Search results for: lateness gene

Authors: Asma Lamin, Anna H. Kaksonen, Ivan Cole, Paul White, Xiao-Bo Chen

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Microbiologically influenced corrosion (MIC) is a process in which microorganisms initiate, facilitate, or accelerate the electrochemical corrosion reactions of metallic components. Several reports documented that MIC accounts for about 20 to 40 % of the total cost of corrosion. Biofilm formation due to the presence of microorganisms on the surface of metal components is known to play a vital role in MIC, which can lead to severe consequences in various environmental and industrial settings. Quorum sensing (QS) system plays a major role in regulating biofilm formation and control the expression of some microbial enzymes. QS is a communication mechanism between microorganisms that involves the regulation of gene expression as a response to the microbial cell density within an environment. This process is employed by both Gram-positive and Gram-negative bacteria to regulate different physiological functions. QS involves production, detection, and responses to signalling chemicals, known as auto-inducers. QS controls specific processes important for the microbial community, such as biofilm formation, virulence factor expression, production of secondary metabolites and stress adaptation mechanisms. The use of QS inhibitors (QSIs) has been proposed as a possible solution to biofilm related challenges in many different applications. Although QSIs have demonstrated some strength in tackling biofouling, QSI-based strategies to control microbially influenced corrosion have not been thoroughly investigated. As such, our research aims to target the QS mechanisms as a strategy for mitigating MIC on metal surfaces in engineered systems.

Keywords: quorum sensing, quorum quenching, biofilm, biocorrosion

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Authors: Pejman Taghibeikzadehbadr

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Muscle contraction stimulates a transient change of myogenic factors, partly related to the mode of contractions. Here, we assessed the response of Insulin-like growth factor 1Ea (IGF-1Ea), Insulin-like growth factor 1Eb (IGF-1Eb), Insulin-like growth factor 1Ec (IGF-1Ec), Peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC1α-1), Peroxisome proliferator-activated receptor gamma coactivator 4-alpha (PGC1α-4), and myostatin to the eccentric Vs the concentric contraction in human skeletal muscle. Ten healthy males were performed an acute eccentric and concentric exercise bout (n = 5 per group). For each contraction type, participants performed 12 sets of 10 repetitions knee extension by the dominant leg. Baseline and post-exercise muscle biopsy were taken 4 weeks before and immediately after experimental sessions from Vastus Lateralis muscle. Genes expression was measured by real-time PCR technique. There was a significant increase in PGC1α-1, PGC1α-4, IGF-1Ea and, IGF-1Eb mRNA after concentric contraction (p ≤ 0.05), while the PGC1α-4 and IGF-1Ec significantly increased after eccentric contraction (p ≤ 0.05). It is intriguing to highlight that; no significant differences between groups were evident for changes in any variables following exercise bouts (p ≥ 0.05). Our results found that concentric and eccentric contractions presented different responses in PGC1α-1, IGF-1Ea, IGF-1Eb, and IGF-1Ec mRNA. However, a similar significant increase in mRNA content was observed in PGC1α-4. Further, no apparent differences could be found between the response of genes to eccentric and concentric contraction.

Keywords: eccentric contraction, concentric contraction, gene expression, PGC-1 alpha, IGF-1 Myostatin

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Authors: Huajuan Shi

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Background: The biopsy of the preimplantation embryo may increase the potential risk and concern of embryo viability. Clinically discarded spent embryo medium (SEM) has entered the view of researchers, sparking an interest in noninvasive embryo screening. However, one of the major restrictions is the extremelty low quantity of cf-RNA, which is difficult to efficiently and unbiased amplify cf-RNA using traditional methods. Hence, there is urgently need to an efficient and low bias amplification method which can comprehensively and accurately obtain cf-RNA information to truly reveal the state of SEM cf-RNA. Result: In this present study, we established an agarose PCR amplification system, and has significantly improved the amplification sensitivity and efficiency by ~90 fold and 9.29 %, respectively. We applied agarose to sequencing library preparation (named AG-seq) to quantify and characterize cf-RNA in SEM. The number of detected cf-RNAs (3533 vs 598) and coverage of 3' end were significantly increased, and the noise of low abundance gene detection was reduced. The increasing percentage 5' end adenine and alternative splicing (AS) events of short fragments (< 400 bp) were discovered by AG-seq. Further, the profiles and characterizations of cf-RNA in spent cleavage medium (SCM) and spent blastocyst medium (SBM) indicated that 4‐mer end motifs of cf-RNA fragments could remarkably differentiate different embryo development stages. Significance: This study established an efficient and low-cost SEM amplification and library preparation method. Not only that, we successfully described the characterizations of SEM cf-RNA of preimplantation embryo by using AG-seq, including abundance features fragment lengths. AG-seq facilitates the study of cf-RNA as a noninvasive embryo screening biomarker and opens up potential clinical utilities of trace samples.

Keywords: cell-free RNA, agarose, spent embryo medium, RNA sequencing, non-invasive detection

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Authors: Karthika Durairaj, Buvaneswari G., Gowdham M., Gilles M., Velmurugan G.

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Background: Hyperglycaemia is the primary cause of metabolic illness. Recently, researchers focused on the possibility that chemical exposure could promote metabolic disease. Hyperglycaemia causes a variety of metabolic diseases dependent on its etiologic conditions. According to animal and population-based research, individual chemical exposure causes health problems through alteration of endocrine function with the influence of microbial influence. We were intrigued by the function of gut microbiota variation in high fat and chemically induced hyperglycaemia. Methodology: C57/Bl6 mice were subjected to two different treatments to generate the etiologic-based diabetes model: I – a high-fat diet with a 45 kcal diet, and II - endocrine disrupting chemicals (EDCs) cocktail. The mice were monitored periodically for changes in body weight and fasting glucose. After 120 days of the experiment, blood anthropometry, faecal metagenomics and metabolomics were performed and analyzed through statistical analysis using one-way ANOVA and student’s t-test. Results: After 120 days of exposure, we found hyperglycaemic changes in both experimental models. The treatment groups also differed in terms of plasma lipid levels, creatinine, and hepatic markers. To determine the influence on glucose metabolism, microbial profiling and metabolite levels were significantly different between groups. The gene expression studies associated with glucose metabolism vary between hosts and their treatments. Conclusion: This research will result in the identification of biomarkers and molecular targets for better diabetes control and treatment.

Keywords: hyperglycaemia, endocrine-disrupting chemicals, gut microbiota, host metabolism

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Authors: Regina R. Gunawan, Indwiani Astuti, H. Raden Danarto

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Cancer is the leading cause of death worldwide. Cancer is caused by mutations that alter the function of normal human genes and give rise to cancer genes. MicroRNA (miRNA) is a small non-coding RNA that regulates the gen through complementary bond towards mRNA target and cause mRNA degradation. miRNA works by either promoting or suppressing cell proliferation. miRNA level expression in cancer may offer another value of miRNA as a biomarker in cancer diagnostic. miRNA-21 is believed to have a role in carcinogenesis by enhancing proliferation, anti-apoptosis, cell cycle progression and invasion of tumor cells. Hsa-miR-21-5p marker has been identified in Prostate Cancer (PCa) and Benign Prostatic Hyperplasia (BPH) patient’s urine. This research planned to explore the diagnostic performance of miR-21 to differentiate PCa and BPH patients. In this study, urine samples were collected from 20 PCa patients and 20 BPH patients. miR-21 relative expression against the reference gene was analyzed and compared between the two. miRNA expression was analyzed using the comparative quantification method to find the fold change. miR-21 validity in identifying PCa patients was performed by quantifying the sensitivity and specificity with the contingency table. miR-21 relative expression against miR-16 in PCa patient and in BPH patient has 12,98 differences in fold change. From a contingency table of Cq expression of miR-21 in identifying PCa patients from BPH patient, Cq miR-21 has 100% sensitivity and 75% specificity. miR-21 relative expression can be used in discriminating PCa from BPH by using a urine sample. Furthermore, the expression of miR-21 has higher sensitivity compared to PSA (Prostate specific antigen), therefore miR-21 has a high potential to be analyzed and developed more.

Keywords: benign prostate hyperplasia, biomarker, miRNA-21, prostate cancer

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Authors: Dipranjan Laha, Parimal Karmakar

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Metal oxide nanoparticles are well known to generate oxidative stress and deregulate normal cellular activities. Among these, transition metals copper oxide nanoparticles (CuO NPs) are more compelling than others and able to modulate different cellular responses. In this work, we have synthesized and characterized CuO NPs by various biophysical methods. These CuO NPs (~30 nm) induce autophagy in human breast cancer cell line, MCF7 in a time and dose-dependent manner. Cellular autophagy was tested by MDC staining, induction of green fluorescent protein light chain 3 (GFP-LC3B) foci by confocal microscopy, transfection of pBABE-puro mCherry-EGFP-LC3B plasmid and western blotting of autophagy marker proteins LC3B, beclin1, and ATG5. Further, inhibition of autophagy by 3-Methyladenine (3-MA) decreased LD50 doses of CuO NPs. Such cell death was associated with the induction of apoptosis as revealed by FACS analysis, cleavage of PARP, dephosphorylation of Bad and increased cleavage product of caspase3. siRNA-mediated inhibition of autophagy-related gene beclin1 also demonstrated similar results. Finally, induction of apoptosis by 3-MA in CuO NPs treated cells were observed by TEM. This study indicates that CuO NPs are a potent inducer of autophagy which may be a cellular defense against the CuO NPs mediated toxicity and inhibition of autophagy switches the cellular response into apoptosis. A combination of CuO NPs with the autophagy inhibitor is essential to induce apoptosis in breast cancer cells. Acknowledgments: The authors would like to acknowledge for financial support for this research work to the Department of Biotechnology (No. BT/PR14661/NNT/28/494/2010), Government of India.

Keywords: nanoparticle, autophagy, apoptosis, siRNA-mediated inhibition

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Authors: Bahadir Batar, Elif Serdal, Berna Erdal, Hasan Ogul

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Micro-ribonucleic acids (miRNAs) are small short non-coding ribonucleic acid molecules about 22 nucleotides long. miRNAs play a key role in response to chemotherapeutic agents. WW domain-containing oxidoreductase (WWOX) gene encodes a tumor suppressor protein. Loss or reduction of Wwox protein is observed in many breast cancer cases. WWOX protein deficiency is increased in triple-negative breast cancer (TNBC). TNBC is a heterogeneous, highly aggressive, and difficult to treat tumor type. WWOX loss contributes to resistance to cisplatin therapy in patients with TNBC. Here, the aim of the study was to investigate the potential role of miRNAs in cisplatin therapy resistance of WWOX-deficient TNBC cells. This was a cell culture study. miRNA expression profiling was analyzed by LightCycler 480 system. miRNA Set Enrichment Analysis tool was used to integrate experimental data with literature-based biological knowledge to infer a new hypothesis. Increased miR-182 and decreased miR-214 were significantly correlated with cisplatin resistance in WWOX-deficient TNBC cells. miR-182 and miR-214 may involve in cisplatin resistance of WWOX-deficient TNBC cells by deregulating the DNA repair, apoptosis, or protein kinase B signaling pathways. These data highlight the mechanism by which WWOX regulates cisplatin resistance of TNBC and the potential use of WWOX as a predictor biomarker for cisplatin resistance.

Keywords: cisplatin, microRNA, triple-negative breast cancer, WWOX

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Authors: Tomáš Nejedlý, Dominika Mrázková, Jitka Viktorová

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The blood-brain barrier (BBB) serves as a protective shield, preventing the entry of harmful substances into the central nervous system. On the other hand, it also restricts the transport of neuroactive drugs, such as antiepileptics, which mitigate epileptic seizures. Drug-resistant epilepsy is often associated with the overexpression of efflux transporters, including P-glycoprotein (P-gp) or multidrug resistance protein 1 (MRP1), on the BBB. The aim of this work is to find P-gp and MRP1 inhibitors derived from phytocannabinoids and terpenes. The work evaluates whether these compounds interact directly with P-gp or MRP1 by rhodamine 123 or fluorescein efflux assay. The effect of phytocannabinoids on the gene expression of these transporters is also studied using qPCR and Western blot. These transporters are found in BBB cells; however, we decided to use the human ovarian cancer cell line (A2780ADR) due to its overproduction of P-gp and malignant glioma cell line (U87) due to its overproduction of MRP1. The results showed that while terpenes suppressed the activity of efflux transporters, phytocannabinoids tended to decrease their expression. Terpenes demonstrated an average inhibition of 65%, surpassing phytocannabinoids, which exhibited an average inhibition of approximately 30%. Particularly noteworthy was the modulating effect of (-)-α-bisabolol with the highest activity among the compounds tested. Based on these findings, phytocannabinoids and terpenes emerge as promising natural candidates for addressing drug resistance linked to efflux transporters. Acknowledgment: The project was funded by the Grant No 22-20860S of The Czech Science Foundation.

Keywords: drug-resistant epilepsy, efflux transporters, multidrug resistance protein 1, P-glycoprotein, phytocannabinoids, terpens

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Authors: Yong Zhang

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Light has proven to be useful in a wide range of biomedical applications such as fluorescence imaging, photoacoustic imaging, optogenetics, photodynamic therapy, photothermal therapy, and light controlled drug/gene delivery. Taking photodynamic therapy (PDT) as an example, PDT has been proven clinically effective in early lung cancer, bladder cancer, head, and neck cancer and is the primary treatment for skin cancer as well. However, clinical use of PDT is severely constrained by the low penetration depth of visible light through thick tissue, limiting its use to target regions only a few millimeters deep. One way to enhance the range is to use invisible near-infrared (NIR) light within the optical window (700–1100nm) for biological tissues, extending the depth up to 1cm with no observable damage to the intervening tissue. We have demonstrated use of NIR-to-visible upconversion fluorescent nanoparticles (UCNPs), emitting visible fluorescence when excited by a NIR light at 980nm, as a nanotransducer for PDT to convert deep tissue-penetrating NIR light to visible light suitable for activating photosensitizers. The unique optical properties of UCNPs enable the upconversion wavelength to be tuned and matched to the activation absorption wavelength of the photosensitizer. At depths beyond 1cm, however, tissue remains inaccessible to light even within the NIR window, and this critical depth limitation renders existing phototherapy ineffective against most deep-seated cancers. We have demonstrated some new treatment modalities for deep-seated cancers based on UCNP hydrogel implants and miniaturized, wirelessly powered optoelectronic devices for light delivery to deep tissues.

Keywords: upconversion, fluorescent, nanoparticle, bioimaging, photodynamic therapy

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Authors: M. F. Miah, K. M. A. Zinnah, M. J. Raihan, H. Ali, M. N. Naser

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As genetic diversity is most important for existing, breeding and production of any fish; this study was undertaken for investigating genetic diversity of freshwater mud eel, Monopterus cuchia at population level where three ecological populations such as flooded area of Sylhet (P1), open water of Moulvibazar (P2) and open water of Sunamganj (P3) districts of Bangladesh were considered. Four arbitrary RAPD primers (OPB-12, C0-4, B-03 and OPB-08) were screened and RAPD banding patterns were analyzed among the populations considering 15 individuals of each population. In total 174, 138 and 149 bands were detected in the populations of P1, P2 and P3 respectively; however, each primer revealed less number of bands in each population. 100% polymorphic loci were recorded in P2 and P3 whereas only one monomorphic locus was observed in P1, recorded 97.5% polymorphism. Different genetic parameters such as inter-individual pairwise similarity, genetic distance, Nei genetic similarity, linkage distances, cluster analysis and allelic information, etc. were considered for measuring genetic diversity. The average inter-individual pairwise similarity was recorded 2.98, 1.47 and 1.35 in P1, P2 and P3 respectively. Considering genetic distance analysis, the highest distance 1 was recorded in P2 and P3 and the lowest genetic distance 0.444 was found in P2. The average Nei genetic similarity was observed 0.19, 0.16 and 0.13 in P1, P2 and P3, respectively; however, the average linkage distance was recorded 24.92, 17.14 and 15.28 in P1, P3 and P2 respectively. Based on linkage distance, genetic clusters were generated in three populations where 6 clades and 7 clusters were found in P1, 3 clades and 5 clusters were observed in P2 and 4 clades and 7 clusters were detected in P3. In addition, allelic information was observed where the frequency of p and q alleles were observed 0.093 and 0.907 in P1, 0.076 and 0.924 in P2, 0.074 and 0.926 in P3 respectively. The average gene diversity was observed highest in P2 (0.132) followed by P3 (0.131) and P1 (0.121) respectively.

Keywords: genetic diversity, Monopterus cuchia, population, RAPD, Bangladesh

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Authors: Belaynesh Tsegay, Teklay Gebrecherkos, Atsebaha Gebrekidan Kahsay, Mahmud Abdulkader

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Background: Hepatitis B and C viruses are important health and socioeconomic problem across the globe, with a remarkable number of diseases and deaths in sub-Saharan African countries. The burden of hepatitis is unknown in the prison settings of Tigray. Therefore, we aimed to describe the seroprevalence and associated factors of hepatitis B and C viruses among prisoners in Tigray, Ethiopia. Methods: A cross-sectional study was carried out from February 2020 to May 2020 at the prison facilities of Tigray. Demographics and associated factors were collected from 315 prisoners prospectively. Five milliliters of blood were collected and tested using rapid tests kits of HBsAg (Zhejiang orient Gene Biotech Co., Ltd., China) and HCV antibodies (Volkan Kozmetik Sanayi Ve Ticaret Ltd. STI, Turkey). Positive samples were confirmed using ELISA (Beijing Wantai Biological Pharmacy Enterprise Co. Ltd). Data were analyzed using the SPSS version 20, and p<0.05 was considered statistically significant. Results: The overall seroprevalence of HBV and HCV were 25 (7.9%) and 1 (0.3%), respectively. The majority of hepatitis B viral infections were identified from the age groups of 18–25 years (10.7%) and unmarried prisoners (11.8%). Prisoners greater than 100 per cell (AOR=3.95, 95% CI=1.15–13.6, p=0.029) and with a history of alcohol consumption (AOR=3.01, 95% CI=1.17–7.74, p=0.022) were significantly associated with HBV infections. Conclusion: The seroprevalence of HBV among prisoners was nearly high or borderline, with a very low HCV prevalence. HBV was most prevalent among young adults, those housed with a large number of prisoners per cell, and those who had a history of alcohol consumption. This study recommends that there should be prison-focused intervention, including regular health education, with the emphasis on the mode of transmission and introducing HBV screening policy for prisoners, especially when they enter the prison.

Keywords: seroprevalence, HBV, HCV, prisoners, tigray

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Authors: Negar Zaheddoust, Negin Zaheddoust, Abbas Asoudeh-Fard

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Probiotic bacteria have been employed as a novel and less side-effect strategy for anticancer therapy. Since the oral cavity is a host for probiotic and pathogen bacteria to colonize, more investigation is needed to evaluate the effectiveness of this novel adjunctive treatment for oral cancer. We considered Lactobacillus plantarum as a probiotic and Streptococcus mutans as a pathogen bacterium in our study. The aim of this study is to examine the effect of Lactobacillus plantarum and Streptococcus mutans on Casp3, AKT / PTEN, and MAPK signaling pathway, which is involved in apoptosis or survival of oral cancer KB cells. On the other hand, to study the effects of these bacteria on normal cells, we used HUVEC cells. The KB and HUVEC cell lines were co-cultured with Lactobacillus plantarum and Streptococcus mutans isolated from traditional Iranian dairy and dental plaque, respectively. The growth-inhibitory effects of these two bacteria on KB and HUVEC cells were determined by (3-(4, 5-dimethylthiazolyl-2)-2,5diphenyltetrazolium bromide) MTT assay. MTT results demonstrated that the proliferation of KB cells was affected in a time, dose, and strain-dependent manner. In the following, the examination of induced apoptosis or necrosis in co-cultured KB cells with the best IC50 concentration of the Lactobacillus plantarum and Streptococcus mutans will be analyzed by FACS flow cytometry, and the changes in gene expression of Casp3, AKT / PTEN, MAPK genes will be evaluated using real-time polymerase chain reaction.

Keywords: cancer therapy, induced apoptosis, oral cancer, probiotics

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Authors: Lata Kumari Dhanesh Tiwary, Pradeep Kumar Mishra

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The effluent coming from various industries such as textile, carpet, food, pharmaceutical and many other industries is big challenge due to its recalcitrant and xenobiotiocs in nature. Recently, biodegradation of dye wastewater through biological means was widely used due to eco-friendly and cost effective with the higher percentage of removal of dye from wastewater. The present study deals with the biodegradation and decolourization of Direct Red 23 dye using indigenously isolated bacterial consortium. The bacterial consortium was isolated from soil sample from dye contaminated site near a cluster of Carpet industries of Bhadohi, Uttar Pradesh, India. The bacterial strain formed consortia were identified and characterized by morphological, biochemical and 16S rRNA gene sequence analysis. The bacterial strain mainly Staphylococcus saprophyticus strain BHUSS X3 (KJ439576), Microbacterium sp. BHUMSp X4 (KJ740222) and Staphylococcus saprophyticus strain BHUSS X5 (KJ439576) were used as consortia for further studies of dye decolorization. Experimental investigations were made in a Sequencing Air- lift bioreactor using the synthetic solution of Direct Red 23 dye by optimizing various parameters for efficient degradation of dye. The effect of several operating parameters such as flow rate, pH, temperature, initial dye concentration and inoculums size on removal of dye was investigated. The efficiency of isolated bacterial consortia from dye contaminated area in Sequencing Air- lift Bioreactor with different concentration of dye between 100-1200 mg/l at different hydraulic rate (HRTs) 26h and 10h. The maximum percentage of dye decolourization 98% was achieved when operated at HRT of 26h. The percentage of decolourization of dye was confirmed by using UV-Vis spectrophotometer and HPLC.

Keywords: carpet industry, bacterial consortia, sequencing air-lift bioreactor

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Authors: Syeda Fahria Hoque Mimmi, Aparna Islam

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Genetically engineered (GE) plants are the need of time for increased demand for food. A complete set of regulations need to be followed from the development of a GE plant to its release into the environment. The whole regulation system is categorized into separate stages for maintaining the proper biosafety. Environmental risk assessment (ERA) is one of such crucial stages in the whole process. ERA identifies potential risks and their impacts through science-based evaluation where it is done in a case-by-case study. All the countries which deal with GE plants follow specific guidelines to conduct a successful ERA. In this study, ERA guidelines of 4 developing and 4 developed countries, including Bangladesh, were compared. ERA guidelines of countries such as India, Canada, Australia, the European Union, Argentina, Brazil, and the US were considered as a model to conduct the comparison study with Bangladesh. Initially, ten parameters were detected to compare the required data and information among all the guidelines. Surprisingly, an adequate amount of data and information requirements (e.g., if the intended modification/new traits of interest has been achieved or not, the growth habit of GE plants, consequences of any potential gene flow upon the cultivation of GE plants to sexually compatible plant species, potential adverse effects on the human health, etc.) matched between all the countries. However, a few differences in data requirement (e.g., agronomic conventions of non-transformed plants, applicants should clearly describe experimental procedures followed, etc.) were also observed in the study. Moreover, it was found that only a few countries provide instructions on the quality of the data used for ERA. If these similarities are recognized in a more framed manner, then the approval pathway of GE plants can be shared.

Keywords: GE plants, ERA, harmonization, ERA guidelines, Information and data requirements

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Authors: A. Reum Son, Jin Seon Kwon, Seung Hun Park, Hai Bang Lee, Moon Suk Kim

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These days, stem cell therapy is focused on for promising source of treatment in clinical human disease. As a supporter of stem cells, in situ-forming hydrogels with growth factors and cells appear to be a promising approach in tissue engineering. To examine osteogenic differentiation of hTMSCs which is one of mesenchymal stem cells in vivo in an injectable hydrogel, we use a methoxy polyethylene glycol-polycaprolactone blockcopolymer (MPEG-PCL) solution with osteogenic factors. We synthesized MPEG-PCL hydrogel and measured viscosity to check sol-gel transition. In order to demonstrate osteogenic ability of hTMSCs, we conducted in vitro osteogenesis experiment. Then, to confirm the cell cytotoxicity, we performed WST-1 with hTMSCs and MPEG-PCL. As the result of in vitro experiment, we implanted cell and hydrogel mixture into animal model and checked degree of osteogenesis with histological analysis and amount of expression genes. Through these experimental data, MPEG-PCL hydrogel has sol-gel transition in temperature change and is biocompatible with stem cells. In histological analysis and gene expression, hTMSCs are very good source of osteogenesis with hydrogel and will use it to tissue engineering as important treatment method. hTMSCs could be a good adult stem cell source for usability of isolation and high proliferation. When hTMSCs are used as cell therapy method with in situ-formed hydrogel, they may provide various benefits like a noninvasive alternative for bone tissue engineering applications.

Keywords: injectable hydrogel, stem cell, osteogenic differentiation, tissue engineering

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Authors: Rashad R. Al-Hindi

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The aim of this study was to isolate and identify lactic acid bacteria and evaluate their therapeutic and food preservation importance. Ninety-three suspected lactic acid bacteria (LAB) were isolated from thirteen different raw and fermented milk of indigenous sources in the Kingdom of Saudi Arabia. The identification of forty-six selected LAB strains and genetic relatedness were performed based on 16S rDNA gene sequence comparison. The LAB counts in certain samples were higher under microaerobic than anaerobic conditions. The identified LAB belonged to genera Enterococcus (16 strains), Lactobacillus (9 strains), Weissella (10 strains), Streptococcus (8 strains) and Lactococcus (3 strains). Phylogenetic tree generated from the full-length (~1.6 kb) sequences confirmed previous findings. Utilization of shorter 16S rDNA sequences (~1.0 kb) also discriminated among strains of which V2 region was the most effective. None of the strains exhibited resistance to clinically relevant antibiotics or undesirable hemolytic activity, while they differed in other probiotic characteristics, e.g., tolerance to acidic pH, resistance to bile salt, and antibacterial activity. In conclusion, the isolates Lactobacillus casei MSJ1, Lactobacillus casei Dwan5, Lactobacillus plantarum EyLan2 and Enterococcus faecium Gail-BawZir8 are likely the best probiotic LAB and we speculate that studying the synergistic effects of bacterial combinations might result in the occurrence of more effective probiotic potential. We argue that the raw and fermented milk of animals hosted in Saudi Arabia, especially stirred yogurt (Laban) made from camel milk, are rich in LAB with promising probiotics potential.

Keywords: fermented foods, lactic acid bacteria, probiotics, Saudi Arabia

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Authors: Arefeh Jafarian, Mohammad Taghikani, Saied Abroun, Amir Allahverdi, Masoud Soleimani

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Introduction: The miRNAs have key roles in control of pancreatic islet development and insulin secretion. In this regards, current study investigated the pancreatic differentiation of human bone marrow mesenchymal stem cells (hBM-MSCs) by up-regulation of miR-375 and down-regulation of miR-9 by lentiviruses containing miR-375 and anti-miR-9. Findings: After 21 days of induction, islet-like clusters containing insulin producing cells (IPCs) were confirmed by dithizone (DTZ) staining. The IPCs and β cell specific related genes and proteins were detected using qRT-PCR and immunofluorescence on days 7, 14 and 21 of differentiation. Glucose challenge test was performed at different concentrations of glucose as well as extracellular and intracellular insulin and C-peptide were assayed using ELISA kit. In derived IPCs by miR-375 alone are capable to express insulin and other endocrine specific transcription factors, the cells lack the machinery to respond to glucose. The differentiated hMSCs by miR-375 and anti-miR-9 lentiviruses could secrete insulin and c-peptide in a glucose-regulated manner. Conclusion: It was found that over-expression of miR-375 led to a reduction in levels of Mtpn protein in derived IPCs, while treatment with anti-miR-9 following miR-375 over-expression had synergistic effects on MSCs differentiation and insulin secretion in a glucose-regulated manner. The researchers reported that silencing of miR-9 increased OC-2 protein in IPCs that may contribute to the observed glucose-regulated insulin secretion. These findings highlight miRNAs functions in stem cells differentiation and suggest that they could be used as therapeutic tools for gene-based therapy in diabetes mellitus.

Keywords: diabetes, differentiation, MSCs, insulin producing cells, miR-375, miR-9

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Authors: Yves Mann Elate Lea Mbassi, Marie Solange Evehe Bebandoue, Wilfred Fon Mbacham

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Improving the catalytic performance of enzymes has been a long-standing theme of analytical biochemistry research. Induction of peroxidase activity by metals is a common reaction in higher plants. We thought that this increase in peroxidase activity may be due, on the one hand, to the stimulation of the gene expression of these enzymes but also to a modification of their chemical reactivity following the binding of some metal ions on their active site. We tested the effect of some metal salts (MgCl₂, MnCl₂, ZnCl₂, CaCl₂ and CuSO₄) on the activity and thermostability of peroxidase A6, a thermostable peroxidase that we discovered and purified in a previous study. The chromogenic substrate used was 3,3′,5,5′-tetramethylbenzidine. Of all the metals tested for their effect on A6, only magnesium and copper had a significant effect on the activity of the enzyme at room temperature. The Mann-Whitney test shows a slight inhibitory effect of activity by the magnesium salt (P = 0.043), while the activity of the enzyme is 5 times higher in the presence of the copper salt (P = 0.002). Moreover, the thermostability of peroxidase A6 is increased when calcium and copper salts are present. The activity in the presence of CaCl₂ is 8 times higher than the residual activity of the enzyme alone after incubation at 80°C for 10 min and 35 times higher in the presence of CuSO4 under the same conditions. In addition, manganese and zinc salts slightly reduce the thermostability of the enzyme. The activity and structural stability of peroxidase A6 can clearly be activated by Cu₂+, which therefore enhance the oxidation of 3,3′,5,5′-tetramethylbenzidine, which was used in this study as a chromogenic substrate. Ca₂+ likely has a more stabilizing function for the catalytic site.

Keywords: peroxidase activity, copper ions, calcium ions, thermostability

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Authors: Gyati Shilakari Asthana, Abhay Asthana

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Purpose: The therapeutic potential of oligonucleotide (ODN) is primarily dependent upon its safe and efficient delivery to specific cells overcoming degradation and maximizing cellular uptake in vivo. The present study is focused to design low molecular weight chitosan nanoconstructs to meet the requirements of safe and effectual delivery of ODNs. LMW-chitosan is a biodegradable, water soluble, biocompatible polymer and is useful as a non-viral vector for gene delivery due to its better stability in water. Methods: LMW chitosan ODN nanoparticles (CHODN NPs) were formulated by self assembled method using various N/P ratios (moles ratio of amine groups of CH to phosphate moieties of ODNs; 0.5:1, 1:1, 3:1, 5:1 and 7:1) of CH to ODN. The developed CHODN NPs were evaluated with respect to gel retardation assay, particle size, zeta potential and cytotoxicity and transfection efficiency. Results: Complete complexation of CH/ODN was achieved at the charge ratio of 0.5:1 or above and CHODN NPs displayed resistance against DNase I. On increasing the N/P ratio of CH/ODN, particle size of the NPs decreased whereas zeta potential (ZV) value increased. No significant toxicity was observed at all CH concentrations. The transfection efficiency was increased on increasing N/P ratio from 1:1 to 3:1, whereas it was decreased with further increment in N/P ratio upto 7:1. Maximum transfection of CHODN NPs with both the cell lines (Raw 267.4 cells and Hela cells) was achieved at N/P ratio of 3:1. The results suggest that transfection efficiency of CHODN NPs is dependent on N/P ratio. Conclusion: Thus the present study states that LMW chitosan nanoparticulate carriers would be acceptable choice to improve transfection efficiency in vitro as well as in vivo delivery of oligonucleotide.

Keywords: LMW-chitosan, chitosan nanoparticles, biocompatibility, cytotoxicity study, transfection efficiency, oligonucleotide

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Authors: B. S. Yadav, A. Mani, S. Srivastava

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Chickpea (Cicer arietinum L.) is an annual, self-pollinating, diploid (2n = 2x = 16) pulse crop that ranks second in world legume production after common bean (Phaseolus vulgaris). ICC 4958 flowers approximately 39 days after sowing under peninsular Indian conditions and the crop matures in less than 90 days in rained environments. The estimated collective yield losses due to abiotic stresses (6.4 million t) have been significantly higher than for biotic stresses (4.8 million t). Most legumes are known to be salt sensitive, and therefore, it is becoming increasingly important to produce cultivars tolerant to high-salinity in addition to other abiotic and biotic stresses for sustainable chickpea production. Our aim was to identify the genes that are involved in the defence mechanism against heavy metal toxicity in chickpea and establish the biological network of heavy metal toxicity in chickpea. ICC4958 variety of chick pea was taken and grown in normal condition and 150µM concentration of different heavy metal salt like CdCl₂, K₂Cr2O₇, NaAsO₂. At 15th day leave samples were collected and stored in RNA Later solution microarray was performed for checking out differential gene expression pattern. Our studies revealed that 111 common genes that involved in defense mechanism were up regulated and 41 genes were commonly down regulated during treatment of 150µM concentration of CdCl₂, K₂Cr₂O₇, and NaAsO₂. Biological network study shows that the genes which are differentially expressed are highly connected and having high betweenness and centrality.

Keywords: abiotic stress, biological network, chickpea, microarray

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Authors: Tereza Branyšová, Martina Kračmarová, Kateřina Demnerová, Michal Ďurovič, Hana Stiborová

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Microbial deterioration threatens all objects of cultural heritage, including audio-visual materials. Fungi are commonly known to be the main factor in audio-visual material deterioration. However, although being neglected, bacteria also play a significant role. In addition to microbial contamination of materials, it is also essential to analyse air as a possible contamination source. This work aims to identify bacterial species in the archives of the Czech Republic that occur on audio-visual materials as well as in the air in the archives. For sampling purposes, the smears from the materials were taken by sterile polyurethane sponges, and the air was collected using a MAS-100 aeroscope. Metagenomic DNA from all collected samples was immediately isolated and stored at -20 °C. DNA library for the 16S rRNA gene was prepared using two-step PCR and specific primers and the concentration step was included due to meagre yields of the DNA. After that, the samples were sent to the University of Fairbanks, Alaska, for Illumina MiSeq sequencing. Subsequently, the analysis of the sequences was conducted in R software. The obtained sequences were assigned to the corresponding bacterial species using the DADA2 package. The impact of air contamination and the impact of different photosensitive layers that audio-visual materials were made of, such as gelatine, albumen, and collodion, were evaluated. As a next step, we will take a deeper focus on air contamination. We will select an appropriate culture-dependent approach along with a culture-independent approach to observe a metabolically active species in the air. Acknowledgment: This project is supported by grant no. DG18P02OVV062 of the Ministry of Culture of the Czech Republic.

Keywords: cultural heritage, Illumina MiSeq, metagenomics, microbial identification

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Authors: Vivek Vaishnav, Shamim Akhtar Ansari

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Teak (Tectona grandis L.f.) belonging to plant family Lamiaceae and the most commercialized timber species is endemic to South-Asia. The adaptive fitness of the species metapopulation was evaluated through its genetic differentiation and assessing the influence of geo-climatic conditions. 290 genotypes were sampled from 29 locations of its natural distribution and the genetic data was incorporated with geo-climatic parameters. Through Bayesian approach based analysis of 43 highly polymorphic ISSR markers, six homogeneous clusters (0.8% genetic variability) were identified. The six clusters were found with the various regimes of the temperature range, i.e., I - 9.10±1.35⁰C, II -6.35±0.21⁰C, III -12.21±0.43⁰C, IV - 10.8±1.06⁰C, V - 11.67±3.04⁰C, and VI - 12.35±0.21⁰C. The population had a very high percentage of LD (21.48%) among the amplified loci possibly due to experiencing restricted gene flow as well as co-adaptation and association of distant/diverse loci/alleles as a result of the stabilized climatic conditions and countless cycles of historical recombination events on a large geological timescale. The same possibly accounts for the narrow distribution of teak as a climax species in the tropical deciduous forests of the country. The regions of strong LD in teak genome significantly associated with climatic parameters also reflect that the species is tolerant to the wide regimes of the temperature range and may possibly withstand global warming and climate change in the coming millennium.

Keywords: Bayesian analysis, inter simple sequence repeat, linkage disequilibrium, marker-geoclimatic association

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Authors: M. Sangeetha, D. Chamundeeswari, C. Saravanababu, C. Rose, V. Gopal

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Rheumatoid arthritis (RA) is a chronic, progressive, systemic inflammatory disorder affecting the synovial joints and typically producing symmetrical arthritis that leads to joint destruction, which is responsible for the deformity and disability. Despite improvements in the treatment of RA over the past decade, there still is a need for new therapeutic agents that are efficacious, less expensive, and free of severe adverse reactions. The present study aimed to investigate role of inflammatory markers in arthritic rats treated with ethanolic bark extract of Albizia procera. The protective effect of ethanolic bark extract of Albizia procera against complete Freund’s adjuvant (CFA) induced arthritis in rats. Arthritis was induced by an intradermal injection of 0.1 ml FCA in the foot pad of left hind limb of rats. ETBE (100 and 200 mg/kg b.wt./p.o) and the reference drug diclofenac (25 mg/kg b.wt./p.o) were administered to arthritic rats. Paw volume was measured for all the animals before inducing arthritis and thereafter once in seven days by using plethysmometer for 42 days. Gene expression of inflammatory markers such as IL-1β and IL-10 were investigated in paw tissues. Up regulation of IL-1β and Down regulation IL-10 were observed in CFA injected rats when compared to normal rats. ETBE attenuated these alterations dose dependently when compared to the vehicle treated rats. These results provide insights into the mechanism of anti-arthritic activity, and unravel potential therapeutic use of Albizia procera in arthritis.

Keywords: CFA-Complete Freund’s adjuvant, ETBE – ethanolic bark extract, IL- interleukins, RA-rheumatoid arthritis

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Authors: Benchouala Amira, Bojic Clement, Poupin Pascal, Cossu Leguille-carole

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The objective of this study is to evaluate in vitro the cytotoxic and androgenic potential of several antifungal molecules (amphotericin B, econazole, ketoconazole and miconazole) on MDA-Kb2 cell lines. This biological model is an effective tool for the detection of endocrine disruptors because it responds well to the main agonist of the androgen receptor (testosterone) and also to an antagonist: flutamide. The cytotoxicity of each chemical compound tested was measured using an MTT assay (tetrazolium salt, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) which measures the activity of the reductase function of mitochondrial succinate dehydrogenase enzymes of cultured cells. This complementary cytotoxicity test is essential to ensure that the effects of reduction in luminescence intensity observed during androgenic tests are only attributable to the anti-androgenic action of the compounds tested and not to their possible cytotoxic properties. Tests of the androgenic activity of antifungals show that these compounds do not have the capacity to induce transcription of the luciferase gene. These compounds do not exert an androgenic effect on MDA-Kb2 cells in culture for the environmental concentrations tested. The addition of flutamide for the same tested concentrations of antifungal molecules reduces the luminescence induced by amphotericin B, econazole and miconazole, which is explained by a strong interaction of these molecules with flutamide which may have a greater toxic effect than when tested alone. The cytotoxicity test shows that econazole and ketoconazole can cause cell death at certain concentrations tested. This cell mortality is perhaps induced by a direct or indirect action on deoxyribonucleic acid (DNA), ribonucleic acid (RNA) or proteins necessary for cell division.

Keywords: cytotoxicity, androgenic potential, antifungals, MDA-Kb2

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Authors: Simona Pileviciene

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On 24th January 2014, Lithuania notified two primary cases of African swine fever (ASF) in wild boars. The animals were tested positive for ASF virus (ASFV) genome by real-time PCR at the National Reference Laboratory for ASF in Lithuania (NRL), results were confirmed by the European Union Reference Laboratory for African swine fever (CISA-INIA). Intensive wild and domestic animal monitoring program was started. During the period of 2014-2017 ASF was confirmed in two large commercial pig holding with the highest biosecurity. Pigs were killed and destroyed. Since 2014 ASF outbreak territory from east and south has expanded to the middle of Lithuania. Diagnosis by PCR is one of the highly recommended diagnostic methods by World Organization for Animal Health (OIE) for diagnosis of ASF. The aim of the present study was to compare singleplex real-time PCR assays to a duplex assay allowing the identification of ASF and internal control in a single PCR tube and to compare primers, that target the p72 gene (ASF 250 bp and ASF 75 bp) effectivity. Multiplex real-time PCR assays prove to be less time consuming and cost-efficient and therefore have a high potential to be applied in the routine analysis. It is important to have effective and fast method that allows virus detection at the beginning of disease for wild boar population and in outbreaks for domestic pigs. For experiments, we used reference samples (INIA, Spain), and positive samples from infected animals in Lithuania. Results show 100% sensitivity and specificity.

Keywords: African swine fewer, real-time PCR, wild boar, domestic pig

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Authors: R. Gounina-Allouane, N. Ouali, F. Z. Berrabah, A. Bentaleb

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For a long time, the Gram-positive spore-forming bacteria Bacillus thuringiensis (Bt) has been widely used in biological control against devastating and disease vectors insects. This is due to the insecticidal activity of its crystalline parasporal inclusion (crystals) predominantly comprised of one or more proteins (Cry and Cyt proteins) also called δ-endotoxins, produced during sporulation. The shape and composition of Bt crystals vary among strains and crystalline proteins are extremely varied (more than 475 cry gene were discovered). The insecticidal activity of Bt crystals is very well studied, thus their insecticidal mode of action is well established, however, their antimicrobial effect is largely unknown. The lack of data on the antimicrobial effect of crystalline proteins of Bt and the need for searching new antimicrobial molecules encouraged us to carried out this study. The antibacterial effect of δ-endotoxines produced by two Bt stains; a strain isolated from soil at northern of Algeria (Bt 7.2.B), and a strain isolated from a bioinsecticide (Bacillus thuringiensis var aizawai), activated by proteolysis, was assayed on clinical bacterial strains and ATCC collection ones respectively. Gram positive and negative clinical bacterial strains (Escherichia coli, Klebsiella pneumonaie, Pseudomonas aeruginosa, Staphylococcus aureus) were sensitive to activated Bt 72B endotoxins. Similarly, bacterial strains from ATCC collection (Escherichia coli ATCC 25922, Pseudomonas aerugenosa ATCC 27853, Staphylococcus aureus ATCC 25923) were sensitive to activated B. thuringiensis var aizawai δ-endotoxines. The activated δ-endotoxins were separated by SDS-PAGE.

Keywords: Bacillus thuringiensis, crystals, cry proteins, δ-endotoxins, antibacterial activity

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Authors: Ibrahim Iddrisu, Joseph Ayariga, Junhuan Xu, Ayomide Adebanjo, Boakai K. Robertson, Michelle Samuel-Foo, Olufemi Ajayi

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The rise of antimicrobial resistance is a global public health crisis that threatens the effective control and prevention of infections. Due to the emergence of pan drug-resistant bacteria, most antibiotics have lost their efficacy. Bacteriophages or their components are known to target bacterial cell walls, cell membranes, and lipopolysaccharides (LPS) and hydrolyze them. Bacteriophages, being the natural predators of pathogenic bacteria, are inevitably categorized as ‘human friends’, thus fulfilling the adage that ‘the enemy of my enemy is my friend’. Leveraging on their lethal capabilities against pathogenic bacteria, researchers are searching for more ways to overcome the current antibiotic resistance challenge. In this study, we expressed and purified epsilon 34 phage tail spike protein (E34 TSP) from the E34 TSP gene, then assessed the ability of this bacteriophage protein in the killing of two CBD-resistant strains of Salmonella spp. We also assessed the ability of the tail spike protein to cause bacteria membrane disruption and dehydrogenase depletion. We observed that the combined treatment of CBD-resistant strains of Salmonella with CBD and E34 TSP showed poor killing ability, whereas the mono treatment with E34 TSP showed considerably higher killing efficiency. This study demonstrates that the inhibition of the bacteria by E34 TSP was due in part to membrane disruption and dehydrogenase inactivation by the protein. The results of this work provide an interesting background to highlight the crucial role phage proteins such as E34 TSP could play in pathogenic bacterial control.

Keywords: cannabidiol, resistance, Salmonella, antimicrobials, phages

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Authors: Zoe M. G. Skalkos, Sam N. Dowland, James U. Van Dyke, Camilla. M. Whittington

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Syngnathid fishes (seahorses, pipefishes, and seadragons) are unique because embryos develop on or in the male in a specialised brooding structure. Many seahorse species are endangered or vulnerable, while others are popular in the ornamental fish trade. Seahorses are capable of nutrient provisioning (patrotrophy) of lipids during pregnancy via their fully enclosed brood pouch. Protein is vital for gene regulation and tissue growth during embryogenesis. We tested the hypothesis that protein is paternally transported to developing embryos during pregnancy in the Australian Pot-bellied seahorse, Hippocampus abdominalis. We compared the dry masses and nitrogen content in recently fertilised H. abdominalis embryos and newborns. We calculated an updated patrotrophy index, 1.34, but without a significant difference in dry mass between the two developmental stages. There was, however, a significant increase in total protein content from recently fertilised embryos to neonates. This suggests paternal protein transport is essential for H. abdominalis embryogenesis because protein yolk reserves are depleted by embryonic metabolism, and supplementation is required. This study is the first to provide evidence for paternal protein transport during pregnancy in seahorses. It furthers our understanding of the paternal influence on embryonic development in male pregnancy and how a protein-deficient diet during pregnancy may limit the allocation of resources to embryos, reducing offspring fitness. This research contributes to a deeper understanding of the fundamental reproductive biology of seahorses, which can help improve conservation and farming production outcomes.

Keywords: brood pouch, embryonic provisioning, nitrogen, parentotrophy, paternal investment, reproduction

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Authors: Omar Alhamd, Peter A. Thomas, Trevor J. Greenhough, Annette K. Shrive

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The Pseudomonas syringae species complex includes many plant pathogenic strains with highly specific interactions with varied host species and cultivars. The rapid spread of these bacteria over the last ten years has become a cause for concern. Nanoparticles have previously shown promise in microbiological action. We have therefore investigated in vitro and in vivo the effects of different types and sizes of nanoparticles in order to provide quantitative information about their effect on the bacteria. The effects of several different nanoparticles against several bacteria strains were investigated. The effect of NP on bacterial growth was studied by measuring the optical density, biochemical and nutritional tests, and transmission electron microscopy (TEM) to determine the shape and size of NP. Our results indicate that their effects varied, with either a negative or a positive impact on both bacterial and plant growth. Additionally, the methods of exposure to nanoparticles have a crucial role in accumulation, translocation, growth response and bacterial growth. The results of our studies on the behaviour and effects of nanoparticles in model plants showed. Cerium oxide (CeO₂) and silver (Ag) NP showed significant antibacterial activity against several pathogenic bacteria. It was found that titanium nanoparticles (TiO₂) can have either a negative or a positive impact, according to concentration and size. It is also thought that environmental conditions can have a major influence on bacterial growth. Studies were therefore also carried out under some environmental stress conditions to test bacterial survival and to assess bacterial virulence. All results will be presented including information about the effects of different nanoparticles on Pseudomonas syringae bacteria.

Keywords: plant microbiome, nanoparticles, 16S rRNA gene sequencing, bacterial survival

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Authors: Ngonidzashe Mangoma, Caroline Marigold Sebata

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The release of cyanide-rich effluents from gold mines, and other industries, into the environment, is a global concern considering the well-known metabolic effects of cyanide in all forms of life. Such effluents need to be treated to remove cyanide, among other pollutants, before their disposal. This study aimed at investigating the possible use of bacteria in the biological removal of cyanide from cyanide-rich effluents. Firstly, cyanide-degrading bacteria were isolated from gold mine effluents and characterised. The isolates were then tested for their ability to grow in the presence of cyanide and their tolerance to increasing levels of the compound. To evaluate each isolate’s cyanide-degrading activities, isolates were grown in the simulated and actual effluent, and a titrimetric method was used to quantify residual cyanide over a number of days. Cyanide degradation efficiency (DE) was then calculated for each isolate. Identification of positive isolates involved 16S rRNA gene amplification and sequence analysis through BLAST. Six cyanide-utilising bacterial strains were isolated. Two of the isolates were identified as Klebsiella spp. while the other two were shown to be different strains of Clostridium bifermentans. All isolates showed normal growth in the presence of cyanide, with growth being inhibited at 700 mg/L cyanide and beyond. Cyanide degradation efficiency for all isolates in the simulated effluent ranged from 79% to 97%. All isolates were able to remove cyanide from actual gold mine effluent with very high DE values (90 – 94%) being recorded. Isolates obtained in this study were able to efficiently remove cyanide from both simulated and actual effluent. This observation clearly demonstrates the feasibility of the biological removal of cyanide from cyanide-rich gold mine effluents and should, therefore, motivate research towards the possible large-scale application of this technology.

Keywords: cyanide effluent, bioremediation, Clostridium bifermentans, Klebsiella spp, environment

Procedia PDF Downloads 146