 per cent respectively. The isolates S39RFI-47, S22RFI-28, and S9RFI-11 have shown 83.77, 83.54 and 82.37 per cent respectively. The minimum inhibition of 65.76 and 69.15 per cent over control was recorded by isolates S32RFI-36 and S12RFI14 respectively by isolate S36RFI-42 i.e., 86.32 per cent. The isolates S24RFI-30, S39RFI-47, S2RFI-2, have shown 84.65, 84.27 and 84.19 per cent respectively. Among the fifteen isolates tested in dual culture, isolates S14RBI-34 and S2RBI 4 recorded the maximum inhibition of 74.05 and 71.80 per cent over control respectively followed by isolates S38RBI-107 (69.26 %), S42RBI-121 (68.03 %), S15RBI-40 (64.83 %), S21RBI-54 (64.77 %), S36RBI-97 (57.15%), S34RBI-91 (53.49 %), S35RBI 95 (47.55%), S39RBI-111(39.71 %), S10RBI-26 (37.27%), S17RBI-37 (30.79 %), S37RBI-104(23.16 %), S28RBI-73(19.17%). The minimum inhibition of 12.80 per cent over control was recorded by isolate S27RBI-69. Eight potential bacterial biocontrol isolates (S2RBI-4, S14RBI-34, S38RBI-107, S42RBI-121, S15RBI-40, S21RBI-54, S36RBI-97, S34RBI-91) were screened for various plant growth-promoting characters including IAA production, Siderophore production, HCN production, Ammonia production, Phosphate solubilization, under invitro conditions. Further potential bacterial isolate S14RBI-34 (Pseudomonas aeruginosa) as well as fungal isolate S36RFI-42 (Trichoderma viride) were used in pot culture studies to induce defense reaction in four pigeon pea genotypes viz., ICP-87119 (Resistant), WRGE-124 (Moderately resistant), WRGE-121 (Moderately resistant) and ICP-2376 (Susceptible). Defense enzymes viz., peroxidases (Pos), poly phenol oxidases (PPO), phenyl alanine ammonia lyase (PAL) activity five different intervals were measured at five different intervals 0, 3, 6, 9 and 12 days after inoculation of pathogen. Per cent disease incidence was measured at 45, 60 days after sowing. Secondary metabolites of these potential isolates were also evaluated against F. udum. Plant parameters including emergence of seedlings (vigour index), length of root, length of shoot was calculated and found an increase in vigour index, length of root and shoot after application of bio control agents. Soil parameters such as pH, Organic carbon per cent, available N, P and K (Kg/ha) and Agronomic parameters: C:N ratio and germination percentage before and after application of bio control agents were calculated and slight improvement in soil and agronomic parameters were found.
  • ThesisItemOpen Access
    DEVELOPMENT OF MICROENCAPSULATION TECHNIQUE OF BIOCONTROL AGENT AGAINST FUSARIUM WILT DISEASE IN BENGALGRAM (CICER ARIETINUM)
    (PROFFESSOR JAYASHANKAR TELANGANA STATE AGRICULTURAL UNIVERSITY, 2022-07-02) Lohitha, B.; Ameer Basha, S.
    Bengalgram (Cicer arietinum L.) is an important legume crop grown in India after dry beans. In Telangana, it is cultivated in an area of 1.30 lakh ha, with a production of 1.99 lakh tonnes and with a productivity of 1532 Kg/ha. The major bengalgram growing districts in Telangana are Adilabad, Kama Reddy, Gadwal, Nirmal and Nizamabad. This crop is affected by various biotic and abiotic stresses. Among biotic stresses fungal diseases are predominantly affecting the crop among which Fusarium wilt caused by Fusarium oxysporum f. sp. ciceris is the major cause for the low productivity of bengalgram in India. During Rabi 2020, a roving survey was conducted to know the wilt incidence caused by F. oxysporum f. sp. ciceris in 19 villages of 3 districts of Telangana i.e., Rangareddy, Siddipet and Nirmal. In which, the maximum (44.25 %) wilt incidence was recorded in Hinapally village of Nirmal district, followed by Pragnapur village (35.32 %) of Siddipet district and minimum (16 %) wilt incidence was recorded at Kistapur village of Ranga Reddy district. A total of seven isolates of F. oxysporum f. sp. ciceris were isolated from different areas surveyed and morphological and cultural variability among them were studied. The rhizospheric bacterial biocontrol agents were isolated from bengalgram fields and morphological characteristics of the isolates were recorded. Out of thirty-nine bacterial biocontrol agents, only four bacterial isolates have shown antagonistic activity against seven isolates of Fusarium by dual culture technique. In this, the maximum (64.72 %) mycelial growth inhibition was reported by isolate BS-9 against virulent isolate (FOC 3) of F. oxysporum f. sp. ciceris, whereas minimum (51.50 %) mycelial inhibition was reported by isolate BS-32 against isolate FOC 7. The spore germination test done against isolates of Fusarium revealed that isolate BS-9 bacterial pellet suspension effectively reduced the germination of spores in isolate FOC 3. The molecular identification of isolate BS-9 was done using 16S rRNA gene sequencing w hich was identified as Bacillus subtilis and it was used in our research work. In preliminary screening, before capsule preparation B. subtilis (BS-9, OM019320.1) was tested against Fusarium wilt disease under glass house conditions. It was observed that, isolate (BS-9, OM019320.1) effectively reduced the wilt incidence Author : B. LOHITHA Title of the thesis : DEVELOPMENT OF MICROENCAPSULATION TECHNIQUE OF BIOCONTROL AGENT AGAINST FUSARIUM WILT DISEASE IN BENGALGRAM (CICER ARIETINUM) Degree : MASTER OF SCIENCE Faculty : AGRICULTURE Discipline : PLANT PATHOLOGY Chairperson : Dr. S. AMEER BASHA University : PROFESSOR JAYASHANKAR TELANGANA STATE AGRICULTURAL UNIVERSITY Year of submission : 2022 18 and increased growth promotion activity in bengalgram plants. The per cent disease incidence of wilt was less (13 %, 20%) in treatment T8 (seed treatment and soil drenching with B. subtilis @ 10 ml + F. oxysporum f. sp. ciceris) at 30 and 45 DAS compared to other treatments. Plant growth promotional activity was also more pronounced in seeds treated with B. subtilis compared to the control. Based on this, B. subtilis (BS-9, OM019320.1) was selected for the preparation of microcapsule. The microencapsulation of B. subtilis (BS-9, OM019320.1) with biodegradable polymers (sodium alginate and chitosan) was done using the extrusion technique. For the preparation of monolayer microcapsule, sodium alginate was used as a polymer, whereas for double layer microcapsule, chitosan was used. Biodegradable polymers are non-toxic, sterile, consistent in quality and easily available. These polymer formulations effectively protect the bioagents against many environmental stresses and release beneficial microbes gradually into the soil. Different concentrations of sodium alginate (0%, 1%, 1.5%, 2%, 2.5%, 3% and 4%) and chitosan (0%, 0.2%, 0.4%, 0.8%, 1.2% and 1.5%) were used for preparation of microcapsules. Out of these, microcapsules prepared with 3 % sodium alginate and 0.8 % chitosan were used in glass house studies. The surface morphology of 3 % sodium alginate monolayer and 0.8 % chitosan double layer microcapsules were observed through scanning electron microscopy (SEM) which revealed that, microcapsules formed were round in structure with shallow grooves and few cavities on its rough surface. Moreover, multiplication of the bacteria inside the capsule resulted in the formation of numerous bulges over the surface area of capsule. Further examination of an bulges revealed that, it was formed of a bacterial microcolony covered with a solidified matrix. The effect of Na-alginate monolayer and Na-alginate-chitosan double layer microcapsule embedded with B. subtilis (BS-9, OM019320.1) against Fusarium wilt disease in bengalgram was studied under glass house conditions. The seed treatment with Na-alginate monolayer + Na-alginate-chitosan double layer microcapsules embedded with B. subtilis (BS-9, OM019320.1) @ 20 g has shown maximum (96.67%) seed germination percentage and more growth promotion activity compared to other treatments. The defense end products such as PAL, PPO, total phenolics and individual phenolic acids in the treated plants were estimated at 24, 48 and 72 h after 21 days of plant growth. The accumulation of PAL, PPO, total phenolics and individual phenolics were more pronounced in T13 (seed treatment with Na-alginate monolayer + Na-alginate chitosan double layer microcapsules embedded with B. subtilis @ 20 g + F. oxysporum f. sp. ciceris) compared to other treatments. Moreover, the seed treated with monolayer and double layer microcapsule embedded with B. subtilis (BS-9, OM019320.1) challenged with pathogen effectively reduced the wilt incidence compared to the other treatments. Despite intensive investigation, many potentially useful bacteria or fungi, reported in literature, never appeared in the commercial market. Microorganisms applied in the field encompass many problems with respect to unpredictable environmental conditions. They have to face the competition of better adapted indigenous microorganism’s variable. Therefore, for commercial use, microbial inoculum should be supported by appropriate formulations preventing a rapid decline of introduced microorganisms and extending their shelf life. Based on ground reality, with the help of microencapsulation technique potential biocontrol agents can be entrapped in microcapsules and applied to plants in field conditions
  • ThesisItemOpen Access
    BIOLOGICAL MANAGEMENT OF REDGRAM WILT CAUSED BY FUSARIUM UDUM
    (PROFFESSOR JAYASHANKAR TELANGANA STATE AGRICULTURAL UNIVERSITY, 2022-07-02) Srikala, J.; Vidya Sagar, B.
    Pigeonpea (Cajanus cajan L var. Manak) is one of the important pulse crops and very popular food in developing tropical countries. India is a principal pigeonpea growing country contributing nearly 90 per cent of the total world's production. Pigeon pea is attacked by more than 100 pathogens including fungi, bacteria, viruses, nematodes, and mycoplasma-like organisms, but only a few of them cause economic losses. Among them, wilt caused by Fusarium udum is considered the most important soilborne pathogen of pigeon pea. The wilt causing pathogen Fusarium udum was isolated from the wilt infected redgram plants and it was tested for its ability to cause disease under glass house conditions and pathogenicity was proved. A total of fifteen rhizobacteria and fifteen rhizospheric fungi were isolated from the rhizosphere of the healthy redgram plant. The isolates were purified and colony characters were recorded such as conidial characteristics like macroconidia, microconidia, and chlamydospore structures. All the isolated bioagents were evaluated in vitro by dual culture technique, against Fusarium f. sp. udum of pigeon pea and the maximum inhibition was shown by Bacillus subtilis RB-4 (73.31 per cent) and Penicillium citrinum RF-15 (75.03 per cent). The isolates of fungi (RF-14, RF-8, RF-15, RF-2, RF-4) and isolates of bacteria (RB-4, RB-8, RB-13, RB-2, RB-5) which proved effective under dual culture experiment were further studied for the effect of volatile and non-volatile compounds against Fusarium udum under in vitro. The Five selected potential bacterial isolates (RB-4, RB-8, RB-13, RB-2, RB 5) from dual culture were tested for biochemical characters like siderophore, catalase, oxidase, IAA, HCN, PSB production under in vitro conditions. All five bacterial isolates (RB-4, RB-8, RB-13, RB-2, and RB-5) were found positive for catalase, oxidase, and grams staining. Among these five isolates, two isolates (RB- 2, RB-4) showed positive to IAA production, all five isolates (RB-4, RB-8, RB-13, RB-2, RB 5) were found positive to HCN, siderophore production and pectinase production, two isolates (RB-4, RB-13) have shown positive for cellulase. ` Under in vitro evaluation, the effect of volatile substances emanating from the cultures of the test microorganisms inhibited the radial growth of Fusarium udum. Volatiles from isolate (RB-4) Bacillus subtilis showed the highest inhibition of (77.04 per cent) and (RF-15) Penicillium citrinum showed the highest inhibition (83.33 per cent). The effect of non-volatile metabolites was more pronounced than the volatile ones. The culture filtrates of isolate (RB-4) Bacillus subtilis (95.65 per cent) and isolate (RF-15) Penicillium citrinum (90.26 per cent) were most effective in inhibiting the radial growth of Fusarium udum. From the above experiments isolate (RB-4) Bacillus subtilis and isolate (RB 15) Penicillium citrinum were observed to be highly effective against test pathogen Fusarium udum. Consideringly, a total of 12 treatments were formulated with potential bio control agents alone and in combination with Neem cake, FYM and VAM impact on Defence enzyme activity, plant growth parameters and wilt incidence under glass house conditions from the formulated treatments. Estimation of defense end product viz., PAL, PO, PPO and TPC activity in redgram leaf samples were analyzed at respective time intervals. Which reported that maximum PO and PPO activity is seen in leaf with treatment T11 (Seed treatment of Penicillium citrinum (BC1) + Bacillus subtilis (BC2)) Whereas, PAL, TPC activity is maximum in T10 (Seed treatment with Carbendazim) followed by treatment T11 (Seed treatment of Penicillium citrinum (BC1) + Bacillus subtilis (BC2)) at 24, 48, and 72 h after inoculation of the pathogen. Data on plant growth parameters of redgram revealed that out of twelve treatments, treatment T11 (Seed treatment of Penicillium citrinum (BC1) + Bacillus subtilis (BC2)) recorded the highest germination (93.31 per cent), root length (19.24 cm), shoot length (81.62 cm), and vigor index (9411.58), while minimum germination (59.53 per cent), root length (14.25 cm), shoot length (62.22 cm) and vigor index (4552.46) were shown by T1 (Seed treatment Penicillium citrinum (BC1)). Pre, post-emergence, and total mortality revealed that all the test amendments non-significantly influenced pre-emergence mortality and significantly influenced post-emergence mortality except treatment T1 (Seed treatment of Penicillium citrinum (BC1)) and treatment T3 (Soil application Penicillium citrinum (BC1) +FYM). The pre-emergence mortality recorded with all the test amendments ranged from (1.22 to 6.85 per cent), as against 6.65 per cent treated control. The post emergence mortality recorded in all the test amendments ranged from (26.53 to 57.24 per cent), as against 100.00 per cent in treated control. Whereas, total mortality recorded in all the treatments ranged from 27.43 to 61.78 per cent as against 100.00 per cent in treated control. Percent reduction of wilt incidence over control results revealed that among all the treatments maximum reduction of wilt incidence over control was recorded in treatment T11 (Seed treatment of Penicillium citrinum (BC1) + Bacillus subtilis (BC2)) 72.56 per cent.
  • ThesisItemOpen Access
    STUDIES ON INTERACTION OF LEAF AND NECK BLAST CAUSED BY Magnaporthe oryzae (Hebert) Barr AND ITS MANAGEMENT
    (PROFFESSOR JAYASHANKAR TELANGANA STATE AGRICULTURAL UNIVERSITY, 2022-11-21) Rajeswari, E.; Vidyasagar, B.
    The present investigation was undertaken with an aim to know the interaction of leaf blast and neck blast pathogen population to host and the variability of isolates using cultural, morphological, pathological, molecular characteristics and avirulent gene distribution. Further efforts were also made to identify the effective R-gene for both leaf blast and neck blast resistance. Attempts were also made to manage the disease with non-chemical and biocontrol agent T. asperellum in integration. A survey on incidence of rice blast disease was conducted in 13 major rice growing districts of Telangana. The incidence varied from 8.24 per cent in Vikarabad district to 58.26 per cent in Warangal district. Disease incidence was recorded among all the prominent cultivars grown in their particular regions and it was observed that different cultivars showed different levels of incidence irrespective of the regions. Among the different cultivars, HMT Sona showed the highest incidence of 71.33 per cent whereas, cultivar Tellahamsa with 49.16 per cent incidence. The maximum isolation rate of the pathogen M. oryzae was recorded from Siddipet with 75 per cent followed by Mehbubnagar with 50 per cent. The disease was observed from nursery to grain hardening stage. A total of 40 monoconidial isolates of Magnaporthe oryzae were established. Out of forty isolates, 36 were leaf blast and 4 were neck blast isolates. The Principal Component Analysis (PCA) was performed for analyzing all forty isolates of M. oryzae growth pattern in 10 different media by considering different components (sectors, zonations, wrinkles, elevation, colony colour, Radial growth and growth rate) for studying cultural variability in M. oryzae population of Telangana. The components elevation, sectors, zonations and wrinkles have been observed to be in strong association among each other (F2: 23.59%). From the eigen value data, it was observed that three components (wrinkles, zonations and sectors) accounts for eigen values more than one and highest variance as indicated in PCA analysis. In present investigation it was ascertained that even in presence of different carbon compounds, host extracts as media source to the isolates their effect was insignificant i.e., irrespective of the carbon source and host extracts the above three parameters were not affected significantly. Oat meal agar was found best for supporting the growth of M. oryzae. However, ragi leaf extract agar media recorded as best for sporulation. Sucrose as carbon source and ammonium nitrate favours the growth of M. oryzae. Morphological variability among M. oryzae isolates was studied through the conidial morphology. Among the isolates, the overall size of the conidia was 14.43-26.80 µm × 3.68- 8.76 µm (Length × Width). A total of 25 rice host differentials with different combinations of resistance genes were tested in for rice blast resistance. Among 25 genotypes tested, NILA57, RamanadStr3, Shia-tia-tsao, Tetep, Tadukan and Dular were found resistant. Genotypes CO 39 and HR 12 were highly susceptible genotypes. A total of 18 races were identified among them the most frequently occurred race was ID-14 followed by IB-41. M. oryzae isolates were identified further to species level with ITS4 and ITS5 universal primers. Actin and TEF 1 α gene primers were also used for further accuracy of identification of conserved sequence, in order to support the morphological identification of M. oryzae isolates. In case of ITS region, all the isolates were appeared as identical by producing the amplified fragment of 560 bp. PCR amplification with actin gene and TEF 1 α gene produced the 336 bp and 550 bp amplicon with DNA from all fourty isolates. These results further confirmed by sequencing of ITS region and identified the isolates as M. oryzae, the pathogen is same that causes both leaf and neck blast. The genetic diversity of M. oryzae isolates was studied using a total of 8 SSR markers and 10 RAPD markers. The polymorphic information content value was high for SSR primers i.e. 0.85 whereas RAPD primers showed only 0.64. Hence the study of genetic diversity among M. oryzae isolates using RAPD primers was more useful than MGM primers. The RAPD analysis formed two main clusters at the Jaccard similarity coefficient of 0.50. This grouping revealed that, all the isolates were clustered in accordance of geographical location but not in the specificity of leaf blast and neck blast isolates, because all the neck blast isolates were clustered in all groups along with the leaf blast isolates. Mating type distribution of M. oryzae isolates was studied by using mating type specific SSR primers. Most of the isolates were proved as MAT1-1. We have identified 8 haplotype groups where, haplotype 2 was predominant (n=27) in the population. M. oryzae isolates were grouped into 8 haplotypes. Majority of the isolates (n=27) were under the Hap-2 group, found to be older haplotype with broad geographical distribution and quite older than other haplotypes as evidenced with its multiple connections with other haplotypes in the study area followed by Hap-4 (n=5) and Hap-1 (n=3) respectively. Haplotype 2 (Hp-2) has the broader genetic base in the entire study area with its swift distribution in the areas of Siddipet, Suryapet, Mehbubnagar, Karimnagar, Medak and Rangareddy. The broader genetic base of DNA sequence polymorphism analysis of Haplotypes was illustrated in the present study using PopART (Population Analysis Reticulate Tree). In the present investigation, Tajima’s D test (-2.660) Fu’s Fs statistics (2.931) supported presence of higher rare alleles in the population. To identify the diversity of the 14 avirulence genes, specific primer pairs were used in this study. The highest frequency of avirulent gene prominent in M. oryzae population was Avr Pi54 (97.5%) followed by Avr Pizt with (95%) and Avr Pi9 with (92.5%). The avirulent genes Avr Pik, ACE1, PWL2, Avr Pib, Avr Pii were able to amplify with a frequency of 90 per cent, 90 per cent, 80 per cent, 80 per cent and 17.5 per cent respectively. The avirulent genes with lowest prominence were PWL1, Avr Pita and Avr C039 with frequency of 7.5 per cent, 5 per cent and 5 per cent respectively. Five artificial inoculation methods, Spraying, smearing, cotton string, cotton wool and Injection were evaluated and out these, injection method gave 96.00 per cent PDI whereas, cotton wool method and string method showed 58 per cent PDI and 24 per cent PDI, respectively on HR 12 cultivar. In cross inoculation studies, a pool of isolates consists of two leaf blast isolates i.e., Mo-31, Mo-28 and two neck blast isolates i.e., Mo-3 and Mo-6 were used. The results indicated that all the isolates (both leaf and neck blast isolates) showed both leaf blast and neck blast infection. In cross inoculation studies on other cereal hosts the rice blast pathogen can infect ragi rather than wheat. In the present study, the interaction between 29 monogenic lines, carrying at least one major R gene, with 16 races commonly found. The monogenic lines carrying Piz5, Pi1, Pi2 and Pi54 provided broad spectrum resistance and their resistance frequencies were 62.5 per cent, 97.5 per cent, 97.5 per cent and 77.5 per cent respectively. The races of M. oryzae from Telangana were able to overcome 12 resistance genes (Pib, Piks, Pik, Pik-h, Pik-m, Pik-p, Pi1, Pi7(t), Pish, Pita, Pita-2, Pita-3 and Piz-t). This information is useful for development of durable resistant variety to blast disease in Telangana. In present investigation, non-chemical agents viz., neem oil, panchagavya, beejamrutham, vermiwash and buttermilk were tested against M. oryzae using Poisoned food technique. All the non-chemical agents at different concentrations were significantly superior to control in checking the mycelial growth of the fungus. Among the tested agents, panchagavya and neem oil showed 81.67 and 79.63 per cent inhibition of mycelium growth. The antagonistic microflora shown a antagonism percentage of 64.71 to 72.22 against M. oryzae under in vitro conditions, upon seed treatment they exhibited highest seed vigor index ranging from 1561.35 (B7) to 2183.68 (TAIK 1) when compared with control. Further, they were identified through internal transcribed spacer (ITS) and 16S rRNA gene sequences analysis for bacteria and fungi respectively and they were identified as Bacillus cereus, Bacillus xiamenensis, Bacillus subtilis and Trichoderma asperellum. Bio-efficacy tests were carried out in field conditions during the two consecutive years Kharif 2019-20 and Kharif 2021-22 by spraying different combinations of the non-chemical agents. From the two years data, the treatments T2 (Seed treatment with Beejamrutha+ Three foliar sprays of neem oil at 40, 60 DAT and panicle initiation) T5 (Seed treatment with T. asperellum + Three foliar spray of neem oil at 40, 60 DAT and panicle initiation) and T8 (Seed treatment with T. asperellum + Three foliar sprays of Panchagavya) were proved superior for control of blast disease with higher yield. The results showing significant improvement in the seedlings treated with different concentrations of non-chemical agents in terms of disease incidence reduction, lower AUDPC and lower r value during two consecutive years