Search results for: sequences of AMR gene
465 Genetic Diversity Based Population Study of Freshwater Mud Eel (Monopterus cuchia) in Bangladesh
Authors: M. F. Miah, K. M. A. Zinnah, M. J. Raihan, H. Ali, M. N. Naser
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As genetic diversity is most important for existing, breeding and production of any fish; this study was undertaken for investigating genetic diversity of freshwater mud eel, Monopterus cuchia at population level where three ecological populations such as flooded area of Sylhet (P1), open water of Moulvibazar (P2) and open water of Sunamganj (P3) districts of Bangladesh were considered. Four arbitrary RAPD primers (OPB-12, C0-4, B-03 and OPB-08) were screened and RAPD banding patterns were analyzed among the populations considering 15 individuals of each population. In total 174, 138 and 149 bands were detected in the populations of P1, P2 and P3 respectively; however, each primer revealed less number of bands in each population. 100% polymorphic loci were recorded in P2 and P3 whereas only one monomorphic locus was observed in P1, recorded 97.5% polymorphism. Different genetic parameters such as inter-individual pairwise similarity, genetic distance, Nei genetic similarity, linkage distances, cluster analysis and allelic information, etc. were considered for measuring genetic diversity. The average inter-individual pairwise similarity was recorded 2.98, 1.47 and 1.35 in P1, P2 and P3 respectively. Considering genetic distance analysis, the highest distance 1 was recorded in P2 and P3 and the lowest genetic distance 0.444 was found in P2. The average Nei genetic similarity was observed 0.19, 0.16 and 0.13 in P1, P2 and P3, respectively; however, the average linkage distance was recorded 24.92, 17.14 and 15.28 in P1, P3 and P2 respectively. Based on linkage distance, genetic clusters were generated in three populations where 6 clades and 7 clusters were found in P1, 3 clades and 5 clusters were observed in P2 and 4 clades and 7 clusters were detected in P3. In addition, allelic information was observed where the frequency of p and q alleles were observed 0.093 and 0.907 in P1, 0.076 and 0.924 in P2, 0.074 and 0.926 in P3 respectively. The average gene diversity was observed highest in P2 (0.132) followed by P3 (0.131) and P1 (0.121) respectively.Keywords: genetic diversity, Monopterus cuchia, population, RAPD, Bangladesh
Procedia PDF Downloads 496464 Speaker Identification by Atomic Decomposition of Learned Features Using Computational Auditory Scene Analysis Principals in Noisy Environments
Authors: Thomas Bryan, Veton Kepuska, Ivica Kostanic
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Speaker recognition is performed in high Additive White Gaussian Noise (AWGN) environments using principals of Computational Auditory Scene Analysis (CASA). CASA methods often classify sounds from images in the time-frequency (T-F) plane using spectrograms or cochleargrams as the image. In this paper atomic decomposition implemented by matching pursuit performs a transform from time series speech signals to the T-F plane. The atomic decomposition creates a sparsely populated T-F vector in “weight space” where each populated T-F position contains an amplitude weight. The weight space vector along with the atomic dictionary represents a denoised, compressed version of the original signal. The arraignment or of the atomic indices in the T-F vector are used for classification. Unsupervised feature learning implemented by a sparse autoencoder learns a single dictionary of basis features from a collection of envelope samples from all speakers. The approach is demonstrated using pairs of speakers from the TIMIT data set. Pairs of speakers are selected randomly from a single district. Each speak has 10 sentences. Two are used for training and 8 for testing. Atomic index probabilities are created for each training sentence and also for each test sentence. Classification is performed by finding the lowest Euclidean distance between then probabilities from the training sentences and the test sentences. Training is done at a 30dB Signal-to-Noise Ratio (SNR). Testing is performed at SNR’s of 0 dB, 5 dB, 10 dB and 30dB. The algorithm has a baseline classification accuracy of ~93% averaged over 10 pairs of speakers from the TIMIT data set. The baseline accuracy is attributable to short sequences of training and test data as well as the overall simplicity of the classification algorithm. The accuracy is not affected by AWGN and produces ~93% accuracy at 0dB SNR.Keywords: time-frequency plane, atomic decomposition, envelope sampling, Gabor atoms, matching pursuit, sparse dictionary learning, sparse autoencoder
Procedia PDF Downloads 285463 Genomic Characterisation of Equine Sarcoid-derived Bovine Papillomavirus Type 1 and 2 Using Nanopore-Based Sequencing
Authors: Lien Gysens, Bert Vanmechelen, Maarten Haspeslagh, Piet Maes, Ann Martens
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Bovine papillomavirus (BPV) types 1 and 2 play a central role in the etiology of the most common neoplasm in horses, the equine sarcoid. The unknown mechanism behind the unique variety in a clinical presentation on the one hand and the host-dependent clinical outcome of BPV-1 infection, on the other hand, indicate the involvement of additional factors. Earlier studies have reported the potential functional significance of intratypic sequence variants, along with the existence of sarcoid-sourced BPV variants. Therefore, intratypic sequence variation seems to be an important emerging viral factor. This study aimed to give a broad insight in sarcoid-sourced BPV variation and explore its potential association with disease presentation. In order to do this, a nanopore sequencing approach was successfully optimized for screening a wide spectrum of clinical samples. Specimens of each tumour were initially screened for BPV-1/-2 by quantitative real-time PCR. A custom-designed primer set was used on BPV-positive samples to amplify the complete viral genome in two multiplex PCR reactions, resulting in a set of overlapping amplicons. For phylogenetic analysis, separate alignments were made of all available complete genome sequences for BPV-1/-2. The resulting alignments were used to infer Bayesian phylogenetic trees. We found substantial genetic variation among sarcoid-derived BPV-1, although this variation could not be linked to disease severity. Several of the BPV-1 genomes had multiple major deletions. Remarkably, the majority of the cluster within the region coding for late viral genes. Together with the extensiveness (up to 603 nucleotides) of the described deletions, this suggests an altered function of L1/L2 in disease pathogenesis. By generating a significant amount of complete-length BPV genomes, we succeeded in introducing next-generation sequencing into veterinary research focusing on the equine sarcoid, thus facilitating the first report of both nanopore-based sequencing of complete sarcoid-sourced BPV-1/-2 and the simultaneous nanopore sequencing of multiple complete genomes originating from a single clinical sample.Keywords: Bovine papillomavirus, equine sarcoid, horse, nanopore sequencing, phylogenetic analysis
Procedia PDF Downloads 173462 Seroprevalence and Associated Factors of Hepatitis B and Hepatitis C Viral Infections Among Prisoners in Tigray, Northern Ethiopia
Authors: Belaynesh Tsegay, Teklay Gebrecherkos, Atsebaha Gebrekidan Kahsay, Mahmud Abdulkader
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Background: Hepatitis B and C viruses are important health and socioeconomic problem across the globe, with a remarkable number of diseases and deaths in sub-Saharan African countries. The burden of hepatitis is unknown in the prison settings of Tigray. Therefore, we aimed to describe the seroprevalence and associated factors of hepatitis B and C viruses among prisoners in Tigray, Ethiopia. Methods: A cross-sectional study was carried out from February 2020 to May 2020 at the prison facilities of Tigray. Demographics and associated factors were collected from 315 prisoners prospectively. Five milliliters of blood were collected and tested using rapid tests kits of HBsAg (Zhejiang orient Gene Biotech Co., Ltd., China) and HCV antibodies (Volkan Kozmetik Sanayi Ve Ticaret Ltd. STI, Turkey). Positive samples were confirmed using ELISA (Beijing Wantai Biological Pharmacy Enterprise Co. Ltd). Data were analyzed using the SPSS version 20, and p<0.05 was considered statistically significant. Results: The overall seroprevalence of HBV and HCV were 25 (7.9%) and 1 (0.3%), respectively. The majority of hepatitis B viral infections were identified from the age groups of 18–25 years (10.7%) and unmarried prisoners (11.8%). Prisoners greater than 100 per cell (AOR=3.95, 95% CI=1.15–13.6, p=0.029) and with a history of alcohol consumption (AOR=3.01, 95% CI=1.17–7.74, p=0.022) were significantly associated with HBV infections. Conclusion: The seroprevalence of HBV among prisoners was nearly high or borderline, with a very low HCV prevalence. HBV was most prevalent among young adults, those housed with a large number of prisoners per cell, and those who had a history of alcohol consumption. This study recommends that there should be prison-focused intervention, including regular health education, with the emphasis on the mode of transmission and introducing HBV screening policy for prisoners, especially when they enter the prison.Keywords: seroprevalence, HBV, HCV, prisoners, tigray
Procedia PDF Downloads 75461 Survey of the Effect of the Probiotic Bacterium Lactobacillus plantarum and Streptococcus mutans on Casp3, AKT/PTEN, and MAPK Signaling Pathways at Co-Culture with KB Oral Cancer Cell Line and HUVEC Cells
Authors: Negar Zaheddoust, Negin Zaheddoust, Abbas Asoudeh-Fard
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Probiotic bacteria have been employed as a novel and less side-effect strategy for anticancer therapy. Since the oral cavity is a host for probiotic and pathogen bacteria to colonize, more investigation is needed to evaluate the effectiveness of this novel adjunctive treatment for oral cancer. We considered Lactobacillus plantarum as a probiotic and Streptococcus mutans as a pathogen bacterium in our study. The aim of this study is to examine the effect of Lactobacillus plantarum and Streptococcus mutans on Casp3, AKT / PTEN, and MAPK signaling pathway, which is involved in apoptosis or survival of oral cancer KB cells. On the other hand, to study the effects of these bacteria on normal cells, we used HUVEC cells. The KB and HUVEC cell lines were co-cultured with Lactobacillus plantarum and Streptococcus mutans isolated from traditional Iranian dairy and dental plaque, respectively. The growth-inhibitory effects of these two bacteria on KB and HUVEC cells were determined by (3-(4, 5-dimethylthiazolyl-2)-2,5diphenyltetrazolium bromide) MTT assay. MTT results demonstrated that the proliferation of KB cells was affected in a time, dose, and strain-dependent manner. In the following, the examination of induced apoptosis or necrosis in co-cultured KB cells with the best IC50 concentration of the Lactobacillus plantarum and Streptococcus mutans will be analyzed by FACS flow cytometry, and the changes in gene expression of Casp3, AKT / PTEN, MAPK genes will be evaluated using real-time polymerase chain reaction.Keywords: cancer therapy, induced apoptosis, oral cancer, probiotics
Procedia PDF Downloads 241460 Biodegradation of Direct Red 23 by Bacterial Consortium Isolated from Dye Contaminated Soil Using Sequential Air-lift Bioreactor
Authors: Lata Kumari Dhanesh Tiwary, Pradeep Kumar Mishra
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The effluent coming from various industries such as textile, carpet, food, pharmaceutical and many other industries is big challenge due to its recalcitrant and xenobiotiocs in nature. Recently, biodegradation of dye wastewater through biological means was widely used due to eco-friendly and cost effective with the higher percentage of removal of dye from wastewater. The present study deals with the biodegradation and decolourization of Direct Red 23 dye using indigenously isolated bacterial consortium. The bacterial consortium was isolated from soil sample from dye contaminated site near a cluster of Carpet industries of Bhadohi, Uttar Pradesh, India. The bacterial strain formed consortia were identified and characterized by morphological, biochemical and 16S rRNA gene sequence analysis. The bacterial strain mainly Staphylococcus saprophyticus strain BHUSS X3 (KJ439576), Microbacterium sp. BHUMSp X4 (KJ740222) and Staphylococcus saprophyticus strain BHUSS X5 (KJ439576) were used as consortia for further studies of dye decolorization. Experimental investigations were made in a Sequencing Air- lift bioreactor using the synthetic solution of Direct Red 23 dye by optimizing various parameters for efficient degradation of dye. The effect of several operating parameters such as flow rate, pH, temperature, initial dye concentration and inoculums size on removal of dye was investigated. The efficiency of isolated bacterial consortia from dye contaminated area in Sequencing Air- lift Bioreactor with different concentration of dye between 100-1200 mg/l at different hydraulic rate (HRTs) 26h and 10h. The maximum percentage of dye decolourization 98% was achieved when operated at HRT of 26h. The percentage of decolourization of dye was confirmed by using UV-Vis spectrophotometer and HPLC.Keywords: carpet industry, bacterial consortia, sequencing air-lift bioreactor
Procedia PDF Downloads 330459 A Comparative Study of Environment Risk Assessment Guidelines of Developing and Developed Countries Including Bangladesh
Authors: Syeda Fahria Hoque Mimmi, Aparna Islam
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Genetically engineered (GE) plants are the need of time for increased demand for food. A complete set of regulations need to be followed from the development of a GE plant to its release into the environment. The whole regulation system is categorized into separate stages for maintaining the proper biosafety. Environmental risk assessment (ERA) is one of such crucial stages in the whole process. ERA identifies potential risks and their impacts through science-based evaluation where it is done in a case-by-case study. All the countries which deal with GE plants follow specific guidelines to conduct a successful ERA. In this study, ERA guidelines of 4 developing and 4 developed countries, including Bangladesh, were compared. ERA guidelines of countries such as India, Canada, Australia, the European Union, Argentina, Brazil, and the US were considered as a model to conduct the comparison study with Bangladesh. Initially, ten parameters were detected to compare the required data and information among all the guidelines. Surprisingly, an adequate amount of data and information requirements (e.g., if the intended modification/new traits of interest has been achieved or not, the growth habit of GE plants, consequences of any potential gene flow upon the cultivation of GE plants to sexually compatible plant species, potential adverse effects on the human health, etc.) matched between all the countries. However, a few differences in data requirement (e.g., agronomic conventions of non-transformed plants, applicants should clearly describe experimental procedures followed, etc.) were also observed in the study. Moreover, it was found that only a few countries provide instructions on the quality of the data used for ERA. If these similarities are recognized in a more framed manner, then the approval pathway of GE plants can be shared.Keywords: GE plants, ERA, harmonization, ERA guidelines, Information and data requirements
Procedia PDF Downloads 182458 Osteogenesis in Thermo-Sensitive Hydrogel Using Mesenchymal Stem Cell Derived from Human Turbinate
Authors: A. Reum Son, Jin Seon Kwon, Seung Hun Park, Hai Bang Lee, Moon Suk Kim
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These days, stem cell therapy is focused on for promising source of treatment in clinical human disease. As a supporter of stem cells, in situ-forming hydrogels with growth factors and cells appear to be a promising approach in tissue engineering. To examine osteogenic differentiation of hTMSCs which is one of mesenchymal stem cells in vivo in an injectable hydrogel, we use a methoxy polyethylene glycol-polycaprolactone blockcopolymer (MPEG-PCL) solution with osteogenic factors. We synthesized MPEG-PCL hydrogel and measured viscosity to check sol-gel transition. In order to demonstrate osteogenic ability of hTMSCs, we conducted in vitro osteogenesis experiment. Then, to confirm the cell cytotoxicity, we performed WST-1 with hTMSCs and MPEG-PCL. As the result of in vitro experiment, we implanted cell and hydrogel mixture into animal model and checked degree of osteogenesis with histological analysis and amount of expression genes. Through these experimental data, MPEG-PCL hydrogel has sol-gel transition in temperature change and is biocompatible with stem cells. In histological analysis and gene expression, hTMSCs are very good source of osteogenesis with hydrogel and will use it to tissue engineering as important treatment method. hTMSCs could be a good adult stem cell source for usability of isolation and high proliferation. When hTMSCs are used as cell therapy method with in situ-formed hydrogel, they may provide various benefits like a noninvasive alternative for bone tissue engineering applications.Keywords: injectable hydrogel, stem cell, osteogenic differentiation, tissue engineering
Procedia PDF Downloads 442457 Refusal Speech Acts in French Learners of Mandarin Chinese
Authors: Jui-Hsueh Hu
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This study investigated various models of refusal speech acts among three target groups: French learners of Mandarin Chinese (FM), Taiwanese native Mandarin speakers (TM), and native French speakers (NF). The refusal responses were analyzed in terms of their options, frequencies, and sequences and the contents of their semantic formulas. This study also examined differences in refusal strategies, as determined by social status and social distance, among the three groups. The difficulties of refusal speech acts encountered by FM were then generalized. The results indicated that Mandarin instructors of NF should focus on the different reasons for the pragmatic failure of French learners and should assist these learners in mastering refusal speech acts that rely on abundant cultural information. In this study, refusal policies were mainly classified according to the research of Beebe et al. (1990). Discourse completion questionnaires were collected from TM, FM, and NF, and their responses were compared to determine how refusal policies differed among the groups. This study not only emphasized the dissimilarities of refusal strategies between native Mandarin speakers and second-language Mandarin learners but also used NF as a control group. The results of this study demonstrated that regarding overall strategies, FM were biased toward NF in terms of strategy choice, order, and content, resulting in pragmatic transfer under the influence of social factors such as 'social status' and 'social distance,' strategy choices of FM were still closer to those of NF, and the phenomenon of pragmatic transfer of FM was revealed. Regarding the refusal difficulties among the three groups, the F-test in the analysis of variance revealed statistical significance was achieved for Role Playing Items 13 and 14 (P < 0.05). A difference was observed in the average number of refusal difficulties between the participants. However, after multiple comparisons, it was found that item 13 (unrecognized heterosexual junior colleague requesting contacts) was significantly more difficult for NF than for TM and FM; item 14 (contacts requested by an unrecognized classmate of the opposite sex) was significantly more difficult to refuse for NF than for TM. This study summarized the pragmatic language errors that most FM often perform, including the misuse or absence of modal words, hedging expressions, and empty words at the end of sentences, as the reasons for pragmatic failures. The common social pragmatic failures of FM include inaccurately applying the level of directness and formality.Keywords: French Mandarin, interlanguage refusal, pragmatic transfer, speech acts
Procedia PDF Downloads 246456 The Generation of Insulin Producing Cells from Human Mesenchymal Stem Cells by miR-375 and Anti-miR-9
Authors: Arefeh Jafarian, Mohammad Taghikani, Saied Abroun, Amir Allahverdi, Masoud Soleimani
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Introduction: The miRNAs have key roles in control of pancreatic islet development and insulin secretion. In this regards, current study investigated the pancreatic differentiation of human bone marrow mesenchymal stem cells (hBM-MSCs) by up-regulation of miR-375 and down-regulation of miR-9 by lentiviruses containing miR-375 and anti-miR-9. Findings: After 21 days of induction, islet-like clusters containing insulin producing cells (IPCs) were confirmed by dithizone (DTZ) staining. The IPCs and β cell specific related genes and proteins were detected using qRT-PCR and immunofluorescence on days 7, 14 and 21 of differentiation. Glucose challenge test was performed at different concentrations of glucose as well as extracellular and intracellular insulin and C-peptide were assayed using ELISA kit. In derived IPCs by miR-375 alone are capable to express insulin and other endocrine specific transcription factors, the cells lack the machinery to respond to glucose. The differentiated hMSCs by miR-375 and anti-miR-9 lentiviruses could secrete insulin and c-peptide in a glucose-regulated manner. Conclusion: It was found that over-expression of miR-375 led to a reduction in levels of Mtpn protein in derived IPCs, while treatment with anti-miR-9 following miR-375 over-expression had synergistic effects on MSCs differentiation and insulin secretion in a glucose-regulated manner. The researchers reported that silencing of miR-9 increased OC-2 protein in IPCs that may contribute to the observed glucose-regulated insulin secretion. These findings highlight miRNAs functions in stem cells differentiation and suggest that they could be used as therapeutic tools for gene-based therapy in diabetes mellitus.Keywords: diabetes, differentiation, MSCs, insulin producing cells, miR-375, miR-9
Procedia PDF Downloads 312455 Positive effect of Cu2+ and Ca2+ on the Thermostability of Bambara Groundnut Peroxidase A6, and its Catalytic Efficiency Toward the Oxidation of 3,3,5,5 -Tetramethyl Benzidine
Authors: Yves Mann Elate Lea Mbassi, Marie Solange Evehe Bebandoue, Wilfred Fon Mbacham
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Improving the catalytic performance of enzymes has been a long-standing theme of analytical biochemistry research. Induction of peroxidase activity by metals is a common reaction in higher plants. We thought that this increase in peroxidase activity may be due, on the one hand, to the stimulation of the gene expression of these enzymes but also to a modification of their chemical reactivity following the binding of some metal ions on their active site. We tested the effect of some metal salts (MgCl₂, MnCl₂, ZnCl₂, CaCl₂ and CuSO₄) on the activity and thermostability of peroxidase A6, a thermostable peroxidase that we discovered and purified in a previous study. The chromogenic substrate used was 3,3′,5,5′-tetramethylbenzidine. Of all the metals tested for their effect on A6, only magnesium and copper had a significant effect on the activity of the enzyme at room temperature. The Mann-Whitney test shows a slight inhibitory effect of activity by the magnesium salt (P = 0.043), while the activity of the enzyme is 5 times higher in the presence of the copper salt (P = 0.002). Moreover, the thermostability of peroxidase A6 is increased when calcium and copper salts are present. The activity in the presence of CaCl₂ is 8 times higher than the residual activity of the enzyme alone after incubation at 80°C for 10 min and 35 times higher in the presence of CuSO4 under the same conditions. In addition, manganese and zinc salts slightly reduce the thermostability of the enzyme. The activity and structural stability of peroxidase A6 can clearly be activated by Cu₂+, which therefore enhance the oxidation of 3,3′,5,5′-tetramethylbenzidine, which was used in this study as a chromogenic substrate. Ca₂+ likely has a more stabilizing function for the catalytic site.Keywords: peroxidase activity, copper ions, calcium ions, thermostability
Procedia PDF Downloads 68454 Formulation Development and Characterization of Oligonucleotide Containing Chitosan Nanoparticles
Authors: Gyati Shilakari Asthana, Abhay Asthana
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Purpose: The therapeutic potential of oligonucleotide (ODN) is primarily dependent upon its safe and efficient delivery to specific cells overcoming degradation and maximizing cellular uptake in vivo. The present study is focused to design low molecular weight chitosan nanoconstructs to meet the requirements of safe and effectual delivery of ODNs. LMW-chitosan is a biodegradable, water soluble, biocompatible polymer and is useful as a non-viral vector for gene delivery due to its better stability in water. Methods: LMW chitosan ODN nanoparticles (CHODN NPs) were formulated by self assembled method using various N/P ratios (moles ratio of amine groups of CH to phosphate moieties of ODNs; 0.5:1, 1:1, 3:1, 5:1 and 7:1) of CH to ODN. The developed CHODN NPs were evaluated with respect to gel retardation assay, particle size, zeta potential and cytotoxicity and transfection efficiency. Results: Complete complexation of CH/ODN was achieved at the charge ratio of 0.5:1 or above and CHODN NPs displayed resistance against DNase I. On increasing the N/P ratio of CH/ODN, particle size of the NPs decreased whereas zeta potential (ZV) value increased. No significant toxicity was observed at all CH concentrations. The transfection efficiency was increased on increasing N/P ratio from 1:1 to 3:1, whereas it was decreased with further increment in N/P ratio upto 7:1. Maximum transfection of CHODN NPs with both the cell lines (Raw 267.4 cells and Hela cells) was achieved at N/P ratio of 3:1. The results suggest that transfection efficiency of CHODN NPs is dependent on N/P ratio. Conclusion: Thus the present study states that LMW chitosan nanoparticulate carriers would be acceptable choice to improve transfection efficiency in vitro as well as in vivo delivery of oligonucleotide.Keywords: LMW-chitosan, chitosan nanoparticles, biocompatibility, cytotoxicity study, transfection efficiency, oligonucleotide
Procedia PDF Downloads 490453 Genome-Wide Expression Profiling of Cicer arietinum Heavy Metal Toxicity
Authors: B. S. Yadav, A. Mani, S. Srivastava
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Chickpea (Cicer arietinum L.) is an annual, self-pollinating, diploid (2n = 2x = 16) pulse crop that ranks second in world legume production after common bean (Phaseolus vulgaris). ICC 4958 flowers approximately 39 days after sowing under peninsular Indian conditions and the crop matures in less than 90 days in rained environments. The estimated collective yield losses due to abiotic stresses (6.4 million t) have been significantly higher than for biotic stresses (4.8 million t). Most legumes are known to be salt sensitive, and therefore, it is becoming increasingly important to produce cultivars tolerant to high-salinity in addition to other abiotic and biotic stresses for sustainable chickpea production. Our aim was to identify the genes that are involved in the defence mechanism against heavy metal toxicity in chickpea and establish the biological network of heavy metal toxicity in chickpea. ICC4958 variety of chick pea was taken and grown in normal condition and 150µM concentration of different heavy metal salt like CdCl₂, K₂Cr2O₇, NaAsO₂. At 15th day leave samples were collected and stored in RNA Later solution microarray was performed for checking out differential gene expression pattern. Our studies revealed that 111 common genes that involved in defense mechanism were up regulated and 41 genes were commonly down regulated during treatment of 150µM concentration of CdCl₂, K₂Cr₂O₇, and NaAsO₂. Biological network study shows that the genes which are differentially expressed are highly connected and having high betweenness and centrality.Keywords: abiotic stress, biological network, chickpea, microarray
Procedia PDF Downloads 186452 Sequence Stratigraphy and Petrophysical Analysis of Sawan Gas Field, Central Indus Basin, Pakistan
Authors: Saeed Ur Rehman Chaudhry
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The objectives of the study are to reconstruct sequence stratigraphic framework and petrophysical analysis of the reservoir marked by using sequence stratigraphy of Sawan Gas Field. The study area lies in Central Indus Basin, District Khairpur, Sindh province, Pakistan. The study area lies tectonically in an extensional regime. Lower Goru Formation and Sembar Formation act as a reservoir and source respectively. To achieve objectives, data set of seismic lines, consisting of seismic lines PSM96-114, PSM96-115, PSM96-133, PSM98-201, PSM98-202 and well logs of Sawan-01, Sawan-02 and Gajwaro-01 has been used. First of all interpretation of seismic lines has been carried out. Interpretation of seismic lines shows extensional regime in the area and cut entire Cretaceous section. Total of seven reflectors has been marked on each seismic line. Lower Goru Formation is thinning towards west. Seismic lines also show eastward tilt of stratigraphy due to uplift at the western side. Sequence stratigraphic reconstruction has been done by integrating seismic and wireline log data. Total of seven sequence boundaries has been interpreted between the top of Chiltan Limestone to Top of Lower Goru Formation. It has been observed on seismic lines that Sembar Formation initially generated shelf margin profile and then ramp margin on which Lower Goru deposition took place. Shelf edge deltas and slope fans have been observed on seismic lines, and signatures of slope fans are also observed on wireline logs as well. Total of six sequences has been interpreted. Stratigraphic and sequence stratigraphic correlation has been carried out by using Sawan 01, Sawan 02 and Gajwaro 01 and a Low Stand Systems tract (LST) within Lower Goru C sands has been marked as a zone of interest. The petrophysical interpretation includes shale volume, effective porosity, permeability, saturation of water and hydrocarbon. On the basis of good effective porosity and hydrocarbon saturation petrophysical analysis confirms that the LST in Sawan-01 and Sawan-02 has good hydrocarbon potential.Keywords: petrophysical analysis, reservoir potential, Sawan Gas Field, sequence stratigraphy
Procedia PDF Downloads 256451 Evaluation of Adaptive Fitness of Indian Teak (Tectona grandis L. F.) Metapopulation through Inter Simple Sequence Repeat Markers
Authors: Vivek Vaishnav, Shamim Akhtar Ansari
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Teak (Tectona grandis L.f.) belonging to plant family Lamiaceae and the most commercialized timber species is endemic to South-Asia. The adaptive fitness of the species metapopulation was evaluated through its genetic differentiation and assessing the influence of geo-climatic conditions. 290 genotypes were sampled from 29 locations of its natural distribution and the genetic data was incorporated with geo-climatic parameters. Through Bayesian approach based analysis of 43 highly polymorphic ISSR markers, six homogeneous clusters (0.8% genetic variability) were identified. The six clusters were found with the various regimes of the temperature range, i.e., I - 9.10±1.35⁰C, II -6.35±0.21⁰C, III -12.21±0.43⁰C, IV - 10.8±1.06⁰C, V - 11.67±3.04⁰C, and VI - 12.35±0.21⁰C. The population had a very high percentage of LD (21.48%) among the amplified loci possibly due to experiencing restricted gene flow as well as co-adaptation and association of distant/diverse loci/alleles as a result of the stabilized climatic conditions and countless cycles of historical recombination events on a large geological timescale. The same possibly accounts for the narrow distribution of teak as a climax species in the tropical deciduous forests of the country. The regions of strong LD in teak genome significantly associated with climatic parameters also reflect that the species is tolerant to the wide regimes of the temperature range and may possibly withstand global warming and climate change in the coming millennium.Keywords: Bayesian analysis, inter simple sequence repeat, linkage disequilibrium, marker-geoclimatic association
Procedia PDF Downloads 258450 Role of Inflammatory Markers in Arthritic Rats Treated with Ethanolic Bark Extract of Albizia procera
Authors: M. Sangeetha, D. Chamundeeswari, C. Saravanababu, C. Rose, V. Gopal
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Rheumatoid arthritis (RA) is a chronic, progressive, systemic inflammatory disorder affecting the synovial joints and typically producing symmetrical arthritis that leads to joint destruction, which is responsible for the deformity and disability. Despite improvements in the treatment of RA over the past decade, there still is a need for new therapeutic agents that are efficacious, less expensive, and free of severe adverse reactions. The present study aimed to investigate role of inflammatory markers in arthritic rats treated with ethanolic bark extract of Albizia procera. The protective effect of ethanolic bark extract of Albizia procera against complete Freund’s adjuvant (CFA) induced arthritis in rats. Arthritis was induced by an intradermal injection of 0.1 ml FCA in the foot pad of left hind limb of rats. ETBE (100 and 200 mg/kg b.wt./p.o) and the reference drug diclofenac (25 mg/kg b.wt./p.o) were administered to arthritic rats. Paw volume was measured for all the animals before inducing arthritis and thereafter once in seven days by using plethysmometer for 42 days. Gene expression of inflammatory markers such as IL-1β and IL-10 were investigated in paw tissues. Up regulation of IL-1β and Down regulation IL-10 were observed in CFA injected rats when compared to normal rats. ETBE attenuated these alterations dose dependently when compared to the vehicle treated rats. These results provide insights into the mechanism of anti-arthritic activity, and unravel potential therapeutic use of Albizia procera in arthritis.
Keywords: CFA-Complete Freund’s adjuvant, ETBE – ethanolic bark extract, IL- interleukins, RA-rheumatoid arthritis
Procedia PDF Downloads 277449 Cytotoxicity and Androgenic Potential of Antifungal Drug Substances on MDA-KB2 Cells
Authors: Benchouala Amira, Bojic Clement, Poupin Pascal, Cossu Leguille-carole
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The objective of this study is to evaluate in vitro the cytotoxic and androgenic potential of several antifungal molecules (amphotericin B, econazole, ketoconazole and miconazole) on MDA-Kb2 cell lines. This biological model is an effective tool for the detection of endocrine disruptors because it responds well to the main agonist of the androgen receptor (testosterone) and also to an antagonist: flutamide. The cytotoxicity of each chemical compound tested was measured using an MTT assay (tetrazolium salt, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) which measures the activity of the reductase function of mitochondrial succinate dehydrogenase enzymes of cultured cells. This complementary cytotoxicity test is essential to ensure that the effects of reduction in luminescence intensity observed during androgenic tests are only attributable to the anti-androgenic action of the compounds tested and not to their possible cytotoxic properties. Tests of the androgenic activity of antifungals show that these compounds do not have the capacity to induce transcription of the luciferase gene. These compounds do not exert an androgenic effect on MDA-Kb2 cells in culture for the environmental concentrations tested. The addition of flutamide for the same tested concentrations of antifungal molecules reduces the luminescence induced by amphotericin B, econazole and miconazole, which is explained by a strong interaction of these molecules with flutamide which may have a greater toxic effect than when tested alone. The cytotoxicity test shows that econazole and ketoconazole can cause cell death at certain concentrations tested. This cell mortality is perhaps induced by a direct or indirect action on deoxyribonucleic acid (DNA), ribonucleic acid (RNA) or proteins necessary for cell division.Keywords: cytotoxicity, androgenic potential, antifungals, MDA-Kb2
Procedia PDF Downloads 37448 African Swine Fewer Situation and Diagnostic Methods in Lithuania
Authors: Simona Pileviciene
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On 24th January 2014, Lithuania notified two primary cases of African swine fever (ASF) in wild boars. The animals were tested positive for ASF virus (ASFV) genome by real-time PCR at the National Reference Laboratory for ASF in Lithuania (NRL), results were confirmed by the European Union Reference Laboratory for African swine fever (CISA-INIA). Intensive wild and domestic animal monitoring program was started. During the period of 2014-2017 ASF was confirmed in two large commercial pig holding with the highest biosecurity. Pigs were killed and destroyed. Since 2014 ASF outbreak territory from east and south has expanded to the middle of Lithuania. Diagnosis by PCR is one of the highly recommended diagnostic methods by World Organization for Animal Health (OIE) for diagnosis of ASF. The aim of the present study was to compare singleplex real-time PCR assays to a duplex assay allowing the identification of ASF and internal control in a single PCR tube and to compare primers, that target the p72 gene (ASF 250 bp and ASF 75 bp) effectivity. Multiplex real-time PCR assays prove to be less time consuming and cost-efficient and therefore have a high potential to be applied in the routine analysis. It is important to have effective and fast method that allows virus detection at the beginning of disease for wild boar population and in outbreaks for domestic pigs. For experiments, we used reference samples (INIA, Spain), and positive samples from infected animals in Lithuania. Results show 100% sensitivity and specificity.Keywords: African swine fewer, real-time PCR, wild boar, domestic pig
Procedia PDF Downloads 160447 Antibacterial Activity of Bacillus thuringiensis Activated Delta-endotoxins
Authors: R. Gounina-Allouane, N. Ouali, F. Z. Berrabah, A. Bentaleb
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For a long time, the Gram-positive spore-forming bacteria Bacillus thuringiensis (Bt) has been widely used in biological control against devastating and disease vectors insects. This is due to the insecticidal activity of its crystalline parasporal inclusion (crystals) predominantly comprised of one or more proteins (Cry and Cyt proteins) also called δ-endotoxins, produced during sporulation. The shape and composition of Bt crystals vary among strains and crystalline proteins are extremely varied (more than 475 cry gene were discovered). The insecticidal activity of Bt crystals is very well studied, thus their insecticidal mode of action is well established, however, their antimicrobial effect is largely unknown. The lack of data on the antimicrobial effect of crystalline proteins of Bt and the need for searching new antimicrobial molecules encouraged us to carried out this study. The antibacterial effect of δ-endotoxines produced by two Bt stains; a strain isolated from soil at northern of Algeria (Bt 7.2.B), and a strain isolated from a bioinsecticide (Bacillus thuringiensis var aizawai), activated by proteolysis, was assayed on clinical bacterial strains and ATCC collection ones respectively. Gram positive and negative clinical bacterial strains (Escherichia coli, Klebsiella pneumonaie, Pseudomonas aeruginosa, Staphylococcus aureus) were sensitive to activated Bt 72B endotoxins. Similarly, bacterial strains from ATCC collection (Escherichia coli ATCC 25922, Pseudomonas aerugenosa ATCC 27853, Staphylococcus aureus ATCC 25923) were sensitive to activated B. thuringiensis var aizawai δ-endotoxines. The activated δ-endotoxins were separated by SDS-PAGE.Keywords: Bacillus thuringiensis, crystals, cry proteins, δ-endotoxins, antibacterial activity
Procedia PDF Downloads 444446 Cannabidiol (CBD) Resistant Salmonella Strains Are Susceptible to Epsilon 34 Phage Tailspike Protein
Authors: Ibrahim Iddrisu, Joseph Ayariga, Junhuan Xu, Ayomide Adebanjo, Boakai K. Robertson, Michelle Samuel-Foo, Olufemi Ajayi
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The rise of antimicrobial resistance is a global public health crisis that threatens the effective control and prevention of infections. Due to the emergence of pan drug-resistant bacteria, most antibiotics have lost their efficacy. Bacteriophages or their components are known to target bacterial cell walls, cell membranes, and lipopolysaccharides (LPS) and hydrolyze them. Bacteriophages, being the natural predators of pathogenic bacteria, are inevitably categorized as ‘human friends’, thus fulfilling the adage that ‘the enemy of my enemy is my friend’. Leveraging on their lethal capabilities against pathogenic bacteria, researchers are searching for more ways to overcome the current antibiotic resistance challenge. In this study, we expressed and purified epsilon 34 phage tail spike protein (E34 TSP) from the E34 TSP gene, then assessed the ability of this bacteriophage protein in the killing of two CBD-resistant strains of Salmonella spp. We also assessed the ability of the tail spike protein to cause bacteria membrane disruption and dehydrogenase depletion. We observed that the combined treatment of CBD-resistant strains of Salmonella with CBD and E34 TSP showed poor killing ability, whereas the mono treatment with E34 TSP showed considerably higher killing efficiency. This study demonstrates that the inhibition of the bacteria by E34 TSP was due in part to membrane disruption and dehydrogenase inactivation by the protein. The results of this work provide an interesting background to highlight the crucial role phage proteins such as E34 TSP could play in pathogenic bacterial control.Keywords: cannabidiol, resistance, Salmonella, antimicrobials, phages
Procedia PDF Downloads 57445 Evidence of Paternal Protein Provisioning During Male Pregnancy in the Seahorse, Hippocampus Abdominalis
Authors: Zoe M. G. Skalkos, Sam N. Dowland, James U. Van Dyke, Camilla. M. Whittington
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Syngnathid fishes (seahorses, pipefishes, and seadragons) are unique because embryos develop on or in the male in a specialised brooding structure. Many seahorse species are endangered or vulnerable, while others are popular in the ornamental fish trade. Seahorses are capable of nutrient provisioning (patrotrophy) of lipids during pregnancy via their fully enclosed brood pouch. Protein is vital for gene regulation and tissue growth during embryogenesis. We tested the hypothesis that protein is paternally transported to developing embryos during pregnancy in the Australian Pot-bellied seahorse, Hippocampus abdominalis. We compared the dry masses and nitrogen content in recently fertilised H. abdominalis embryos and newborns. We calculated an updated patrotrophy index, 1.34, but without a significant difference in dry mass between the two developmental stages. There was, however, a significant increase in total protein content from recently fertilised embryos to neonates. This suggests paternal protein transport is essential for H. abdominalis embryogenesis because protein yolk reserves are depleted by embryonic metabolism, and supplementation is required. This study is the first to provide evidence for paternal protein transport during pregnancy in seahorses. It furthers our understanding of the paternal influence on embryonic development in male pregnancy and how a protein-deficient diet during pregnancy may limit the allocation of resources to embryos, reducing offspring fitness. This research contributes to a deeper understanding of the fundamental reproductive biology of seahorses, which can help improve conservation and farming production outcomes.Keywords: brood pouch, embryonic provisioning, nitrogen, parentotrophy, paternal investment, reproduction
Procedia PDF Downloads 96444 The Effect of Different Metal Nanoparticles on Growth and Survival of Pseudomonas syringae Bacteria
Authors: Omar Alhamd, Peter A. Thomas, Trevor J. Greenhough, Annette K. Shrive
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The Pseudomonas syringae species complex includes many plant pathogenic strains with highly specific interactions with varied host species and cultivars. The rapid spread of these bacteria over the last ten years has become a cause for concern. Nanoparticles have previously shown promise in microbiological action. We have therefore investigated in vitro and in vivo the effects of different types and sizes of nanoparticles in order to provide quantitative information about their effect on the bacteria. The effects of several different nanoparticles against several bacteria strains were investigated. The effect of NP on bacterial growth was studied by measuring the optical density, biochemical and nutritional tests, and transmission electron microscopy (TEM) to determine the shape and size of NP. Our results indicate that their effects varied, with either a negative or a positive impact on both bacterial and plant growth. Additionally, the methods of exposure to nanoparticles have a crucial role in accumulation, translocation, growth response and bacterial growth. The results of our studies on the behaviour and effects of nanoparticles in model plants showed. Cerium oxide (CeO₂) and silver (Ag) NP showed significant antibacterial activity against several pathogenic bacteria. It was found that titanium nanoparticles (TiO₂) can have either a negative or a positive impact, according to concentration and size. It is also thought that environmental conditions can have a major influence on bacterial growth. Studies were therefore also carried out under some environmental stress conditions to test bacterial survival and to assess bacterial virulence. All results will be presented including information about the effects of different nanoparticles on Pseudomonas syringae bacteria.Keywords: plant microbiome, nanoparticles, 16S rRNA gene sequencing, bacterial survival
Procedia PDF Downloads 198443 Evaluating Cyanide Biodegradation by Bacteria Isolated from Gold Mine Effluents in Bulawayo, Zimbabwe
Authors: Ngonidzashe Mangoma, Caroline Marigold Sebata
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The release of cyanide-rich effluents from gold mines, and other industries, into the environment, is a global concern considering the well-known metabolic effects of cyanide in all forms of life. Such effluents need to be treated to remove cyanide, among other pollutants, before their disposal. This study aimed at investigating the possible use of bacteria in the biological removal of cyanide from cyanide-rich effluents. Firstly, cyanide-degrading bacteria were isolated from gold mine effluents and characterised. The isolates were then tested for their ability to grow in the presence of cyanide and their tolerance to increasing levels of the compound. To evaluate each isolate’s cyanide-degrading activities, isolates were grown in the simulated and actual effluent, and a titrimetric method was used to quantify residual cyanide over a number of days. Cyanide degradation efficiency (DE) was then calculated for each isolate. Identification of positive isolates involved 16S rRNA gene amplification and sequence analysis through BLAST. Six cyanide-utilising bacterial strains were isolated. Two of the isolates were identified as Klebsiella spp. while the other two were shown to be different strains of Clostridium bifermentans. All isolates showed normal growth in the presence of cyanide, with growth being inhibited at 700 mg/L cyanide and beyond. Cyanide degradation efficiency for all isolates in the simulated effluent ranged from 79% to 97%. All isolates were able to remove cyanide from actual gold mine effluent with very high DE values (90 – 94%) being recorded. Isolates obtained in this study were able to efficiently remove cyanide from both simulated and actual effluent. This observation clearly demonstrates the feasibility of the biological removal of cyanide from cyanide-rich gold mine effluents and should, therefore, motivate research towards the possible large-scale application of this technology.Keywords: cyanide effluent, bioremediation, Clostridium bifermentans, Klebsiella spp, environment
Procedia PDF Downloads 168442 Nitrification Efficiency and Community Structure of Municipal Activated Sewage Sludge
Authors: Oluyemi O. Awolusi, Abimbola M. Enitan, Sheena Kumari, Faizal Bux
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Nitrification is essential to biological processes designed to remove ammonia and/or total nitrogen. It removes the excess nitrogenous compound in wastewater which could be very toxic to the aquatic fauna or cause a serious imbalance of such aquatic ecosystem. Efficient nitrification is linked to an in-depth knowledge of the structure and dynamics of the nitrifying community structure within the wastewater treatment systems. In this study, molecular technique was employed for characterizing the microbial structure of activated sludge [ammonia oxidizing bacteria (AOB) and nitrite oxidizing bacteria (NOB)] in a municipal wastewater treatment with intention of linking it to the plant efficiency. PCR-based phylogenetic analysis was also carried out for. The average operating and environmental parameters, as well as specific nitrification rate of a plant, was investigated during the study. During the investigation, the average temperature was 23±1.5oC. Other operational parameters such as mixed liquor suspended solids and chemical oxygen demand inversely correlated with ammonia removal. The dissolved oxygen level in the plant was constantly lower than the optimum (between 0.24 and 1.267 mg/l) during this study. The plant was treating wastewater with the influent ammonia concentration of 31.69 and 24.47 mg/l. The influent flow rates (ML/day) was 96.81 during the period. The dominant nitrifiers include: Nitrosomonas spp. Nitrobacter spp. and Nitrospira spp. The AOB had a correlation with nitrification efficiency and temperature. This study shows that the specific ammonia oxidizing rate and the specific nitrate formation rates can serve as a good indicator of the plant overall nitrification performance.Keywords: Ammonia monooxygenase α-subunit gene, amoA, ammonia-oxidizing bacteria, AOB, nitrite-oxidizing bacteria, NOB, specific nitrification rate
Procedia PDF Downloads 452441 Hypervirulent Klebsiella Pneumoniae in a South African Tertiary Hospital – Clinical Profile, Genetic Determinants and Virulence in Caenorhabditis Elegans
Authors: Dingiswayo Likhona, Arko-Cobbah Emmanuel, Carolina Pohl, Nthabiseng Z. Mokoena, Jolly Musoke
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A distinct strain of Klebsiella pneumoniae (K. pneumoniae), referred to as hypervirulent (hvKp), is associated with invasive infections such as an invasive pyogenic liver abscess in young and healthy individuals. In South Africa, limited information is known about the prevalence and virulence of this hvKp strain. Thus, this study aimed to determine the prevalence of hvKp and virulence-associated factors in K. pneumoniae isolates from one of the largest Tertiary hospitals in a South African province. A total of 74 K. pneumoniae isolates were received from Pelonomi National Health Laboratory Services (NHLS), Bloemfontein. Virulence-associated genes (rmpA, capsule serotype K1/K2, iroB, and irp2) were screened, and the virulence of hvKp vs. classical Klebsiella pneumoniae (cKp) was investigated using Caenorhabditis elegans nematode model. The iutA (aerobactin transporter) gene was used as a primary biomarker of hvKp. An average of 12% (9/74) of cases were defined as hvKp. Moreover, hvKp was found to be significantly more virulent in vivo Caenorhabditis elegans relative to cKp. The virulence-associated genes (rmpA, iroB, hmv phenotype, and capsule K1/K2) were significantly (p< 0.05) associated with hvKp. Findings from this study confirm the presence of hvKp in one large Tertiary hospital in South Africa. However, the low prevalence and mild to moderate clinical presentation suggest a marginal threat to public health. Further studies in different settings are required to establish the true potential impact of hvKp in developing countries.Keywords: hypervirulent klebsiella pneumoniae, virulence, caenorhabditis elegans, aerobactin (iutA)
Procedia PDF Downloads 75440 Antibacterial Activity of Bacillus thuringiensis Cristalline Parasporal Proteins
Authors: R. Gounina-Allouane, N. Ouali, F. Z. Berrabah, A. Bentaleb
Abstract:
For a long time, the Gram-positive spore-forming bacteria Bacillus thuringiensis (Bt) has been widely used in biological control against devastating and disease vectors insects. This is due to the insecticidal activity of its crystalline parasporal inclusion (crystals) predominantly comprised of one or more proteins (Cry and Cyt proteins) also called δ-endotoxins, produced during sporulation. The shape and composition of Bt crystals vary among strains and crystalline proteins are extremely varied (more than 475 cry gene were discovered). The insecticidal activity of Bt crystals is very well studied, thus their insecticidal mode of action is well established, however, their antimicrobial effect is largely unknown. The lack of data on the antimicrobial effect of crystalline proteins of Bt and the need for searching new antimicrobial molecules encouraged us to carried out this study. The antibacterial effect of δ-endotoxines produced by two Bt stains; a strain isolated from soil at northern of Algeria (Bt 7.2.B), and a strain isolated from a bioinsecticide (Bacillus thuringiensis var aizawai), activated by proteolysis, was assayed on clinical bacterial strains and ATCC collection ones respectively. Gram positive and negative clinical bacterial strains (Escherichia coli, Klebsiella pneumonaie, Pseudomonas aeruginosa, Staphylococcus aureus) were sensitive to activated Bt 72B endotoxins. Similarly, bacterial strains from ATCC collection (Escherichia coli ATCC 25922, Pseudomonas aerugenosa ATCC 27853, Staphylococcus aureus ATCC 25923) were sensitive to activated B. thuringiensis var aizawai δ-endotoxines. The activated δ-endotoxins were separated by SDS-PAGE.Keywords: Bacillus thuringiensis, crystals, cry proteins, δ-endotoxins, antibacterial activity
Procedia PDF Downloads 428439 Epigenomic Analysis of Lgr5+ Stem Cells in Gastrointestinal Tract
Authors: Hyo-Min Kim, Seokjin Ham, Mi-Joung Yoo, Minseon Kim, Tae-Young Roh
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The gastrointestinal (GI) tract of most animals, including murine, is highly compartmentalized epithelia which also provide distinct different functions of its own tissue. Nevertheless, these epithelia share certain characteristics that enhance immune responses to infections and maintain the barrier function of the intestine. GI tract epithelia also undergo regeneration not only in homeostatic conditions but also in a response to the damage. A full turnover of the murine gastrointestinal epithelium occurs every 4-5 day, a process that is regulated and maintained by a minor population of Lgr5+ adult stem cell that commonly conserved in the bottom of crypts through GI tract. Maintenance of the stem cell is somehow regulated by epigenetic factors according to recent studies. Chromatin vacancy, remodelers, histone variants and histone modifiers could affect adult stem cell fate. In this study, Lgr5-EGFP reporter mouse was used to take advantage of exploring the epigenetic dynamics among Lgr5 positive mutual stem cell in GI tract. Cells were isolated by fluorescence-activated cell sorting (FACS), gene expression levels, chromatin accessibility changes and histone modifications were analyzed. Some notable chromatin structural related epigenetic variants were detected. To identify the overall cell-cell interaction inside the stem cell niche, an extensive genome-wide analysis should be also followed. According to the results, nevertheless, we expected a broader understanding of cellular niche maintaining stem cells and epigenetic barriers through conserved stem cell in GI tract. We expect that our study could provide more evidence of adult stem cell plasticity and more chances to understand each stem cell that takes parts in certain organs.Keywords: adult stem cell, epigenetics, LGR5 stem cell, gastrointestinal tract
Procedia PDF Downloads 223438 Functional Role of Tyr12 in the Catalytic Activity of Zeta-Like Glutathione S-Transferase from Acidovorax sp. KKS102
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Glutathione S-transferases (GSTs) are family of enzymes that function in the detoxification of variety of electrophilic substrates. In the present work, we report a novel zeta-like GST (designated as KKSG9) from the biphenyl/polychlorobiphenyl degrading organism Acidovorax sp. KKS102. KKSG9 possessed low sequence similarity but similar biochemical properties to zeta class GSTs. The gene for KKSG9 was cloned, purified and biochemically characterized. Functional analysis showed that the enzyme exhibits wider substrate specificity compared to most zeta class GSTs by reacting with 1-chloro-2,4-dinitrobenzene (CDNB), p-nitrobenzyl chloride (NBC), ethacrynic acid (EA), hydrogen peroxide, and cumene hydroperoxide (CuOOH). The enzyme also displayed dehalogenation function against dichloroacetate (a common substrate for zeta class GSTs) in addition to permethrin, and dieldrin. The functional role of Tyr12 was also investigated by site-directed mutagenesis. The mutant (Y12C) displayed low catalytic activity and dehalogenation function against all the substrates when compared with the wild type. Kinetic analysis using NBC and GSH as substrates showed that the mutant (Y12C) displayed a higher affinity for NBC when compared with the wild type, however, no significant change in GSH affinity was observed. These findings suggest that the presence of tyrosine residue in the motif might represent an evolutionary trend toward improving the catalytic activity of the enzyme. The enzyme as well could be useful in the bioremediation of various types of organochlorine pollutants.Keywords: Acidovorax sp. KKS102, bioremediation, glutathione s-transferase, site-directed mutagenesis, zeta
Procedia PDF Downloads 148437 Orientia Tsutsugamushi an Emerging Etiology of Acute Encephalitis Syndrome in Northern Part of India
Authors: Amita Jain, Shantanu Prakash, Suruchi Shukla
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Introduction: Acute encephalitis syndrome (AES) is a complex multi etiology syndrome posing a great public health problem in the northern part of India. Japanese encephalitis (JE) virus is an established etiology of AES in this region. Recently, Scrub typhus (ST) is being recognized as an emerging aetiology of AES in JE endemic belt. This study was conducted to establish the direct evidence of Central nervous system invasion by Orientia tsutsugamushi leading to AES. Methodology: A total of 849 cases with clinical diagnosis of AES were enrolled from six districts (Deoria and its adjoining area) of the traditional north Indian Japanese encephalitis (JE) belt. Serum and Cerebrospinal fluid samples were collected and tested for major agent causing acute encephalitis. AES cases either positive for anti-ST IgM antibodies or negative for all tested etiologies were investigated for ST-DNA by real-time PCR. Results: Of these 505 cases, 250 patients were laboratory confirmed for O. tsutsugamushi infection either by anti-ST IgM antibodies positivity (n=206) on serum sample or by ST-DNA detection by real-time PCR assay on CSF sample (n=2) or by both (n=42).Total 29 isolate could be sequenced for 56KDa gene. Conclusion: All the strains were found to cluster with Gilliam strains. The majority of the isolates showed a 97–99% sequence similarity with Thailand and Cambodian strains. Gilliam strain of O.tsusugamushi is an emerging as one of the major aetiologies leading to AES in northern part of India.Keywords: acute encephalitis syndrome, O. tsutsugamushi, Gilliam strain, North India, cerebrospinal fluid
Procedia PDF Downloads 241436 Structure, Bioinformatics Analysis and Substrate Specificity of a 6-Phospho-β-Glucosidase Glycoside Hydrolase 1 Enzyme from Bacillus licheniformis
Authors: Wayde Veldman, Ozlem T. Bishop, Igor Polikarpov
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In bacteria, mono and disaccharides are phosphorylated during uptake into the cell via the widely used phosphoenolpyruvate (PEP)-dependent phosphotransferase transport system. As an initial step in the phosphorylated disaccharide metabolism pathway, certain glycoside hydrolase family 1 (GH1) enzymes play a crucial role in releasing phosphorylated and non-phosphorylated monosaccharides. However, structural determinants for the specificity of these enzymes still need to be clarified. GH1 enzymes are known to have a wide array of functions. According to the CAZy database, there are twenty-one different enzymatic activities in the GH1 family. Here, the structure and substrate specificity of a GH1 enzyme from Bacillus licheniformis, hereafter known as BlBglH, was investigated. The sequence of the enzyme BlBglH was compared to the sequences of other characterized GH1 enzymes using sequence alignment, sequence identity calculations, phylogenetic analysis, and motif discovery. Through these various analyses, BlBglH was found to have sequence features characteristic of the 6-phospho-β-glucosidase activity enzymes. Additionally, motif and structure comparisons of the three most commonly studied GH1 enzyme-activities revealed a shared loop amongst the different structures that consist of different sequence motifs – this loop is thought to guide specific substrates (depending on activity) towards the active-site. To further affirm BlBglH enzyme activity, molecular docking and molecular dynamics simulations were performed. Docking was carried out using 6-phospho-β-glucosidase enzyme-activity positive (p-Nitrophenyl-beta-D-glucoside-6-phosphate) and negative (p-Nitrophenyl-beta-D-galactoside-6-phosphate) control ligands, followed by 400 ns molecular dynamics simulations. The positive-control ligand maintained favourable interactions within the active site until the end of the simulation. The negative-control ligand was observed exiting the enzyme at 287 ns. Binding free energy calculations showed that the positive-control complex had a substantially more favourable binding energy compared to the negative-control complex. Jointly, the findings of this study suggest that the BlBglH enzyme possesses 6-phospho-β-glucosidase enzymatic activity.Keywords: 6-P-β-glucosidase, glycoside hydrolase 1, molecular dynamics, sequence analysis, substrate specificity
Procedia PDF Downloads 126