World Academy of Science, Engineering and Technology
[Bioengineering and Life Sciences]
Online ISSN : 1307-6892
1347 Ameliorating Effects of Silver Nanoparticles Synthesized Using Chlorophytum borivillianum against Gamma Radiation Induced Oxidative Stress in Testis of Swiss Albino Mice
Authors: Ruchi Vyas, Sanjay Singh, Rashmi Sisodia
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Chlorophytum borivillianum root extract (CBE) was chosen as a reducing agent to fabricate silver nanoparticles with the aim of studying its radioprotective efficacy. The formation of synthesized nanoparticles was characterized by UV–visible analysis (UV–vis), Fourier transform infra-red (FT-IR), Transmission electron microscopy (TEM), Scanning electron microscope (SEM). TEM analysis showed particles size in the range of 20-30 nm. For this study, Swiss albino mice were selected from inbred colony and were divided into 4 groups: group I- control (irradiated-6 Gy), group II- normal (vehicle treated), group III- plant extract alone and group IV- CB-AgNPs (dose of 50 mg/kg body wt./day) administered orally for 7 consecutive days before irradiation to serve as experimental. CB-AgNPs pretreatment rendered significant increase in body weight and testes weight at various post irradiation intervals in comparison to irradiated group. Supplementation of CB-AgNPs reversed the adverse effects of gamma radiation on biochemical parameters as it notably ameliorated the elevation in lipid peroxidation and decline in glutathione concentration in testes. These observations indicate the radio-protective potential of CB-AgNPs in testicular constituents against gamma irradiation in mice.Keywords: Chlorophytum borivillianum, gamma radiation, radioprotective, silver nanoparticles
Procedia PDF Downloads 1481346 A Cross Culture Analysis of Medicinal Plants and Phytotherapies: Highly Effective for Gastropathic Disorders among Three Ethnic Communities of South West Pakistan
Authors: Sheikh Z. Ul Abidin, Raees Khan, Rainer W. Bussmann, Mushtaq Ahmad, Shayan Jamshed, Humera Jabeen, Ajmal Khan
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Gastropathic disorders are increasing rapidly and millions patients are reported every years across the world. Herbal medicines and traditional phytotherapies are very effective for many diseases including gastropathic ailments. Many communities and study region have their own unique remedies for such diseases. The current study was aimed to investigate and document high valued medicinal plants and folk remedies for different gastropathic disorders among the three ethnic groups of three regions in South West Pakistan. A total of 104 semi-structured interviews involving experts of traditional knowledge in 21 localities of the three regions (D.I. Khan, Zhob and Mianwali) were conducted. The interviews were especially focused on the documentation of folk herbal remedies. The collected data was analyzed using different quantitative methods. The highly effective plants from all localities were identified with the help of local interviewers and collected for proper taxonomic identification. A total of 56 medicinal plants and 33 effective recipes for 12 gastropathic diseases were documented from all the three ethnic groups in 21 localities. Fabaceae and Asteraceae were most prominently used for different gastropathic diseases. Diarrhea, vomiting and dysentery were the most commonly diseases treated with herbal remedies. It was observed that the three communities shared knowledge about the use of medicinal plants, 35 species were commonly reported from all three areas. However, each community had also their own unique uses of medicinal plants, e.g. 23 plants species were only used in Zhob, 20 plant species were only reported in D.I. Khan and 16 species in Mianwali. The present study reveals that different communities and ethnic groups share some traditional knowledge and also have their own unique knowledge of plants utilization. Gastropathic disorder is increasing very rapidly and the traditional cross-cultural knowledge of medicinal plants use can be very effective for its cure.Keywords: cross cultural, ethnic groups, gastropathy, phytotherapies, South West Pakistan
Procedia PDF Downloads 2951345 Phytogeography and Regional Conservation Status of Gymnosperms in Pakistan
Authors: Raees Khan, Mir A. Khan, Sheikh Z. Ul Abidin, Abdul S. Mumtaz
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In the present study, phytogeography and conservation status of gymnosperms of Pakistan were investigated. 44 gymnosperms species of 18 genera and 9 families were collected from 66 districts of the country. Among the 44 species, 20 species were native (wild) and 24 species were exotic (cultivated). Ephedra sarocarpa of Ephedraceae was not collected in this study from its distribution area and most probably it may be Nationally Extinct now from this area. Previously in Gymnosperms Flora of Pakistan 34 species was reported. 12 new gymnosperms species were recorded for the first time. Pinus wallichiana (40 districts), Cedrus deodara (39 districts) Pinus roxburghii (36 districts), Picea smithiana (36 districts) and Abies pindrow (34 districts) have the maximum ecological amplitude. Juniperus communis (17districts) and Juniperus excelsa (14 districts) were the widely distributed among the junipers. Ephedra foliata (23 districts), Ephedra gerardiana (20 districts) and Ephedra intermedia (19 districts) has the widest distribution range. Taxus fuana was also wider distribution range and recorded in 19 districts but its population was not very stable. These species was recorded to support local flora and fuana, especially endemics. PCORD version 5 clustered all gymnosperms species into 4 communities and all localities into 5 groups through cluster analyses. The Two Way Cluster Analyses of 66 districts (localities) resulted 4 various plant communities. The Gymnosperms in Pakistan are distributed in 3 floristic regions i.e. Western plains of the country, Northern and Western mountainous regions and Western Himalayas. The assessment of the National conservation status of these species, 10 species were found to be threatened, 6 species were endangered, 4 species were critically endangered and 1 species have become extinct (Ephedra sarcocarpa). The population of some species i.e. Taxus fuana, Ephedra gerardiana, Ephedra monosperma, Picea smithiana and Abies spectabilis is decreasing at an alarming rate.Keywords: conservation status, gymnosperms, phytogeography, Pakistan
Procedia PDF Downloads 2611344 Palyno-Morphological Characteristics of Gymnosperm Flora of Pakistan and Its Taxonomic Implications with Light Microscope and Scanning Electron Microscopy Methods
Authors: Raees Khan, Sheikh Z. Ul Abidin, Abdul S. Mumtaz, Jie Liu
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The present study is intended to assess gymnosperms pollen flora of Pakistan using Light Microscope (LM) and Scanning Electron Microscopy (SEM) for its taxonomic significance in identification of gymnosperms. Pollens of 35 gymnosperm species (12 genera and five families) were collected from its various distributional sites of gymnosperms in Pakistan. LM and SEM were used to investigate different palyno-morphological characteristics. Five pollen types (i.e., Inaperturate, Monolete, Monoporate, Vesiculate-bisaccate, and Polyplicate) were observed. In equatorial view seven types of pollens were observed, in which ten species were sub-angular, nine species were triangular, six species were perprolate, three species were rhomboidal, three species were semi-angular, two species were rectangular and two species were prolate. While five types of pollen were observed in polar view, in which ten species were spheroidal, nine species were angular, eight were interlobate, six species were circular, and two species were elliptic. Eighteen species have rugulate and 17 species has faveolate ornamentation. Eighteen species have verrucate and 17 have gemmate type sculpturing. The data was analysed through cluster analysis. The study showed that these palyno-morphological features have significance value in classification and identification of gymnosperms. Based on these different palyno-morphological features, a taxonomic key was proposed for the accurate and fast identifications of gymnosperms from Pakistan.Keywords: gymnosperms, palynology, Pakistan, taxonomy
Procedia PDF Downloads 2211343 A Brief History of Kampo Extract Formulations for Prescription in Japan
Authors: Kazunari Ozaki, Mitsuru Kageyama, Kenki Miyazawa, Yoshio Nakamura
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Background: Kampo (Japanese Traditional medicine) is a medicine traditionally practiced in Japan, based on ancient Chinese medicine. Most Kampo doctors have used decoction of crude drug pieces for treatment. 93% of the Kampo drugs sold in Japan are Kampo products nowadays. Of all Kampo products, 81% of them are Kampo extract formulations for prescription, which is prepared in powdered or granulated form from medicinal crude drug extracts mixed with appropriate excipient. Physicians with medical license for Western medicine prescribe these Kampo extract formulations for prescription in Japan. Objectives: Our study aims at presenting a brief history of Kampo extract formulations for prescription in Japan. Methods: Systematic searches for relevant studies were conducted using not only printed journals but also electronic journals from the bibliographic databases, such as PubMed/Medline, Ichushi-Web, and university/institutional websites, as well as search engines, such as Google and Google Scholar. Results: The first commercialization of Kampo extract formulations for general use (or OTC (over-the-counter) Kampo extract formulation) was achieved after 1957. The number of drugs has been subsequentially increased, reaching 148 Kampo extract formulation for prescription currently. Conclusion: We provide a history of Kampo extract formulations for prescription in Japan. The originality of this research is that it analyzes the background history of Kampo in parallel with relevant transitions in the government and insurance systems.Keywords: health insurance system, history, Kampo, Kampo extract formulation for prescription, OTC Kampo extract formulation, pattern corresponding prescription (Ho-sho-so-tai) system
Procedia PDF Downloads 2861342 Collagen/Hydroxyapatite Compositions Doped with Transitional Metals for Bone Tissue Engineering Applications
Authors: D. Ficai, A. Ficai, D. Gudovan, I. A. Gudovan, I. Ardelean, R. Trusca, E. Andronescu, V. Mitran, A. Cimpean
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In the last years, scientists struggled hardly to mimic bone structures to develop implants and biostructures which present higher biocompatibility and reduced rejection rate. One way to obtain this goal is to use similar materials as that of bone, namely collagen/hydroxyapatite composite materials. However, it is very important to tailor both compositions but also the microstructure of the bone that would ensure both the optimal osteointegartion and the mechanical properties required by the application. In this study, new collagen/hydroxyapatites composite materials doped with Cu, Li, Mn, Zn were successfully prepared. The synthesis method is described below: weight the Ca(OH)₂ mass, i.e., 7,3067g, and ZnCl₂ (0.134g), CuSO₄ (0.159g), LiCO₃ (0.133g), MnCl₂.4H₂O (0.1971g), and suspend in 100ml distilled water under magnetic stirring. The solution thus obtained is added a solution of NaH₂PO₄*H2O (8.247g dissolved in 50ml distilled water) under slow dropping of 1 ml/min followed by adjusting the pH to 9.5 with HCl and finally filter and wash until neutral pH. The as-obtained slurry was dried in the oven at 80°C and then calcined at 600°C in order to ensure a proper purification of the final product of organic phases, also inducing a proper sterilization of the mixture before insertion into the collagen matrix. The collagen/hydroxyapatite composite materials are tailored from morphological point of view to optimize their biocompatibility and bio-integration against mechanical properties whereas the addition of the dopants is aimed to improve the biological activity of the samples. The addition of transitional metals can improve the biocompatibility and especially the osteoblasts adhesion (Mn²⁺) or to induce slightly better osteoblast differentiation of the osteoblast, Zn²⁺ being a cofactor for many enzymes including those responsible for cell differentiation. If the amount is too high, the final material can become toxic and lose all of its biocompatibility. In order to achieve a good biocompatibility and not reach the cytotoxic effect, the amount of transitional metals added has to be maintained at low levels (0.5% molar). The amount of transitional metals entering into the elemental cell of HA will be verified using inductively-coupled plasma mass spectrometric system. This highly sensitive technique is necessary, because, at such low levels of transitional metals, the difference between biocompatible and cytotoxic is a very thin line, thus requiring proper and thorough investigation using a precise technique. In order to determine the structure and morphology of the obtained composite materials, IR spectroscopy, X-Ray diffraction (XRD), scanning electron microscopy (SEM), and Energy Dispersive X-Ray Spectrometry (EDS) were used. Acknowledgment: The present work was possible due to the EU-funding grant POSCCE-A2O2.2.1-2013-1, Project No. 638/12.03.2014, code SMIS-CSNR 48652. The financial contribution received from the national project “Biomimetic porous structures obtained by 3D printing developed for bone tissue engineering (BIOGRAFTPRINT), No. 127PED/2017 is also highly acknowledged.Keywords: collagen, composite materials, hydroxyapatite, bone tissue engineering
Procedia PDF Downloads 2061341 Combining in vitro Protein Expression with AlphaLISA Technology to Study Protein-Protein Interaction
Authors: Shayli Varasteh Moradi, Wayne A. Johnston, Dejan Gagoski, Kirill Alexandrov
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The demand for a rapid and more efficient technique to identify protein-protein interaction particularly in the areas of therapeutics and diagnostics development is growing. The method described here is a rapid in vitro protein-protein interaction analysis approach based on AlphaLISA technology combined with Leishmania tarentolae cell-free protein production (LTE) system. Cell-free protein synthesis allows the rapid production of recombinant proteins in a multiplexed format. Among available in vitro expression systems, LTE offers several advantages over other eukaryotic cell-free systems. It is based on a fast growing fermentable organism that is inexpensive in cultivation and lysate production. High integrity of proteins produced in this system and the ability to co-express multiple proteins makes it a desirable method for screening protein interactions. Following the translation of protein pairs in LTE system, the physical interaction between proteins of interests is analysed by AlphaLISA assay. The assay is performed using unpurified in vitro translation reaction and therefore can be readily multiplexed. This approach can be used in various research applications such as epitope mapping, antigen-antibody analysis and protein interaction network mapping. The intra-viral protein interaction network of Zika virus was studied using the developed technique. The viral proteins were co-expressed pair-wise in LTE and all possible interactions among viral proteins were tested using AlphaLISA. The assay resulted to the identification of 54 intra-viral protein-protein interactions from which 19 binary interactions were found to be novel. The presented technique provides a powerful tool for rapid analysis of protein-protein interaction with high sensitivity and throughput.Keywords: AlphaLISA technology, cell-free protein expression, epitope mapping, Leishmania tarentolae, protein-protein interaction
Procedia PDF Downloads 2371340 Comparative Production of Secondary Metabolites by Prunus africana (Hook. F.) Kalkman Provenances in Cameroon and Some Associated Endophytic Fungi
Authors: Gloria M. Ntuba-Jua, Afui M. Mih, Eneke E. T. Bechem
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Prunus africana (Hook. F.) Kalkman, commonly known as Pygeum or African cherry belongs to the Rosaceae family. It is a medium to large, evergreen tree with a spreading crown of 10 to 20 m. It is used by the traditional medical practitioners for the treatment of over 45ailments in Cameroon and sub-Sahara Africa. In modern medicine, it is used in the treatment of benign prostrate hyperplasia (BPH), prostate gland hypertrophy (enlarged prostate glands). This is possible because of its ability to produce some secondary metabolites which are believed to have bioactivity against these ailments. The ready international market for the sale of Prunus bark, uncontrolled exploitation, illegal harvesting using inappropriate techniques and poor timing of harvesting have contributed enormously to making the plant endangered. It is known to harbor a large number of endophytic fungi with the potential to produce similar secondary metabolites as the parent plant. Alternative sourcing of medicinal principles through endophytic fungi requires succinct knowledge of the endophytic fungi. This will serve as a conservation measure for Prunus africana by reducing dependence on Prunus bark for such metabolites. This work thus sought to compare the production of some major secondary metabolites produced by P. africana and some of its associated endophytic fungi. The leaves and stem bark of the plant from different provenances were soaked in methanol for 72 hrs to yield the methanolic crude extract. The phytochemical screening of the methanolic crude extracts using different standard procedures revealed the presence of tannins, flavonoids, terpenoids, saponins, phenolics and steroids. Pure cultures of some predominantly isolated endophyte species from the difference Prunus provenances such as Curvularia sp, and Morphospecies P001 were also grown in Potato Dextrose Broth (PDB) for 21 days and later extracted with Methylene dichloride (MDC) solvent after 24hrs to produce crude culture extracts. Qualitative assessment of crude culture extracts showed the presence of tannins, terpenoids, phenolics and steroids particularly β-Sitosterol, (a major bioactive metabolite) as did the plant tissues. Qualitative analysis by thin layer chromatography (TLC) was done to confirm and compare the production of β-Sitosterol (as marker compounds) in the crude extracts of the plant and endophyte. Samples were loaded on TLC silica gel aluminium barked plate (Kieselgel 60 F254, 0.2 mm, Merck) using acetone/hexane, (3.0:7.0) solvent system. They were visualized under an ultra violet lamp (UV254 and UV360). TLC revealed that leaves had a higher concentration of β-sitosterol in terms of band intensity than stem barks from the different provenances. The intensity of β-sitosterol bands in the culture extracts of endophytes was comparable to the plant extracts except for Curvularia sp (very minute) whose band was very faint. The ability of these fungi to make β-sitosterol was confirmed by TLC analysis with the compound having chromatographic properties (retention factor) similar to those of β-sitosterol standard. The ability of these major endophytes to produce secondary metabolites similar to the host has therefore been demonstrated. There is, therefore, the potential of developing the in vitro production system of Prunus secondary metabolites thereby enhancing its conservation.Keywords: Caneroon, endophytic fungi, Prunus africana, secondary metabolite
Procedia PDF Downloads 2311339 Mesoporous Titania Thin Films for Gentamicin Delivery and Bone Morphogenetic Protein-2 Immobilization
Authors: Ane Escobar, Paula Angelomé, Mihaela Delcea, Marek Grzelczak, Sergio Enrique Moya
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The antibacterial capacity of bone-anchoring implants can be improved by the use of antibiotics that can be delivered to the media after the surgery. Mesoporous films have shown great potential in drug delivery for orthopedic applications, since pore size and thickness can be tuned to produce different surface area and free volume inside the material. This work shows the synthesis of mesoporous titania films (MTF) by sol-gel chemistry and evaporation-induced self-assembly (EISA) on top of glass substrates. Pores with a diameter of 12nm were observed by Transmission Electron Microscopy (TEM). A film thickness of 100 nm was measured by Scanning Electron Microscopy (SEM). Gentamicin was used to study the antibiotic delivery from the film by means of High-performance liquid chromatography (HPLC). The Staphilococcus aureus strand was used to evaluate the effectiveness of the penicillin loaded films toward inhibiting bacterial colonization. MC3T3-E1 pre-osteoblast cell proliferation experiments proved that MTFs have a good biocompatibility and are a suitable surface for MC3T3-E1 cell proliferation. Moreover, images taken by Confocal Fluorescence Microscopy using labeled vinculin, showed good adhesion of the MC3T3-E1 cells to the MTFs, as well as complex actin filaments arrangement. In order to improve cell proliferation Bone Morphogenetic Protein-2 (BMP-2) was adsorbed on top of the mesoporous film. The deposition of the protein was proved by measurements in the contact angle, showing an increment in the hydrophobicity while the protein concentration is higher. By measuring the dehydrogenase activity in MC3T3-E1 cells cultured in dually functionalized mesoporous titatina films with gentamicin and BMP-2 is possible to find an improvement in cell proliferation. For this purpose, the absorption of a yellow-color formazan dye, product of a water-soluble salt (WST-8) reduction by the dehydrogenases, is measured. In summary, this study proves that by means of the surface modification of MTFs with proteins and loading of gentamicin is possible to achieve an antibacterial effect and a cell growth improvement.Keywords: antibacterial, biocompatibility, bone morphogenetic protein-2, cell proliferation, gentamicin, implants, mesoporous titania films, osteoblasts
Procedia PDF Downloads 1631338 Evaluation of Two DNA Vaccine Constructs in Labeo rohita against Edwardsiella tarda
Authors: Ranjeeta Kumari, Makesh M, Gayatri Tripathi, K V Rajendran, Megha Bedekar
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A comparative study on DNA immunization with recombinant glyceraldehyde-3-phosphate dehydrogenase (GAPDH) construct of Edwardsiella tarda (pGPD group) and a bicistronic construct expressing GAPDH plus IFN-γ of Labeo rohita as adjuvant (pGPD+IFN group) was undertaken in Labeo rohita along with the control animals. Successful co-expression of two genes that is GAPDH and IFN-γ was confirmed in SSN-1 cells line by RT-qPCR and western blot. The protective immune response of host to DNA vaccine construct was determined by RPS and specific antibody production. Fishes immunized with plasmids via intramuscular injection (I/M) exhibited a considerable relative percentage survivability of 66.66% in pGPD+IFN immunized group and 53.34% in pGPD immunized group after challenge with E. tarda. Antibody response was also significantly high in pGPD+IFN group at all time points under study. This was analysed by competitive ELISA, using anti GAPDH monoclonal antibodies. The experiment revealed that the GAPDH gene of E. tarda is one of the ideal candidates for generating protective immune response in L. rohita. Further addition of Interferon gamma to DNA vaccine construct can enhance the immune response in host.Keywords: DNA vaccine, Edwardsiella tarda, Labeo rohita, zoonosis, immune response
Procedia PDF Downloads 2031337 Impact of Mixed Prey Population on Predation Potential and Food Preference of a Predaceous Ladybird, Coccinella septempunctata
Authors: Ahmad Pervez
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We investigated predation potential and food preference of different life stages of a predaceous ladybird Coccinella septempunctata L. (Coleptera: Coccinellidae) using a nutritive food (mustard aphid, Lipaphis erysimi) and a toxic food (cabbage aphid, Brevicoryne brassicae). We gave monotypic prey, L. erysimi, then B. brassicae to all life stages and found that second, third and fourth instars and adult female C. septempunctata daily consumed greater number of former prey. However, the first instar and the adult male equally consumed both the prey. In choice condition, each larva, adult male and female consumed mixed aphid diet separately in three proportions (i.e. low: high, equal: equal and high: low densities of L. erysimi: B. brassicae). We hypothesized that life stages of C. septempunctata will prefer L. erysimi regardless of its proportions. Laboratory experiment supported this hypothesis only at the adult level showing high values of β and C preference indices. However, it rejects this hypothesis at the larval level, as larvae preferred B. brassicae in certain combinations and showed no preference in a few combinations. We infer that mixing of nutritive diet in a toxic diet may possibly overcome the probable nutritive deficiency and/or reduces the toxicity of toxic diet, especially to the larvae of C. septempunctata. Consumption of high proportion of B. brassicae mixed with fewer L. erysimi suggests that mixed diet could be better for the development of immature stages of C. septempunctata.Keywords: Coccinella septempunctata, predatory potential, prey preference, Lipaphis erysimi, Brevicoryne brassicae
Procedia PDF Downloads 1961336 Analysis of Replication Protein A (RPA): The Role of Homolog Interaction and Recombination during Meiosis
Authors: Jeong Hwan Joo, Keun Pil Kim
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During meiosis, meiotic recombination is initiated by Spo11-mediated DSB formation and exonuclease-mediated DSB resection occurs to expose single stranded DNA formation. RPA is further required to inhibit secondary structure formation of ssDNA that can be formed Watson-Crick pairing. Rad51-Dmc1, RecA homologs in eukaryote and their accessory factors involve in searching homolog templates to mediate strand exchange. In this study, we investigate the recombinational roles of replication protein A (RPA), which is heterotrimeric protein that is composed of RPA1, RPA2, and RPA3. Here, we investigated meiotic recombination using DNA physical analysis at the HIS4LEU2 hot spot. In rfa1-119 (K45E, N316S) cells, crossover (CO) and non-crossover (NCO) products reduced than WT. rfa1-119 delayed in single end invasion-to-double holiday junction (SEI-to-dHJ) transition and exhibits a defect in second-end capture that is also modulated by Rad52. In the further experiment, we observed that in rfa1-119 mutant, RPA could not be released in timely manner. Furthermore, rfa1-119 exhibits failure in the second end capture, implying reduction of COs and NCOs. In this talk, we will discuss more detail how RPA involves in chromatin axis association via formation of axis-bridge and why RPA is required for Rad52-mediated second-end capture progression.Keywords: homolog interaction, meiotic recombination, replication protein A, RPA1
Procedia PDF Downloads 2011335 Applying Massively Parallel Sequencing to Forensic Soil Bacterial Profiling
Authors: Hui Li, Xueying Zhao, Ke Ma, Yu Cao, Fan Yang, Qingwen Xu, Wenbin Liu
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Soil can often link a person or item to a crime scene, which makes it a valuable evidence in forensic casework. Several techniques have been utilized in forensic soil discrimination in previous studies. Because soil contains a vast number of microbiomes, the analyse of soil microbiomes is expected to be a potential way to characterise soil evidence. In this study, we applied massively parallel sequencing (MPS) to soil bacterial profiling on the Ion Torrent Personal Genome Machine (PGM). Soils from different regions were collected repeatedly. V-region 3 and 4 of Bacterial 16S rRNA gene were detected by MPS. Operational taxonomic units (OTU, 97%) were used to analyse soil bacteria. Several bioinformatics methods (PCoA, NMDS, Metastats, LEfse, and Heatmap) were applied in bacterial profiles. Our results demonstrate that MPS can provide a more detailed picture of the soil microbiomes and the composition of soil bacterial components from different region was individualistic. In conclusion, the utility of soil bacterial profiling via MPS of the 16S rRNA gene has potential value in characterising soil evidences and associating them with their place of origin, which can play an important role in forensic science in the future.Keywords: bacterial profiling, forensic, massively parallel sequencing, soil evidence
Procedia PDF Downloads 5641334 Performance of a Lytic Bacteriophage Cocktail against Pseudomonas aeruginosa in Conditions That Simulate the Cystic Fibrosis Lung Environment
Authors: Isaac Martin, Abigail Lark, Sandra Morales, Eric W. Alton, Jane C. Davies
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Objectives: The cystic fibrosis (CF) lung is a unique microbiological niche, wherein harmful bacteria persist for many years despite antibiotic therapy. Pseudomonas aeruginosa (Pa), the major culprit leading to lung decline and increased mortality, thrives in the lungs of patients with CF due to several factors that have been linked with poor antibiotic performance. Our group is investigating alternative therapies including bacteriophage cocktails with which we have previously demonstrated efficacy against planktonic organisms. In this study, we explored the effects of a 4-phage cocktail on Pa grown in two different conditions, intended to mirror the CF lung: a) alongside standard antibiotic treatment in pre-formed biofilms (structures formed by Pa-secreted exopolysaccharides which provide both physical and cell division barriers to antimicrobials and host defenses and b) in an acidic environment postulated to be present in the CF airway due both to the primary defect in bicarbonate secretion and secondary effects of inflammation. Methods: 16 Pa strains from CF patients at the Royal Brompton Hospital were selected based on sensitivity to a) ceftazidime/ tobramycin and b) the phage cocktail in a conventional plaque assay. To assess efficacy of phage in biofilms, 96 well plates with Pa (5x10⁷ CFU/ ml) were incubated in static conditions, allowing adherent bacterial colonies to form for 24 hr. Ceftazidime and tobramycin (both at 2 × MIC) were added, +/- bacteriophage (4x10⁸ PFU/mL) for a further 24 hr. Cell viability and biomass were estimated using fluorescent resazurin and crystal violet assays, respectively. To evaluate the effect of pH, strains were grown planktonically in shaking 96 well plates at pH 6.0, 6.6, 7.0 and 7.5 with tobramycin or phage, at varying concentrations. Cell viability was quantified by fluorescent resazurin assay. Results: For the biofilm assay, treatment groups were compared with untreated controls and expressed as percent reduction in cell viability and biomass. Addition of the 4-phage cocktail resulted in a 1.3-fold reduction in cell viability and 1.7-fold reduction in biomass (p < 0.001) when compared to standard antibiotic treatment alone. Notably, there was a 50 ± 15% reduction in cell viability and 60 ± 12% reduction in biomass (95% CI) for the 4 biofilms demonstrating the most resistance to antibiotic treatment. 83% of strains tested (n=6) showed decreased bacterial killing by tobramycin at acidic pHs (p < 0.01). However, 25% of strains (n=12) showed improved phage killing at acidic pHs (p < 0.05), with none showing the pattern of reduced efficacy at acidic pH demonstrated by tobramycin. Conclusion: The 4-phage anti-Pa cocktail tested against Pa performs well in pre-formed biofilms and in acidic environments; two conditions intended to mimic the CF lung. To our knowledge, these are the first data looking at the effects of subtle pH changes on phage-mediated bacterial killing in the context of Pa infection. These findings contribute to a growing body of evidence supporting the use of nebulised lytic bacteriophage as a treatment in the context of lung infection.Keywords: biofilm, cystic fibrosis, pH, Pseudomonas aeruginosa, lytic bacteriophage
Procedia PDF Downloads 1731333 Molecular Defects Underlying Genital Ambiguity in Egyptian Patients: A Systematic Review
Authors: Y. Z. Gad
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Disorders of Sex Development (DSD) are defined as congenital conditions in which development of chromosomal, gonadal or anatomical sex is atypical. The DSD are relatively prevalent in Egypt. In spite of that, the relative rarity of the individual disease types or their molecular pathologies frequently resulted in reporting on single or few cases. This augmented the challenging nature of phenotype-genotype correlation in this disease group and its utilization in the management of such medical emergency. Through critical assessment of the published DSD reports, the current review aims at analyzing the clinical characteristics of the various DSD forms in relation to the underlying molecular pathologies. A systematic literature search was done in Pubmed, using relevant keywords (Egypt versus DSD, genital ambiguity or ambiguous genitalia, the old terms of 'intersex, hermaphroditism and pseudohermaphroditism', and a list of the DSD entities and their related genes). The search yielded 24 reports of molecular data in Egyptian patients presenting with ambiguous genitalia. However, only 21 publications fulfilled the criteria of inclusion of detailed clinical descriptions and definitive molecular diagnoses of individual patients. Curation of the data yielded a total of 53 cases that were ascertained from 40 families. Fifty-one patients present with ambiguous genitalia only while 2 had multiple congenital anomalies. Parental consanguinity was noted in 60% of cases. Sex of rearing at initial presentation was female in 75% and 60% in 46,XY and 46,XX DSD cases, respectively. The external genital phenotype in 2/3 of the 46,XY DSD cases showed moderate undermasculinization [Quigley scores 3 & 4] and 1/3 had severe presentations [scores 5 & 6]. For 46,XX subjects, 1 had severe virilization of the external genitalia while 8 had moderate phenotype. Hormonal data were inconclusive or contradictory to final diagnosis in a forth of cases. Collectively, 31 families [31/40, 77.5%] with 46,XY DSD had molecular defects in the genes, 5 alpha reductase 2 (SRD5A2) [12/31], 17 beta-hydroxysteroid dehydrogenase 3 [8/31], androgen receptor [7/31], Steroidogenic factor 1 [2/31], luteinizing hormone receptor [1/31], and fibroblast growth factor receptor 1 [1/31]. In a multiethnic study, 9 families afflicted with 46,XX DSD due to 11 beta hydroxylase (CYP11B1) deficiency were documented. Two recurrent mutations, G34R and N160D, in SRD5A2 were present, respectively, in 42 and 17% of cases. Similarly, 4 recurrent mutations resulted in 89% of the CYP11B1 presentations. In conclusion, this analysis highlights the importance of autosomal recessive inheritance and inbreeding among DSD presentations, the importance of founder effect in at least 2 disorders, the difficulties in relating the genotype with the indeterminate genital phenotype, the under-reporting of some DSD subtypes, and the notion that the reported mutational profiles among Egyptian DSD cases are relatively different from those reported in other ethnic groups.Keywords: disorders of sex development, genital ambiguity, mutation, molecular diagnosis, Egypt
Procedia PDF Downloads 1371332 A Review on Biological Control of Mosquito Vectors
Authors: Asim Abbasi, Muhammad Sufyan, Iqra, Hafiza Javaria Ashraf
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The share of vector-borne diseases (VBDs) in the global burden of infectious diseases is almost 17%. The advent of new drugs and latest research in medical science helped mankind to compete with these lethal diseases but still diseases transmitted by different mosquito species, including filariasis, malaria, viral encephalitis and dengue are serious threats for people living in disease endemic areas. Injudicious and repeated use of pesticides posed selection pressure on mosquitoes leading to development of resistance. Hence biological control agents are under serious consideration of scientific community to be used in vector control programmes. Fish have a history of predating immature stages of different aquatic insects including mosquitoes. The noteworthy examples in Africa and Asia includes, Aphanius discolour and a fish in the Panchax group. Moreover, common mosquito fish, Gambusia affinis predates mostly on temporary water mosquitoes like anopheline as compared to permanent water breeders like culicines. Mosquitoes belonging to genus Toxorhynchites have a worldwide distribution and are mostly associated with the predation of other mosquito larvae habituating with them in natural and artificial water containers. These species are harmless to humans as their adults do not suck human blood but feeds on floral nectar. However, their activity is mostly temperature dependent as Toxorhynchites brevipalpis consume 359 Aedes aegypti larvae at 30-32 ºC in contrast to 154 larvae at 20-26 ºC. Although many bacterial species were isolated from mosquito cadavers but those belonging to genus Bacillus are found highly pathogenic against them. The successful species of this genus include Bacillus thuringiensis and Bacillus sphaericus. The prime targets of B. thuringiensis are mostly the immatures of genus Aedes, Culex, Anopheles and Psorophora while B. sphaericus is specifically toxic against species of Culex, Psorophora and Culiseta. The entomopathogenic nematodes belonging to family, mermithidae are also pathogenic to different mosquito species. Eighty different species of mosquitoes including Anopheles, Aedes and Culex proved to be highly vulnerable to the attack of two mermithid species, Romanomermis culicivorax and R. iyengari. Cytoplasmic polyhedrosis virus was the first described pathogenic virus, isolated from the cadavers of mosquito specie, Culex tarsalis. Other viruses which are pathogenic to culicine includes, iridoviruses, cytopolyhedrosis viruses, entomopoxviruses and parvoviruses. Protozoa species belonging to division microsporidia are the common pathogenic protozoans in mosquito populations which kill their host by the chronic effects of parasitism. Moreover, due to their wide prevalence in anopheline mosquitoes and transversal and horizontal transmission from infected to healthy host, microsporidia of the genera Nosema and Amblyospora have received much attention in various mosquito control programmes. Fungal based mycopesticides are used in biological control of insect pests with 47 species reported virulent against different stages of mosquitoes. These include both aquatic fungi i.e. species of Coelomomyces, Lagenidium giganteum and Culicinomyces clavosporus, and the terrestrial fungi Metarhizium anisopliae and Beauveria bassiana. Hence, it was concluded that the integrated use of all these biological control agents can be a healthy contribution in mosquito control programmes and become a dire need of the time to avoid repeated use of pesticides.Keywords: entomopathogenic nematodes, protozoa, Toxorhynchites, vector-borne
Procedia PDF Downloads 2671331 Real Time Activity Recognition Framework for Health Monitoring Support in Home Environments
Authors: Shaikh Farhad Hossain, Liakot Ali
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Technology advances accelerate the quality and type of services provided for health care and especially for monitoring health conditions. Sensors have turned out to be more effective to detect diverse physiological signs and can be worn on the human body utilizing remote correspondence modules. An assortment of programming devices have been created to help in preparing a difference rundown of essential signs by examining and envisioning information produced by different sensors. In this proposition, we presented a Health signs and Activity acknowledgment monitoring system. Utilizing off-the-rack sensors, we executed a movement location system for identifying five sorts of action: falling, lying down, sitting, standing, and walking. The framework collects and analyzes sensory data in real-time, and provides different feedback to the users. In addition, it can generate alerts based on the detected events and store the data collected to a medical server.Keywords: ADL, SVM, TRIL , MEMS
Procedia PDF Downloads 3981330 Varietal Screening of Watermelon against Powdery Mildew Disease and Its Management
Authors: Asim Abbasi, Amer Habib, Sajid Hussain, Muhammad Sufyan, Iqra, Hasnain Sajjad
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Except for few scattered cases, powdery mildew disease was not a big problem for watermelon in the past but with the outbreaks of its pathotypes, races 1W and 2W, this disease becomes a serious issue all around the globe. The severe outbreak of this disease also increased the rate of fungicide application for its proper management. Twelve varieties of watermelon were screened in Research Area of Department of Plant pathology, University of Agriculture, Faisalabad to check the incidence of powdery mildew disease. Disease inoculum was prepared and applied with the help of foliar spray method. Fungicides and plants extracts were also applied after the disease incidence. Percentage leaf surface area diseased was assessed visually with a modified Horsfall-Barratt scale. The results of the experiment revealed that among all varieties, WT2257 and Zcugma F1 were highly resistant showing less than 5% disease incidence while Anar Kali and Sugar baby were highly susceptible with disease incidence of more than 65%. Among botanicals neem extract gave best results with disease incidence of less than 20%. Besides neem, all other botanicals also gave significant control of powdery mildew disease than the untreated check. In case of fungicides, Gemstar showed least disease incidence i.e. < 10%, however besides control maximum disease incidence was observed in Curzate (> 30%).Keywords: botanicals, fungicides, pathotypes, powdery mildew
Procedia PDF Downloads 2971329 In vitro Modeling of Aniridia-Related Keratopathy by the Use of Crispr/Cas9 on Limbal Epithelial Cells and Rescue
Authors: Daniel Aberdam
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Haploinsufficiency of PAX6 in humans is the main cause of congenital aniridia, a rare eye disease characterized by reduced visual acuity. Patients have also progressive disorders including cataract, glaucoma and corneal abnormalities making their condition very challenging to manage. Aniridia-related keratopathy (ARK), caused by a combination of factors including limbal stem-cell deficiency, impaired healing response, abnormal differentiation, and infiltration of conjunctival cells onto the corneal surface, affects up to 95% of patients. It usually begins in the first decade of life resulting in recurrent corneal erosions, sub-epithelial fibrosis with corneal decompensation and opacification. Unfortunately, current treatment options for aniridia patients are currently limited. Although animal models partially recapitulate this disease, there is no in vitro cellular model of AKT needed for drug/therapeutic tools screening and validation. We used genome editing (CRISPR/Cas9 technology) to introduce a nonsense mutation found in patients into one allele of the PAX6 gene into limbal stem cells. Resulting mutated clones, expressing half of the amount of PAX6 protein and thus representative of haploinsufficiency were further characterized. Sequencing analysis showed that no off-target mutations were induced. The mutated cells displayed reduced cell proliferation and cell migration but enhanced cell adhesion. Known PAX6 targets expression was also reduced. Remarkably, addition of soluble recombinant PAX6 protein into the culture medium was sufficient to activate endogenous PAX6 gene and, as a consequence, rescue the phenotype. It strongly suggests that our in vitro model recapitulates well the epithelial defect and becomes a powerful tool to identify drugs that could rescue the corneal defect in patients. Furthermore, we demonstrate that the homeotic transcription factor Pax6 is able to be uptake naturally by recipient cells to function into the nucleus.Keywords: Pax6, crispr/cas9, limbal stem cells, aniridia, gene therapy
Procedia PDF Downloads 2071328 Ectoine: A Compatible Solute in Radio-Halophilic Stenotrophomonas sp. WMA-LM19 Strain to Prevent Ultraviolet-Induced Protein Damage
Authors: Wasim Sajjad, Manzoor Ahmad, Sundas Qadir, Muhammad Rafiq, Fariha Hasan, Richard Tehan, Kerry L. McPhail, Aamer Ali Shah
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Aim: This study aims to investigate the possible radiation protective role of a compatible solute in the tolerance of radio-halophilic bacterium against stresses, like desiccation and exposure to ionizing radiation. Methods and Results: Nine different radio-resistant bacteria were isolated from desert soil, where strain WMA-LM19 was chosen for detailed studies on the basis of its high tolerance for ultraviolet radiation among all these isolates. 16S rRNA gene sequencing indicated that the bacterium was closely related to Stenotrophomonas sp. (KT008383). A bacterial milking strategy was applied for extraction of intracellular compatible solutes in 70% (v/v) ethanol, which were purified by high-performance liquid chromatography (HPLC). The compound was characterized as ectoine by 1H and 13C nuclear magnetic resonance (NMR), and mass spectrometry (MS). Ectoine demonstrated more efficient preventive activity (54.80%) to erythrocyte membranes and also inhibited oxidative damage to proteins and lipids in comparison to the standard ascorbic acid. Furthermore, a high level of ectoine-mediated protection of bovine serum albumin against ionizing radiation (1500-2000 Jm-2) was observed, as indicated by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) analysis. Conclusion: The results indicated that ectoine can be used as a potential mitigator and radio-protective agent to overcome radiation- and salinity-mediated oxidative damage in extreme environments. Significance and Impact of the Study: This study shows that ectoine from radio-halophiles can be used as a potential source in topical creams as sunscreen. The investigation of ectoine as UV protectant also changes the prospective that radiation resistance is specific only to molecular adaptation.Keywords: ectoine, anti-oxidant, stenotrophomonas sp., ultraviolet radiation
Procedia PDF Downloads 2091327 Comparative Demography of Lady Beetle, Coccinella septempunctata Linnaeus (Coleoptera: Coccinellidae) with Respect to Different Aphid Species
Authors: Muhammad Farooq, Muhammad R. Shahid, M. Shakeel, A. Iftikhar, M. Sagheer, Riaz A. Kainth
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Comparative demography of Coccinella septempunctata Linnaeus (Coleoptera: Coccinellidae) was studied with respect to four host aphid species viz; Rhopalosiphum padi, Rhopalosiphum maidis, Sitobion avenae, and Shizaphis graminum under laboratory conditions using Two-sex Age-stage life table instead of traditional age specific life table which considers only female. Results revealed that developmental period from egg to adult of C. septempunctata were shorter on R. padi (16.49 days) whereas longer on R. maidis (22.83 days). Net reproductive rate varied from 110.01 offspring on R. maidis to 288.78 offspring on R. padi. Mean generation time (T) ranged from 29.02 d on R. padi to 39.788 d on R. maidis. Highest to lowest values of intrinsic rate of increase (rm) were recorded on R. padi, S. graminum, S. avenae, and R. maidis (0.194, 0.143, 0.140 and 0.117 d⁻¹, respectively). Highest finite rate of increase was observed on R. padi (1.214 d⁻¹) followed by S. graminum (1.154 d⁻¹) whereas lowest values were obtained on R. maidis and S. avenae (1.124 and 1.150 d⁻¹, respectively). In this study, the data on the life table of both predator and prey provide useful information in the mass rearing and practical application of a natural agent to biological control systems.Keywords: C. septempunctata, two-sex age-stage life table, population parameters, aphid species
Procedia PDF Downloads 1981326 Identification of Microbial Community in an Anaerobic Reactor Treating Brewery Wastewater
Authors: Abimbola M. Enitan, John O. Odiyo, Feroz M. Swalaha
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The study of microbial ecology and their function in anaerobic digestion processes are essential to control the biological processes. This is to know the symbiotic relationship between the microorganisms that are involved in the conversion of complex organic matter in the industrial wastewater to simple molecules. In this study, diversity and quantity of bacterial community in the granular sludge taken from the different compartments of a full-scale upflow anaerobic sludge blanket (UASB) reactor treating brewery wastewater was investigated using polymerase chain reaction (PCR) and real-time quantitative PCR (qPCR). The phylogenetic analysis showed three major eubacteria phyla that belong to Proteobacteria, Firmicutes and Chloroflexi in the full-scale UASB reactor, with different groups populating different compartment. The result of qPCR assay showed high amount of eubacteria with increase in concentration along the reactor’s compartment. This study extends our understanding on the diverse, topological distribution and shifts in concentration of microbial communities in the different compartments of a full-scale UASB reactor treating brewery wastewater. The colonization and the trophic interactions among these microbial populations in reducing and transforming complex organic matter within the UASB reactors were established.Keywords: bacteria, brewery wastewater, real-time quantitative PCR, UASB reactor
Procedia PDF Downloads 2601325 Glycan Analyzer: Software to Annotate Glycan Structures from Exoglycosidase Experiments
Authors: Ian Walsh, Terry Nguyen-Khuong, Christopher H. Taron, Pauline M. Rudd
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Glycoproteins and their covalently bonded glycans play critical roles in the immune system, cell communication, disease and disease prognosis. Ultra performance liquid chromatography (UPLC) coupled with mass spectrometry is conventionally used to qualitatively and quantitatively characterise glycan structures in a given sample. Exoglycosidases are enzymes that catalyze sequential removal of monosaccharides from the non-reducing end of glycans. They naturally have specificity for a particular type of sugar, its stereochemistry (α or β anomer) and its position of attachment to an adjacent sugar on the glycan. Thus, monitoring the peak movements (both in the UPLC and MS1) after application of exoglycosidases provides a unique and effective way to annotate sugars with high detail - i.e. differentiating positional and linkage isomers. Manual annotation of an exoglycosidase experiment is difficult and time consuming. As such, with increasing sample complexity and the number of exoglycosidases, the analysis could result in manually interpreting hundreds of peak movements. Recently, we have implemented pattern recognition software for automated interpretation of UPLC-MS1 exoglycosidase digestions. In this work, we explain the software, indicate how much time it will save and provide example usage showing the annotation of positional and linkage isomers in Immunoglobulin G, apolipoprotein J, and simple glycan standards.Keywords: bioinformatics, automated glycan assignment, liquid chromatography, mass spectrometry
Procedia PDF Downloads 2001324 Effect of Silicon on Tritrophic Interaction of Cotton, Whitefly and Chrysoperla carnea
Authors: Asim Abbasi, Muhammad Sufyan
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The present experiment was carried out to examine the effects of silicon dioxide on tritrophic interaction of cotton, whitefly, and the predator Chrysoperla carnea. Population of whitefly was maintained on silicon treated and non-treated cotton for two generations in greenhouse net cages exposed to outside temperature and luminosity. The cotton was treated with silicon dioxide twice after 15 days intervals with 200 ppm concentration. A stock rearing of the natural predator was developed in the laboratory conditions. In the bioassay eggs of the predator all at the same age were individualized in glass petri plates that will be pierced with a pin to allow aeration and maintained in an incubator at 28 ± 2°C, 70 ± 10% relative humidity and 12h photo phase. Population of whitefly stayed on silicon treated, and non-treated cotton were offered to newly hatched chrysopid larvae until the end of the larval stage, assuring a permanent supply. Feeding preference of C. carnea along with longevity, survival of each instar larvae, pupation, adult emergence, and fecundity was checked. The results revealed that there was no significant difference in the feeding preference of C. carnea among both treatments. Durations of 1st and 2nd larval instar were also at par in both treatments. However overall longevity and adult emergence were a bit lower in silicon treated whitefly treatment. This may be due to the fact that silicon reduces the nutritional quality of host because of reduced whitefly feeding on silicon treated cotton. No significant difference in 1st and 2nd larval instars and then increased larval duration in later instars suggested that the effect of silicon treated host should be checked on more than 1 generation of C. carnea to get better findings.Keywords: Chrysoperla carnea, silicon, tritrophic, whitefly
Procedia PDF Downloads 1801323 Digitization and Morphometric Characterization of Botanical Collection of Indian Arid Zones as Informatics Initiatives Addressing Conservation Issues in Climate Change Scenario
Authors: Dipankar Saha, J. P. Singh, C. B. Pandey
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Indian Thar desert being the seventh largest in the world is the main hot sand desert occupies nearly 385,000km2 and about 9% of the area of the country harbours several species likely the flora of 682 species (63 introduced species) belonging to 352 genera and 87 families. The degree of endemism of plant species in the Thar desert is 6.4 percent, which is relatively higher than the degree of endemism in the Sahara desert which is very significant for the conservationist to envisage. The advent and development of computer technology for digitization and data base management coupled with the rapidly increasing importance of biodiversity conservation resulted in the invention of biodiversity informatics as discipline of basic sciences with multiple applications. Aichi Target 19 as an outcome of Convention of Biological Diversity (CBD) specifically mandates the development of an advanced and shared biodiversity knowledge base. Information on species distributions in space is the crux of effective management of biodiversity in the rapidly changing world. The efficiency of biodiversity management is being increased rapidly by various stakeholders like researchers, policymakers, and funding agencies with the knowledge and application of biodiversity informatics. Herbarium specimens being a vital repository for biodiversity conservation especially in climate change scenario the digitization process usually aims to improve access and to preserve delicate specimens and in doing so creating large sets of images as a part of the existing repository as arid plant information facility for long-term future usage. As the leaf characters are important for describing taxa and distinguishing between them and they can be measured from herbarium specimens as well. As a part of this activity, laminar characterization (leaves being the most important characters in assessing climate change impact) initially resulted in classification of more than thousands collections belonging to ten families like Acanthaceae, Aizoaceae, Amaranthaceae, Asclepiadaceae, Anacardeaceae, Apocynaceae, Asteraceae, Aristolochiaceae, Berseraceae and Bignoniaceae etc. Taxonomic diversity indices has also been worked out being one of the important domain of biodiversity informatics approaches. The digitization process also encompasses workflows which incorporate automated systems to enable us to expand and speed up the digitisation process. The digitisation workflows used to be on a modular system which has the potential to be scaled up. As they are being developed with a geo-referencing tool and additional quality control elements and finally placing specimen images and data into a fully searchable, web-accessible database. Our effort in this paper is to elucidate the role of BIs, present effort of database development of the existing botanical collection of institute repository. This effort is expected to be considered as a part of various global initiatives having an effective biodiversity information facility. This will enable access to plant biodiversity data that are fit-for-use by scientists and decision makers working on biodiversity conservation and sustainable development in the region and iso-climatic situation of the world.Keywords: biodiversity informatics, climate change, digitization, herbarium, laminar characters, web accessible interface
Procedia PDF Downloads 2291322 Detection, Isolation, and Raman Spectroscopic Characterization of Acute and Chronic Staphylococcus aureus Infection in an Endothelial Cell Culture Model
Authors: Astrid Tannert, Anuradha Ramoji, Christina Ebert, Frederike Gladigau, Lorena Tuchscherr, Jürgen Popp, Ute Neugebauer
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Staphylococcus aureus is a facultative intracellular pathogen, which by entering host cells may evade immunologic host response as well as antimicrobial treatment. In that way, S. aureus can cause persistent intracellular infections which are difficult to treat. Depending on the strain, S. aureus may persist at different intracellular locations like the phagolysosome. The first barrier invading pathogens from the blood stream that they have to cross are the endothelial cells lining the inner surface of blood and lymphatic vessels. Upon proceeding from an acute to a chronic infection, intracellular pathogens undergo certain biochemical and structural changes including a deceleration of metabolic processes to adopt for long-term intracellular survival and the development of a special phenotype designated as small colony variant. In this study, the endothelial cell line Ea.hy 926 was used as a model for acute and chronic S. aureus infection. To this end, Ea.hy 926 cells were cultured on QIAscout™ Microraft Arrays, a special graded cell culture substrate that contains around 12,000 microrafts of 200 µm edge length. After attachment to the substrate, the endothelial cells were infected with GFP-expressing S. aureus for 3 weeks. The acute infection and the development of persistent bacteria was followed by confocal laser scanning microscopy, scanning the whole Microraft Array for the presence and for detailed determination of the intracellular location of fluorescent intracellular bacteria every second day. After three weeks of infection representative microrafts containing infected cells, cells with protruded infections and cells that did never show any infection were isolated and fixed for Raman micro-spectroscopic investigation. For comparison, also microrafts with acute infection were isolated. The acquired Raman spectra are correlated with the fluorescence microscopic images to give hints about a) the molecular alterations in endothelial cells during acute and chronic infection compared to non-infected cells, and b) metabolic and structural changes within the pathogen when entering a mode of persistence within host cells. We thank Dr. Ruth Kläver from QIAGEN GmbH for her support regarding QIAscout technology. Financial support by the BMBF via the CSCC (FKZ 01EO1502) and from the DFG via the Jena Biophotonic and Imaging Laboratory (JBIL, FKZ PO 633/29-1, BA 1601/10-1) is highly acknowledged.Keywords: correlative image analysis, intracellular infection, pathogen-host adaption, Raman micro-spectroscopy
Procedia PDF Downloads 1811321 Multivariate Genome-Wide Association Studies for Identifying Additional Loci for Myopia
Authors: Qiao Fan, Xiaobo Guo, Junxian Zhu, Xiaohu Ding, Ching-Yu Cheng, Tien-Yin Wong, Mingguang He, Heping Zhang, Xueqin Wang
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A systematic, simultaneous analysis of multiple phenotypes in genome-wide association studies (GWASs) draws a great attention to integrate the signals from single phenotypes with increased power. However, lacking an interpretable and efficient multivariate GWAS analysis impede the application of such approach. In this study, we propose to decompose the multivariate model into a series of simple univariate models. This transformation illuminates what exactly the individual trait contributes to the significant signals from the multivariate analyses. By employing our approach in the analysis of three myopia-related endophenotypes from the Singapore Malay Eye Study (SIMES), we identify novel candidate loci which were successfully validated in an independent Guangzhou Twin Eye Study (GTES).Keywords: GWAS multivariate, multiple traits, myopia, association
Procedia PDF Downloads 2241320 Evaluation of Neonicotinoids Against Sucking Insect Pests of Cotton in Laboratory and Field Conditions
Authors: Muhammad Sufyan, Muhammad D. Gogi, Muhammad Arshad, Ahmad Nawaz, Muhammad Usman
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Cotton (Gossypium hirsutum) universally known as silver fiber and is one of the most important cash crop of Pakistan. A wide array of pests constraints cotton production among which sucking insect pests cause serious losses. Mostly new chemistry insecticides used to control a wide variety of insect pests including sucking insect pests. In the present study efficacy of different neonicotinoids was evaluated against sucking insect pests of cotton in the field and in laboratory for red and dusky cotton bug. The experiment was conducted at Entomology Research Station, University of Agriculture Faisalabad, in a Randomized Complete Block Design (RCBD). Field trial was conducted to evaluate the efficacy of Confidence Ultra (Imidacloprid) 70% SL, Confidor (Imidacloprid) 20% SL, Kendo (Lambda cyhalothrin) 24.7 SC, Actara (Thiamethoxam) 25% WG, Forcast (Tebufenozide+ Emamectin benzoate) 8.8 EW and Timer (Emamectin benzoate) 1.9 EC at their recommended doses. The data was collected on per leaf basis of thrips, aphid, jassid and whitefly before 24 hours of spray. The post treatment data was recorded after 24, 48 and 72 hours. The fresh, non-infested and untreated cotton leaves was collected from the field and brought to the laboratory to assess the efficacy of neonicotinoids against red and dusky cotton bug. After data analysis all the insecticides were found effective against sucking pests. Confidence Ultra was highly effective against the aphid, jassid, and whitefly and gave maximum mortality, while showed non-significant results against thrips. In case of aphid plot which was treated with Kando 24.7 SC showed significant mortality after 72 hours of pesticide application. Similar trends were found in laboratory conditions with all these treatments by making different concentrations and had significant impact on dusky cotton bug and red cotton bug population after 24, 48 and 72 hours after application.Keywords: cotton, laboratory and field conditions, neonicotinoids, sucking insect pests
Procedia PDF Downloads 2421319 Impact of Foliar Formulations of Macro and Micro Nutrients on the Tritrophic Association of Wheat Aphid and Entomophagous Insects
Authors: Muhammad Sufyan, Muhammad J. Arif, Muhammad Arshad, Usman Shoukat
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In Pakistan, wheat (Triticum aestivum L.) is seriously attacked by the wheat aphid. Naturally, bio control agents play an important role in managing wheat aphid. However, association among pest, natural enemies and host plant is highly affected by food resource concentration and predator/parasitoid factor of any ecosystem. The present study was conducted to estimate the effect of different dose levels of macro and micronutrients on the aphid population and its entomophagous insect on wheat and their tri-trophic association. The experiment was laid out in RCBD with six different combinations of macro and micronutrients and a control treatment. The data was initiated from the second week of the February till the maturity of the crop. Data regarding aphid population and coccinellids counts were collected on weekly basis and subjected to analysis of variance and mean comparison. The data revealed that aphid population was at peak in the last week of March. Coccinellids population increased side by side with aphid population and declined after second week of April. Aphid parasitism was maximum 25% on recommended dose of Double and Flasher and minimum 8.67% on control treatment. Maximum aphid population was observed on first April with 687.2 specimens. However, this maximum population was shown against the application of Double + Flasher treatment. The minimum aphid population was recorded after the application of HiK Gold + Flasher recommended dose on 15th April. The coccinellids population was at peak level at on 8th April and against the treatment double recommended dose of HiK gold + Flasher. Amount of nitrogen, phosphorus and potassium percentage dry leaves components was maximum (2.33, 0.18 and 2.62 % dry leaves. respectively) in plots treated with recommended double dose mixture of Double + Flasher and Hi-K Gold + Flasher while it was minimum (1.43, 0.12 and 1.77 dry leaves respectively) in plots where no nutrients applied. The result revealed that maximum parasitism was at recommended level of micro and macro nutrients application. Maximum micro nutrients zinc, copper, manganese, iron and boron found with values 46.67 ppm, 21.81 ppm, 62.35 ppm, 152.69 ppm and 36.78 respectively. The result also showed that Over application of macro and micro nutrients should be avoided because it do not help in pest control, conversely it may cause stress on plant. The treatment Double and Flasher recommended dose ratio is almost comparable with recommended dose and present studies confirm its usefulness on wheat.Keywords: entomophagous insects, macro and micro nutrients, tri-trophic, wheat aphid
Procedia PDF Downloads 2301318 Risks for Cyanobacteria Harmful Algal Blooms in Georgia Piedmont Waterbodies Due to Land Management and Climate Interactions
Authors: Sam Weber, Deepak Mishra, Susan Wilde, Elizabeth Kramer
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The frequency and severity of cyanobacteria harmful blooms (CyanoHABs) have been increasing over time, with point and non-point source eutrophication and shifting climate paradigms being blamed as the primary culprits. Excessive nutrients, warm temperatures, quiescent water, and heavy and less regular rainfall create more conducive environments for CyanoHABs. CyanoHABs have the potential to produce a spectrum of toxins that cause gastrointestinal stress, organ failure, and even death in humans and animals. To promote enhanced, proactive CyanoHAB management, risk modeling using geospatial tools can act as predictive mechanisms to supplement current CyanoHAB monitoring, management and mitigation efforts. The risk maps would empower water managers to focus their efforts on high risk water bodies in an attempt to prevent CyanoHABs before they occur, and/or more diligently observe those waterbodies. For this research, exploratory spatial data analysis techniques were used to identify the strongest predicators for CyanoHAB blooms based on remote sensing-derived cyanobacteria cell density values for 771 waterbodies in the Georgia Piedmont and landscape characteristics of their watersheds. In-situ datasets for cyanobacteria cell density, nutrients, temperature, and rainfall patterns are not widely available, so free gridded geospatial datasets were used as proxy variables for assessing CyanoHAB risk. For example, the percent of a watershed that is agriculture was used as a proxy for nutrient loading, and the summer precipitation within a watershed was used as a proxy for water quiescence. Cyanobacteria cell density values were calculated using atmospherically corrected images from the European Space Agency’s Sentinel-2A satellite and multispectral instrument sensor at a 10-meter ground resolution. Seventeen explanatory variables were calculated for each watershed utilizing the multi-petabyte geospatial catalogs available within the Google Earth Engine cloud computing interface. The seventeen variables were then used in a multiple linear regression model, and the strongest predictors of cyanobacteria cell density were selected for the final regression model. The seventeen explanatory variables included land cover composition, winter and summer temperature and precipitation data, topographic derivatives, vegetation index anomalies, and soil characteristics. Watershed maximum summer temperature, percent agriculture, percent forest, percent impervious, and waterbody area emerged as the strongest predictors of cyanobacteria cell density with an adjusted R-squared value of 0.31 and a p-value ~ 0. The final regression equation was used to make a normalized cyanobacteria cell density index, and a Jenks Natural Break classification was used to assign waterbodies designations of low, medium, or high risk. Of the 771 waterbodies, 24.38% were low risk, 37.35% were medium risk, and 38.26% were high risk. This study showed that there are significant relationships between free geospatial datasets representing summer maximum temperatures, nutrient loading associated with land use and land cover, and the area of a waterbody with cyanobacteria cell density. This data analytics approach to CyanoHAB risk assessment corroborated the literature-established environmental triggers for CyanoHABs, and presents a novel approach for CyanoHAB risk mapping in waterbodies across the greater southeastern United States.Keywords: cyanobacteria, land use/land cover, remote sensing, risk mapping
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