Search results for: brine shrimp assay (BSA)

Authors: Yagahira E. Castro-Sesquen, Chloe Kim, Robert H. Gilman, David J. Sullivan, Peter C. Searson

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Diagnosis of severe malaria is particularly important in highly endemic regions since most patients are positive for parasitemia and treatment differs from non-severe malaria. Diagnosis can be challenging due to the prevalence of diseases with similar symptoms. Accurate diagnosis is increasingly important to avoid overprescribing antimalarial drugs, minimize drug resistance, and minimize costs. A nanoparticle-based assay for detection and quantification of Plasmodium falciparum histidine-rich protein 2 (HRP2) in urine and serum is reported. The assay uses magnetic beads conjugated with anti-HRP2 antibody for protein capture and concentration, and antibody-conjugated quantum dots for optical detection. Western Blot analysis demonstrated that magnetic beads allows the concentration of HRP2 protein in urine by 20-fold. The concentration effect was achieved because large volume of urine can be incubated with beads, and magnetic separation can be easily performed in minutes to isolate beads containing HRP2 protein. Magnetic beads and Quantum Dots 525 conjugated to anti-HRP2 antibodies allows the detection of low concentration of HRP2 protein (0.5 ng mL-1), and quantification in the range of 33 to 2,000 ng mL-1 corresponding to the range associated with non-severe to severe malaria. This assay can be easily adapted to a non-invasive point-of-care test for classification of severe malaria.

Keywords: HRP2 protein, malaria, magnetic beads, Quantum dots

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Authors: Hassan Sbiga

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Laboratory measurements methods were undertaken on core samples selected from three different fields (A, B, and C) from the Nubian Sandstone Formation of the central graben reservoirs in Libya. These measurements were conducted in order to determine the factors which affect resistivity parameters, and to investigate the effect of rock heterogeneity and wettability on these parameters. This included determining the saturation exponent (n) in the laboratory at two stages. The first stage was before wettability measurements were conducted on the samples, and the second stage was after the wettability measurements in order to find any effect on the saturation exponent. Another objective of this work was to quantify experimentally pores and porosity types (macro- and micro-porosity), which have an affect on the electrical properties, by integrating capillary pressure curves with other routine and special core analysis. These experiments were made for the first time to obtain a relation between pore size distribution and saturation exponent n. Changes were observed in the formation resistivity factor and cementation exponent due to ambient conditions and changes of overburden pressure. The cementation exponent also decreased from GHE-5 to GHE-8. Changes were also observed in the saturation exponent (n) and water saturation (Sw) before and after wettability measurement. Samples with an oil-wet tendency have higher irreducible brine saturation and higher Archie saturation exponent values than samples with an uniform water-wet surface. The experimental results indicate that there is a good relation between resistivity and pore type depending on the pore size. When oil begins to penetrate micro-pore systems in measurements of resistivity index versus brine saturation (after wettability measurement), a significant change in slope of the resistivity index relationship occurs.

Keywords: part of thesis, cementation, wettability, resistivity

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Authors: Khaled M. Mohamed

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Background: P-Phenylenediamine (PPD) is used in the manufacture of hair dyes and skin decoration. In some developing countries, suicidal, homicidal and accidental cases by PPD were recorded. In this work, a sensitive LC-MS/MS method for determination of PPD and its metabolites N-acetyl-p-phenylenediamine (MAPPD) and N,N-diacetyl-p-phenylenediamine (DAPPD) in human urine has been developed and validated. Methods: PPD, MAPPD and DAPPD were extracted from urine by methylene chloride at alkaline pH. Acetanilide was used as internal standard (IS). The analytes and IS were separated on an Eclipse XDB- C18 column (150 X 4.6 mm, 5 µm) using a mobile phase of acetonitrile-1% formic acid in gradient elution. Detection was performed by LC-MS/MS using electrospray positive ionization under multiple reaction-monitoring mode. The transition ions m/z 109 → 92, m/z 151 → 92, m/z 193 → 92, and m/z 136 → 77 were selected for the quantification of PPD, MAPPD, DAPPD, and IS, respectively. Results: Calibration curves were linear in the range 10–2000 ng/mL for all analytes. The mean recoveries for PPD, MAPPD and DAPPD were 57.62, 74.19 and 50.99%, respectively. Intra-assay and inter-assay imprecisions were within 1.58–9.52% and 5.43–9.45% respectively for PPD, MAPPD and DAPPD. Inter-assay accuracies were within -7.43 and 7.36 for all compounds. PPD, MAPPD and DAPPD were stable in urine at –20 degrees for 24 hours. Conclusions: The method was successfully applied to the analysis of PPD, MAPPD and DAPPD in urine samples collected from suicidal cases.

Keywords: p-Phenylenediamine, metabolites, urine, LC-MS/MS, validation

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Authors: A. Mohammad Abdul Motalib Momin, B. Sheikh Mohammad Adil Uddin, C. Md Mamunur Rashid, D. Sheikh Arman Mahbub, E. Mohammad Sazzad Rahman, F. Abdullah Faruque

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The present study was designed to investigate the antioxidant and cytotoxicity potential of methanol and pet ether extracts of the Lagenaria siceraria (LM, LP), Cucumis sativus (CSM, CSP), Cucurbita maxima (CMM, CMP) plants. For the phytochemical screening, crude extract was tested for the presence of different chemical groups. In Lagenaria siceraria the following groups were identified: alkaloids, steroids, glycosides and saponins for methanol extract and alkaloids, steroids, glycosides, tannins and saponins are for pet ether extract. Glycosides, steroids, alkaloids, saponins and tannins are present in the methanol extract of Cucumis sativus; the pet ether extract has the alkaloids, steroids and saponins. Glycosides, steroids, alkaloids, saponins and tannins are present in both the methanolic and pet ether extract of Cucurbita maxima. In vitro antioxidant activity of the extracts were performed using DPPH radical scavenging, nitric oxide (NO) scavenging, total antioxidant capacity, total phenol content, total flavonoid content, and Cupric Reducing Antioxidant Capacity assays. The most prominent antioxidant activity was observed with the CSM in the DPPH free radical scavenging test with an IC50 value of 1667.23±11.00271 μg/ml as opposed to that of standard ascorbic acid (IC50 value of 15.707± 1.181 μg/ml.) In total antioxidant capacity method, CMP showed the highest activity (427.81±11.4 mg ascorbic acid/g). The total phenolic and flavonoids content were determined by Folin-Ciocalteu Reagent and aluminium chloride colorimetric method, respectively. The highest total phenols and total flavonoids content were found in CMM and LP with the value of 79.06±16.06 mg gallic acid/g & 119.0±1.41 mg quercetin/g, respectively. In nitric oxide (NO) scavenging the most prominent antioxidant activity was observed in CMM with an IC50 value of 8.119± 0.0036 μg/ml. The Cupric reducing capacity of the extracts was strong and dose dependent manner and CSM showed lowest reducing capacity. The cytotoxicity was determined by Brine shrimp lethality test and among these extracts most potent cytotoxicity was shown by CMM with LC50 value 16.98 µg/ml. The obtained results indicate that the investigated plants could be potential sources of natural antioxidants and can be used for various types of diseases.

Keywords: antioxidant, cytotoxicity, methanol, petroleum ether

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Authors: Canan Türeli̇, İrem Nur Yeşi̇lyurt

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A total of nine stomatopod crustaceans have been reported from the eastern Mediterranean, with the eight from the Turkey coasts and three from the Levantine coast of Turkey (Squilla massavensis (Linnaeus,1758), Erugosquilla massavensis (Kossmann, 1880), Clorida albolitura (Ahyong&Naiyanetr, 2000). E. massavensis is the most successful Red Sea immigrant, and it is now the dominant eastern Levantine stomatopod. At this research, carapace length/total length- weight relationship of 2 erythrean mantis shrimp, E. massavensis and C. albolitura was studied in Yumurtalık Cove, in the Northeastern Mediterranean of Turkey. Samples were collected monthly from July 2014 to June 2015 (except February), using bottom trawl net into three different depth strata; 0- 10 m, 10-20 m, 20-50m. E. massavensis samples were caught all studied time, except winter, in the three depth range but C. albolitura samples were caught only in July, October and May's months. A total of 572 specimens of the E. massavensis and 12 specimens of the C. albolitura were collected and analyzed. Morphometric equations for the conversions of total length and weight, carapace lengths were constructed for males, females, combined sexes for E. massavensis and C. albolitura. The b-values for total length (TL)-weight (W) in males and females for E. massavensis were 2.738 and 2.725 respectively and for carapace length (CL)-total length (TL) they were 1.041 and 0.978, relatively. The b-values (total length-weight) were determined for combined sexes of E. massavensis and C. albolitura as: 2.771, 1.852 relatively. In both species, the relative growth of weight and total length was found highly positively allometric. However, carapace length showed negative allometry in females of E. Massavensis.

Keywords: Clorida albolitura, Erugosquilla massavensis, Erythrean stomatopod, length/length-weight relationships

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Authors: Chanasit Chaocharoenphat

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Hibiscus sabdariffa is a herbal plant that is commonly used for home remedies in Thailand. This study aims to determine the antioxidant activity of polyphenols, as oxidative stress plays a vital role in the development of cancer, and H. sabdariffa was used in this study. The total flavonoids content was determined using the aluminium chloride colourimetric method and expressed as quercetin equivalents (QE)/g and the antioxidant capacity of the flavonoids using the 2,2-diphenyl-1-picrylhydrazyl (DPPH) and 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) radical scavenging capacity assays. The IC50 values of H. sabdariffa extract were 167.14 μg/mL ± 0.843 and 77.59 μg/mL ± 0.798, respectively. In the DPPH assay, vitamin C was used as a positive control, whereas Trolox was used as a positive control in the ABTS assay. To summarise, H. sabdariffa extract contains a high concentration of total flavonoids and exhibits potent antioxidant activity. However, additional antioxidant activity assays such as superoxide dismutase (SOD), reactive oxygen species (ROS), and reactive nitrogen species (RNS) scavenging assays and in vitro antioxidant experiments should be carried out to investigate the molecular mechanism of the compound.

Keywords: ABTS assay, antioxidant activity, Gracilaria fisheri, DPPH assays, total flavonoid content

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Authors: Hinda Berghiche, Hamida Benradia, Noureddine Soltani

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Pesticides are widely used in crop production and are known to induce a major contamination of ecosystems especially in aquatic environments. The leaching of a large amount of pollutants derived from agricultural activities (fertilizers, pesticides) might contaminate rivers which diverse into the likes and estuarine and coastal environments affecting several organisms such as crustacean species. In this context, there is searched for new selective insecticides with minimal toxic effects on the environment and human health such as growth insect regulators (GIRs). The current study aimed to examine the impact of novaluron (CE 20%), a potent benzoylphenylurea derivative insecticide on mosquito larvae, against non-target shrimp, Palaemon adspersus (Decapoda, Palaemonidae). The compound was tested at two concentrations (0.91 mg/L and 4.30 mg/L) corresponding respectively to the LC50 and LC90 determined against fourth-instar larvae of Culiseta longiareolata (Diptera, Culicidae). The molting hormone titer was determined in the haemolymph by an enzyme-immunoassay, while chitin was measured in peripheral integument at different stages during the molting cycle. Under normal conditions, the haemolymphatic ecdysteroid concentrations increased during the molting cycle to reach peak at stage D. In the treated series, we note absence of the peak at stage D and an increase at stages B, C and D as compared to the controls. Concerning the chitin amounts, we observe an increase from stage A to stage C followed by a decrease at stage D. Exposition of shrimps to novaluron resulted in a significant decrease of values at all molting stages with a dose-response effect. Thus, the insecticide can present secondary effects on this non-target arthropod species.

Keywords: toxicology, novaluron, crustacean, palaemon adspersus, ecdysteroids, cuticle, chitin

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Authors: Mengjie Qu, Yi Wang, Jiawei Ding, Siyu Chen, Yanan Di

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Marine ecosystem is facing intensified multiple stresses caused by environmental contaminants from human activities. Xenobiotics, such as benzo(α)pyrene (BaP) have been discharged into marine environment and cause hazardous impacts on both marine organisms and human beings. As a filter-feeder, marine mussels, Mytilus spp., has been extensively used to monitor the marine environment. However, their genomic alterations induced by such xenobiotics are still kept unknown. In the present study, gills, as the first defense barrier in mussels, were selected to evaluate the genetic instability alterations induced by the exposure to BaP both in vivo and in vitro. Both random amplified polymorphic DNA (RAPD) assay and comet assay were applied as the rapid tools to assess the environmental stresses due to their low money- and time-consumption. All mussels were identified to be the single species of Mytilus coruscus before used in BaP exposure at the concentration of 56 μg/l for 1 & 3 days (in vivo exposure) or 1 & 3 hours (in vitro). Both RAPD and comet assay results were showed significantly increased genomic instability with time-specific altering pattern. After the recovery period in 'in vivo' exposure, the genomic status was as same as control condition. However, the relative higher genomic instabilities were still observed in gill cells after the recovery from in vitro exposure condition. Different repair mechanisms or signaling pathway might be involved in the isolated gill cells in the comparison with intact tissues. The study provides the robust and rapid techniques to exam the genomic stability in marine organisms in response to marine environmental changes and provide basic information for further mechanism research in stress responses in marine organisms.

Keywords: genotoxic impacts, in vivo/vitro exposure, marine mussels, RAPD and comet assay

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Authors: Pallavi Srivastava, Ajay Singh

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Hematological assay has used for evaluation of blood changes according to its environment. It’s studies employed to evaluate possible eco-toxic risk due to the exposure of chemicals and pesticides in aquatic organisms. Fishes serve as a sensitive bio-indicator, as changes occur in its surrounding environment. The aim of present study has two-folds first we observed that after exposure of two doses of each class of fungicide i.e. 1.11mg/l, 2.23mg/l for Propiconazole and 11.43mg/l, 22.87mg/l for Mancozeb show maximum blood changes. Second we conclude that toxic effects and blood changes induced by Propiconazole is greater than Mancozeb.

Keywords: hematological assay, fungicides, bio-indicator, eco-toxic risk

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Authors: Jichan Jang

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Mycobacterium abscessus is a rapidly growing life-threatening mycobacterium with multiple drug-resistance mechanisms. In this study, we screened the library to identify active molecules targeting Mycobacterium abscessus using resazurin live/dead assays. In this screening assay, the Z-factor was 0.7, as an indication of the statistical confidence of the assay. A cut-off of 80% growth inhibition in the screening resulted in the identification of four different compounds at a single concentration (20 μM). Dose-response curves identified three different hit candidates, which generated good inhibitory curves. All hit candidates were expected to have different molecular targets. Thus, we found that compound X, identified, may be a promising candidate in the M. abscessus drug discovery pipeline.

Keywords: Mycobacterium abscessus, antibiotics, drug discovery, emerging Pathogen

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Authors: Olawale Babatunde Olatinsu, Deborah Oluwaseun Olorode

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Industrial rocks were collected from a quarry site at Ewekoro in south-western Nigeria and analysed using Nuclear Magnetic Resonance (NMR) technique. NMR measurement was conducted on the samples in partial water-saturated and full brine-saturated conditions. Raw NMR data were analysed with the aid of T2 curves and T2 spectra generated by inversion of raw NMR data using conventional regularized least-squares inversion routine. Results show that NMR transverse relaxation (T2) signatures fairly adequately distinguish between the rock types. Similar T2 curve trend and rates at partial saturation suggests that the relaxation is mainly due to adsorption of water on micropores of similar sizes while T2 curves at full saturation depict relaxation decay rate as: 1/T2(shale)>1/ T2(glauconite)>1/ T2(limestone) and 1/T2(sandstone). NMR T2 distributions at full brine-saturation show: unimodal distribution in shale; bimodal distribution in sandstone and glauconite; and trimodal distribution in limestone. Full saturation T2 distributions revealed the presence of well-developed and more abundant micropores in all the samples with T2 in the range, 402-504 μs. Mesopores with amplitudes much lower than those of micropores are present in limestone, sandstone and glauconite with T2 range: 8.45-26.10 ms, 6.02-10.55 ms, and 9.45-13.26 ms respectively. Very low amplitude macropores of T2 values, 90.26-312.16 ms, are only recognizable in limestone samples. Samples with multiple peaks showed well-connected pore systems with sandstone having the highest degree of connectivity. The difference in T2 curves and distributions for the rocks at full saturation can be utilised as a potent diagnostic tool for discrimination of these rock types found at Ewekoro.

Keywords: Ewekoro, NMR techniques, industrial rocks, characterization, relaxation

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Authors: Ceyda Okudu, Secil Eroglu, Khandakar A. S. M. Saadat, Sibel O. Balci

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Ring Finger Protein 2 (RNF2) is a member of polycomb repressive complex 1 (PRC1), which is one of the epigenetic regulators in the genome. When RNF2 combines with other PRC1 members, it mediates the mono-ubiquitination of Histon2A (H2A). In breast cancer, RNF2 is commonly overexpressed, and also it promotes metastasis and invasion in other aggressive tumors like melanoma, prostate, and hepatocarcinoma. The role of RNF2 in the metastasis and invasion of breast cancer has not yet been elucidated. Our aim is to observe the role of RNF2 in metastasis and invasion in this study by miRNA mediated RNF2 gene silencing in breast cancer cell lines. We selected miRNAs, targeting to RNF2 by searching online databases. miR-17-5p, miR20a-5p, and miR-106b-5p were transfected to breast cancer cell lines (MCF-7, MDA-MB-231, SK-BR-3, and ZR-75-1), and also we used normal breast epithelial cell line (hTERT-HME1) to compare RNF2 gene expression level. After 48-72 hours post-transfection, mRNAs were isolated from the cells, and gene expressions were measured by RT-qPCR after from cDNA syntheses. We observed that RNF2 was highly expressed in SK-BR-3 and MDA-MB-231 cell lines opposite to MCF-7 and ZR-75-1 cell lines. RNF2 was downregulated 5, 5 and 7 fold by miR17-5p, miR20a-5p and miR106b-5p respectively in MCF-7. However, in SK-BR-3 and ZR-75-1 cell lines, miRNAs did not affect significantly RNF2 gene expression level. miR20a-5p decreased RNF2 3 fold and miR17-5p and miR106b-5p did not affect MDA-MB-231. After gene expression analysis, we performed metastasis and invasion assay in MCF-7 cells. For metastasis, we used both wound healing assay and Transwell Cell Migration Assay, and we used Transwell Cell Invasion Assay for invasion. The data of this assay showed that miR17-5p and miR20a-5p decreased both invasion and metastasis level, but miR106b-5p has no effect. We would like to conclude that RNF2 can be targeted by miR17-5p, miR20a-5p and miR106b-5p in MCF-7 cells and also RNF2, which is one of the upregulated genes in aggressive tumor, can be decreased by using these miRNAs. In future, we would like to confirm these results at the protein level and also whether these miRNAs are direct target of RNF2 or not.

Keywords: breast cancer, epigenetic, microRNAs, RNF2

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Authors: A. Majeed, M.K. Abbasi, S. Hameed, A. Imran, T. Naqqash, M. K. Hanif

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Plant growth-promoting rhizobacteria (PGPR) are free-living soil bacteria that aggressively colonize the rhizosphere/plant roots, and enhance the growth and yield of plants when applied to seed or crops. Root associated (endophytic and rhizospheric) PGPR were isolated from Sunflower (Helianthus anus) grown in soils collected from 16 different sites of sub division Dhirkot, Poonch, Azad Jammu & Kashmir, Pakistan. A total of 150 bacterial isolates were isolated, purified, screened in vitro for their plant growth promoting (PGP) characteristics. 11 most effective isolates were selected on the basis of biochemical assays (nitrogen fixation, phosphate solubilization, growth hormone production, biocontrol assay, and carbon substrates utilization assay through gas chromatography (GCMS), spectrophotometry, high performance liquid chromatography HPLC, fungal and bacterial dual plate assay and BIOLOG GN2/GP2 microplate assay respectively) and were tested on the crop under controlled and field conditions. From the inoculation assay, the most promising 4 strains (on the basis of increased root/shoot weight, root/shoot length, seed oil content, and seed yield) were than selected for colonization studies through confocal laser scanning and transmission electron microscope. 16Sr RNA gene analysis showed that these bacterial isolates belong to Pseudononas, Enterobacter, Azospirrilum, and Citobacter genera. This study is the clear evident that such isolates have the potential for application as inoculants adapted to poor soils and local crops to minimize the chemical fertilizers harmful for soil and environment

Keywords: PGPR, nitrogen fixation, phosphate solubilization, colonization

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Authors: Yun-Hsiang Wang, Huai-Yi Chen, Kin Fong Lei

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Deoxyribonucleic acid (DNA) hybridization is a common technique used in genetic assay widely. However, the hybridization ratio and rate are usually limited by the diffusion effect. Here, microfluidic electrode platform producing electroosmosis generated by alternating current signal has been proposed to enhance the hybridization ratio and rate. The electrode was made of aurum fabricated by microfabrication technique. Thiol-modified oligo probe was immobilized on the electrode for specific capture of target, which is modified by fluorescent tag. Alternative electroosmosis can induce local microfluidic vortexes to accelerate DNA hybridization. This study provides a strategy to enhance the rate of DNA hybridization in the genetic assay.

Keywords: DNA hybridization, electroosmosis, electrical enhancement, hybridization ratio

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Authors: Muhammad Farooq Baig, Saleem Qadeer

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Background: The frequency of hepatitis C virus infection along with tuberculosis has not been widely investigated and very low statistics on rates of hepatitis C virus co-infection in tuberculosis patients. Hepatotoxicity is the major side effect of anti-tuberculosis therapy hepatitis HCVliver disease elevates the chances of hepatotoxicity up-to five folds. Objectives & Aim: To see the frequency of Hepatitis Cvirus infection amongst people with diagnosed Tuberculosis using gene X-pert technique. To evaluate the factors associated with HCVinfection in patients with MTBtuberculosis and to determine sensitivity and specificity of the tests. Study design: Comparative analytical study. Methodology: Three hundred and thirteen patients of tuberculosis diagnosed by Genexpert included while testing hepatitis C virus using immunochromotography rapid test technique, enzyme linked immunosorbent assay method and polymerase chain reaction test for confirmation. Results:Higher frequency of tuberculosis infection in males 57.8%, 42.5% between 20-39 years and 22% of hepatitis C virus infection in tuberculosis patients.The sensitivity of rapid test and enzyme-linked immunosorbent assay was 79% and 96% respectively while the specificity of rapid test and enzyme-linked immunosorbent assay was 91% and 99% respectively.

Keywords: Mycobactrium Tuberculosis, PC'R, Gene x pert, Hepatitis C virus

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Authors: H. Bouaziz, M. Sefi, J. de Lapuente, M. Borras, N. Zeghal

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Although arsenic trioxide has been the subject of toxicological research, in vitro cytotoxicity and genotoxicity studies using relevant cell models and uniform methodology are not well elucidated. Hence, the aim of the present study was to evaluate the cytotoxicity and genotoxicity induced by arsenic trioxide in human keratinocytes (HaCaT) using the MTT [3-(4, 5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] and alkaline single cell gel electrophoresis (Comet) assays, respectively. Human keratinocytes were treated with different doses of arsenic trioxide for 4 h prior to cytogenetic assessment. Data obtained from the MTT assay indicated that arsenic trioxide significantly reduced the viability of HaCaT cells in a dose-dependent manner, showing a IC50 value of 34.18 ± 0.6 µM. Data generated from the comet assay also indicated a significant dose-dependent increase in DNA damage in HaCaT cells associated with arsenic trioxide exposure. We observed a significant increase in comet tail length and tail moment, showing an evidence of arsenic trioxide -induced genotoxic damage in HaCaT cells. This study confirms that the comet assay is a sensitive and effective method to detect DNA damage caused by arsenic.

Keywords: arsenic trioxide, cytotoxixity, genotoxicity, HaCaT

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Authors: Wenxiao Liu, Kun Zhang, Yongqing Li

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Bovine herpesvirus 1, a member of the genus Variellovirus of the subfamily Alphaherpesvirinae, has caused severe economic cost to the bovine industry. In this study, BoHV-1 glycerol protein gD was expressed in insect cells, and the purified gD was immunized in the Balb/C mice to generate monoclonal antibodies. Based on hybridoma cell fusion techniques, 20 monoclonal antibodies against Bovine herpesvirus 1 have been obtained. Further, mAb 3F8 with neutralizing activity and gD were applied to develop a blocking enzyme-linked immunosorbent assay (Elisa) for detecting neutralizing antibodies against BoHV-1, which shows a significant correlation between the blocking Elisa and VNT. The sensitivity and specificity of the test were estimated to be 94.59% and 93.42%, respectively. Furthermore, antibody pairing tests revealed that mAb 1B6 conjugated to fluorescence microspheres was used as the capture antibody, and mAb 3F9 was used as the detectable antibody to establish the immunochromatographic assay (ICS). The ICS was conducted to detect BoHV-1 in bovine samples with high sensitivity, specificity, and good stability. Clinical sample testing revealed that the results of ICS and real-time PCR have a coincidence rate of 95.42%. Our research confirmed that the ICS is a rapid and reliable method for the diagnosis of BoHV-1. In conclusion, our results lay a solid foundation for the prevention and control of BoHV-1 infection.

Keywords: bovine disease, BoHV-1, ELISA, ICS assay

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Authors: Juan Carlos Aguirre Pabon, Stephen Sabatino, James Morris, Khor Waiho, Antonio Murias

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The tiger shrimp Penaeus monodon is one of the most important species in aquaculture and is native to the Indo-Pacific Ocean. During its greatest success in world production (70s and 80s) was introduced in many Atlantic Ocean countries for cultivation purposes and is currently reported as established in several countries of this area. Because there are no studies to understand the magnitude of the invasion process, this is an exciting opportunity to test evolutionary hypotheses in the context of marine invasions mediated by culture systems; therefore, the purpose of this study was to reconstruct the scenario of invasion of P. monodon in the Atlantic Ocean, by using mitochondrial DNA and eight loci microsatellites. In addition, samples of the invasion area in the Atlantic Ocean (US, Colombia, Venezuela, Brazil, Guienne Bissau, Senegal), the Indo-Pacific Ocean (Indonesia, India, Mozambique), and some cultivation systems (India, Bangladesh, Madagascar) were collected; and analysis of phylogenetic relationships (using some species of the family), genetic diversity, structure population, and demographic changes were performed. High intraspecific divergence in P. semisulcatus and P. monodon were found, high genetic variability in all sites (especially with microsatellites) and the presence of three clusters or populations. In addition, signs of demographic expansion in the culture population and bottlenecks in the invasive and native populations were found, as well as evidence of gene mixtures from all of the populations studied, implying that cropping systems play an essential role in mitigating the negative effects of the founder effect and providing a source of genetic variability that can ensure the success of the invasion.

Keywords: species introduction, increased variability, demographic changes, promoting invasion.

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Authors: Neha Singh, Anuj Ranjan, Tanu Jindal

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Over the last decade, the exponential growth of mobile communication has been accompanied by a parallel increase in density of electromagnetic fields (EMF). The continued expansion of mobile phone usage raises important questions as EMF, especially radio frequency (RF), have long been suspected of having biological effects. In the present experiments, we studied the effects of RF-EMF on cell death (apoptosis) and DNA damage of a well- tested biological model, Drosophila melanogaster exposed to 2400 MHz frequency for different time duration i.e. 2 hrs, 4 hrs, 6 hrs,8 hrs, 10 hrs, and 12 hrs each day for five continuous days in ambient temperature and humidity conditions inside an exposure chamber. The flies were grouped into control, sham-exposed, and exposed with 100 flies in each group. In this study, well-known techniques like Comet Assay and TUNEL (Terminal deoxynucleotide transferase dUTP Nick End Labeling) Assay were used to detect DNA damage and for apoptosis studies, respectively. Experiments results showed DNA damage in the brain cells of Drosophila which increases as the duration of exposure increases when observed under the observed when we compared results of control, sham-exposed, and exposed group which indicates that EMF radiation-induced stress in the organism that leads to DNA damage and cell death. The process of apoptosis and mutation follows similar pathway for all eukaryotic cells; therefore, studying apoptosis and genotoxicity in Drosophila makes similar relevance for human beings as well.

Keywords: cell death, apoptosis, Comet Assay, DNA damage, Drosophila, electromagnetic fields, EMF, radio frequency, RF, TUNEL assay

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Authors: Laura Boschis, Elena D. Ozzello, Enzo Mastromatteo

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Point of care diagnostic finds growing interest in medicine and agri-food because of faster intervention and prevention. EliChip is a microfluidic platform to perform Point of Care immunoenzymatic assay based on ready-to-use kits and a portable instrument to manage fluidics and read reliable quantitative results. Thanks to miniaturization, analyses are faster and more sensible than conventional ELISA. EliChip is one of the crucial assets of the Europen-founded Flamingo project for in-line measuring inflammatory markers.

Keywords: lab on chip, point of care, immunoenzymatic analysis, synovial arthritis

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Authors: Nurjannatul Naim Kamaruddin, Tengku Sifziuzl Tengku Muhammad, Aina Farahiyah Abdul Manan, Habsah Mohamad

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Atherosclerosis which leads to cardiovascular diseases such as myocardial infarction, unstable angina (ischemic heart pain), sudden cardiac death and stroke is the principal cause of death worldwide. It has been a very critical issue as current common drug treatment, statin therapy has left bad side effects like rhabdomyolysis, atrial fibrillation, liver disease, abdominal and chest pain. Interestingly, the discoveries of proprotein convertase subtilisin-kexin type 9 have paved a new way in the treatment of atherosclerosis. This serine protease is believed to involve in the regulation of LDL- uptake by LDL-receptor. Therefore, this study was conducted to evaluate the potential of Acanthaster plancii fractions to reduce the transcriptional activity of the PCSK9 promoter. In this study, the marine organism which is Acanthaster plancii has been used as the source for marine compounds in inhibiting PCSK9. The cytotoxicity activity of ten fractions from the methanol extracts of Acanthaster plancii was investigated on HepG2 cell lines using MTS assay and dual glo luciferase assay was carried out later to analyses the effects of the samples in reducing the transcriptional activity of the PCSK9 promoter. Both assays used fractions with five different concentrations, 3.13µg/mL, 6.25µg/mL, 12.5µg/mL, 25µg/mL, and 50µg/mL. MTS assay indicated that the fractions are non-cytotoxic towards HepG2 cell lines as their IC50 value is greater than 30µg/mL. Whilst, for the dual glo luciferase assay, among all the fractions, Enhance Fraction 2 (EF2) showed the best potential in reducing the transcriptional activity of the PCSK9 promoter. The results indicated that this EF2 gave the lowest PCSK9 promoter expression at low concentration which is 0.2 fold change at 6.25µg/mL. This finding suggested that further analysis should be done to validate the potential of Acanthaster plancii as the source of anti-atherosclerotic agent.

Keywords: Acanthaster plancii, atherosclerosis, luciferase assay, PCSK9

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Authors: Brizita Djordjevic, Bojana Vidovic, Milica Zrnic, Uros Cakar, Ivan Stankovic, Davor Korcok, Sladjana Sobajic

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Histamine is a biogenic amine, which is formed by enzymatic decarboxylation from the amino acid histidine. It can be found in foods such as fish and fish products, meat and fermented meat products, cheese, wine and beer. The presence of histamine in these foods can indicate microbiological spoilage or poor manufacturing processes. The consumption of food containing large amounts of histamine can have toxicological consequences. In 62 food products (31 canned fish products, 19 wines and 12 cheeses) from the market of Serbia the content of histamine was determined using enzyme-linked immunosorbent assay (ELISA) test kit according to the manufacturer's instructions (Immunolab GmbH, Kassel, Germany). The detection limits of this assay were 20 µg/kg for fish and cheese and 4 µg/L for wine. The concentration of histamine varied between 0.16-207 mg/kg in canned fish products, 0.03-1.47 mg/kg in cheeses and 0.01- 0.18 mg/L in wines. In all analyzed canned fish products the results obtained for the histamine were below the limits set by European and national legislation, so they can be considered acceptable and safe for the health consumers. The levels of histamine in analyzed cheeses and wines were very low and did not pose safety concerns.

Keywords: cheese, enzyme-linked immunosorbent assay, histamine, fish products, wine

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Authors: Shishen Xie, Yingda L. Xie

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Tuberculosis (TB) is a potentially serious infectious disease that remains a health concern. The Interferon Gamma Release Assay (IGRA) is a blood test to find out if an individual is tuberculous positive or negative. This study applies statistical analysis to the clinical data of interferon-gamma levels of seventy-three subjects who diagnosed pulmonary TB in an anti-tuberculous treatment. Data analysis is performed to determine if there is a significant decline in interferon-gamma levels for the subjects during a period of six months, and to infer if the anti-tuberculous treatment is effective.

Keywords: data analysis, interferon gamma release assay, statistical methods, tuberculosis infection

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Authors: Benyapa Suksuwan

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Introduction: Oxidative stress is identified as the root cause of the development and progression of several diseases as the disproportion of free radicals in the body leads to tissue or cell damage. Polyphenols are the most common antioxidant found in plants and are efficient in capturing oxidative free radicals. Aim of the Study: This study focused on the antioxidant activity of polyphenols extracted from Phyllanthus Emblica L. as oxidative stress plays a vital role in developing and progressing many diseases, including cardiovascular diseases and cancer. Materials and Methods: The plant was extracted using a mixture solvent (ethyl alcohol: water in ratio 8:2). The total phenolic content of P. Emblica extract was determined using the Folin-Cioucalteu method and calculated as gallic acid equivalents (GAE) and various antioxidant assays DPPH and ABTS radical scavenging capacity assays. Results and Discussion: The findings exhibited a strong correlation between antioxidant activity and the total phenol contents. In addition, the IC₅₀ of P. Emblica extract via DPPH and ABTS assays were 68.10 μg/mL ± 0.455, and 49.24 μg/mL ± 0.716, respectively. Furthermore, P. Emblica extract showed antioxidant activities in a concentration-dependent manner. Vitamin C was used as a positive control in the DPPH assay, while Trolox was used as a positive control in the ABTS assay. Conclusions: In conclusion, P. Emblica extract consisted of a high amount of total phenolic content, which possesses potent antioxidant activity. However, further antioxidant activity assays using human cell lines such as SOD, ROS, and RNS scavenging assays and in vitro antioxidant experiments should be performed in order.

Keywords: antioxidant, ABTS scavenging, DPPH scavenging assay, total phenol contents assay, Phyllanthus Emblica L

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Authors: Mohammed Falalu Hamza

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A phytochemical investigation conducted on the leaves extract of Cryptocarya latifolia (Lauraceae) revealed the presence of two major essential oils; Nerolidol (1) and Copaene (2) with the aid of gas chromatography-mass spectrometry (GC-MS). The compounds exhibited good anti-oxidant capacity using 2,2-diphenyl-1-picryl-hydrazyl (DPPH) radical scavenging assay. The result shows that the anti-oxidant capacity of the compounds is dependent on concentration similar to the standard (ascorbic acid). This study shows that the leaves extract of C. latifolia is a good source of important natural antioxidants.

Keywords: broad-leaved quince, phytochemical, anti-oxidant, essential oils

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Authors: Abdolreza Esmaeilzadeh, Maryam Erfanmanesh, Sousan Ghasemi, Farzaneh Mohammadi

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Background: Hepatitis C Virus (HCV), a public health problem, is an enveloped, single-stranded RNA virus and a member of the Hepacivirus genus of the Flaviviridae family. Liver cancer, cirrhosis, and end-stage liver are the outcomes of chronic infection with HCV. HCV isolates show significant heterogeneity in genetics around the world. Therefore, determining HCV genotypes is a vital step in determining prognosis and planning therapeutic strategies. Materials and Methods: Serum samples of 136 patients were collected and analyzed for anti-HCV antibodies using ELISA (The enzyme-linked immunosorbent assay) method. Then, positive samples were exposed to RT-PCR, which was performed under standard condition. Afterwards, they investigated for genotyping using allele-specific PCR (AS-PCR), and HCV genotype 2.0 line probe assay (LiPA). Results: Samples indicated 216 bp bands on 2% agarose gel. Analyses of the results demonstrated that the most dominant subtype was 3a with frequency of 38.26% in Zanjan Province followed by subtypes of 1b, 1a, 2, and 4 with frequencies of 25.73%, 22.05%, 5.14%, and 4.41%, respectively. The frequency of unknown HCV genotypes was 4.41%. Conclusions: According to the results, it was found that HCV high prevalent genotype in Zanjan is subtype 3a. Analysis of the results provides identification of certain HCV genotypes, and these valuable findings could affect the type and duration of the treatment.

Keywords: anti-HCV antibody, Hepatitis C Virus (HCV), genotype, RT-PCR, AS-PCR

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Authors: Suharni Mohamad Suharni Mohamad, Nurul Izzati Hamzan Nurul Izzati Hamzan, Norhayu Abdul Rahman Norhayu Abdul Rahman, Siti Suraiya Md Noor Siti Suraiya Md Noor

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Human papillomavirus (HPV) is an important risk factor for development of oral cancer. HPV16 is the most common type found in HPV-positive squamous cell carcinoma. In the present study, we established a real-time loop-mediated isothermal amplification (real-time LAMP) for detection of HPV16. A set of six primers was specially designed to recognize eight distinct sequences of HPV16-E6. Detection and quantification was achieved by real-time monitoring using a real-time turbidimeter based on threshold time required for turbidity in the LAMP reaction. LAMP reagents (MgSO4, dNTPs, Bst polymerase concentrations) and various incubation times and temperatures were optimized. The sensitivity was determined using 10-fold serial dilutions of HPV16 standard strain. The specificity of was evaluated using other HPV genotypes. The optimized method was established with specifically designed primers by real-time detection in approximately 30 min at 65°C. The limit of detection of HPV16 using the LAMP assay was 10 pg/ml that could be detected in 30 min. The LAMP assay was 10 times more sensitive than the conventional PCR in detecting HPV16. No cross-reactivity with other HPV genotypes was observed. This quantitative real-time LAMP assay may improve diagnostic potential for the detection and quantification of HPV16 in clinical samples and epidemiological studies due to its rapidity, simplicity, high sensitivity and specificity. This assay will be further evaluated with HPV DNAs of saliva from patients with oral squamous cell carcinoma. Acknowledgement: This study was financially supported by the ScienceFund Grant, Ministry of Science, Technology and Innovation (305/PPSG/6113219).

Keywords: Oral Squamous Cell Carcinoma (OSCC), Human Papillomavirus 16 (HPV16), Loop-Mediated Isothermal Amplification (LAMP), rapid detection

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Authors: E. Chiam, E. Koh, W. Teh, M. Prabhakaran

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Fruits and vegetables (F&V) are widely eaten for their nutritional value and associated health benefits being an immense source of bioactive compounds. However, F&V peels are often discarded, and it accounts for a higher proportion of food waste. Incorporation of F&V peels as functional ingredients can add more value to food due to the higher amounts of phytochemicals present in them. In this research, methanolic extracts of different F&V peels, namely apple, orange, kiwi, grapefruit, dragon fruit, pomelo, and pumpkin are investigated for their total phenolic content (TPC) by Folin-Ciocalteau (FC) assay and the antioxidant capacity was evaluated by 2,2-diphenyl-1-picrylhydrazyl (DPPH) and phosphomolybdenum assay using UV-Vis spectroscopy. Evaluation of the α-glucosidase inhibitory assay was carried out during this study to determine the antidiabetic potential of F&V peels. Results of our study showed that grapefruit peels contained the highest total phenolic content of 477.81 ± 0.01 mg gallic acid equivalent per gram dry weight of the sample, and kiwi peel had the highest antioxidant capacity (90.51 ± 0.10 % inhibition of DPPH radical) among the different F&V peels studied. Fruit peels exhibited high α-glucosidase inhibitory activity. Comparing fruit peels with vegetable peels, it was found that fruit peels had high total phenolic content, antioxidant capacity and anti-diabetic potential compared to vegetable peels.

Keywords: polyphenolics, fruit peels, antioxidant, antidiabetic

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Authors: S. Techaoei, K. Jarmkom, P. Eakwaropas, W. Khobjai

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The objective of this research was focused on investigating in vitro antimicrobial activity of Phellinus linteus fruiting body extracts on Pseudomonas aeruginosa, Escherichia coli, Staphylococcus aureus and Methicillin-resistant Staphylococcus aureus. Phellinus linteus fruiting body was extracted with ethanol and ethyl acetate and was vaporized. The disc diffusion assay was used to assess antimicrobial activity against tested bacterial strains. Primary screening of chemical profile of crude extract was determined by using thin layer chromatography. The positive control and the negative control were used as erythromycin and dimethyl sulfoxide, respectively. Initial screening of Phellinus linteus crude extract with the disc diffusion assay demonstrated that only ethanol had greater antimicrobial activity against Pseudomonas aeruginosa, Escherichia coli, Staphylococcus aureus and Methicillin-resistant Staphylococcus aureus. The MIC assay showed that the lower MIC was observed with 0.5 mg/ml of Pseudomonas aeruginosa and Methicillin-resistant Staphylococcus aureus and 0.25 mg/ml. of Escherichia coli and Staphylococcus aureus, respectively. TLC chemical profile of extract was represented at R_f ≈ 0.71-0.76.

Keywords: Staphylococcus aureus, Escherichia coli, Phellinus linteus, Methicillin-resistant Staphylococcus aureus, antimicrobial activity

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Authors: Xiaoqian Cheng, Jon Kleppe, Ole Torsaeter

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One objective of this work is to analyze the effects of surfactant properties (viscosity, concentration, and adsorption) on surfactant enhanced oil recovery at laboratory scale. The other objective is to obtain the functional relationships between surfactant properties and the ultimate oil recovery and oil recovery rate. A core is cut into two parts from the middle to imitate the matrix with a horizontal fracture. An injector and a producer are at the left and right sides of the fracture separately. The middle slice of the core is used as the model in this paper, whose size is 4cm x 0.1cm x 4.1cm, and the space of the fracture in the middle is 0.1 cm. The original properties of matrix, brine, oil in the base case are from Ekofisk Field. The properties of surfactant are from literature. Eclipse is used as the simulator. The results are followings: 1) The viscosity of surfactant solution has a positive linear relationship with surfactant oil recovery time. And the relationship between viscosity and oil production rate is an inverse function. The viscosity of surfactant solution has no obvious effect on ultimate oil recovery. Since most of the surfactant has no big effect on viscosity of brine, the viscosity of surfactant solution is not a key parameter of surfactant screening for surfactant flooding in fractured reservoirs. 2) The increase of surfactant concentration results a decrease of oil recovery rate and an increase of ultimate oil recovery. However, there are no functions could describe the relationships. Study on economy should be conducted because of the price of surfactant and oil. 3) In the study of surfactant adsorption, assume that the matrix wettability is changed to water-wet when the surfactant adsorption is to the maximum at all cases. And the ratio of surfactant adsorption and surfactant concentration (Cads/Csurf) is used to estimate the functional relationship. The results show that the relationship between ultimate oil recovery and Cads/Csurf is a logarithmic function. The oil production rate has a positive linear relationship with exp(Cads/Csurf). The work here could be used as a reference for the surfactant screening of surfactant enhanced oil recovery from fractured reservoirs. And the functional relationships between surfactant properties and the oil recovery rate and ultimate oil recovery help to improve upscaling methods.

Keywords: fractured reservoirs, surfactant adsorption, surfactant concentration, surfactant EOR, surfactant viscosity

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