Search results for: leaf extracts

Authors: Umara Nissar Rafiqi, Irum Gul, Nazima Nasrullah, Monica Saifi, Malik Z. Abdin

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Background: Stevia rebaudiana, a member of Asteraceae family is an important medicinal plant and produces a commercially used non-caloric natural sweetener, which is also an alternate herbal cure for diabetes. Steviol glycosides are the main sweetening compounds present in these plants. Secondary metabolites are crucial to the adaption of plants to the environment and its overcoming stress conditions. In agricultural procedures, the abiotic stresses like salinity, high metal toxicity and drought, in particular, are responsible for the majority of the reduction that differentiates yield potential from harvestable yield. Salt stress and heavy metal toxicity lead to increased production of reactive oxygen species (ROS). To avoid oxidative damage due to ROS and osmotic stress, plants have a system of anti-oxidant enzymes along with several stress induced enzymes. This helps in scavenging the ROS and relieve the osmotic stress in different cell compartments. However, whether stress induced toxicity modulates the activity of these enzymes in Stevia rebaudiana is poorly understood. Aim: The present study focussed on the effect of salinity, heavy metal toxicity (lead and mercury) on physiological traits and transcriptional profiling of Stevia rebaudiana. Method: Stevia rebaudiana plants were collected from the Central Institute of Medicinal and Aromatic plants (CIMAP), Patnagar, India and maintained under controlled conditions in a greenhouse at Hamdard University, Delhi, India. The plants were subjected to different concentrations of salt (0, 25, 50 and 75 mM respectively) and heavy metals, lead and mercury (0, 100, 200 and 300 µM respectively). The physiological traits such as shoot length, root numbers, leaf growth were evaluated. The samples were collected at different developmental stages and analysed for transcription profiling by RT-PCR. Transcriptional studies in stevia rebaudiana involves important antioxidant enzymes: catalase (CAT), superoxide dismutase (SOD), cytochrome P450 monooxygenase (CYP) and stress induced aquaporin (AQU), auxin repressed protein (ARP-1), Ndhc gene. The data was analysed using GraphPad Prism and expressed as mean ± SD. Result: Low salinity and lower metal toxicity did not affect the fresh weight of the plant. However, this was substantially decreased by 55% at high salinity and heavy metal treatment. With increasing salinity and heavy metal toxicity, the values of all studied physiological traits were significantly decreased. Chlorosis in treated plants was also observed which could be due to changes in Fe:Zn ratio. At low concentrations (upto 25 mM) of NaCl and heavy metals, we did not observe any significant difference in the gene expressions of treated plants compared to control plants. Interestingly, at high salt concentration and high metal toxicity, a significant increase in the expression profile of stress induced genes was observed in treated plants compared to control (p < 0.005). Conclusion: Stevia rebaudiana is tolerant to lower salt and heavy metal concentration. This study also suggests that with the increase in concentrations of salt and heavy metals, harvest yield of S. rebaudiana was hampered.

Keywords: Stevia rebaudiana, natural sweetener, salinity, heavy metal toxicity

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Authors: Hülya Torun, Ondřej Novák, Jaromír Mikulík, Miroslav Strnad, Faik A. Ayaz

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World climate is changing. Millions of people in the world still face chronic undernourishment for conducting a healthy life and the world’s population is growing steadily. To meet this growing demand, agriculture and food systems must adapt to the adverse effects of climate change and become more resilient, productive and sustainable. From this perspective, to determine tolerant cultivars for undesirable environmental conditions will be necessary food production for sustainable development. Among abiotic stresses, soil salinity is one of the most detrimental global fact restricting plant sources. Development of salt-tolerant lines is required in order to increase the crop productivity and quality in salt-treated lands. Therefore, the objective of this study was to investigate the morphological and physiological responses of barley cultivars accessions to salinity stress by NaCl. For this purpose, it was aimed to determine the crosstalk between some endogenous phytohormones and exogenous salicylic acid (SA) in two different vegetative parts (leaves and roots) of barley (Hordeum vulgare L.; Poaceae; 2n=14; Ince-04) which is detected salt-tolerant. The effects of SA on growth parameters, leaf relative water content (RWC), endogenous phytohormones; including indole-3-acetic acid (IAA), cytokinins (CKs), abscisic acid (ABA), jasmonic acid (JA) and ethylene were investigated in barley cultivars under salinity stress. SA was applied to 17-day-old seedlings of barley in two different ways including before (pre-treated for 24 h) and simultaneously with NaCl stress treatment. NaCl (0, 150, 300 mM) exposure in the hydrophonic system was associated with a rapid decrease in growth parameters and RWC, which is an indicator of plant water status, resulted in a strong up-regulation of ABA as a stress indicator. Roots were more dramatically affected than leaves. Water conservation in 150 mM NaCl treated-barley plants did not change, but decreased in 300 mM NaCl treated plants. Pre- and simultaneously treatment of SA did not significantly alter growth parameters and RWC. ABA, JA and ethylene are known to be related with stress. In the present work, ethylene also increased, similarly to ABA, but not with the same intensity. While ABA and ethylene increased by the increment of salt concentrations, JA levels rapidly decreased especially in roots. Both pre- and simultaneously SA applications alleviated salt-induced decreases in 300 mM NaCl resulted in the increment of ABA levels. CKs and IAA are related to cell growth and development. At high salinity (300 mM NaCl), CKs (cZ+cZR) contents increased in both vegetative organs while IAA levels stayed at the same level with control groups. However, IAA increased and cZ+cZR rapidly decreased in leaves of barley plants with SA treatments before salt applications (in pre- SA treated groups). Simultaneously application of SA decreased CKs levels in both leaves and roots of the cultivar. Due to increasing concentrations of NaCl in association with decreasing ABA, JA and ethylene content and increments in CKs and IAA were recorded with SA treatments. As results of the study, in view of all the phytohormones that we tested, exogenous SA induced greater tolerance to salinity particularly when applied before salinity stress.

Keywords: Barley, Hordeum vulgare, phytohormones, salicylic acid, salinity

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Authors: Adrian Szczyrba, Aneta Halka-Grysinska, Tomasz Baj, Tadeusz H. Dzido

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Since prehistory, plants were constituted as an essential source of biologically active substances in folk medicine. One of the examples of medicinal plants is Ruta graveolens L. For a long time, Ruta g. herb has been famous for its spasmolytic, diuretic, or anti-inflammatory therapeutic effects. The wide spectrum of secondary metabolites produced by Ruta g. includes flavonoids (eg. rutin, quercetin), coumarins (eg. bergapten, umbelliferone) phenolic acids (eg. rosmarinic acid, chlorogenic acid), and limonoids. Unfortunately, the presence of produced substances is highly dependent on environmental factors like temperature, humidity, or soil acidity; therefore standardization is necessary. There were many attempts of characterization of various phytochemical groups (eg. coumarins) of Ruta graveolens using the normal – phase thin-layer chromatography (TLC). However, due to the so-called general elution problem, usually, some components remained unseparated near the start or finish line. Therefore Ruta graveolens is a very good model plant. Methanol and petroleum ether extract from its aerial parts were used to demonstrate the capabilities of the new device for gradient thin-layer chromatogram development. The development of gradient thin-layer chromatograms in the reversed-phase system in conventional horizontal chambers can be disrupted by problems associated with an excessive flux of the mobile phase to the surface of the adsorbent layer. This phenomenon is most likely caused by significant differences between the surface tension of the subsequent fractions of the mobile phase. An excessive flux of the mobile phase onto the surface of the adsorbent layer distorts the flow of the mobile phase. The described effect produces unreliable, and unrepeatable results, causing blurring and deformation of the substance zones. In the prototype device, the mobile phase solution is delivered onto the surface of the adsorbent layer with controlled velocity (by moving pipette driven by 3D machine). The delivery of the solvent to the adsorbent layer is equal to or lower than that of conventional development. Therefore chromatograms can be developed with optimal linear mobile phase velocity. Furthermore, under such conditions, there is no excess of eluent solution on the surface of the adsorbent layer so the higher performance of the chromatographic system can be obtained. Directly feeding the adsorbent layer with eluent also enables to perform convenient continuous gradient elution practically without the so-called gradient delay. In the study, unique fingerprints of methanol and petroleum ether extracts of Ruta graveolens aerial parts were obtained with stepwise gradient reversed-phase thin-layer chromatography. Obtained fingerprints under different chromatographic conditions will be compared. The advantages and disadvantages of the proposed approach to chromatogram development with controlled solvent velocity will be discussed.

Keywords: fingerprints, gradient thin-layer chromatography, reversed-phase TLC, Ruta graveolens

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Authors: Inês Amorim Leitão, Loes van Shaick, António Dinis Ferreira, Violette Geissen

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Plastic pollution is a growing concern worldwide: plastics are commercialized in large quantities and it takes a long time for them to degrade. When in the environment, plastic is fragmented into microplastics (<5mm), which have been found in all environmental compartments at different locations. Microplastics contribute to the environmental pollution in water, air and soil and are linked to human health problems. The progressive increase of population living in cities led to the aggravation of the pollution problem worldwide, especially in urban environments. Urban areas represent a strong source of pollution, through the roads, industrial production, wastewater, landfills, etc. It is expected that pollutants such as microplastics are transported diffusely from the sources through different pathways such as wind and rain. Therefore, it is very complex to quantify, control and treat these pollutants, designated current problematic issues by the European Commission. Green areas are pointed out by experts as natural filters for contaminants in cities, through their capacity of retention by vegetation. These spaces have thus the capacity to control the load of pollutants transported. This study investigates the spatial distribution of microplastics in urban soils of different land uses, their transport through atmospheric deposition, wind erosion, runoff and streams, as well as their deposition in vegetation like grass and tree leaves in urban environment. Coimbra, a medium large city located in the central Portugal, is the case-study. All the soil, sediments, water and vegetation samples were collected in Coimbra and were later analyzed in the Wageningen University & Research laboratory. Microplastics were extracted through the density separation using Sodium Phosphate as solution (~1.4 g cm−3) and filtration methods, visualized under a stereo microscope and identified using the u-FTIR method. Microplastic particles were found in all the different samples. In terms of soils, higher concentrations of microplastics were found in green parks, followed by landfills and industrial places, and the lowest concentrations in forests and pasture land-uses. Atmospheric deposition and streams after rainfall events seems to represent the strongest pathways of microplastics. Tree leaves can retain microplastics on their surfaces. Small leaves such as needle leaves seem to present higher amounts of microplastics per leaf area than bigger leaves. Rainfall episodes seem to reduce the concentration of microplastics on leaves surface, which suggests the wash of microplastics down to lower levels of the tree or to the soil. When in soil, different types of microplastics could be transported to the atmosphere through wind erosion. Grass seems to present high concentrations of microplastics, and the enlargement of the grass cover leads to a reduction of the amount of microplastics in soil, but also of the microplastics moved from the ground to the atmosphere by wind erosion. This study proof that vegetation can help to control the transport and dispersion of microplastics. In order to control the entry and the concentration of microplastics in the environment, especially in cities, it is essential to defining and evaluating nature-based land-use scenarios, considering the role of green urban areas in filtering small particles.

Keywords: microplastics, cities, sources, pathways, vegetation

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Authors: Prabhjot Kaur, B.K. Kang, Balwinder Singh

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The tobacco caterpillar, Spodoptera litura (Fabricius) (Lepidoptera: Noctuidae) is an agricultural important pest species. S. litura has a wide host range of approximately recorded 150 plant species worldwide. In Punjab, this pest attains sporadic status primarily on cauliflower, Brassica oleracea (L.). This pest destroys vegetable crop and particularly prefers the cruciferae family. However, it is also observed feeding on other crops such as arbi, Colocasia esculenta (L.), mung bean, Vigna radiata (L.), sunflower, Helianthus annuus (L.), cotton, Gossypium hirsutum (L.), castor, Ricinus communis (L.), etc. Larvae of this pest completely devour the leaves of infested plant resulting in huge crop losses which ranges from 50 to 70 per cent. Indiscriminate and continuous use of insecticides has contributed in development of insecticide resistance in insects and caused the environmental degradation as well. Moreover, a base line data regarding the toxicity of the newer insecticides would help in understanding the level of resistance developed in this pest and any possible cross-resistance there in, which could be assessed in advance. Therefore, present studies on development of resistance in S. litura against four new chemistry insecticides (emamectin benzoate, chlorantraniliprole, indoxacarb and spinosad) were carried out in the Toxicology laboratory, Department of Entomology, Punjab Agricultural University, Ludhiana, Punjab, India during the year 2011-12. Various stages of S. litura (eggs, larvae) were collected from four different locations (Malerkotla, Hoshiarpur, Amritsar and Samrala) of Punjab. Resistance is developed in third instars of lepidopterous pests. Therefore, larval bioassays were conducted to estimate the response of field populations of thirty third-instar larvae of S. litura under laboratory conditions at 25±2°C and 65±5 per cent relative humidity. Leaf dip bioassay technique with diluted insecticide formulations recommended by Insecticide Resistance Action Committee (IRAC) was performed in the laboratory with seven to ten treatments depending on the insecticide class, respectively. LC50 values were estimated by probit analysis after correction to record control mortality data which was used to calculate the resistance ratios (RR). The LC50 values worked out for emamectin benzoate, chlorantraniliprole, indoxacarb, spinosad are 0.081, 0.088, 0.380, 4.00 parts per million (ppm) against pest populations collected from Malerkotla; 0.051, 0.060, 0.250, 3.00 (ppm) of Amritsar; 0.002, 0.001, 0.0076, 0.10 ppm for Samrala and 0.000014, 0.00001, 0.00056, 0.003 ppm against pest population of Hoshiarpur, respectively. The LC50 values for populations collected from these four locations were in the order Malerkotla>Amritsar>Samrala>Hoshiarpur for the insecticides (emamectin benzoate, chlorantraniliprole, indoxacarb and spinosad) tested. Based on LC50 values obtained, emamectin benzoate (0.000014 ppm) was found to be the most toxic among all the tested populations, followed by chlorantraniliprole (0.00001 ppm), indoxacarb (0.00056 ppm) and spinosad (0.003 ppm), respectively. The pairwise correlation coefficients of LC50 values indicated that there was lack of cross resistance for emamectin benzoate, chlorantraniliprole, spinosad, indoxacarb in populations of S. litura from Punjab. These insecticides may prove to be promising substitutes for the effective control of insecticide resistant populations of S. litura in Punjab state, India.

Keywords: Spodoptera litura, insecticides, toxicity, resistance

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Authors: Andrés Quiroz, Emilio Hormazábal, Ana Mutis, Fernando Ortega, Loreto Méndez, Leonardo Parra

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Red clover root borer, Hylastinus obscurus Marsham (Coleoptera: Curculionidae), is the main insect pest associated to red clover, Trifolium pratense L. An average of 1.5 H. obscurus per plant can cause 5.5% reduction in forage yield in pastures of two to three years old. Moreover, insect attack can reach 70% to 100% of the plants. To our knowledge, there is no a chemical strategy for controlling this pest. Therefore alternative strategies for controlling H. obscurus are a high priority for red clover producers. One of this alternative is related to the study of secondary metabolites involved in intrinsic chemical defenses developed by plants, such as isoflavonoids. The isoflavonoids formononetin and daidzein have elicited an antifeedant and phagostimult effect on H. obscurus respectively. However, we do not know how is the dynamic variation of these isoflavonoids under field conditions. The main objective of this work was to evaluate the variation of the antifeedant isoflavonoids formononetin, the phagostimulant isoflavonoids daidzein, and their respective glycosides over time in different ecotypes of red clover. Fourteen red clover ecotypes (8 cultivars and 6 experimental lines), were collected at INIA-Carillanca (La Araucanía, Chile). These plants were established in October 2015 under irrigated conditions. The cultivars were distributed in a randomized complete block with three replicates. The whole plants were sampled in four times: 15th October 2016, 12th December 2016, 27th January 2017 and 16th March 2017 with sufficient amount of soil to avoid root damage. A polar fraction of isoflavonoid was obtained from 20 mg of lyophilized root tissue extracted with 2 mL of 80% MeOH for 16 h using an orbital shaker in the dark at room temperature. After, an aliquot of 1.4 mL of the supernatant was evaporated, and the residue was resuspended in 300 µL of 45% MeOH. The identification and quantification of isoflavonoid root extracts were performed by the injection of 20 µL into a Shimadzu HPLC equipped with a C-18 column. The sample was eluted with a mobile phase composed of AcOH: H₂O (1:9 v/v) as solvent A and CH₃CN as solvent B. The detection was performed at 260 nm. The results showed that the amount of aglycones was higher than the respective glycosides. This result is according to the biosynthetic pathway of flavonoids, where the formation of glycoside is further to the glycosides biosynthesis. The amount of formononetin was higher than daidzein. In roots, where H. obscurus spent the most part of its live cycle, the highest content of formononetin was found in G 27, Pawera, Sabtoron High, Redqueli-INIA and Superqueli-INIA cvs. (2.1, 1.8, 1.8, 1.6 and 1.0 mg g⁻¹ respectively); and the lowest amount of daidzein were found Superqueli-INIA (0.32 mg g⁻¹) and in the experimental line Sel Syn Int4 (0.24 mg g⁻¹). This ecotype showed a high content of formononetin (0.9 mg g⁻¹). This information, associated with cultural practices, could help farmers and breeders to reduce H. obscurus in grassland, selecting ecotypes with high content of formononetin and low amount of daidzein in the roots of red clover plants. Acknowledgements: FONDECYT 1141245 and 11130715.

Keywords: daidzein, formononetin, isoflavonoid glycosides, trifolium pratense

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Authors: Karolina Wieczorek, Sophie Wiliams

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Introduction: Patient data is often collected in Electronic Health Record Systems (EHR) for purposes such as providing care as well as reporting data. This information can be re-used to validate data models in clinical trials or in epidemiological studies. Manual validation of automated tools is vital to pick up errors in processing and to provide confidence in the output. Mentioning a disease in a discharge letter does not necessarily mean that a patient suffers from this disease. Many of them discuss a diagnostic process, different tests, or discuss whether a patient has a certain disease. The COVID-19 dataset in this study used natural language processing (NLP), an automated algorithm which extracts information related to COVID-19 symptoms, complications, and medications prescribed within the hospital. Free-text patient clinical patient notes are rich sources of information which contain patient data not captured in a structured form, hence the use of named entity recognition (NER) to capture additional information. Methods: Patient data (discharge summary letters) were exported and screened by an algorithm to pick up relevant terms related to COVID-19. Manual validation of automated tools is vital to pick up errors in processing and to provide confidence in the output. A list of 124 Systematized Nomenclature of Medicine (SNOMED) Clinical Terms has been provided in Excel with corresponding IDs. Two independent medical student researchers were provided with a dictionary of SNOMED list of terms to refer to when screening the notes. They worked on two separate datasets called "A” and "B”, respectively. Notes were screened to check if the correct term had been picked-up by the algorithm to ensure that negated terms were not picked up. Results: Its implementation in the hospital began on March 31, 2020, and the first EHR-derived extract was generated for use in an audit study on June 04, 2020. The dataset has contributed to large, priority clinical trials (including International Severe Acute Respiratory and Emerging Infection Consortium (ISARIC) by bulk upload to REDcap research databases) and local research and audit studies. Successful sharing of EHR-extracted datasets requires communicating the provenance and quality, including completeness and accuracy of this data. The results of the validation of the algorithm were the following: precision (0.907), recall (0.416), and F-score test (0.570). Percentage enhancement with NLP extracted terms compared to regular data extraction alone was low (0.3%) for relatively well-documented data such as previous medical history but higher (16.6%, 29.53%, 30.3%, 45.1%) for complications, presenting illness, chronic procedures, acute procedures respectively. Conclusions: This automated NLP algorithm is shown to be useful in facilitating patient data analysis and has the potential to be used in more large-scale clinical trials to assess potential study exclusion criteria for participants in the development of vaccines.

Keywords: automated, algorithm, NLP, COVID-19

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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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Authors: Suneet Jagdev, Hrishabh Amrodia, Siddharth Menon, Abhishek Singh, Mansi Shivhare

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Delhi, the National Capital, has undergone a surge in development rate, consequently engendering an unprecedented increase in population. Over the years the city has transformed into a car-centric infrastructure with high-speed corridors, flyovers and fast lanes. A considerable section of the population is hankering to rehabilitate to the good old cycling days, in order to contribute towards a green environment as well as to maintain their physical well-being. Furthermore, an extant section of Delhi’s population relies on cycles as their primary means of commuting in the city. Delhi has the highest number of cyclists and second highest number of pedestrians in the country. However, the tumultuous problems of unregulated traffic, inadequate space on roads, adverse weather conditions stifle them to opt for cycling. Lately, the city has been facing a conglomeration of problems such as haphazard traffic movement, clogged roads, congestion, pollution, accidents, safety issues, etc. In 1957, Delhi’s cyclists accounted for 36 per cent of trips which dropped down to a mere 4 per cent in 2008. The declining rate is due to unsafe roads and lack of proper cycle lanes. Now as the 10 percent of the city has cycle tracks. There is also a lack of public recreational activities in the city. These conundrums incite the need of a covered elevated cycling bridge track to facilitate the safe and smooth cycle commutation in the city which would also serve the purpose of an alternate urban public space over the cycle bridge reducing the cost as well as the space requirement for the same, developing a user–friendly transportation and public interaction system for urban areas in the city. Based on the archival research methodologies, the following research draws information and extracts records from the data accounts of the Delhi Metro Rail Corporation Ltd. as well as the Centre for Science and Environment, India. This research will predominantly focus on developing a prototype design for high speed elevated bicycle lanes based on different road typologies, which can be replicated with minor variations in similar situations, all across the major cities of our country including the proposed smart cities. Furthermore, how these cycling lanes could be utilized for the place making process accommodating cycle parking and renting spaces, public recreational spaces, food courts as well as convenient shopping facilities with appropriate optimization. How to preserve and increase the share of smooth and safe cycling commute cycling for the routine transportation of the urban community of the polluted capital which has been on a steady decline over the past few decades.

Keywords: bicycle track, prototype, road safety, urban spaces

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Authors: Dipankar Saha, J. P. Singh, C. B. Pandey

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Indian Thar desert being the seventh largest in the world is the main hot sand desert occupies nearly 385,000km2 and about 9% of the area of the country harbours several species likely the flora of 682 species (63 introduced species) belonging to 352 genera and 87 families. The degree of endemism of plant species in the Thar desert is 6.4 percent, which is relatively higher than the degree of endemism in the Sahara desert which is very significant for the conservationist to envisage. The advent and development of computer technology for digitization and data base management coupled with the rapidly increasing importance of biodiversity conservation resulted in the invention of biodiversity informatics as discipline of basic sciences with multiple applications. Aichi Target 19 as an outcome of Convention of Biological Diversity (CBD) specifically mandates the development of an advanced and shared biodiversity knowledge base. Information on species distributions in space is the crux of effective management of biodiversity in the rapidly changing world. The efficiency of biodiversity management is being increased rapidly by various stakeholders like researchers, policymakers, and funding agencies with the knowledge and application of biodiversity informatics. Herbarium specimens being a vital repository for biodiversity conservation especially in climate change scenario the digitization process usually aims to improve access and to preserve delicate specimens and in doing so creating large sets of images as a part of the existing repository as arid plant information facility for long-term future usage. As the leaf characters are important for describing taxa and distinguishing between them and they can be measured from herbarium specimens as well. As a part of this activity, laminar characterization (leaves being the most important characters in assessing climate change impact) initially resulted in classification of more than thousands collections belonging to ten families like Acanthaceae, Aizoaceae, Amaranthaceae, Asclepiadaceae, Anacardeaceae, Apocynaceae, Asteraceae, Aristolochiaceae, Berseraceae and Bignoniaceae etc. Taxonomic diversity indices has also been worked out being one of the important domain of biodiversity informatics approaches. The digitization process also encompasses workflows which incorporate automated systems to enable us to expand and speed up the digitisation process. The digitisation workflows used to be on a modular system which has the potential to be scaled up. As they are being developed with a geo-referencing tool and additional quality control elements and finally placing specimen images and data into a fully searchable, web-accessible database. Our effort in this paper is to elucidate the role of BIs, present effort of database development of the existing botanical collection of institute repository. This effort is expected to be considered as a part of various global initiatives having an effective biodiversity information facility. This will enable access to plant biodiversity data that are fit-for-use by scientists and decision makers working on biodiversity conservation and sustainable development in the region and iso-climatic situation of the world.

Keywords: biodiversity informatics, climate change, digitization, herbarium, laminar characters, web accessible interface

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Authors: Jia Li, Xing Wei, Yuchen Hong, Yang Lu

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Detecting vehicle behavior has always been the focus of intelligent transportation, but with the explosive growth of the number of vehicles and the complexity of the road environment, the vehicle behavior videos captured by traditional surveillance have been unable to satisfy the study of vehicle behavior. The traditional method of manually labeling vehicle behavior is too time-consuming and labor-intensive, but the existing object detection and tracking algorithms have poor practicability and low behavioral location detection rate. This paper proposes a vehicle behavior detection algorithm based on the dual-stream convolution network and the multi-dimensional video dynamic detection network. In the videos, the straight-line behavior of the vehicle will default to the background behavior. The Changing lanes, turning and turning around are set as target behaviors. The purpose of this model is to automatically mark the target behavior of the vehicle from the untrimmed videos. First, the target behavior proposals in the long video are extracted through the dual-stream convolution network. The model uses a dual-stream convolutional network to generate a one-dimensional action score waveform, and then extract segments with scores above a given threshold M into preliminary vehicle behavior proposals. Second, the preliminary proposals are pruned and identified using the multi-dimensional video dynamic detection network. Referring to the hierarchical reinforcement learning, the multi-dimensional network includes a Timer module and a Spacer module, where the Timer module mines time information in the video stream and the Spacer module extracts spatial information in the video frame. The Timer and Spacer module are implemented by Long Short-Term Memory (LSTM) and start from an all-zero hidden state. The Timer module uses the Transformer mechanism to extract timing information from the video stream and extract features by linear mapping and other methods. Finally, the model fuses time information and spatial information and obtains the location and category of the behavior through the softmax layer. This paper uses recall and precision to measure the performance of the model. Extensive experiments show that based on the dataset of this paper, the proposed model has obvious advantages compared with the existing state-of-the-art behavior detection algorithms. When the Time Intersection over Union (TIoU) threshold is 0.5, the Average-Precision (MP) reaches 36.3% (the MP of baselines is 21.5%). In summary, this paper proposes a vehicle behavior detection model based on multi-dimensional dynamic detection network. This paper introduces spatial information and temporal information to extract vehicle behaviors in long videos. Experiments show that the proposed algorithm is advanced and accurate in-vehicle timing behavior detection. In the future, the focus will be on simultaneously detecting the timing behavior of multiple vehicles in complex traffic scenes (such as a busy street) while ensuring accuracy.

Keywords: vehicle behavior detection, convolutional neural network, long short-term memory, deep learning

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Authors: L. Abubakar, B. M. Sokoto, I. U. Mohammed, M. S. Na’allah, A. Mohammad, A. N. Garba, T. S. Bubuche

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Onion (Allium cepa var. cepa L.), is the most important species of the Allium group belonging to family Alliaceae and genus Allium. It can be regarded as the single important vegetable species in the world after tomatoes. Despite the similarities, which bring the species together, the genus is a strikingly diverse one, with more than five hundred species, which are perennial and mostly bulbous plants. Out of these, only seven species are in cultivation, and five are the most important species of the cultivated Allium. However, Allium cepa (onion) and Allium sativum (Garlic) are the two major cultivated species grown all over the world of which the onion crop is the most important. Heritability defined as the proportion of the observed total variability that is genetic, and its estimates from variance components give more useful information of genotypic variation from the total phenotypic differences and environmental effects on the individuals or families. It therefore guide the breeder with respect to the ease with which selection of traits can be carried out. Heritability estimates guide the breeder with respect to ease of selection of traits while correlations suggest how selection among characters can be practiced. Correlations explain relationship between characters and suggest how selection among characters can be practiced in breeding programmes. Highly significant correlations have been reported, between yield, maturity, rings/bulb and storage loss in onions. Similarly significant positive correlation exists between total bulb yield and plant height, leaf number/plant, bulb diameter and bulb yield/plant. Moderate positive correlations have been observed between maturity date and yield, dry matter content was highly correlated with soluble solids, and higher correlations were also observed between storage loss and soluble solids. The objective of the study is to determine heritability estimates and correlations for characters among onion cultivars of North Western Nigeria. This is envisaged will assist in the breeding of superior onion cultivars within the zone. Thirteen onion cultivars were collected during an expedition covering north western Nigeria and Southern part of Niger Republic during 2013, which are areas noted for onion production. The cultivars were evaluated at two locations; Sokoto, in Sokoto State and Jega in Kebbi State all in Nigeria during the 2013/14 onion season (dry season) under irrigation. Combined analysis of the results revealed fresh bulb yield is highly significantly positively correlated with bulb height and cured bulb yield, and significant positive correlation with plant height and bulb diameter. It also recorded significant negative correlation with mean No. of leaves/plant and non significant negative correlation with bolting %. Cured bulb yield (marketable yield) had highly significant positive correlation with mean bulb weight and fresh bulb yield/ha, with significant positive correlation with bulb height. It also recorded highly significant negative correlation with No. of leaves/plant and significant negative correlation with bolting % and non significant positive correlation with plant height and non significant negative correlation with bulb diameter. High broad sense heritability estimates were recorded for plant height, fresh bulb yield, number of leaves/plant, bolting % and cured bulb yield. Medium to low broad sense heritabilities were also observed for mean bulb weight, plant height and bulb diameter.

Keywords: correlation, heritability, onions, North Western Nigeria

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Authors: S. F. Afzali, W. L. Wong

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The epizootic ulcerative syndrome (EUS) causes by the oomycete fungus, Aphanomyces invadans; known to be one of the infectious fish diseases for farmed and wild fishes in fresh and brackish-water from the Asia-pacific region, America and Africa. Although, EUS had been documented by the Office International des Epizooties (OIE) since 1995, hitherto, there is neither standard chemical agents that can be used for successful treatment of this destructive infection in the time of outbreak; nor available vaccine for prevention. Plant-based remedies in controlling fish diseases are gaining much attention recently as an alternative to chemical treatments, which possess negative effects to the environment and human. In present study, Sonneratia alba, a mangrove plant belongs to the Sonneratiaceae family, was screened in vitro and in vivo for its antifungal activity against A. invadans mycelium growth and its effects on fish innate immune system and disease resistant. The in vitro tests was performed using the disc diffusion methods with measurements of minimum inhibitory concentration (MIC) and inhibition zone. For in vivo study, the S. alba extract supplemented diets were administrated at 0.0, 1.0%, 3.0%, and 5.0% on healthy goldfish, Carassius auratus, which challenged with A. invadans zoospores (100 spores/ml). To compare the significant differences in the hematological and immunological parameters obtained from the experiments, the data were analysed using the SPSS. The methanol extract of S. alba effectively inhibited the mycelial growth of A. invadans at a minimum concentration of 1000 ppm for agar and filter paper diffusion experiments. In the agar diffusion test, 500 ppm of the extract inhibited the fungus mycelial growth up to 96 hours after exposure. The mycelial growth from the edge of the pre-inoculated A. invadans agar discs treated with S. alba extracts at concentrations of 100, 500 and 1000 ppm were 15, 8 and 0 mm respectively. The results of the filter paper disc test showed that the S. alba extract at its minimal inhibitory concentration (1000 ppm) has similar qualitative inhibitory effect as malachite green at 1 ppm and formalin at 250 ppm. According to the in vivo tests findings, in the infected fish fed with 3.0% and 5.0% supplementation diet, the numbers of white blood cell and myeloperoxidase activity significantly increased after the second week of treatment. Whilst the numbers of red blood cell significantly decreased in the infected fish fed with 0.0 and 1.0% supplementation diet. After the third week of feeding, significant increases in the total protein, albumin level, lysozyme activity were recorded in the infected fish fed with 3.0% and 5.0% supplementation diet. Also, the enriched diets increased the survival rate as compared to the untreated group that suffered from 90% mortality. The present study indicated that S. alba extract may inhibit the mycelial growth of A. invadans effectively, suggesting an alternative to other chemotherapeutic agents, which brought much environmental and health concerns to the public, for EUS treatment.

Keywords: fungal pathogen, goldfish, organic extract, treatment

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Authors: Okolie Pius Ifeanyi, Emerenini Emilymary Chima

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Aims: The study investigated the diversity and identities of Lactic Acid Bacteria (LAB) isolated from different fresh fruits using Molecular Nested PCR analysis and the efficacy of cell free supernatants from Lactic Acid Bacteria (LAB) isolated from fresh fruits for in vitro control of some tomato pathogens. Study Design: Nested PCR approach was used in this study employing universal 16S rRNA gene primers in the first round PCR and LAB specific Primers in the second round PCR with the view of generating specific Nested PCR products for the LAB diversity present in the samples. The inhibitory potentials of supernatant obtained from LAB isolates of fruits origin that were molecularly characterized were investigated against some tomato phytopathogens using agar-well method with the view to develop biological agents for some tomato disease causing organisms. Methodology: Gram positive, catalase negative strains of LAB were isolated from fresh fruits on Man Rogosa and Sharpe agar (Lab M) using streaking method. Isolates obtained were molecularly characterized by means of genomic DNA extraction kit (Norgen Biotek, Canada) method. Standard methods were used for Nested Polymerase Chain Reaction (PCR) amplification targeting the 16S rRNA gene using universal 16S rRNA gene and LAB specific primers, agarose gel electrophoresis, purification and sequencing of generated Nested PCR products (Macrogen Inc., USA). The partial sequences obtained were identified by blasting in the non-redundant nucleotide database of National Center for Biotechnology Information (NCBI). The antimicrobial activities of characterized LAB against some tomato phytopathogenic bacteria which include (Xanthomonas campestries, Erwinia caratovora, and Pseudomonas syringae) were obtained by using the agar well diffusion method. Results: The partial sequences obtained were deposited in the database of National Centre for Biotechnology Information (NCBI). Isolates were identified based upon the sequences as Weissella cibaria (4, 18.18%), Weissella confusa (3, 13.64%), Leuconostoc paramensenteroides (1, 4.55%), Lactobacillus plantarum (8, 36.36%), Lactobacillus paraplantarum (1, 4.55%) and Lactobacillus pentosus (1, 4.55%). The cell free supernatants of LAB from fresh fruits origin (Weissella cibaria, Weissella confusa, Leuconostoc paramensenteroides, Lactobacillus plantarum, Lactobacillus paraplantarum and Lactobacillus pentosus) can inhibits these bacteria by creating clear zones of inhibition around the wells containing cell free supernatants of the above mentioned strains of lactic acid bacteria. Conclusion: This study shows that potentially LAB can be quickly characterized by molecular methods to specie level by nested PCR analysis of the bacteria isolate genomic DNA using universal 16S rRNA primers and LAB specific primer. Tomato disease causing organisms can be most likely biologically controlled by using extracts from LAB. This finding will reduce the potential hazard from the use of chemical herbicides on plant.

Keywords: nested pcr, molecular characterization, 16s rRNA gene, lactic acid bacteria

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Authors: Euripides Antoniades

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In 1821 the Greek Revolution movement, under the political influence that arose from the French revolution, and the corresponding movements in Italy, Germany and America, requested the liberation of the nation and the establishment of an independent national state. Published topics in the British press regarding the Greek Revolution, focused on : a) the right of the Greeks to claim their freedom from Turkish domination in order to establish an independent state based on the principle of national autonomy, b) criticism regarding Turkish rule as illegal and the power of the Ottoman Sultan as arbitrary, c) the recognition of the Greek identity and its distinction from the Turkish one and d) the endorsement Greeks as the descendants of ancient Greeks. The advantage of newspaper as a media is sharing information and ideas and dealing with issues in greater depth and detail, unlike other media, such as radio or television. The London Times is a print publication that presents, in chronological or thematic order, the news, opinions or announcements about the most important events that have occurred in a place during a specified period of time. This paper employs the rich archive of The London Times archive by quoting extracts from publications of that period, to convey the British public perspective regarding the Greek Revolution from its beginning until the London Protocol of 1828. Furthermore, analyses the publications of the British newspaper in terms of the number of references to the Greek revolution, front page and editorial references as well as the size of publications on the revolution during the period 1821-1828. A combination of qualitative and quantitative content analysis was applied. An attempt was made to record Greek Revolution references along with the usage of specific words and expressions that contribute to the representation of the historical events and their exposure to the reading public. Key finds of this research reveal that a) there was a frequency of passionate daily articles concerning the events in Greece, their length, and context in The London Times, b) the British public opinion was influenced by this particular newspaper and c) the newspaper published various news about the revolution by adopting the role of animator of the Greek struggle. For instance, war events and the battles of Wallachin and Moldavia, Hydra, Crete, Psara, Mesollogi, Peloponnese were presented not only for informing the readers but for promoting the essential need for freedom and the establishment of an independent Greek state. In fact, this type of news was the main substance of the The London Times’ structure, establishing a positive image about the Greek Revolution contributing to the European diplomatic development such as the standpoint of France, - that did not wish to be detached from the conclusions regarding the English loans and the death of Alexander I of Russia and his succession by the ambitious Nicholas. These factors offered a change in the attitude of the British and Russians respectively assuming a positive approach towards Greece. The Great Powers maintained a neutral position in the Greek-Ottoman conflict, same time they engaged in Greek power increasement by offering aid.

Keywords: Greece, revolution, newspaper, the London times, London, great britain, mass media

Procedia PDF Downloads 63

Authors: Khiem M. Nguyen, Ming C. Yang

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Chloroplast pigments are extremely important during photosynthesis since they play essential roles in light absorption and energy transfer. Therefore, understanding the efficiency of chlorophyll (Chl) biosynthesis could facilitate enhancement in photo-assimilates accumulation, and ultimately, in crop yield. The Chl-deficient mutants have been used extensively to study the Chl biosynthetic pathways and the biogenesis of the photosynthetic apparatus. Rice (Oryza sativa L.) is one of the most leading food crops, serving as staple food for many parts of the world. To author’s best knowledge, Chl b–lacking rice has been found; however the molecular mechanism of Chl biosynthesis still remains unclear compared to wild-type rice. In this study, the ultrastructure analysis, photosynthetic properties, and transcriptome profile of wild-type rice (Norin No.8, N8) and its Chl b-lacking mutant (Chlorina 1, C1) were examined. The finding concluded that total Chl content and Chl b content in the C1 leaves were strongly reduced compared to N8 leaves, suggesting that reduction in the total Chl content contributes to leaf color variation at the physiological level. Plastid ultrastructure of C1 possessed abnormal thylakoid membranes with loss of starch granule, large number of vesicles, and numerous plastoglobuli. The C1 rice also exhibited thinner stacked grana, which was caused by a reduction in the number of thylakoid membranes per granum. Thus, the different Chl a/b ratio of C1 may reflect the abnormal plastid development and function. Transcriptional analysis identified 23 differentially expressed genes (DEGs) and 671 transcription factors (TFs) that were involved in Chl metabolism, chloroplast development, cell division, and photosynthesis. The transcriptome profile and DEGs revealed that the gene encoding PsbR (PSII core protein) was down-regulated, therefore suggesting that the lower in light-harvesting complex proteins are responsible for the lower photosynthetic capacity in C1. In addition, expression level of cell division protein (FtsZ) genes were significantly reduced in C1, causing chloroplast division defect. A total of 19 DEGs were identified based on KEGG pathway assignment involving Chl biosynthesis pathway. Among these DEGs, the GluTR gene was down-regulated, whereas the UROD, CPOX, and MgCH genes were up-regulated. Observation through qPCR suggested that later stages of Chl biosynthesis were enhanced in C1, whereas the early stages were inhibited. Plastid structure analysis together with transcriptomic analysis suggested that the Chl a/b ratio was amplified both by the reduction in Chl contents accumulation, owning to abnormal chloroplast development, and by the enhanced conversion of Chl b to Chl a. Moreover, the results indicated the same Chl-cycle pattern in the wild-type and C1 rice, indicating another Chl b degradation pathway. Furthermore, the results demonstrated that normal grana stacking, along with the absence of Chl b and greatly reduced levels of Chl a in C1, provide evidence to support the conclusion that other factors along with LHCII proteins are involved in grana stacking. The findings of this study provide insight into the molecular mechanisms that underlie different Chl a/b ratios in rice.

Keywords: Chl-deficient mutant, grana stacked, photosynthesis, RNA-Seq, transcriptomic analysis

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Authors: Ani Kojoyan

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Linguistic studies have been investigating a connection between gender and linguistic development since 1970s. Scholars claim that gender differences in first and second language learning are socially constructed. Recent studies to language learning and gender reveal that second language acquisition is also a social phenomenon directly influencing one’s gender identity. Those responsible for designing language learning-teaching materials should be encouraged to understand the importance of and address the gender sensitivity accurately in textbooks. Writing or compiling a textbook is not an easy task; it requires strong academic abilities, patience, and experience. For a long period of time Armenia has been involved in the compilation process of a number of foreign language textbooks. However, there have been very few discussions or evaluations of those textbooks which will allow specialists to theorize that practice. The present paper focuses on the analysis of gender sensitivity issues and policy aspects involved in an EFL textbook. For the research the following material has been considered – “A Basic English Grammar: Morphology”, first printed in 2011. The selection of the material is not accidental. First, the mentioned textbook has been widely used in university teaching over years. Secondly, in Armenia “A Basic English Grammar: Morphology” has considered one of the most successful English grammar textbooks in a university teaching environment and served a source-book for other authors to compile and design their textbooks. The present paper aims to find out whether an EFL textbook is gendered in the Armenian teaching environment, and whether the textbook compilers are aware of gendered messages while compiling educational materials. It also aims at investigating students’ attitude toward the gendered messages in those materials. And finally, it also aims at increasing the gender sensitivity among book compilers and educators in various educational settings. For this study qualitative and quantitative research methods of analyses have been applied, the quantitative – in terms of carrying out surveys among students (45 university students, 18-25 age group), and the qualitative one – by discourse analysis of the material and conducting in-depth and semi-structured interviews with the Armenian compilers of the textbook (interviews with 3 authors). The study is based on passive and active observations and teaching experience done in a university classroom environment in 2014-2015, 2015-2016. The findings suggest that the discussed and analyzed teaching materials (145 extracts and examples) include traditional examples of intensive use of language and role-modelling, particularly, men are mostly portrayed as active, progressive, aggressive, whereas women are often depicted as passive and weak. These modeled often serve as a ‘reliable basis’ for reinforcing the traditional roles that have been projected on female and male students. The survey results also show that such materials contribute directly to shaping learners’ social attitudes and expectations around issues of gender. The applied techniques and discussed issues can be generalized and applied to other foreign language textbook compilation processes, since those principles, regardless of a language, are mostly the same.

Keywords: EFL textbooks, gender policy, gender sensitivity, qualitative and quantitative research methods

Procedia PDF Downloads 176

Authors: A. K. Tursunova, O. V. Chebonenko, A. Zh. Amirkulova, A. O. Abaildayev, O. A. Sapko, Y. M. Dyo, A. Sh. Utarbaeva

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Fusarium solani and its variants cause root and stem rot of plants. Dry rot is the most common disease of potato tubers during storage. The causative agents of fusariosis in contact with plants behave as antagonists, growth stimulants or parasites. The diversity of host-parasite relationships is explained by the parasite’s ability to produce a wide spectrum of biologically active compounds including toxins, enzymes, oligosaccharides, antibiotic substances, enniatins and gibberellins. Many of these metabolites contribute to the creation of compatible relations; others behave as elicitors, inducing various protective responses in plants. An important part of the strategy for developing plant resistance against pathogens is the activation of protein synthesis to produce protective ‘pathogenesis-related’ proteins. The family of PR-proteins known to confer the most protective response is chitinases (EC 3.2.1.14, Cht) and β-1,3-glucanases (EC 3.2.1.39, Glu). PR-proteins also include a large multigene family of peroxidases (EC 1.11.1.7, Pod), and increased activity of Pod and expression of the Pod genes leads to the development of resistance to a broad class of pathogens. Despite intensive research on the role of PR-proteins, the question of their participation in the mechanisms of formation of the F.solani–S.tuberosum pathosуstem is not sufficiently studied. Our aim was to investigate the effect of different classes of F. solani metabolites on the activity of chitinase, β-1,3-glucanases and peroxidases in tubers of Solanum tuberosum. Metabolite culture filtrate (CF) and cytoplasmic components were fractionated by extraction of the mycelium with organic solvents, salting out techniques, dialysis, column chromatography and ultrafiltration. Protein, lipid, carbohydrate and polyphenolic fractions of fungal metabolites were derived. Using enzymatic hydrolysis we obtained oligo glycans from fungal cell walls with different molecular weights. The activity of the metabolites was tested using potato tuber discs (d = 16mm, h = 5mm). The activity of PR-proteins of tubers was analyzed in a time course of 2–24 hours. The involvement of the analysed metabolites in the modulation of both early non-specific and late related to pathogenesis reactions was demonstrated. The most effective inducer was isolated from the CF (fraction of total phenolic compounds including naphtazarins). Induction of PR-activity by this fraction was: chitinase - 340-360%, glucanase - 435-450%, soluble forms of peroxidase - 400-560%, related forms of peroxidase - 215-237%. High-inducing activity was observed by the chloroform and acetonitrile extracts of the mycelium (induction of chitinase and glucanase activity was 176-240%, of soluble and bound forms of peroxidase - 190-400%). The fraction of oligo glycans mycelium cell walls of 1.2 kDa induced chitinase and β-1,3-glucanase to 239-320%; soluble forms and related peroxidase to 198-426%. Oligo glycans cell walls of 5-10 kDa had a weak suppressor effect - chitinase (21-25%) and glucanase (25-28%) activity; had no effect on soluble forms of peroxidase, but induced to 250-270% activity related forms. The CF polysaccharides of 8.5 kDa and 3.1 kDa inhibited synchronously the glucanase and chitinase specific response in step (after 24 hours at 42-50%) and the step response induced nonspecific peroxidase activity: soluble forms 4.8 -5.2 times, associated forms 1.4-1.6 times.

Keywords: fusarium solani, PR-proteins, peroxidase, solanum tuberosum

Procedia PDF Downloads 180

Authors: Maryam Hajrezaie, Mahmood Ameen Abdulla, Nazia Abdul Majid, Hapipa Mohd Ali, Pouya Hassandarvish, Maryam Zahedi Fard

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Background: Colon cancer is one of the most prevalent cancers in the world and is the third leading cause of death among cancers in both males and females. The incidence of colon cancer is ranked fourth among all cancers but varies in different parts of the world. Cancer chemoprevention is defined as the use of natural or synthetic compounds capable of inducing biological mechanisms necessary to preserve genomic fidelity. Andrographolide is the major labdane diterpenoidal constituent of the plant Andrographis paniculata (family Acanthaceae), used extensively in the traditional medicine. Extracts of the plant and their constituents are reported to exhibit a wide spectrum of biological activities of therapeutic importance. Laboratory animal model studies have provided evidence that Andrographolide play a role in inhibiting the risk of certain cancers. Objective: Our aim was to evaluate the chemopreventive efficacy of the Andrographolide in the AOM induced rat model. Methods: To evaluate inhibitory properties of andrographolide on colonic aberrant crypt foci (ACF), five groups of 7-week-old male rats were used. Group 1 (control group) were fed with 10% Tween 20 once a day, Group 2 (cancer control) rats were intra-peritoneally injected with 15 mg/kg Azoxymethan, Gropu 3 (drug control) rats were injected with 15 mg/kg azoxymethan and 5-Flourouracil, Group 4 and 5 (experimental groups) were fed with 10 and 20 mg/kg andrographolide each once a day. After 1 week, the treatment group rats received subcutaneous injections of azoxymethane, 15 mg/kg body weight, once weekly for 2 weeks. Control rats were continued on Tween 20 feeding once a day and experimental groups 10 and 20 mg/kg andrographolide feeding once a day for 8 weeks. All rats were sacrificed 8 weeks after the azoxymethane treatment. Colons were evaluated grossly and histopathologically for ACF. Results: Administration of 10 mg/kg and 20 mg/kg andrographolide were found to be effectively chemoprotective, as evidenced microscopily and biochemically. Andrographolide suppressed total colonic ACF formation up to 40% to 60%, respectively, when compared with control group. Pre-treatment with andrographolide, significantly reduced the impact of AOM toxicity on plasma protein and urea levels as well as on plasma aspartate aminotransferase (AST), alanine aminotransferase (ALT), lactate dehydrogenase (LDH) and gamma-glutamyl transpeptidase (GGT) activities. Grossly, colorectal specimens revealed that andrographolide treatments decreased the mean score of number of crypts in AOM-treated rats. Importantly, rats fed andrographolide showed 75% inhibition of foci containing four or more aberrant crypts. The results also showed a significant increase in glutathione (GSH), superoxide dismutase (SOD), nitric oxide (NO), and Prostaglandin E2 (PGE2) activities and a decrease in malondialdehyde (MDA) level. Histologically all treatment groups showed a significant decrease of dysplasia as compared to control group. Immunohistochemical staining showed up-regulation of Hsp70 and down-regulation of Bax proteins. Conclusion: The current study demonstrated that Andrographolide reduce the number of ACF. According to these data, Andrographolide might be a promising chemoprotective activity, in a model of AOM-induced in ACF.

Keywords: chemopreventive, andrographolide, colon cancer, aberrant crypt foci (ACF)

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Authors: Massimiliano Biagini, Marco Spinsanti, Gabriella De Angelis, Sara Tomei, Ilaria Ferlenghi, Maria Scarselli, Alessia Biolchi, Alessandro Muzzi, Brunella Brunelli, Silvana Savino, Marzia M. Giuliani, Isabel Delany, Paolo Costantino, Rino Rappuoli, Vega Masignani, Nathalie Norais

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The gram-negative bacterium Neisseria meningitidis serogroup B (MenB) is an exclusively human pathogen representing the major cause of meningitides and severe sepsis in infants and children but also in young adults. This pathogen is usually present in the 30% of healthy population that act as a reservoir, spreading it through saliva and respiratory fluids during coughing, sneezing, kissing. Among surface-exposed protein components of this diplococcus, factor H binding protein is a lipoprotein proved to be a protective antigen used as a component of the recently licensed Bexsero vaccine. fHbp is a highly variable meningococcal protein: to reflect its remarkable sequence variability, it has been classified in three variants (or two subfamilies), and with poor cross-protection among the different variants. Furthermore, the level of fHbp expression varies significantly among strains, and this has also been considered an important factor for predicting MenB strain susceptibility to anti-fHbp antisera. Different methods have been used to assess fHbp expression on meningococcal strains, however, all these methods use anti-fHbp antibodies, and for this reason, the results are affected by the different affinity that antibodies can have to different antigenic variants. To overcome the limitations of an antibody-based quantification, we developed a quantitative Mass Spectrometry (MS) approach. Selected Reaction Monitoring (SRM) recently emerged as a powerful MS tool for detecting and quantifying proteins in complex mixtures. SRM is based on the targeted detection of ProteoTypicPeptides (PTPs), which are unique signatures of a protein that can be easily detected and quantified by MS. This approach, proven to be highly sensitive, quantitatively accurate and highly reproducible, was used to quantify the absolute amount of fHbp antigen in total extracts derived from 105 clinical isolates, evenly distributed among the three main variant groups and selected to be representative of the fHbp circulating subvariants around the world. We extended the study at the genetic level investigating the correlation between the differential level of expression and polymorphisms present within the genes and their promoter sequences. The implications of fHbp expression on the susceptibility of the strain to killing by anti-fHbp antisera are also presented. To date this is the first comprehensive fHbp expression profiling in a large panel of Neisseria meningitidis clinical isolates driven by an antibody-independent MS-based methodology, opening the door to new applications in vaccine coverage prediction and reinforcing the molecular understanding of released vaccines.

Keywords: quantitative mass spectrometry, Neisseria meningitidis, vaccines, bexsero, molecular epidemiology

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Authors: Adriana Muñoz-Talavera, Ana Rosa Rincón-Sánchez, Abraham Escobedo-Moratilla, María Cristina Islas-Carbajal, Miguel Ángel Gómez-Lim

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The highest rates of diabetes incidence and prevalence worldwide will increase the number of diabetic patients requiring insulin or insulin analogues. Then, current production systems would not be sufficient to meet the future market demands. Therefore, developing efficient expression systems for insulin and insulin analogues are needed. In addition, insulin analogues with better pharmacokinetics and pharmacodynamics properties and without mitogenic potential will be required. SCI-57 (single chain insulin-57) is an insulin analogue having 10 times greater affinity to the insulin receptor, higher resistance to thermal degradation than insulin, native mitogenicity and biological effect. Plants as expression platforms have been used to produce recombinant proteins because of their advantages such as cost-effectiveness, posttranslational modifications, absence of human pathogens and high quality. Immunoglobulin production with a yield of 50% has been achieved by transient expression in Nicotiana benthamiana (Nb). The aim of this study is to produce SCI-57 by transient expression in Nb. Methodology: DNA sequence encoding SCI-57 was cloned in pICH31070. This construction was introduced into Agrobacterium tumefaciens by electroporation. The resulting strain was used to infiltrate leaves of Nb. In order to isolate SCI-57, leaves from transformed plants were incubated 3 hours with the extraction buffer therefore filtrated to remove solid material. The resultant protein solution was subjected to anion exchange chromatography on an FPLC system and ultrafiltration to purify SCI-57. Detection of SCI-57 was made by electrophoresis pattern (SDS-PAGE). Protein band was digested with trypsin and the peptides were analyzed by Liquid chromatography tandem-mass spectrometry (LC-MS/MS). A purified protein sample (20µM) was analyzed by ESI-Q-TOF-MS to obtain the ionization pattern and the exact molecular weight determination. Chromatography pattern and impurities detection were performed using RP-HPLC using recombinant insulin as standard. The identity of the SCI-57 was confirmed by anti-insulin ELISA. The total soluble protein concentration was quantified by Bradford assay. Results: The expression cassette was verified by restriction mapping (5393 bp fragment). The SDS-PAGE of crude leaf extract (CLE) of transformed plants, revealed a protein of about 6.4 kDa, non-present in CLE of untransformed plants. The LC-MS/MS results displayed one peptide with a high score that matches SCI-57 amino acid sequence in the sample, confirming the identity of SCI-57. From the purified SCI-57 sample (PSCI-57) the most intense charge state was 1069 m/z (+6) on the displayed ionization pattern corresponding to the molecular weight of SCI-57 (6412.6554 Da). The RP-HPLC of the PSCI-57 shows the presence of a peak with similar retention time (rt) and UV spectroscopic profile to the insulin standard (SCI-57 rt=12.96 and insulin rt=12.70 min). The collected SCI-57 peak had ELISA signal. The total protein amount in CLE from transformed plants was higher compared to untransformed plants. Conclusions: Our results suggest the feasibility to produce insulin analogue SCI-57 by transient expression in Nicotiana benthamiana. Further work is being undertaken to evaluate the biological activity by glucose uptake by insulin-sensitive and insulin-resistant murine and human cultured adipocytes.

Keywords: insulin analogue, mass spectrometry, Nicotiana benthamiana, transient expression

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Authors: D. Heinemann, S. Schramm, S. Knabner, D. Baumgarten

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Purpose: Applying solder paste to printed circuit boards (PCB) with stencils has been the method of choice over the past years. A new method uses a jet printer to deposit tiny droplets of solder paste through an ejector mechanism onto the board. This allows for more flexible PCB layouts with smaller components. Due to the viscosity of the solder paste, air blisters can be trapped in the cartridge. This can lead to missing solder joints or deviations in the applied solder volume. Therefore, a built-in and real-time inspection of the printing process is needed to minimize uncertainties and increase the efficiency of the process by immediate correction. The objective of the current study is the design of an optimal imaging system and the development of an automatic algorithm for the detection of applied solder joints from optical from the captured images. Methods: In a first approach, a camera module connected to a microcomputer and LED strips are employed to capture images of the printed circuit board under four different illuminations (white, red, green and blue). Subsequently, an improved system including a ring light, an objective lens, and a monochromatic camera was set up to acquire higher quality images. The obtained images can be divided into three main components: the PCB itself (i.e., the background), the reflections induced by unsoldered positions or screw holes and the solder joints. Non-uniform illumination is corrected by estimating the background using a morphological opening and subtraction from the input image. Image sharpening is applied in order to prevent error pixels in the subsequent segmentation. The intensity thresholds which divide the main components are obtained from the multimodal histogram using three probability density functions. Determining the intersections delivers proper thresholds for the segmentation. Remaining edge gradients produces small error areas which are removed by another morphological opening. For quantitative analysis of the segmentation results, the dice coefficient is used. Results: The obtained PCB images show a significant gradient in all RGB channels, resulting from ambient light. Using different lightings and color channels 12 images of a single PCB are available. A visual inspection and the investigation of 27 specific points show the best differentiation between those points using a red lighting and a green color channel. Estimating two thresholds from analyzing the multimodal histogram of the corrected images and using them for segmentation precisely extracts the solder joints. The comparison of the results to manually segmented images yield high sensitivity and specificity values. Analyzing the overall result delivers a Dice coefficient of 0.89 which varies for single object segmentations between 0.96 for a good segmented solder joints and 0.25 for single negative outliers. Conclusion: Our results demonstrate that the presented optical imaging system and the developed algorithm can robustly detect solder joints on printed circuit boards. Future work will comprise a modified lighting system which allows for more precise segmentation results using structure analysis.

Keywords: printed circuit board jet-printing, inspection, segmentation, solder paste detection

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Authors: Monica Racha, Siva Chandrasekaran, Alex Stojcevski

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The engineering programs focus on promoting students' personal and professional development by ensuring that students acquire technical and professional competencies during four-year studies. The traditional engineering laboratory provides an opportunity for students to "practice by doing," and laboratory facilities aid them in obtaining insight and understanding of their discipline. Due to rapid technological advancements and the current COVID-19 outbreak, the traditional labs were transforming into virtual learning environments. Aim: To better understand the limitations of the physical laboratory, this research study aims to use a Machine Learning (ML) algorithm that interfaces with the Augmented Reality HoloLens and predicts the image behavior to classify and detect the electronic components. The automated electronic components error classification and detection automatically detect and classify the position of all components on a breadboard by using the ML algorithm. This research will assist first-year undergraduate engineering students in conducting laboratory practices without any supervision. With the help of HoloLens, and ML algorithm, students will reduce component placement error on a breadboard and increase the efficiency of simple laboratory practices virtually. Method: The images of breadboards, resistors, capacitors, transistors, and other electrical components will be collected using HoloLens 2 and stored in a database. The collected image dataset will then be used for training a machine learning model. The raw images will be cleaned, processed, and labeled to facilitate further analysis of components error classification and detection. For instance, when students conduct laboratory experiments, the HoloLens captures images of students placing different components on a breadboard. The images are forwarded to the server for detection in the background. A hybrid Convolutional Neural Networks (CNNs) and Support Vector Machines (SVMs) algorithm will be used to train the dataset for object recognition and classification. The convolution layer extracts image features, which are then classified using Support Vector Machine (SVM). By adequately labeling the training data and classifying, the model will predict, categorize, and assess students in placing components correctly. As a result, the data acquired through HoloLens includes images of students assembling electronic components. It constantly checks to see if students appropriately position components in the breadboard and connect the components to function. When students misplace any components, the HoloLens predicts the error before the user places the components in the incorrect proportion and fosters students to correct their mistakes. This hybrid Convolutional Neural Networks (CNNs) and Support Vector Machines (SVMs) algorithm automating electronic component error classification and detection approach eliminates component connection problems and minimizes the risk of component damage. Conclusion: These augmented reality smart glasses powered by machine learning provide a wide range of benefits to supervisors, professionals, and students. It helps customize the learning experience, which is particularly beneficial in large classes with limited time. It determines the accuracy with which machine learning algorithms can forecast whether students are making the correct decisions and completing their laboratory tasks.

Keywords: augmented reality, machine learning, object recognition, virtual laboratories

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Authors: Jayanwita Sarkar, Usha Chakraborty, Bishwanath Chakraborty

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Plants are constantly interacting with various abiotic and biotic stresses. In changing climate scenario plants are continuously modifying physiological processes to adapt to changing environmental conditions which profoundly affect plant-pathogen interactions. Spot blotch in wheat is a fast-rising disease in the warmer plains of South Asia where the rise in minimum average temperature over most of the year already affecting wheat production. Hence, the study was undertaken to explore the role of elevated temperature in spot blotch disease development and modulation of antioxidative responses by plant growth promoting rhizobacteria (PGPR) for biocontrol of spot blotch at high temperature. Elevated temperature significantly increases the susceptibility of wheat plants to spot blotch causing pathogen Bipolaris sorokiniana. Two PGPR Bacillus safensis (W10) and Ochrobactrum pseudogrignonense (IP8) isolated from wheat (Triticum aestivum L.) and blady grass (Imperata cylindrical L.) rhizophere respectively, showing in vitro antagonistic activity against Bipolaris sorokiniana were tested for growth promotion and induction of resistance against spot blotch in wheat. GC-MS analysis showed that Bacillus safensis (W10) and Ochrobactrum pseudogrignonense (IP8) produced antifungal and antimicrobial compounds in culture. Seed priming with these two bacteria significantly increase growth, modulate antioxidative signaling and induce resistance and eventually reduce disease incidence in wheat plants at optimum as well as elevated temperature which was further confirmed by indirect immunofluorescence assay using polyclonal antibody raised against Bipolaris sorokiniana. Application of the PGPR led to enhancement in activities of plant defense enzymes- phenylalanine ammonia lyase, peroxidase, chitinase and β-1,3 glucanase in infected leaves. Immunolocalization of chitinase and β-1,3 glucanase in PGPR primed and pathogen inoculated leaf tissue was further confirmed by transmission electron microscopy using PAb of chitinase, β-1,3 glucanase and gold labelled conjugates. Activity of ascorbate-glutathione redox cycle related enzymes such as ascorbate peroxidase, superoxide dismutase and glutathione reductase along with antioxidants such as carotenoids, glutathione and ascorbate and osmolytes like proline and glycine betain accumulation were also increased during disease development in PGPR primed plant in comparison to unprimed plants at high temperature. Real-time PCR analysis revealed enhanced expression of defense genes- chalcone synthase and phenyl alanineammonia lyase. Over expression of heat shock proteins like HSP 70, small HSP 26.3 and heat shock factor HsfA3 in PGPR primed plants effectively protect plants against spot blotch infection at elevated temperature as compared with control plants. Our results revealed dynamic biochemical cross talk between elevated temperature and spot blotch disease development and furthermore highlight PGPR mediated array of antioxidative and molecular alterations responsible for induction of resistance against spot blotch disease at elevated temperature which seems to be associated with up-regulation of defense genes, heat shock proteins and heat shock factors, less ROS production, membrane damage, increased expression of redox enzymes and accumulation of osmolytes and antioxidants.

Keywords: antioxidative enzymes, defense enzymes, elevated temperature, heat shock proteins, PGPR, Real-Time PCR, spot blotch, wheat

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Authors: Fani Sakellariadou, Danae Antivachis

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Purpose of this paper is to evaluate the environmental status of a coastal Mediterranean lake, named Koumoundourou, located in the northeastern coast of Elefsis Bay, in the western region of Attiki in Greece, 15 km far from Athens. It is preserved from ancient times having an important archaeological interest. Koumoundourou lake is also considered as a valuable wetland accommodating an abundant flora and fauna, with a variety of bird species including a few world’s threatened ones. Furthermore, it is a heavily modified lake, affected by various anthropogenic pollutant sources which provide industrial, urban and agricultural contaminants. The adjacent oil refineries and the military depot are the major pollution providers furnishing with crude oil spills and leaks. Moreover, the lake accepts a quantity of groundwater leachates from the major landfill of Athens. The environmental status of the lake results from the intensive land uses combined with the permeable lithology of the surrounding area and the existence of karstic springs which discharge calcareous mountains. Sediment samples were collected along the shoreline of the lake using a Van Veen grab stainless steel sampler. They were studied for the determination of the total metal content and the metal fractionation in geochemical phases as well as the characterization of the dissolved organic matter (DOM). These constituents have a significant role in the ecological consideration of the lake. Metals may be responsible for harmful environmental impacts. The metal partitioning offers comprehensive information for the origin, mode of occurrence, biological and physicochemical availability, mobilization and transport of metals. Moreover, DOM has a multifunctional importance interacting with inorganic and organic contaminants leading to biogeochemical and ecological effects. The samples were digested using microwave heating with a suitable laboratory microwave unit. For the total metal content, the samples were treated with a mixture of strong acids. Then, a sequential extraction procedure was applied for the removal of exchangeable, carbonate hosted, reducible, organic/sulphides and residual fractions. Metal content was determined by an ICP-MS (Perkin Elmer, ICP MASS Spectrophotometer NexION 350D). Furthermore, the DOM was removed via a gentle extraction procedure and then it was characterized by fluorescence spectroscopy using a Perkin-Elmer LS 55 luminescence spectrophotometer equipped with the WinLab 4.00.02 software for data processing (Agilent, Cary Eclipse Fluorescence). Mono dimensional emission, excitation, synchronous-scan excitation and total luminescence spectra were recorded for the classification of chromophoric units present in the aqueous extracts. Total metal concentrations were determined and compared with those of the Elefsis gulf sediments. Element partitioning showed the anthropogenic sources and the contaminant bioavailability. All fluorescence spectra, as well as humification indices, were evaluated in detail to find out the nature and origin of DOM. All the results were compared and interpreted to evaluate the environmental quality of Koumoundourou lake and the need for environmental management and protection.

Keywords: anthropogenic contaminant, dissolved organic matter, lake, metal, pollution

Procedia PDF Downloads 128

Authors: Jing Jin

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The incorporation of artificial intelligence (AI) and machine learning (ML) in aircraft manufacturing and aerospace supply chains leads to the generation of a substantial amount of data among various tiers of suppliers and OEMs. Identifying the high-quality information challenges decision-makers. The application of AI/ML models necessitates access to 'high-quality' information to yield desired outputs. However, the process of information sharing introduces complexities, including distortion through various communication channels and biases introduced by both human and AI entities. This phenomenon significantly influences the quality of information, impacting decision-makers engaged in configuring supply chain systems. Traditionally, distorted information is categorized as 'low-quality'; however, this study challenges this perception, positing that distorted information, contributing to stakeholder goals, can be deemed high-quality within supply chains. The main aim of this study is to identify and evaluate the dimensions of information quality crucial to the UK aerospace supply chain. Guided by a central research question, "What information quality dimensions are considered when defining information quality in the UK aerospace supply chain?" the study delves into the intricate dynamics of information quality in the aerospace industry. Additionally, the research explores the nuanced impact of information distortion on stakeholders' decision-making processes, addressing the question, "How does the information distortion phenomenon influence stakeholders’ decisions regarding information quality in the UK aerospace supply chain system?" This study employs deductive methodologies rooted in positivism, utilizing a cross-sectional approach and a mono-quantitative method -a questionnaire survey. Data is systematically collected from diverse tiers of supply chain stakeholders, encompassing end-customers, OEMs, Tier 0.5, Tier 1, and Tier 2 suppliers. Employing robust statistical data analysis methods, including mean values, mode values, standard deviation, one-way analysis of variance (ANOVA), and Pearson’s correlation analysis, the study interprets and extracts meaningful insights from the gathered data. Initial analyses challenge conventional notions, revealing that information distortion positively influences the definition of information quality, disrupting the established perception of distorted information as inherently low-quality. Further exploration through correlation analysis unveils the varied perspectives of different stakeholder tiers on the impact of information distortion on specific information quality dimensions. For instance, Tier 2 suppliers demonstrate strong positive correlations between information distortion and dimensions like access security, accuracy, interpretability, and timeliness. Conversely, Tier 1 suppliers emphasise strong negative influences on the security of accessing information and negligible impact on information timeliness. Tier 0.5 suppliers showcase very strong positive correlations with dimensions like conciseness and completeness, while OEMs exhibit limited interest in considering information distortion within the supply chain. Introducing social network analysis (SNA) provides a structural understanding of the relationships between information distortion and quality dimensions. The moderately high density of ‘information distortion-by-information quality’ underscores the interconnected nature of these factors. In conclusion, this study offers a nuanced exploration of information quality dimensions in the UK aerospace supply chain, highlighting the significance of individual perspectives across different tiers. The positive influence of information distortion challenges prevailing assumptions, fostering a more nuanced understanding of information's role in the Industry 4.0 landscape.

Keywords: information distortion, information quality, supply chain configuration, UK aerospace industry

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Authors: Luisa Barreira, Viana Castaneda, Maria J. Rodrigues, Florinda Gama, Tamara Santos, Marta Oliveira, Catarina Pereira, Maribela Pestana, Pedro Correia, Miguel Salazar, Carla Nunes, Luisa Custodio, Joao Varela

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In the last century, the world witnessed an exponential demographic increase that has put an enormous pressure on agriculture and food production. Associated also with climate changes, there has been a decrease in the amount of available freshwater and an increased salinization of soils which can affect the production of most food crops. Halophytes, however, are plants able to withstand high salinities while maintaining a good growth productivity. To cope with the excess salt, they produce secondary metabolites (e.g. vitamins and phenolic compounds) which, along with the natural presence of some minerals, makes them not only nutritionally rich but also functional foods. Some halophytes, as quinoa or salicornia, are already used in some countries, mostly as gourmet food. Hydroponic cultivation of halophytes using seawater or diluted seawater for watering can decrease the pressure on freshwater resources while producing a nutritional and functional food. The XtremeGourmet project funded by the EU aims to develop and optimize the production of different halophytes by hydroponics. One of the more specific objectives of this project is the study of halophytes’ productivity and chemical composition under different abiotic conditions, e.g. salt and nutrient concentration and light intensity. Three species of halophytes commonly occurring in saltmarshes of the South of Portugal (Inula chrithmoides, Salicornia ramosissima and Mesembryanthemum nodiflorum) were cultivated using hydroponics under different salinities, ranging from 5 to 45 dS/m. For each condition, several parameters were assessed namely: total and commercial productivity, electrical conductivity, total soluble solids, proximal composition, mineral profile, total phenolics, flavonoids and condensed tannins content and antioxidant activity. Results show that productivity was significantly reduced for all plants with increasing salinity up to salinity 29 dS/m and remained low onwards. Oppositely, the electrical conductivity and the total soluble solids content of the produced plants increased with salinity, reaching a plateau at 29 dS/m. It seems that plants reflect the salt concentration of the water up to some point, being able to regulate their salt content for higher salinities. The same tendency was observed for the ash content of these plants, which is related to the mineral uptake from the cultivating media and the plants’ capacity to both accumulate and regulate ions’ concentration in their tissues. Nonetheless, this comes with a metabolic cost which is observed by a decrease in productivity. The mineral profile of these plants shows high concentrations of sodium but also high amounts of potassium. In what concerns the microelements, these plants appear to be a good source of manganese and iron and the low amounts of toxic metals account for their safe consumption in moderate amounts. Concerning the phenolics composition, plants presented moderate concentrations of phenolics but high amounts of condensed tannins, particularly I. crithmoides which accounts for its characteristic sour and spicy taste. Contrary to some studies in which higher amounts of phenolics were found in plants cultivated under higher salinities, in this study, the highest amount of phenolic compounds were found in plants grown at the lowest or intermediate salinities. Nonetheless, there was a positive correlation between the concentration of these compounds and the antioxidant capacity of the plants’ extracts.

Keywords: functional properties, halophytes, hydroponics, nutritional composition, salinity effect

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Authors: Achraf Al Faraj, Asma Sultana Shaik, Baraa Al Sayed

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Specific and effective targeting of drug delivery systems (DDS) to cancerous sites remains a major challenge for a better diagnostic and therapy. Recently, SWCNTs with their unique physicochemical properties and the ability to cross the cell membrane show promising in the biomedical field. The purpose of this study was first to develop a biocompatible iron oxide tagged SWCNTs as diagnostic nanoprobes to allow their noninvasive detection using MRI and their preferential targeting in a breast cancer murine model by placing an optimized flexible magnet over the tumor site. Magnetic targeting was associated to specific antibody-conjugated SWCNTs active targeting. The therapeutic efficacy of doxorubicin-conjugated SWCNTs was assessed, and the superiority of diffusion-weighted (DW-) MRI as sensitive imaging biomarker was investigated. Short Polyvinylpyrrolidone (PVP) stabilized water soluble SWCNTs were first developed, tagged with iron oxide nanoparticles and conjugated with Endoglin/CD105 monoclonal antibodies. They were then conjugated with doxorubicin drugs. SWCNTs conjugates were extensively characterized using TEM, UV-Vis spectrophotometer, dynamic light scattering (DLS) zeta potential analysis and electron spin resonance (ESR) spectroscopy. Their MR relaxivities (i.e. r1 and r2*) were measured at 4.7T and their iron content and metal impurities quantified using ICP-MS. SWCNTs biocompatibility and drug efficacy were then evaluated both in vitro and in vivo using a set of immunological assays. Luciferase enhanced bioluminescence 4T1 mouse mammary tumor cells (4T1-Luc2) were injected into the right inguinal mammary fat pad of Balb/c mice. Tumor bearing mice received either free doxorubicin (DOX) drug or SWCNTs with or without either DOX or iron oxide nanoparticles. A multi-pole 10x10mm high-energy flexible magnet was maintained over the tumor site during 2 hours post-injections and their properties and polarity were optimized to allow enhanced magnetic targeting of SWCNTs toward the primary tumor site. Tumor volume was quantified during the follow-up investigation study using a fast spin echo MRI sequence. In order to detect the homing of SWCNTs to the main tumor site, susceptibility-weighted multi-gradient echo (MGE) sequence was used to generate T2* maps. Apparent diffusion coefficient (ADC) measurements were also performed as a sensitive imaging biomarker providing early and better assessment of disease treatment. At several times post-SWCNT injection, histological analysis were performed on tumor extracts and iron-loaded SWCNT were quantified using ICP-MS in tumor sites, liver, spleen, kidneys, and lung. The optimized multi-poles magnet revealed an enhanced targeting of magnetic SWCNTs to the primary tumor site, which was found to be much higher than the active targeting achieved using antibody-conjugated SWCNTs. Iron-loading allowed their sensitive noninvasive tracking after intravenous administration using MRI. The active targeting of doxorubicin through magnetic antibody-conjugated SWCNTs nanoprobes was found to considerably decrease the primary tumor site and may have inhibited the development of metastasis in the tumor-bearing mice lung. ADC measurements in DW-MRI were found to significantly increase in a time-dependent manner after the injection of DOX-conjugated SWCNTs complexes.

Keywords: single-walled carbon nanotubes, nanomedicine, magnetic resonance imaging, cancer diagnosis and therapy

Procedia PDF Downloads 299