Thumbnail Image

Theses

Browse

2016 - 201835
Reset filters
Subject: Plant Pathology × End date: 2018 × Start date: 2016 × Type: Thesis ×
Reset filters

Search Results

Now showing 1 - 9 of 35
  • Thumbnail Image
    ThesisItemOpen 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, D
    The 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.
  • Thumbnail Image
    ThesisItemOpen 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 V
    The 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.
  • Thumbnail Image
    ThesisItemOpen Access
    Integrated management of foliar fungal disease of culinary melon (Cucumis meloL. var. acidulus Naudin)
    (Department of Plant Pathology, College of Agriculture, Vellayani, 2016) Narmadhavathy, S; KAU; Kamala Nayar
    The project entitled “Integrated management of foliar fungal disease of culinary melon (Cucumis melo L. var. acidulus Naudin)” was undertaken with the objective of making a comparative evaluation of the efficacy of foliar application of fertilizers, micronutrients, bio-control agents and newer fungicide for the management of Colletotrichum leaf spot (Colletotrichum sp.) disease of culinary melon. Surveys conducted during September 2013 to December 2013, in ten culinary melon fields located at Instructional Farm (IF), College of Agriculture (CoA), Vellayani as well as in farmers’ fields near, CoA, Vellayani, in order to assess the prevalence of major diseases such as Colletotrichum leaf spot and downy mildew disease affecting the crop. Highest disease incidence (DI) and percentage disease index (PDI) of Colletotrichum leaf spot were observed, 75 days after sowing, at Chavadinada (70.00 per cent and 64.44 per cent respectively). Incidence and index of downy mildew disease were recorded in four out of the ten locations surveyed (Palapoor, Papanchani, Kalliyoor and Punjakari). Maximum disease incidence and percentage disease index of downy mildew disease (36 per cent and 33.33 per cent respectively) were observed at Papanchani. The most virulent isolate of anthracnose leaf spot pathogen (IF, Vellayani isolate), obtained during the survey was identified as Colletotrichum fructicola by molecular characterization. The treatment NPK 19:19:19 (0.5 per cent) combined with the fungicide mancozeb (0.4 per cent) and adjuvant was most effective in inhibiting the mycelia growth of the pathogen C. fructicola, in vitro, (100 per cent) over control as well as in suppressing artificially induced anthracnose disease and improving the growth parameters of the plants, in the two greenhouse experiments conducted at the CoA, Vellayani during March to June 2014 and August to October, 2014. Results of two field trials conducted at CoA, Vellayani, during January to March, 2015 and April to June, 2015 for testing four most effective treatments screened from the greenhouse experiments, indicated that NPK 19:19:19 (0.5 per cent) + azoxystrobin (0.15 ml/l) + adjuvant (DI 40.00 and PDI 13.05 respectively) and NPK 19:19:19 (0.5 per cent) + mancozeb (0.4 per cent) + adjuvant (DI 40.00 and PDI 13.47 respectively) were most effective in managing the disease and also increasing total yield of plants, when compared to the remaining treatments. Trials were conducted in farmers’ fields at three locations (Venganoor, Vavamoola and Venjaramoodu) for confirming the efficacy of the two most effective treatments screened from the field trials conducted at CoA, Vellayani and pooled analysis of the results indicated that the lowest PDI (12.22) and DI (28.50) were obtained in plants treated with NPK 19:19:19 (0.5 per cent) + azoxystrobin (0.15ml/l) + adjuvant, which was significantly superior to the other treatments. Results of the microbial studies indicated that there was decline in fungal flora of the plants treated with foliar fertilizer NPK 19:19:19 (0.5 per cent) + azoxystrobin (0.15 ml/l) + adjuvant, days after application of treatments whereas bacterial population was higher in plants applied with the same treatment when compared to the application of combination of foliar fertilizer NPK 19:19:19 (0.5 per cent) + mancozeb (0.4 per cent) + adjuvant. There was indication of higher induction of systemic resistance in plants treated with NPK 19:19:19 (0.5 per cent) + azoxystrobin (0.15 ml/l) + adjuvant due to the higher activity of defense related enzymes, such as phenylalanine ammonia lyase (PAL), peroxidase (PO), polyphenol oxidase (PPO), β-1,3glucanase, super oxide dismutase (SOD) and the compound phenol, all of which, reached maximum level on the 15th day after treatment. Leaf samples obtained from plants treated with foliar fertilizer NPK 19:19:19 (0.5 per cent) + azoxystrobin (0.15 ml/l) + adjuvant indicated highest nutrient use efficiency in all three locations of the confirmation trials while highest pigment status due to this treatment was observed in the trial conducted at Venganoor. Relative water content was generally high in leaf samples collected from all plants irrespective of the treatments, although it was comparatively low, in leaf samples obtained from plants of absolute control plot. Epicuticular wax content was slightly lower in the plants treated with combination of the foliar fertilizer NPK 19:19:19 (0.5 per cent) and fungicides, either azoxystrobin (0.15 ml/l) or mancozeb (0.4 per cent) + adjuvant. Stomatal frequency on the upper and lower surfaces of leaves was not much affected by application of foliar fertilizer NPK 19:19:19 (0.5 per cent) combined with the fungicides. B:C estimated ratio revealed that the highest returns were obtained from the plants treated with foliar spray of NPK 19:19:19 (0.5 per cent) + azoxystrobin (0.15 ml/l) + adjuvant, in all three locations of the farmers’ field trials. This study presents the first report of the pathogen Colletotrichum fructicola causing anthracnose leaf spot disease of culinary melon in India. In field conditions, combination of the foliar fertilizer NPK 19:19:19 (0.5%) and azoxystrobin (0.15 ml/l) along with adjuvant applied twice at 15 days’ interval was most effective in controlling anthracnose leaf spot disease of culinary melon and also increasing the yield of the crop.
  • Thumbnail Image
    ThesisItemOpen Access
    Strain improvement of oyster mushrooms- pleurotus cystidiosus O.K. Mill and pleurotus opuntiae (Durieu and LEV.) SACC.
    (Department of Plant Pathology, College of Agriculture, Vellayani, 2018) Krishnapriya, P J; KAU; Geetha, D
    The present study entitled “Strain improvement of oyster mushrooms: Pleurotus cystidiosus O.K.Mill and Pleurotus opuntiae (Durieu and Lev.) Sacc.” was carried out in College of Agriculture, Vellayani during 2015-2018, with the objective to standardize the techniques for production of oyster mushrooms: P. cystidiosus and P. opuntiae; and to study their morphological, physiological and cultural characteristics as well as nutritional and organoleptic qualities; and to undertake genetic improvement by protoplast fusion. The mushrooms were collected from two locations of Thiruvananthapuram and three fast growing isolates of Pleurotus spp. viz., PC2 (Vellayani), PNC1 (Chirayinkeezhu) and PO1 (Vellayani) were selected for the study. These isolates were identified as P. cystidiosus subsp. abalonus, P. cystidiosus and P. opuntiae using internal transcribed spacer (ITS) primers and subsequent sequencing; and registered at Genbank database with accession numbers KY214254, KY887023 and KY214255 respectively. The fast growing isolates of P. cystidiosus (coremial), P. cystidiosus (non-coremial) and P. opuntiae recorded maximum growth on PDPA amended with one per cent yeast under dark condition. The optimum temperatures for the growth were 30 0C, 25 to 30 0C and 25 0C respectively whereas, the optimum pH were 8, 8 and 7 to 8 respectively. Studies with different substrates and amendments for spawn production revealed that sorghum with one per cent yeast was the best for P. cystidiosus (coremial) and P. opuntiae whereas, paddy grains with one per cent yeast for P. cystidiosus (non-coremial). Experiments with different substrates and amendments for mushroom production revealed that rubber wood sawdust sprayed with 2.5 per cent of 1 M potassium dihydrogen phosphate recorded the maximum BE for P. cystidiosus (non-coremial) (192.76 per cent). P. opuntiae recorded the maximum BE in rubber wood sawdust amended either with 4 per cent neem cake (91.38 per cent) or wheat bran (91.37 per cent). Major insect pests observed in the beds of Pleurotus spp. were phorid flies, spring tails, black ants and staphylinid beetles. The competitor moulds observed were different species of Coprinus, Aspergillus, Penicillium and Trichoderma. Sporocarps soaked in one per cent CA for 15 minutes followed by mechanical drying and powdering was the best post harvest treatment for both P. cystidiosus (non-coremial) and P. opuntiae. Mycelium of P. cystidiosus (coremial) showed black coremial structures, representing its asexual stage (Antromycopsis broussonetiae Pat. & Trab.). The coremia comprised of elliptical (16.31 µm x 7.48 µm) and round conidia (8.06 to 8.49 µm). The black colour of coremia was due to melanin which was extracted (255.56 mg l-1) and characterized. The performance of long duration P. cystidiosus (non-coremial) and short duration P. opuntiae was compared with two ruling mushrooms of Kerala viz., long duration P. florida (Mont.) Singer and short duration P. eous (Berk.) Sacc. The study revealed that P. cystidiosus (non-coremial) and P. opuntiae showed higher BE compared to P. florida and P. eous, respectively. P. cystidiosus (non-coremial) recorded maximum moisture (94.05 per cent), starch (200.55 mg g-1), protein (30.2 mg g-1), fat (4.25 per cent), antioxidants (485.45 μg equivalent gram of ascorbic acid-1), beta-carotene (25.69 µg 100 mg-1), polyphenols (7.55 mg g-1) and energy (359.45 Kcal) compared to other Pleurotus spp. Sensory evaluation of mushroom products made from the species of Pleurotus was done and masala curry prepared from P. cystidiosus (non-coremial) scored the maximum value for overall acceptability. Shelf life of P. cystidiosus (non-coremial) was higher (5 days) compared to P. opuntiae, P. florida and P. eous (3 days each) in perforated poly propylene covers stored under refrigeration. Vanillin (0.05 per cent) and carbendazim (1 mM) were selected as dual biochemical markers for the PEG mediated protoplast fusion. Three days old P. cystidiosus (non-coremial) and four days old P. opuntiae recorded the maximum protoplast yield at five and four hours after incubation respectively with 0.6 M KCl and 30 mg ml-1 of enzyme consortium. Eight fusant lines with varied mycelial characters were obtained. Among fusants, F6 and F8 did not segregate in the second generation whereas, F4 segregated. F6 and F8 recorded higher BE of 168.05 and 99.95 per cent respectively compared to the parental lines and other fusants. Sporocarp of F6 and F8 was morphologically similar to P. cystidiosus (non-coremial) and P. opuntiae respectively; and F8 also exhibited low temperature adaptability. The present investigation indicated the exploitability of two promising isolates viz. P. opuntiae for tropical areas and P. cystidiosus (non-coremial) for cooler regions of Kerala using locally available materials and the standardized cultivation practices. The present study also standardized the protoplast fusion technique between P. cystidiosus (non-coremial) and P. opuntiae; and two fusant lines (F6 and F8) recorded higher BE which can be used for future breeding programmes.
  • Thumbnail Image
    ThesisItemOpen Access
    Integrated management of anthracnose of snake gourd (Trichosanthes cucumerina L.)
    (Department of Plant Pathology,College of Agriculture, Vellayani, 2016) Aswani Devi; KAU; Kamala Nayar
    The objective of present study entitled “Integrated management of anthracnose of snake gourd (Trichosanthes cucumerina L.)” was to make a comparative evaluation of foliar application of newer fungicides and biocontrol agents for the management of anthracnose disease of snake gourd. Surveys were conducted during October 2015 in snakegourd fields of five different locationsof Thiruvananthapuram district , viz., Instructional Farm, College of Agriculture,Vellayani, Department of Olericulture, College of Agriculture, Vellayani, Kalliyoor, Kakkamoola and Palapoor. Maximum disease incidence (90.00 per cent) and disease severity/ percentage disease index (44.22) were recorded in the snake gourd fields of Instructional Farm, Vellayani whereas disease parameters were minimum (21.89 per cent and 70.00 per cent) in the fields of Palapoor Pathogenicity tests revealed that the isolate C1 obtained from Instructional Farm, Vellayani produced maximum lesion size both on detatched leaf (2.53 cm2) as well as and on intact leaves of 30-days old potted plant (10.06 cm2). The smallest lesion size of 1.33 cm2 on detatched leaf and 1.53 cm2 on 30 days old potted plant was produced by the isolate C4 from Kakkamoola. Based on lesion size and virulence rating, C1 was screened as the most virulent isolate. Results of the cultural studies among the five isolates, showed that potato dextrose medium, oat meal medium and Richard’s medium were screened as the best media both in solid and liquid states for the growth of the tested isolates. Effect of different light sources (Fluorescent light: 253.7 - 185 nm, L.E.D light: 365 nm and UV light: 10 - 380 nm) on growth and sporulation of anthracnose pathogen indicated that the cultures exposed to fluorescent light and darkness for alternate cycles of 12 h of each day resulted in maximum mycelial growth and minimum days for sporulation for all the isolates. Growth was less when the cultures of the isolates were exposed to UV light for a period of 45 min. The morphological characters studied indicated that average conidial size of isolate C1, C2, C3 and C5 ranged from 11.40μm- 13.14μm x 4.48μm-4.82 µm and that they were cylindrical in shape with obtuse ends. The isolate C4 was ellipsoidal in shape and had a conidial size of 10.52μm x 4.40 µm. The isolates C1, C2, C3 and C5 were further identified at National Fungal Collection Culture India (NFCCI) - Pune, as C. coffeanum F.Noack and the isolate C4 was identified as C. musae (Berk. & M.A. Curtis) Arx. Isolate C1 of C. coffeanum which was earlier screened as the most virulent isolate was used for the subsequent studies conducted for the management of anthracnose disease in snake gourd. In the invitro assayconducted for screening of newer fungicides and bio control agents, effective for the inhibition of the isolate C1 of C. coffeanum, the nutrient potassium silicate (0.5 per cent) and fungicide mancozeb (0.4 per cent) resulted in hundred per cent mycelial inhibition while, KAU talc based formulations of the bio control agents viz., P. fluorescens (2 per cent) and T. viride (2 per cent) resulted in inhibition of 88.33 per cent and 87.33 per cent respectively and were selected for further evaluation in green house study. Minimum spore germination was recorded due to amendment of medium with T. viride (1.33 per cent). Conidia were also reduced in size when growth medium (PDA) were amended with tested chemicals and bio-control agents. In the green house studies using the snake gourd variety Kaumudi, maximum disease suppression was observed when plants were sprayed with 0.5 per cent potassium silicate (89.12 per cent) and 0.4 per cent of fungicide mancozeb (84.99 per cent) at fifteen days interval. Biometric parameters observed were also maximum for plants sprayed with 0.5 per cent potassium silicate followed by mancozeb and KAU talc based formulation of bio-control agents P. fluorescens and T. viride. Anthracnose affecting the foliage of snake gourd is a serious disease in snakegourd fields of Thiruvanathapuram district. The nutrient potassium silicate which was tested for the first time in Kerala against a plant pathogen, Colletotrichum sp. is found to be a promising chemical for management of the disease. The present study has also revealed the prospects of utilizing the mineral nutrient, potassium silicate for control of anthracnose disease as well as attainment growth promotion, flowering behavior, yield components and increases in yield of snake gourd which is an important vegetable crop of Kerala. Besides, this study also has highlighted the scope for integrating the fungicide mancozeb with bio-control agents like P. fluorescens and T. viride which were was also highly beneficial for the disease management as well as in improving the growth parameters of the crop under green house conditions. Such integration of fungicide mancozeb with bio-control agents will also be useful in minimizing the detrimental effects of continous and intensive use of this fungicide which is otherwise very effective in disease management. In the background of the promising results obtained from this thesis project especially from the use of potassium silicate, trials for confirming the beneficial effects of the treatments in field grown snake gourd plants would be useful for the vegetable growers in Kerala.
  • Thumbnail Image
    ThesisItemOpen Access
    Strain evaluation and production technology of shittake mushroom ( Lentinula edodes ( Berk. ) pegler)
    (Department of Plant Pathology, College of Agriculture, Vellayani, 2016) Deepa Rani, C V; KAU; Lulu Das
    The present investigation on "Strain evaluation and production technology of Shiitake mushroom (Lentinula edodes (Berk.) Pegler' was conducted at Department of Plant Pathology, College of Agriculture, Vellayani, Thiruvananthapuram during the period 2012-2015. The aim of the experiment was to exploit various strains of Lentinula spp. for novel production technology and their phylogeny analysis through physiological and molecular studies. Surveys were collected during pre and post monsoon periods of May to December from different parts of Thiruvananthapuram, Kollam, Wayanad, Idukki, Pathanamthitta, Kannur and Kasargode districts. Six isolates of sp. (VLYN- 1 to VLYN-13) obtained during the survey were identified and compared with procured reference strains of Lentinula edodes (LE-1 to LE-5 from GB Pant University of Agricultural and Technology, Pantnagar, Uttarakhand) and LE-6 strain (Maharana Pratap University of Agriculture and Technology, Udaipur) . Morphologically the native isolates of Lentinus spp. had concave, funnel and convex pileus with varying colors and were leathery in nature.L. edodes strains in contrast had convex pileus with chocolate brown and golden yellow sporocarps which were fleshy and edible. Phylogenetic analysis of all six strains of L. edodes using RAPD markers confirmed the variability between the strains. Maximum similarity coefficient of 74.10 per cent was observed between LE-2 and LE-6 strains while LE-2 and LE-4 strains showed a minimum similarity coefficient of 35.70 per cent. Further studies by ITS sequencing showed that all the L. edodes strains tested in the study showed 99- 100 per cent similarity with the known sequences off L. edodes available in NCBI database while that of native isolates showed 99- 100 per cent similarity to Lentinus tuber-regium and Lentinus connatus thus confirming the variability between Lentinus and Lentinula sp. All the six strains of L. edodes, showed maximum mycelial growth in malt extract peptone dextrose agar in solid and oat meal broth in liquid medium. L. edodes strains preferred temperature of 20 °C with an acidic pH of 6. Dark and ambient light conditions favored maximum mycelial growth and biomass production for L. edodes culture. Although a minimum period of 16.33 days was required for full mycelial run in maize grains but due to comparatively less contamination rate in paddy grains which took 18.33 days for completion of mycelial run were selected as best substrate for further studies. Different substrates were evaluated for the development of a cultivation package for shiitake mushroom. Results showed that LE-1 strain took minimum of 71.00 days for initiation of sporocarp in sawdust supplemented with 20 per cent wheat bran. Hard wood sawdust especially of teakwood was used in the study. The substrate based on paddy straw and banana pseudo stem were not found effective for pinhead initiation and thus failed to produce sporocarps. LE-1 produced maximum sporocarp (11.33) in sawdust + 20 per cent wheat bran which was followed by LE-3 (10.63) in sawdust + 20 per cent rice bran. Maximum yield of 290.66 g/ 500 g substrate was obtained in sawdust + 20 per cent wheat bran by LE-6 strain. Maximum biological efficiency of 58.13 per cent was also recorded in LE-6 in sawdust supplemented with 20 per cent wheat bran substrate. Substrates like paddy straw and sawdust amended with 20 per cent wheat bran substrates were evaluated for the development of native isolates of Lentinus tuberregium and Lentinus connatus . Results showed that maximum biological efficiency of 58.00 per cent was obtained by Lentinus tuber-regium whereas 36.60per cent biological efficiency by Lentinus connatus in sawdust amended with 20 per cent wheat bran substrate. Nutrient analysis of all the six strains showed that carbohydrate content ranged between 35.29 per cent to 40.23 per cent, protein 18.33 per cent to 21.66 per cent, crude fibre 22.33 per cent to 27.33 per cent, Vitamin- C 2.53 per cent to 3.50 per cent, ash 2.70 per cent to 4.40 per cent and lipid 2.46 per cent to 3.60 per cent. Mineral content of L. edodes included Ca (11.00 mg to 19.00 mg/ 100 g), Mg (0.46 to 1.10 mg/ 100 g), Fe (1.36 mg to 1.80 mg/ 100 g), Mn (1.53 mg to 2.63 mg), P (1.65 mg to 2.87 mg), K (16.33 mg to 25.20 mg), Na (13.00 mg to 23.66 mg) and Zn (19.66 mg to 28.33 mg/ 100 g). Sensory evaluation of mushroom products made from L. edodes was carried out by a panel of judges for various characters of which mushroom masala scored maximum for texture, taste, flavor and overall acceptability when compared to other recipes like mushroom cutlet, scramble, soup, baji and biscuit. As part of the study, paddy grain was found to be the most suitable substrate for spawn production of L. edodes and teakwood sawdust amended with 20 per cent wheat bran was the most efficient bed substrate. LE-6 strain was superior in terms of yield and biological efficiency. Therefore findings of the above investigation recommends the adoption of a suitable cultivation package for shiitake mushroom by using low cost substrates (hardwood sawdust) available in Kerala in plains and hilly regions.
  • Thumbnail Image
    ThesisItemOpen Access
    Integrated management of viral diseases of bittergourd (momordica charantia L.)
    (Department of Plant Pathology, College of Agriculture, Vellayani, 2018) Radhika, N S; KAU; Umamaheswaran, K
    The present research work entitled ‘Integrated management of viral diseases of bitter gourd (Momordica charantia L.) was carried out in the College of Agriculture, Vellayani during 2014-2017, with the objectives to study the occurrence and distribution of viruses in bitter gourd in Thiruvananthapuram, Idukki and Palakkad, immunomolecular characterization of the viruses, and screening of antiviral chemicals, antiviral principles of animal, plant and microbial origin for the management of the disease. In the suvey conducted at five locations in Thiruvanaanthapuram district, Pappanchani area recorded highest incidence of viral disease (60%) while highest Vulnerability Index (V.I) was recorded from Vellayani (56.00). In Idukki district, six major bitter gourd cultivating areas were surveyed among which Rajakumary area recorded the highest disease incidence (100%) and V.I (82.00). In Palakkad district, five locations were surveyed, among which panackatri and Thekkepotta recorded highest disease incidence of 88% and highest V.I (69.00). The major insects associated with the crop were whitefly (Bemisia tabaci (Genadius) with an incidence of 10-25%, aphids (Aphis gossypii glover) with an incidence of 10-40%, Jassids (Empoasca (Empoasca) motti Pruthi) with an incidence of 10-30% and mites with an incidence of 10-50%. Phyllody and little leaf symtoms (20% incidence) were also recorded in bittetgourd form Rajakumary and Rajakkad areas in Idukki. Flat limb and multiple proliferation of shoot tip were observed at many fields in Idukki. Symptoms associated with the disease include yellow mottle, mosaic,blistering, leaf curl and reduction in leaf size. Yellow mosaic and blistering is seen in severe infection finally leading to stunting of the plant, reduced flowering an fruiting and hairyness on stem. Mechanical transmission of the virus on Datura stramonium produced yellow lacal lesions indicating the presenceof Bean Golden mosaic virus (Begomo) in the infected leaf extract. This leaf extract also produced local lesions on othe indicator hosts like Chenopodium amaranticolor and Gomphrena globosa indicating the presence of Cucumber mosaic virus (CMV) or Potato virus Y (PVY). The viruses were transmitted by whiteflies (20%) and aphids (30%) from infected bittetgourd plants to healthy seedlings. Whiteflies (Bemisia tabaci Gennadius)) and aphids (Aphis gossypii Glover) are the vectors of the respective viruses Wedge grafting diseases scion on to 3-5 leaf stage healthy seedling of bittergourd produced symptoms of infection within ten days. KAU varieties Preethi and Priyanka were found to be susceptible to infection with preethi expressing a V.I of 70.80 and Priyanka expressing a V.I of 62.50 respectively. Ensyme linked immunosorbent assay (ELISA) and Dot immunobinding assay (DIBA) revealed the presence of three viruses belonging to Begomo, CMV and PVY group causing an mixed infection in bittergourd. The presence of all the three viruses were also confirmed in electron micrograph, Begomovirus as twin particles of size 18-20 X 30nm,CMVas single particles of 18nm and PVY as lonog flexuous rod of size 750nm. PCR amplification of coat protein gene (cp gene) of virus isolates from all the three districts yielded an amplicon of size approximately equal to 570 bp. Idukki and Palakkad isolates showed 94% identity to Tomato leaf Curl Virus isolate TNUDU BGI Coat Protein (AVI) gene while Trivandrum isolate showed 95% identity to Tomato leaf Curl Virus isolate TNPDU BG4 Coat Protein (AV1) gene . Phylogenetic tree constructed using multiple sequence alignment programme showed close relation between Begomo viruses identified in bittergourd from different districts. Studies on defense related enzymes such as peroxidase (PO), polyphenol oxidase (PPO) and phenyl alanine ammonialyase PAL) showed significant activity of PO and PPO in diseased plants than in healthy plants and the activity was on par in healthy and diseased for PAL. Protien profile of healthy and diseased at different days after virus inoculation through grafting indicated the production of novel proteins in diseased. There was no difference in the native profile of peroxidase in healthy and diseased at 15 days after virus inoculation. An additional isozyme band with a Rm value of 0.5 was observed in diseased at 45 days after virus inoculation. Management of the disease with antiviral chemicals and antiviral principles of plant, animal and microbial origin was undertaken as pot culture studies with pre and post inoculation of treatments. Twelve treatments with three replications each were laid out in completely randomized design for the evaluation. The treatments included Aspirin at two levels of 100 and 150 ppm, Salicylic acid (SA) at two levels of 100 and 150 ppm and Acibenzolar S methyl (ASM) at 50 and 75 ppm concentration, and two commercial formulations viz., Perfect and virus –Ex at 0.5 and 1.0 ml concentrations. The treatments were applied three times at 10 days interval. Pre application of thrice sprapying of Acibenzolar S methyl (ASM), 75 ppm concentration (V.I-35.00) at ten days interval was statistically significant over other treatments followed by ASM-50 ppm (V.I-41.33). Post application of antiviral chemicals also showed a statistically significant effect of three times spraying ASM-50 ppm(V.I-25.00) at ten days interval followed by spraying of Virus Ex 1ml L-1 (VThe best eight treatments with control was laid out as Randomised Block Design at the Instructional Farm, College of Agriculture, Vellayani during February to May 2017 as a field trial to study the effect of treatments on natural incidence of the viruses in the susceptible variety Preethi. The treatment, three sprays of ASM-50 ppm (V.I-28.33) at ten days interval ws on par with buttermilk (Three times dilution of curd) (V.I-39.16). Yield was also significantly high in ASM-50 ppm (437g plant-1) followed by Pseudomonas fluorescens talc based formulation (2%) (233 g plant-1)among the treatments.
  • Thumbnail Image
    ThesisItemOpen Access
    Etiology and management of mosaic disease in ginger (Zingiber officinale Roscoe)
    (Department of Plant Pathology, College of Agriculture, Vellayani, 2018) Ananthu, N; KAU; Umamaheswaran, K
    The study entitled ‘Etiology and management of mosaic disease in ginger (Zingiber officinale Roscoe)’ was conducted at the Department of Plant Pathology, College of Agriculture, Vellayani during 2015-2018 with the objectives to identify, characterize and sequence the genes of Ginger mosaic virus infecting ginger along with the management of the disease. As part of the study, the symptoms produced by the virus in ginger plants collected from the field and grown in glass house were observed. The symptoms on the leaves appeared as small light green flecks. These flecks eventually increased in size and formed streaks. The streaks were arranged parallel to the veins. The appearance of too many streaks on the leaves led to severe chlorosis and the leaves showed necrotic symptoms in the advanced stage. Transmission of the virus was tested in rhizome (seed material collected from infected plants) and mechanical inoculation was done using infected leaf sap to the healthy plants. The infected rhizomes resulted in 100% transmission and mechanical transmission failed to transmit the virus. Changes in total carbohydrates, chlorophyll, phenol, total soluble proteins and defense related enzymes namely peroxidase, polyphenol oxidase and phenylalanine ammonialyase were carried out at 30, 60, 90 and 120 days after infection. The study revealed an increase in the content of phenols and defense related enzymes in infected plants. An increase in carbohydrates, chlorophyll and protein in healthy plants was also observed. Protein profile study using sodium dodecyl sulphate- polyacrylamide gel electrophoresis (SDS- PAGE) indicated the presence of a novel protein with molecular weight of 20 kDa in the infected plant sample. The immunological detection techniques direct antigen coating- enzyme linked immunosorbant assay (DAC- ELISA) and dot immunobinding assay (DIBA) were carried out. Since the etiology of the virus was unknown, seven suspected viruses were tested in DAC- ELISA and antibodies specific to two viruses namely Banana bract mosaic virus (BBrMV), Cucumber mosaic virus (CMV) gave an absorbance value of 0.15 and 0.19 respectively which was three times more than the absorbance shown by the healthy leaf (0.05 and 0.07). The infected leaf tested for the presence of African cassava mosaic virus (ACMV) by triple antibody sandwich ELISA (TAS-ELISA) gave an absorbance of 0.0425 while the healthy gave an absorbance of 0.017. DIBA analysis gave positive reaction to BBrMV. Polymerase chain reaction ( PCR) was carried out with both total DNA and RNA isolated from infected ginger leaf sample. The PCR experiment was positive to Begomoviruses and negative to PVY (Potato virus Y) and BBrMV isolates. An amplicon of size 550 bp was obtained for the sample DNA using begomo degenerate primer. The sequence was subjected to BLAST analysis which indicated 74 per cent similarity to Tomato leaf curl virus Bangalore isolate. The management studies were conducted with antiviral principles like botanicals, chemicals and that of microbial origin against the virus. The experiment was conducted in completely randomized design (CRD) with 13 treatments and three replications. Karthika was the variety used for the study. Perfekt (a botanical extract-76%) at 0.5 ml L-1 and 1ml L-1, chemicals namely aspirin, salicylic acid, barium chloride, at 100 and 150 ppm concentrations and botanicals namely ten per cent leaf extracts of Mirabilis jalapa and Bougainvillea spectabilis, two per cent neem oil garlic emulsion and two per cent PGPR mix II were used in the experiment. The treatments were given at fortnightly interval. Before each spray the efficacy of the treatments were evaluated using Vulnerability Index (V.I) developed by Bos (1982). The treatments with Perfekt at the rate of 0.5 ml L-1 and 1.0 ml L-1 and ten per cent leaf extract of Mirabilis jalapa were found effective for the management of the disease. The study indicates that the mosaic disease in ginger is transmitted through rhizomes, the virus has been molecularly characterized and shows only 74% identity with Tomato leaf curl virus (TLCV) and management of the disease can be done by application of Perfekt at 0.5 ml L-1 or 10% leaf extract of Mirabilis jalapa at fortnightly interval.
  • Thumbnail Image
    ThesisItemOpen Access
    Identification of graft transmissible resistant factors and development of si RNA mediated resistance in cassava against cassava mosaic geminivirus
    (Department of Plant Pathology, College of Agriculture, Vellayani, 2017) Asha, B Nair; KAU; Umamaheswaran, K
    The present study entitled ‘Identification of graft transmissible resistant factors and development of siRNA mediated resistance in Cassava against Cassava mosaic virus’ was carried out during the period 2012-2017 at the Department of Plant Pathology, College of Agriculture, Vellayani. The study was carried out with the objective of identification of transferability of resistance factor from resistant cassava, Sree Padmanabha to susceptible, Vellayani Hraswa by grafting and to develop siRNA mediated technology for the development of cassava plant resistant to Cassava mosaic geminivirus. Grafting experiments were conducted using resistant Sree Padmanabha as root stock and susceptible Vellayani Hraswa as scion. Symptoms like leaf mosaic, chlorotic spots, reduction in leaflet size and stunting of plants were noticed in susceptible variety. Virus concentration was found to be less in grafted plants. Grafting experiments showed the expression of an extra protein by SDS-PAGE and Coomassie staining in grafted plants which is around 38 kDa. Molecular weight of the new protein revealed the presence of extracellular protein in grafted samples. The extra proteins found in the grafted plants are assumed to be transferred from Sree Padmanabha to Vellayani Hraswa by the process of grafting. The study also involved the development of an intron hairpin RNA vector against replicase gene of SriLankan cassava mosaic virus and introduction of this construct into embryogenic cells via Agrobacterium mediated transformation. A protocol for somatic embryogenesis in cassava variety, Vellayani Hraswa was developed by using immature leaf lobes as explants. The young leaf lobes from tissue culture plantlets produced through meristem culture was used for embryogenic callus formation. Cremish white calli was initiated in Murashige and Skooge (MS) medium supplemented with picloram 12 mg L-1 in dark. For embryogenesis, the calli were transferred to MS medium supplemented with BA 2µM and NAA 1µM which resulted in the production of glassy elongated somatic embryos. The germinated cotyledonary embryos were then regenerated into plantlets by culturing in MS medium supplemented with BA 1mg L-1. Effort was taken to construct an intron hairpin RNA vector and the gene targeted for silencing was the replicase gene of SriLankan cassava mosaic virus (SLCMV). Total DNA was extracted from virus infected plants and the whole replicase gene was isolated using gene specific primers. Sequencing of the whole gene was done. BLAST analysis showed 98% similarity to replicase gene of various isolates of SLCMV. The sequence was then subjected to miRNA target prediction and restriction mapping to select suitable region for the construct. Based on this information, a fragment of 397 bp towards the 5’ end was amplified by designing a set of primers with anchored restriction sites. The primers anchored with Xho and Kpn 1sites were used for the amplification of sense strand and the primers anchored with Xba and Cla 1sites were used for amplification of antisense strand. Selected region was amplified to form sense and anti-sense fragments and cloned to pTZ57R/T cloning vector. Inserts were then released from pTZ57R/T using the corresponding restriction enzymes. The sense and anti-sense fragments were then integrated in the primary vector pHANNIBAL on either side of the pdk intron which facilitated the formation of intron hairpin RNA construct. The intron hairpin RNA construct in pHANNIBAL contained CaMV35S promoter, sense strand, pdk intron, antisense strand and OCS terminator in the order with Not 1 restriction sites. After confirmation of integration by restriction digestion, the Not1 fragment with sense and anti-sense strand were released from pHANNIBAL and ligated to the digested Not1 site in the lacZ gene of binary vector pART27 containing antibiotic resistant marker nptII and spec. the binary vector was confirmed for the presence of insert by transferring to DH5α cells and colony selection by blue white screening. Plasmid DNA isolated from transformed colonies grown on Luria agar medium supplemented with 100 mg L -1 spectinomycin were confirmed for the presence of insert. After confirmation of insert in the binary vector, it was transformed to Agrobacterium tumefaciens strain LBA4404 via freeze thaw method. Transformed colonies were selected on kanamycin selection medium at 100 mg L -1 and confirmed for the presence of binary vector and ihpRNA insert using nptII primers and primer for sense and antisense strands by PCR reaction. Cotyledons excised from the somatic embryos were transformed with LBA4404 having pART 27 by co-cultivation and the transformed embryos were selected with antibiotic pressure (Kanamycin 100 mg L-1). DNA was isolated from the transformed somatic embryos and confirmed for the presence of insert using forward primer of sense fragment and reverse primer of antisense fragments. Transformed embryos were subjected to regeneration.