1. KAUTIR (Kerala Agricultural University Theses Information and Retrieval)
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Item Physiological and molecular assessment of abiotic stress tolerance in selectively fertilized coconut hybrids (Cocos nucifera L.)(Department of Plant Physiology, College of Agriculture,Vellayani, 2024-03-14) Afna Mol, O P; KAU; Roy Stephen; Gladis, RThe present study entitled “Physiological and molecular assessment of abiotic stress tolerance in selectively fertilized coconut hybrids (Cocos nucifera L.) was carried out in the Department of Plant Physiology, College of Agriculture, Vellayani during 2018-2023. The objective of the study was physiological and molecular evaluation of selectively fertilized coconut hybrids for water stress and temperature stress tolerance and to assess genetic variability for temperature stress tolerance through pollen selection at critical temperature. The study consisted of four experiments. In the first experiment, 10 year old field grown coconut palms of the following varieties Kerasree (WCT X MYD)- 198 selectively fertilized, Keraganga (WCT X GB)-selectively fertilized, Kerasree (WCT X MYD) Hybrid, Keraganga (WCT X GB) Hybrid and West Coast Tall at Regional Agricultural Research Station, Pilicode, Kasaragod were used as the experimental material for the study. The experiment was carried out in Randomized Block Design (RBD) with four replications during 2019 March to 2019 April. Water stress was imposed by withdrawing irrigation from the field for one month and physiological and biochemical observations were made at 15 days interval .Then the palms were irrigated on 30th day and recovery responses were analysed on 15th day after the irrigation rescheduling. On physiological analysis, selectively fertilized Kerasree showed higher relative water content and epicuticular wax of the leaves. Keraganga selectively fertilized showed maximum photosynthetic rate and cell membrane stability index. Leaf temperature was minimum in Kerasree selectively fertilized hybrids followed by Keraganga selectively fertilized hybrid. WCT recorded higher transpiration rate followed by Kerasree selectively fertilized hybrid. Stomatal conductance was higher in WCT and then in Kerasree selectively fertilized hybrids. In all the parameters tested both the selectively fertilized hybrids responded positively towards drought than their normal hybrids and this was an indication of higher water use efficiency in them. Recovery response after rewatering was also higher in selectively fertilized hybrids than their normal hybrids. The biochemical parameters like chlorophyll content, antioxidant enzymes viz SOD and peroxidase activities were found maximum in Kerasree selectively fertilized hybrids under water stress condition. Recovery response of these parameters were also found elevated in Kerasree selectively fertilized hybrids. Also Kerasree selectively fertilized hybrids showed lowest membrane damage (lipid peroxidation value) compared with normal hybrids and WCT. Amount of proline in leaves was found higher in WCT, which was on par with Kerasree selectively fertilized hybrid and Keraganga selectively fertilized hybrid. 199 In the second experiment, molecular characterization was carried out in the above five treatments. Protein profiling of leaves was done in all the five treatments of experiment one, using SDS-PAGE, in which, there was variation in the intensity of large subunit (55 kDa) as well as small subunit (around16 kDa) of RuBisCO, among the treatments. The intensity of those bands were found higher in WCT and Kerasree selectively fertilized hybrid .The band expressed in selectively fertilized hybrid in between 26 kDa to 36 kDa was absent in other treatments.. SSR markers CncirE2, CncirE10, CncirE12 and CncirH4 were screened in five treatments and polymorphism was observed in CncirE2, CncirE12, and CncirE10. RNA-seq tranascriptomic analysis of Kerasree selectively fertilized hybrid in reference with Kerasree was done to identify the key differentially expressed genes (DEGs) responding to water stress tolerance in Kerasree selectively fertilized hybrid. A total of 16671 genes were found expressed in Kerasree SF differing from Kerasree. 9635 genes were upregulated and remaining 7036 genes were down regulated in Kerasree SF.Abiotic stress related genes like late embryogenesis abundant protein Lea5 and HSPs were found differently expressed in Kerasree S.F hybrid in response to water stress. Gene ontology of the identified genes revealed that in terms of biological processes the following were the most represented GO terms (GO:0006749, 24 proteins, GO:0006468, 23 proteins,GO:0009415,12). In cellular component (GO: 0016020, 31 proteins) and in molecular function (GO: 0005524, 56 proteins, GO: 0009408, 21 proteins) the above shown were the most represented GO terms. In experiment three, physiological assessment of temperature stress tolerance of selectively fertilized coconut hybrids was done. Pollen grains were collected from the selectively fertilized coconut palms during summer season. The pollen grains were incubated in specific pollen germination media and exposed to different temperature ranging from 35oC to 45oC for a period of 2 hours. The temperature at which only 20-30 % of pollen germinated was identified as the critical temperature stress for pollen germination. From the results obtained, both the selectively fertilized hybrid of Kerasree and Keraganga were noted with maximum critical temperature for pollen germination (42oC) followed by WCT (40oC).This indicated 200 the temperature stress tolerance of selectively fertilized hybrids over their normal hybrids. Pollen viability was found maximum in WCT followed by Kerasree selectively fertilized hybrid. Accumulation of starch was found significantly higher in Kerasree selectively fertilized hybrid and the protein in WCT. The fourth experiment was conducted to screen temperature stress tolerance of 30 coconut genotypes in Regional Agricultural Research Station, Pilicode for recognizing the temperature stress tolerance through critical temperature for pollen germination. Significant genetic variation in critical temperature was discerned among the treatments. Andaman ordinary and Tanjore coconut genotypes were observed with higher critical temperature (42oC) for pollen germination. Hence they can be used as a pollen parent in hybridization programs for developing temperature stress tolerance. The present study revealed that Kerasree selectively fertilized and Keraganga selectively fertilized hybrids of coconut were more tolerant to water stress than normal hybrids in terms physiological, biochemical and molecular parameters. Selectively fertilized hybrids developed through pollen selection at critical water potential also have temperature stress tolerance. By selective fertilization techniques it may be possible to add water stress and temperature stress tolerance traits to high yielding coconut hybrids.Item Abiotic stress tolerance in native isolates of Beauveria bassiana (Balsamo) Vuillemin (Hypocreales :Cordycipitaceae)(Department of Agricultural Entomology, College of Agriculture,Vellanikkara, 2024-07-22) Nimisha, T; KAU; Deepthy, K BEnvironmental factors such as high temperature, low humidity and soil acidity often limit the biological control potential of the entomopathogenic fungus (EPF), Beauveria bassiana. Identifying abiotic stress-tolerant B. bassiana isolates represents a possible strategy to overcome this problem. Native isolates of EPF tend to adapt more to the environmental stress conditions in the locality than the exotic isolates, which are exposed to a different microclimate and microbiota. Thus, this study aimed at identification of the abiotic stress tolerant native isolates of B. bassiana and elucidation of the biochemical and molecular mechanism of stress tolerance. Survey was conducted in ten districts of Kerala, across different agroclimatic zones (159 locations). Soil samples as well as field infected cadavers were collected during the survey. The physicochemical properties of the soil samples were analysed to understand prevailing abiotic stress conditions in the area of sample collection, from which the entomopathogenic fungi isolated. For soil isolation of EPF insect bait method and serial dilution and plating method were followed. Beauveria bassiana was not obtained from any of these soil samples. However, out of the 12 field infected cadavers collected, three were infected with B. bassiana. Beauveria bassiana was identified based on the morphological characters and later confirmed by molecular characterization. Sequencing of the ITS region (550 bp) revealed genetic differences among the isolates. The sequences were submitted to NCBI GenBank (National Center for Biotechnology Information) and the accession numbers were generated (BTL1 - OP271760, BTL2 - OP290199 and PKDE - OP292066). A maximum likelihood phylogenetic tree was built, and the evolutionary relationship among the isolates were also studied. Beauveria bassiana isolates (BTL1, BTL2 and PKDE) were grouped in a single cluster confirming their genetic relationship. Bioassay against third instar nymphs of cowpea aphid (Aphis craccivora Koch) revealed that at lower concentration of 105 spores/ml, only the PKDE isolate recorded cent percent mortality compared to other two native isolates as well as NBAIR (National Bureau of Agricultural Insect Resources) strain (Bb13). As the concentration of spore suspension increased to 107 spores/ml the PKDE and BTL2 isolates were on par with NBAIR strain in terms of LT50 values. The growth and biochemical parameters of the three native isolates of B. bassiana were studied under different abiotic stress conditions. The effects of temperature (28 - 40 oC), pH (2 - 6), salinity (0.5 - 2 M) and water stress induced by polyethylene glycol (PEG 10 - 45 %) on the growth of B. bassiana were assessed. Beauveria bassiana isolate PKDE (collected from Palakkad district) tolerated a temperature stress upto 40 oC. It also survived the extreme acidity (pH 2) and salinity (1.5 M) conditions. The B. bassiana isolate, PKDE was compatible with most of the commonly used insecticides viz., chlorantraniliprole, imidacloprid, thiamethoxam and spinosad. Among the fungicides tested, hexaconazole and carbendazim completely inhibited the growth of all the three isolates, while copper oxychloride showed 89 per cent compatibility with the isolate PKDE. The PKDE isolate of B. bassiana isolated from Palakkad district has shown exceptional resistance to the effects of temperature and drought stresses. Hence, biochemical characterization of this isolate was performed to confirm their ability of stress tolerance. Significant levels of trehalose content were recorded on exposure to heat (40 oC) and drought stress (45 % PEG) (20.33 mg/g of mycelia and 20.43 mg/g of mycelia, respectively) in the multiple stress tolerant PKDE isolate. A significant activity of catalase and peroxidase was also observed in response to heat stress at 40 oC in PKDE isolate (0.0072 EU/min/mg protein and 0.0602 EU/min/g tissue weight respectively), while activity was not significant with respect to drought. In PKDE isolte the mannitol dehydrogenase (MTD) and mannitol -1-phosphate dehydrogenase (MPD) displayed significantly increased activity upon exposure to temperature stress of 40 oC (0.363 and 0.317 EU/ min/ mg protein respectively) and drought stress (0.289 and 0.364 EU/min/ mg protein respectively) induced by 45% polyethylene glycol concentration compared to the control. Field studies concluded that two sprays of B. bassiana (PKDE, BTL2 and NBAIR strain) at a spore concentration of 1x108 spores/ml at 10 days interval, suppressed the cowpea aphid incidence. No mortality of natural enemies (coccinellid beetles and spiders) was observed in the treated plots. The protein profiling of PKDE isolate under stress conditions and without stress (control) was carried out to identify the molecular basis of stress tolerance. The results revealed that there was over-expression of proteins at high-temperature stress, and the molecular weight of proteins ranged between 11-17, 35-48, 48-63 and 100- 135 kDa. The relationship between heat shock proteins and thermotolerance in fungal biocontrol agents suggests a new approach for improving entomopathogenicity by enhancing the expression of thermotolerance-related proteins in conidia. This can be achieved by identifying fungal isolates with greater thermotolerance or by optimising the components of substrate for the growth of fungi to produce more thermotolerant conidia. The nucleotide sequence analysis in the neutral trehalase gene (Bb Nth1) and high osmolarity glycerol gene (Hog), which are known to be associated with multiple stress tolerance, revealed that Bb Nth1gene sequence of the three native isolates of B. bassiana contains a coding sequence (CDS) of 2232 bp which codes for 743 amino acids and the hog gene consists of 1077 bp which codes for 358 amino acids. The polymorphism analysis in the Bb Nth1 gene revealed that seven single nucleotide polymorphisms (SNPs) in the exon region and six SNPs in the intron region, in the BTL1 isolate. In the exon3 region of the BTL1 isolate one SNP was observed with G➔C transition and codon changed from GAG to GAC. The non-synonymous variation resulted in the substitution of glutamic acid to aspartic acid at 363rd position of amino acid sequence. In BTL2 isolate eight SNPs were found in the exon region and seven SNPs in the intron region. BTL2 isolate displayed two non-synonymous variations at the 363rd position (glutamic acid to aspartic acid substitution) and at 542th position (lysine to glutamic acid) of amino acid sequence. The second SNP observed in BTL2 isolate was with G➔A transition and codon changed from GAG to AAG. In the PKDE isolate, six SNPs were found in the exon region and none of which were non-synonymous variations. Only five SNPs were found in the intron region of PKDE isolate. Variant Effect Predictor software was used to determine the functional consequences of the observed SNPs. The two mutations observed in the Bb Nth1 gene resulted in a missense variant. The Protparam and HOPE software results also revealed that the mutation resulted in protein instability. While analysing the multiple sequence alignment of susceptible and multiple stress tolerant isolates, there was no non-synonymous variation in the Hog gene. The results of bioinformatics software such as Variant Effect Predictor and Protparam supported the above findings. The present study identified multiple stress tolerant isolate of B. bassiana (PKDE isolate) isolated from Palakkad district of Kerala which is safer to natural enemies and compatible with synthetic pesticides. This isolate may be successfully integrated as a microbial control agent in IPM programme.Item Assesment of multiple abiotic stress tolerance mechanisms in rice (Oryza sativa L.)(Department of Plant Biotechnology, College of Agriculture,Vellayani, 2019) Alif Ali, B S; Beena, RThe study entitled “Assessment of multiple abiotic stress tolerance mechanisms in rice (Oryza sativa L.)” undertaken at the Department of Plant Physiology, College of Agriculture, Vellayani during 2018-19. The objective was to study the multiple abiotic viz. drought, salinity and high temperature stress tolerance mechanisms in rice and to validate the identified QTLs for stress tolerance in rice. The investigation comprises four experiments, In experiment I initial screening of 20 rice genotypes for single abiotic stress tolerance was studied. Stresses were induced using different concentrations of PEG6000, NaCl and temperature controlled incubator for providing drought, salinity and temperature stresses respectively. Germination study was carried out using paper towel method. In the first experiment drought stress were given at concentrations -1bar, -3bar, 5bar and -7bar water potentials of PEG6000, salinity stress was given at 100mM, 150mM, 200mM, 250mM NaCl and temperature stress were given at 350C, 400C, 450C and 500C for all 20 rice varieties with two replications. The physio-morphological and biochemical parameters were studied on 14th day of germination. The highest level drought, salinity and temperature stresses at which germination occurred was selected as Dh (-5 bar), Sh (250mM NaCl) and temperature (Th) (350C) respectively. Among 20 rice varieties, PTB-7, PTB-60 and PTB-35 showed maximum seedling vigour at highest level of drought stress (Dh) stress condition Vyttila-9, MO-18 and Vyttila-3 recorded maximum seedling vigour index at highest tolerated level of salinity (Sh) stress condition and N-22, NL-44 and Vyttila -6 showed maximum seedling vigour index at highest tolerated level of temperature stress (Th) stress condition. These nine genotypes were selected for the evaluation of combination stress treatment In the second experiment, The combination stress treatments given were Dh x Sh, Dh x Th, Th x Sh and Dh x Sh x Th. Rice varieties did not germinated at Dh x Sh and Dh x Th. The maximum seedling vigour index at Dh x Sh and combination stress treatment was observed in PTB-7, Vyttila-9, PTB-35 and at Th x Sh was observed in NL-44, MO-18 and N-22 respectively. These rice varieties were selected as tolerant varieties. In experiment III six rice varieties selected from combination stress treatment were evaluated for yield parameters in pot culture experiment. The design of the experiment was CRD with two replications and one control. Drought and salt stress were imposed during reproductive stage for 5 days by applying -5bar PEG6000 and 250mM NaCl solutions respectively into the pots containing rice varieties, Temperature stress was induced using a temperature controlled polyhouse from panicle initiation to maturity stage. Physio-morphological, biochemical and yield parameters were studied under the combination stress treatments. Highest yield under the combination stress treatment of Dh x Sh was observed in PTB-7 and highest yield under the combination stress treatment Th x Sh was observed in N-22. Based on morpho-physiological and yield parameters PTB-7 was selected as the tolerant variety under drought and saline condition and N-22 was selected as the tolerant variety under temperature and salinity condition. In experiment IV all the 20 genotypes were analyzed for the identification of reported markers linked to stress tolerance such as drought salinity and temperature. Reported microsatellite markers linked to drought, salinity and temperature were used to screen 20 rice varieties. Among the markers distinct polymorphism for temperature tolerance between temperature tolerant (N-22 and NL-44) and susceptible varieties was shown by RM 6100. RM 7076 showed distinct polymorphism in tolerant varieties PTB-7 and NL-44 . RM 1287 showed distinct polymorphism for salinity tolerance in PTB-7 and N-22. Drought tolerance between drought tolerant (PTB-7) and susceptible varieties was Shown by RM 490.Item Identification of saline tolerant rice hybrids for pokkali tract(Department of Plant Breeding and Genetics, Vellanikkara, 2019) Nikhil Narayanan; Biju, SVarious abiotic stresses causing a decline in global rice production are drought, salinity and chillness. Among these, salinity is the major factor that affects crop productivity. The traditional rice varieties grown under Pokkali area show remarkable tolerance to salinity and floods making it unique. However, the yield potential of these varieties is low. In India, out of ninety three rice hybrids developed, a few viz. DRRH 28, KRH 4, PSD 3, etc. were reported to have tolerance to salinity. Considering the potentials of hybrids, identification of salt tolerant hybrids suitable for Pokkali tract of Kerala is an urgent need of the hour. Hence, the study was designed to explore the adaptability of eight rice hybrids to the unique tract of Pokkali and evaluation of traits associated with salinity tolerance. The study comprised of two experiments, the first experiment was Laboratory screening for salinity tolerance and the second experiment was Field evaluation of rice hybrids in Pokkali tract. The laboratory screening was done following a completely randomised design and the genotypes were grown in Yoshida's solution and evaluated for various seedling characters. High shoot length, root length, high root CEC and high root and shoot proline content were seen in Vytilla-6 followed by Ezhome-2. Low root and shoot proline content were observed in IR-29 confirming its susceptibility to salinity. Among the hybrids, a high value for shoot length, root length, total number of roots, root CEC, root proline, shoot proline and survival per cent was observed in the hybrids CORH-3 and KRH-4. Vigour index was high in Sahyadri 4 followed by KRH-4 and low in MRP- 5401. Vytilla-6 and Ezhome-2 showed the high survival per cent while low survival per cent was noticed in Sahyadri 2 and MRP-5401. Vytilla-6 exhibited high chlorophyll content and RWC while low RWC was recorded in MRP-5401, Sahyadri- 2 and Sahyadri-4. Ca in root and shoot along with Na content in shoot were high in IR-29 while Vytilla-6 had low Ca and Na content. High Na in root was seen in Sahyadri 4 followed by Sahyadri 3. High amount of root Cl2 was recorded in Sahyadri-3, Sahyadri-1, Sahyadri-4, IR-29 and Sahyadri-2 while, the lowest amount of root and shoot Cl2 was in Vytilla-6. High amount of shoot Cl2 was seen in Ezhome-2 and IR-29. High amount of root and shoot SO4 content was seen in IR-29, Sahyadri-3, Sahyadri-4, Sahyadri-2 and Sahyadri-1. The lowest root and shoot SO4 content was recorded in Ezhome-2, followed by Vytilla-6. Low visual score of 3 was recorded in Vytilla-6 and Ezhome-2; a score of 5 was recorded for CORH-3, MRP-5401, TNAU RH-4 and KRH-4. Sahyadri-2, Sahyadri-3 and Sahyadri-4 were observed to have scored more than seven. Score 9 was noticed in Sahyadri-1 and IR-29, indicating that these genotypes are more susceptible to salinity. Visual score under salinity was found to be negatively correlated with shoot and root length, number of roots, vigour, proline content and RWC, while it was positively correlated with content of Ca, Na, Cl2, SO4 and K in the plant tissue. Field evaluation of rice hybrids in Pokkali tract was done during Kharif season of 2016-17, following a randomised block design. Days to fifty per cent flowering and maturity showed that all hybrids became early under salinity. Tall plants were observed in Vyttila-6 and Ezhome-2 and short plants in IR-29. High number of productive tillers was observed in CORH-3 and low in Sahyadri-2. Ezhome-2, IR-29 and Vyttila-6 had high number of spikelets per panicle while seed setting percentage and long panicles were observed in Ezhome-2. Spikelet sterility was high in Sahyadri-2 and low Ezhome-2. High 1000 grain weight was observed in Ezhome-2 and low in IR-29. Based on visual scoring Vyttila-6 was classified as highly tolerant and IR-29 as highly susceptible. Correlation studies indicated that grain yield per plant was positively correlated with productive tillers per plant and seed setting per cent, while it was found to be negatively correlated with sterility per cent. Visual scoring was negatively correlated with plant height, uppermost internodal length, days to maturity and seed setting per cent. The hybrid CORH-3 was found to be moderately tolerant based on visual scoring and it produced highest yield per plant indicating that this hybrid is better than saline tolerant traditional varieties.Item Validation of identified genes for water stress in rice (Oryza sativa L.) mediated by Pseudomonas fluorescens(Centre for Plant Biotechnology and Molecular Biology, College of Horticulture, Vellanikkara, 2016) Saakre Manjesh; Abida, P SRice is the staple food for an expansive part of the world, particularly Asia. It is grown under diverse water regimes ranging from poorly drained waterlogged situations to well drain or water stressed upland conditions. Drought is one of the major abiotic stresses that adversely affect crop plants limiting their growth and yield potential. It disrupts the ionic and osmotic equilibrium of the cell. Susceptibility to water stress in rice is more pronounced at the reproductive stage and causes the greatest reduction in yield when stress coincides with the irreversible reproductive processes. Transcriptome analysis was carried out in Matta Triveni (PTB 45) to understand the influence of PGPR under water stress tolerance. Matta Triveni is a popular short duration (100-105 days) rice variety of Kerala, recommended for rainfed ecosystem especially in lowlands during the first crop and irrigated conditions during summers (third crop). With growing emphasis on expanding area under upland rice in the state, the need for an early, profusely tillering, drought tolerant variety is on the rise. Matta Triveni though high yielding is found to be adversely affected by drought. Hence, the present study was performed to assess the impact of application of Pseudomonas fluorescens in inducing tolerance to drought in variety Matta Triveni. In the present study the already identified differentially expressed genes such as Cytochrome oxidase subunit I (CO I), Protein kinase domain protein (PKDP), bZIPl (basic Leucine Zipper), AP2-EREBP (APETALA2 and ethylene-responsive element binding protein), Hsp20 (Heat shock protein 20) and COC1 (Circadian oscillator component) which are involved in ABA dependent and independent signal transduction pathways under water stress were validated using real-time PCR. Seedlings of rice variety Matta Triveni were given three treatments. Unstressed plants were maintained as absolute control. Water was withheld in second treatment and in the third treatment water was withheld in plants for which Pseudomonas fluorescens Pf1(KAU strain) was applied. Plants were given two applications of Pseudomonas fluorescens(pf) i.e., seed treatment and foliar spray. Water was withheld continuously for 15 days during reproductive stage (panicle initiation stage) after foliar spray. Plants exhibited leaf rolling which was scored as per IRRI, SES and was used for total RNA isolation. Biometric parameters were recorded after harvest. Total RNA was isolated from 65 days old rice plants after drought induction using TRIzol reagent. The first strand cDNA was synthesized by reverse transcriptase PCR from the above RNA samples. The cDNA was normalized using actin, which is a housekeeping gene. The real-time PCR primers were designed for six differentially expressed genes such as COI, PKDP, bZIPl, AP2-EREBP, Hsp20 and COC1 using IDT PrimerQuest software and the designed primers were validated using IDT OligoAnalyzer 3.1 tool. The target genes were analyzed by keeping actin as endogenous control. Comparative Δ Ct method was used to analyze the expression levels of genes. Results revealed that all these genes were upregulated in water stressed + Pf treated plants. COI,PKDP, bZIPl, AP2-EREBP, Hsp20 and COC1 genes were found to have 2.3, 2, 6, 2.8, 4.5 and 4 fold increases in relative expression levels respectively in the same treatment as compared to control. In-silico analysis was performed to identify the Transcription Factor Binding Sites (TFBSs) and combinatorial cis-regulatory elements for COI, PKDP, bZIP, AP2- EREBP, Hsp20 and MYB genes which are expressed in treatments. TFBSs were identified by AthaMap database, a genome-wide map of TFBSs in Arabidopsis thaliana, and STIFDB2 database which is a comprehensive collection of biotic and abiotic stress responsive genes in Arabidopsis and Oryza sativa. The significant TFBSs were analyzed based on the parameters provided by databases and were cross validated. The results revealed that the WRKY, MYB, HSF, bZIP, ARF, AP2/EREBP, bHLH, Trihelix and ABI3/VP1 TF families and their respective regulatory elements were predicted as functionally significant. The combinatorial cis- regulatory elements for three genes viz, CO 7, AP2-EREBP and Hsp20 were also identified. The transcription factors belonging to MYB, WRKY, bHLH and Trihelix families which are mainly responsible for drought stress signals and were predicted as co-associated. The study had shown that gene expression in plants lacking adequate water can be remarkably influenced by microbial colonization. The activation of the ABA dependent signaling pathway induced by colonization of P. fluorescens might be a key element for induced systemic tolerance. The predicted TFBSs and combinatorial cis-regulatory elements would be responsible for the upregulation of genes under water stress.Item Assessment of soil temperature and soil parameters on the population and functional efficiency of pseudomonas fluorescens in the rhizosphere of pokkali rice (Oryza sative L.)(Department of Agricultural Microbiology, College of Agriculture, Vellayani, 2018) Reshma Francis; Surendra Gopal, KPokkali is a umque variety of rice that is cultivated organically in coastal wetlands of Kerala. Pokkali fields are situated close to the sea and therefore prone to flooding and salinity. Pokkali rice has enormous potential for tolerating most of the abiotic stresses. It is the only economic crop which can be grown in waterlogged environment while tolerating salinity. The present study on “Assessment of soil temperature and soil parameters on the population and functional efficiency of Pseudomonas fluorescens in the rhizosphere of Pokkali rice (Oryza sativa L.)”, was conducted in Rice Research Station, Vyttila, under Kerala Agricultural University during 2016 to 2018. Two plots (40 m2) were used for the study. One of the plots was maintained as control (without application of Pseudomonas fluorescens) and another plot with rice was treated with P. fluorescens. P. fluorescens (KAU) was applied as seed treatment (10g kg-1) just before sowing and soil application (2.5 kg ha-1) at 1 week after dismantling. The main objectives were to study the effect of soil temperature and soil parameters on the population and functional efficiency of P. fluorescens / fluorescent Pseudomonas in Pokkali rice. The rhizosphere soils of Pokkali rice were collected at monthly interval from June, 2017 to October, 2017. The population of fluorescent Pseudomonas sp. in the rhizosphere soils of Pokkali rice were recorded at monthly interval. The population was absent before the start of the experiment in treated plot. However, the highest population 3.3x102 cfu g-l) was recorded at 90 DAS (August, 2017). However, the lowest population was recorded at 60 DAS (July, 2017). The population was not found at 30 DAS (June, 2017), 120 DAS (September, 2017) and 150 DAS (October, 2017). In the case of control plot, population of fluorescent Pseudomonas was absent before the start of experiment. At 30 DAS (June, 2017) population of fluorescent Pseudomonas was 3.3 x 102 cfu g-1. However, no fluorescent Pseudomonas were found at 60 DAS (July, 2017), 90 DAS (August, 2017), 120 DAS (September, 2017) and 150 DAS (October, 2017). Out of the six isolates (VPJU, VPJL, VPAU1, VPAU2, VPAU3 and VPAU4) of fluorescent Pseudomonas obtained, all the isolates produced IAA and ammonia with varied intensity. Three isolates (VPAU1, VPAU3 and VPAU4) produced HCN and none of the isolates showed siderophore production. The correlation studies between soil temperature, soil pH, EC and C02 evolution with population, revealed that the population of fluorescent Pseudomonas was affected by soil pH and soil EC. However, soil temperature and C02 evolution did not affect the population of fluorescent Pseudomonas. The functional efficiency of the fluorescent Pseudomonas were correlated with soil temperature, soil pH, EC and C02 evolution It was found that soil temperature, soil pH, EC and C02 evolution did not affect the functional efficiency of fluorescent Pseudomonas. All the six isolates obtained m the study were identified and confirmed through 168 rDNA sequencing. The isolates VPAJU, VPAU1 and VPAU2 were identified as Pseudomonas sp. and VPAJL, VPAU3 and VPAU4 isolates were found to be Pseudomonas aeruginosa. The present studies indicated that the inoculated P.fluorescens did not survive in Pokkali fields. The population of fluorescent Pseudomonas showed negative correlation with soil pH and EC. However, soil temperature and C02 evolution did not had any effect on population. The functional efficiency of fluorescent Pseudomonas was not affected by soil temperature, soil pH, EC and C02 evolution. However, further studies on influence of soil edaphic factors and weather variables on host plant and P. fluorescens needs to be studied and develop a suitable PGPR for Pokkali rice.Item Exploitation of abiotic stress tolerant strains of trichoderma spp. for the management of soil borne fungal pathogens(Department of Plant Pathology, College of Horticulture, Vellanikkara, 2018) Stella Doncy, P P; Reshmy VijayaraghavanSoil borne phytopathogenic fungi are known to cause severe yield loss of several crops. To mitigate these crop ailments, farmers mostly rely on chemical methods of disease control such as fungicides and other pesticides which are deleterious to the environment. To date, biological control is an ecofriendly approach for the effective management of crop diseases. The fungi belonging to the genus Trichoderma are one among the most exploited biocontrol agents in the field of agriculture. However, the performance of Trichoderma spp. gets reduced when it is exposed to abiotic stressed conditions such as drought, high temperature, salinity, acidity and fungicides. Hence, this study was proposed to identify and exploit stress tolerant isolates of Trichoderma spp. with antagonistic potential in Kerala. Intensive soil sampling surveys were conducted across different stressed ecosystems of Kerala viz., Palakkad, Alappuzha, Vytilla, Kumarakom, Wayanad and Thrissur for the isolation and enumeration of native Trichoderma spp. A total of 24 isolates were obtained from 52 soil samples collected from different locations. Based on the number of Trichoderma spp. obtained from each district, they were serially numbered and abbreviated according to the name of the location. Accordingly, PAT 1 to PAT 6 represents number of isolates of Trichoderma spp. from Palakkad district, ALT 1 to ALT 3 from Alappuzha, VYT 1 and VYT 2 from Ernakulam district, KUT 1 from Kottayam district, WAT 1 to WAT 8 from Wayanad and THT 1 to THT 4 from Thrissur district. Cultural and morphological identification of these isolates were carried out under in vitro conditions. Isolates of Trichoderma spp. were subjected to in vitro screening for abiotic stress tolerance such as high temperature, drought, acidity, salinity and also to test their sensitivity towards copper fungicides. The isolates PAT6 and WAT2 were found as thermotolerant, VYT2 and ALT 1 as drought tolerant, ALT 3 and ALT 1 as acid tolerant and saline tolerant and the isolates ALT1, ALT3 and PAT 1 as copper fungicide tolerant. The selected six isolates were further subjected to biochemical tests and the study showed that the isolates VYT 2, ALT 3 and ALT 1 showed highest cellulase, β- 1, 3 glucanase and protease activity. Likewise, isolates PAT 1, ALT 1 and ALT 3 were found as best producers of ACC deaminase and PAT 1 and ALT 1 as the best cytokinin producer. The best performing isolates (ALT 1, ALT 3, PAT 1, PAT 6 and VYT 2) after enzyme study were subjected to dual culture experiment with five major soil borne pathogens to test their antagonistic potential. The isolates ALT 3, ALT 1, PAT 6 and PAT 1 showed more than 70 per cent inhibition of R. solani whereas, isolates ALT1 and VYT 2 showed only 57.78 and 55 per cent inhibition of S. rolfsii respectively. However, no significant difference was noticed among the isolates when grown against F. solani. Cent per cent inhibition of P. aphanidermatum was noticed with Trichoderma isolates PAT 1 and ALT 1. All five isolates showed 100 per cent inhibition on the growth of pathogen, P. capsici. Among the five, four isolates viz., ALT 1, ALT 3, PAT 1 and PAT 6 with best antagonistic potential were subjected to molecular characterization at Rajiv Gandhi Centre for Biotechnology, Thiruvananthapuram and the isolates showed homology with the nucleotide sequence of Trichoderma asperellum. A pot culture experiment was laid with the selected isolates viz., ALT 1, ALT 3, PAT 1 and PAT 6 to test the growth promotion of cowpea and biocontrol efficacy against Rhizoctonia solani. It was observed that the isolate PAT 6 coated seeds showed 100 per cent germination and also recorded better biometric characters and yield. Moreover, the lowest per cent disease incidence of 11.11 per cent was only recorded with both the isolates ALT 3 and PAT 6. Thus, the study has enlightened our knowledge on the existence of abiotic stress tolerant isolates of T. asperellum which can be employed in future for the biocontrol of soil borne pathogens in such conditions.Item Oxidative stress and protein profiling in sweet potato (Ipomoeabatatas (L.) Lam.) under abiotic stresses(Department of Plant Biotechnology, College of Agriculture, Vellayani, 2017) Aiswarya, N S; Ravi, VThe present study an “Oxidative stress and protein profiling in sweet potato (Ipomoeabatatas (L.) Lam.) under abiotic stresses” was conducted during the period 2014-2015 in the Division of Crop Production, Central Tuber Crops Research Institute (CTCRI) Thiruvananthapuram. The objective of the study was to elicit information on the antioxidative enzyme activity, protein profiling, growth parameter and yield parameter of five varieties / genotypes of sweet potato under irrigated high temperature stress (HTS) (40oC during the day time) and water deficit stress (WDS) ( 2nd and 3rd month after planting) conditions and identify marker physiological parameter which can be used for identifying / developing sweet potato varieties for tolerance to abiotic stress conditions. The study indicated that the vein length of five different sweet potato varieties/ genotypes was not significantly affected by HTS and WDS. The leaf production of five sweet potato varieties was significantly affected by both HTS and WDS. The leaf retention of five varieties / genotype was differentially affected by both heat and water stress. There was negligible difference in yield parameters between stress and control condition. However, there was significant difference in yield observed for parameters among five varieties. The effect of abiotic stress on various physiological parameters like Relative water content (RWC), chlorophyll and carotenoid content significantly differed in control plant. Among the WDS and HTS treatments, the chlorophyll and carotenoid content decreased in all the five varieties. Chlorophyll and carotenoid content were significantly reduced in plants exposed to HTS. RWC of leaves varied under both WDS and HTS conditions. A higher RWC was observed in plants exposed to HTS condition compared to plants was WDS condition. Total soluble protein content of leaf was found to be lower due to presence of high mucilage in leaves. Total soluble protein in tuber is increased in control and WDS conditions. Total soluble protein in tuber forming roots had increased under stress free condition. The effect of abiotic stress on biological parameter like proline status of plants under stress conditions followed an increasing trend compared to stress free condition. The outcome of abiotic stress on various enzymatic antioxidant such as superoxide dismutase (SOD), catalase (CAT), ascorbate peroxidase (APX), ascorbate reductase, glutathione reductase (GR) was studied. Among the five different sweet potato varieties/ genotypes (Sree Arun, S-1464, S-1466, Sree Kanaka and Kanjangad) S-1466 and Sree Kanaka had maximum GR activity under HTS compared to WDS and control condition. Catalase activity of five varieties/ genotypes of sweet potato plant varied among different stress condition (WDS and HTS) compared to control. The GR activity was significantly reduced due to stress condition and SOD activity increased under stress conditions. The APX enzyme activity is more under WDS conditions compared to HTS and control conditions. Ascorbate reductase become there was no change in absorbance therefore the enzyme activity could not be measured. The AA content in sweet potato leaves increased under WDS condition compared to HTS and control. The glutathione content in sweet potato leaves was negligible or below detectable amount under both control and stress condition. Under HTS condition SOD activity increased in Sree Arun and S-1466 alone whereas other varieties/genotypes had increased SOD activity under WDS. Kanjangad had maximum APX activity in HTS whereas S-1464 had higher APX activity in WDS. From the present study, it is inferred that SOD activity can be utilised as biochemical marker for developing HTS tolerant varieties as it showed consistent changes under both WDS and HTS. APX activity can be utilised as marker for identifying WDS tolerant varieties. Detailed molecular biochemical and physiological information will be necessary using different varieties for getting appropriate activities as the marker to screen abiotic stress tolerant varieties.Item Climate change adaptation through improved water use efficiency in rice (Oryza sativa L.)(Academy of Climate Change Education and Research Vellanikkara, 2016) Anjaly C Bose; Santhoshkumar, A VThe food security of more than half of the world population depends on rice. Studies suggest that global climate change is going to affect the food production through temperature and water stress and this affect the rice production around the globe. The present study tried to elucidate the influence of varying soil moisture status on rice productivity and evaluate the strategies for increased water use efficiency in a climate change adaptation strategy. The study was conducted during May 2016-September 2016 at RARS, Pattambi in variety Jyothi. The treatment combination included the presence or absence of hydrogel along with 4 different levels of irrigation (IW/CPE=2, IW/CPE=1.5, IW/CPE=1 and IW/CPE=0.5). The results showed that the various irrigation levels and hydrogel application had a significant impact on the physiology of rice. Hydrogel application improved the soil moisture availability and increased plant establishment. The maximum plant height was observed for the treatment IW/CPE=2 (105.30 cm) without hydrogel. The hydrogel effect on plant height was significant only up to the booting stage. Hydrogel had its significance on number of tillers only at the vegetative stage of the plant, while, interaction was significant at the vegetative, reproductive and ripening stages. The higher value (19.67) of tiller number was recorded for the treatment IW/CPE=1.5 with hydrogel. LAI was not affected by the application of hydrogel. Only the irrigation treatments had a significant effect on LAI, of which the treatments IW/CPE=2 (2.72) and IW/CPE=1.5 (2.61) recorded the maximum LAI. Higher number of primary branches per panicle was recorded for plants with hydrogel (10.25). The number of panicle per hill was more for the treatment IW/CPE=1.5 without hydrogel (9.20). The number of filled grains produced per panicle is more for plants with hydrogel (86.00). 1000 grain weight observed was higher for the treatment IW/CPE=2 (27.23 g) without hydrogel. Hydrogel did not have any significant effect on the plants physiological parameters like booting, heading, flowering, number of days taken for active tillering and panicle initiation. The more stressed plants took the maximum number of days to booting, heading, flowering and panicle initiation. For the treatment IW/CPE=0.5, there seen no sign of 50 percent flowering and consequently, it did not attained physiological maturity. Hydrogel and irrigation had a significant impact on grain yield. Even though the higher yield (7014.63 kg ha-1) was observed for the irrigation level IW/CPE=2 without hydrogel, the mean average value of grain yield of plants treated with hydrogel is higher than plants treated without hydrogel (4455.03 kg ha-1 and 3951.80 kg ha-1 for with and without hydrogel). It can be concluded that hydrogel had significance only when the irrigation level was low (IW/CPE=1.5 and IW/CPE=1). However, at extreme low water level (IW/CPE=0.5) and high water level (IW/CPE=2), hydrogel failed to exhibit any beneficial role. Under the projected climate scenario using RCP 4.5, it was found for the year 2030 the maximum yield was observed for the treatment IW/CPE=2 (6010 kg ha-1), followed by comparable yield in the treatment IW/CPE=1.5 (5997 kg ha- 1 ). The production was found to be less in the treatment IW/CPE=1 (3504 kg ha-1) and nil to the treatment IW/CPE=0.5. For the year 2050 and 2080, the maximum yield was for the treatment IW/CPE=2.Item Efficacy of arbuscular mycorrhizal fungi for drought tolerance in Swietenia macrophylla King. seedlings(Department of Tree Physiology and Breeding, College of Forestry, Vellanikkara, 2016) Lakshmy J Rajan; Santhoshkumar, A VAn experiment was carried out to analyze the influence of four different levels of water stress ((IW (irrigation water)/ET(cumulative crop evapotranspiration)=1, 0.8, 0.6 and 0.4) and three different species of arbuscular mycorrhizal fungi (AMF), viz; Funneliformis mossae, Glomus etunicatum, and Rhizophagus intraradices on the growth and development of Swietenia macrophylla seedlings in nursery. Seedlings that were irrigated daily and the seedlings not inoculated with AMF were maintained as controls. Drought stress was found to affect the growth and physiology of mahogany seedlings significantly.