Search results for: gene bank

Authors: Isabel N. Sierra-Garcia, Maria J. Ferreira, Sandro Figuereido, Newton Gomes, Helena Silva, Angela Cunha

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Plant-associated microbial communities are considered crucial in the adaptation of halophytes to coastal environments. The plant microbiota can be horizontally acquired from the environment or vertically transmitted from generation to generation via seeds. Recruiting of the microbial communities by the plant is affected by geographical location, soil source, host genotype, and cultivation practice. There is limited knowledge reported on the microbial communities in halophytes the influence of biotic and abiotic factors. In this work, the microbiota associated with the halophyte Salicornia ramosissima was investigated to determine whether the structure of bacterial communities is influenced by host genotype or soil source. For this purpose, two contrasting sites where S. ramosissima is established in the estuarine system of the Ria de Aveiro were investigated. One site corresponds to a natural salt marsh where S. ramosissima plants are present (wild plants), and the other site is a former salt pan that nowadays are subjected to intensive crop production of S. ramosissima (crop plants). Bacterial communities from the rhizosphere, seeds and root endosphere of S. ramossisima from both sites were investigated by sequencing bacterial 16S rRNA gene using the Illumina MiSeq platform. The analysis of the sequences showed that the three plant-associated compartments, rhizosphere, root endosphere, and seed endosphere, harbor distinct microbiomes. However, bacterial richness and diversity were higher in seeds of wild plants, followed by rhizosphere in both sites, while seeds in the crop site had the lowest diversity. Beta diversity measures indicated that bacterial communities in root endosphere and seeds were more similar in both wild and crop plants in contrast to rhizospheres that differed by local, indicating that the recruitment of the similar bacterial communities by the plant genotype is active in regard to the site. Moreover, bacterial communities from the root endosphere and rhizosphere were phylogenetically more similar in both sites, but the phylogenetic composition of seeds in wild and crop sites was distinct. These results indicate that cultivation practices affect the seed microbiome. However, minimal vertical transmission of bacteria from seeds to adult plants is expected. Seeds from the crop site showed higher abundances of Kushneria and Zunongwangia genera. Bacterial members of the classes Alphaprotebacteria and Bacteroidia were the most ubiquitous across sites and compartments and might encompass members of the core microbiome. These findings indicate that bacterial communities associated with S. ramosissima are more influenced by host genotype rather than local abiotic factors or cultivation practices. This study provides a better understanding of the composition of the plant microbiota in S. ramosissima , which is essential to predict the interactions between plant and associated microbial communities and their effects on plant health. This knowledge is useful to the manipulations of these microbial communities to enhance the health and productivity of this commercially important plant.

Keywords: halophytes, plant microbiome, Salicornia ramosissima, agriculture

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Authors: Milu T. Cherian, Sergio C. Chai, Morgan A. Casal, Taosheng Chen

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This study aims to use CINPA1, a recently discovered small-molecule inhibitor of the xenobiotic receptor CAR (constitutive androstane receptor) for understanding the binding modes of CAR and to guide CAR-mediated gene expression profiling studies in human primary hepatocytes. CAR and PXR are xenobiotic sensors that respond to drugs and endobiotics by modulating the expression of metabolic genes that enhance detoxification and elimination. Elevated levels of drug metabolizing enzymes and efflux transporters resulting from CAR activation promote the elimination of chemotherapeutic agents leading to reduced therapeutic effectiveness. Multidrug resistance in tumors after chemotherapy could be associated with errant CAR activity, as shown in the case of neuroblastoma. CAR inhibitors used in combination with existing chemotherapeutics could be utilized to attenuate multidrug resistance and resensitize chemo-resistant cancer cells. CAR and PXR have many overlapping modulating ligands as well as many overlapping target genes which confounded attempts to understand and regulate receptor-specific activity. Through a directed screening approach we previously identified a new CAR inhibitor, CINPA1, which is novel in its ability to inhibit CAR function without activating PXR. The cellular mechanisms by which CINPA1 inhibits CAR function were also extensively examined along with its pharmacokinetic properties. CINPA1 binding was shown to change CAR-coregulator interactions as well as modify CAR recruitment at DNA response elements of regulated genes. CINPA1 was shown to be broken down in the liver to form two, mostly inactive, metabolites. The structure-activity differences of CINPA1 and its metabolites were used to guide computational modeling using the CAR-LBD structure. To rationalize how ligand binding may lead to different CAR pharmacology, an analysis of the docked poses of human CAR bound to CITCO (a CAR activator) vs. CINPA1 or the metabolites was conducted. From our modeling, strong hydrogen bonding of CINPA1 with N165 and H203 in the CAR-LBD was predicted. These residues were validated to be important for CINPA1 binding using single amino-acid CAR mutants in a CAR-mediated functional reporter assay. Also predicted were residues making key hydrophobic interactions with CINPA1 but not the inactive metabolites. Some of these hydrophobic amino acids were also identified and additionally, the differential coregulator interactions of these mutants were determined in mammalian two-hybrid systems. CINPA1 represents an excellent starting point for future optimization into highly relevant probe molecules to study the function of the CAR receptor in normal- and pathophysiology, and possible development of therapeutics (for e.g. use for resensitizing chemoresistant neuroblastoma cells).

Keywords: antagonist, chemoresistance, constitutive androstane receptor (CAR), multi-drug resistance, structure activity relationship (SAR), xenobiotic resistance

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Authors: Sevelina Ndlovu

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Financial Technology (fintech) advancement is recognised as one of the most transformational innovations in the financial industry. Fintech has given rise to internet-only digital banking, a novel financial technology advancement, and innovation that allows banking services through internet applications with no need for physical branches. This technology is becoming a new banking normal among consumers for its ubiquitous and real-time access advantages. There is evident switching and migration from traditional banking towards these fintech facilities, which could possibly pose a systemic risk if not properly understood and monitored. Fintech advancement has also brought about the emergence and escalation of financial technology consumption themes such as trust, security, perceived risk, and sustainability within the banking industry, themes scarcely covered in existing theoretic literature. To that end, the objective of this research is to investigate factors that determine fintech adoption and propose an integrated adoption model. This study aims to establish what the significant drivers of adoption are and develop a conceptual model that integrates technological, behavioral, and environmental constructs by extending the Unified Theory of Acceptance and Use of Technology 2 (UTAUT2). It proposes integrating constructs that influence financial consumption themes such as trust, perceived risk, security, financial incentives, micro-investing opportunities, and environmental consciousness to determine the impact of these factors on the adoption and intention to use digital banking apps. The main advantage of this conceptual model is the consolidation of a greater number of predictor variables that can provide a fuller explanation of the consumer's adoption of digital banking Apps. Moderating variables of age, gender, and income are incorporated. To the best of author’s knowledge, this study is the first that extends the UTAUT2 model with this combination of constructs to investigate user’s intention to adopt internet-only digital banking apps in the UK context. By investigating factors that are not included in the existing theories but are highly pertinent to the adoption of internet-only banking services, this research adds to existing knowledge and extends the generalisability of the UTAUT2 in a financial services adoption context. This is something that fills a gap in knowledge, as highlighted to needing further research on UTAUT2 after reviewing the theory in 2016 from its original version of 2003. To achieve the objectives of this study, this research assumes a quantitative research approach to empirically test the hypotheses derived from existing literature and pilot studies to give statistical support to generalise the research findings for further possible applications in theory and practice. This research is explanatory or casual in nature and uses cross-section primary data collected through a survey method. Convenient and purposive sampling using structured self-administered online questionnaires is used for data collection. The proposed model is tested using Structural Equation Modelling (SEM), and the analysis of primary data collected through an online survey is processed using Smart PLS software with a sample size of 386 digital bank users. The results are expected to establish if there are significant relationships between the dependent and independent variables and establish what the most influencing factors are.

Keywords: banking applications, digital banking, financial technology, technology adoption, UTAUT2

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Authors: Yana Posmitnaya, Galina Rudnitskaya, Tatyana Lukashenko, Anton Bukatin, Anatoly Evstrapov

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An important area of biological and medical research is the study of genetic mutations and polymorphisms that can alter gene function and cause inherited diseases and other diseases. The following methods to analyse DNA fragments are used: capillary electrophoresis and electrophoresis on microfluidic chip (MFC), mass spectrometry with electrophoresis on MFC, hybridization assay on microarray. Electrophoresis on MFC allows to analyse small volumes of samples with high speed and throughput. A soft lithography in polydimethylsiloxane (PDMS) was chosen for operative fabrication of MFCs. A master-form from silicon and photoresist SU-8 2025 (MicroChem Corp.) was created for the formation of micro-sized structures in PDMS. A universal topology which combines T-injector and simple cross was selected for the electrophoretic separation of the sample. Glass K8 and PDMS Sylgard® 184 (Dow Corning Corp.) were used for fabrication of MFCs. Electroosmotic flow (EOF) plays an important role in the electrophoretic separation of the sample. Therefore, the estimate of the quantity of EOF and the ways of its regulation are of interest for the development of the new methods of the electrophoretic separation of biomolecules. The following methods of surface modification were chosen to change EOF: high-frequency (13.56 MHz) plasma treatment in oxygen and argon at low pressure (1 mbar); 1% aqueous solution of polyvinyl alcohol; 3% aqueous solution of Kolliphor® P 188 (Sigma-Aldrich Corp.). The electroosmotic mobility was evaluated by the method of Huang X. et al., wherein the borate buffer was used. The influence of physical and chemical methods of treatment on the wetting properties of the PDMS surface was controlled by the sessile drop method. The most effective way of surface modification of MFCs, from the standpoint of obtaining the smallest value of the contact angle and the smallest value of the EOF, was the processing with aqueous solution of Kolliphor® P 188. This method of modification has been selected for the treatment of channels of MFCs, which are used for the separation of mixture of oligonucleotides fluorescently labeled with the length of chain with 10, 20, 30, 40 and 50 nucleotides. Electrophoresis was performed on the device MFAS-01 (IAI RAS, Russia) at the separation voltage of 1500 V. 6% solution of polydimethylacrylamide with the addition of 7M carbamide was used as the separation medium. The separation time of components of the mixture was determined from electropherograms. The time for untreated MFC was ~275 s, and for the ones treated with solution of Kolliphor® P 188 – ~ 220 s. Research of physical-chemical methods of surface modification of MFCs allowed to choose the most effective way for reducing EOF – the modification with aqueous solution of Kolliphor® P 188. In this case, the separation time of the mixture of oligonucleotides decreased about 20%. The further optimization of method of modification of channels of MFCs will allow decreasing the separation time of sample and increasing the throughput of analysis.

Keywords: electrophoresis, microfluidic chip, modification, nucleic acid, polydimethylsiloxane, soft lithography

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Authors: Tracey Woods

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In the ever-evolving landscape of academia, the traditional role of the librarian has undergone a remarkable transformation. Once considered as custodians of books and gatekeepers of information, librarians have the potential to take on the vital role of facilitators of cross and inter-disciplinary projects. This shift is driven by the growing recognition of the value of interdisciplinary collaboration in addressing complex research questions in pursuit of novel solutions to real-world problems. This paper shall explore the potential of the academic librarian’s role in facilitating innovative, multi-disciplinary projects, both recognising and validating the vital role that the librarian plays in a somewhat underplayed profession. Academic libraries support teaching, the strengthening of knowledge discourse, and, potentially, the development of innovative practices. As the role of the library gradually morphs from a quiet repository of books to a community-based information hub, a potential opportunity arises. The academic librarian’s role is to build knowledge across a wide span of topics, from the advancement of AI to subject-specific information, and, whilst librarians are generally not offered the research opportunities and funding that the traditional academic disciplines enjoy, they are often invited to help build research in support of the academic. This identifies that one of the primary skills of any 21st-century librarian must be the ability to collaborate and facilitate multi-disciplinary projects. In universities seeking to develop research diversity and academic performance, there is an increasing awareness of the need for collaboration between faculties to enable novel directions and advancements. This idea has been documented and discussed by several researchers; however, there is not a great deal of literature available from recent studies. Having a team based in the library that is adept at creating effective collaborative partnerships is valuable for any academic institution. This paper outlines the development of such a project, initiated within and around an identified library-specific need: the replication of fragile special collections for object-based learning. The research was developed as a multi-disciplinary project involving the faculties of engineering (digital twins lab), architecture, design, and education. Centred around methods for developing a fragile archive into a series of tactile objects furthers knowledge and understanding in both the role of the library as a facilitator of projects, chairing and supporting, alongside contributing to the research process and innovating ideas through the bank of knowledge found amongst the staff and their liaising capabilities. This paper shall present the method of project development from the initiation of ideas to the development of prototypes and dissemination of the objects to teaching departments for analysis. The exact replication of artefacts is also balanced with the adaptation and evolutionary speculations initiated by the design team when adapted as a teaching studio method. The dynamic response required from the library to generate and facilitate these multi-disciplinary projects highlights the information expertise and liaison skills that the librarian possesses. As academia embraces this evolution, the potential for groundbreaking discoveries and innovative solutions across disciplines becomes increasingly attainable.

Keywords: Liaison librarian, multi-disciplinary collaborations, library innovations, librarian stakeholders

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Authors: Diego Santibañez Oyarce, Fernanda Bravo Cornejo, Camilo Cerda Sarabia, Belén Díaz Díaz, Esteban Gómez Terán, Hugo Osses Prado, Raúl Caulier-Cisterna, Jorge Vergara-Quezada, Ana Moya-Beltrán

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The advent of high-throughput sequencing technologies has revolutionized genomics, generating vast amounts of genetic data that challenge traditional bioinformatics methods. Machine learning addresses these challenges by leveraging computational power to identify patterns and extract information from large datasets. However, biological sequence data, being symbolic and non-numeric, must be converted into numerical formats for machine learning algorithms to process effectively. So far, some encoding methods, such as one-hot encoding or k-mers, have been explored. This work proposes additional approaches for encoding DNA sequences in order to compare them with existing techniques and determine if they can provide improvements or if current methods offer superior results. Data from the 16S rRNA gene, a universal marker, was used to analyze eight bacterial groups that are significant in the pulmonary environment and have clinical implications. The bacterial genes included in this analysis are Prevotella, Abiotrophia, Acidovorax, Streptococcus, Neisseria, Veillonella, Mycobacterium, and Megasphaera. These data were downloaded from the NCBI database in Genbank file format, followed by a syntactic analysis to selectively extract relevant information from each file. For data encoding, a sequence normalization process was carried out as the first step. From approximately 22,000 initial data points, a subset was generated for testing purposes. Specifically, 55 sequences from each bacterial group met the length criteria, resulting in an initial sample of approximately 440 sequences. The sequences were encoded using different methods, including one-hot encoding, k-mers, Fourier transform, and Wavelet transform. Various machine learning algorithms, such as support vector machines, random forests, and neural networks, were trained to evaluate these encoding methods. The performance of these models was assessed using multiple metrics, including the confusion matrix, ROC curve, and F1 Score, providing a comprehensive evaluation of their classification capabilities. The results show that accuracies between encoding methods vary by up to approximately 15%, with the Fourier transform obtaining the best results for the evaluated machine learning algorithms. These findings, supported by the detailed analysis using the confusion matrix, ROC curve, and F1 Score, provide valuable insights into the effectiveness of different encoding methods and machine learning algorithms for genomic data analysis, potentially improving the accuracy and efficiency of bacterial classification and related genomic studies.

Keywords: DNA encoding, machine learning, Fourier transform, Fourier transformation

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Authors: Zhihao Peng, Qiong Zhang, Xiyao Li, Yongzhen Peng

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Nowadays, conventional nitrogen removal process (nitrification and denitrification) was adopted in most wastewater treatment plants, but many problems have occurred, such as: high aeration energy consumption, extra carbon sources dosage and high sludge treatment costs. The emergence of anammox has bring about the great revolution to the nitrogen removal technology, and only the ammonia and nitrite were required to remove nitrogen autotrophically, no demand for aeration and sludge treatment. However, there existed many challenges in anammox applications: difficulty of biomass retention, insufficiency of nitrite substrate, damage from complex organic etc. Much effort was put into the research in overcoming the above challenges, and the payment was rewarded. It was also imperative to establish an innovative process that can settle the above problems synchronously, after all any obstacle above mentioned can cause the collapse of anammox system. Therefore, in this study, a two-stage process was established that the sequencing batch reactor (SBR) and upflow anaerobic sludge blanket (UASB) were used in the pre-stage and post-stage, respectively. The domestic wastewater entered into the SBR first and went through anaerobic/aerobic/anoxic (An/O/A) mode, and the draining at the aerobic end of SBR was mixed with domestic wastewater, the mixture then entering to the UASB. In the long term, organic and nitrogen removal performance was evaluated. All along the operation, most COD was removed in pre-stage (COD removal efficiency > 64.1%), including some macromolecular organic matter, like: tryptophan, tyrosinase and fulvic acid, which could weaken the damage of organic matter to anammox. And the An/O/A operating mode of SBR was beneficial to the achievement and maintenance of partial nitrification (PN). Hence, sufficient and steady nitrite supply was another favorable condition to anammox enhancement. Besides, the flexible mixing ratio helped to gain a substrate ratio appropriate to anammox (1.32-1.46), which further enhance the anammox. Further, the UASB was used and gas recirculation strategy was adopted in the post-stage, aiming to achieve granulation by the selection pressure. As expected, the granules formed rapidly during 38 days, which increased from 153.3 to 354.3 μm. Based on bioactivity and gene measurement, the anammox metabolism and abundance level rose evidently, by 2.35 mgN/gVss·h and 5.3 x109. The anammox bacteria mainly distributed in the large granules (>1000 μm), while the biomass in the flocs (<200 μm) and microgranules (200-500 μm) barely displayed anammox bioactivity. Enhanced anammox promoted the advanced autotrophic nitrogen removal, which increased from 71.9% to 93.4%, even when the temperature was only 12.9 ℃. Therefore, it was feasible to enhance anammox in the multiple favorable conditions created, and the strategy extended the application of anammox to the full-scale mainstream, enhanced the understanding of anammox in the aspects of culturing conditions.

Keywords: anammox, granules, nitrite accumulation, nitrogen removal efficiency

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Authors: Mala Mukherjee

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Background: India is urbanizing very fast and urbanisation in India is treated as one of the most crucial components of economic growth. Though the pace of urbanisation (31.6 per cent in 2011) is however slower and lower than the average for Asia but the absolute number of people residing in cities and towns has increased substantially. Rapid urbanization leads to urban poverty and it is well represented in slums. Currently India has four metropolises and 53 million plus cities. All of them have significant slum population but the standard of living and success of slum development programmes varies across regions. Objectives: Objectives of the paper are to show how urbanisation and slum development varies across space; to show spatial variation in the standard of living in Indian slums; to analyse how the implementation of slum development policies like JNNURM, Rajiv Awas Yojana varies across cities and bring different results in different regions and what are the factors responsible for such variation. Data Sources and Methodology: Census 2011 data on urban population and slum households and amenities have been used for analysing the regional variation of urbanisation in 53 million plus cities of India. Special focus has been put on Kolkata Metropolitan Area. Statistical techniques like z-score and PCA have been employed to work out Standard of Living Deprivation score for all the slums of 53 metropolises. ARC-GIS software is used for making maps. Standard of living has been measured in terms of access to basic amenities, infrastructure and assets like drinking water, sanitation, housing condition, bank account, and so on. Findings: 1. The first finding reveals that migration and urbanization is very high in Greater Mumbai, Delhi, Bangaluru, Chennai, Hyderabad and Kolkata; but slum population is high in Greater Mumbai (50% population live in slums), Meerut, Faridabad, Ludhiana, Nagpur, Kolkata etc. Though the rate of urbanization is high in southern and western states but the percentage of slum population is high in northern states (except Greater Mumbai). 2. Standard of Living also varies widely. Slums of Greater Mumbai and North Indian Cities score fairly high in the index indicating the fact that standard of living is high in those slums compare to the slums in eastern India (Dhanbad, Jamshedpur, Kolkata). Therefore, though Kolkata have relatively lesser percentage of slum population compare to north and south Indian cities but the standard of living in Kolkata’s slums is deplorable. 3. It is interesting to note that even within Kolkata Metropolitan Area slums located in the southern and eastern municipal towns like Rajpur-Sonarpur, Pujali, Diamond Harbour, Baduria and Dankuni have lower standard of living compare to the slums located in the Hooghly Industrial belt like Titagarh, Rishrah, Srerampore etc. Slums of the Hooghly Industrial Belt are older than the slums located in eastern and southern part of the urban agglomeration. 4. Therefore, urban development and emergence of slums should not be the only issue of urban governance but standard of living should be the main focus. Slums located in the main cities like Delhi, Mumbai, Kolkata get more attention from the urban planners and similarly, older slums in a city receives greater political attention compare to the slums of smaller cities and newly emerged slums of the peripheral parts.

Keywords: urbanisation, slum, spatial variation, India

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Authors: Mingxin Li, Liya Ni

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Autonomous parking is a valuable feature applicable to many robotics applications such as tour guide robots, UV sanitizing robots, food delivery robots, and warehouse robots. With auto-parking, the robot will be able to park at the charging zone and charge itself without human intervention. As compared to self-driving vehicles, auto-parking is more challenging for a small-scale mobile robot only equipped with a front camera due to the camera view limited by the robot’s height and the narrow Field of View (FOV) of the inexpensive camera. In this research, auto-parking of a small-scale mobile robot with a front camera only was achieved in a four-step process: Firstly, transfer learning was performed on the AlexNet, a popular pre-trained convolutional neural network (CNN). It was trained with 150 pictures of empty parking slots and 150 pictures of occupied parking slots from the view angle of a small-scale robot. The dataset of images was divided into a group of 70% images for training and the remaining 30% images for validation. An average success rate of 95% was achieved. Secondly, the image of detected empty parking space was processed with edge detection followed by the computation of parametric representations of the boundary lines using the Hough Transform algorithm. Thirdly, the positions of the entrance point and center of available parking space were predicted based on the robot kinematic model as the robot was driving closer to the parking space because the boundary lines disappeared partially or completely from its camera view due to the height and FOV limitations. The robot used its wheel speeds to compute the positions of the parking space with respect to its changing local frame as it moved along, based on its kinematic model. Lastly, the predicted entrance point of the parking space was used as the reference for the motion control of the robot until it was replaced by the actual center when it became visible again by the robot. The linear and angular velocities of the robot chassis center were computed based on the error between the current chassis center and the reference point. Then the left and right wheel speeds were obtained using inverse kinematics and sent to the motor driver. The above-mentioned four subtasks were all successfully accomplished, with the transformed learning, image processing, and target prediction performed in MATLAB, while the motion control and image capture conducted on a self-built small scale differential drive mobile robot. The small-scale robot employs a Raspberry Pi board, a Pi camera, an L298N dual H-bridge motor driver, a USB power module, a power bank, four wheels, and a chassis. Future research includes three areas: the integration of all four subsystems into one hardware/software platform with the upgrade to an Nvidia Jetson Nano board that provides superior performance for deep learning and image processing; more testing and validation on the identification of available parking space and its boundary lines; improvement of performance after the hardware/software integration is completed.

Keywords: autonomous parking, convolutional neural network, image processing, kinematics-based prediction, transfer learning

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Authors: Yuriko Takayama, Norihiro Kato

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Quorum sensing (QS) is a cell-to-cell communication system in bacteria to regulate expression of target genes. In gram-negative bacteria, activation on QS is controlled by a concentration increase of N-acylhomoserine lactone (AHL), which can diffuse in and out of the cell. Effective control of QS is expected to avoid virulence factor production in infectious pathogens, biofilm formation, and antibiotic production because various cell functions in gram-negative bacteria are controlled by AHL-mediated QS. In this research, we applied cyclodextrins (CDs) as artificial hosts for the AHL signal to reduce the AHL concentration in the culture broth below its threshold for QS activation. The AHL-receptor complex induced under the high AHL concentration activates transcription of the QS-target gene. Accordingly, artificial reduction of the AHL concentration is one of the effective strategies to inhibit the QS. A hydrophobic cavity of the CD can interact with the acyl-chain of the AHL due to hydrophobic interaction in aqueous media. We studied N-hexanoylhomoserine lactone (C6HSL)-mediated QS in Serratia marcescens; accumulation of C6HSL is responsible for regulation of the expression of pig cluster. Inhibitory effects of added CDs on QS were demonstrated by determination of prodigiosin amount inside cells after reaching stationary phase, because production of prodigiosin depends on the C6HSL-mediated QS. By adding approximately 6 wt% hydroxypropyl-β-CD (HP-β-CD) in Luria-Bertani (LB) medium prior to inoculation of S. maecescens AS-1, the intracellularly accumulated prodigiosin was drastically reduced to 7-10%, which was determined after the extraction of prodigiosin in acidified ethanol. The AHL retention ability of HP-β-CD was also demonstrated by Chromobacterium violacuem CV026 bioassay. The CV026 strain is an AHL-synthase defective mutant that activates QS solely by adding AHLs from outside of cells. A purple pigment violacein is induced by activation of the AHL-mediated QS. We demonstrated that the violacein production was effectively suppressed when the C6HSL standard solution was spotted on a LB agar plate dispersing CV026 cells and HP-β-CD. Physico-chemical analysis was performed to study the affinity between the immobilized CD and added C6HSL using a quartz crystal microbalance (QCM) sensor. The COOH-terminated self-assembled monolayer was prepared on a gold electrode of 27-MHz AT-cut quartz crystal. Mono(6-deoxy-6-N, N-diethylamino)-β-CD was immobilized on the electrode using water-soluble carbodiimide. The C6HSL interaction with the β-CD cavity was studied by injecting the C6HSL solution to a cup-type sensor cell filled with buffer solution. A decrement of resonant frequency (ΔFs) clearly showed the effective C6HSL complexation with immobilized β-CD and its stability constant for MBP-SpnR-C6HSL complex was on the order of 102 M-1. The CD has high potential for engineered control of QS because it is safe for human use.

Keywords: acylhomoserine lactone, cyclodextrin, intracellular signaling, quorum sensing

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Authors: Almudena Espin Perez, Tyler Risom, Carl Pelz, Isabel English, Robert M. Angelo, Rosalie Sears, Andrew J. Gentles

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Pancreatic ductal adenocarcinoma (PDAC) is one of the most deadly malignancies. The role of the tumor microenvironment (TME) is gaining significant attention in cancer research. Despite ongoing efforts, the nature of the interactions between tumors, immune cells, and stromal cells remains poorly understood. The cell-intrinsic properties that govern cell lineage plasticity in PDAC and extrinsic influences of immune populations require technically challenging approaches due to the inherently heterogeneous nature of PDAC. Understanding the cell lineage plasticity of PDAC will improve the development of novel strategies that could be translated to the clinic. Members of the team have demonstrated that the acquisition of ductal to neuroendocrine lineage plasticity in PDAC confers therapeutic resistance and is a biomarker of poor outcomes in patients. Our approach combines computational methods for deconvolving bulk transcriptomic cancer data using CIBERSORTx and high-throughput single-cell imaging using Multiplexed Ion Beam Imaging (MIBI) to study lineage plasticity in PDAC and its relationship to the infiltrating immune system. The CIBERSORTx algorithm uses signature matrices from immune cells and stroma from sorted and single-cell data in order to 1) infer the fractions of different immune cell types and stromal cells in bulked gene expression data and 2) impute a representative transcriptome profile for each cell type. We studied a unique set of 300 genomically well-characterized primary PDAC samples with rich clinical annotation. We deconvolved the PDAC transcriptome profiles using CIBERSORTx, leveraging publicly available single-cell RNA-seq data from normal pancreatic tissue and PDAC to estimate cell type proportions in PDAC, and digitally reconstruct cell-specific transcriptional profiles from our study dataset. We built signature matrices and optimized by simulations and comparison to ground truth data. We identified cell-type-specific transcriptional programs that contribute to cancer cell lineage plasticity, especially in the ductal compartment. We also studied cell differentiation hierarchies using CytoTRACE and predict cell lineage trajectories for acinar and ductal cells that we believe are pinpointing relevant information on PDAC progression. Collaborators (Angelo lab, Stanford University) has led the development of the Multiplexed Ion Beam Imaging (MIBI) platform for spatial proteomics. We will use in the very near future MIBI from tissue microarray of 40 PDAC samples to understand the spatial relationship between cancer cell lineage plasticity and stromal cells focused on infiltrating immune cells, using the relevant markers of PDAC plasticity identified from the RNA-seq analysis.

Keywords: deconvolution, imaging, microenvironment, PDAC

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Authors: Gary J. Pickering, Sarah Lucas, Catherine E. Klodnicki, Nicole J. Gaudette

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Two taste pheno types that are of interest in the study of habitual diet-related risk factors and disease are 6-n-propylthiouracil (PROP) responsiveness and thermal tasting. Individuals differ considerable in how intensely they experience the bitterness of PROP, which is partially explained by three major single nucleotide polymorphisms associated with the TAS2R38 gene. Importantly, this variable responsiveness is a useful proxy for general taste responsiveness, and links to diet-related disease risk, including body mass index, in some studies. Thermal tasting - a newly discovered taste phenotype independent of PROP responsiveness - refers to the capacity of many individuals to perceive phantom tastes in response to lingual thermal stimulation, and is linked with TRPM5 channels. Thermal tasters (TTs) also experience oral sensations more intensely than thermal non-tasters (TnTs), and this was shown to associate with differences in self-reported food preferences in a previous survey from our lab. Here we report on two related studies, where we sought to determine whether PROP responsiveness and thermal tasting would associate with perceptual differences in the oral sensations elicited by sampled energy-dense foods, and whether in turn this would influence liking. We hypothesized that hyper-tasters (thermal tasters and individuals who experience PROP intensely) would (a) rate sweet and high-fat foods more intensely than hypo-tasters, and (b) would differ from hypo-tasters in liking scores. (Liking has been proposed recently as a more accurate measure of actual food consumption). In Study 1, a range of energy-dense foods and beverages, including table cream and chocolate, was assessed by 25 TTs and 19 TnTs. Ratings of oral sensation intensity and overall liking were obtained using gVAS and gDOL scales, respectively. TTs and TnTs did not differ significantly in intensity ratings for most stimuli (ANOVA). In a 2nd study, 44 female participants sampled 22 foods and beverages, assessing them for intensity of oral sensations (gVAS) and overall liking (9-point hedonic scale). TTs (n=23) rated their overall liking of creaminess and milk products lower than did TnTs (n=21), and liked milk chocolate less. PROP responsiveness was negatively correlated with liking of food and beverages belonging to the sweet or sensory food grouping. No other differences in intensity or liking scores between hyper- and hypo-tasters were found. Taken overall, our results are somewhat unexpected, lending only modest support to the hypothesis that these taste phenotypes associate with energy-dense food liking and consumption through differences in the oral sensations they elicit. Reasons for this lack of concordance with expectations and some prior literature are discussed, and suggestions for future research are advanced.

Keywords: taste phenotypes, sensory evaluation, PROP, thermal tasting, diet-related health risk

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Authors: Jeet Chakraborty, Sanjay Dutta

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Antibiotic resistance by bacteria has proved to be a severe threat to mankind in recent times, and this fortifies an urgency to design and develop potent antibacterial small molecules/compounds with nonconventional mechanisms than the conventional ones. DNA carries the genetic signature of any organism, and bacteria maintain their genomic DNA inside the cell in a well-regulated compact form with the help of various nucleoid associated proteins like HU, HNS, etc. These proteins control various fundamental processes like gene expression, replication, etc., inside the cell. Alteration of the native DNA structure of bacteria can lead to severe consequences in cellular processes inside the bacterial cell that ultimately result in the death of the organism. The change in the global DNA structure by small molecules initiates a plethora of cellular responses that have not been very well investigated. Echinomycin and Triostin-A are biologically active Quinoxaline small molecules that typically consist of a quinoxaline chromophore attached with an octadepsipeptide ring. They bind to double-stranded DNA in a sequence-specific way and have high activity against a wide variety of bacteria, mainly against Gram-positive ones. To date, few synthetic quinoxaline scaffolds were synthesized, displaying antibacterial potential against a broad scale of pathogenic bacteria. QNOs (Quinoxaline N-oxides) are known to target DNA and instigate reactive oxygen species (ROS) production in bacteria, thereby exhibiting antibacterial properties. The divergent role of Quinoxaline small molecules in medicinal research qualifies them for the evaluation of their antimicrobial properties as a potential candidate. The previous study from our lab has given new insights on a 6-nitroquinoxaline derivative 1d as an intercalator of DNA, which induces conformational changes in DNA upon binding.7 The binding event observed was dependent on the presence of a crucial benzyl substituent on the quinoxaline moiety. This was associated with a large induced CD (ICD) appearing in a sigmoidal pattern upon the interaction of 1d with dsDNA. The induction of DNA superstructures by 1d at high Drug:DNA ratios was observed that ultimately led to DNA condensation. Eviction of invitro-assembled nucleosome upon treatment with a high dose of 1d was also observed. In this work, monoquinoxaline derivatives of 1d were synthesized by various modifications of the 1d scaffold. The set of synthesized 6-nitroquinoxaline derivatives along with 1d were all subjected to antibacterial evaluation across five different bacteria species. Among the compound set, 3a displayed potent antibacterial activity against Staphylococcus aureus bacteria. 3a was further subjected to various biophysical studies to check whether the DNA structural alteration potential was still intact. The biological response of S. aureus cells upon treatment with 3a was studied using various cell biology processes, which led to the conclusion that 3d can initiate DNA damage in the S. aureus cells. Finally, the potential of 3a in disrupting preformed S.aureus and S.epidermidis biofilms was also studied.

Keywords: DNA structural change, antibacterial, intercalator, DNA superstructures, biofilms

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Authors: Supriya Ambawat, Subaran Singh, C. Tara Satyavathi, B. S. Rajpurohit, Ummed Singh, Balraj Singh

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Pearl millet [Pennisetum glaucum (L.) R. Br.] is an important staple food of the arid and semiarid tropical regions of Asia, Africa, and Latin America. It is rightly termed as nutricereal as it has high nutrition value and a good source of carbohydrate, protein, fat, ash, dietary fiber, potassium, magnesium, iron, zinc, etc. Pearl millet has low prolamine fraction and is gluten free which is useful for people having a gluten allergy. It has several health benefits like reduction in blood pressure, thyroid, diabe¬tes, cardiovascular and celiac diseases but its direct consumption as food has significantly declined due to several reasons. Keeping this in view, it is important to reorient the ef¬forts to generate demand through value-addition and quality improvement and create awareness on the nutritional merits of pearl millet. In India, through Indian Council of Agricultural Research-All India Coordinated Research Project on Pearl millet, multilocational coordinated trials for developed hybrids were conducted at various centers. The gene banks of pearl millet contain varieties with high levels of iron and zinc which were used to produce new pearl millet varieties with elevated iron levels bred with the high‐yielding varieties. Thus, using breeding approaches and biochemical analysis, a total of 167 hybrids and 61 varieties were identified and released for cultivation in different agro-ecological zones of the country which also includes some biofortified hybrids rich in Fe and Zn. Further, using several biotechnological interventions such as molecular markers, next-generation sequencing (NGS), association mapping, nested association mapping (NAM), MAGIC populations, genome editing, genotyping by sequencing (GBS), genome wide association studies (GWAS) advancement in millet improvement has become possible by identifying and tagging of genes underlying a trait in the genome. Using DArT markers very high density linkage maps were constructed for pearl millet. Improved HHB67 has been released using marker assisted selection (MAS) strategies, and genomic tools were used to identify Fe-Zn Quantitative Trait Loci (QTL). The draft genome sequence of millet has also opened various ways to explore pearl millet. Further, genomic positions of significantly associated simple sequence repeat (SSR) markers with iron and zinc content in the consensus map is being identified and research is in progress towards mapping QTLs for flour rancidity. The sequence information is being used to explore genes and enzymatic pathways responsible for rancidity of flour. Thus, development and application of several biotechnological approaches along with biofortification can accelerate the genetic gain targets for pearl millet improvement and help improve its quality.

Keywords: Biotechnological approaches, genomic tools, malnutrition, MAS, nutricereal, pearl millet, sequencing.

Procedia PDF Downloads 185

Authors: Rongrong Liu, Li Wang, Hui Xu, Jianpei Fang, Sixi Liu, Xiaolin Yin, Junbin Liang, Gaohui Yan, Yaoyun Li, Yali Zhou, Xinyu Li, Yue Li, Lei Shi, Yongrong Lai, Junjiu Huang, Xinhua Zhang

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Background: Beta-Thalassemia is caused by reduced (β+) or absent (β0) synthesis of the β-globin chains of hemoglobin. Transfusions and oral iron chelation therapy have improved the quality of life for patients with Transfusion-Dependent thalassemia (TDT). Recent advances in genome editing platforms of CRISPR-Cas9 have paved the way for induction of HbF by reactivating expression of γ-chain.Aims: We performed CRISPR-Cas9-mediated genome editing of hematopoietic stem cells to mutate HBG1/HBG2 promoter sequence, thereby representing a naturally occurring HPFH-liked mutation, producing RM-001. Here, we present an initial assessment of safety and efficacy of RM-001 in patients with TDT. Methods: Patients (6–35 y of age) with TDT receiving packed red blood cell (pRBC) transfusions of ≥100 mL/kg/y or ≥10 units/y in the previous 2 y were eligible. CD34+ cells were edited with CRISPR-Cas9 using a guide RNA specific for the binding site of BCL11A on the HBG1/2 promoter. Prior to RM-001 product infusion (day 0), patients received myeloablative conditioning with Busulfan from day-7 to day-4. Patients were monitored for AEs Hb expression.Results: Data cut as of 28 Feb 2024, 16 TDT patients have been treated with RM-001 and followed ≥3 months. 5 of these 16 patients had finished their 24 months follow up. Eleven patients have β0/β0 genotype and five patients have β0/β+ genotype. In addition to β-thalassemia, two patients had α- deletion with the genotype of --/αα. Efficacy:All patients received a single dose intravenous infusion of RM-001 cells. 5 of them had been followed 24 months or longer. All patients achieved transfusion-independent (TI, total Hb continued ≥ 9g/dL) (Figure1). Patients demonstrated sustained and clinically meaningful increases in HbF levels since 4 month post-RM-001 infusion (Figure.2). Total hemoglobin in all patients was stable at 10-12g/dL during the follow-up period. Safety:The adverse events observed after RM-001 infusion were consistent with those that are typical of Busulfan-based myeloablation. The allelic editing analysis at 6-month visit showed that the on-target allelic editing frequency in bone marrow cells was 73.44% (64.65% to 84.6%, n=13).Summary/Conclusion: This interim analysis, in which all the 19 patients age from 7.9 to 25yo met the success criteria for the trial with respect to transfusion independence, showed that autologous HBG1/2 promoter-modified CD34+ HSPCs gene therapy resulted in an adequate amount of HbF as early as 2 months after infusion led to near-normal hemoglobin levels, remained transfusion-free through the reported period without product related SAE. After RM-001 infusion, high levels of HbF proportion and on-target editing in bone marrow cells were maintained. Submitted on behalf of the RM-001 Investigators.

Keywords: thalassemian, genetherapy, CRISPR/Cas9, HbF

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Authors: Parashram Jakappa Patil

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India is the global leader in world cashew business and cashew-nut industry is one of the important food processing industries in world. However India is the largest producer, processor, exporter and importer eschew in the world. India is providing cashew to the rest of the world. India is meeting world demand of cashew. India has a tremendous potential of cashew production and export to other countries. Every year India earns more than 2000 cores rupees through cashew trade. Cashew industry is one of the important small scale industries in the country which is playing significant role in rural development. It is generating more than 400000 jobs at remote area and 95% cashew worker are women, it is giving income to poor cashew farmers, majority cashew processing units are small and cottage, it is helping to stop migration from young farmers for employment opportunities, it is motivation rural entrepreneurship development and it is also helping to environment protection etc. Hence India cashew business is very important agribusiness in India which has potential make inclusive development. World Bank and IMF recognized cashew-nut industry is one the important tool for poverty eradication at global level. It shows important of cashew business and its strong existence in India. In spite of such huge potential cashew processing industry is facing different problems such as lack of infrastructure ability, lack of supply of raw cashew, lack of availability of finance, collection of raw cashew, unavailability of warehouse, marketing of cashew kernels, lack of technical knowledge and especially processing technology and packaging of finished products. This industry has great prospects such as scope for more cashew cultivation and cashew production, employment generation, formation of cashew processing units, alcohols production from cashew apple, shield oil production, rural development, poverty elimination, development of social and economic backward class and environment protection etc. This industry has domestic as well as foreign market; India has tremendous potential in this regard. The cashew is a poor men’s crop but rich men’s food. The cashew is a source of income and livelihood for poor farmers. Cashew-nut industry may play very important role in the development of hilly region. The objectives of this paper are to identify problems of cashew processing and use of processing technology, problems of cashew kernel packaging, evolving of cashew processing technology over the year and its impact on final product and impact of good processing by adopting appropriate technology packaging on international trade of cashew-nut. The most important problem of cashew processing industry is that is processing and packaging. Bad processing reduce the quality of cashew kernel at large extent especially broken of cashew kernel which has very less price in market compare to whole cashew kernel and not eligible for export. On the other hand if there is no good packaging of cashew kernel will get moisture which destroy test of it. International trade of cashew-nut is depend of two things one is cashew processing and other is packaging. This study has strong relevance because cashew-nut industry is the labour oriented, where processing technology is not playing important role because 95% processing work is manual. Hence processing work was depending on physical performance of worker which makes presence of large workforce inevitable. There are many cashew processing units closed because they are not getting sufficient work force. However due to advancement in technology slowly this picture is changing and processing work get improve. Therefore it is interesting to explore all the aspects in context of cashew processing and packaging of cashew business.

Keywords: cashew, processing technology, packaging, international trade, change

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Authors: Shaista Suhail

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As cancer populations is increasing sharply, the incidence of oral squamous cell carcinoma (OSCC) has also been expected to increase. Oral carcinogenesis is a highly complex, multistep process which involves accumulation of genetic alterations that lead to the induction of proteins promoting cell growth (encoded by oncogenes), increased enzymatic (telomerase) activity promoting cancer cell proliferation. The global increase in frequency and mortality, as well as the poor prognosis of oral squamous cell carcinoma, has intensified current research efforts in the field of prevention and early detection of this disease. The advances in the understanding of the molecular basis of oral cancer should help in the identification of new markers. The study of the carcinogenic process of the oral cancer, including continued analysis of new genetic alterations, along with their temporal sequencing during initiation, promotion and progression, will allow us to identify new diagnostic and prognostic factors, which will provide a promising basis for the application of more rational and efficient treatments. Telomerase activity has been readily found in most cancer biopsies, in premalignant lesions or germ cells. Activity of telomerase is generally absent in normal tissues. It is known to be induced upon immortalization or malignant transformation of human cells such as in oral cancer cells. Maintenance of telomeres plays an essential role during transformation of precancer to malignant stage. Mammalian telomeres, a specialized nucleoprotein structures are composed of large conctamers of the guanine-rich sequence 5_-TTAGGG-3_. The roles of telomeres in regulating both stability of genome and replicative immortality seem to contribute in essential ways in cancer initiation and progression. It is concluded that activity of telomerase can be used as a biomarker for diagnosis of malignant oral cancer and a target for inactivation in chemotherapy or gene therapy. Its expression will also prove to be an important diagnostic tool as well as a novel target for cancer therapy. The activation of telomerase may be an important step in tumorgenesis which can be controlled by inactivating its activity during chemotherapy. The expression and activity of telomerase are indispensable for cancer development. There are no drugs which can effect extremely to treat oral cancers. There is a general call for new emerging drugs or methods that are highly effective towards cancer treatment, possess low toxicity, and have a minor environment impact. Some novel natural products also offer opportunities for innovation in drug discovery. Natural compounds isolated from medicinal plants, as rich sources of novel anticancer drugs, have been of increasing interest with some enzyme (telomerase) blockage property. The alarming reports of cancer cases increase the awareness amongst the clinicians and researchers pertaining to investigate newer drug with low toxicity.

Keywords: oral carcinoma, telomere, telomerase, blockage

Procedia PDF Downloads 175

Authors: Nourhan Hussein Fanaki, Hoda Mohamed Gamal El-Din Omar, Nihal Kadry Moussa, Eva Adel Edward Farid

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The resistance of staphylococci to various antibiotics has become a major concern for health care professionals. The efficacy of the combinations of selected glycopeptides (vancomycin and teicoplanin) with gentamicin or rifampicin, as well as that of gentamicin/rifampicin combination, was studied against selected pathogenic staphylococcus isolated from Egypt. The molecular distribution of genes conferring resistance to these four antibiotics was detected among tested clinical isolates. Antibiotic combinations were studied using the checkerboard technique and the time-kill assay (in both the stationary and log phases). Induction of resistance to glycopeptides in staphylococci was tried in the absence and presence of diclofenac sodium as inducer. Transmission electron microscopy was used to study the effect of glycopeptides on the ultrastructure of the cell wall of staphylococci. Attempts were made to cure gentamicin resistance plasmids and to study the transfer of these plasmids by conjugation. Trials for the transformation of the successfully isolated gentamicin resistance plasmid to competent cells were carried out. The detection of genes conferring resistance to the tested antibiotics was performed using the polymerase chain reaction. The studied antibiotic combinations proved their efficacy, especially when tested during the log phase. Induction of resistance to glycopeptides in staphylococci was more promising in presence of diclofenac sodium, compared to its absence. Transmission electron microscopy revealed the thickening of bacterial cell wall in staphylococcus clinical isolates due to the presence of tested glycopeptides. Curing of gentamicin resistance plasmids was only successful in 2 out of 9 tested isolates, with a curing rate of 1 percent for each. Both isolates, when used as donors in conjugation experiments, yielded promising conjugation frequencies ranging between 5.4 X 10-2 and 7.48 X 10-2 colony forming unit/donor cells. Plasmid isolation was only successful in one out of the two tested isolates. However, low transformation efficiency (59.7 transformants/microgram plasmid DNA) of such plasmids was obtained. Negative regulators of autolysis, such as arlR, lytR and lrgB, as well as cell-wall associated genes, such as pbp4 and/or pbp2, were detected in staphylococcus isolates with reduced susceptibility to the tested glycopeptides. Concerning rifampicin resistance genes, rpoBstaph was detected in 75 percent of the tested staphylococcus isolates. It could be concluded that in vitro studies emphasized the usefulness of the combination of vancomycin or teicoplanin with gentamicin or rifampicin, as well as that of gentamicin with rifampicin, against staphylococci showing varying resistance patterns. However, further in vivo studies are required to ensure the safety and efficacy of such combinations. Diclofenac sodium can act as an inducer of resistance to glycopeptides in staphylococci. Cell-wall thickness is a major contributor to such resistance among them. Gentamicin resistance in these strains could be chromosomally or plasmid mediated. Multiple mutations in the rpoB gene could mediate staphylococcus resistance to rifampicin.

Keywords: glycopeptides, combinations, induction, diclofenac, transmission electron microscopy, polymerase chain reaction

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Authors: Razieh Karimi Aghcheh, Christian Kubicek, Joseph Strauss, Gerhard Braus

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Filamentous fungi are of considerable economic and social significance for human health, nutrition and in white biotechnology. These organisms are dominant producers of a range of primary metabolites such as citric acid, microbial lipids (biodiesel) and higher unsaturated fatty acids (HUFAs). In particular, they produce also important but structurally complex secondary metabolites with enormous therapeutic applications in pharmaceutical industry, for example: cephalosporin, penicillin, taxol, zeranol and ergot alkaloids. Several fungal secondary metabolites, which are significantly relevant to human health do not only include antibiotics, but also e.g. lovastatin, a well-known antihypercholesterolemic agent produced by Aspergillus. terreus, or aflatoxin, a carcinogen produced by A. flavus. In addition to their roles for human health and agriculture, some fungi are industrially and commercially important: Species of the ascomycete genus Hypocrea spp. (teleomorph of Trichoderma) have been demonstrated as efficient producer of highly active cellulolytic enzymes. This trait makes them effective in disrupting and depolymerization of lignocellulosic materials and thus applicable tools in number of biotechnological areas as diverse as clothes-washing detergent, animal feed, and pulp and fuel productions. Fungal LaeA/LAE1 (Loss of aflR Expression A) homologs their gene products act at the interphase between secondary metabolisms, cellulase production and development. Lack of the corresponding genes results in significant physiological changes including loss of secondary metabolite and lignocellulose degrading enzymes production. At the molecular level, the encoded proteins are presumably methyltransferases or demethylases which act directly or indirectly at heterochromatin and interact with velvet domain proteins. Velvet proteins bind to DNA and affect expression of secondary metabolites (SMs) genes and cellulases. The dynamic interplay between LaeA/LAE1, velvet proteins and additional interaction partners is the key for an understanding of the coordination of metabolic and morphological functions of fungi and is required for a biotechnological control of the formation of desired bioactive products. Aspergilli and Trichoderma represent different biotechnologically significant species with significant differences in the LaeA/LAE1-Velvet protein machinery and their target proteins. We, therefore, performed a comparative study of the interaction partners of this machinery and the dynamics of the various protein-protein interactions using our robust proteomic and mass spectrometry techniques. This enhances our knowledge about the fungal coordination of secondary metabolism, cellulase production and development and thereby will certainly improve recombinant fungal strain construction for the production of industrial secondary metabolite or lignocellulose hydrolytic enzymes.

Keywords: cellulases, LaeA/1, proteomics, secondary metabolites

Procedia PDF Downloads 270

Authors: N. Manjunatha, K. T. Rangswamy, N. Nagaraju, H. A. Prameela, P. Rudraswamy, M. Krishnareddy

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The identification of virus in cowpea especially potyviruses is confusing. Even though there are several studies on viruses causing diseases in cowpea, difficult to distinguish based on symptoms and serological detection. The differentiation of potyviruses considering as a constraint, the present study is initiated for molecular characterization, host plant resistance and epidemiology of the BCMV infecting cowpea. The etiological agent causing cowpea mosaic was identified as Bean Common Mosaic Virus (BCMV) on the basis of RT-PCR and electron microscopy. An approximately 750bp PCR product corresponding to coat protein (CP) region of the virus and the presence of long flexuous filamentous particles measuring about 952 nm in size typical to genus potyvirus were observed under electron microscope. The characterized virus isolate genome had 10054 nucleotides, excluding the 3’ terminal poly (A) tail. Comparison of polyprotein of the virus with other potyviruses showed similar genome organization with 9 cleavage sites resulted in 10 functional proteins. The pairwise sequence comparison of individual genes, P1 showed most divergent, but CP gene was less divergent at nucleotide and amino acid level. A phylogenetic tree constructed based on multiple sequence alignments of the polyprotein nucleotide and amino acid sequences of cowpea BCMV and potyviruses showed virus is closely related to BCMV-HB. Whereas, Soybean variant of china (KJ807806) and NL1 isolate (AY112735) showed 93.8 % (5’UTR) and 94.9 % (3’UTR) homology respectively with other BCMV isolates. This virus transmitted to different leguminous plant species and produced systemic symptoms under greenhouse conditions. Out of 100 cowpea genotypes screened, three genotypes viz., IC 8966, V 5 and IC 202806 showed immune reaction in both field and greenhouse conditions. Single marker analysis (SMA) was revealed out of 4 SSR markers linked to BCMV resistance, M135 marker explains 28.2 % of phenotypic variation (R2) and Polymorphic information content (PIC) value of these markers was ranged from 0.23 to 0.37. The correlation and regression analysis showed rainfall, and minimum temperature had significant negative impact and strong relationship with aphid population, whereas weak correlation was observed with disease incidence. Path coefficient analysis revealed most of the weather parameters exerted their indirect contributions to the aphid population and disease incidence except minimum temperature. This study helps to identify specific gaps in knowledge for researchers who may wish to further analyse the science behind complex interactions between vector-virus and host in relation to the environment. The resistant genotypes identified are could be effectively used in resistance breeding programme.

Keywords: cowpea, epidemiology, genotypes, virus

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Authors: Akanksha Bhatnagar, Sandhya Kortegare, Felice Elefant

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Context: Alzheimer's disease (AD) is a neurodegenerative disorder that is characterized by progressive cognitive decline and memory loss. The cause of AD is not fully understood, but it is thought to be caused by a combination of genetic, environmental, and lifestyle factors. One of the hallmarks of AD is the loss of neurons in the hippocampus, a brain region that is important for memory and learning. This loss of neurons is thought to be caused by a decrease in histone acetylation, which is a process that regulates gene expression. Research Aim: The research aim of the study was to develop mall molecule compounds that can enhance the activity of Tip60, a histone acetyltransferase that is important for memory and learning. Methodology/Analysis: The researchers used in silico structural modeling and a pharmacophore-based virtual screening approach to design and synthesize small molecule compounds strongly predicted to target and enhance Tip60’s HAT activity. The compounds were then tested in vitro and in vivo to assess their ability to enhance Tip60 activity and rescue cognitive deficits in AD models. Findings: The researchers found that several of the compounds were able to enhance Tip60 activity and rescue cognitive deficits in AD models. The compounds were also developed to cross the blood-brain barrier, which is an important factor for the development of potential AD therapeutics. Theoretical Importance: The findings of this study suggest that Tip60 HAT activators have the potential to be developed as therapeutic agents for AD. The compounds are specific to Tip60, which suggests that they may have fewer side effects than other HDAC inhibitors. Additionally, the compounds are able to cross the blood-brain barrier, which is a major hurdle for the development of AD therapeutics. Data Collection: The study collected data from a variety of sources, including in vitro assays and animal models. The in vitro assays assessed the ability of compounds to enhance Tip60 activity using histone acetyltransferase (HAT) enzyme assays and chromatin immunoprecipitation assays. Animal models were used to assess the ability of the compounds to rescue cognitive deficits in AD models using a variety of behavioral tests, including locomotor ability, sensory learning, and recognition tasks. The human clinical trials will be used to assess the safety and efficacy of the compounds in humans. Questions: The question addressed by this study was whether Tip60 HAT activators could be developed as therapeutic agents for AD. Conclusions: The findings of this study suggest that Tip60 HAT activators have the potential to be developed as therapeutic agents for AD. The compounds are specific to Tip60, which suggests that they may have fewer side effects than other HDAC inhibitors. Additionally, the compounds are able to cross the blood-brain barrier, which is a major hurdle for the development of AD therapeutics. Further research is needed to confirm the safety and efficacy of these compounds in humans.

Keywords: Alzheimer's disease, cognition, neuroepigenetics, drug discovery

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Authors: Jong H. Kim, Kathleen L. Chan, Luisa W. Cheng

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Control of fungal pathogens, such as foodborne mycotoxin producers, is problematic as effective antimycotic agents are often very limited. Mycotoxin contamination significantly interferes with the safe production of foods or crops worldwide. Moreover, expansion of fungal resistance to commercial drugs or fungicides is a global human health concern. Therefore, there is a persistent need to enhance the efficacy of commercial antimycotic agents or to develop new intervention strategies. Disruption of the cellular antioxidant system should be an effective method for pathogen control. Such disruption can be achieved with safe, redox-active compounds. Natural phenolic derivatives are potent redox cyclers that inhibit fungal growth through destabilization of the cellular antioxidant system. The goal of this study is to identify novel, redox-active compounds that disrupt the fungal antioxidant system. The identified compounds could also function as sensitizing agents to conventional antimycotics (i.e., chemosensitization) to improve antifungal efficacy. Various benzo derivatives were tested against fungal pathogens. Gene deletion mutants of the yeast Saccharomyces cerevisiae were used as model systems for identifying molecular targets of benzo analogs. The efficacy of identified compounds as potent antifungal agents or as chemosensitizing agents to commercial drugs or fungicides was examined with methods outlined by the Clinical Laboratory Standards Institute or the European Committee on Antimicrobial Susceptibility Testing. Selected benzo derivatives possessed potent antifungal or antimycotoxigenic activity. Molecular analyses by using S. cerevisiae mutants indicated antifungal activity of benzo derivatives was through disruption of cellular antioxidant or cell wall integrity system. Certain benzo analogs screened overcame tolerance of Aspergillus signaling mutants, namely mitogen-activated protein kinase mutants, to fludioxonil fungicide. Synergistic antifungal chemosensitization greatly lowered minimum inhibitory or fungicidal concentrations of test compounds, including inhibitors of mitochondrial respiration. Of note, salicylaldehyde is a potent antimycotic volatile that has some practical application as a fumigant. Altogether, benzo derivatives targeting cellular antioxidant system of fungi (along with cell wall integrity system) effectively suppress fungal growth. Candidate compounds possess the antifungal, antimycotoxigenic or chemosensitizing capacity to augment the efficacy of commercial antifungals. Therefore, chemogenetic approaches can lead to the development of novel antifungal intervention strategies, which enhance the efficacy of established microbe intervention practices and overcome drug/fungicide resistance. Chemosensitization further reduces costs and alleviates negative side effects associated with current antifungal treatments.

Keywords: antifungals, antioxidant system, benzo derivatives, chemosensitization

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Authors: Faris Mohsin Alabeedi

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Pemphigus Vulgaris (PV) is a life-threatening autoimmune blistering disease characterized by the presence of autoantibodies directed against the epidermis's surface proteins. There are two forms of PV, mucocutaneous and mucosal-dominant PV. Disruption of the cell junctions is a hallmark of PV due to the autoantibodies targeting the desmosomal cadherins, desmoglein-3 (Dsg3) and desmoglein-1, leading to acantholysis in the skin and mucous membrane. Although the pathogenesis of PV is known, the detailed molecular events remain not fully understood. Our recent study has shown that both the PV sera and pathogenic anti-Dsg3 antibody AK23 can induce ROS and cause oxidative stress in cultured keratinocytes. In line with our finding, other independent studies also demonstrate oxidative stress in PV. Since Nrf2 plays a crucial role in cellular anti-oxidative stress response, we hypothesize that the expression of Nrf2 may alter in PV. Thus, treatment of cells with PV sera or AK23 may cause changes in Nrf2 expression and distribution. The purpose of this study was to examine the effect of AK23 and PV sera on Nrf2 in a normal human keratinocyte cell line, such as NTERT cells. Both a time-course and dose-dependent experiments with AK23, alongside the matched isotype control IgG, were performed in keratinocyte cultures and analysed by immunofluorescence for Nrf2 and Dsg3. Additionally, the same approach was conducted with the sera from PV patients and healthy individuals that served as a control in this study. All the fluorescent images were analysed using ImageJ software. Each experiment was repeated twice. In general, variations were observed throughout this study. In the dose-response experiments, although enhanced Dsg3 expression was consistently detected in AK23 treated cells, the expression of Nrf2 showed no consistent findings between the experiments, although changes in its expression were noticeable in cells treated with AK23. In the time-course study, a trend with induction of Nrf2 over time was shown in control cells treated with mouse isotype IgG. Treatment with AK23 showed a reduction of Nrf2 in a time-dependent manner, especially at the 24-hour time point. However, the earlier time points, such as 2 hours and 6 hours with AK23 treatments, detected somewhat variations. Finally, PV sera caused a decrease of Dsg3, but on the other hand, variations were observed in Nrf2 expression in PV sera treated cells. In general, PV sera seemed to cause a reduction of Nrf2 in the majority of PV sera treated samples. In addition, more pronounced cytoplasmic expression of Nrf2 has been observed in PV sera treated cells than those treated with AK23, suggesting that polyclonal and monoclonal IgG might induce a different effect on Nrf2 expression and distribution. Further experimental studies are crucial to obtain a more coincide global view of Nrf2-mediated gene regulation. In particular, Pemphigus Voulgaris studies assessing how the Nrf2-dependent network changes from a physiological to a pathological condition can provide insight into disease mechanisms and perhaps initiate further treatment approaches.

Keywords: pemphigus vulgaris, monoclonal antibody against desmoglein-3, Nrf2 oxidative stress, keratinocyte cultures

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Authors: Varinder Kaur Gambhir, Neema Gupta, Sonal Tickoo Chaudhuri

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Bengaluru, a rapidly growing metropolis in India, with a population of 12.5 million citizens, generates 5,757 metric tonnes of solid waste per day. Despite their invaluable contribution to waste management, society and the economy, waste pickers face significant stigma, suspicion and contempt and are left with a sense of shame about their work. In this context, BBC Media Action was funded by the H&M Foundation to develop a 3-year multi-phase social media campaign to shift perceptions of waste picking and informal waste pickers amongst the Bengaluru population. Research has been used to inform project strategy and adaptation, at all stages. Formative research to inform campaign strategy used mixed methods– 14 focused group discussions followed by 406 online surveys – to explore people’s knowledge of, and attitudes towards waste pickers, and identify potential barriers and motivators to changing perceptions. Use of qualitative techniques like metaphor maps (using bank of pictures rather than direct questions to understand mindsets) helped establish the invisibility of informal waste pickers, and the quantitative research enabled audience segmentation based on attitudes towards informal waste pickers. To pretest the campaign idea, eight I-GDs (individual interaction followed by group discussions) were conducted to allow interviewees to first freely express their feelings individually, before discussing in a group. Robert Plucthik’s ‘wheel of emotions’ was used to understand audience’s emotional response to the content. A robust monitoring and evaluation is being conducted (baseline and first phase of monitoring already completed) using a rotating longitudinal panel of 1,800 social media users (exposed and unexposed to the campaign), recruited face to face and representative of the social media universe of Bengaluru city. In addition, qualitative in-depth interviews are being conducted after each phase to better understand change drivers. The research methodology and ethical protocols for impact evaluation have been independently reviewed by an Institutional Review Board. Formative research revealed that while waste on the streets is visible and is of concern to the public, informal waste pickers are virtually ‘invisible’, for most people in Bengaluru Pretesting research revealed that the creative outputs evoked emotions like acceptance and gratitude towards waste-pickers, suggesting that the content had the potential to encourage attitudinal change. After the first phase of campaign, social media analytics show that #Invaluables content reached at least 2.6 million unique people (21% of the Bengaluru population) through Facebook and Instagram. Further, impact monitoring results show significant improvements in spontaneous awareness of different segments of informal waste pickers ( such as sorters at scrap shops or dry waste collection centres -from 10% at baseline to 16% amongst exposed and no change amongst unexposed), recognition that informal waste pickers help the environment (71% at baseline to 77% among exposed and no change among unexposed) and greater discussion about informal waste pickers among those exposed (60%) as against not exposed (49%). Using the insights from this research, the planned social media intervention is designed to increase the visibility of and appreciation for the work of waste pickers in Bengaluru, supporting a more inclusive society.

Keywords: awareness, discussion, discrimination, informal waste pickers, invisibility, social media campaign, waste management

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Authors: M. Reza Hashemi, Chris Small, Scott Hayward

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The Northeast Coast of the US faces damaging effects of coastal flooding and winds due to Atlantic tropical and extratropical storms each year. Historically, several large storm events have produced substantial levels of damage to the region; most notably of which were the Great Atlantic Hurricane of 1938, Hurricane Carol, Hurricane Bob, and recently Hurricane Sandy (2012). The objective of this study was to develop an integrated modeling system that could be used as a forecasting/hindcasting tool to evaluate and communicate the risk coastal communities face from these coastal storms. This modeling system utilizes the ADvanced CIRCulation (ADCIRC) model for storm surge predictions and the Simulating Waves Nearshore (SWAN) model for the wave environment. These models were coupled, passing information to each other and computing over the same unstructured domain, allowing for the most accurate representation of the physical storm processes. The coupled SWAN-ADCIRC model was validated and has been set up to perform real-time forecast simulations (as well as hindcast). Modeled storm parameters were then passed to a coastal risk assessment tool. This tool, which is generic and universally applicable, generates spatial structural damage estimate maps on an individual structure basis for an area of interest. The required inputs for the coastal risk model included a detailed information about the individual structures, inundation levels, and wave heights for the selected region. Additionally, calculation of wind damage to structures was incorporated. The integrated coastal risk assessment system was then tested and applied to Charlestown, a small vulnerable coastal town along the southern shore of Rhode Island. The modeling system was applied to Hurricane Sandy and a synthetic storm. In both storm cases, effect of natural dunes on coastal risk was investigated. The resulting damage maps for the area (Charlestown) clearly showed that the dune eroded scenarios affected more structures, and increased the estimated damage. The system was also tested in forecast mode for a large Nor’Easters: Stella (March 2017). The results showed a good performance of the coupled model in forecast mode when compared to observations. Finally, a nearshore model XBeach was then nested within this regional grid (ADCIRC-SWAN) to simulate nearshore sediment transport processes and coastal erosion. Hurricane Irene (2011) was used to validate XBeach, on the basis of a unique beach profile dataset at the region. XBeach showed a relatively good performance, being able to estimate eroded volumes along the beach transects with a mean error of 16%. The validated model was then used to analyze the effectiveness of several erosion mitigation methods that were recommended in a recent study of coastal erosion in New England: beach nourishment, coastal bank (engineered core), and submerged breakwater as well as artificial surfing reef. It was shown that beach nourishment and coastal banks perform better to mitigate shoreline retreat and coastal erosion.

Keywords: ADCIRC, coastal flooding, storm surge, coastal risk assessment, living shorelines

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Authors: Huseyin Apdik, Aysegul Dogan, Selami Demirci, Ezgi Avsar Apdik, Fikrettin Sahin

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Mesenchymal stem cells (MSCs) are crucial cell types for bone maintenance and growth along with resident bone progenitor cells providing bone tissue integrity during osteogenesis and skeletal growth. Any deficiency in this regulation would result in vital bone diseases. Of those, osteoporosis, characterized by a reduction in bone mass and mineral density, is a critical skeletal disease for especially elderly people. The commonly used drugs for the osteoporosis treatment are bisphosphonates (BPs). The most prominent role of BPs is to prevent bone resorption arisen from high osteoclast activity. However, administrations of bisphosphonates may also cause bisphosphonate-induced osteonecrosis of the jaw (BIONJ). Up to the present, the researchers have proposed several circumstances for BIONJ. However, effects of long-term and/or high dose usage of BPs on stem cell’s proliferation, survival, differentiation or maintenance capacity have not been evaluated yet. The present study will be held to; figure out BPs’ effects on MSCs in vitro in the aspect of cell proliferation and toxicity, migration, angiogenic activity, lineage specific gene and protein expression levels, mesenchymal stem cell properties and potential signaling pathways affected by BP treatment. Firstly, mesenchymal stem cell characteristics of Dental Pulp Stem Cells (DPSCs) and Periodontal Ligament Stem Cells (PDLSCs) were proved using flow cytometry analysis. Cell viability analysis was completed to determine the cytotoxic effects of BPs (Zoledronate (Zol), Alendronate (Ale) and Risedronate (Ris)) on DPSCs and PDLSCs by the 3-(4,5-di-methyl-thiazol-2-yl)-5-(3-carboxymethoxy-phenyl)-2-(4-sulfo-phenyl)-2H-tetrazolium (MTS) assay. Non-toxic concentrations of BPs were determined at 24 h under growth condition, and at 21 days under osteogenic differentiation condition for both cells. The scratch assay was performed to evaluate their migration capacity under the usage of determined of BPs concentrations at 24 h. The results revealed that while the scratch closure is 70% in the control group for DPSCs, it was 57%, 66% and 66% in Zol, Ale and Ris groups, respectively. For PDLSs, while wound closure is 71% in control group, it was 65%, 66% and 66% in Zol, Ale and Ris groups, respectively. As future experiments, tube formation assay and aortic ring assay will be done to determinate angiogenesis abilities of DPSCs and PDLSCs treated with BPs. Expression levels of osteogenic differentiation marker genes involved in bone development will be determined using real time-polymerase change reaction (RT-PCR) assay and expression profiles of important proteins involved in osteogenesis will be evaluated using western blotting assay for osteogenically differentiated MSCs treated with or without BPs. In addition to these, von Kossa staining will be performed to measure calcium mineralization status of MSCs.

Keywords: bisphosphonates, bisphosphonate-induced osteonecrosis of the jaw, mesenchymal stem cells, osteogenesis

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Authors: Katarzyna Drela, Miroslaw Wielgos, Mikolaj Wrobel, Barbara Lukomska

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Mesenchymal stem cells (MSCs) have a great potential in regenerative medicine. Since the initial number of isolated MSCs is limited, in vitro propagation is often required to reach sufficient numbers of cells for therapeutic applications. During long-term culture MSCs may undergo genetic or epigenetic alterations that subsequently increase the probability of spontaneous malignant transformation. Thus, factors that influence genomic stability of MSCs following long-term expansions need to be clarified before cultured MSCs are employed for clinical application. The aim of our study was to investigate the potential for spontaneous transformation of human neonatal cord blood (HUCB-MSCs) and adult bone marrow (BM-MSCs) derived MSCs. Materials and Methods: HUCB-MSCs and BM-MSCs were isolated by standard Ficoll gradient centrifugations method. Isolated cells were initially plated in high density 106 cells per cm2. After 48 h medium were changed and non-adherent cells were removed. The malignant transformation of MSCs in vitro was evaluated by morphological changes, proliferation rate, ability to enter cell senescence, the telomerase expression and chromosomal abnormality. Proliferation of MSCs was analyzed with WST-1 reduction method and population doubling time (PDT) was calculated at different culture stages. Then the expression pattern of genes characteristic for mesenchymal or epithelial cells, as well as transcriptions factors were examined by RT-PCR. Concomitantly, immunocytochemical analysis of gene-related proteins was employed. Results: Our studies showed that MSCs from all bone marrow isolations ultimately entered senescence and did not undergo spontaneous malignant transformation. However, HUCB-MSCs from one of the 15 donors displayed an increased proliferation rate, failed to enter senescence, and exhibited an altered cell morphology. In this sample we observed two different cell phenotypes: one mesenchymal-like exhibited spindle shaped morphology and express specific mesenchymal surface markers (CD73, CD90, CD105, CD166) with low proliferation rate, and the second one with round, densely package epithelial-like cells with significantly increased proliferation rate. The PDT of epithelial-like populations was around 1day and 100% of cells were positive for proliferation marker Ki-67. Moreover, HUCB-MSCs showed a positive expression of human telomerase reverse transcriptase (hTERT), cMYC and exhibit increased number of CFU during the long-term culture in vitro. Furthermore, karyotype analysis revealed chromosomal abnormalities including duplications. Conclusions: Our studies demonstrate that HUCB-MSCs are susceptible to spontaneous malignant transformation during long-term culture. Spontaneous malignant transformation process following in vitro culture has enormous effect on the biosafety issues of future cell-based therapies and regenerative medicine regimens.

Keywords: mesenchymal stem cells, spontaneous, transformation, long-term culture

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Authors: M. Kazou, A. Gavriil, O. Kalagkatsi, T. Paschos, E. Tsakalidou

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Kopanisti cheese is a Greek soft Protected Designation of Origin (PDO) cheese made of raw cow, sheep or goat milk, or mixtures of them, with similar organoleptic characteristics to that of Roquefort cheese. Traditional manufacturing of Kopanisti cheese is limited in small-scale dairies, without the addition of starter cultures. Instead, an amount of over-mature Kopanisti cheese, called Mana Kopanisti, is used to initiate ripening. Therefore, the selection of proper starter cultures and the understanding of the contribution of various microbial groups to its overall quality is crucial for the production of a high-quality final product with standardized organoleptic and physicochemical characteristics. Taking the above into account, the aim of the present study was the investigation of Kopanisti cheese microbiota and its role in cheese quality. For this purpose, four different types of Kopanisti were produced in triplicates, all with pasteurized cow milk, with the addition of (A) the typical mesophilic species Lactococcus lactis and Lactobacillus paracasei used as starters in the production of soft spread cheeses, (B) strains of Lactobacillus acidipiscis and Lactobacillus rennini previously isolated from Kopanisti and Mana Kopanisti, (C) all the species from (A) and (B) as inoculum, and finally (D) the species from (A) and Mana Kopanisti. Physicochemical and microbiological analysis was performed for milk and cheese samples during ripening. Enumeration was performed for major groups of lactic acid bacteria (LAB), total mesophilic bacteria, yeasts as well as hygiene indicator microorganisms. Bacterial isolates from all the different LAB groups, apart from enterococci, alongside yeasts isolates, were initially grouped using repetitive sequence-based polymerase chain reaction (rep-PCR) and then identified at the species level using 16S rRNA gene and internal transcribed spacer (ITS) DNA region sequencing, respectively. Sensory evaluation was also performed for final cheese samples at the end of the ripening period (35 days). Based on the results of the classical microbiological analysis, the average counts of the total mesophilic bacteria and LAB, apart from enterococci, ranged between 7 and 10 log colony forming unit (CFU) g⁻¹, phychrotrophic bacteria, and yeast extract glucose chloramphenicol (YGC) isolates between 4 and 8 log CFU g⁻¹, while coliforms and enterococci up to 2 log CFU g⁻¹ throughout ripening in cheese samples A, C and D. In contrast, in cheese sample B, the average counts of the total mesophilic bacteria and LAB, apart from enterococci, phychrotrophic bacteria, and YGC isolates ranged between 0 and 10 log CFU g⁻¹ and coliforms and enterococci up to 2 log CFU g⁻¹. Although the microbial counts were not that different among samples, identification of the bacterial and yeasts isolates revealed the complex microbial community structure present in each cheese sample. Differences in the physicochemical characteristics among the cheese samples were also observed, with pH ranging from 4.3 to 5.3 and moisture from 49.6 to 58.0 % in the final cheese products. Interestingly, the sensory evaluation also revealed differences among samples, with cheese sample B ranking first based on the total score. Overall, the combination of these analyses highlighted the impact of different starter cultures on the Kopanisti microbiota as well as on the physicochemical and sensory characteristics of the final product.

Keywords: Kopanisti cheese, microbiota, classical microbiological analysis, physicochemical analysis

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Authors: Lena Payati, Maria Kazou, Effie Tsakalidou

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Table olives are one of the most popular fermented vegetables worldwide, which along with olive oil, have a crucial role in the world economy. They are highly appreciated by the consumers for their characteristic taste and pleasant aromas, while several health and nutritional benefits have been reported as well. Until recently, microbial biogeography, i.e., the study of microbial diversity over time and space, has been mainly associated with wine. However, nowadays, the term 'terroir' has been extended to other crops and food products so as to link the geographical origin and environmental conditions to quality aspects of fermented foods. Taking the above into consideration, the present study focuses on the microbial fingerprinting of the most important olive varieties of Greece with the state-of-the-art amplicon-based metagenomics analysis. Towards this, in 2019, 61 samples from 38 different olive varieties were collected at the final stage of ripening from 13 well spread geographical regions in Greece. For the metagenomics analysis, total DNA was extracted from the olive samples, and the 16S rRNA gene and ITS DNA region were sequenced and analyzed using bioinformatics tools for the identification of bacterial and yeasts/fungal diversity, respectively. Furthermore, principal component analysis (PCA) was also performed for data clustering based on the average microbial composition of all samples from each region of origin. According to the composition, results obtained, when samples were analyzed separately, the majority of both bacteria (such as Pantoea, Enterobacter, Roserbergiella, and Pseudomonas) and yeasts/fungi (such as Aureobasidium, Debaromyces, Candida, and Cladosporium) genera identified were found in all 61 samples. Even though interesting differences were observed at the relative abundance level of the identified genera, the bacterial genus Pantoea and the yeast/fungi genus Aureobasidium were the dominant ones in 35 and 40 samples, respectively. Of note, olive samples collected from the same region had similar fingerprint (genera identified and relative abundance level) regardless of the variety, indicating a potential association between the relative abundance of certain taxa and the geographical region. When samples were grouped by region of origin, distinct bacterial profiles per region were observed, which was also evident from the PCA analysis. This was not the case for the yeast/fungi profiles since 10 out of the 13 regions were grouped together mainly due to the dominance of the genus Aureobasidium. A second cluster was formed for the islands Crete and Rhodes, both of which are located in the Southeast Aegean Sea. These two regions clustered together mainly due to the identification of the genus Toxicocladosporium in relatively high abundances. Finally, the Agrinio region was separated from the others as it showed a completely different microbial fingerprinting. However, due to the limited number of olive samples from some regions, a subsequent PCA analysis with more samples from these regions is expected to yield in a more clear clustering. The present study is part of a bigger project, the first of its kind in Greece, with the ultimate goal to analyze a larger set of olive samples of different varieties and from different regions in Greece in order to have a reliable olives’ microbial biogeography.

Keywords: amplicon-based metagenomics analysis, bacteria, microbial biogeography, olive microbiota, yeasts/fungi

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Authors: Enoch Ocran

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Film Pertinence maintains consistency in storytelling by sustaining the natural flow of action while evoking a particular feeling or emotion from the viewers with selected motion pictures. This study presents a thorough investigation of "Film Pertinence" in videography that examines its influence in Africa and around the world. This research delves into the dynamic realm of visual storytelling through film, with a specific focus on the concept of Film Pertinence (FP). The study’s primary objectives are to conduct a comparative analysis of videography tools and techniques employed in both African and international contexts, examining how they contribute to the achievement of organizational goals and the enhancement of cultural awareness. The research methodology includes a comprehensive literature review, interviews with videographers from diverse backgrounds in Africa and the international arena, and the examination of pertinent case studies. The investigation aims to elucidate the multifaceted nature of videographic practices, with particular attention to equipment choices, visual storytelling techniques, cultural sensitivity, and adaptability. This study explores the impact of cultural differences on videography choices, aiming to promote understanding between African and foreign filmmakers and create more culturally sensitive films. It also explores the role of technology in advancing videography practices, resource allocation, and the influence of globalization on local filmmaking practices. The research also contributes to film studies by analyzing videography's impact on storytelling, guiding filmmakers to create more compelling narratives. The findings can inform film education, tailoring curricula to regional needs and opportunities. The study also encourages cross-cultural collaboration in the film industry by highlighting convergence and divergence in videography practices. At its core, this study seeks to explore the implications of film pertinence as a framework for videographic practice. It scrutinizes how cultural expression, education, and storytelling transcend geographical boundaries on a global scale. By analyzing the interplay between tools, techniques, and context, the research illuminates the ways in which videographers in Africa and worldwide apply film Pertinence principles to achieve cross-cultural communication and effectively capture the objectives of their clients. One notable focus of this paper is on the techniques employed by videographers in West Africa to emphasize storytelling and participant engagement, showcasing the relevance of FP in highlighting cultural awareness in visual storytelling. Additionally, the study highlights the prevalence of film pertinence in African agricultural documentaries produced for esteemed organizations such as the Roundtable on Sustainable Palm Oil (RSPO), Proforest, World Food Program, Fidelity Bank Ghana, Instituto BVRio, Aflatoun International, and the Solidaridad Network. These documentaries serve to promote prosperity, resilience, human rights, sustainable farming practices, community respect, and environmental preservation, underlining the vital role of film in conveying these critical messages. In summary, this research offers valuable insights into the evolving landscape of videography in different contexts, emphasizing the significance of film pertinence as a unifying principle in the pursuit of effective visual storytelling and cross-cultural communication.

Keywords: film pertinence, Africa, cultural awareness, videography tools

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