Search results for: protein complementation assay

Authors: Susan Hall, Gary D. Grant, Catherine McDermott, Devinder Arora

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Introduction /Aim: The kynurenine pathway is thought to play an important role in the pathophysiology of numerous neurodegenerative diseases including depression, Alzheimer’s disease, and Parkinson’s disease. Numerous neuroactive compounds, including the neurotoxic 3-hydroxyanthranilic acid, 3-hydroxykynurenine and quinolinic acid and the neuroprotective kynurenic acid and picolinic acid, are produced through the metabolism of kynurenine and are thought to be the causative agents responsible for neurodegeneration. The toxicity of 3-hydroxykynurenine, 3-hydroxyanthranilic acid and quinolinic acid has been widely evaluated and demonstrated in primary cell cultures but to date only 3-hydroxykynurenine and 3-hydroxyanthranilic acid have been shown to cause toxicity in immortal tumour cells. The aim of this study was to evaluate the toxicity of kynurenine metabolites, both individually and in combination, on SH-SY5Y neuroblastoma cells after 24 and 72 h exposure in order to explore a cost-effective model to study their neurotoxic effects and potential protective agents. Methods: SH-SY5Y neuroblastoma cells were exposed to various concentrations of the neuroactive kynurenine metabolites, both individually and in combination, for 24 and 72 h, and viability was subsequently evaluated using the Resazurin (Alamar blue) proliferation assay. Furthermore, the effects of these compounds, alone and in combination, on specific death pathways including apoptosis, necrosis and free radical production was evaluated using various assays. Results: Consistent with literature, toxicity was shown with short-term 24-hour treatments at 1000 μM concentrations for both 3-hydroxykynurenine and 3-hydroxyanthranilic acid. Combinations of kynurenine metabolites showed modest toxicity towards SH-SY5Y neuroblastoma cells in a concentration-dependent manner. Specific cell death pathways, including apoptosis, necrosis and free radical production were shown to be increased after both 24 and 72 h exposure of SH-SY5Y neuroblastoma cells to 3-hydroxykynurenine and 3-hydroxyanthranilic acid and various combinations of neurotoxic kynurenine metabolites. Conclusion: It is well documented that neurotoxic kynurenine metabolites show toxicity towards primary human neurons in the nanomolar to low micromolar concentration range. Results show that the concentrations required to show significant cell death are in the range of 1000 µM for 3-hydroxykynurenine and 3-hydroxyanthranilic acid and toxicity of quinolinic acid towards SH-SY5Y was unable to be shown. This differs significantly from toxicities observed in primary human neurons. Combinations of the neurotoxic metabolites were shown to have modest toxicity towards these cells with increased toxicity and activation of cell death pathways observed after 72 h exposure. This study suggests that the 24 h model is unsuitable for use in neurotoxicity studies, however, the 72 h model better represents the observations of the studies using primary human neurons and may provide some benefit in providing a cost-effective model to assess possible protective agents against kynurenine metabolite toxicities.

Keywords: kynurenine metabolites, neurotoxicity, quinolinic acid, SH-SY5Y neuroblastoma

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Authors: Venencia Albert, Arulselvi Subramanian, Hara Prasad Pati, Asok K. Mukhophadhyay

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Acute traumatic coagulopathy in an endogenous dysregulation of the intrinsic coagulation system in response to the injury, associated with three-fold risk of poor outcome, and is more amenable to corrective interventions, subsequent to early identification and management. Multiple definitions for stratification of the patients' risk for early acute coagulopathy have been proposed, with considerable variations in the defining criteria, including several trauma-scoring systems based on prehospital data. We aimed to develop a clinically relevant definition for acute coagulopathy of trauma based on conventional coagulation assays and to assess its efficacy in comparison to recently established prehospital prediction models. Methodology: Retrospective data of all trauma patients (n = 490) presented to our level I trauma center, in 2014, was extracted. Receiver operating characteristic curve analysis was done to establish cut-offs for conventional coagulation assays for identification of patients with acute traumatic coagulopathy was done. Prospectively data of (n = 100) adult trauma patients was collected and cohort was stratified by the established definition and classified as "coagulopathic" or "non-coagulopathic" and correlated with the Prediction of acute coagulopathy of trauma score and Trauma-Induced Coagulopathy Clinical Score for identifying trauma coagulopathy and subsequent risk for mortality. Results: Data of 490 trauma patients (average age 31.85±9.04; 86.7% males) was extracted. 53.3% had head injury, 26.6% had fractures, 7.5% had chest and abdominal injury. Acute traumatic coagulopathy was defined as international normalized ratio ≥ 1.19; prothrombin time ≥ 15.5 s; activated partial thromboplastin time ≥ 29 s. Of the 100 adult trauma patients (average age 36.5±14.2; 94% males), 63% had early coagulopathy based on our conventional coagulation assay definition. Overall prediction of acute coagulopathy of trauma score was 118.7±58.5 and trauma-induced coagulopathy clinical score was 3(0-8). Both the scores were higher in coagulopathic than non-coagulopathic patients (prediction of acute coagulopathy of trauma score 123.2±8.3 vs. 110.9±6.8, p-value = 0.31; trauma-induced coagulopathy clinical score 4(3-8) vs. 3(0-8), p-value = 0.89), but not statistically significant. Overall mortality was 41%. Mortality rate was significantly higher in coagulopathic than non-coagulopathic patients (75.5% vs. 54.2%, p-value = 0.04). High prediction of acute coagulopathy of trauma score also significantly associated with mortality (134.2±9.95 vs. 107.8±6.82, p-value = 0.02), whereas trauma-induced coagulopathy clinical score did not vary be survivors and non-survivors. Conclusion: Early coagulopathy was seen in 63% of trauma patients, which was significantly associated with mortality. Acute traumatic coagulopathy defined by conventional coagulation assays (international normalized ratio ≥ 1.19; prothrombin time ≥ 15.5 s; activated partial thromboplastin time ≥ 29 s) demonstrated good ability to identify coagulopathy and subsequent mortality, in comparison to the prehospital parameter-based scoring systems. Prediction of acute coagulopathy of trauma score may be more suited for predicting mortality rather than early coagulopathy. In emergency trauma situations, where immediate corrective measures need to be taken, complex multivariable scoring algorithms may cause delay, whereas coagulation parameters and conventional coagulation tests will give highly specific results.

Keywords: trauma, coagulopathy, prediction, model

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Authors: Menna Ewida, Dalal Abou El-Ella, Dina Lasheen, Huessin El-Subbagh

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Introduction: Vascular Endothelial Growth Factor receptor (VEGF) is a signal protein produced by cells that stimulates vasculogenesis to create new blood vessels. VEGF family binds to three trans-membrane tyrosine kinase receptors,Dihydrofolate reductase (DHFR) is an enzyme of crucial importance in medicinal chemistry. DHFR catalyzes the reduction 7,8 dihydro-folate to tetrahydrofolate and intimately couples with thymidylate synthase which is a pivotal enzyme that catalysis the reductive methylation of deoxyuridine monophosphate (dUMP) to deoxythymidine monophosphate (dTMP) utilizing N5,N10-methylene tetrahydrofolate as a cofactor which functions as the source of the methyl group. Purpose: Novel substituted Thiazole agents were designed as DHFR and VEGF-TK inhibitors with increased synergistic activity and decreased side effects. Methods: Five series of compounds were designed with a rational that mimic the pharmacophoric features present in the reported active compounds that target DHFR & VEGFR. These molecules were docked against Methotrexate & Sorafenib as controls. An in silico ADMET study was also performed to validate the bioavailability of the newly designed compounds. The in silico molecular docking & ADMET study were also applied to the non-classical antifolates for comparison. The interaction energy comparable to that of MTX for DHFRI and Sorafenib for VEGF-TKI activity were recorded. Results: Compound 5 exhibited the highest interaction energy when docked against Sorafenib, While Compound 9 showed the highest interaction energy when docked against MTX with the perfect binding mode. Comparable results were also obtained for the ADMET study. Most of the compounds showed absorption within (95-99) zone which varies according to the type of substituents. Conclusions: The Substituted Thiazole Analogues could be a suitable template for antitumor drugs that possess enhanced bioavailability and act as DHFR and VEGF-TK inhibitors.

Keywords: anti-tumor agents, DHFR, drug design, molecular modeling, VEGFR-TKIs

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Authors: Min Liu, Jirui Gong, Qinpu Luo, Lili Yang, Bo Yang, Zihe Zhang, Yan Pan, Zhanwei Zhai

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Grazing disturbance is one of the important land-use types that affect plant growth and ecosystem processes. In order to study the responses of dominant species to grazing in the semiarid temperate grassland of Inner Mongolia, we set five grazing intensity plots: a control and four levels of grazing (light (LG), moderate (MG), heavy (HG) and extreme heavy grazing (EHG)) to test the morphological and physiological responses of Stipa grandis, Leymus chinensis at the individual levels. With the increase of grazing intensity, Stipa grandis and Leymus chinensis both exhibited reduced plant height, leaf area, stem length and aboveground biomass, showing a significant dwarf phenomenon especially in HG and EHG plots. The photosynthetic capacity decreased along the grazing gradient. Especially in the MG plot, the two dominant species have lowest net photosynthetic rate (Pn) and water use efficiency (WUE). However, in the HG and EHG plots, the two species had high light saturation point (LSP) and low light compensation point (LCP), indicating they have high light-use efficiency. They showed a stimulation of compensatory photosynthesis to the remnant leaves as compared with grasses in MG plot. For Leymus chinensis, the lipid peroxidation level did not increase with the low malondialdehyde (MDA) content even in the EHG plot. It may be due to the high enzymes activity of superoxide dismutase (SOD) and peroxidase (POD) to reduce the damage of reactive oxygen species. Meanwhile, more carbohydrate was stored in the leaf of Leymus chinensis to provide energy to the plant regrowth. On the contrary, Stipa grandis showed the high level of lipid peroxidation especially in the HG and EHG plots with decreased antioxidant enzymes activity. The soluble protein content did not change significantly in the different plots. Therefore, with the increase of grazing intensity, plants changed morphological and physiological traits to defend themselves effectively to herbivores. Leymus chinensis is more resistant to grazing than Stipa grandis in terms of tolerance traits, particularly under heavy grazing pressure.

Keywords: antioxidant enzymes activity, grazing density, morphological responses, photosynthesis

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Authors: O. A. Adebiyi, A. O. Oni, A. O. K. Adeshehinwa, I. O. Adejumo

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In forty nine (49) days, twenty four (24) growing pigs (Landrace x Large white) with an average weight of 17 ±2.1kg were allocated to four experimental treatments T1 (dry mash without probiotics), T2 (wet feed without probiotics), T3 (dry mash + Saccaromyces cereviseae probiotics) and T4 (wet feed + Saccaromyces cereviseae probiotics) which were replicated three times with two pigs per replicate in a completely randomised design. The basal feed (dry feed) was formulated to meet the nutritional requirement of the animal with crude protein of 18.00% and metabolisable energy of 2784.00kcal/kgME. Growth pattern, faecal microbial load and behavioural activities (eating, drinking, physical pen interaction and frequency of visiting the drinking troughs) were accessed. Pigs fed dry mash without probiotics (T1) had the highest daily feed intake among the experimental animals (1.10kg) while pigs on supplemented diets (T3 and T4) had an average daily feed intake of 0.95kg. However, the feed conversion ratio was significantly (p < 0.05) affected with pigs on T3 having least value of 6.26 compared those on T4 (wet feed + Saccaromyces cereviseae) with means of 7.41. Total organism counts varied significantly (p < 0.05) with pigs on T1, T2, T3 and T4 with mean values of 179.50 x106cfu; 132.00 x 106cfu; 32.00 x 106cfu and 64.50 x 106cfu respectively. Coliform count was also significantly (p < 0.05) different among the treatments with corresponding values of 117.50 x 106cfu; 49.00 x 106cfu, 8.00 x 106cfu for pigs in T1, T2 and T4 respectively. The faecal Saccaromyces cereviseae was significantly lower in pigs fed supplemented diets compared to their counterparts on unsupplemented diets. This could be due to the inability of yeast organisms to be voided easily through feaces. The pigs in T1 spent the most time eating (7.88%) while their counterparts on T3 spent the least time eating. The corresponding physical pen interaction times expressed in percentage of a day for pigs in T1, T2, T3 and T4 are 6.22%, 5.92%, 4.04% and 4.80% respectively. These behavioural responses exhibited by these pigs (T3) showed that little amount of dry feed supplemented with probiotics is needed for better performance. The water intake increases as a result of the dryness of the feed with consequent decrease in pen interaction and more time was spent resting than engaging in other possible vice-habit like fighting or tail biting. Pigs fed dry feed (T3) which was supplemented with Saccaromyces cereviseae probiotics had a better overall performance, least faecal microbial load than wet fed pigs either supplemented with Saccaromyces cereviseae or non-supplemented.

Keywords: behaviour, feed forms, feed utilization, growth, microbial

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Authors: Manal Elsayed Elkheshin, Rasha Saleh Mansour, Mohamed Fawzy Mohamed Eldnasury, Mamdouh Elbadry Mohamed

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This research Aims to identify the main factors affecting the production and the fish consumption in Egypt, through the econometric estimation for various forms functions of fish production and fish consumption during the period (1991-2014), as the aim of this research to forecast the production and the fish consumption in Egypt until 2020, through determine the best standard methods using (ARIMA).This research also aims to the economic feasibility of the production of fish in aquaculture farms study; investment cost and represents the value of land, buildings, equipment and irrigation. Aquaculture requires three types of fish (Tilapia, carp fish, and mullet fish), and the total area of the farm, about an acre. The annual Fish production from this project about 3.5 tons. The annual investment costs of about 50500 pounds, Find conclude that the project can repay the cost of their investments after about 4 years and 5 months, and therefore recommend the implementation of the project, and internal rate of return reached (IRR) of about 22.1%, where it is clear that the rate of large internal rate of return, and achieves pound invested in this project annual return is estimated at 22.1 pounds, more than the opportunity cost, so we recommend the need to implement the project.Recommendations:1. Increasing the fish agriculture to decrease the gap of animal protein. 2.Increasing the number of mechanism fishing boats, and the provision of transport equipped to maintain the quality of fish production. 3.Encourage and attract the local and foreign investments, providing advice to the investor on the aquaculture field. 4. Action newsletters awareness of the importance of these projects where these projects resulted in a net profit after recovery in less than five years, IRR amounted to about 23%, which is much more than the opportunity cost of a bank interest rate is about 7%, helping to create work and graduates opportunities, and contribute to the reduction of imports of the fish, and improve the performance of the food trade balance.

Keywords: equation model, individual share, red meat, consumption, production, endogenous variable, exogenous variable, financial performance evaluates fish culture, feasibility study, fish production, aquaculture

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Authors: Francesca Lombardi, Francesca Rosaria Augello, Benedetta Cinque, Maria Grazia Cifone, Paola Palumbo

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Glioblastoma multiforme (GBM) is a high-grade primary brain tumor refractory to current forms of treatment. The survival benefits of patients with GBM remain unsatisfactory due to the intrinsic or acquired resistance to temozolomide (TMZ), an alkylating agent, used as the first-line chemotherapeutic drug to treat GBM patients. Its cytotoxic effect is visualized by the induction of O6-methylguanine (O6MeG) within DNA. Cyclooxygenase-2 (COX-2), an inflammation-associated enzyme, has been implicated in tumorigenesis and progression of GBM, its inhibition shows anticancer activities. In the present study, it was verified if the combination of a COX-2 selective inhibitor, NS398, with TMZ could counteract the TMZ resistance. In particular, the effect of NS398 mixed with TMZ was investigated in the GBM TMZ-resistant cell line, T98G. Cells were pretreated with NS398 (100µM, 24 hours) and then exposed to TMZ alone (200µM), NS398 alone, or both for 72 hours, after which cell growth rate and cycle phases, as well as apoptosis level, were evaluated. Coadministration of NS398 and TMZ caused a significant decrease in cell growth and a progressive increase of dead cells detected by trypan blue staining. Moreover, a significant level of apoptotic cell percentage and alteration of cell cycle phases were observed in T98G treated with TMZ-NS398 combination when compared to untreated cells or TMZ-treated cells. TMZ-resistant tumors, as GBM, express elevated levels of DNA repair enzyme O6-methylguanine-DNA methyltransferase (MGMT). The mixture drastically reduced MGMT expression in the TMZ-resistant cell line T98G, known to express high levels of MGMT basically. Moreover, while TMZ alone did not influence the COX-2 protein expression, the combination successfully reduced it. In conclusion, these results demonstrated that NS398 enhanced the efficacy of TMZ through cell number reduction, apoptosis induction, and decreased MGMT levels, suggesting the ability of drug combination to reduce the chemoresistance. This drug combination deserves attention and could be considered as a promising therapeutic strategy for GBM patients.

Keywords: COX-2, COX-2 inhibitor, glioblastoma, NS398, T98G, temozolomide

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Authors: Delgado-Meza M., Minor-Pérez H.

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Protease is the generic name of enzymes that hydrolyze proteins. These are classified in the subgroup EC3.4.11-99X of the classification enzymes. In food technology the proteolysis is used to modify functional and nutritional properties of food, and in some cases this proteolysis may cause food spoilage. In general, seafood and rainbow trout have accelerated decomposition process once it has done its capture, due to various factors such as the endogenous enzymatic activity that can result in loss of structure, shape and firmness, besides the release of amino acid precursors of biogenic amines. Some studies suggest the use of protease inhibitors from legume as biological regulators of proteolytic activity. The enzyme inhibitors are any substance that reduces the rate of a reaction catalyzed by an enzyme. The objective of this study was to evaluate the reduction of the proteolytic activity of enzymes in extracts of rainbow trout with protease inhibitors obtained from chickpea flour. Different proportions of rainbow trout enzyme extract (75%, 50% and 25%) and extract chickpea enzyme inhibitors were evaluated. Chickpea inhibitors were obtained by mixing 5 g of flour in 30 mL of pH 7.0 phosphate buffer. The sample was centrifuged at 8000 rpm for 10 min. The supernatant was stored at -15°C. Likewise, 20 g of rainbow trout were ground in 20 mL of phosphate buffer solution at pH 7.0 and the mixture was centrifuged at 5000 rpm for 20 min. The supernatant was used for the study. In each treatment was determined the specific enzymatic activity with the technique of Kunitz, using hemoglobin as substrate for the enzymes acid fraction and casein for basic enzymes. Also biuret protein was quantified for each treatment. The results showed for fraction of basic enzymes in the treatments evaluated, that were inhibition of endogenous enzymatic activity. Inhibition values compared to control were 51.05%, 56.59% and 59.29% when the proportions of endogenous enzymes extract rainbow trout were 75%, 50% and 25% and the remaining volume used was extract with inhibitors. Treatments with acid enzymes showed no reduction in enzyme activity. In conclusion chickpea flour reduced the endogenous enzymatic activity of rainbow trout, which may favor its application to increase the half-life of this food. The authors acknowledge the funding provided by the CONACYT for the project 131998.

Keywords: rainbouw trout, enzyme inhibitors, proteolysis, enzyme activity

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Authors: Rosa Figueroa, Christopher Flores

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Text categorization is the problem of assigning a new document to a set of predetermined categories, on the basis of a training set of free-text data that contains documents whose category membership is known. To train a classification model, it is necessary to extract characteristics in the form of tokens that facilitate the learning and classification process. In text categorization, the feature extraction process involves the use of word sequences also known as N-grams. In general, it is expected that documents belonging to the same category share similar features. The Smith-Waterman (SW) algorithm is a dynamic programming algorithm that performs a local sequence alignment in order to determine similar regions between two strings or protein sequences. This work explores the use of SW algorithm as an alternative to feature extraction in text categorization. The dataset used for this purpose, contains 2,610 annotated documents with the classes Obese/Non-Obese. This dataset was represented in a matrix form using the Bag of Word approach. The score selected to represent the occurrence of the tokens in each document was the term frequency-inverse document frequency (TF-IDF). In order to extract features for classification, four experiments were conducted: the first experiment used SW to extract features, the second one used unigrams (single word), the third one used bigrams (two word sequence) and the last experiment used a combination of unigrams and bigrams to extract features for classification. To test the effectiveness of the extracted feature set for the four experiments, a Support Vector Machine (SVM) classifier was tuned using 20% of the dataset. The remaining 80% of the dataset together with 5-Fold Cross Validation were used to evaluate and compare the performance of the four experiments of feature extraction. Results from the tuning process suggest that SW performs better than the N-gram based feature extraction. These results were confirmed by using the remaining 80% of the dataset, where SW performed the best (accuracy = 97.10%, weighted average F-measure = 97.07%). The second best was obtained by the combination of unigrams-bigrams (accuracy = 96.04, weighted average F-measure = 95.97) closely followed by the bigrams (accuracy = 94.56%, weighted average F-measure = 94.46%) and finally unigrams (accuracy = 92.96%, weighted average F-measure = 92.90%).

Keywords: comorbidities, machine learning, obesity, Smith-Waterman algorithm

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Authors: Yee-Seul So, Guiseppe Ianiri, Alex Idnurm, Yong-Sun Bahn

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Tor1 is a serine/threonine protein kinase that is widely conserved across eukaryotic species. Tor1 was first identified in Saccharomyces cerevisiae as a target of rapamycin (TOR). The TOR pathway has been implicated in regulating cellular responses to nutrients, proliferation, translation, transcription, autophagy, and ribosome biogenesis. Here we identified two homologues of S. cerevisiae Tor proteins, CNAG_06642 (Tor1) and CNAG_05220 (Tlk1, TOR-like kinase 1), in Cryptococcus neoformans causing a life-threatening fungal meningoencephalitis. Both Tor1 and Tlk1 have rapamycin-binding (RB) domains but Tlk1 has truncated RB form. To study the TOR-signaling pathway in the fungal pathogen, we attempt to construct the tor1Δ and tlk1Δ mutants and phenotypically analyze them. Although we failed to construct the tor1Δ mutant, we successfully construct the tlk1Δ mutant. The tlk1Δ mutant does not exhibit any discernable phenotypes, suggesting that Tlk1 is dispensable in C. neoformans. The essentiality of TOR1 is independently confirmed by constructing the TOR1 promoter replacement strain by using a copper transporter 4 (CTR4) promoter and the TOR1/tor1 heterozygous mutant in diploid C. neoformans strain background followed by sporulation analysis. To further analyze the function of Tor1, we construct TOR1 overexpression mutant using a constitutively active histone H3 in C. neoformans. We find that the Tor1 overexpression mutant is resistant to rapamycin but the tlk1Δ mutant does not exhibit any altered resistance to rapamycin, further confirming that Tor1, but not Tlk1, is critical for TOR signaling. Furthermore, we found that Tor1 is involved in response to diverse stresses, including genotoxic stress, oxidative stress, thermo-stress, antifungal drug treatment, and production of melanin. To identify any TOR-related transcription factors, we screened C. neoformans transcription factor library that we constructed in our previous study and identified several potential downstream factors of Tor1, including Atf1, Crg1 and Bzp3. In conclusion, the current study provides insight into the role of the TOR signaling pathway in human fungal pathogens as well as C. neoformans.

Keywords: fungal pathogen, serine/threonine kinase, target of rapamycin, transcription factor

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Authors: E. Obreque-Slier, P. Osorio-Umaña, G. Vidal-Acevedo, A. Peña-Neira, M. Medel-Marabolí

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Sensory and nutraceutical characteristics of a wine are determined by different chemical compounds, such as organic acids, sugars, alcohols, polysaccharides, aromas, and polyphenols. The polyphenols correspond to secondary metabolites that are associated with the prevention of several pathologies, and those are responsible for color, aroma, bitterness, and astringency in wines. These compounds come from grapes and wood during aging in barrels, which correspond to the format of wood most widely used in wine production. However, the barrels is a high-cost input with a limited useful life (3-4 years). For this reason, some oenological products have been developed in order to renew the barrels and increase their useful life in some years. These formats are being used slowly because limited information exists about the effect on the wine chemical characteristics. The objective of the study was to evaluate the effect of different laubarrel renewal systems (staves and zigzag) on the polyphenolic characteristics of a Carménère wine (Vitis vinifera), an emblematic cultivar of Chile. For this, a completely randomized experimental design with 5 treatments and three replicates per treatment was used. The treatments were: new barrels (T0), used barrels during 4 years (T1), scraped used barrels (T2), used barrels with staves (T3) and used barrels with zigzag (T4). The study was performed for 12 months, and different spectrophotometric parameters (phenols, anthocyanins, and total tannins) and HPLC-DAD (low molecular weight phenols) were evaluated. The wood inputs were donated by Toneleria Nacional and corresponded to products from the same production batch. The total phenols content increased significantly after 40 days, while the total tannin concentration decreased gradually during the study. The anthocyanin concentration increased after 120 days of the assay in all treatments. Comparatively, it was observed that the wine of T2 presented the lowest values of these polyphenols, while the T0 and T4 presented the highest total phenol contents. Also, T1 presented the highest values of total tannins in relation to the rest of the treatments in some samples. The low molecular weight phenolic compounds identified by HPLC-DAD were 7 flavonoids (epigallocatechin, catechin, procyanidin gallate, epicatechin, quercetin, rutin and myricetin) and 14 non-flavonoids (gallic, protocatechuic, hydroxybenzoic, trans-cutaric, vanillinic, caffeic, syringic, p-coumaric and ellagic acids; tyrosol, vanillin, syringaldehyde, trans-resveratrol and cis-resveratrol). Tyrosol was the most abundant compound, whereas ellagic acid was the lowest in the samples. Comparatively, it was observed that the wines of T2 showed the lowest concentrations of flavonoid and non-flavonoid phenols during the study. In contrast, wines of T1, T3, and T4 presented the highest contents of non-flavonoid polyphenols. In summary, the use of barrel renovators (zig zag and staves) is an interesting alternative which would emulate the contribution of polyphenols from the barrels to the wine.

Keywords: barrels, oak wood aging, polyphenols, red wine

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Authors: Hamdy Khamis Korayem, Manal Yehia Tayel, Abeer Shawky El Hadedy, Emmanuel Kamal Aziz Saba, Shimaa Badr Abdelnaby Badr

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The aim of the current study was to demonstrate whether serum Interleukin-18 (IL-18) is increased in rheumatoid arthritis (RA) and its correlation with disease activity, functional disability and quality of life in RA patients. The study included 30 RA patients and 20 healthy normal control subjects. The RA patients were diagnosed according to the 2010 ACR/EULAR classification criteria for RA with the exclusion of those who had diabetes mellitus, endocrine disorders, associated rheumatologic diseases, viral hepatitis B or C and other diseases with increased serum IL-18 level. All patients were subjected to clinical evaluation of the musculoskeletal system. Disease activity was assessed by disease activity score 28 with 4 variables (DAS 28). Functional disability was assessed by health assessment questionnaire disability index (HAQ-DI). The quality of life was assessed by Short form-36 (SF-36) questionnaire. Radiological assessment of both hands and feet by Sharp/van der Heijde (SvH) scoring method. Laboratory parameters including erythrocyte sedimentation rate (ESR), C-reactive protein (CRP), rheumatoid factor (RF) and anti-cyclic citrullinated peptide antibody (ACPA) were assessed in patients and serum level of IL-18 in both patients and control subjects. There was no statistically significant difference between patient and control group as regards age and sex. Among patients, 29 % were females and the age range was between 25 to 55 years. Extra-articular manifestations were presented in 56.7% of the patients. The mean of DAS 28 score was 5.73±1.46 and that of HAQ-DI was 1.22±0.72 while that of SF-36 was 40.03±13.96. The level of serum IL-18 was significantly higher in patients than in the control subjects (P= 0.030). Serum IL-18 was correlated with ACPA among the patient group. There were no statistically significant correlations between serum IL-18 and DAS28, HAQ-DI, SF-36, total SvH score and the other laboratory results. In conclusion, IL-18 is significantly higher in RA patient than in healthy control subjects and positively correlated with ACPA level. IL-18 is associated with extra-articular manifestations. However, it is not correlated with other laboratory parameters, disease activity, functional disability, quality of life nor radiological severity.

Keywords: disease activity score, Interleukin-18, quality of life assessment, rheumatoid arthritis

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Authors: Iwona Cieślik, Władysław Migdał, Kinga Topolska, Ewa Cieślik

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The consumption of fish is recommended as a means of preventing serious diseases, especially cardiovascular problems. Fish is known to be a valuable source of protein (rich in essential amino acids), unsaturated fatty acids, fat-soluble vitamins, macro- and microelements. However, it can also contain several contaminants (e.g. pesticides, heavy metals) that may pose considerable risks for humans. Among others, pesticide are of special concern. Their widespread use has resulted in the contamination of environmental compartments, including water. The occurrence of pesticides in the environment is a serious problem, due to their potential toxicity. Therefore, a systematic monitoring is needed. The aim of the study was to determine the organochlorine and organophosphate pesticide residues in fish muscle tissues of the pike (Esox lucius, L.) and the rainbow trout (Oncorhynchus mykkis, Walbaum) by a modified QuEChERS (Quick, Easy, Cheap, Effective, Rugged and Safe) method, using Gas Chromatography Quadrupole Mass Spectrometry (GC/Q-MS), working in selected-ion monitoring (SIM) mode. The analysis of α-HCH, β-HCH, lindane, diazinon, disulfoton, δ-HCH, methyl parathion, heptachlor, malathion, aldrin, parathion, heptachlor epoxide, γ-chlordane, endosulfan, α-chlordane, o,p'-DDE, dieldrin, endrin, 4,4'-DDD, ethion, endrin aldehyde, endosulfan sulfate, 4,4'-DDT, and metoxychlor was performed in the samples collected in the Carp Valley (Malopolska region, Poland). The age of the pike (n=6) was 3 years and its weight was 2-3 kg, while the age of the rainbow trout (n=6) was 0.5 year and its weight was 0.5-1.0 kg. Detectable pesticide (HCH isomers, endosulfan isomers, DDT and its metabolites as well as metoxychlor) residues were present in fish samples. However, all these compounds were below the limit of quantification (LOQ). The other examined pesticide residues were below the limit of detection (LOD). Therefore, the levels of contamination were - in all cases - below the default Maximum Residue Levels (MRLs), established by Regulation (EC) No 396/2005 of the European Parliament and of the Council. The monitoring of pesticide residues content in fish is required to minimize potential adverse effects on the environment and human exposure to these contaminants.

Keywords: contaminants, fish, pesticides residues, QuEChERS method

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Authors: Amna Shoaib, Sidrah Hanif, Rashid Mehmood

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Current research work was carried out to check influence of farmyard manure (FYM) in Lycopersicon esculentum L. against Fusarium oxysporum f. sp. lycopersici (FO) in copper polluted soil. Silt-loam soil naturally enriched with 70 ppm of Cu was inoculated with 1 x 106 spore suspensions of FO and incorporated with 0%, 1%, 1.5% or 2% FYM. The multilateral interaction of host-pathogen-metal-organic amendment was assessed in terms of morphology, growth, yield, physiology, biochemistry and metal uptake in tomato plant after 30 and 60 days of sowing. When soil was inoculated with FO, plant growth and biomass were significantly increased during vegetative stage, while declining during flowering stage with substantial increase in productivity over control. Infected plants exhibited late wilting and disease severity was found on 26-50% of plant during reproductive stage. Incorporation of up to 1% FYM suppressed disease severity, improved plant growth and biomass, while it decreased yield. Rest of manure doses was found ineffective in suppressing disease. Content of total chlorophyll, sugar and protein were significantly declined in FO inoculated plants and incorporation of FYM caused significant reduction or no influence on sugar and chlorophyll content, and no pronounced difference among different FYM doses were observed. On the other hand, proline, peroxidase, catalase and nitrate reductase activity were found to be increased in infected plants and incorporation of 1-2% FYM further enhanced the activity of these enzymes. Tomato plant uptake of 30-40% of copper naturally present in the soil and incorporation of 1-2% FYM markedly decreased plant uptake of metal by 15-30%, while increased Cu retention in soil. Present study concludes that lower dose (1%) of FYM could be used to manage disease, increase growth and biomass, while being ineffective for yield and productivity in Cu-polluted soil. Altered physiology/biochemistry of plant in response to any treatment could be served as basis for resistant against pathogen and metal homeostasis in plants.

Keywords: Lycopersicon esculentum, copper, Fusarium wilt, farm yard manure

Procedia PDF Downloads 386

Authors: Elisabetta Ceretti, Silvia Bonizzoni, Alberto Bonetti, Milena Villarini, Marco Verani, Maria Antonella De Donno, Sara Bonetta, Umberto Gelatti

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Introduction: Air pollution is a global problem. In 2013, the International Agency for Research on Cancer (IARC) classified air pollution and particulate matter as carcinogenic to human. The study of the health effects of air pollution in children is very important because they are a high-risk group in terms of the health effects of air pollution and early exposure during childhood can increase the risk of developing chronic diseases in adulthood. The MAPEC_LIFE (Monitoring Air Pollution Effects on Children for supporting public health policy) is a project founded by EU Life+ Programme which intends to evaluate the associations between air pollution and early biological effects in children and to propose a model for estimating the global risk of early biological effects due to air pollutants and other factors in children. Methods: The study was carried out on 6-8-year-old children living in five Italian towns in two different seasons. Two biomarkers of early biological effects, primary DNA damage detected with the comet assay and frequency of micronuclei, were investigated in buccal cells of children. Details of children diseases, socio-economic status, exposures to other pollutants and life-style were collected using a questionnaire administered to children’s parents. Child exposure to urban air pollution was assessed by analysing PM0.5 samples collected in the school areas for PAHs and nitro-PAHs concentration, lung toxicity and in vitro genotoxicity on bacterial and human cells. Data on the chemical features of the urban air during the study period were obtained from the Regional Agency for Environmental Protection. The project created also the opportunity to approach the issue of air pollution with the children, trying to raise their awareness on air quality, its health effects and some healthy behaviors by means of an educational intervention in the schools. Results: 1315 children were recruited for the study and participate in the first sampling campaign in the five towns. The second campaign, on the same children, is still ongoing. The preliminary results of the tests on buccal mucosa cells of children will be presented during the conference as well as the preliminary data about the chemical composition and the toxicity and genotoxicity features of PM0.5 samples. The educational package was tested on 250 children of the primary school and showed to be very useful, improving children knowledge about air pollution and its effects and stimulating their interest. Conclusions: The associations between levels of air pollutants, air mutagenicity and biomarkers of early effects will be investigated. A tentative model to calculate the global absolute risk of having early biological effects for air pollution and other variables together will be proposed and may be useful to support policy-making and community interventions to protect children from possible health effects of air pollutants.

Keywords: air pollution exposure, biomarkers of early effects, children, public health policy

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Authors: Djeddi Khaled, Houssou Hind, Miloudi Abdellatif, Rabah Siham

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In the field of veterinary medicine, there is a growing application of artificial intelligence (AI) for diagnosing bovine mastitis, a prevalent inflammatory disease in dairy cattle. AI technologies, such as automated milking systems, have streamlined the assessment of key metrics crucial for managing cow health during milking and identifying prevalent diseases, including mastitis. These automated milking systems empower farmers to implement automatic mastitis detection by analyzing indicators like milk yield, electrical conductivity, fat, protein, lactose, blood content in the milk, and milk flow rate. Furthermore, reports highlight the integration of somatic cell count (SCC), thermal infrared thermography, and diverse systems utilizing statistical models and machine learning techniques, including artificial neural networks, to enhance the overall efficiency and accuracy of mastitis detection. According to a review of 15 publications, machine learning technology can predict the risk and detect mastitis in cattle with an accuracy ranging from 87.62% to 98.10% and sensitivity and specificity ranging from 84.62% to 99.4% and 81.25% to 98.8%, respectively. Additionally, machine learning algorithms and microarray meta-analysis are utilized to identify mastitis genes in dairy cattle, providing insights into the underlying functional modules of mastitis disease. Moreover, AI applications can assist in developing predictive models that anticipate the likelihood of mastitis outbreaks based on factors such as environmental conditions, herd management practices, and animal health history. This proactive approach supports farmers in implementing preventive measures and optimizing herd health. By harnessing the power of artificial intelligence, the diagnosis of bovine mastitis can be significantly improved, enabling more effective management strategies and ultimately enhancing the health and productivity of dairy cattle. The integration of artificial intelligence presents valuable opportunities for the precise and early detection of mastitis, providing substantial benefits to the dairy industry.

Keywords: artificial insemination, automatic milking system, cattle, machine learning, mastitis

Procedia PDF Downloads 32

Authors: Jan Masak, Eva Kvasnickova, Vladimir Scholtz, Olga Matatkova, Marketa Valkova, Alena Cejkova

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Microbial colonization of medical instruments, catheters, implants, etc. is a serious problem in the spread of nosocomial infections. Biofilms exhibit enormous resistance to environment. The resistance of biofilm populations to antibiotic or biocides often increases by two to three orders of magnitude in comparison with suspension populations. Subjects of interests are substances or physical processes that primarily cause the destruction of biofilm, while the released cells can be killed by existing antibiotics. In addition, agents that do not have a strong lethal effect do not cause such a significant selection pressure to further enhance resistance. Non-thermal plasma (NTP) is defined as neutral, ionized gas composed of particles (photons, electrons, positive and negative ions, free radicals and excited or non-excited molecules) which are in permanent interaction. In this work, the effect of NTP generated by the cometary corona with a metallic grid on the formation and stability of biofilm and metabolic activity of cells in biofilm was studied. NTP was applied on biofilm populations of Staphylococcus epidermidis DBM 3179, Pseudomonas aeruginosa DBM 3081, DBM 3777, ATCC 15442 and ATCC 10145, Escherichia coli DBM 3125 and Candida albicans DBM 2164 grown on solid media on Petri dishes and on the titanium alloy (Ti6Al4V) surface used for the production joint replacements. Erythromycin (for S. epidermidis), polymyxin B (for E. coli and P. aeruginosa), amphotericin B (for C. albicans) and ceftazidime (for P. aeruginosa) were used to study the combined effect of NTP and antibiotics. Biofilms were quantified by crystal violet assay. Metabolic activity of the cells in biofilm was measured using MTT (3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyl tetrazolium bromide) colorimetric test based on the reduction of MTT into formazan by the dehydrogenase system of living cells. Fluorescence microscopy was applied to visualize the biofilm on the surface of the titanium alloy; SYTO 13 was used as a fluorescence probe to stain cells in the biofilm. It has been shown that biofilm populations of all studied microorganisms are very sensitive to the type of used NTP. The inhibition zone of biofilm recorded after 60 minutes exposure to NTP exceeded 20 cm², except P. aeruginosa DBM 3777 and ATCC 10145, where it was about 9 cm². Also metabolic activity of cells in biofilm differed for individual microbial strains. High sensitivity to NTP was observed in S. epidermidis, in which the metabolic activity of biofilm decreased after 30 minutes of NTP exposure to 15% and after 60 minutes to 1%. Conversely, the metabolic activity of cells of C. albicans decreased to 53% after 30 minutes of NTP exposure. Nevertheless, this result can be considered very good. Suitable combinations of exposure time of NTP and the concentration of antibiotic achieved in most cases a remarkable synergic effect on the reduction of the metabolic activity of the cells of the biofilm. For example, in the case of P. aeruginosa DBM 3777, a combination of 30 minutes of NTP with 1 mg/l of ceftazidime resulted in a decrease metabolic activity below 4%.

Keywords: anti-biofilm activity, antibiotic, non-thermal plasma, opportunistic pathogens

Procedia PDF Downloads 162

Authors: Mohammad Nasb, Muhammad Rehan, Chen Hong

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Background Meniscus defects critically alter knee function and lead to degenerative changes. Regenerative medicine applications including stem cell transplantation have showed a promising efficacy in finding alternatives to overcome traditional treatment limitations. However, stem cell therapy remains limited due to the substantially reduced viability and inhibitory microenvironment. Since tissue growth and repair are under the control of biochemical and mechanical signals, several approaches have recently been investigated (e.g., low intensity pulsed ultrasound [LIPUS]) to promote the regeneration process. This study employed LIPUS to improve growth and osteogenic differentiation of mesenchymal stem cells derived from human embryonic stem cells to improve the regeneration of meniscus tissue. Methodology: The Mesenchymal stromal cells (MSCs) were transplanted into the epicenter of the injured meniscus in rabbits, which were randomized into two main groups: a treatment group (n=32 New Zealand rabbits) including 4 subgroups of 8 rabbits in each subgroup (LIPUS treatment, MSC treatment, LIPUS with MSC and control), and a second group (n=9) to track implanted cells and their progeny using green fluorescence protein (GFP). GFP consists of the MSC and LIPUS-MSC combination subgroups. Rabbits were then subjected to histological, immunohistochemistry, and MRI assessment. Results: The quantity of the newly regenerated tissue in the combination treatment group that had Ultrasound irradiation after mesenchymal stem cells were better at all end points. Likewise, Tissue quality scores were also greater in knees treated with both approaches compared with controls and single treatment at all end points, achieving significance at twelve and twenty-four weeks [p < 0.05], and [p = 0.008] at twelve weeks. Differentiation into type-I and II collagen-expressing cells were higher in the combination group at up to twenty-four weeks. Conclusions: the combination of mesenchymal stem cells and LIPUS showed greater adhering to the sites of meniscus injury, differentiate into cells resembling meniscal fibrochondrocytes, and improve both quality and quantity of meniscal regeneration.

Keywords: stem cells, regenerative medicine, osteoarthritis, knee

Procedia PDF Downloads 90

Authors: Yu-Ju Hsieh, Lin-Hung Lin, Wen-Hui Chang

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This study was designed as an interventional research and intended to validate whether the introduction of drinking vinegar every day can reduce and even prevent urinary tract infection in Taiwan home stayed disabilities who using indwelling catheter. The data was collected from the subjects who have received home care case at northern Taiwan, according to the questionnaire and a medical records retroactive methodology, the subjects were informed and consent to drink 15ml of table vinegar in a daily diet, and through routine urine testing and culture study. Home care nurses would assist collecting urine at the point of before and after a meal from total 35 studied subjects per month, and total collected 4 times for testing. The results showed that when the average age of study subjects was 65.46 years and catheter indwelling time was 15 years, drinking table vinegar could inhibit the activity of E. coli O157: H7 and reduce its breeding. Before drinking table vinegar daily, the subjects’ urine pH value was 7.0-8.0, and the average was 7.5, and the urine PH value dropped to 6.5 after drinking table vinegar for a month. There were two purple urine cases whose urine were changed from purple to normal color after two weeks of drinking, and the protein and bacteria values of urine gradually improved. Urine smell unpleasant before attending to this study, and the symptom improved significantly only after 1 week, and the urine smell returned to normal ammonia and became clean after 1 month later. None of these subjects received treatment in a hospital due to urinary tract infection, and there were no signs of bleeding in all cases during this study. The subjects of this study are chronic patients with a long-term bedridden catheterization; drinking cranberry juice is an economic burden for them, and also highly prohibited for diabetes patients. By adapting to use cheaper table vinegar to acidified urine and improve its smell and ease Purple Urine Syndrome, to furthermore, proven urinary tract infection, it can also to reduce the financial burden on families, the cost of social resources and the rate of re-admission.

Keywords: table vinegar, urinary tract infection, disability patients, long-term indwelling catheter

Procedia PDF Downloads 235

Authors: Uraib Sharaha, Ahmad Salman, Eladio Rodriguez-Diaz, Elad Shufan, Klaris Riesenberg, Irving J. Bigio, Mahmoud Huleihel

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Antimicrobial drugs have played an indispensable role in controlling illness and death associated with infectious diseases in animals and humans. However, the increasing resistance of bacteria to a broad spectrum of commonly used antibiotics has become a global healthcare problem. Many antibiotics had lost their effectiveness since the beginning of the antibiotic era because many bacteria have adapted defenses against these antibiotics. Rapid determination of antimicrobial susceptibility of a clinical isolate is often crucial for the optimal antimicrobial therapy of infected patients and in many cases can save lives. The conventional methods for susceptibility testing require the isolation of the pathogen from a clinical specimen by culturing on the appropriate media (this culturing stage lasts 24 h-first culturing). Then, chosen colonies are grown on media containing antibiotic(s), using micro-diffusion discs (second culturing time is also 24 h) in order to determine its bacterial susceptibility. Other methods, genotyping methods, E-test and automated methods were also developed for testing antimicrobial susceptibility. Most of these methods are expensive and time-consuming. Fourier transform infrared (FTIR) microscopy is rapid, safe, effective and low cost method that was widely and successfully used in different studies for the identification of various biological samples including bacteria; nonetheless, its true potential in routine clinical diagnosis has not yet been established. The new modern infrared (IR) spectrometers with high spectral resolution enable measuring unprecedented biochemical information from cells at the molecular level. Moreover, the development of new bioinformatics analyses combined with IR spectroscopy becomes a powerful technique, which enables the detection of structural changes associated with resistivity. The main goal of this study is to evaluate the potential of the FTIR microscopy in tandem with machine learning algorithms for rapid and reliable identification of bacterial susceptibility to antibiotics in time span of few minutes. The UTI E.coli bacterial samples, which were identified at the species level by MALDI-TOF and examined for their susceptibility by the routine assay (micro-diffusion discs), are obtained from the bacteriology laboratories in Soroka University Medical Center (SUMC). These samples were examined by FTIR microscopy and analyzed by advanced statistical methods. Our results, based on 700 E.coli samples, were promising and showed that by using infrared spectroscopic technique together with multivariate analysis, it is possible to classify the tested bacteria into sensitive and resistant with success rate higher than 90% for eight different antibiotics. Based on these preliminary results, it is worthwhile to continue developing the FTIR microscopy technique as a rapid and reliable method for identification antibiotic susceptibility.

Keywords: antibiotics, E.coli, FTIR, multivariate analysis, susceptibility, UTI

Procedia PDF Downloads 152

Authors: Rabia Göçmen, Gülşah Kanbur, Sinan Sefa Parlat

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Aim of this study is to determine the effects of cellular insulin receptor stimulators on performance, plasma glucose, high density lipoprotein (HDL), low density lipoprotein (LDL), total cholesterol, triglyceride, triiodothyronine (T3) and thyroxine (T4) hormone levels, and incubation features in the breeding Japanese quails (Coturnix japonica). In the study, a total of 84 breeding quails was used, 6 weeks’ age, 24 are male and 60, female. Rations used in experiment are 2900 kcal/kg metabolic energy and 20% crude protein. Water pH is calibrated to 7.45. Ration and water were administered ad-libitum to the animals. As metformin source, metformin-HCl was used and as chrome resource, chromium picolinate was used. Trial groups were formed as control group (basal ration), metformin group (basal ration, added metformin at the level of feed of 20 mg/kg), and chromium picolinate (basal ration, added feed of 1500 ppb Cr) group. When regarded to the results of performance at the end of experiment, it is seen that live weight gain, feed consumption, egg weight, feed conversion ratio (Feed consumption/ egg weight), and egg production were affected at the significant level (p < 0.05). When the results are evaluated in terms of incubation features, hatchability and hatchability of fertile egg ratio were not affected from the treatments. Fertility ratio was significantly affected by metformin and chromium picolinate treatments and fertility rose at the significant level compared to control group (p < 0.05). According to results of experiment, plasma glucose level was not affected by metformin and chromium picolinate treatments. Plasma, total cholesterol, HDL, LDL, and triglyceride levels were significantly affected from insulin receptor stimulators added to ration (p < 0.05). Hormone level of Plasma T3 and T4 were also affected at the significant level from insulin receptor stimulators added to ration (p < 0.05).

Keywords: chromium picolinate, cholesterol, hormone, metformin, quail

Procedia PDF Downloads 195

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

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

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

Procedia PDF Downloads 255

Authors: Junichi Hosoi

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Epidermal Langerhans cells reside in upper layer of epidermis and play a role in immune surveillance. The finding of the close association of nerve endings to Langerhans cells triggered the research on systemic regulation of Langerhans cells. They disappear from epidermis after exposure to environmental and internal stimuli and reappear about a week later. Myeloid progenitor cells are assumed to be one of the sources of Langerhans cells. We examined the effects of cortisol on the reappearance of Langerhans cells in vitro. Cord-blood derived CD34-positive cells were cultured in the medium supplemented with stem cell factor/Flt3 ligand/granulocyte macrophage-colony stimulating factor/tumor necrosis factor alpha/bone morphologic protein 7/transforming growth factor beta in the presence or absence of cortisol. Cells were analyzed by flow cytometry for CD1a (cluster differentiation 1a), a marker of Langerhans cells and dermal dendritic cells, and CD39 (cluster differentiation factor 39), extracellular adenosine triphosphatase. Both CD1a-positive cells and CD39-positive cells were decreased by treatment with cortisol (suppression by 35% and 22% compared to no stress hormone, respectively). Differentiated Langerhans cells are attracted to epidermis by chemokines that are secreted from keratinocytes. Epidermal keratinocytes were cultured in the presence or absence of cortisol and analyzed for the expression of CCL2 (C-C motif chemokine ligand 2) and CCL20 (C-C motif chemokine ligand 20), which are typical attractants of Langerhans cells, by quantitative reverse transcriptase polymerase chain reaction. The expression of both chemokines, CCL2 and CCL20, were suppressed by treatment with cortisol (suppression by 38% and 48% compared to no stress hormone, respectively). We examined the possible regulation of the suppression by cortisol with plant extracts. The extracts of Ganoderma lucidum and Iris protected the suppression of the differentiation to CD39-positive cells and also the suppression of the gene expression of LC-chemoattractants. These results suggest that cortisol, which is either systemic or locally produced, blocks the supply of epidermal Langerhans cells at 2 steps, differentiation from the precursor and attraction to epidermis. The suppression is possibly blocked by some plant extracts.

Keywords: Langerhans cell, stress, CD39, chemokine

Procedia PDF Downloads 157

Authors: Farahnaz Sadat Golestan Hashemi, Mohd Razi Ismail, Mohd Y. Rafii

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A huge revolution has emerged in genome engineering by the discovery of CRISPR (clustered regularly interspaced palindromic repeats) and CRISPR-associated system genes (Cas) in bacteria. The function of type II Streptococcus pyogenes (Sp) CRISPR/Cas9 system has been confirmed in various species. Other S. thermophilus (St) CRISPR-Cas systems, CRISPR1-Cas and CRISPR3-Cas, have been also reported for preventing phage infection. The CRISPR1-Cas system interferes by cleaving foreign dsDNA entering the cell in a length-specific and orientation-dependant manner. The S. thermophilus CRISPR3-Cas system also acts by cleaving phage dsDNA genomes at the same specific position inside the targeted protospacer as observed in the CRISPR1-Cas system. It is worth mentioning, for the effective DNA cleavage activity, RNA-guided Cas9 orthologs require their own specific PAM (protospacer adjacent motif) sequences. Activity levels are based on the sequence of the protospacer and specific combinations of favorable PAM bases. Therefore, based on the specific length and sequence of PAM followed by a constant length of target site for the three orthogonals of Cas9 protein, a well-organized procedure will be required for high-throughput and accurate mining of possible target sites in a large genomic dataset. Consequently, we created a reliable procedure to explore potential gRNA sequences for type I (Streptococcus thermophiles), II (Streptococcus pyogenes), and III (Streptococcus thermophiles) CRISPR/Cas systems. To mine CRISPR target sites, four different searching modes of sgRNA binding to target DNA strand were applied. These searching modes are as follows: i) coding strand searching, ii) anti-coding strand searching, iii) both strand searching, and iv) paired-gRNA searching. The output of such procedure highlights the power of comparative genome mining for different CRISPR/Cas systems. This could yield a repertoire of Cas9 variants with expanded capabilities of gRNA design, and will pave the way for further advance genome and epigenome engineering.

Keywords: CRISPR/Cas systems, gRNA mining, Streptococcus pyogenes, Streptococcus thermophiles

Procedia PDF Downloads 222

Authors: Richa Dhingra, Monika, Manav Malhotra, Tilak Raj Bhardwaj, Neelima Dhingra

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5-Alpha-reductases (5AR), a membrane bound, NADPH dependent enzyme and convert male hormone testosterone (T) into more potent androgen dihydrotestosterone (DHT). DHT is the required for the development and function of male sex organs, but its overproduction has been found to be associated with physiological conditions like Benign Prostatic Hyperplasia (BPH). Thus the inhibition of 5ARs could be a key target for the treatment of BPH. In present study, 2D and 3D Quantitative Structure Activity Relationship (QSAR) pharmacophore models have been generated for 5AR based on known inhibitory concentration (IC₅₀) values with extensive validations. The four featured 2D pharmacophore based PLS model correlated the topological interactions (–OH group connected with one single bond) (SsOHE-index); semi-empirical (Quadrupole2) and physicochemical descriptors (Mol. wt, Bromines Count, Chlorines Count) with 5AR inhibitory activity, and has the highest correlation coefficient (r² = 0.98, q² =0.84; F = 57.87, pred r² = 0.88). Internal and external validation was carried out using test and proposed set of compounds. The contribution plot of electrostatic field effects and steric interactions generated by 3D-QSAR showed interesting results in terms of internal and external predictability. The well validated 2D Partial Least Squares (PLS) and 3D k-nearest neighbour (kNN) models were used to search novel 5AR inhibitors with different chemical scaffold. To gain more insights into the molecular mechanism of action of these steroidal derivatives, molecular docking and in silico absorption, distribution, metabolism, and excretion (ADME) studies were also performed. Studies have revealed the hydrophobic and hydrogen bonding of the ligand with residues Alanine (ALA) 63A, Threonine (THR) 60A, and Arginine (ARG) 456A of 4AT0 protein at the hinge region. The results of QSAR, molecular docking, in silico ADME studies provide guideline and mechanistic scope for the identification of more potent 5-Alpha-reductase inhibitors (5ARI).

Keywords: 5α-reductase inhibitor, benign prostatic hyperplasia, ligands, molecular docking, QSAR

Procedia PDF Downloads 137

Authors: Cangir Uyarlar, Eyup Eren Gultepe, Mustafa Kabu, Hacı Ahmet Celik

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This study aimed to evaluate the effects of orally Immunoglobulin (Ig) Y treatments to calves were fed with colostrum having insufficient antibodies before first suckling. A total of 28 Holstein calves were fed assigned into control and treatment groups. The calves were fed fresh colostrum from their respective mother for the first 4 days. The treatment group calves were orally administered IgLock (10 g/d/calf) immediately before the first colostrum feeding and IgLock was administered just one time in treatment group calves. Then, the calves were offered normal milk until weaning. After weaning, all calves kept same paddock and were fed same ration. Diarrhea and respiratoric diseases were recorded for one year. Blood was collected from all calves in the study on birth day (0 day) before vaccination and IgLock administration, then, collected for the following 2 days in all groups. Albumin (ALB), Total Protein (TP), Aspartate Aminotransferase (AST), Alanine Aminotrasferase (ALT), Gamma-Glutamyl Transferase (GGT), Serum Amyloid A (SAA), Haptoglobin (HPT) and Ig G analyses were performed on all samples. Although serum ALB, ALT, GGT and Ig G levels were not shown a time dependent-change within control group; serum TP, AST, HPT and SAA levels were significantly changed by the time within mentioned group. Serum TP level was steady at first 2 days, then, it was increased significantly at 3rd day. Also, serum AST level was significantly increased at 2nd day, then it was descended to first day levels again at 3rd day. Although serum HPT levels were shown a significant gradually decreasing within control group, serum SAA levels were decreased rapidly after first day and there were no significance differences between 2nd and 3rd day in SAA levels. Serum ALB, ALT, HPT and SAA levels were not shown a time dependent-change within treatmet group. After first day Serum TP, GGT, AST and Ig G levels were shown an significant increasing at 2nd day. Serum TP, GGT and Ig G levels were higher as compared to 1st day within treatment group at 3rd day. But, serum AST level was less significantly 3rd day as compared to 2nd day values. The total numbers of calves suffered from diarrhea were significantly less in treatment group as compared to control group (p < 0,05). The pneumonia reappear ratio in calves suffered the diseases is 33,3% in control group and 11,11% in treatment group. Total cost of diseases and additives was 2339,36 $ for control and 1276,4 $ for treatment. As a conclusion, the immunity enhancers like IgLock are important and cost-effective to boost up immunity status in the early age which would be having positive effects on calves were received colostrum included insufficient antibodies.

Keywords: dairy calves, Ig Y, pneumonia, scours

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Authors: N. G. Klivleyeva, T. I. Glebova, G. V. Lukmanova, S. B. Bayseit, S. Z. Taubaeva, M. K. Kalkozhaeva

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The role of viral diseases in the general infectious disease incidence increases every year and requires special attention to the problem of interpreting the etiology of infectious agents. Influenza and acute respiratory viral infections are one of the most pressing public health issues. In the period 2012-2014, collection of 419 nasal swabs and 150 blood sera has been carried out in the patient care institutions of the various Kazakhstan regions from patients with symptoms of ARVI and pneumonia. Primary identification of biosamples for the presence of influenza viral antigens in enzyme immunoassay on nitrocellulose membrane gave positive results in 125 swabs (29.8%). Biosample screening in immunofluorescence test revealed the presence of influenza viral antigens against A/H1 in 63 samples (15.0%), A/H3 – in 70 samples (16.7%) and type B – in 9 samples (2.1%). As a result of primary infection, and successive passages in chick embryos and MDCK cell cultures, 38 HAAg were isolated from 419 samples with a clear cytopathic effect and hemagglutination titre in MDCK cell culture within 1:2-1:4, in CE - 1:8-1:256. The infectivity of isolates in chicken embryos were 3.5-6.5 lg EID50/0.2, in MDCK cell culture – 2.5-6.5 lg PFU/ml. Identification of 28 isolates was carried out in inhibition reactions of hemagglutinating activity and neuraminidase activity, showed their belonging to the influenza virus: 26 strains to A/H1N1, one - to A/H3N2, and one - to type B. Serological examination of blood sera for the presence of specific antibodies being an indirect evidence of the performed isolation and contributing to the timely interpretation of the disease etiology in the epidemics takes an important place in the comprehensive study of influenza viruses circulating among people. Serological analyzes were carried out in HAI assay using a kit consisting of 12 reference strains obtained from the WHO centre for reference and research on Influenza (CDC, Atlanta, USA) and three Kazakhstan (A/Almaty/347/09 (H1N1v), A/Almaty/462/11 (H3N2) and B/Almaty/414/10) human influenza viruses that are stored in the laboratory collection. The results of serological analysis of 150 blood sera showed that antihaemagglutinins against the A/H3N2 virus serosubtype were found in 46 samples (49.4%) out of 93 sera collected in 2012-2013. The antibody titres were within 1:160-1:320. 19 sera (20.4%) were seropositive against influenza A/H1N1 virus, the antibodies were observed in titres of 1:20-1:40. Six sera (6.4%) were positive against the influenza A/H1N1+A/H3N2 virus (mixed infection); the antibodies were recorded in titres of 1:20-1:40. Antihaemagglutinins against influenza type B virus were detected only in five sera (5.4%). The results of analysis of 57 sera collected in 2014 showed that antihaemagglutinins against A/H3N2 virus subtype were detected in 32 blood sera (56.1%) in titres of 1:160-1:640. Ten sera (17.5%) were seropositive against A/H1N1 virus; antihaemagglutinins against influenza type B virus were not detected. Therefore, virological and serological studies have shown that in Kazakhstan, as well as in the world, the influenza viruses A/H1N1, A/H3N2 and influenza B viruses were actively circulating during the epidemic seasons in 2012-2014.

Keywords: influenza, MDCK cell, serological analysis, virus

Procedia PDF Downloads 151

Authors: Fakunle Egbo, Kammalchukwu A., Akinremi T.

Abstract:

Physical activity and nutritional status influence the health status and cognition of young adults. Lack of physical activity increases the likelihood of developing obesity which leads to the risk of heart diseases and other risk factors like high blood pressure, high blood cholesterol, diabetes etc. The study employed a cross-sectional study design. The study used a multi stage sampling technique multi- stage sampling technique; Purposive, for the selection of colleges that would be used, stratified random sampling for stratifying the colleges into departments and the simple random sampling for the selection of each respondent from the departments. Structured questionnaires were used to obtain data from the respondents and pre-tested anthropometric instruments were used to get the weight and height of the respondents and statistically analyzed using SPSS version 22.0 and the TDA (Total dietary allowance) software which was used to analyze the nutrient intake of the respondents. This study showed that they comprised of 50.1% males and 40.9% females. Slightly above average 51.8% were between ages of 15-19 with mean age being 19.57 years; ages 20-24 were slightly below average at 45.7%. The male students 58.7% had vigorous physical activity, whereas majority of females 76.5% had light physical activity level. 39.1% of the male students carried out physical activity 2-3 times per week while One third of the female students (38.3%) carried out physical activity 6-7 times per week. Majority of the respondents had Inadequate Protein- 63.8%, Carbohydrate- 60.2%, and Dietary fiber- 88.8. 36% eat rice 4-6 times per week. Majority of the respondents had inadequate fruit and vegetables (Efo, Banana,) at 47.7%, 40.6% respectively. Using Body mass index, (63.2%) have normal weight. 22.9% are overweight, 6.8% are underweight, 5.4% have grade 1 obesity and 1.6% have grade II obesity. There was a statistically significant association between the physical activity of the respondents with their nutritional status (p=0.037), physical activity and sex (p=0.000), nutritional status and amount spent on food daily (p=0.007). The study concluded that the physical activity level of the respondents, most especially the females were low; One third of the students were malnourished therefore, there should be an urgent need for improving the overall health status of students by providing the students with well-equipped gyms and other sporting equipment’s that would make them participate actively and keep fit.

Keywords: physical activity, nutritional status, undergraduates, dietary pattern

Procedia PDF Downloads 43

Authors: Oqba Basal, András Szabó

Abstract:

Legume crops are able to fix atmospheric nitrogen by the symbiotic relation with specific bacteria, which allows the use of the mineral nitrogen-fertilizer to be reduced, or even excluded, resulting in more profit for the farmers and less pollution for the environment. Soybean (Glycine max (L.) Merrill) is one of the most important legumes with its high content of both protein and oil. However, it is recommended to combine the two nitrogen sources under stress conditions in order to overcome its negative effects. Drought stress is one of the most important abiotic stresses that increasingly limits soybean yields. A precise rate of mineral nitrogen under drought conditions is not confirmed, as it depends on many factors; soybean yield-potential and soil-nitrogen content to name a few. An experiment was conducted during 2017 growing season in Debrecen, Hungary to investigate the effects of nitrogen source on the physiology and the yield of the soybean cultivar 'Boglár'. Three N-fertilizer rates including no N-fertilizer (0 N), 35 kg ha^-1 of N-fertilizer (35 N) and 105 kg ha^-1 of N-fertilizer (105 N) were applied under three different irrigation regimes; severe drought stress (SD), moderate drought stress (MD) and control with no drought stress (ND). Half of the seeds in each treatment were pre-inoculated with Bradyrhizobium japonicum inoculant. The overall results showed significant differences associated with fertilization and irrigation, but not with inoculation. Increasing N rate was mostly accompanied with increased chlorophyll content and leaf area index, whereas it positively affected the plant height only when the drought was waived off. Plant height was the lowest under severe drought, regardless of inoculation and N-fertilizer application and rate. Inoculation increased the yield when there was no drought, and a low rate of N-fertilizer increased the yield furthermore; however, the high rate of N-fertilizer decreased the yield to a level even less than the inoculated control. On the other hand, the yield of non-inoculated plants increased as the N-fertilizer rate increased. Under drought conditions, adding N-fertilizer increased the yield of the non-inoculated plants compared to their inoculated counterparts; moreover, the high rate of N-fertilizer resulted in the best yield. Regardless of inoculation, the mean yield of the three fertilization rates was better when the water amount increased. It was concluded that applying N-fertilizer to provide the nitrogen needed by soybean plants, with the absence of N₂-fixation process, is very important. Moreover, adding relatively high rate of N-fertilizer is very important under severe drought stress to alleviate the drought negative effects. Further research to recommend the best N-fertilizer rate to inoculated soybean under drought stress conditions should be executed.

Keywords: drought stress, inoculation, N-fertilizer, soybean physiology, yield

Procedia PDF Downloads 121

Authors: Saeed Qaisrani

Abstract:

The study was conducted to observe carbon and nutrient loss by burning of rice residues on rice-wheat cropping system The rice crop was harvested to conduct the experiment in a randomized complete block design (RCBD) with factors and 4 replications with a net plot size of 10 m x 20 m. Rice stubbles were managed by two methods i.e. Incorporation & burning of rice residues. Soil samples were taken to a depth of 30 cm before sowing & after harvesting of wheat. Wheat was sown after harvesting of rice by three practices i.e. Conventional tillage, Minimum tillage and Zero tillage to observe best tillage practices. Laboratory and field experiments were conducted on wheat to assess best tillage practice and residues management method with estimation of carbon losses. Data on the following parameters; establishment count, plant height, spike length, number of grains per spike, biological yield, fat content, carbohydrate content, protein content, and harvest index were recorded to check wheat quality & ensuring food security in the region. Soil physico-chemical analysis i.e. pH, electrical conductivity, organic matter, nitrogen, phosphorus, potassium, and carbon were done in soil fertility laboratory. Substantial results were found on growth, yield and related parameters of wheat crop. The collected data were examined statistically with economic analysis to estimate the cost-benefit ratio of using different tillage techniques and residue management practices. Obtained results depicted that Zero tillage method have positive impacts on growth, yield and quality of wheat, Moreover, it is cost effective methodology. Similarly, Incorporation is suitable and beneficial method for soil due to more nutrients provision and reduce the need of fertilizers. Burning of rice stubbles has negative impact including air pollution, nutrient loss, microbes died and carbon loss. Recommended the zero tillage technology to reduce carbon losses along with food security in Pakistan.

Keywords: agricultural agronomy, food security, carbon sequestration, rice-wheat cropping system

Procedia PDF Downloads 253