Search results for: extracellular proteins

Authors: Chowon Kim, Yoo-Kyung Kim, Kyeong Yeon Park, Hyun-Shik Lee

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Perfluoroalkyl compounds (PFCs) are globally distributed synthetic compounds that are known to adversely affect human health. Developmental toxicity assessment of PFCs is important to facilitate the evaluation of their environmental impact. In the present study, we assessed the developmental toxicity and teratogenicity of PFCs with different numbers of carbon atoms on Xenopus embryogenesis. An initial frog embryo teratogenicity assay-Xenopus (FETAX) assay was performed that identified perfluorohexanoic (PFHxA) and perfluoroheptanoic (PFHpA) acids as potential teratogens and developmental toxicants. The mechanism underlying this teratogenicity was also investigated by measuring the expression of tissue-specific biomarkers such as phosphotyrosine‑binding protein, xPTB (liver); NKX2.5 (heart); and Cyl18 (intestine). Whole‑mount in situ hybridization, reverse transcriptase‑polymerase chain reaction (RT-PCR), and histologic analyses detected severe defects in the liver and heart following exposure to PFHxA or PFHpA. In addition, immunoblotting revealed that PFHpA significantly increased the phosphorylation of extracellular signal-regulated kinase (ERK) and c-Jun N-terminal kinase (JNK), while PFHxA slightly increased these, as compared with the control. These results suggest that PFHxA and PFHpA are developmental toxicants and teratogens, with PFHpA producing more severe effects on liver and heart development through the induction of ERK and JNK phosphorylation.

Keywords: PFCs, ERK, JNK, xenopus

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Authors: Nirupama Dixit, Anyaa Mittal, Neeru Sood

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Alzheimer’s disease (AD) is a progressive form of dementia, contributing to up to 70% of cases, mostly observed in elderly but is not restricted to old age. The pathophysiology of the disease is characterized by specific pathological changes in brain. The changes (i.e. accumulation of metal ions in brain, formation of extracellular β-amyloid (Aβ) peptide aggregates and tangle of hyper phosphorylated Tau protein inside neurons) damage the neuronal connections irreversibly. The current issues in improvement of life quality of Alzheimer's patient lies in the fact that the diagnosis is made at a late stage of the disease and the medications do not treat the basic causes of Alzheimer's. The targeted delivery of drug through the blood brain barrier (BBB) poses several limitations via traditional approaches for treatment. To overcome these drug delivery limitation, nanoparticles provide a promising solution. This review focuses on current strategies for efficient targeted drug delivery using nanoparticles and improving the quality of therapy provided to the patient. Nanoparticles can be used to encapsulate drug (which is generally hydrophobic) to ensure its passage to brain; they can be conjugated to metal ion chelators to reduce the metal load in neural tissue thus lowering the harmful effects of oxidative damage; can be conjugated with drug and monoclonal antibodies against BBB endogenous receptors. Finally this review covers how the nanoparticles can play a role in diagnosing the disease.

Keywords: Alzheimer's disease, β-amyloid plaques, blood brain barrier, metal chelators, nanoparticles

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Authors: Fatima Bouazza, Khadija Khedid, Lamiae Amallah, Aziz Mouhaddach, Basma Boukour, Jihane Ennadir, Rachida Hassikou

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In order to combine milk and its derived products conservation and flavoring, Moroccans often used aromatic and medicinal plants. These plant extracts are endowed with several nutritive and therapeutic properties. This study constitutes a first national assessment of physico-chemical quality of sheep’s milk from moroccan Sardi breed and the evaluation of the antibacterial effect of three medicinal plants extracts: Aloe barbadensis Miller, Thymus satureioides and Mentha pulegium on flora isolated from this sheep's milk. 100 milk samples were collected in four regions of Morocco. The bacteria isolated were identified by classical and molecular methods (16S rRNA sequencing) and tested, according to the disk method, for their sensitivity to several antibiotics. The physico-chemical analyzes of sheep’s milk concerned the pH, titratable acidity, density, dry extract, freezing point and contents of: fat, proteins, lactose and calcium. The essential oils (EOs) of T. satureioides and M .pulegium were extracted by hydrodistillation and analyzed by GC / MS, while the Aloe vera leaf pulp was analyzed by the methods of Harborne and HPLC. A total number of 125 bacteria have been identified. Significant resistance to chemical antibiotics has been noted in LABs. The average temperature value of milk is around 57.15 °C, the pH is 6.56, the titratable acidity is around 3.4 ° D, the density is 1.035g / cm³ , the total dry extract is around 169.5g / l, the ash (9.8g / l), the freezing point (- 0.556 °C) while the average fat content is 67.85g / l . The samples richest in fat belong to the region of Settat, cradle of the Sardi breed, with a maximum average value of 74.4g / l. The average protein is 56g / l, lactose (39.92g / l), and calcium (1.855g / l). Analysis of the major components of EOs revealed the dominance of borneol in the case of T. satureioides and of pulegone in M. pulegium. Aloe vera gel contains alkaloids, flavonoids, catechic tannins, saponins and 1.60 µg / ml of aloin. The plant extracts have a bactericidal effect on E. coli, Klebsiellaoxytoca and Staphylococci and bacteriostatic effect on LABs of technological interest (Lactobacillus). As a result of this study, it is believed that the consumption of sardi sheep’s milk would be of nutritional benefit. Its richness in fat and proteins predisposes it for biotechnological development in the manufacture of cheese and yogurt. Also, the use of aromatic and medicinal plants, as natural additives would be of great benefit to flavor and maintain its quality.

Keywords: sheep’s milk, lactic flora, antimicrobial power, aloe barbadensis miller, thymus satureioides, mentha pulegium

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Authors: José Maria, Vânia Laranjo, Luís Abrunhosa, António Inês

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The occurrence of mycotoxigenic moulds such as Aspergillus, Penicillium and Fusarium in food and feed has an important impact on public health, by the appearance of acute and chronic mycotoxicoses in humans and animals, which is more severe in the developing countries due to lack of food security, poverty and malnutrition. This mould contamination also constitutes a major economic problem due the lost of crop production. A great variety of filamentous fungi is able to produce highly toxic secondary metabolites known as mycotoxins. Most of the mycotoxins are carcinogenic, mutagenic, neurotoxic and immunosuppressive, being ochratoxin A (OTA) one of the most important. OTA is toxic to animals and humans, mainly due to its nephrotoxic properties. Several approaches have been developed for decontamination of mycotoxins in foods, such as, prevention of contamination, biodegradation of mycotoxins-containing food and feed with microorganisms or enzymes and inhibition or absorption of mycotoxin content of consumed food into the digestive tract. Some group of Gram-positive bacteria named lactic acid bacteria (LAB) are able to release some molecules that can influence the mould growth, improving the shelf life of many fermented products and reducing health risks due to exposure to mycotoxins. Some LAB are capable of mycotoxin detoxification. Recently our group was the first to describe the ability of LAB strains to biodegrade OTA, more specifically, Pediococcus parvulus strains isolated from Douro wines. The pathway of this biodegradation was identified previously in other microorganisms. OTA can be degraded through the hydrolysis of the amide bond that links the L-β-phenylalanine molecule to the ochratoxin alpha (OTα) a non toxic compound. It is known that some peptidases from different origins can mediate the hydrolysis reaction like, carboxypeptidase A an enzyme from the bovine pancreas, a commercial lipase and several commercial proteases. So, we wanted to have a better understanding of this OTA degradation process when LAB are involved and identify which molecules where present in this process. For achieving our aim we used some bioinformatics tools (BLAST, CLUSTALX2, CLC Sequence Viewer 7, Finch TV). We also designed specific primers and realized gene specific PCR. The template DNA used came from LAB strains samples of our previous work, and other DNA LAB strains isolated from elderberry fruit, silage, milk and sausages. Through the employment of bioinformatics tools it was possible to identify several proteins belonging to the carboxypeptidase family that participate in the process of OTA degradation, such as serine type D-Ala-D-Ala carboxypeptidase and membrane carboxypeptidase. In conclusions, this work has identified carboxypeptidase proteins being one of the molecules present in the OTA degradation process when LAB are involved.

Keywords: carboxypeptidase, lactic acid bacteria, mycotoxins, ochratoxin a.

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Authors: Aleksandra Twarda, Dobrosława Krzemień, Grzegorz Dubin, Tad A. Holak

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Stabilin-2 belongs to the group of scavenger receptors and plays a crucial role in clearance of more than 10 ligands from the bloodstream, including hyaluronic acid, products of degradation of extracellular matrix and metabolic products. The Link domain, a defining feature of stabilin-2, has a sequence similar to Link domains in other hyaluronic acid receptors, such as CD44 or TSG-6, and is responsible for most of ligands binding. Present knowledge of signal transduction by stabilin-2, as well as ligands’ recognition and binding mechanism, is limited. Until now, no experimental structures have been solved for any segments of stabilin-2. It has recently been demonstrated that the stabilin-2 knock-out or blocking of the receptor by an antibody effectively opposes cancer metastasis by elevating the level of circulating hyaluronic acid. Moreover, loss of expression of stabilin-2 in a peri-tumourous liver correlates with increased survival. Solving of the crystal structure of stabilin-2 and elucidation of the binding mechanism of hyaluronic acid could enable the precise characterization of the interactions in the binding site. These results may allow for designing specific small-molecule inhibitors of stabilin-2 that could be used in cancer therapy. To carry out screening for crystallization of stabilin-2, we cloned constructs of the Link domain of various lengths with or without surrounding domains. The folding properties of the constructs were checked by nuclear magnetic resonance (NMR). It is planned to show the binding of hyaluronic acid to the Link domain using several biochemical methods, i.a. NMR, isothermal titration calorimetry and fluorescence polarization assay.

Keywords: stabilin-2, Link domain, X-ray crystallography, NMR, hyaluronic acid, cancer

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Authors: Isabella Jabbour

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The world's population is expected to reach over 10 billion by 2050, creating a significant demand for food production, particularly in the agricultural industry. Cellular agriculture presents a solution to this challenge by producing meat that resembles traditionally produced meat, but with significantly less land use. Decellularized plant scaffolds, such as spinach leaves, have been shown to be a suitable edible scaffold for growing animal muscle, enabling cultured cells to grow and organize into three-dimensional structures that mimic the texture and flavor of conventionally produced meat. However, the use of freshwater to remove the intact extracellular material from these plants remains a concern, particularly when considering scaling up the production process. In this study, two protocols were used, 1X SDS and Boom Sauce, to decellularize spinach leaves with both distilled water and ocean water. The decellularization process was confirmed by histology, which showed an absence of cell nuclei, DNA and protein quantification. Results showed that spinach decellularized with ocean water contained 9.9 ± 1.4 ng DNA/mg tissue, which is comparable to the 9.2 ± 1.1 ng DNA/mg tissue obtained with DI water. These findings suggest that decellularized spinach leaves using ocean water hold promise as an eco-friendly and cost-effective scaffold for laboratory-grown meat production, which could ultimately transform the meat industry by providing a sustainable alternative to traditional animal farming practices while reducing freshwater use.

Keywords: cellular agriculture, plant scaffold, decellularization, ocean water usage

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Authors: Lingling Shui, Shuting Xie

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As alternatives to electronic devices, optically active structures from responsive nanomaterials offer great opportunity buildup smart functional sensors. Hereby, we report on droplet microfluidics enabled construction and application of photonic microcapsules (PMCs) for colorimetric temperature microsensors, enabling miniaturization for injectable local micro-area sensing and integration for large-area sensing. Monodispersed PMCs are produced by in-situ photopolymerization of hydrogel shells of cholesteric liquid crystal (CLC)-in-water-in-oil double emulsion droplets prepared using microfluidic devices, with controllable physical structures and chemical compositions. Constructed PMCs exhibit thermal responsive structural color according to the selective Bragg reflection of CLC’s periodical helical structures within the microdroplet’s spherical confinement. Constructed PMCs with tunable size and composition have been successfully applied for monitoring the living cell extracellular temperature via co-incubation with cell suspension, and for detecting human body temperature via a flexible device from assembled PMCs. These PMCs could be flexibly applied in either micro-environment or large-area surface, enabling wide applications for precision temperature monitoring biological activities (e.g. cells or organs), optoelectronic devices working conditions (e.g. temperature indicators under extreme conditions), and etc.

Keywords: droplet, microfluidics, assembly, soft materials, microsensor

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Authors: Sergejs Kolesovs, Armands Vigants

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The use of dairy industry by-products and wastewater as a cheap substrate for microalgal growth is gaining recognition. However, the mechanisms of lactose utilization remain understudied, limiting the potential of successful microalgal biomass production using various dairy by-products, such as whey and permeate. The necessity for microalgae to produce a specific enzyme, β-galactosidase, requires the selection of suitable strains. This study focuses on a freshwater microalgal isolate's ability to grow on a semi-synthetic medium supplemented with lactose. After 10 days of agitated cultivation, an axenic microalgal isolate achieved significantly higher biomass production under mixotrophic growth conditions (0.86 ± 0.07 g/L, dry weight) than heterotrophic growth (0.46 ± 0.04 g/L). Moreover, mixotrophic cultivation had significantly higher biomass production compared to photoautotrophic growth (0.67 ± 0.05 g/L). The activity of β-galactosidase was detected in both supernatant and microalgal biomass under mixotrophic and heterotrophic growth conditions, showing the potential of extracellular and intracellular mechanisms of enzyme production. However, the main limiting factor in this study was the increase of pH values during the cultivation, significantly reducing the activity of the β-galactosidase enzyme after 3rd day of cultivation. It highlights the need for stricter control of growth parameters to ensure the enzyme's activity. Further research will assess the isolate's suitability for dairy by-product bioconversion and biomass composition.

Keywords: microalgae, lactose, whey, permeate, beta-galactosidase, mixotrophy, heterotrophy

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Authors: S. P. Gomes, D. C. Goncalves, E. Ribeiro, M. C. L. Seelaender

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Skin is susceptible to photo damage induced by exposure to sunlight, or ultraviolet (UV) radiation, which induces breakdown of extracellular matrix, DNA degradation, skin cell lesion and apoptosis, and development of cancer. Phytonutrients demonstrate protective effects against UV damage. The purpose of this study was evaluating the effect of an antioxidant juice (AJ) contaning Brazilian natural products upon skin damage. The juice was produced by Metabolics®. Male Wistar rats were divided in 4 groups: Animals receiving the antioxidant juice (AJ): orange, carrot, honey, tomato extract, avocado, ginger and camu-camu (Brazilian fruit, a major source of vitamin C) ad libitum for 21 days; or water (C), subdivided in groups exposed or not to UV radiation for 2 non consecutive days, during five hours each day, after 15 days of juice supplementation. On the 22nd day, rats were killed by decapitation and epithelium samples from the dorsal skin removed, fixed in bouin and embedded in paraffin. The sections were stained with hematoxylin and eosin or mallory and picrosirius red. Isolated DNA was submitted to electrophoresis (1.8% agarose gel, 0.5% ethidium bromide). UV radiation significantly induced sunburn of superficial epithelial cells of C, AJ treatment reduced this effect. Collagen changes were observed in UV groups, yet AJ treatment prevented collagen degradation. UV radiation induced significant DNA degradation, in C, which was prevented by AJ treatment. The antioxidant juice consumed chronically protected against acute skin damage.

Keywords: nutraceuticals, antioxidants, photoprotection, uv radiation

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Authors: Guo Wenyu, Qu Youli

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A practical and simple self-indexing data structure, Partitioned Elias-Fano (PEF) - Compressed Suffix Arrays (CSA), is built in linear time for the CSA based on PEF indexes. Moreover, the PEF-CSA is compared with two classical compressed indexing methods, Ferragina and Manzini implementation (FMI) and Sad-CSA on different type and size files in Pizza & Chili. The PEF-CSA performs better on the existing data in terms of the compression ratio, count, and locates time except for the evenly distributed data such as proteins data. The observations of the experiments are that the distribution of the φ is more important than the alphabet size on the compression ratio. Unevenly distributed data φ makes better compression effect, and the larger the size of the hit counts, the longer the count and locate time.

Keywords: compressed suffix array, self-indexing, partitioned Elias-Fano, PEF-CSA

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Authors: Ramesh Subramani, Mathivanan Narayanasamy, William Aalbersberg

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It is well accepted by last 35 years of research and on-going programmes that marine environment harbours abundant and unique biodiversity, which is currently playing as an important source in bioprospecting. It has become apparent that marine microorganisms are lead in the biodiscovery. Among marine organisms, actinobacteria are a target phylum for discovering novel antibiotics against increasing the multi-drug resistant human pathogens because of these taxa representing for novel genera and species. Marine actinomycetes are a proven source of new antibiotic leads and novel enzymes with important industrial applications. A total of 183 streptomycete and 25 non-streptomycete strains were isolated from different marine samples collected from north-eastern part of the Indian Ocean. Among them, 111 isolates displayed antibacterial activity against human pathogens and 151 exhibited antifungal activity against phytopathogens. Importantly, most of them produced various extracellular enzymes and 58 of them produced exopolysaccharides. Totally eight small bioactive compounds and a thermostable alkaline protease have been purified from a selected strain, Streptomyces fungicidicus. Besides, our on-going studies on non-streptomycete strains (rare actinomycetes) are most likely promising resource for new and unique compounds against current emerging drug-resistant pathogens. We have just recognised the chemical diversity in marine microorganisms. Therefore it is worthwhile to continue the exploration of marine microorganisms for new drug leads, novel enzymes and other bioprospecting research.

Keywords: bioactive compounds, industrial enzymes, marine actinobacteria, microbial metabolites, marine natural products

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Authors: Walter Vera-Ortega, Idoia Busnadiego, Sam J. Wilson

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Introduction: Human Immunodeficiency Virus type 1 (HIV-1) is responsible for the acquired immunodeficiency syndrome (AIDS), a leading cause of death worldwide infecting millions of people each year. Despite intensive research in vaccine development, therapies against HIV-1 infection are not curative, and the huge genetic variability of HIV-1 challenges to drug development. Current animal models for HIV-1 research present important limitations, impairing the progress of in vivo approaches. Macaques require a CD8+ depletion to progress to AIDS, and the maintenance cost is high. Mice are a cheaper alternative but need to be 'humanized,' and breeding is not possible. The development of an HIV-1 clone able to replicate in mice is a challenging proposal. The lack of human co-factors in mice impedes the function of the HIV-1 accessory proteins, Tat and Rev, hampering HIV-1 replication. However, Tat and Rev function can be replaced by constitutive/chimeric promoters, codon-optimized proteins and the constitutive transport element (CTE), generating a novel HIV-1 clone able to replicate in mice without disrupting the amino acid sequence of the virus. By minimally manipulating the genomic 'identity' of the virus, we propose the generation of an HIV-1 clone able to replicate in mice to assist in antiviral drug development. Methods: i) Plasmid construction: The chimeric promoters and CTE copies were cloned by PCR using lentiviral vectors as templates (pCGSW and pSIV-MPCG). Tat mutants were generated from replication competent HIV-1 plasmids (NHG and NL4-3). ii) Infectivity assays: Retroviral vectors were generated by transfection of human 293T cells and murine NIH 3T3 cells. Virus titre was determined by flow cytometry measuring GFP expression. Human B-cells (AA-2) and Hela cells (TZMbl) were used for infectivity assays. iii) Protein analysis: Tat protein expression was determined by TZMbl assay and HIV-1 capsid by western blot. Results: We have determined that NIH 3T3 cells are able to generate HIV-1 particles. However, they are not infectious, and further analysis needs to be performed. Codon-optimized HIV-1 constructs are efficiently made in 293T cells in a Tat and Rev independent manner and capable of packaging a competent genome in trans. CSGW is capable of generating infectious particles in the absence of Tat and Rev in human cells when 4 copies of the CTE are placed preceding the 3’LTR. HIV-1 Tat mutant clones encoding different promoters are functional during the first cycle of replication when Tat is added in trans. Conclusion: Our findings suggest that the development of an HIV-1 Tat-Rev independent clone is challenging but achievable aim. However, further investigations need to be developed prior presenting our HIV-1 clone as a candidate model for research.

Keywords: codon-optimized, constitutive transport element, HIV-1, long terminal repeats, research model

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Authors: Fabienne Vozy, Anick Lepage

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Insects are displaying high nutritional value, low greenhouse gas emissions, low land use requirements and high food conversion efficiency. They can contribute to the food chain and be one of many solutions to protein shortages. Currently, in North America, nutritional entomology is under-developed and the needs to better understand its benefits remain to convince large-scale producers and consumers (both for human and agricultural needs). As such, large-scale production of mealworms offers a promising alternative to replacing traditional sources of protein and fatty acids. To proceed orderly, it is required to collect more data on the nutritional values of insects such as, a) Evaluate the diets of insects to improve their dietary value; b) Test the breeding conditions to optimize yields; c) Evaluate the use of by-products and organic residues as sources of food. Among the featured technical parameters, relative humidity (RH) percentage and temperature, optimal substrates and hydration sources are critical elements, thus establishing potential benchmarks for to optimize conversion rates of protein and fatty acids. This research is to establish the combination of the most influential rearing parameters with local food residues, to correlate the findings with the nutritional value of the larvae harvested. 125 same-monthly old adults/replica are randomly selected in the mealworm breeding pool then placed to oviposit in growth chambers preset at 26°C and 65% RH. Adults are removed after 7 days. Larvae are harvested upon the apparition of the first nymphosis signs and batches, are analyzed for their nutritional values using wet chemistry analysis. The first samples analyses include total weight of both fresh and dried larvae, residual humidity, crude proteins (CP%), and crude fats (CF%). Further analyses are scheduled to include soluble proteins and fatty acids. Although they are consistent with previous published data, the preliminary results show no significant differences between treatments for any type of analysis. Nutritional properties of each substrate combination have yet allowed to discriminate the most effective residue recipe. Technical issues such as the particles’ size of the various substrate combinations and larvae screen compatibility are to be investigated since it induced a variable percentage of lost larvae upon harvesting. To address those methodological issues are key to develop a standardized efficient procedure. The aim is to provide producers with easily reproducible conditions, without incurring additional excessive expenditure on their part in terms of equipment and workforce.

Keywords: entomophagy, nutritional value, rearing parameters optimization, Tenebrio molitor

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Authors: Leonard Lipovich, Pattaraporn Thepsuwan, Anton-Scott Goustin, Juan Cai, Donghong Ju, James B. Brown

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Two-thirds of human genes do not encode any known proteins. Aside from long non-coding RNA (lncRNA) genes with recently-discovered functions, the ~40,000 non-protein-coding human genes remain poorly understood, and a role for their transcripts as de-facto unconventional messenger RNAs has not been formally excluded. Ribosome profiling (Riboseq) predicts translational potential, but without independent evidence of proteins from lncRNA open reading frames (ORFs), ribosome binding of lncRNAs does not prove translation. Previously, we mass-spectrometrically documented translation of specific lncRNAs in human K562 and GM12878 cells. We now examined lncRNA translation in human MCF7 cells, integrating strand-specific Illumina RNAseq, Riboseq, and deep mass spectrometry in biological quadruplicates performed at two core facilities (BGI, China; City of Hope, USA). We excluded known-protein matches. UCSC Genome Browser-assisted manual annotation of imperfect (tryptic-digest-peptides)-to-(lncRNA-three-frame-translations) alignments revealed three peptides hypothetically explicable by 'stop-to-nonstop' in-frame replacement of stop codons by amino acids in two ORFs of the lncRNA MMP24-AS1. To search for this phenomenon genomewide, we designed and implemented a novel pipeline, matching tryptic-digest spectra to wildcard-instead-of-stop versions of repeat-masked, six-frame, whole-genome translations. Along with singleton putative stop-to-nonstop events affecting four other lncRNAs, we identified 24 additional peptides with stop-to-nonstop in-frame substitutions from multiple positive-strand MMP24-AS1 ORFs. Only UAG and UGA, never UAA, stop codons were impacted. All MMP24-AS1-matching spectra met the same significance thresholds as high-confidence known-protein signatures. Targeted resequencing of MMP24-AS1 genomic DNA and cDNA from the same samples did not reveal any mutations, polymorphisms, or sequencing-detectable RNA editing. This unprecedented apparent gene-specific violation of the genetic code highlights the importance of matching peptides to whole-genome, not known-genes-only, ORFs in mass-spectrometry workflows, and suggests a new mechanism enhancing the combinatorial complexity of the proteome. Funding: NIH Director’s New Innovator Award 1DP2-CA196375 to LL.

Keywords: genetic code, lncRNA, long non-coding RNA, mass spectrometry, proteogenomics, ribo-seq, ribosome, RNAseq

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Authors: Sofia Papadimitriou

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INTRODUCTION: The differential diagnosis between acute viral and bacterial infections is an important cost-effectiveness parameter at the stage of the treatment process in order to achieve the maximum benefits in therapeutic intervention by combining the minimum cost to ensure the proper use of antibiotics.The discovery of sensitive and robust molecular diagnostic tests in response to the role of the host in infections has enhanced the accurate diagnosis and differentiation of infections. METHOD: The study used a sample of six independent blood samples (total=756) which are associated with human proteins-proteins, each of which at the transcription stage expresses a different response in the host network between viral and bacterial infections.Τhe individual blood samples are subjected to a sequence of computer filters that identify a gene panel corresponding to an autonomous diagnostic score. The data set and the correspondence of the gene panel to the diagnostic patents a new Bangalore -Viral Bacterial (BL-VB). FINDING: We use a biomarker based on the blood of 10 genes(Panel-VB) that are an important prognostic value for the detection of viruses from bacterial infections with a weighted average AUROC of 0.97(95% CL:0.96-0.99) in eleven independent samples (sets n=898). We discovered a base with a patient score (VB 10 ) according to the table, which is a significant diagnostic value with a weighted average of AUROC 0.94(95% CL: 0.91-0.98) in 2996 patient samples from 56 public sets of data from 19 different countries. We also studied VB 10 in a new cohort of South India (BL-VB,n=56) and found 97% accuracy in confirmed cases of viral and bacterial infections. We found that VB 10 (a)accurately identifies the type of infection even in unspecified cases negative to the culture (b) shows its clinical condition recovery and (c) applies to all age groups, covering a wide range of acute bacterial and viral infectious, including non-specific pathogens. We applied our VB 10 rating to publicly available COVID 19 data and found that our rating diagnosed viral infection in patient samples. RESULTS: Τhe results of the study showed the diagnostic power of the biomarker VB 10 as a diagnostic test for the accurate diagnosis of acute infections in recovery conditions. We look forward to helping you make clinical decisions about prescribing antibiotics and integrating them into your policies management of antibiotic stewardship efforts. CONCLUSIONS: Overall, we are developing a new property of the RNA-based biomarker and a new blood test to differentiate between viral and bacterial infections to assist a physician in designing the optimal treatment regimen to contribute to the proper use of antibiotics and reduce the burden on antimicrobial resistance, AMR.

Keywords: acute infections, antimicrobial resistance, biomarker, blood transcriptome, systems biology, classifier diagnostic score

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Authors: Agnė Mikalauskaitė, Renata Karpicz, Vitalijus Karabanovas, Arūnas Jagminas

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The use of biocompatible precursors for the synthesis and stabilization of fluorescent gold nanoclusters (NCs) with strong red photoluminescence creates an important link between natural sciences and nanotechnology. Herein, we report the cost-effective synthesis of Au nanoclusters by templating and reduction of chloroauric acid with the cheap amino acid food supplements. This synthesis under the optimized conditions leads to the formation of biocompatible Au NCs having good stability and intense red photoluminescence, peaked at 680 to 705 nm, with a quantum yield (QY) of ≈7% and the average lifetime of up to several µs. The composition and luminescent properties of the obtained NCs were compared with ones formed via well-known bovine serum albumin reduction approach. Our findings implied that synthesized Au NCs tend to accumulate in more tumorigenic breast cancer cells (line MDA-MB-213) and after dialysis can be prospective for bio imagining.

Keywords: gold nanoclusters, proteins, materials chemistry, red-photoluminescence, bioimaging

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Authors: Yu Chen, Weiwei Ren, Xiaojing Mu, Feng Zhang, Yi Xu

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The superhydrophobic surface is widely used to reduce friction for the flow inside micro-channel and can be used to control/manipulate fluid, cells and even proteins in lab-on-chip. Fabricating micro grooves on hydrophobic surfaces is a common method to obtain such superhydrophobic surface. This study utilized the numerical method to investigate the effect of eccentric micro-grooves on the friction of flow inside micro-channel. A detailed parametric study was conducted to reveal how the eccentricity of micro-grooves affects the micro-channel flow under different grooves sizes, channel heights, Reynolds number. The results showed that the superhydrophobic surface with eccentric micro-grooves induces less friction than the counter part with aligning micro-grooves, which means requiring less power for pumps.

Keywords: eccentricity, micro-channel, micro-grooves, superhydrophobic surface

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Authors: Sachin Latiyan, T. S. Sampath Kumar, Mukesh Doble

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Natural polymer based nanofibrous wound dressings have gained increased attention because of their high surface area, bioactivity, biodegradability and resemblance to extracellular matrix. Agarose (a natural polymer) have been used largely for angiogenesis, cartilage formation and wound healing applications. However, electrospinning of agarose is tedious thereby rendering limited studies on fabrication and evaluation of agarose based nanofibrous wound dressings. Thus, present study focuses on the fabrication of agarose (10% w/v)/ polyvinyl alcohol (12% w/v) based multifunctional nanofibrous scaffolds. Zinc citrate (1, 3 and 5% w/w of the polymer) was added as a potential antibacterial agent to combat wound infections. The fabricated scaffolds exhibit ~500% swelling (in phosphate buffer saline) with enhanced mechanical strength which is suitable for most of the wound healing applications. In vitro studies were found to reveal an increased migration and proliferation of L929 mouse fibroblasts with agarose blends w.r.t to the control. The fabricated dressings were found to be effective against both Escherichia coli (Gram-negative) and Staphylococcus aureus (Gram-positive) bacterial strains. Hence, a multifunctional (as provides effective swelling and mechanical support along with antibacterial property), natural product based, eco-friendly scaffold was successfully fabricated to serve as a potential wound dressing material.

Keywords: antibacterial dressings, benign solvent, nanofibrous agarose, biocompatibility, enhanced swelling and mechanical strength, biopolymeric dressings

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Authors: M. Caruffo, P. Navarrete, C. G. Feijoo, L. Sáenz

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Disease prevention through the use of vaccines has been crucial to achieve the current level of production in the salmon industry. However, vaccines have been developed based largely on inactivated bacterial formulations, using the whole pathogen. These formulations have demonstrated excellent efficacy against extracellular bacterial pathogens. However diseases with the greatest economic impacts correspond to intracellular bacterial and viral pathogens, vaccines based on these types of agents have shown a discrete effectiveness. It is for these reasons that the development of subunit vaccines based on defined antigens offers a promising solution. The main problem is that subunit vaccines offer a low immunogenicity, since they lack immunostimulatory elements, so that the development of new adjuvants platforms becomes an important challenge for this type of formulations. We evaluate the effect of a formulation based on proteoliposomes of Vibrio anguillarum administered by immersion as a new adjuvant strategy, allowing efficient stimulation of the innate immune system. Proteoliposomes physicochemical properties were evaluated in its ability to produce an inflammatory process. Using zebrafish (Danio rerio) larvae as a model species and the transgenic line (Tg(mpx: GFP)i114) allowed us to track the neutrophil migration in real time. Additionally we evaluated the gene expression of some molecular markers involved in the development of the innate immune response characterizing the adjuvant capacity of the formulation.

Keywords: adjuvants, vaccine development, zebrafish, innate immunity

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Authors: Sudhir Kumar Sharma, Abiy D. Woldetsadik, Mazin Magzoub, Ramesh Jagannathan

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It is well known that bioactive ceramics exhibit specific biological affinities, especially in the area of tissue re-generation. In this context, we report the development of an eminently scalable, novel, supercritical CO₂ based process for the fabrication of hierarchically porous 'soft' CaCO₃ scaffolds. Porosity at the macro, micro, and nanoscales was obtained through process optimization of the so-called 'coffee-ring effect'. Exposure of these CaCO₃ scaffolds to monocytic THP-1 cells yielded negligible levels of tumor necrosis factor-alpha (TNF-α) thereby confirming the lack of immunogenicity of the scaffolds. ECM protein treatment of the scaffolds showed enhanced adsorption comparable to standard control such as glass. In vitro studies using osteoblast precursor cell line, MC3T3, also demonstrated the cytocompatibility of hierarchical porous CaCO₃ scaffolds.

Keywords: supercritical CO2, CaCO3 scaffolds, coffee-ring effect, ECM proteins

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Authors: M. Yusuf

Abstract:

The recent discovery of the phenomenon of RNA interference has led to its application in various aspects of plant improvement. Crops can be modified by engineering novel RNA interference pathways that create small RNA molecules to alter gene expression in crops or plant pests. RNA interference can generate new crop quality traits or provide protection against insects, nematodes and pathogens without introducing new proteins into food and feed products. This is an advantage in contrast with conventional procedures of gene transfer. RNA interference has been used to develop crop varieties resistant to diseases, pathogens and insects. Male sterility has been engineered in plants using RNA interference. Better quality crops have been developed through the application of RNA interference etc. The objective of this paper is to highlight the application of RNA interference in crop improvement and to project its potential future use to solve problems of agricultural production in relation to plant breeding.

Keywords: RNA interference, application, crop Improvement, agricultural production

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Authors: Fatima Zohra Bissaad, Farid Bounaceur, Nassima Behidj, Nadjiba Chebouti, Fatma Halouane, Bahia Doumandji-Mitiche

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Effect of Beauveria bassiana and Metarhizium anisopliae var. acridum on the 5^th instar nymphs of Schistocerca gregaria was studied in the laboratory. Infection by these both entomopathogenic fungi caused reduction in the hemolymph total protein. The average amounts of total proteins were 2.3, 2.07, 2.09 µg/100 ml of haemolymph in the control and M. anisopliae var. acridum, and B. bassiana based-treatments, respectively. Three types of haemocytes were recognized and identified as prohaemocytes, plasmatocytes and granulocytes. The treatment caused significant reduction in the total haemocyte count and in each haemocyte type on the 9^th day after its application.

Keywords: Beauveria bassiana, haemolymph picture, haemolymph protein, Metarhizium anisopliae var. acridum, Schistocerca gregaria

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Authors: Hu-Jiang Shi, Li-Juan Zhu

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The implication of 5-hydroxytryptamine 2C receptor (5-HT2CR) in affective behaviors is a topic of debate, and the underlying mechanisms remain largely unclear. Here, we elucidate that the interaction between hippocampal neuronal nitric oxide synthase (nNOS) and carboxy-terminal PDZ ligand of nNOS (CAPON) contributes to the disruption of hippocampal excitation-inhibition (E/I) balance, which is responsible for the anxiety- and depressive-like behaviors caused by chronic stress-related 5-HT2CR signaling deficiency. In detail, activation or inhibition of 5-HT2CR by CP809101 or SB242084 modulates nNOS-CAPON interaction by influencing intracellular Ca²⁺ release. Notably, the dissociation of nNOS-CAPON abolishes SB242084-induced anxiety- and depressive-like behaviors, as well as the reduction in extracellular signal-regulated kinase (ERK)/cAMP-response element binding protein (CREB)/synapsin signaling and SNARE complex assembly. Furthermore, nNOS-CAPON blockers restore the impairments caused by SB242084, including the reduction in SNARE assembly-mediated γ-aminobutyric acid (GABA) vesicle release and a consequent shift of the E/I balance toward excitation in CA3 pyramidal neurons. Conclusively, our findings disclose the regulatory role of 5-HT2CR in anxiety- and depressive-like behaviors and highlight the hippocampal nNOS-CAPON coupling-triggered E/I imbalance as a pivotal cellular event underpinning the behavioral consequences of 5-HT2CR inhibition.

Keywords: 5-HT2CR, anxiety, depression, nNOS-CAPON coupling, excitation-inhibition balance, neurotransmitter release

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Authors: Amna Abdalla Mohammed Khalid, Pietro Parisse, Silvia Onesti, Loredana Casalis

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Basic cellular processes as DNA replication are crucial to cell life. Understanding at the molecular level the mechanisms that govern DNA replication in proliferating cells is fundamental to understand disease connected to genomic instabilities, as a genetic disease and cancer. A key step for DNA replication to take place, is unwinding the DNA double helix and this carried out by proteins called helicases. The archaeal MCM (minichromosome maintenance) complex from Methanothermobacter thermautotrophicus have being studied using Atomic Force Microscopy (AFM), imaging in air and liquid (Physiological environment). The accurate analysis of AFM topographic images allowed to understand the static conformations as well the interaction dynamic of MCM and DNA double helix in the present of ATP.

Keywords: DNA, protein-DNA interaction, MCM (mini chromosome manteinance) complex, atomic force microscopy (AFM)

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Authors: M. Chňapek, M. Tomka, R. Peroutková, Z. Gálová

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Processes of plant breeding, testing and licensing of new varieties, patent protection in seed production, relations in trade and protection of copyright are dependent on identification, differentiation and characterization of plant genotypes. Therefore, we focused our research on utilization of wheat storage proteins as genetic markers suitable not only for differentiation of individual genotypes, but also for identification and characterization of their considerable properties. We analyzed a collection of 102 genotypes of bread wheat (Triticum aestivum L.), 41 genotypes of spelt wheat (Triticum spelta L.), and 35 genotypes of durum wheat (Triticum durum Desf.), in this study. Our results show, that genotypes of bread wheat and durum wheat were homogenous and single line, but spelt wheat genotypes were heterogenous. We observed variability of HMW-GS composition according to environmental factors and level of breeding and predict technological quality on the basis of Glu-score calculation.

Keywords: genotype identification, HMW-GS, wheat quality, polymorphism

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Authors: Abdurzag Kerban, Mostfa M. Abou-Ahmed, Abdelrof M. Ghallab, Mona H. Shaker

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Preservation of semen had a major impact on sheep genetic breeding. The aim of this study was to evaluate the effect of protected fat, probiotic and zinc-enriched diets on semen freezability. Twenty two Barki rams were randomly assigned into four groups; Group I (n=5) was fed the basal diet enriched with 3.7% of dry fat/kg concentration/day, Group II (n=5) was fed a basal diet-enriched with 10gm of probiotic /head/day, Group III (n=6) was fed on the basal diet enriched with 100 ppm of 10% zinc chelated with methionine/kg dry matter/day and Group IV (n=6) was served as control. A pool of three to four ejaculates were pooled from rams within a period of ten weeks. Semen was diluted in egg yolk-Tris diluent and processed in 0.25 ml straw. Motility was evaluated after dilution, before freezing and post-thawing at 0, 1, 2 and 3 hour incubation. Viability index, acrosome integrity and leakage of intracellular enzymes (Aspartat aminotransferase and Alkline phosphatase) were also evaluated. Spermatozoa exhibited highly significant (P<0.01) percentages of motility at 0, 1, 2, and 3 hours incubation after thawing, viability index and acrosome integrity in rams fed a diet enriched with protected fat and zinc groups as compared with probiotic and control groups. Also, the mean value of extracellular leakage of AST was significantly lower in fat and zinc group as compared with probiotic and control groups. In conclusion, semen freezability was improved in animals fed a diet fortified with fat and zinc with no significant improvement in animals fed the probiotic-enriched diet.

Keywords: Barki ram semen, freezing, straw, feed additives

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Authors: N. Bouzouita, A. Snoussi , H. Ben Haj Koubaier, I. Essaidi, M. M. Chaabouni, S. Zgoulli, P. Thonart

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Color is one of the most important quality attributes for the food industry. Grape marc, a complex lignocellulosic material is one of the most abundant and worth less byproduct, generated after the pressing process. The development of the process of purification by micro filtration, ultra filtration, nano filtration and drying by atomization of the anthocyanins of Tunisian origin is the aim of this work. Locust bean gum is the ground endosperm of the seeds of carob fruit; owing to its remarkable water-binding properties, it is widely used to improve the texture of food and largely employed in food industry. The purification of LGB causes drastically reduced ash and proteins contents but important increase for galactomannan.

Keywords: Carob, food additives, grape pomace, locust bean gum, natural colorant, nano filtration, thickener, ultra filtration

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Authors: K. Rasool

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Sulfate-reducing bacteria (SRBs) induced biofilm formation is a global industrial concern due to its role in the development of microbial-induced corrosion (MIC). Herein, we have developed a biodegradable chitosan/lignosulfonate nanocomposite (CS@LS) as an efficient green biocide for the inhibition of SRBs biofilms. We investigated in detail the inhibition mechanism of SRBs by CS@LS in seawater media. Stable CS@LS-1:1 with 150–200 nm average size and zeta potential of + 34.25 mV was synthesized. The biocidal performance of CS@LS was evaluated by sulfate reduction profiles coupled with analysis of extracted extracellular polymeric substances (EPS) and lactate dehydrogenase (LDH) release assays. As the nanocomposite concentration was increased from 50 to 500 µg/mL, the specific sulfate reduction rate (SSRR) decreased from 0.278 to 0.036 g-sulfate/g-VSS*day showing a relative sulfate reduction inhibition of 86.64% as compared to that of control. Similarly, the specific organic uptake rate (SOUR) decreased from 0.082 to 0.039 0.036 g-TOC/g-VSS*day giving a relative co-substrate oxidation inhibition of 52.19% as compared to that of control. The SRBs spiked with 500 µg/mL CS@LS showed a reduction in cell viability to 1.5 × 106 MPN/mL. To assess the biosafety of the nanocomposite on the marine biota, the 72-hours acute toxicity assays using the zebrafish embryo model revealed that the LC50 for the CS@LS was 103.3 µg/mL. Thus, CS@LS can be classified as environmentally friendly. The nanocomposite showed long-term stability and excellent antibacterial properties against SRBs growth and is thus potentially useful for combating the problems of biofilm growth in harsh marine and aquatic environments.

Keywords: green biocides, chitosan/lignosulfonate nanocomposite, SRBs, toxicity

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Authors: V. T. Gerov, D. I. Gerova, I. D. Micheva, N. F. Nazifova-Tasinova, M. N. Nikolova, M. G. Pasheva, B. T. Galunska

Abstract:

Multiple myeloma (MM) is the most frequent plasma cell (PC) dyscrasia that involves the skeleton. Myeloma bone disease (MBD) is characterized by osteolytic bone lesions as a result of increased osteoclasts activity not followed by reactive bone formation due to osteoblasts suppression. Skeletal complications cause significant adverse effects on quality of life and lead to increased morbidity and mortality. Last decade studies revealed the implication of different proteins in osteoclast activation and osteoblast inhibition. The aim of the present study was to determine serum levels of periostin, sRANKL and osteopontin and to evaluate their role as bone markers in MBD. Materials and methods. Thirty-two newly diagnosed MM patients (mean age: 62.2 ± 10.7 years) and 33 healthy controls (mean age: 58.9 ± 7.5 years) were enrolled in the study. According to IMWG criteria 28 patients were with symptomatic MM and 4 with monoclonal gammopathy of undetermined significance (MGUS). In respect to their bone involvement all symptomatic patients were divided into two groups (G): 9 patients with 0-3 osteolytic lesions (G1) and 19 patients with >3 osteolytic lesions and/or pathologic fractures (G2). Blood samples were drawn for routine laboratory analysis and for measurement of periostin, sRANKL and osteopontin serum levels by ELISA kits (Shanghai Sunred Biological Technology, China). Descriptive analysis, Mann-Whitney test for assessment the differences between groups and non-parametric correlation analysis were performed using GraphPad Prism v8.01. Results. The median serum levels of periostin, sRANKL and osteopontin of ММ patients were significantly higher compared to controls (554.7pg/ml (IQR=424.0-720.6) vs 396.9pg/ml (IQR=308.6-471.9), p=0.0001; 8.9pg/ml (IQR=7.1-10.5) vs 5.6pg/ml (IQR=5.1-6.4, p<0.0001 and 514.0ng/ml (IQR=469.3-754.0) vs 387.0ng/ml (IQR=335.9-441.9), p<0.0001, respectively). for assessment of differences between groups and non-parametric correlation analysis were performed using GraphPad Prism v8.01. Statistical significance was found for all tested bone markers between symptomatic MM patients and controls: G1 vs controls (p<0.03), G2 vs controls (p<0.0001) for periostin; G1 vs controls (p<0.0001), G2 vs controls (p<0.0001) for sRANKL; G1 vs controls (p=0.002), G2 vs controls (p<0.0001) for osteopontin, as well between symptomatic MM patients and MGUS patients: G1 vs MGUS (p<0.003), G2 vs MGUS (p=0.003) for periostin; G1 vs MGUS (p<0.05), G2 vs MGUS (p<0.001) for sRANKL; G1 vs MGUS (p=0.011), G2 vs MGUS (p=0.0001) for osteopontin. No differences were detected between MGUS and controls and between patients in G1 and G2 groups. Spearman correlation analysis revealed moderate positive correlation between periostin and beta-2-microglobulin (r=0.416, p=0.018), percentage bone marrow myeloma PC (r=0.432, p=0.014), and serum total protein (r=0.427, p=0.015). Osteopontin levels were also positively related to beta-2-microglobulin (r=0.540, p=0.0014), percentage bone marrow myeloma PC (r=0.423, p=0.016), and serum total protein (r=0.413, p=0.019). Serum sRANKL was only related to beta-2-microglobulin levels (r=0.398, p=0.024). Conclusion: In the present study, serum levels of periostin, sRANKL and osteopontin in newly diagnosed MM patients were evaluated. They gradually increased from MGUS to more advanced stages of MM reflecting the severity of bone destruction. These results support the idea that some new protein markers could be used in monitoring the MBD as a most severe complication of MM.

Keywords: myeloma bone disease, periostin, sRANKL, osteopontin

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Authors: E. Akdur Öztürk, F. İrvasa Bilgiç, A. Ludovisi , O. Gülbahar, D. Dirim Erdoğan, M. Korkmaz, M. Á. Gómez Morales

Abstract:

Anisakidosis is a zoonotic human fish-borne parasitic disease caused by accidental ingestion of anisakid third-stage larvae (L3) of members of the Anisakidae family present in infected marine fish or cephalopods. Infection with anisakid larvae can lead to gastric, intestinal, extra-gastrointestinal and gastroallergic forms of the disease. Anisakid parasites have been reported in almost all seas, particularly in the Mediterranean Sea. There is a remarkably high level of risk exposure to these zoonotic parasites as they are present in economically and ecologically important fish of Europe. Anisakid L3 larvae have been also detected in several fish species from the Aegean Sea. Turkey is a peninsular country surrounded by Black, Aegean and the Mediterranean Sea. In this country, fishing habit and fishery product consumption are highly common. In recent years, there was also an increase in the consumption of raw fish due to the increasing interest in the cuisine of the Far East countries. In different regions of Turkey, A. simplex (inMerluccius Merluccius Scomber japonicus, Trachurus mediterraneus, Sardina pilchardus, Engraulis encrasicolus, etc.), Anisakis spp., Contraceucum spp., Pseudoterronova spp. and, C. aduncum were identified as well. Although it is accepted both the presence of anisakid parasites in fish and fishery products in Turkey and the presence of Turkish people with allergic manifestations after fish consumption, there are no reports of human anisakiasis in this country. Given the high prevalence of anisakid parasites in the country, the absence of reports is likely not due to the absence of clinical cases rather to the unavailability of diagnostic tools and the low awareness of the presence of this infection. The aim of the study was to set up an IgE-Western Blot (WB) based test to detect the anisakidosis sensitization among Turkish people with a history of allergic manifestation related to fish consumption. To this end, crude worm antigens (CWA) and allergen enriched fraction (50-66% ) were prepared from L3 of A. simplex (s.l.) collected from Lepidopus caudatus fished in the Mediterranean Sea. These proteins were electrophoretically separated and transferred into the nitrocellulose membranes. By WB, specific proteins recognized by positive control serum samples from sensitized patients were visualized on nitrocellulose membranes by a colorimetric reaction. The CWA and 50–66% fraction showed specific bands, mainly due to Ani s 1 (20-22 kD) and Ani s 4 (9-10 kD). So far, a total of 7 serum samples from people with allergic manifestation and positive skin prick test (SPT) after fish consumption, have been tested and all of them resulted negative by WB, indicating the lack of sensitization to anisakids. This preliminary study allowed to set up a specific test and evidence the lack of correlation between both tests, SPT and WB. However, the sample size should be increased to estimate the anisakidosis burden in Turkish people.

Keywords: anisakidosis, fish parasite, serodiagnosis, Turkey

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