Search results for: cap-dependent protein translation

Authors: Adnan A. Kadi, Nasser S. Al-Shakliah, Haitham Al-Rabiah

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Zorifertinib is a novel, potent, oral, a small molecule used to treat non-small cell lung cancer (NSCLC). zorifertinib is an Epidermal Growth Factor Receptor (EGFR) inhibitor and has good blood–brain barrier permeability for (NSCLC) patients with EGFR mutations. zorifertinibis currently at phase II/III clinical trials. The current research reports the characterization and identification of in vitro, in vivo and reactive intermediates of zorifertinib. Prediction of susceptible sites of metabolism and reactivity pathways (cyanide and GSH) of zorifertinib were performed by the Xenosite web predictor tool. In-vitro metabolites of zorifertinib were performed by incubation with rat liver microsomes (RLMs) and isolated perfused rat liver hepatocytes. Extraction of zorifertinib and it's in vitro metabolites from the incubation mixtures were done by protein precipitation. In vivo metabolism was done by giving a single oral dose of zorifertinib(10 mg/Kg) to Sprague Dawely rats in metabolic cages by using oral gavage. Urine was gathered and filtered at specific time intervals (0, 6, 12, 18, 24, 48, 72,96and 120 hr) from zorifertinib dosing. A similar volume of ACN was added to each collected urine sample. Both layers (organic and aqueous) were injected into liquid chromatography ion trap mass spectrometry(LC-IT-MS) to detect vivozorifertinib metabolites. N-methyl piperizine ring and quinazoline group of zorifertinib undergoe metabolism forming iminium and electro deficient conjugated system respectively, which are very reactive toward nucleophilic macromolecules. Incubation of zorifertinib with RLMs in the presence of 1.0 mM KCN and 1.0 Mm glutathione were made to check reactive metabolites as it is often responsible for toxicities associated with this drug. For in vitro metabolites there were nine in vitro phase I metabolites, four in vitro phase II metabolites, eleven reactive metabolites(three cyano adducts, five GSH conjugates metabolites, and three methoxy metabolites of zorifertinib were detected by LC-IT-MS. For in vivo metabolites, there were eight in vivo phase I, tenin vivo phase II metabolitesofzorifertinib were detected by LC-IT-MS. In vitro and in vivo phase I metabolic pathways wereN- demthylation, O-demethylation, hydroxylation, reduction, defluorination, and dechlorination. In vivo phase II metabolic reaction was direct conjugation of zorifertinib with glucuronic acid and sulphate.

Keywords: in vivo metabolites, in vitro metabolites, cyano adducts, GSH conjugate

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Authors: S. Fröhlich, M. Herold, M. Allmer

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Early stage quantitative analysis of host cell protein (HCP) variations is challenging yet necessary for comprehensive bioprocess development. High resolution mass spectrometry (HRMS) provides a high-end technology for accurate identification alongside with quantitative information. Hereby we describe a flexible HRMS assay platform to quantify HCPs relevant in microbial expression systems such as E. Coli in both up and downstream development by means of MVDA tools. Cell pellets were lysed and proteins extracted, purified samples not further treated before applying the SMART tryptic digest kit. Peptides separation was optimized using an RP-UHPLC separation platform. HRMS-MSMS analysis was conducted on an Orbitrap Velos Elite applying CID. Quantification was performed label-free taking into account ionization properties and physicochemical peptide similarities. Results were analyzed using SIEVE 2.0 (Thermo Fisher Scientific) and SIMCA (Umetrics AG). The developed HRMS platform was applied to an E. Coli expression set with varying productivity and the corresponding downstream process. Selected HCPs were successfully quantified within the fmol range. Analysing HCP networks based on pattern analysis facilitated low level quantification and enhanced validity. This approach is of high relevance for high-throughput screening experiments during upstream development, e.g. for titer determination, dynamic HCP network analysis or product characterization. Considering the downstream purification process, physicochemical clustering of identified HCPs is of relevance to adjust buffer conditions accordingly. However, the technology provides an innovative approach for label-free MS based quantification relying on statistical pattern analysis and comparison. Absolute quantification based on physicochemical properties and peptide similarity score provides a technological approach without the need of sophisticated sample preparation strategies and is therefore proven to be straightforward, sensitive and highly reproducible in terms of product characterization.

Keywords: process analytical technology, mass spectrometry, process characterization, MVDA, pattern recognition

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Authors: Obuzor Eze Obuzor, Ekpoke Okurube Sliver

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The study was carried out at Debee farms at Ahoada West Local Government area, Rivers State, Nigeria. To evaluate the impact of two different cost-effective available feed composition on growth performance of weaned piglets. Thirty weaned uncontrolled cross bred (Large white x pietrain) piglets of average initial weight of 3.04 Kg weaned at 30days were assigned to three dietary treatments, comprising three replicates of 10 weaned piglets each, piglets were kept at 7 °C in different pens with dimensions of 4.50 × 4.50 m. The design of the experiment was completely randomized design, data from the study were subjected to one-way analysis of variance (ANOVA) and significant means were separated using Duncan's Multiple Range Test using Statistical Analysis System (SAS) software for windows (2 0 0 3), statistical significance was assessed at P < 0.05 (95% confidence interval) while survival rate was calculated using simple percentage. A standard diet was prepared to meet the nutrient requirements of weaned piglets at (20.8% crude protein). The three diets were fed to the animals in concrete feeding trough, control diet (C) had soybean meal while first treatment had spent grain (T1) and the second treatment had wheat offal (T2) respectively. The experiment was partitioned into four weeks periods (days 1-7, 8-14, 15-21 and 22-28). Feed and water were given unrestrictedly throughout the period of the experiment. The feed intake and weights of the pigs were recorded on weekly basis. Feed conversion ratio and daily weight gain were calculated and the study lasted for four weeks. There was no significant (P>0.05) effect of diet on survival rate, final body weight, average daily weight gain, daily feed intake and feed conversion ratio. The overall performance showed that treatment one (T1) had survival rate (93%), improved daily weight gain (36.21 g), average daily feed intake (120.14 g) and had the best feed conversion ratio (0.29) similar high mean value with the control while treatment two (T2) had lowest and negative response to all parameters. It could be concluded that feed formulated with spent grain is cheaper than control (soybean meal) and also improved the growth performance of weaned piglets.

Keywords: piglets, weaning, feed conversions ratio, daily weight gain

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Authors: Nilgün Öztürk, Hakan Sabahtin Ali, Hülya Tuba Kıyan

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The genus Chenopodium belongs to Amaranthaceae, is represented by approximately 250 species in the world and 15 species and three subspecies in Turkey. Chenopodium species are traditionally used to treat chest and abdominal pain, shortness of breath, cough and neurological disorders. Chenopodium quinoa Willd. (Quinoa) is native to Andes region of South America (especially Peru and Bolivia) and cultivated in many countries include also Turkey in the world nowadays. The seeds of quinoa are rich in protein, and the phytochemical composition consists of antioxidant substances such as polyphenolic compounds, flavonoids, vitamins, and minerals; anticancer and neuroprotective compounds such as tocotrienols; anti-inflammatory compounds such as carotenoids and anthocyanins and also saponins and starch. Food products of quinoa such as quinoa cereal bar, pasta and cornflakes are used in the diet made during many disorders like obesity, cardiovascular disorder, hypertension and Celiac disease. Also quinoa seems to have antimicrobial, anti-inflammatory and cholesterol-lowering properties because of its bioactive compounds. In this present study, the aqueous ethanolic extracts of the seeds of three different coloured genotypes of quinoa were investigated for their antioxidant activities using 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging activity, ferrous ion-chelating effect, ferric-reducing antioxidant power, ABTS radical cation decolorization assays and total phenolic contents using Folin-Ciocalteu assay. Among the three genotypes of quinoa; the aqueous ethanolic extract of the red genotype had the highest total phenolic content (83.54 ± 2.12 mg gallic acid/100 g extract) whereas the extract of the white genotype had the lowest total phenolic content (70.66 ± 0.25 mg gallic acid/100 g). According to the antioxidant activity results; the extracts showed moderate reducing power effect whereas weak ABTS radical cation decolorization and ferrous ion-chelating effect and also too weak DPPH radical scavenging activity when compared to the positive standards.

Keywords: amaranthaceae, antioxidant activity, Chenopodium quinoa willd., total phenolic content

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Authors: Ja-Keoung Koo, Kensuke Nakamura, Hyohun Kim, Dongwha Shin, Yeonseok Kim, Ji-Su Ahn, Byung-Woo Hong

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The machine learning techniques based on a convolutional neural network (CNN) have been actively developed and successfully applied to a variety of image analysis tasks including reconstruction, noise reduction, resolution enhancement, segmentation, motion estimation, object recognition. The classical visual information processing that ranges from low level tasks to high level ones has been widely developed in the deep learning framework. It is generally considered as a challenging problem to derive visual interpretation from high dimensional imagery data. A CNN is a class of feed-forward artificial neural network that usually consists of deep layers the connections of which are established by a series of non-linear operations. The CNN architecture is known to be shift invariant due to its shared weights and translation invariance characteristics. However, it is often computationally intractable to optimize the network in particular with a large number of convolution layers due to a large number of unknowns to be optimized with respect to the training set that is generally required to be large enough to effectively generalize the model under consideration. It is also necessary to limit the size of convolution kernels due to the computational expense despite of the recent development of effective parallel processing machinery, which leads to the use of the constantly small size of the convolution kernels throughout the deep CNN architecture. However, it is often desired to consider different scales in the analysis of visual features at different layers in the network. Thus, we propose a CNN model where different sizes of the convolution kernels are applied at each layer based on the random projection. We apply random filters with varying sizes and associate the filter responses with scalar weights that correspond to the standard deviation of the random filters. We are allowed to use large number of random filters with the cost of one scalar unknown for each filter. The computational cost in the back-propagation procedure does not increase with the larger size of the filters even though the additional computational cost is required in the computation of convolution in the feed-forward procedure. The use of random kernels with varying sizes allows to effectively analyze image features at multiple scales leading to a better generalization. The robustness and effectiveness of the proposed CNN based on random kernels are demonstrated by numerical experiments where the quantitative comparison of the well-known CNN architectures and our models that simply replace the convolution kernels with the random filters is performed. The experimental results indicate that our model achieves better performance with less number of unknown weights. The proposed algorithm has a high potential in the application of a variety of visual tasks based on the CNN framework. Acknowledgement—This work was supported by the MISP (Ministry of Science and ICT), Korea, under the National Program for Excellence in SW (20170001000011001) supervised by IITP, and NRF-2014R1A2A1A11051941, NRF2017R1A2B4006023.

Keywords: deep learning, convolutional neural network, random kernel, random projection, dimensionality reduction, object recognition

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Authors: Fana Woldetsadik

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Ethiopia has a huge livestock population, including sheep, that has been contributing a considerable portion to the economy of the country and still promising to rally around the economic advancement of the country. However, feed shortage is a limiting factor in the production and productivity of sheep among Ethiopian smallholder farmers. Therefore, the aim of this study was to prove the role of the locally available brewery by-products called dried Atella as a supplement in feed intake, digestibility, live weight gain, carcass yield, and economic benefit in comparison with commercially purchased supplements known as niger seed cake (NSC). This on-station feeding experiment was conducted on the Zenzelma Campus of Bahir Dar University animal farm. The experimental design used for this research was a completely randomized design (CRD) with five replications. The crude protein (CP) content of dried Atella, wheat bran (WB), natural pasture hay (NPH) and NSC were about 25.07%, 16.57%, 4.48% and 38.04%, respectively, while the neutral detergent fibre (NDF),acid detergent fibre (ADF) and acid detergent lignin (ADL) content of dried Atella, WB, NPH and NSC were around 31.75%, 8.31%, 8.14%; 42.05%, 22.64%, 4.04%; 74.21%, 50.81%, 8.66%; 42.31%, 26.95% and 6.9%, respectively. The result depicted that a higher(P < 0.001) feed intake, nutrient intake, and digestibility for lambs supplemented with Atella than those supplemented with NSC. Furthermore, daily body weight gain and carcass characteristics were better (P < 0.05) for the sheep supplemented with dried Atella than NSC. On the other hand, in terms of profitability, although there was no substantial difference (P > 0.05) between T2 (animals fed NPH,NSC and WB) and T3 (animals fed NPH, Atella and WB), slightly better benefit was recorded in T3 groups. However, loss of money was recorded in T1 (animals fed NPH and WB). Hence, from the biological performance of lambs, it was concluded that Atella could be a potential supplementary feed for sheep fattening among smallholder farmers than NSC despite no profitability difference. Nevertheless, further investigation is recommended to examine the consequence of supplementation of NPH with NSC and NPH with Atella on fatty acid profile analysis, the physicochemical composition of meat, and meat composition.

Keywords: Attela, Bahir Dar university, Carcass yield, digestibility, natural pasture hay, Niger seed cake, smallholder farmers, weight gain, Ethiopia

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Authors: Shveta Bathla, Preeti Rawat, Sudarshan Kumar, Rubina Baithalu, Jogender Singh Rana, Tushar Kumar Mohanty, Ashok Kumar Mohanty

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Pregnancy is a complex process which includes series of events such as fertilization, formation of blastocyst, implantation of embryo, placental formation and development of fetus. The success of these events depends on various interactions which are synchronized by endocrine interaction between a receptive dam and competent embryo. These interactions lead to change in expression of hormones and proteins. But till date no protein biomarker is available which can be used to detect successful completion of these events. We employed quantitative proteomics approach to develop putative serological biomarker which has diagnostic applicability for early detection of pregnancy in cows. For this study, sera were collected from control (non-pregnant, n=6) and pregnant animals on successive days of pregnancy (7, 19, 45, n=6). The sera were subjected to depletion for removal of albumin using Norgen depletion kit. The tryptic peptides were labeled with iTRAQ. The peptides were pooled and fractionated using bRPLC over 80 min gradient. Then 12 fractions were injected to nLC for identification and quantitation in DDA mode using ESI. Identification using Mascot search revealed 2056 proteins out of which 352 proteins were differentially expressed. Twenty proteins were upregulated and twelve proteins were down-regulated with fold change > 1.5 and < 0.6 respectively (p < 0.05). The gene ontology studies of DEPs using Panther software revealed that majority of proteins are actively involved in catalytic activities, binding and enzyme regulatory activities. The DEP'S such as NF2, MAPK, GRIPI, UGT1A1, PARP, CD68 were further subjected to pathway analysis using KEGG and Cytoscape plugin Cluego that showed involvement of proteins in successful implantation, maintenance of pluripotency, regulation of luteal function, differentiation of endometrial macrophages, protection from oxidative stress and developmental pathways such as Hippo. Further efforts are continuing for targeted proteomics, western blot to validate potential biomarkers and development of diagnostic kit for early pregnancy diagnosis in cows.

Keywords: bRPLC, Cluego, ESI, iTRAQ, KEGG, Panther

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Authors: Norhidayu Muhamad Zain

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Gelatin, a purified protein derived mostly from porcine and bovine sources, is used widely in food manufacturing, pharmaceutical, and cosmetic industries. However, the presence of any porcine-related products are strictly forbidden for Muslim and Jewish consumption. Therefore, analytical methods offering reliable results to differentiate the sources of gelatin are needed. The aim of this study was to differentiate the sources of gelatin (porcine and tilapia fish) using Fourier transform infrared- attenuated total reflection (FTIR-ATR) combined with two dimensional infrared (2DIR) correlation analysis. Porcine gelatin (PG) and tilapia fish gelatin (FG) samples were diluted in distilled water at concentrations ranged from 4-20% (w/v). The samples were then analysed using FTIR-ATR and 2DIR correlation software. The results showed a significant difference in the pattern map of synchronous spectra at the region of 1000 cm⁻¹ to 1100 cm⁻¹ between PG and FG samples. The auto peak at 1080 cm⁻¹ that attributed to C-O functional group was observed at high intensity in PG samples compared to FG samples. Meanwhile, two auto peaks (1080 cm⁻¹ and 1030 cm⁻¹) at lower intensity were identified in FG samples. In addition, using 2D correlation analysis, the original broad water OH bands in 1D IR spectra can be effectively differentiated into six auto peaks located at 3630, 3340, 3230, 3065, 2950 and 2885 cm⁻¹ for PG samples and five auto peaks at 3630, 3330, 3230, 3060 and 2940 cm⁻¹ for FG samples. Based on the rule proposed by Noda, the sequence of the spectral changes in PG samples is as following: NH₃⁺ amino acid > CH₂ and CH₃ aliphatic > OH stretch > carboxylic acid OH stretch > NH in secondary amide > NH in primary amide. In contrast, the sequence was totally in the opposite direction for FG samples and thus both samples provide different 2D correlation spectra ranged from 2800 cm-1 to 3700 cm⁻¹. This method may provide a rapid determination of gelatin source for application in food, pharmaceutical, and cosmetic products.

Keywords: 2 dimensional infrared (2DIR) correlation analysis, Fourier transform infrared- attenuated total reflection (FTIR-ATR), porcine gelatin, tilapia fish gelatin

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Authors: Fahad Al Yahyaey, Ihab Shaat, Russell Bush

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This study was conducted at the Livestock Research Centre, Ministry of Agriculture and Fisheries, Oman, on two local goat breeds (Jabbali and Sahrawi) due to their importance to Omani livestock production and food security. The Jabbali is characterized by increased growth rates and a higher twinning rate, while the Sahrawi has increased milk production. The aim of the study was to investigate the effect of Spirulina supplementation on live weight (BWT), average daily gain (ADG), and body conformation measurements; chest girth (CG), wither height (WH), body length (BL), and body condition score (BCS). Thirty-six males (approximately nine-months-old and 16.44 ± 0.33 kg average of initial body weight) were used across an eleven-week study from November–February 2019-2020. Each breed was divided into three groups (n = 6/group) and fed one of three rations: (1) concentrate mixture (Control) with crude protein 14% and energy 11.97% MJ/kg DM; (2) the same concentrate feed with the addition of 2 gm /capita daily Spirulina platensis (Treatment 1) and (3) the same concentrate feed with the addition of 4 gm /capita daily Spirulina platensis (Treatment 2). Analysis of weekly data collections for all traits indicated a significant effect of feeding Spirulina on all the studied traits except WH and BL. Analysis of variance for fixed effects in this study (damage and kid birth type i.e., single, twin or triple) were not significant for all studied traits. However, the breed effect was highly significant (P < 0.001) on BWT, ADG, BCS, and CG traits. On the other hand, when the analysis was done for the treatment effect within breeds for ADG, the Sahrawi breed had a significant effect (P < 0.05) at 56.52, 85.51, and 85.50 g/day for control, treatment 1 and treatment 2, respectively. This is a 51% difference between the control and treatment 1 (2 gm /capita). Whereas for the Jabbali breed, the treatment effect was not significant for ADG (P =0.55), and the actual ADG was 104.59, 118.84, and 114.25 g/day for control, treatment 1, and treatment 2, respectively, providing a 14% difference between the control group and the treated group (4 gm /capita). These findings indicate using Spirulina supplementation in Omani goat diets is recommended at 2 gm per capita as there was no benefit in feeding at 4 gm per capita for either breed. Farmers feeding Spirulina supplementation to kids after weaning at six-months could increase their herd performance and growth rate and facilitate buck selection at an earlier age.

Keywords: body conformation, goats, live weight, spirulina

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Authors: Man-Yun Liu, Emily Chia-Yu Su

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Alzheimer's disease (AD) is the public health crisis of the 21st century. AD is a degenerative brain disease and the most common cause of dementia, a costly disease on the healthcare system. Unfortunately, the cause of AD is poorly understood, furthermore; the treatments of AD so far can only alleviate symptoms rather cure or stop the progress of the disease. Currently, there are several ways to diagnose AD; medical imaging can be used to distinguish between AD, other dementias, and early onset AD, and cerebrospinal fluid (CSF). Compared with other diagnostic tools, blood (plasma) test has advantages as an approach to population-based disease screening because it is simpler, less invasive also cost effective. In our study, we used blood biomarkers dataset of The Alzheimer’s disease Neuroimaging Initiative (ADNI) which was funded by National Institutes of Health (NIH) to do data analysis and develop a prediction model. We used independent analysis of datasets to identify plasma protein biomarkers predicting early onset AD. Firstly, to compare the basic demographic statistics between the cohorts, we used SAS Enterprise Guide to do data preprocessing and statistical analysis. Secondly, we used logistic regression, neural network, decision tree to validate biomarkers by SAS Enterprise Miner. This study generated data from ADNI, contained 146 blood biomarkers from 566 participants. Participants include cognitive normal (healthy), mild cognitive impairment (MCI), and patient suffered Alzheimer’s disease (AD). Participants’ samples were separated into two groups, healthy and MCI, healthy and AD, respectively. We used the two groups to compare important biomarkers of AD and MCI. In preprocessing, we used a t-test to filter 41/47 features between the two groups (healthy and AD, healthy and MCI) before using machine learning algorithms. Then we have built model with 4 machine learning methods, the best AUC of two groups separately are 0.991/0.709. We want to stress the importance that the simple, less invasive, common blood (plasma) test may also early diagnose AD. As our opinion, the result will provide evidence that blood-based biomarkers might be an alternative diagnostics tool before further examination with CSF and medical imaging. A comprehensive study on the differences in blood-based biomarkers between AD patients and healthy subjects is warranted. Early detection of AD progression will allow physicians the opportunity for early intervention and treatment.

Keywords: Alzheimer's disease, blood-based biomarkers, diagnostics, early detection, machine learning

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Authors: Xiaojie Cheng, Ulrike Frank, Feng Zhao, Karin Pritsch

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Ragweed (Ambrosia artemisiifolia) is an invasive weed that has become an increasing global problem. In addition to affecting land use and crop yields, ragweed has a strong impact on human health as it produces highly allergenic pollen. Global warming will result in an earlier and longer pollen season enhanced pollen production and an increase in pollen allergenicity with a negative effect on atopic patients. The aims of this study were to investigate the effects of increasing temperature, the future climate scenario in the Munich area, southern Germany, predicted on the basis of RCP8.5 until the end of 2050s, or/and NO₂, a major air pollutant, 1) on the vegetative and reproductive characteristics of ragweed plants, 2) on the total allergenicity of ragweed pollen, 3) on the total pollen proteomic patterns. Ragweed plants were cultivated for the whole plant vegetation period under controlled conditions either under ambient climate conditions or 4°C higher temperatures with or without additional NO₂. Higher temperature resulted in bigger plant sizes, longer male inflorescences, and longer pollen seasons. The total allergenic potential of the pollen was accessed by dot blot using serum from ragweed pollen sensitized patients. The comparative immunoblot analysis revealed that the in vivo fumigation of ragweed plants with elevated NO₂-concentrations significantly increased the allergenic potential of the pollen, and in combination with increased temperature, the allergenic potential was even higher. On the other hand, label-free protein quantification by liquid chromatography-tandem mass spectrometry (LC-MS/MS) was performed. The results showed that more proteins were significantly up- and down-regulated under higher temperatures with/without elevated NO₂ conditions. Most of the highly expressed proteins were participating intensively in the metabolic process, the cellular process, and the stress defense process. These findings suggest that rising temperature and elevated NO₂ are important environmental factors for higher abiotic stress activities, catalytic activities, and thus higher allergenic potential observed in pollen proteins.

Keywords: climate change, NO₂, pollen proteome, ragweed, temperature

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Authors: Sze Wing Ho, Nagendra P. Shah

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Lactic acid bacteria (LAB) have shown the beneficial effects on human gastrointestinal tract, such as protects diarrhea induced by lactose intolerance or enteric pathogens. Citrulline is a non-protein amino acid and also the precursors of arginine and nitric oxide, it has shown to enhance intestinal barrier function. Citrulline has shown to improve the growth of some strains of LAB, it is important for LAB to have a sufficient cell concentration to contribute the effects. Therefore, the aims of this study were to investigate the effect of combining citrulline with LAB on the anti-adhesion effect against pathogens and the effect on the cell integrity. The effect of citrulline on selected LAB was determined by incubating in 0%, 0.1% or 0.2% citrulline enriched MRS broth for 18 h. The adhesion ability of LAB and the anti-adhesion effect of LAB and citrulline against pathogens were performed on IPEC-J2 cell line. Transepithelial electrical resistance (TEER) assay was used to measure the tight junction (TJ) integrity. TJ proteins (claudin-1, occludin and zonula occluden-1 (ZO-1)) were determined by western blot analysis. It found that the growth of Lactobacillus helveticus ASCC 511 was significantly stimulated by 0.2% citrulline compared with control during 18 h fermentation. The adhesion of L. helveticus ASCC 511 and Lactobacillus delbrueckii ssp. bulgaricus (L. bulgaricus) ASCC 756 was increased when supplemented with citrulline. Citrulline has shown significant inhibitory effect on the adhesion of Escherichia coli PELI0480 (O157:H7), Shigella sonnei ATCC 25931, Staphyloccocus aureus CMCC26003 and Cronobacter sakazakii ATCC 29544. The anti-adhesion effect of L. helveticus ASCC 511, L. bulgaricus ASCC 756 and Lactobacillus paracasei ASCC 276 against Cronobacter sakazakii ATCC 29544 was significantly enhanced with citrulline supplementation. Treatments with citrulline and LAB were able to maintain the TEER of IPEC-J2 cell and shown the positive effect on the TJ proteins. In conclusion, citrulline had stimulating effect on some strains of LAB and determined to improve the adhesion of LAB on intestinal epithelial cell, to enhance the inhibitory effect on enteric pathogens adhesion as well as had beneficial effects on maintaining cell integrity. It implied LAB supplemented with citrulline might have advantageous effects on gastrointestinal tracts.

Keywords: citrulline, lactic acid bacteria, amino acid, anti-adhesion effect, cell integrity

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Authors: Galuh Asri Bestari, Hajar Shofiyya

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Double burden of malnutrition (DBM) has been a global problem in these last decades occurs in both developed and developing countries. Overweight in adults and stunting among preschool children have dramatically increased and become the main problems of malnutrition that should be solved immediately since they are directly related with the health status and productivity. Reformulation of food product by using the local sea resources called terasi shrimp (Acetes indicus) has a potential possibility in facing the DBM. A study was carried out in Indonesia to determine the acceptability of terasi shrimp powder through sensory evaluation. Terasi shrimps were processed into powder form through sun drying and pounding methods. The powder form was directly added in food as alternative seasonings and tested among stunted and normal preschool children. Meanwhile, a further processing method is given to the shrimp powder tested in overweight and normal-weighed adults. The shrimp powder was mixed with sago flour and formed into balls, then steamed for 15-20 minutes, and finally served as alternative snacks. Based on the sensory evaluation, the shrimp powder has a good acceptance in taste (54%), shape (60%), and color properties (63%), while the shrimp balls has a good acceptance in size (65%), shape (50%), color (48%), taste (40%), and texture (36%). Terasi shrimp powder can be stored for a month in room temperature. In addition, carried out chemical analysis revealed that terasi shrimp (Acetes indicus) has higher percentage of protein, calcium, and iron than other animal sources, but conversely contains zero sodium and very low percentage of fat. Terasi shrimp’s shell also contains a substance called chitosan which acts by forming gels in the intestinal tract to entrap lipids, thus interfering with their absorption. After going through some processing methods, the shrimp powder and balls did not show any significant changes in their nutrient contents. So that, terasi shrimp powder is good to be consumed not only by overweight adults, but also by children to support their optimum growth. Intervention of terasi shrimp powder should be implemented step by step from national up to global governance program to face the DBM.

Keywords: Acetes indicus, alternative food, double burden of malnutrition, sensory evaluation

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Authors: P. Joris, E. Lombard, X. Cameleyre, G. Navarro, A. Paillet, N. Gorret, S. E. Guillouet

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Today, on the international space station, multiple supplies are needed per year to supply food and spare parts and to take out waste. But as it is planned to go longer and further into space these supplies will no longer be possible. The astronaut life support system must be able of continuously transform waste into valuable compounds. Two types of production were identified as critical and could be be supplemented by microorganisms. On the one hand, since microgravity causes rapid muscle loss, single cell proteins (SCPs) could be used as protein rich feed or food. On the other hand, having enough building materials to build an advanced habitat will not be possible only by transporting space goods from earth to mars for example. The bacterium Cupriavidus. necator is well known for its ability to produce a large amount of proteins or of polyhydroxyalkanoate biopolymers (PHAs) depending on its implementation. By coupling the life support system to a 3D-printer, astronauts could be supplied with an unlimited amount of building materials. Additionally, based on the design of the life support system, waste streams have been identified: urea from the crew urine and volatile fatty acids (VFAs) from a first stage of organic waste (excrement and food waste) treatment through anaerobic digestion. Thus, the objective of this, within the Spaceship.Fr project, was to demonstrate the feasibility of producing SCPs and PHAs from VFAs and urea in bioreactor. Because life support systems operate continuously as loops, continuous culture experiments were chosen and the effect of the bioreactor dilution rate on biomass composition was investigated. Total transformation of the carbon source into biomass with high SCP or PHA content was achieved in all cases. We will present the transformation performances of VFAs and urea by the bacteria in bioreactor in terms of titers, yields and productivities but also in terms of the quality of SCP and PHA produced, nucleic acid content. We will further discuss the envisioned integration of our process within life support systems.

Keywords: life support system, space travel, waste treatment, single cell proteins, polyhydroxyalkanoates, bioreactor

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Authors: Himalaya Patir, Bitupon Baruah, Sanjay Gayary, Subhajit Ray

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In recent age significant importance is given for the development of nutritious and health beneficial foods. Fruit juices collected from different fruits when blended that improves not only the physicochemical and nutritional properties but also enhance the sensorial or organoleptic properties. The study was carried out to determine the physico-chemical, nutritional, microbiological analysis and sensory evaluation of mixed fruit juice blend. Juice of orange (Citrus sinensis), apple (Malus domestica), mosambi (Citrus limetta) were blended in the ratio of sample-I (30% apple:30% orange:40% mosambi), sample-II ( 40% apple :30% orange :30% mosambi), sample-III (30% apple :40% orange :30% mosambi) , sample-IV (50% apple :30% orange :20% mosambi), sample-V (30% apple:20% orange:50% mosambi), sample-VI (20% apple :50% orange :30% mosambi) to evaluate all quality characteristics. Their colour characteristics in terms of hue angle, chroma and colour difference (∆E) were evaluated. The physico-chemical parameters analysis carried out were total soluble solids (TSS), total titratable acidity (TTA), pH, acidity (FA), volatile acidity (VA), pH, and vitamin C. There were significant differences (p˂0.05) in the TSS of the samples. However, sample-V (30% apple: 20% orange: 50% mosambi) provides the highest TSS of 9.02gm and significantly differed from other samples (p˂0.05). Sample-IV (50% apple: 30% orange: 20% mosambi) was shown the highest titratable acidity (.59%) in comparison to other samples. The highest value of pH was found as 5.01 for sample-IV (50% apple: 30% orange: 20% mosambi). Sample-VI (20% apple: 50% orange :30% mosambi) blend has the highest hue angle, chroma and colour changes of 72.14,25.29 and 54.48 and vitamin C, i.e. Ascorbic acid (.33g/l) content compared to other samples. The nutritional compositions study showed that, sample- VI (20% apple: 50% orange: 30% mosambi) has the significantly higher carbohydrate (51.67%), protein (.78%) and ash (1.24%) than other samples, while sample-V (30% apple: 20% orange: 50% mosambi) has higher dietary fibre (12.84%) and fat (2.82%) content. Microbiological analysis of all samples in terms of total plate count (TPC) ranges from 44-60 in 101 dilution and 4-5 in 107 dilutions and was found satisfactory. Moreover, other pathogenic bacterial count was found nil. The general acceptability of the mixed fruit juice blend samples were moderately liked by the panellists, and sensorial quality studies showed that sample-V (30% apple: 20% orange: 50% mosambi) contains highest overall acceptability of 8.37 over other samples and can be considered good for consumption.

Keywords: microbiological, nutritional, physico-chemical, sensory properties

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Authors: Lee Yujin, Han Jiye, Oh Jin-Woo

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The adverse effects of endocrine disrupting chemicals (EDCs) has attracted considerable public interests. The benzene-like EDCs structure mimics the mechanisms of hormones naturally occurring in vivo, and alters physiological function of the endocrine system. Although, some of the most representative EDCs such as polychlorinated biphenyls (PCBs) and phthalates compounds already have been prohibited to produce and use in many countries, however, PCBs and phthalates in plastic products as flame retardant and plasticizer are still circulated nowadays. EDCs can be released from products while using and discarding, and it causes serious environmental and health issues. Here, we developed virus-based structurally coloured nanostructure that can detect minute EDCs concentration sensitively and selectively. These structurally coloured nanostructure exhibits characteristic angel-independent colors due to the regular virus bundle structure formation through simple pulling technique. The designed number of different colour bands can be formed through controlling concentration of virus solution and pulling speed. The virus, M-13 bacteriophage, was genetically engineered to react with specific ECDs, typically PCBs and phthalates. M-13 bacteriophage surface (pVIII major coat protein) was decorated with benzene derivative binding peptides (WHW) through phage library method. In the initial assessment, virus-based color sensor was exposed to several organic chemicals including benzene, toluene, phenol, chlorobenzene, and phthalic anhydride. Along with the selectivity evaluation of virus-based colour sensor, it also been tested for sensitivity. 10 to 300 ppm of phthalic anhydride and chlorobenzene were detected by colour sensor, and showed the significant sensitivity with about 90 of dissociation constant. Noteworthy, all measurements were analyzed through principal component analysis (PCA) and linear discrimination analysis (LDA), and exhibited clear discrimination ability upon exposure to 2 categories of EDCs (PCBs and phthalates). Because of its easy fabrication, high sensitivity, and the superior selectivity, M-13 bacteriophage-based color sensor could be a simple and reliable portable sensing system for environmental monitoring, healthcare, social security, and so on.

Keywords: M-13 bacteriophage, colour sensor, genetic engineering, EDCs

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Authors: Zahra Shokrolahi, Mohammad Reza Atashzar

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The goals of treating patients with breast cancer are to cure the disease, prolong survival, and improve quality of life. Immune cells in the tumor microenvironment have an important role in regulating tumor progression. The term of cellular immunotherapy refers to the administration of living cells to a patient; this type of immunotherapy can be active, such as a dendritic cell (DC) vaccine, in that the cells can stimulate an anti-tumour response in the patient, or the therapy can be passive, whereby the cells have intrinsic anti-tumour activity; this is known as adoptive cell transfer (ACT) and includes the use of autologous or allogeneic lymphocytes that may, or may not, be modified. The most important breast cancer cellular immunotherapies involving the use of T cells and natural killer (NK) cells in adoptive cell transfer, as well as dendritic cells vaccines. T cell-based therapies including tumour-infiltrating lymphocytes (TILs), engineered TCR-T cells, chimeric antigen receptor (CAR T cell), Gamma-delta (γδ) T cells, natural killer T (NKT) cells. NK cell-based therapies including lymphokine-activated killers (LAK), cytokine-induced killer (CIK) cells, CAR-NK cells. Adoptive cell therapy has some advantages and disadvantages some. TILs cell strictly directed against tumor-specific antigens but are inactive against tumor changes due to immunoediting. CIK cell have MHC-independent cytotoxic effect and also need concurrent high dose IL-2 administration. CAR T cell are MHC-independent; overcome tumor MHC molecule downregulation; potent in recognizing any cell surface antigen (protein, carbohydrate or glycolipid); applicable to a broad range of patients and T cell populations; production of large numbers of tumor-specific cells in a moderately short period of time. Meanwhile CAR T cells capable of targeting only cell surface antigens; lethal toxicity due to cytokine storm reported. Here we present the most popular cancer cellular immunotherapy approaches and discuss their clinical relevance referring to data acquired from clinical trials .To date, clinical experience and efficacy suggest that combining more than one immunotherapy interventions, in conjunction with other treatment options like chemotherapy, radiotherapy and targeted or epigenetic therapy, should guide the way to cancer cure.

Keywords: breast cancer , cell therapy , CAR T cell , CIK cells

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Authors: Evgeniya Y. Yuzbasheva, Tigran V. Yuzbashev, Natalia I. Perkovskaya, Elizaveta B. Mostova

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Cell-surface display of lipase is of great interest as it has many applications in the field of biotechnology owing to its unique advantages: simplified product purification, and cost-effective downstream processing. One promising area of application for whole-cell biocatalysts with surface displayed lipase is biodiesel synthesis. Biodiesel is biodegradable, renewable, and nontoxic alternative fuel for diesel engines. Although the alkaline catalysis method has been widely used for biodiesel production, it has a number of limitations, such as rigorous feedstock specifications, complicated downstream processes, including removal of inorganic salts from the product, recovery of the salt-containing by-product glycerol, and treatment of alkaline wastewater. Enzymatic synthesis of biodiesel can overcome these drawbacks. In this study, Lip2p lipase was displayed on Yarrowia lipolytica cells via C- and N-terminal fusion variant. The active site of lipase is located near the C-terminus, therefore to prevent the activity loosing the insertion of glycine-serine linker between Lip2p and C-domains was performed. The hydrolytic activity of the displayed lipase reached 12,000–18,000 U/g of dry weight. However, leakage of enzyme from the cell wall was observed. In case of C-terminal fusion variant, the leakage was occurred due to the proteolytic cleavage within the linker peptide. In case of N-terminal fusion variant, the leaking enzyme was presented as three proteins, one of which corresponded to the whole hybrid protein. The calculated number of recombinant enzyme displayed on the cell surface is approximately 6–9 × 105 molecules per cell, which is close to the theoretical maximum (2 × 106 molecules/cell). Thus, we attribute the enzyme leakage to the limited space available on the cell surface. Nevertheless, cell-bound lipase exhibited greater stability to short-term and long-term temperature treatment than the native enzyme. It retained 74% of original activity at 60°C for 5 min of incubation, and 83% of original activity after incubation at 50°C during 5 h. Cell-bound lipase had also higher stability in organic solvents and detergents. The developed whole-cell biocatalyst was used for recycling biodiesel synthesis. Two repeated cycles of methanolysis yielded 84.1–% and 71.0–% methyl esters after 33–h and 45–h reactions, respectively.

Keywords: biodiesel, cell-surface display, lipase, whole-cell biocatalyst

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Authors: Ethel E. Phiri, Lionel Hartzenberg, Percy Chimwamuromba, Emmanuel Nepolo, Jens Kossmann, James R. Lloyd

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Orphan crops are underresearched and underutilized food plant species that have not been categorized as major food crops, but have the potential to be economically and agronomically significant. They have been documented to have the ability to tolerate extreme environmental conditions. However, limited research has been conducted to uncover their potential as food crop species. The New Partnership for Africa’s Development (NEPAD) has classified Marama bean, Tylosema esculentum, as an orphan crop. The plant is one of the 101 African orphan crops that must have their genomes sequenced, assembled, and annotated in the foreseeable future. Marama bean is a perennial leguminous plant that primarily grows in poor, arid soils in southern Africa. The plants produce large tubers that can weigh as much as 200kg. While the foliage provides fodder, the tuber is carbohydrate rich and is a staple food source for rural communities in Namibia. Also, the edible seeds are protein- and oil-rich. Marama Bean plants respond rapidly to increased temperatures and severe water scarcity without extreme consequences. Advances in molecular biology and biotechnology have made it possible to effectively transfer technologies between model- and major crops to orphan crops. In this research, the aim was to assemble the first transcriptomic analysis of Marama Bean RNA-sequence data. Many model plant species have had their genomes sequenced and their transcriptomes assembled. Therefore the availability of transcriptome data for a non-model crop plant species will allow for gene identification and comparisons between various species. The data has been sequenced using the Ilumina Hiseq 2500 sequencing platform. Data analysis is underway. In essence, this research will eventually evaluate the potential use of Marama Bean as a crop species to improve its value in agronomy. data for a non-model crop plant species will allow for gene identification and comparisons between various species. The data has been sequenced using the Ilumina Hiseq 2500 sequencing platform. Data analysis is underway. In essence, this researc will eventually evaluate the potential use of Marama bean as a crop species to improve its value in agronomy.

Keywords: 101 African orphan crops, RNA-Seq, Tylosema esculentum, underutilised crop plants

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Authors: Martina I. Reinhold, Theresa Bacon-Baguley

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Background: Medical knowledge continuously evolves and to help health care providers to stay up-to-date, evidence-based medicine (EBM) has emerged as a model. The practice of EBM requires new skills of the health care provider, including directed literature searches, the critical evaluation of research studies, and the direct application of the findings to patient care. This paper describes the integration and evaluation of an evidence-based medicine course sequence into a Physician Assistant curriculum. This course sequence teaches students to manage and use the best clinical research evidence to competently practice medicine. A survey was developed to assess the outcomes of the EBM course sequence. Methodology: The cornerstone of the three-semester sequence of EBM are interactive small group discussions that are designed to introduce students to the most clinically applicable skills to identify, manage and use the best clinical research evidence to improve the health of their patients. During the three-semester sequence, the students are assigned each semester to participate in small group discussions that are facilitated by faculty with varying background and expertise. Prior to the start of the first EBM course in the winter semester, PA students complete a knowledge-based survey that was developed by the authors to assess the effectiveness of the course series. The survey consists of 53 Likert scale questions that address the nine objectives for the course series. At the end of the three semester course series, the same survey was given to all students in the program and the results from before, and after the sequence of EBM courses are compared. Specific attention is paid to overall performance of students in the nine course objectives. Results: We find that students from the Class of 2016 and 2017 consistently improve (as measured by percent correct responses on the survey tool) after the EBM course series (Class of 2016: Pre- 62% Post- 75%; Class of 2017: Pre- 61 % Post-70%). The biggest increase in knowledge was observed in the areas of finding and evaluating the evidence, with asking concise clinical questions (Class of 2016: Pre- 61% Post- 81%; Class of 2017: Pre- 61 % Post-75%) and searching the medical database (Class of 2016: Pre- 24% Post- 65%; Class of 2017: Pre- 35 % Post-66 %). Questions requiring students to analyze, evaluate and report on the available clinical evidence regarding diagnosis showed improvement, but to a lesser extend (Class of 2016: Pre- 56% Post- 77%; Class of 2017: Pre- 56 % Post-61%). Conclusions: Outcomes identified that students did gain skills which will allow them to apply EBM principles. In addition, the outcomes of the knowledge-based survey allowed the faculty to focus on areas needing improvement, specifically the translation of best evidence into patient care. To address this area, the clinical faculty developed case scenarios that were incorporated into the lecture and discussion sessions, allowing students to better connect the research studies with patient care. Students commented that ‘class discussion and case examples’ contributed most to their learning and that ‘it was helpful to learn how to develop research questions and how to analyze studies and their significance to a potential client’. As evident by the outcomes, the EBM courses achieved the goals of the course and were well received by the students. 

Keywords: evidence-based medicine, clinical education, assessment tool, physician assistant

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Authors: Adriana Sosa, Susana Rincon, Chérif Ben, Diana Cabañas, Juan E. Ruiz, Alejandro Zepeda

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The complex chemical composition (mixtures of ammonium and recalcitrant compounds) of the effluents from the chemical, pharmaceutical and petrochemical industries represents a challenge in their biological treatment. This treatment involves nitrification process that can suffer an inhibition due to the presence of aromatic compounds giving as a result the decrease of the process efficiency. The inhibitory effects on nitrification in the presence of aromatic compounds have already been studied; however a few studies have considered the presence of phenolic compounds in the form of mixtures, which is the form that they are present in real context. For this reason, we realized a kinetic study on the nitrifying process in the presence of different concentrations of a mixture of phenol, o-cresol, m-cresol and p-cresol (0 - 320 mg C/L) in a sequencing batch reactor (SBR). Firstly, the nitrifying process was evaluated in absence of the phenolic mixture (control 1) in a SBR with 2 L working volume and 176 mg/L of nitrogen of microbial protein. Total oxidation of initial ammonium (efficiency; ENH4+ of 100 %) to nitrate (nitrifying yield; YNO3- of 0.95) were obtained with specific rates of ammonium consumption (qN-NH4+) and nitrate production (qN-NO3-) (of 1.11 ± 0.04 h-1 and 0.67 h-1 ± 0.11 respectively. During the phase of acclimation with 40 mg C/L of the phenolic mixture, an inhibitory effect on the nitrifying process was observed, provoking a decrease in ENH4+ and YNO3- (11 and 54 % respectively) as well as in the specific rates (89 y 46 % respectively), being the ammonia oxidizing bacteria (BAO) the most affected. However, in the next cycles without the phenolic mixture (control 2), the nitrifying consortium was able to recover its nitrifying capacity (ENH4+ = 100% and YNO3-=0.98). Afterwards the SBR was fed with 10 mg C/L of the phenolic mixture, obtaining and ENH4+ of 100%, YNO3- and qN-NH4+ 0.62 ± 0.006 and 0.13 ± 0.004 respectively, while the qN-NO3- was 0.49 ± 0.007. Moreover, with the increase of the phenolic concentrations (10-160 mg C/L) and the number of cycles the nitrifying consortium was able to oxidize the ammonia with ENH4+ of 100 % and YNO3- close to 1. However a decrease in the values of the nitrification specific rates and increase in the oxidation in phenolic compounds (70 to 94%) were observed. Finally, in the presence of 320 mg C/L, the nitrifying consortium was able to simultaneously oxidize the ammonia (ENH4+= 100%) and the phenolic mixture (p-cresol>phenol>m-cresol>o-cresol) being the o-cresol the most recalcitrant compound. In all the experiments the use of a SBR allowed a respiratory adaptation of the consortium to oxidize the phenolic mixture achieving greater adaptation of the nitrite-oxidizing bacteria (NOB) than in the ammonia-oxidizing bacteria (AOB).

Keywords: cresol, inhibition, nitrification, phenol, sequencing batch reactor

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Authors: Iman Farasat, Howard M. Salis

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Engineered genetic circuits reprogram cellular behavior to act as living computers with applications in detecting cancer, creating self-controlling artificial tissues, and dynamically regulating metabolic pathways. Phenemenological models are often used to simulate and design genetic circuit behavior towards a desired behavior. While such models assume that each circuit component’s function is modular and independent, even small changes in a circuit (e.g. a new promoter, a change in transcription factor expression level, or even a new media) can have significant effects on the circuit’s function. Here, we use statistical thermodynamics to account for the several factors that control transcriptional regulation in bacteria, and experimentally demonstrate the model’s accuracy across 825 measurements in several genetic contexts and hosts. We then employ our first principles model to design, experimentally construct, and characterize a family of signal amplifying genetic circuits (genetic OpAmps) that expand the dynamic range of cell sensors. To develop these models, we needed a new approach to measuring the in vivo binding free energies of transcription factors (TFs), a key ingredient of statistical thermodynamic models of gene regulation. We developed a new high-throughput assay to measure RNA polymerase and TF binding free energies, requiring the construction and characterization of only a few constructs and data analysis (Figure 1A). We experimentally verified the assay on 6 TetR-homolog repressors and a CRISPR/dCas9 guide RNA. We found that our binding free energy measurements quantitatively explains why changing TF expression levels alters circuit function. Altogether, by combining these measurements with our biophysical model of translation (the RBS Calculator) as well as other measurements (Figure 1B), our model can account for changes in TF binding sites, TF expression levels, circuit copy number, host genome size, and host growth rate (Figure 1C). Model predictions correctly accounted for how these 8 factors control a promoter’s transcription rate (Figure 1D). Using the model, we developed a design framework for engineering multi-promoter genetic circuits that greatly reduces the number of degrees of freedom (8 factors per promoter) to a single dimensionless unit. We propose the Ptashne (Pt) number to encapsulate the 8 co-dependent factors that control transcriptional regulation into a single number. Therefore, a single number controls a promoter’s output rather than these 8 co-dependent factors, and designing a genetic circuit with N promoters requires specification of only N Pt numbers. We demonstrate how to design genetic circuits in Pt number space by constructing and characterizing 15 2-repressor OpAmp circuits that act as signal amplifiers when within an optimal Pt region. We experimentally show that OpAmp circuits using different TFs and TF expression levels will only amplify the dynamic range of input signals when their corresponding Pt numbers are within the optimal region. Thus, the use of the Pt number greatly simplifies the genetic circuit design, particularly important as circuits employ more TFs to perform increasingly complex functions.

Keywords: transcription factor, synthetic biology, genetic circuit, biophysical model, binding energy measurement

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Authors: Portia Murphy, Danica Vasic, Anoja W. Gunaratne, Encarnita Sitchon, Teresita Tugonon, Marou Ison, Antoinette Le Busque, Christelle Pagonis, Thomas J. Borody

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Irritable bowel syndrome (IBS) is a chronic gastrointestinal disorder that affects an estimated 11% of the population globally with the most predominant symptoms being abdominal pain, bloating and altered bowel movements. All age groups suffer from IBS although the prevalence of IBS decreases for age groups over 50 years. Women are more likely to suffer from IBS than men. IBS can be categorized into 3 groups based on the type of altered bowel movement: diarrhea-predominant IBS (IBS-D), constipation-predominant IBS (IBS-C) and IBS with mixed bowel habit (IBS-M). The contribution of the gut microbiome to the etiology of IBS is becoming increasingly recognized with rising use of anti-microbial agents. Previous studies on vancomycin and rifaximin used as monotherapy or in combination have been conducted mainly on IBS-C and showed marked improvements in the symptoms. According to our knowledge, no studies reported using these two combinations of antibiotics for IBS-D. Here, we report a consecutive cohort of 18 patients treated with both vancomycin and rifaximin for IBS-D. These patients’ records were reviewed retrospectively. In this cohort, patients ages were between 24-74 years (mean 44 years) and 9 were female. Baseline all patients had diarrhea, 4 with mucus and one with blood. Patients reported other symptoms were abdominal pain (n=11) bloating (n=9), flatulence (n=7), fatigue (n=4) and nausea (n=3). Patients treatments were personalized according to their symptom severity and tolerability and were treated with combination of rifaximin (500 - 3000mg/d) and vancomycin (500mg - 1500mg/d) for an ongoing period. Follow-ups were conducted between 2-32 weeks’ time. Of all patients, 89% patients reported improvement of the symptoms, 1 reported no change and 1 patient’s symptoms got worse. The mechanism of action for both vancomycin and rifaximin involves the inhibition of bacterial cell wall and protein synthesis respectively. The role of these medications in improving the symptoms of this cohort suggests that IBS-D may be microbiome infection driven. In this cohort, similar patient presentations to Clostridium difficile, as well as symptom improvement with the use of rifaximin and particularly vancomycin, suggest that the infectious agent may be an unidentified Clostridium. These preliminary results offer an alternative etiology for IBS-D not previously considered and open the avenue for new research.

Keywords: clostridium deficile, diarrhea predominant Irritable Bowel Syndrome, microbiome, vancomycin/rifaximin combination

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Authors: Yudia Mailani Putri, Martalena Purba, B. J. Istiti Kandarina

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Background: Obesity has been recognized as a global health problem. Individuals classified as overweight and obese are increasing at an alarming rate. This condition is associated with psychological and physiological problems. as a person reaches adulthood, somatic growth ceases. At this stage, the human body has developed fully, to a stable state. As the capital of Riau Province in Indonesia, Pekanbaru is dominated by Malay ethnic population habitually consuming cholesterol-rich fatty foods as a daily menu, a trigger to the onset of obesity resulting in high prevalence of degenerative diseases. Research objectives: The aim of this study is elaborating the relationship between high-fat consumption pattern, stress level, physical activity and the incidence of obesity in adult women in Pekanbaru city. Research Methods: Among the combined research methods applied in this study, the first stage is quantitative observational, analytical cross-sectional research design with adult women aged 20-40 living in Pekanbaru city. The sample consists of 200 women with BMI≥25. Sample data is processed with univariate, bivariate (correlation and simple linear regression) and multivariate (multiple linear regression) analysis. The second phase is qualitative descriptive study purposive sampling by in-depth interviews. six participants withdrew from the study. Results: According to the results of the bivariate analysis, there are relationships between the incidence of obesity and the pattern of high fat foods consumption (energy intake (p≤0.000; r = 0.536), protein intake (p≤0.000; r=0.307), fat intake (p≤0.000; r=0.416), carbohydrate intake (p≤0.000; r=0.430), frequency of fatty food consumption (p≤0.000; r=0.506) and frequency of viscera foods consumption (p≤0.000; r=0.535). There is a relationship between physical activity and incidence of obesity (p≤0.000; r=-0.631). However, there is no relationship between the level of stress (p=0.741; r=0.019-) and the incidence of obesity. Physical activity is a predominant factor in the incidence of obesity in adult women in Pekanbaru city. Conclusion: There are relationships between high-fat food consumption pattern, physical activity and the incidence of obesity in Pekanbaru city whereas physical activity is a predominant factor in the occurrence of obesity, supported by the unchangeable pattern of high-fat foods consumption.

Keywords: obesity, adult, high in fat, stress, physical activity, consumption pattern

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Authors: Francielo Vendruscolo

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During industrial processing of raw materials of animal and vegetable origin, large amounts of solid, liquid and gaseous wastes are generated. Solid residues are usually materials rich in carbohydrates, protein, fiber and minerals. Brewer’s spent grain (BSG) is the main waste generated in the brewing industry, representing 85% of the waste generated in this industry. It is estimated that world’s BSG generation is approximately 38.6 x 106 t per year and represents 20-30% (w/w) of the initial mass of added malt, resulting in low commercial value by-product, however, does not have economic value, but it must be removed from the brewery, as its spontaneous fermentation can attract insects and rodents. For every 100 grams in dry basis, BSG has approximately 68 g total fiber, being divided into 3.5 g of soluble fiber and 64.3 g of insoluble fiber (cellulose, hemicellulose and lignin). In addition to dietary fibers, depending on the efficiency of the grinding process and mashing, BSG may also have starch, reducing sugars, lipids, phenolics and antioxidants, emphasizing that its composition will depend on the barley variety and cultivation conditions, malting and technology involved in the production of beer. BSG demands space for storage, but studies have proposed alternatives such as the use of drying, extrusion, pressing with superheated steam, and grinding to facilitate storage. Other important characteristics that enhance its applicability in bioremediation, effluent treatment and biotechnology, is the surface area (SBET) of 1.748 m2 g-1, total pore volume of 0.0053 cm3 g-1 and mean pore diameter of 121.784 Å, characterized as a macroporous and possess fewer adsorption properties but have great ability to trap suspended solids for separation from liquid solutions. It has low economic value; however, it has enormous potential for technological applications that can improve or add value to this agro-industrial waste. Due to its composition, this material has been used in several industrial applications such as in the production of food ingredients, fiber enrichment by its addition in foods such as breads and cookies in bioremediation processes, substrate for microorganism and production of biomolecules, bioenergy generation, and civil construction, among others. Therefore, the use of this waste or by-product becomes essential and aimed at reducing the amount of organic waste in different industrial processes, especially in breweries.

Keywords: brewer’s spent malt, agro-industrial residue, lignocellulosic material, waste generation

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Authors: Kitti Szoke, Laszlo Szoke, Attila Czompa, Arpad Tosaki, Istvan Lekli

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Autophagy plays an important role in cardiac hypertrophy, which is one of the most common causes of heart failure in the world. This self-degradative catabolic process, responsible for protein quality control, balancing sources of energy at critical times, and elimination of damaged organelles. The autophagic activity can be triggered by starvation, oxidative stress, or pharmacological agents, like rapamycin. This induced autophagy can promote cell survival during starvation or pathological stress. In this study, it is investigated the effect of the induced autophagic process on angiotensin induced hypertrophic H9c2 cells. In our study, it is used H9c2 cells as an in vitro model. To induce hypertrophy, cells were treated with 10000 nM angiotensin-II, and to activate autophagy, 100 nM rapamycin treatment was used. The following groups were formed: 1: control, 2: 10000 nM AT-II, 3: 100 nM rapamycin, 4: 100 nM rapamycin pretreatment then 10000 nM AT-II. The cell viability was examined via MTT (cell proliferation assay) assay. The cells were stained with rhodamine-conjugated phalloidin and DAPI to visualize F-actin filaments and cell nuclei then the cell size alteration was examined in a fluorescence microscope. Furthermore, the expression levels of autophagic and apoptotic proteins such as Beclin-1, p62, LC3B-II, Cleaved Caspase-3 were evaluated by Western blot. MTT assay result suggests that the used pharmaceutical agents in the tested concentrations did not have a toxic effect; however, at group 3, a slight decrement was detected in cell viability. In response to AT-II treatment, a significant increase was detected in the cell size; cells became hypertrophic. However, rapamycin pretreatment slightly reduced the cell size compared to group 2. Western blot results showed that AT-II treatment-induced autophagy, because the increased expression of Beclin-1, p62, LC3B-II were observed. However, due to the incomplete autophagy, the apoptotic Cleaved Caspase-3 expression also increased. Rapamycin pretreatment up-regulated Beclin-1 and LC3B-II, down-regulated p62 and Cleaved Caspase-3, indicating that rapamycin-induced autophagy can restore the normal autophagic flux. Taken together, our results suggest that rapamycin activated autophagy reduces angiotensin-II induced hypertrophy.

Keywords: angiotensin-II, autophagy, H9c2 cell line, hypertrophy, rapamycin

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Authors: Sumit Kanwar, Manisha Chand, Gregory Gilot

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Background: Septic arthritis of the shoulder has infrequently been discussed. Focus on infection of the untouched shoulder has not heretofore been described. We present four patients with glenohumeral septic arthritis. Methods: Case 1: A 59 year old male with left shoulder pain in the anterior, posterior and superior aspects. Case 2: A 60 year old male with fever, chills, and generalized muscle aches. Case 3: A 70 year old male with right shoulder pain about the anterior and posterior aspects. Case 4: A 55 year old male with global right shoulder pain, swelling, and limited ROM. Results: In case 1, the left shoulder was affected. Physical examination, swelling was notable, there was global tenderness with a painful range of motion (ROM). The lab values indicated an erythrocyte sedimentation rate (ESR) of 96, and a C-reactive protein (CRP) of 304.30. Imaging studies were performed and MRI indicated a high suspicion for an abscess with osteomyelitis of the humeral head. Our second case’s left arm was affected. He had swelling, global tenderness and painful ROM. His ESR was 38, CRP was 14.9. X-ray showed severe arthritis. Case 3 differed with the right arm being affected. Again, global tenderness and painful ROM was observed. His ESR was 94, and CRP was 10.6. X-ray displayed an eroded glenoid space. Our fourth case’s right shoulder was affected. He had global tenderness and painful, limited ROM. ESR was 108 and CRP was 2.4. X-ray was non-significant. Discussion: Monoarticular septic arthritis of the virgin glenohumeral joint is seldom diagnosed in clinical practice. Common denominators include elevated ESR, painful, limited ROM, and involvement of the dominant arm. The male population is more frequently affected with an average age of 57. Septic arthritis is managed with incision and drainage or needle aspiration of synovial fluid supplemented with 3-6 weeks of intravenous antibiotics. Due to better irrigation and joint visualization, arthroscopy is preferred. Open surgical drainage may be indicated if the above methods fail. Conclusion: If a middle-aged male presents with vague anterior or posterior shoulder pain, elevated inflammatory markers and a low grade fever, an x-ray should be performed. If this displays degenerative joint disease, the complete further workup with advanced imaging, such as an MRI, CT scan, or an ultrasound. If these imaging modalities display anterior space joint effusion with soft tissue involvement, we can suspect septic arthritis of the untouched glenohumeral joint and surgery is indicated.

Keywords: glenohumeral joint, identification, infection, septic arthritis, shoulder

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Authors: Ahmed Bahieldin, Ahmed Atef, Jamal S. M. Sabir, Nour O. Gadalla, Sherif Edris, Ahmed M. Alzohairy, Nezar A. Radhwan, Mohammed N. Baeshen, Ahmed M. Ramadan, Hala F. Eissa, Sabah M. Hassan, Nabih A. Baeshen, Osama Abuzinadah, Magdy A. Al-Kordy, Fotouh M. El-Domyati, Robert K. Jansen

Abstract:

Wild salt-tolerant barley (Hordeum spontaneum) is the ancestor of cultivated barley (Hordeum vulgare or H. vulgare). Although the cultivated barley genome is well studied, little is known about genome structure and function of its wild ancestor. In the present study, RNA-Seq analysis was performed on young leaves of wild barley treated with salt (500 mM NaCl) at four different time intervals. Transcriptome sequencing yielded 103 to 115 million reads for all replicates of each treatment, corresponding to over 10 billion nucleotides per sample. Of the total reads, between 74.8 and 80.3% could be mapped and 77.4 to 81.7% of the transcripts were found in the H. vulgare unigene database (unigene-mapped). The unmapped wild barley reads for all treatments and replicates were assembled de novo and the resulting contigs were used as a new reference genome. This resultedin94.3 to 95.3%oftheunmapped reads mapping to the new reference. The number of differentially expressed transcripts was 9277, 3861 of which were uni gene-mapped. The annotated unigene- and de novo-mapped transcripts (5100) were utilized to generate expression clusters across time of salt stress treatment. Two-dimensional hierarchical clustering classified differential expression profiles into nine expression clusters, four of which were selected for further analysis. Differentially expressed transcripts were assigned to the main functional categories. The most important groups were ‘response to external stimulus’ and ‘electron-carrier activity’. Highly expressed transcripts are involved in several biological processes, including electron transport and exchanger mechanisms, flavonoid biosynthesis, reactive oxygen species (ROS) scavenging, ethylene production, signaling network and protein refolding. The comparisons demonstrated that mRNA-Seq is an efficient method for the analysis of differentially expressed genes and biological processes under salt stress.

Keywords: electron transport, flavonoid biosynthesis, reactive oxygen species, rnaseq

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Authors: Seyedahmad Hosseini, Mohammadjavad Sotoudeheian

Abstract:

Introduction: Allergic asthma is a heterogeneous immuno-inflammatory disease based on Th-2-mediated inflammation. Histopathologic abnormalities of the airways characteristic of asthma include epithelial damage and subepithelial collagen deposition. Objectives: Human bronchial epithelial cell genome expression of TNF‑α, IL‑6, ICAM‑1, VCAM‑1, nuclear factor (NF)‑κB signaling pathways up-regulate during inflammatory cascades. Moreover, immunofluorescence assays confirmed the nuclear translocation of NF‑κB p65 during inflammatory responses. An absolute LDH leakage assays suggestedLPS-inducedcells injury, and the associated mechanisms are co-incident events. LPS-induced phosphorylation of ERKand JNK causes inflammation in epithelial cells through inhibition of ERK and JNK activation and NF-κB signaling pathway. Furthermore, the inhibition of NF-κB mRNA expression and the nuclear translocation of NF-κB lead to anti-inflammatory events. Likewise, activation of SUMF2 which inhibits IL-13 and reduces Th2-cytokines, NF-κB, and IgE levels to ameliorate asthma. On the other hand, TNFα-induced mucus production reduced NF-κB activation through inhibition of the activation status of Rac1 and IκBα phosphorylation. In addition, bradykinin B2 receptor (B2R), which mediates airway remodeling, regulates through NF-κB. Bronchial B2R expression is constitutively elevated in allergic asthma. In addition, certain NF-κB -dependent chemokines function to recruit eosinophils in the airway. Besides, bromodomain containing 4 (BRD4) plays a significant role in mediating innate immune response in human small airway epithelial cells as well as transglutaminase 2 (TG2), which is detectable in saliva. So, the guanine nucleotide-binding regulatory protein α-subunit, Gα16, expresses a κB-driven luciferase reporter. This response was accompanied by phosphorylation of IκBα. Furthermore, expression of Gα16 in saliva markedly enhanced TNF-α-induced κB reporter activity. Methods: The applied method to form NF-κB activation is the electromobility shift assay (EMSA). Also, B2R-BRD4-TG2 complex detection by immunoassay method within saliva with EMSA of NF-κB activation may be a novel biomarker for asthma diagnosis and follow up. Conclusion: This concept introduces NF-κB signaling pathway as potential asthma biomarkers and promising targets for the development of new therapeutic strategies against asthma.

Keywords: NF-κB, asthma, saliva, T-helper

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Authors: Pravina Gurjar, Bothiraja Pour, Vijay Kumbhar, Ganesh Dama

Abstract:

Andrographolide (AG), a bioactive phytoconstituent, has a wider range of pharmacological action. However, due to the intestinal degradation, shows low oral bioavailability. The aim of the present work was to develop Floating In-situ gelling Gastro retentive System (FISGS) for AG in order to enhance its site specific absorption and minimize pH dependent hydrolysis in alkaline environment. Further to increase its therapeutic efficacy for peptic ulcer disease caused by H. pyroli. Gellan based floating in situ gelling system of AG were prepared by using sodium citrate and calcium carbonate. The 32 factorial designs was used to study the effect of gellan and calcium carbonate concentration (independent variables) on dependent variable such as viscosity, floating lag time and drug release. Developed system was evaluated for drug content, floating lag time, viscosity, and drug release studies. Drug content, viscosity, and floating lag time was found to be 81-99%, 67-117 Cps, and 3-5 sec, respectively. The obtained system showed good in vitro floating ability for more than 12 h using 0.1 N HCl as dissolution medium with initial burst release followed by the controlled zero order drug release up to 24 hrs. In vivo testing of FISGS of AG to rats demonstrated significant antiulcer activity that were evaluated by various parameters like pH, volume, total acidity, millimole equivalent of H+ ions/30 min, and protein content of gastric content. The densities of all the formulation batches were found to be near about 0.9 and floating duration above 12 hr. It was observed that with the increase in conc. of gellan there was increase in the viscosity of formulation but all formulations were in optimum range. The drug content of optimized batch was found to be 99.23. In histopathology study of stomach, the villi at the mucosal surface, the intercellular junction, the intestinal lumen were intact; no destruction of the epithelium, and submucosal gland in formulation treated and control group animals as compared to pure drug AG and standard ranitidine. Gellan-based in situ gastro retentive floating system could be advantageous in terms of increased bioavailability of AG to maintain an effective drug conc. in gastric fluid as well as in serum for longer period of time.

Keywords: andrographolide, floating drug delivery, in situ gelling system, gastroretentive system

Procedia PDF Downloads 343