Search results for: biotic and abiotic stresses
Commenced in January 2007
Frequency: Monthly
Edition: International
Paper Count: 891

Search results for: biotic and abiotic stresses

861 Effects of Temperature and the Use of Bacteriocins on Cross-Contamination from Animal Source Food Processing: A Mathematical Model

Authors: Benjamin Castillo, Luis Pastenes, Fernando Cerdova

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The contamination of food by microbial agents is a common problem in the industry, especially regarding the elaboration of animal source products. Incorrect manipulation of the machinery or on the raw materials can cause a decrease in production or an epidemiological outbreak due to intoxication. In order to improve food product quality, different methods have been used to reduce or, at least, to slow down the growth of the pathogens, especially deteriorated, infectious or toxigenic bacteria. These methods are usually carried out under low temperatures and short processing time (abiotic agents), along with the application of antibacterial substances, such as bacteriocins (biotic agents). This, in a controlled and efficient way that fulfills the purpose of bacterial control without damaging the final product. Therefore, the objective of the present study is to design a secondary mathematical model that allows the prediction of both the biotic and abiotic factor impact associated with animal source food processing. In order to accomplish this objective, the authors propose a three-dimensional differential equation model, whose components are: bacterial growth, release, production and artificial incorporation of bacteriocins and changes in pH levels of the medium. These three dimensions are constantly being influenced by the temperature of the medium. Secondly, this model adapts to an idealized situation of cross-contamination animal source food processing, with the study agents being both the animal product and the contact surface. Thirdly, the stochastic simulations and the parametric sensibility analysis are compared with referential data. The main results obtained from the analysis and simulations of the mathematical model were to discover that, although bacterial growth can be stopped in lower temperatures, even lower ones are needed to eradicate it. However, this can be not only expensive, but counterproductive as well in terms of the quality of the raw materials and, on the other hand, higher temperatures accelerate bacterial growth. In other aspects, the use and efficiency of bacteriocins are an effective alternative in the short and medium terms. Moreover, an indicator of bacterial growth is a low-level pH, since lots of deteriorating bacteria are lactic acids. Lastly, the processing times are a secondary agent of concern when the rest of the aforementioned agents are under control. Our main conclusion is that when acclimating a mathematical model within the context of the industrial process, it can generate new tools that predict bacterial contamination, the impact of bacterial inhibition, and processing method times. In addition, the mathematical modeling proposed logistic input of broad application, which can be replicated on non-meat food products, other pathogens or even on contamination by crossed contact of allergen foods.

Keywords: bacteriocins, cross-contamination, mathematical model, temperature

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860 Livability and Growth Performance of Noiler Chickens Fed with Different Biotic Additives

Authors: Idowu Kemi Ruth, Adeyemo Adedayo Akinade, Iyanda Adegboyega Ibukun, Idowu Olubukola Precious Akinade

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Liveability and mortality rate is a germane aspect of product performance that cannot be overlooked in poultry production, while the disease is a major threat in the poultry industry which can cause a major loss for the farmer and a reduction in the total income generated from the stock. Therefore, efforts must be made to enhance the health status of chickens to reduce mortality. The study was conducted to investigate the effect of different biotic additives (prebiotic, probiotic and synbiotic ) on the performance of Noiler females at the growing phase (forty-nine days) till the point of the first egg across the biotic additive. A total of one hundred and twenty-eight female Noiler were used for the experiment. Experimental treatment consisted of prebiotic, probiotic, synbiotic and control at the inclusion rate of a gram into a kilogram of feed. Parameters measured are Feed intake, feed conversion ratio, the weight of the first egg, age of the first egg and livability. Data collected were subjected to a one-way analysis of variance. The result obtained revealed a better growth performance across the treatments than the control group with the least final weight at nineteen weeks of point of lay. Prebiotic treatment had the best age at first lay on day one hundred and thirty seven followed by other treatments on day one hundred and fifty four. However, the size of the eggs was not significantly influenced by the biotic additive. Hence, the experiment can be concluded that the inclusion of different biotic additives influenced the growth performance; likewise, the Prebiotic had a significant effect on the age of first laying in Noiler chicken, and livability was a hundred percent throughout the duration of the experiment.

Keywords: prebiotic, probiotic, synbiotic, noiler

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859 Evolution of DNA-Binding With-One-Finger Transcriptional Factor Family in Diploid Cotton Gossypium raimondii

Authors: Waqas Shafqat Chattha, Muhammad Iqbal, Amir Shakeel

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Transcriptional factors are proteins that play a vital role in regulating the transcription of target genes in different biological processes and are being widely studied in different plant species. In the current era of genomics, plant genomes sequencing has directed to the genome-wide identification, analyses and categorization of diverse transcription factor families and hence provide key insights into their structural as well as functional diversity. The DNA-binding with One Finger (DOF) proteins belongs to C2-C2-type zinc finger protein family. DOF proteins are plant-specific transcription factors implicated in diverse functions including seed maturation and germination, phytohormone signalling, light-mediated gene regulation, cotton-fiber elongation and responses of the plant to biotic as well as abiotic stresses. In this context, a genome-wide in-silico analysis of DOF TF family in diploid cotton species i.e. Gossypium raimondii has enabled us to identify 55 non-redundant genes encoding DOF proteins renamed as GrDofs (Gossypium raimondii Dof). Gene distribution studies have shown that all of the GrDof genes are unevenly distributed across 12 out of 13 G. raimondii chromosomes. The gene structure analysis illustrated that 34 out of 55 GrDof genes are intron-less while remaining 21 genes have a single intron. Protein sequence-based phylogenetic analysis of putative 55 GrDOFs has divided these proteins into 5 major groups with various paralogous gene pairs. Molecular evolutionary studies aided with the conserved domain as well as gene structure analysis suggested that segmental duplications were the principal contributors for the expansion of Dof genes in G. raimondii.

Keywords: diploid cotton , G. raimondii, phylogenetic analysis, transcription factor

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858 Predicting Stem Borer Density in Maize Using RapidEye Data and Generalized Linear Models

Authors: Elfatih M. Abdel-Rahman, Tobias Landmann, Richard Kyalo, George Ong’amo, Bruno Le Ru

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Maize (Zea mays L.) is a major staple food crop in Africa, particularly in the eastern region of the continent. The maize growing area in Africa spans over 25 million ha and 84% of rural households in Africa cultivate maize mainly as a means to generate food and income. Average maize yields in Sub Saharan Africa are 1.4 t/ha as compared to global average of 2.5–3.9 t/ha due to biotic and abiotic constraints. Amongst the biotic production constraints in Africa, stem borers are the most injurious. In East Africa, yield losses due to stem borers are currently estimated between 12% to 40% of the total production. The objective of the present study was therefore to predict stem borer larvae density in maize fields using RapidEye reflectance data and generalized linear models (GLMs). RapidEye images were captured for a test site in Kenya (Machakos) in January and in February 2015. Stem borer larva numbers were modeled using GLMs assuming Poisson (Po) and negative binomial (NB) distributions with error with log arithmetic link. Root mean square error (RMSE) and ratio prediction to deviation (RPD) statistics were employed to assess the models performance using a leave one-out cross-validation approach. Results showed that NB models outperformed Po ones in all study sites. RMSE and RPD ranged between 0.95 and 2.70, and between 2.39 and 6.81, respectively. Overall, all models performed similar when used the January and the February image data. We conclude that reflectance data from RapidEye data can be used to estimate stem borer larvae density. The developed models could to improve decision making regarding controlling maize stem borers using various integrated pest management (IPM) protocols.

Keywords: maize, stem borers, density, RapidEye, GLM

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857 Towards Development of Superior Brassica juncea by Pyramiding of Genes of Diverse Pathways for Value Addition, Stress Alleviation and Human Health

Authors: Deepak Kumar, Ravi Rajwanshi, Mohd. Aslam Yusuf, Nisha Kant Pandey, Preeti Singh, Mukesh Saxena, Neera Bhalla Sarin

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Global issues are leading to concerns over food security. These include climate change, urbanization, increase in population subsequently leading to greater energy and water demand. Futuristic approach for crop improvement involves gene pyramiding for agronomic traits that empower the plants to withstand multiple stresses. In an earlier study from the laboratory, the efficacy of overexpressing γ-tocopherol methyl transferase (γ-TMT) gene from the vitamin E biosynthetic pathway has been shown to result in six-fold increase of the most biologically active form, the α-tocopherol in Brassica juncea which resulted in alleviation of salt, heavy metal and osmoticum induced stress by the transgenic plants. The glyoxalase I (gly I) gene from the glyoxalase pathway has also been earlier shown by us to impart tolerance against multiple abioitc stresses by detoxification of the cytotoxic compound methylglyoxal in Brassica juncea. Recently, both the transgenes were pyramided in Brassica juncea lines through sexual crosses involving two stable Brassica juncea lines overexpressing γ-TMT and gly I genes respectively. The transgene integration was confirmed by PCR analysis and their mRNA expression was evident by RT-PCR analysis. Preliminary physiological investigations showed ~55% increased seed germination under 200 mM NaCl stress in the pyramided line and 81% higher seed germination under 200 mM mannitol stress as compared to the WT control plants. The pyramided lines also retained more chlorophyll content when the leaf discs were floated on NaCl (200, 400 and 600 mM) or mannitol (200, 400 and 600 mM) compared to the WT control plants. These plants had higher Relative Water Content and greater solute accumulation under stress compared to the parental plants having γ-TMT or the glyI gene respectively. The studies revealed the synergy of two components from different metabolic pathways in enhancing stress hardiness of the transgenic B. juncea plants. It was concluded that pyramiding of genes (γ-TMT and glyI) from diverse pathways can lead to enhanced tolerance to salt and mannitol stress (simulating drought conditions). This strategy can prove useful in enhancing the crop yields under various abiotic stresses.

Keywords: abiotic stress, brassica juncea, glyoxalase I, α-tocopherol

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856 The Molecular Analysis of Effect of Phytohormones and Spermidine on Tomato Growth under Biotic Stress

Authors: Rumana Keyani, Haleema Sadia, Asia Nosheen, Rabia Naz, Humaira Yasmin, Sidra Zahoor

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Tomato is a significant crop of the world and is one of the staple foods of Pakistan. A vast number of plant pathogens from simple viruses to complex parasites cause diseases in tomatoes but fungal infection in our country is quite high. Sometimes the symptoms are too harsh destroying the crop altogether. Countries like our own with continuously increasing massive population and limited resources cannot afford such an economic loss. There is an array of morphological, genetic, biochemical and molecular processes involved in plant resistance mechanisms to biotic stress. The study of different metabolic pathways like Jasmonic acid (JA) pathways and most importantly signaling molecules like ROS/RNS and their redoxin enzymes i.e. TRX and NRX is crucial to disease management, contributing to healthy plant growth. So, improving tolerance in crop plants against biotic stresses is a dire need of our country and world as whole. In the current study, fungal pathogenic strains Alternaria solani and Rhizoctonia solani were used to inoculate tomatoes to check the defense responses of tomato plant against these pathogens at molecular as well as phenotypic level with jasmonic acid and spermidine pretreatment. All the growth parameters (root and shoot length, dry and weight root, shoot weight measured 7 days post-inoculation, exhibited that infection drastically declined the growth of the plant whereas jasmonic acid and spermidine assisted the plants to cope up with the infection. Thus, JA and Spermidine treatments maintained comparatively better growth factors. Antioxidant assays and expression analysis through real time quantitative PCR following time course experiment at 24, 48 and 72 hours intervals also exhibited that activation of JA defense genes and a polyamine Spermidine helps in mediating tomato responses against fungal infection when used alone but the two treatments combined mask the effect of each other.

Keywords: fungal infection, jasmonic acid defence, tomato, spermidine

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855 Phylogenetic Relationships between the Whole Sets of Individual Flow Sorted U, M, S and C Chromosomes of Aegilops and Wheat as Revealed by COS Markers

Authors: András Farkas, István Molnár, Jan Vrána, Veronika Burešová, Petr Cápal, András Cseh, Márta Molnár-Láng, Jaroslav Doležel

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Species of Aegilops played a central role in the evolution of wheat and are sources of traits related to yield quality and tolerance against biotic and abiotic stresses. These wild genes and alleles are desirable to use in crop improvement programs via introgressive hybridization. However, the success of chromosome mediated gene transfer to wheat are hampered by the pour knowledge on the genome structure of Aegilops relative to wheat and by the low number of cost-effective molecular markers specific for Aegilops chromosomes. The COS markers specific for genes conserved throughout evolution in both sequence and copy number between Triticeae/Aegilops taxa and define orthologous regions, thus enabling the comparison of regions on the chromosomes of related species. The present study compared individual chromosomes of Aegilops umbellulata (UU), Ae. comosa (MM), Ae. speltoides (SS) and Ae. caudata (CC) purified by flourescent labelling with oligonucleotid SSR repeats and biparametric flow cytometry with wheat by identifying orthologous chromosomal regions by COS markers. The linear order of bin-mapped COS markers along the wheat D chromosomes was identified by the use of chromosome-specific sequence data and virtual gene order. Syntenic regions of wheat identifying genome rearrangements differentiating the U, M, S or C genomes from the D genome of wheat were detected. The conserved orthologous set markers assigned to Aegilops chromosomes promise to accelerate gene introgression by facilitating the identification of alien chromatin. The syntenic relationships between the Aegilops species and wheat will facilitate the targeted development of new markers specific for U, M, S and C genomic regions and will contribute to the understanding of molecular processes related to the evolution of Aegilops.

Keywords: Aegilops, cos-markers, flow-sorting, wheat

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854 Laser Shock Peening of Additively Manufactured Nickel-Based Superalloys

Authors: Michael Munther, Keivan Davami

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One significant roadblock for additively manufactured (AM) parts is the buildup of residual tensile stresses during the fabrication process. These residual stresses are formed due to the intense localized thermal gradients and high cooling rates that cause non-uniform material expansion/contraction and mismatched strain profiles during powder-bed fusion techniques, such as direct metal laser sintering (DMLS). The residual stresses adversely affect the fatigue life of the AM parts. Moreover, if the residual stresses become higher than the material’s yield strength, they will lead to acute geometric distortion. These are limiting the applications and acceptance of AM components for safety-critical applications. Herein, we discuss laser shock peening method as an advanced technique for the manipulation of the residual stresses in AM parts. An X-ray diffraction technique is used for the measurements of the residual stresses before and after the laser shock peening process. Also, the hardness of the structures is measured using a nanoindentation technique. Maps of nanohardness and modulus are obtained from the nanoindentation, and a correlation is made between the residual stresses and the mechanical properties. The results indicate that laser shock peening is able to induce compressive residual stresses in the structure that mitigate the tensile residual stresses and increase the hardness of AM IN718, a superalloy, almost 20%. No significant changes were observed in the modulus after laser shock peening. The results strongly suggest that laser shock peening can be used as an advanced post-processing technique to optimize the service lives of critical components for various applications.

Keywords: additive manufacturing, Inconel 718, laser shock peening, residual stresses

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853 Increase of Quinoa Tolerance to High Salinity Involves Agrophysiological Parameters Improvement by Soil Amendments

Authors: Bourhim Mohammad Redouane, Cheto Said, Qaddoury Ahmed, Hirich Abdelaziz, Ghoulam Cherki

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Several abiotic stresses cause disruptions in the properties of agricultural soils and hence their loss worldwide. Among these abiotic stresses, Salinity to which most crops were exposed caused an important reduction in their productivity. Therefore, in order to deal with this challenging problem, we rely on cultivating alternative plants that can tolerate the adverse salinity stress, such as quinoa (Chenopodium quinoa). Although even it was qualified as tolerant to Salinity, the quinoa’s performance could be negatively affected under high salinity levels. Thus, our study aims to assess the effects of the application of soil amendments to improve quinoa tolerance levels under high Salinity. Thus, three quinoa varieties (Puno, ICBA-Q5, and Titicaca) were grown on agricultural soil under a greenhouse with five amendments; Biochar “Bc,” compost “Cp,” black soldier insect frass “If,” cow manure “Fb” and phosphogypsum “Pg.” Two controls without amendment were adopted consisting of the salinized negative one “T(-)” and the non-salinized positive one “T(+).” After 20 days from sowing, the plants were irrigated with a saline solution of 16 dS/m prepared with NaCl for a period of 60 days. Then plant tolerance was assessed based on agrophysiological parameters. The results showed that salinity stress negatively affected the quinoa plants for all the analyzed agrophysiological parameters in the three varieties compared to their corresponding controls “T(+).” However, most of these parameters were significantly enhanced by the application of soil amendments compared to their negative controls “T(-).” For instance, the biomass was improved by 91.8% and 69.4%, respectively, for Puno and Titicaca varieties amended with “Bc.” The total nitrogen amount was increased by 220% for Titicaca and ICBA-Q5 plants cultivated in the soil amended with “If.” One of the most important improvements was noted for potassium content in Titicaca amended with “Pg,” which was six times higher compared to the negative control. Besides, the plants of Puno amended with “Cp” showed an improvement of 75.9% for the stomatal conductance and 58.5% for nitrate reductase activity. Nevertheless, the pronounced varietal difference was registered between Puno and Titicaca, presenting the highest performances mainly for the soil amended with “If,” “Bc,” and “Pg.”

Keywords: chenopodium quinoa, salinity, soil amendments, growth, nutrients, nitrate reductase

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852 Phi Thickening Induction as a Response to Abiotic Stress in the Orchid Miltoniopsis

Authors: Nurul Aliaa Idris, David A. Collings

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Phi thickenings are specialized secondary cell wall thickenings that are found in the cortex of the roots in a wide range of plant species, including orchids. The role of phi thickenings in the root is still under debate through research have linked environmental conditions, particularly abiotic stresses such as water stress, heavy metal stress and salinity to their induction in the roots. It has also been suggested that phi thickenings may act as a barrier to regulate solute uptake, act as a physical barrier against fungal hyphal penetration due to its resemblance to the Casparian strip and play a mechanical role to support cortical cells. We have investigated phi thickening function in epiphytic orchids of the genus Miltoniopsis through induction experiment against factors such as soil compaction and water stress. The permeability of the phi thickenings in Miltoniopsis was tested through uptake experiments using the fluorescent tracer dyes Calcofluor white, Lucifer yellow and Propidium iodide then viewed with wide-field or confocal microscopy. To test whether phi thickening may prevent fungal colonization in the root cell, fungal re-infection experiment was conducted by inoculating isolated symbiotic fungus to sterile in vitro Miltoniopsis explants. As the movement of fluorescent tracers through the apoplast was not blocked by phi thickenings, and as phi thickenings developed in the roots of sterile cultures in the absence of fungus and did not prevent fungal colonization of cortical cells, the phi thickenings in Miltoniopsis do not function as a barrier. Phi thickenings were found to be absent in roots grown on agar and remained absent when plants were transplanted to moist soil. However, phi thickenings were induced when plants were transplanted to well-drained media, and by the application of water stress in all soils tested. It is likely that phi thickenings stabilize the root cortex during dehydration. Nevertheless, the varied induction responses present in different plant species suggest that the phi thickenings may play several adaptive roles, instead of just one, depending on species.

Keywords: abiotic stress, Miltoniopsis, orchid, phi thickening

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851 Effects of Tensile Pre-Stresses on Corrosion Behavior of AISI 304 Stainless Steel in 1N H2SO4

Authors: Sami Ibrahim Jafar, Israa Abud Alkadir, Samah Abdul Kareem Khashin

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The aim of this work is to assess the influence of tensile pre-stresses on the microstructure and corrosion behavior of the AISI304 stainless steel in 1N H2SO4 austenitic stainless steel. Samples of this stainless steel either with pre-stresses, corresponding to [255, 305, 355, 405, 455, 505, 555, 605 and σf] MPa induced by tensile tests, or without pre-stresses (as received), were characterized regarding their microstructure to investigate the pre-tensile stress effects on the corrosion behavior. The results showed that the corrosion rate of elastic pre-stresses 304 stainless steel was very little increased compared with that of as received specimens. The corrosion rate increases after applying pre-stress between (σ255 - σ 455) MPa. The microstructure showed that the austenitic grains begin to deform in the direction of applied pre-stresses. The maximum hardness at this region was (229.2) Hv, but at higher pre-stress (σ455 – σ 605) MPa unanticipated occurrence, the corrosion rate decreases. The microstructure inspection shows the deformed austenitic grain and ά-martensitic phase needle are appeared inside austenitic grains and the hardness reached the maximum value (332.433) Hv. The results showed that the corrosion rate increases at the values of pre-stresses between (σ605 – σf) MPa., which is inspected the result. The necking of gauge length of specimens occurs in specimens and this leads to deterioration in original properties and the corrosion rate reaches the maximum value.

Keywords: tensile pre-stresses, corrosion rate, austenitic stainless steel, hardness

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850 Impact of Anthropogenic Stresses on Plankton Biodiversity in Indian Sundarban Megadelta: An Approach towards Ecosystem Conservation and Sustainability

Authors: Dibyendu Rakshit, Santosh K. Sarkar

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The study illustrates a comprehensive account of large-scale changes plankton community structure in relevance to water quality characteristics due to anthropogenic stresses, mainly concerned for Annual Gangasagar Festival (AGF) at the southern tip of Sagar Island of Indian Sundarban wetland for 3-year duration (2012-2014; n=36). This prograding, vulnerable and tide-dominated megadelta has been formed in the estuarine phase of the Hooghly Estuary infested by largest continuous tract of luxurious mangrove forest, enriched with high native flora and fauna. The sampling strategy was designed to characterize the changes in plankton community and water quality considering three diverse phases, namely during festival period (January) and its pre - (December) as well as post (February) events. Surface water samples were collected for estimation of different environmental variables as well as for phytoplankton and microzooplankton biodiversity measurement. The preservation and identification techniques of both biotic and abiotic parameters were carried out by standard chemical and biological methods. The intensive human activities lead to sharp ecological changes in the context of poor water quality index (WQI) due to high turbidity (14.02±2.34 NTU) coupled with low chlorophyll a (1.02±0.21 mg m-3) and dissolved oxygen (3.94±1.1 mg l-1), comparing to pre- and post-festival periods. Sharp reduction in abundance (4140 to 2997 cells l-1) and diversity (H′=2.72 to 1.33) of phytoplankton and microzooplankton tintinnids (450 to 328 ind l-1; H′=4.31 to 2.21) was very much pronounced. The small size tintinnid (average lorica length=29.4 µm; average LOD=10.5 µm) composed of Tintinnopsis minuta, T. lobiancoi, T. nucula, T. gracilis are predominant and reached some of the greatest abundances during the festival period. Results of ANOVA revealed a significant variation in different festival periods with phytoplankton (F= 1.77; p=0.006) and tintinnid abundance (F= 2.41; P=0.022). RELATE analyses revealed a significant correlation between the variations of planktonic communities with the environmental data (R= 0.107; p= 0.005). Three distinct groups were delineated from principal component analysis, in which a set of hydrological parameters acted as the causative factor(s) for maintaining diversity and distribution of the planktonic organisms. The pronounced adverse impact of anthropogenic stresses on plankton community could lead to environmental deterioration, disrupting the productivity of benthic and pelagic ecosystems as well as fishery potentialities which directly related to livelihood services. The festival can be considered as multiple drivers of changes in relevance to beach erosion, shoreline changes, pollution from discarded plastic and electronic wastes and destruction of natural habitats resulting loss of biodiversity. In addition, deterioration in water quality was also evident from immersion of idols, causing detrimental effects on aquatic biota. The authors strongly recommend for adopting integrated scientific and administrative strategies for resilience, sustainability and conservation of this megadelta.

Keywords: Gangasagar festival, phytoplankton, Sundarban megadelta, tintinnid

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849 Comparison of Different Methods of Evaluating Nozzle Junction Stresses under External Loads

Authors: Vinod Kumar, Arun Kumar, Surjit Angra

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This paper addresses the junction stress analysis of orthogonally intersecting thin walled cylindrical shell and thin walled cylindrical nozzle subjected to external loading on nozzle. Junction stresses have been calculated theoretically by welding research council (WRC) bulletins 107 and 297 for different nozzle loads. WRC bulletins 107 and 297 have been used by design engineers for calculating nozzle-vessel junction stresses since their publication. They give simple empirical relations and easy in application. Also 3D FEA in which material is elastic has been done in ANSYS software with 8 node solid element model and results of FEA have been compared with WRC results. Stress intensities obtained by WRC 297 are generally slightly higher than obtained by WRC 107. Membrane stresses obtained by FEA are much higher than WRC and membrane plus bending stresses obtained by FEA are lower than WRC.

Keywords: FEA, junction stress, solid element, WRC 107, WRC 297

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848 The Development of a Residual Stress Measurement Method for Roll Formed Products

Authors: Yong Sun, Vladimir Luzin, Zhen Qian, William J. T. Daniel, Mingxing Zhang, Shichao Ding

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The residual stresses in roll formed products are generally very high and un-predictable. This is due to the occurrence of redundant plastic deformation in roll forming process and it can cause various product defects. Although the residual stresses of a roll formed product consist of longitudinal and transverse residual stresses components, but the longitudinal residual stresses plays a key role to the product defects of a roll formed product and therefore, only the longitudinal residual stresses concerned by the roll forming scholars and engineers. However, how to inspect the residual stresses of a product quickly and economically as a routine operation is still a challenge. This paper introduces a residual stresses measurement method called slope cutting method to study the longitudinal residual stresses through layers geometrically to a roll formed products or a product with similar process such as a rolled sheet. The detailed measuring procedure is given and discussed. The residual stresses variation through the layer can be derived based on the variation of curvature in different layers and steps. The slope cutting method has been explored and validated by experimental study on a roll-formed square tube. The neutron diffraction method is applied to validate the accuracy of the newly proposed layering removal materials results. The two set results agree with each other very well and therefore, the method is expected to be a routine testing method to monitor the quality of a product been formed and that is a great impact to roll forming industry.

Keywords: roll forming, residual stress, measurement method, neutron diffraction

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847 Characterization of Transcription Factors Involved in Early Defense Response during Interaction of Oil Palm Elaeis guineensis Jacq. with Ganoderma boninense

Authors: Sakeh N. Mohd, Bahari M. N. Abdul, Abdullah S. N. Akmar

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Oil palm production generates high export earnings to many countries especially in Southeast Asian region. Infection by necrotrophic fungus, Ganoderma boninense on oil palm results in basal stem rot which compromises oil palm production leading to significant economic loss. There are no reliable disease treatments nor promising resistant oil palm variety has been cultivated to eradicate the disease up to date. Thus, understanding molecular mechanisms underlying early interactions of oil palm with Ganoderma boninense may be vital to promote preventive or control measure of the disease. In the present study, four months old oil palm seedlings were infected via artificial inoculation of Ganoderma boninense on rubber wood blocks. Roots of six biological replicates of treated and untreated oil palm seedlings were harvested at 0, 3, 7 and 11 days post inoculation. Next-generation sequencing was performed to generate high-throughput RNA-Seq data and identify differentially expressed genes (DEGs) during early oil palm-Ganoderma boninense interaction. Based on de novo transcriptome assembly, a total of 427,122,605 paired-end clean reads were assembled into 30,654 unigenes. DEGs analysis revealed upregulation of 173 transcription factors on Ganoderma boninense-treated oil palm seedlings. Sixty-one transcription factors were categorized as DEGs according to stringent cut-off values of genes with log2 ratio [Number of treated oil palm seedlings/ Number of untreated oil palm seedlings] ≥ |1.0| (corresponding to 2-fold or more upregulation) and P-value ≤ 0.01. Transcription factors in response to biotic stress will be screened out from abiotic stress using reverse transcriptase polymerase chain reaction. Transcription factors unique to biotic stress will be verified using real-time polymerase chain reaction. The findings will help researchers to pinpoint defense response mechanism specific against Ganoderma boninense.

Keywords: Ganoderma boninense, necrotrophic, next-generation sequencing, transcription factors

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846 Integrated Management of Diseases of Vegetables and Flower Crops Grown in Protected Condition under Organic Production System

Authors: Shripad Kulkarni

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Plant disease is an impairment of the normal state of a plant that interrupts or modifies its vital functions. Disease occurs on different parts of plants and cause heavy losses. Diagnosis of Problem is very important before planning any management practice and this can be done based on appearance of the crop, examination of the root and examination of the soil. There are various types of diseases such as biotic (transmissible) which accounts for ~30% whereas , abiotic (not transmissible) diseases are the major one with ~70% incidence. Plant diseases caused by different groups of organism’s belonging fungi, bacteria, viruses, nematodes and few others have remained important in causing significant losses in different crops indicating the urgent need of their integrated management. Various factors favor disease development and different steps and methods are involved in management of diseases under protected condition. Management of diseases through botanicals and bioagents by modifying root and aerial environment, vector management along with care to be taken while managing the disease are analysed.

Keywords: organic production system, diseases, bioagents and polyhouse, agriculture

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845 Improvement to Abiotic Stress Tolerance in Durum Wheat (Triticum Durum Desf) with the Vegetable Extract Application

Authors: Zemour Kamel, Chouhim Kadda Mohamed Amine

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Salinity is one of the most environmental factors limiting crop productivity. It has a negative effect on both germination and plant growth processes (photosynthesis, respiration, and transpiration), nutrient balance, membrane properties and cellular homeostasis, enzymatic and metabolic activities. Among the strategic crops in the world and more mainly in Algeria, durum wheat is very affected by this abiotic stress. For that, this study focuses on an evaluation of salt stress effect on the germination process of durum wheat as well as its response after application of lavender hydrosol and aqueous pistachio extract. The results have shown that all the physicochemical parameters of germination have been affected by this stress. However, lavender hydrosol and aqueous pistachio extract, considered as organic compounds, significantly improved the germination of wheat seeds. Finally, this study has highlighted the importance of using organic products as an ideal alternative to reduce the effect of abiotic stress on durum wheat productivity.

Keywords: salinity, wheat durum, extract, lavender hydrosol, aqueous pistachio

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844 Effects of Abiotic Stress on the Phytochemical Content and Bioactivity of Pistacia lentiscus L.

Authors: S. Mamoucha, N. Tsafantakis, Α. Ioannidis, S. Chatzipanagiotou, C. Nikolaou, L. Skaltsounis, N. Fokialakis, N. Christodoulakis

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Introduction: Plant secondary metabolites (SM) can be grouped into three chemically distinct groups: terpenes, phenolics, and nitrogen-containing compounds. For many years the adaptive significance of SM was unknown. They were thought to be functionless end-products. Currently it is accepted that many secondary metabolites (also known as natural products) have important ecological roles in plants. For instance, they serve as attractants (odor, color, taste) for pollinators and seed-dispersing animals. Moreover, they protect plants from herbivores, microbial pathogens and from environmental stress (high and low temperatures, drought, alkalinity, salinity, radiation etc). It is well known that both biotic and abiotic stress often increase the accumulation of SM. The local climatic conditions, seasonal changes, external factors such as light, temperature, humidity affect the biosynthesis and composition of secondary metabolites. A well known dioecious evergreen plant, Pistacia lentiscus L. (mastic tree), was selected in order to study the metabolic variations occur in response to the different climate conditions, due to the seasonal variation and its effect on the biosynthesis of bioactive compounds. Materials-methods: Young and mature leaves were collected in January and July 2014, dried and extracted by accelerated solvent extraction (Dionex ASE™ 350) using solvents of increased polarity (DCM, MeOH, and H2O). GC-MS and UHPLC-HRMS analysis were carried out in order to define the nature and the relative abundance of SM. The antibacterial activity was evaluated by using the Agar Disc Diffusion Assay against ATCC and clinical isolates strains: Escherichia coli, Staphylococcus aureus, Pseudomonas aeruginosa, Candida albicans, Streptococcus mutans and Klebsiella pneumoniae. All tests were carried out in duplicate and the average radii of the inhibition zones were calculated for each extract. Results: According to the phytochemical profile obtained from each extract, the biosynthesis of SM varied both qualitatively and quantitatively under the two different types of seasonal stress. With exception of the biologically inactive nonpolar DCM extract of July, all extracts inhibited the growth of most of the investigated microorganisms. A clear positive correlation has been observed between the relative abundance of SM and the bioactivity of the DCM extracts of January and July. Observed changes during phytochemical analysis were mainly focused on the triterpenoid content. On the other hand, the bioactivity of the polar extracts (MeOH and H2O) of January and July resulted practically invariable against most of the microorganisms, besides the significant variation of the SM content due to the seasonal variation. Conclusion: Our results clearly confirmed the hypothesis of abiotic stress as an important regulating factor that significantly affects the biosynthesis of secondary metabolites and thus the presence of bioactive compounds. Acknowledgment: This work was supported by IKY - State Scholarship Foundation, Athens, Greece.

Keywords: antibacterial screening, phytochemical profile, Pistacia lentiscus, abiotic stress

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843 Expression of Micro-RNA268 in Zinc Deficient Rice

Authors: Sobia Shafqat, Saeed Ahmad Qaisrani

Abstract:

MicroRNAs play an essential role in the regulation and development of all processes in most eukaryotes because of their prospective part as mediators controlling cell growth and differentiation towards the exact position of RNAs response in plants under biotic and abiotic factors or stressors. In a few cases, Zn is oblivious poisonous for plants due to its heavy metal status. Some other metals are extremely toxic, like Cd, Hg, and Pb, but these elements require in rice for the programming of genes under abiotic stress resembling Zn stress when micro RNAs268 was importantly introduced in rice. The micro RNAs overexpressed in transgenic plants with an accumulation of a large amount of melanin dialdehyde, hydrogen peroxide, and an excessive quantity of Zn in the seedlings stage. Let out results for rice pliability under Zn stress micro RNAs act as negative controllers. But the role of micro RNA268 act as a modulator in different ecological condition. It has been explained clearly with a long understanding of the role of micro RNA268 under stress conditions; pliability and practically showed outcome to increase plant sufferance under Zn stress because micro RNAs is an intervention technique for gene regulation in gene expression. The proposed study was experimented with by using genetic factors of Zn stress and toxicity effect on rice plants done at District Vehari, Pakistan. The trial was performed randomly with three replications in a complete block design (RCBD). These blocks were controlled with different concentrations of genetic factors. By overexpression of micro RNA268 rice, seedling growth was not stopped under Zn deficiency due to the accumulation of a large amount of melanin dialdehyde, hydrogen peroxide, and an excessive quantity of Zn in their seedlings. Results showed that micro RNA268 act as a negative controller under Zn stress. In the end, under stress conditions, micro RNA268 showed the necessary function in the tolerance of rice plants. The directorial work sketch gave out high agronomic applications and yield outcomes in rice with a specific amount of Zn application.

Keywords: micro RNA268, zinc, rice, agronomic approach

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842 Study on Residual Stress Measurement of Inconel-718 under Different Lubricating Conditions

Authors: M. Sandeep Kumar, Vasu Velagapudi, A. Venugopal

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When machining is carried out on a workpiece, residual stresses are induced in the workpiece due to nonuniform thermal and mechanical loads. These stresses play a vital role in the surface integrity of the final product or the output. Inconel 718 is commonly used in critical structural components of aircraft engines due to its properties at high temperatures. Therefore it is important to keep down the stresses induced due to machining. This can be achieved through proper lubricating conditions. In this work, experiments were carried out to check the influence of the developed nanofluid as cutting fluids on residual stresses developed during the course of machining. The results of MQL/Nanofluids were compared with MQL/Vegetable oil and dry machining lubricating condition. Results indicate the reduction in residual stress with the use of MQL/Nanofluid.

Keywords: nanofluids, MQL, residual stress, Inconel-718

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841 Geographical Parthenogenesis in Plants

Authors: Elvira Hörandl

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The term “Geographical parthenogenesis” describes the phenomenon that asexual organisms usually occupy larger and more northern distribution areas than their sexual relatives and tend to colonize previously glaciated areas. Several case studies in flowering plants confirm the geographical pattern, but the causal factors behind the phenomenon are still unclear. Previous authors regarded predominant polyploidy in asexual (apomictic) plants as the main factor. However, the geographical pattern is not the rule for sexual polyploids. Recent research confirmed a previous hypothesis of the author that a combination of factors is acting: Although uniparental reproduction provides better colonization abilities, it is most efficient in combination with polyploidy. I will present results on case studies in the genus Ranunculus of both autopolyploid and allopolyploid species and species complexes reproducing via facultative apomixis. Polyploidy seems to contribute mainly to a better tolerance of colder climates and temperate extremes, whereby epigenetic flexibility, changes in gene expression, and phenotypic plasticity play an important role in occupying ecological niches under harsh conditions. Phylogenomic studies entangle complex hybrid origins of asexual taxa, which increases intragenomic heterozygosity of asexual plants. Interestingly, our results suggest an association of sexuality with abiotic stresses, specifically with light stress, which might explain that still, most plants in high altitudes and in southern areas retain sexual reproduction despite other climatic conditions that would favor apomictic plants. We conclude that geographical parthenogenesis results from the complex interplay of the genomic constitution, mode of reproduction and environmental factors.

Keywords: apomixis, polyploidy, hybridization, abiotic stress, epigenetics, phylogenomics

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840 Evaluation of Residual Stresses in Human Face as a Function of Growth

Authors: M. A. Askari, M. A. Nazari, P. Perrier, Y. Payan

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Growth and remodeling of biological structures have gained lots of attention over the past decades. Determining the response of living tissues to mechanical loads is necessary for a wide range of developing fields such as prosthetics design or computerassisted surgical interventions. It is a well-known fact that biological structures are never stress-free, even when externally unloaded. The exact origin of these residual stresses is not clear, but theoretically, growth is one of the main sources. Extracting body organ’s shapes from medical imaging does not produce any information regarding the existing residual stresses in that organ. The simplest cause of such stresses is gravity since an organ grows under its influence from birth. Ignoring such residual stresses might cause erroneous results in numerical simulations. Accounting for residual stresses due to tissue growth can improve the accuracy of mechanical analysis results. This paper presents an original computational framework based on gradual growth to determine the residual stresses due to growth. To illustrate the method, we apply it to a finite element model of a healthy human face reconstructed from medical images. The distribution of residual stress in facial tissues is computed, which can overcome the effect of gravity and maintain tissues firmness. Our assumption is that tissue wrinkles caused by aging could be a consequence of decreasing residual stress and thus not counteracting gravity. Taking into account these stresses seems therefore extremely important in maxillofacial surgery. It would indeed help surgeons to estimate tissues changes after surgery.

Keywords: finite element method, growth, residual stress, soft tissue

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839 Gammarus: Asellus Ratio as an Index of Organic Pollution: A Case Study in Markeaton, Kedleston Hall, and Allestree Park Lakes Derby, UK

Authors: Usman Bawa

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Macro-invertebrates have been used to monitor organic pollution in rivers and streams. Several biotic indices based on macro-invertebrates have been developed over the years including the Biological Monitoring Working Party (BMWP). A new biotic index, the Gammarus:Asellus ratio has been recently proposed as an index of organic pollution. This study tested the validity of the Gammarus:Asellus ratio as an index of organic pollution, by examining the relationship between the Gammarus:Asellus ratio and physical-chemical parameters, and other biotic indices such as BMWP and, Average Score Per Taxon (ASPT) from lakes and streams at Markeaton Park, Allestree Park, and Kedleston Hall, Derbyshire. Macro invertebrates were sampled using the standard five-minute kick sampling techniques physical and chemical environmental variables were obtained based on standard sampling techniques. Eighteen sites were sampled, six sites from Markeaton Park (three sites across the stream and three sites across the lake). Six sites each were also sampled from Allestree Park and Kedleston Hall lakes. The Gammarus:Asellus ratio showed an opposite significant positive correlations with parameters indicative of organic pollution such as the level of nitrates, phosphates, and calcium and also revealed a negatively significant correlations with other biotic indices (BMWP/ASPT). The BMWP score correlated positively significantly with some water quality parameters such as dissolved oxygen and flow rate, but revealed no correlations with other chemical environmental variables. The BMWP score was significantly higher in the stream than the lake in Markeaton Park, also The ASPT scores appear to be significantly higher in the upper Lakes than the middle and lower lakes. This study has further strengthened the use of BMWP/ASPT score as an index of organic pollution. But, additional application is required to validate the use of Gammarus:Asellus as a rapid bio monitoring tool.

Keywords: Asellus, biotic index, Gammarus, macro invertebrates, organic pollution

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838 An Analytical Approach to Calculate Thermo-Mechanical Stresses in Integral Abutment Bridge Piles

Authors: Jafar Razmi

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Integral abutment bridges are bridges that do not have joints. If these bridges are subject to large seasonal and daily temperature variations, the expansion and contraction of the bridge slab is transferred to the piles. Since the piles are deep into the soil, displacement induced by slab can cause bending and stresses in piles. These stresses cause fatigue and failure of piles. A complex mechanical interaction exists between the slab, pile, soil and abutment. This complex interaction needs to be understood in order to calculate the stresses in piles. This paper uses a mechanical approach in developing analytical equations for the complex structure to determine the stresses in piles. The solution to these analytical solutions is developed and compared with finite element analysis results and experimental data. Our comparison shows that using analytical approach can accurately predict the displacement in piles. This approach offers a simplified technique that can be utilized without the need for computationally extensive finite element model.

Keywords: integral abutment bridges, piles, thermo-mechanical stress, stress and strains

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837 Solution of S3 Problem of Deformation Mechanics for a Definite Condition and Resulting Modifications of Important Failure Theories

Authors: Ranajay Bhowmick

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Analysis of stresses for an infinitesimal tetrahedron leads to a situation where we obtain a cubic equation consisting of three stress invariants. This cubic equation, when solved for a definite condition, gives the principal stresses directly without requiring any cumbersome and time-consuming trial and error methods or iterative numerical procedures. Since the failure criterion of different materials are generally expressed as functions of principal stresses, an attempt has been made in this study to incorporate the solutions of the cubic equation in the form of principal stresses, obtained for a definite condition, into some of the established failure theories to determine their modified descriptions. It has been observed that the failure theories can be represented using the quadratic stress invariant and the orientation of the principal plane.

Keywords: cubic equation, stress invariant, trigonometric, explicit solution, principal stress, failure criterion

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836 Analysis and Modeling of Stresses and Creeps Resulting from Soil Mechanics in Southern Plains of Kerman Province

Authors: Kourosh Nazarian

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Many of the engineering materials, such as behavioral metals, have at least a certain level of linear behavior. It means that if the stresses are doubled, the deformations would be also doubled. In fact, these materials have linear elastic properties. Soils do not follow this law, for example, when compressed, soils become gradually tighter. On the surface of the ground, the sand can be easily deformed with a finger, but in high compressive stresses, they gain considerable hardness and strength. This is mainly due to the increase in the forces among the separate particles. Creeps also deform the soils under a constant load over time. Clay and peat soils have creep behavior. As a result of this phenomenon, structures constructed on such soils will continue their collapse over time. In this paper, the researchers analyzed and modeled the stresses and creeps in the southern plains of Kerman province in Iran through library-documentary, quantitative and software techniques, and field survey. The results of the modeling showed that these plains experienced severe stresses and had a collapse of about 26 cm in the last 15 years and also creep evidence was discovered in an area with a gradient of 3-6 degrees.

Keywords: Stress, creep, faryab, surface runoff

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835 Induced Systemic Resistance in Tomato Plants against Fusarium Wilt Disease Using Biotic Inducers

Authors: Mostafa A. Amer, I. A. El-Samra, I. I. Abou-ElSeoud, S. M. El-Abd, N. K. Shawertamimi

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Tomato Fusarium wilt disease caused by Fusarium oxysporum f. sp. Lycopercisi (FOL) is considered one of the most destructive diseases in Egypt. Effect of some biotic inducers such as Bacillus megaterium var. phosphaticum, Glomus intraradices and Glomus macrocarpum at seven different mixed treatments, was tested for their ability to induce resistance in tomato plants against the disease. According to pathogenicity tests, all the tested isolates of FOL showed wilt symptoms on both of the tested cultivars; however, they considerably varied in percentages of disease incidence (DI) and disease severity (DS). Castle Rock was more susceptible than Peto 86, which was relatively resistant. Pretreatment of both cultivars, under greenhouse conditions, with the tested biotic inducers alone or in combination with each other's, significantly increased the induction of chitinase, β-1,3-glucanase, peroxidase, and polyphenoloxidase and reduced disease incidence and severity, compared with untreated noninoculated (C1) and untreated inoculated (C2) controls. Application of a combination of BMP, with GI and GM was the most effective in increasing the induction rated of the tested enzymes, compared with the other treatments. Induction of enzymes in most of the tested bioinducers treatments gradually increased, attaining maximum values after 48 or/and 72 hrs after challenging with FOL, then gradually declined. GI was the least effective bioinducer.

Keywords: F. oxysporum f. sp. lycopersici, defense enzymes, induced systemic resistance, ISR, B. megaterium var. phosphaticum, G. macrocarpum, G. intraradices

Procedia PDF Downloads 370
834 Ecobiological Study of Olivier in the Northern Slopes of the Mountains of Tlemcen, Western Algeria

Authors: Hachemi Nouria

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The olive tree is a Mediterranean tree, which belongs to the family Oleaceae. The Olea genus contains various species and subspecies, and the only species bearing edible fruit is Olea europaea. The desired issue in this study is to provide the current status of plant cover and especially the training in Olea europaea currently existing in the major centers of the region of Tlemcen. While based on the flora and biometric aspect of this plant germplasm. In order to make an assessment of the phytomass, we made measurements of the four parameters of the aerial part of the taxon: height, diameter, and canopy density to ten feet of the olive tree per station. The floristic analysis shows a certain floristic difference between the different stations. The vegetal formations reflect the biotic and abiotic conditions including climate affecting the ecosystem. Biometric study on the feet of Olea in the six study sites, has led us to conclude that the four measured parameters provides insight on the development or degradation of Olea feet depending on the layout of the stations and the factors environmental. We find that the terrains are havens for these assets. Also the local microclimate (Oued Thalweg) promotes the healthy development of this species.

Keywords: olivier, ecology, biometrics, Tlemcen, Algeria

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833 Characterization of Soil Microbial Communities from Vineyard under a Spectrum of Drought Pressures in Sensitive Area of Mediterranean Region

Authors: Gianmaria Califano, Júlio Augusto Lucena Maciel, Olfa Zarrouk, Miguel Damasio, Jose Silvestre, Ana Margarida Fortes

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Global warming, with rapid and sudden changes in meteorological conditions, is one of the major constraints to ensuring agricultural and crop resilience in the Mediterranean regions. Several strategies are being adopted to reduce the pressure of drought stress on grapevines at regional and local scales: improvements in the irrigation systems, adoption of interline cover crops, and adaptation of pruning techniques. However, still, more can be achieved if also microbial compartments associated with plants are considered in crop management. It is known that the microbial community change according to several factors such as latitude, plant variety, age, rootstock, soil composition and agricultural management system. Considering the increasing pressure of the biotic and abiotic stresses, it is of utmost necessity to also evaluate the effects of drought on the microbiome associated with the grapevine, which is a commercially important crop worldwide. In this study, we characterize the diversity and the structure of the microbial community under three long-term irrigation levels (100% ETc, 50% ETc and rain-fed) in a drought-tolerant grapevine cultivar present worldwide, Syrah. To avoid the limitations of culture-dependent methods, amplicon sequencing with target primers for bacteria and fungi was applied to the same soil samples. The use of the DNeasy PowerSoil (Qiagen) extraction kit required further optimization with the use of lytic enzymes and heating steps to improve DNA yield and quality systematically across biological treatments. Target regions (16S rRNA and ITS genes) of our samples are being sequenced with Illumina technology. With bioinformatic pipelines, it will be possible to obtain a characterization of the bacterial and fungal diversity, structure and composition. Further, the microbial communities will be assessed for their functional activity, which remains an important metric considering the strong inter-kingdom interactions existing between plants and their associated microbiome. The results of this study will lay the basis for biotechnological applications: in combination with the establishment of a bacterial library, it will be possible to explore the possibility of testing synthetic microbial communities to support plant resistance to water scarcity.

Keywords: microbiome, metabarcoding, soil, vinegrape, syrah, global warming, crop sustainability

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832 Enzymatic Determination of Limonene in Red Clover Genotypes

Authors: Andrés Quiroz, Emilio Hormazabal, Ana Mutis, Fernando Ortega, Manuel Chacón-Fuentes, Leonardo Parra

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Red clover (Trifolium pratense L.) is an important forage species in temperate regions of the world. The main limitation of this species worldwide is a lack of persistence related to the high mortality of plants due to a complex of biotic and abiotic factors, determining a life span of two or three seasons. Because of the importance of red clover in Chile, a red clover breeding program was started at INIA Carillanca Research Center in 1989, with the main objective of improving the survival of plants, forage yield, and persistence. The main selection criteria for selecting new varieties have been based on agronomical parameters and biotic factors. The main biotic factor associated with red clover mortality in Chile is Hylastinus obscurus (Coleoptera: Curculionidae). Both larval and adults feed on the roots, causing weakening and subsequent death of clover plants. Pesticides have not been successful for controlling infestations of this root borer. Therefore, alternative strategies for controlling this pest are a high priority for red clover producers. Currently, the role of semiochemical in the interaction between H. obscurus and red clover plants has been widely studied for our group. Specifically, from the red clover foliage has been identified limonene is eliciting repellency from the root borer. Limonene is generated in the plant from two independent biosynthetic pathways, the mevalonic acid, and deoxyxylulose pathway. Mevalonate pathway enzymes are localized in the cytosol, whereas the deoxyxylulose phosphate pathway enzymes are found in plastids. In summary, limonene can be determinated by enzymatic bioassay using GPP as substrate and by limonene synthase expression. Therefore, the main objective of this work was to study genetic variation of limonene in material provided by INIA´s Red Clover breeding program. Protein extraction was carried out homogenizing 250 mg of leave tissue and suspended in 6 mL of extraction buffer (PEG 1500, PVP-30, 20 mM MgCl2 and antioxidants) and stirred on ice for 20 min. After centrifugation, aliquots of 2.5 mL were desalted on PD-10 columns, resulting in a final volume of 3.5 mL. Protein determination was performed according to Bradford with BSA as a standard. Monoterpene synthase assays were performed with 50 µL of protein extracts transferred into gas-tight 2 mL crimp seal vials after addition of 4 µL MgCl₂ and 41 µL assay buffer. The assay was started by adding 5 µL of a GPP solution. The mixture was incubated for 30 min at 40 °C. Biosynthesized limonene was quantified in a GC equipped with a chiral column and using synthetic R and S-limonene standards. The enzymatic the production of R and S-limonene from different Superqueli-Carillanca genotypes is shown in this work. Preliminary results showed significant differences in limonene content among the genotypes analyzed. These results constitute an important base for selecting genotypes with a high content of this repellent monoterpene towards H. obscurus.

Keywords: head space, limonene enzymatic determination, red clover, Hylastinus obscurus

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