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SAINSTIS
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  This is an Open Access Journal Open Access journal
ISSN (Print) 2089-0699
Published by UIN Maulana Malik Ibrahim Malang Homepage  [16 journals]
  • The Effect of Collagen and Fibrin Hydrogels Encapsulated with Adipose
           Tissue Mesenchymal Stem Cell-Derived Exosomes for Treatment of Spinal Cord
           Injury in a Rat Model

    • Abstract: Background: Mesenchymal stem cell (MSC) derived exosomes (MSC-DE) have been demonstrated to be potential candidates for the treatment of rat spinal cord injury (SCI).Objective: The effect of AD-MSC and AD-MSC-DE encapsulated into collagen and fibrin hydrogels on the treatment of SCI in a rat animal model was investigated for introducing a new effective SCI treatment methodMaterials and Methods: The AD-MSC-DE was isolated using ultra-centrifugation at 100,000×g for 120 min and characterized by different methods. Fibrin and collagen hydrogels were synthesized and then mixed with AD-MSC-DE suspension. the characterized AD-MSC-DE were encapsulated into collagen and fibrin hydrogels. eighteen adult male Wister rats were randomly classified into 3 equal groups (n=6): the control group (SCI rat without treatment), SCI rat treated with either AD-MSC-DE encapsulated in collagen hydrogel or encapsulated in fibrin hydrogel groups. the treatment approaches were evaluated using clinical, histological, and molecular assays.Results: The AD-MSC-DE encapsulated into fibrin and collagen groups showed better clinical function than the control group. The AD-MSC-DE encapsulated into fibrin and collagen also improved SCI-induced polio and leuko-myelomalacia and leads to higher expression of NF protein than the control group. In the AD-MSC-DE encapsulated into collagen and fibrin leads to up-regulation the mean levels of NEFL (23.82 and 24.33, respectively), eNOS (24.31 and 24.53, respectively), and CK19 mRNAs (24.23 and 23.98, respectively) compared to the control group.Conclusion: The AD-MSC-DE encapsulated within ECM-based hydrogel scaffolds such as collagen and fibrin can regenerate the injured nerve in SCI rats and reduce spinal cord lesion-induced central neuropathic pain.
       
  • Dexmedetomidine Suppresses Glutamate-Stimulated Neuronal Injury Via
           MicroRNA-433/Janus Kinase 2/Signal Transducer and Activator of
           Transcription 3 Pathway

    • Abstract: Background: Cerebral ischemia has been a hotpot in the prevention and treatment of cerebral ischemia. Dexmedetomidine (Dex) is a new type of highly selective α2 adrenergic receptor agonist with pharmacological properties.Objective: Quantitative studies have shown that Dex has a protective effect on glutamate (Glu)-induced neuronaldamage. however, its mechanism has not been fully elucidated. The purpose of this study was to explore the underlyingmolecular mechanism by which Dex ameliorates Glu-induced neuronal injury by regulating miR-433/JAK2/STAT3axis.Materials and Methods: A model of neuronal injury was constructed by Glu treatment and intervened with Dex.miRNA expression profiling assay was conducted to screen potential miRNAs affected by Dex. Cell viability, lactatedehydrogenase (LDH) release and apoptosis were detected by MTT assay, LDH kit, and TUNEL staining, respectively.Oxidative stress indicators were assessed by ELISA whereas mitochondrial membrane potential (MMP) was assessedby C11-BODIPY581/591 staining. The targeting relationship between the miR-433 and JAK2 was verified by dualluciferase reporter assay and gene expression was analyzed by quantitative PCR and Western blot.Results: Glu treatment decreased cell viability and MMP and promoted LDH release, apoptosis and oxidative damage.Glu-induced changes in neurons were reversed after Dex treatment through upregulating the miR-433 expression toblock the activation of JAK2/STAT3 pathway.Conclusions: Dex protects against Glu-induced neuronal injury by regulating miR-433/JAK2/STAT3 pathway, whichprovides new insights into the treatment of neuronal injury.
       
  • In vitro Gluten Degradation Using Recombinant Eurygaster Integriceps
           Prolyl Endoprotease: Implications for Celiac Disease

    • Abstract: Background: Celiac disease (CD) is a gluten-sensitive chronic autoimmune enteropathy. A strict life-long gluten-free dietis the only efficient and accepted treatment until now. However, maintaining a truly gluten-free status is both difficult and costly, often resulting in a social burden for the person. Moreover, 2 to 5 percent of patients fail to improve clinically and histologically upon elimination of dietary gluten. Therefore, novel therapeutic approaches, including gluten degrading enzymes, are an unmet need of celiac patients.Objectives: To evaluate the function of sunn pest prolyl endoprotease for gluten and gliadin hydrolysis in vitro.Materials and Methods: The spPEP was expressed as a recombinant protein in E. coli BL21 (DE3), and its catalyticactivity was assessed by SDS-PAGE and RP-HPLC analyses.Results: Production of a 100-kDa spPEP protein was confirmed by SDS-PAGE and western blot analysis. Also, wedemonstrate that spPEP efficiently degrades gluten and α-gliadin (30-40 kDa) in vitro under conditions similar to the GIand is resistant to pepsin and trypsin.Conclusion: The gathered data demonstrated that spPEP might be a novel candidate for Oral Enzymatic Therapy (OET)in CD and other gluten-related disorders.
       
  • MiRNA-145-5p Restrains Malignant Behaviors of Breast Cancer Cells Via
           Downregulating H2AFX Expression

    • Abstract: Background: Breast cancer is a prevalent tumor with high aggressiveness among female populations. MiRNA-145-5pplays an important role in multiple cancers.Materials and Methods: qRT-PCR detected miRNA-145-5p and histone protein family member X (H2AFX) mRNAexpression in breast cancer cells, and western blot determined the protein expression of H2AFX. After predicting the target genes via the bioinformatics methods, the targeting relationship between miRNA-145-5p and H2AFX was verified by dualluciferase, RIP, and RNA pull-down assays. The relationship between H2AFX and clinical indexes was also analyzed. Furthermore, the effects of miRNA-145-5p/H2AFX regulatory axis on breast cancer cell progression were determined by colony formation, wound healing, CCK-8, and Transwell assays.Results: The results suggested that miRNA-145-5p was markedly lowly-expressed in breast cancer tissue and cells,while H2AFX was upregulated, which had a positive correlation with T stages of breast cancer. Besides, overexpressedmiRNA-145-5p was found to remarkably suppress progression of breast cancer cells. As bioinformatic analysis predictedthat H2AFX was the potential target of miRNA-145-5p, the dual-luciferase assay was conducted, which demonstratedthat miRNA-145-5p negatively regulated the expression of H2AFX by targeting its 3’-UTR. The rescue experimentdemonstrated that overexpression of miRNA-145-5p could offset the promotion effects of oe-H2AFX on malignantprogression.Objective: Our study is aimed at exploring how miRNA-145-5p functions in breast cancer cells.Conclusion: Our findings confirmed that miRNA-145-5p hindered malignant progression of breast cancer by negativelyregulating H2AFX. MiRNA-145-5p/H2AFX axis may be a novel therapeutic target for breast cancer.
       
  • SOX2 Overlapping Transcript (SOX2-OT) Enhances the Lung Cancer Malignancy
           Through Interaction with miR-194-5p/SOX5 Axis

    • Abstract: Background: Lung cancer is one of the most common types of cancer and a leading cause of cancer-related deathsworldwide. Therefore, it is useful to know the biomarkers involved in the malignancy of lung cancer.Objectives: This study aimed to show that SOX2-OT as a long non-coding RNA (IncRNA) regulates gene expression viathe SOX2-OT/miR-194-5p/SOX5 axis molecular pathway in lung cancer.Materials and Methods: A549 cells transfected with siRNA-SOX2-OT and the expression of SOX2-OT and miR-194-5pgenes were analyzed by real-time PCR before and after transfection. In addition, the expression of the B-catenin, MMP9,phosphorylated and activated STAT3 (p-STAT3), SOX5, and VEGF proteins before and after transfection was investigatedby Western blotting.Results: After using siRNA-SOX2-OT, an increase in the expression of miR-194-5p and a decrease in the expression ofB-catenin, SOX5, p-STAT3 activated STAT3, VEGF, and MMP9 proteins was observed.Conclusions: According to the results of the present study, an increase in SOX2-OT in lung cancer seems to stimulate theexpression of beta-catenin, SOX5, MMP9, and VEGF thus support the malignancy of lung cancer cells.
       
  • PPI Identification of Immune-Related Biomarkers in Esophageal Cancer on
           the Basis of Gene Co-Expression Network

    • Abstract: Background: The mortality rate of esophageal cancer is on the continuous increase. Fortunately, with the development ofimmunotherapy, the prognosis and survival rate of patients with esophageal cancer have been improved gradually.Objective: Immune markers have a crucial part in immunotherapy. Therefore, it is of great meaning to delve further intoimmune-related biomarkers of esophageal cancer for better treatment.Materials and Methods: In this study, gene co-expression networks were established using weighted gene co-expressionnetwork analysis, thus forming gene modules with different clusters. The tumor immune microenvironment was assessedwith the ESTIMATE algorithm.Results: Analysis of the module Eigen gene -immune score trait indicated that the black module was markedly associatedwith immune score, with the top 80 genes regarding correlation ranking as the candidate hub gene set. Enrichment analysis revealed that genes within the black module were primarily enriched in tumor immune-related functions. To mine the hub genes that were closely connected with immunity, protein-protein interaction networks were constructed by STRING for genes within the black module, and genes with the interaction score top10 were retained. They were intersected with hub genes to finally obtain four hub genes: CCR5, LCP2, PTPRC and TYROBP. The samples were divided into highand low-expression groups by the median expression of hub gene, and survival analysis was performed in combination with clinical information. The results revealed that the high-expression groups of genes LCP2 and PTPRC had a poor prognosis. TIMER immune cell infiltration analysis revealed that the expression levels of the 4 hub genes were positively correlated with immune cell infiltration and negatively correlated with tumor purity. In addition, these 4 hub genes were correlated with the expression of immune checkpoint genes CTLA-4 and PDCD1 positively. Gene set enrichment analysis enrichment analysis demonstrated that there were differences in tumor immunity and cancer-related pathways between high and low expression of 4 hub genes.Conclusion: Altogether, we identified four biomarkers that may have connection with tumor immunity, and speculatedthat these genes may influence patient prognosis by affecting pathways related to esophageal cancer immunity. This study will pave the way for the research of immune mechanisms of esophageal cancer and the analysis of patient’s prognosis.
       
  • Data-Mining of Barley to Identify Salt Stress Hub Genes, Gene Expression
           Analysis and Recombinant Plasmid Construction

    • Abstract: Background: Salinity is one of the major abiotic stresses that limit the production and yields of agricultural cropsworldwide.Objectives: In order to identify key barley genes under salinity stress, the available metadata were examined by twomethods of Cytoscape and R software. Next, the hub expression of the selected gene was evaluated under different salinity stress treatments and finally, this gene was cloned into cloning and expression vector and recombinant plasmid was made.Materials and Methods: In this study, we extracted salinity stress tolerant genes from several kinds of literature and alsomicroarray data related to barley under salinity conditions from various datasets. The list of genes related to literatureanalyzed using string and Cytoscape. The genes from the datasets were first filtered and then the hub genes were identified by Cytoscape and R methods. Next, these hub genes were analyzed for the promoter.Results: Ten hub genes were selected and their promoters were analyzed, the cis-element of which was often cisacting regulatory element involved in the methyl jasmonate -responsiveness, common cis-acting element in promoterand enhancer regions and MYBHv1 binding site. Finally, the sedoheptulose-1,7-bisphosp gene (SBPase), which had thehighest interaction in both gene lists and both types of gene networks, was selected as hub gene. Next, the expression ofSBPase gene was examined in two variety of Youssef variety (salt tolerant) and Fajr variety (salt sensitive) under salinitystress (NaCl 100mM) at 0 (control), 3, 6, 12 and 24 hours after stress. The results showed that the expression of thisgene increased with increasing the duration of stress in both varieties. Comparison of the two varieties showed that theexpression of SBPase gene in the tolerant genotype was twice as high as sensitive. Finally, SBPase gene as a key gene forsalinity stress was cloned in both cloning (pTG19) and expression (pBI121) vectors.Conclusions: According to our results, SBPase gene increased growth and photosynthesis in barley under various abioticstresses, therefore, over-expression of this gene in barley is recommended to produce plants resistant to abiotic stresses.
       
  • Anti-Inflammatory Properties of Extract of the Hairy Root of Native
           Portulaca Oleracea L. and Its Effect on Some Inflammatory Genes Expression
           in Rat Microglial Cells

    • Abstract: Background: Most herbs play significant roles in the treatment of various diseases. Because dopamine functions in theanti-inflammatory process and the presence of this substance in Portulaca Oleracea L. native plant, investigating thisplant’s anti-inflammatory properties in treating neurological diseases is interesting.Objectives: The objective of this study was to estimate the NO production and the expression level of inflammatory genesin lipopolysaccharide (LPS)-treated microglial cells affected by P. oleracea L. extraction.Materials and Methods: P. oleracea L. hairy root extract was isolated, and the primary microglial cell of the rat wasisolated from glial cells and confirmed by immunocytochemistry analysis. Microglial cells were pretreated with differentconcentrations of P. oleracea L. extract and then treated with 1 μg.mL-1 LPS. The control group did not receive anytreatment. The NO level in culture supernatants was measured by the Griess method. The mRNA expression levels ofiNOS (inducible Nitric oxide synthase) and TNF-α (tumor necrosis factor-alpha) in LPS-treated microglial cells wereevaluated using Real-Time PCR.Results: The present study determined that 0.1 mg. mL-1 of the P. oleracea L. extract decreased the NO production in ratmicroglial cells. Different concentrations of the P. oleracea L. extract had no prominent effects on LPS-treated cell viability. The results of real-time PCR indicated that P. oleracea L extracts suppressed the mRNA expression levels of iNOS and TNF-α in LPS-treated cells. MTT assay determined that P. oleracea L. extract was not cytotoxic, and the anti-inflammatory P. oleracea L. extract effects observed were not because of cell death.Conclusion: P. oleracea L. extract might be helpful as an anti-inflammatory agent in treating inflammatory diseases.
       
  • Cloning, Prokaryotic Expression and Functional Characterization of NifH
           Gene from the Associative Nitrogen-Fixing Bacteria Klebsiella Variicola
           DX120E

    • Abstract: Background: Biological nitrogen fixation (BNF) is a unique mechanism in which microorganisms utilize the nitrogenaseenzyme to catalyze the conversion of atmospheric nitrogen (N2) to ammonia (NH3). Fe protein, encoded by the nifHgene, is an essential component of the nitrogenase in Klebsiella variicola DX120E. However, the function of this gene inregulating nitrogen fixing activity is still unclear.Objectives: The objective of this study was to reveal the function of nifH gene in associative nitrogen-fixing bacteriaKlebsiella variicola DX120E and micro-sugarcane system by immunoassay and gene editing.Materials and Methods: In the current investigation, the nifH gene was cloned in a pET-30a (+) vector and expressedin Escherichia coli. The NifH protein was purified and used to immunize rabbit, and then the serum was collected andpurified to obtain rabbit anti-NifH polyclonal antibodies. The CRISPR-Cas9 system was applied to produce nifH mutantstrains, and the nitrogen-fixing enzyme activity, gene, and protein expression were analyzed.Results: Both in vitro and in vivo NifH proteins were detected by Western blotting, which were 43 and 32 kDa respectively. The expression of nifD and nifK genes was decreased, and nitrogenase activity was reduced in the nifH mutant strain.Conclusion: The nifH gene mutant weakened the nitrogenase activity by regulating the expression of Fe protein, whichsuggests a potential strategy to study the nitrogen fixation-related genes and the interactions between endophytic nitrogenfixing bacteria and sugarcane.
       
  • Optimization of Affinity Chromatography Based on Sepharose 4Bchitin for
           Rapid Purification of Urtica dioica Agglutinin

    • Abstract: Background: Today, numerous antimicrobial and anticancer properties have been reported for plant lectins due to theirability to bind to carbohydrates. The Urtica dioica agglutinin (UDA lectin) is a monomeric, small, and low molecularweight glycoprotein. It has attracted the attention of many researchers for identification, treatment, and other clinicalpurposes.Objectives: The aim of this study is the optimization of the chitin affinity chromatography based on Sepharose 4B (CNBractivated Sepharose 4B) for the rapid purification of UDA lectin from Urtica dioica rhizome.Materials and Methods: The chitin ligands were dissolved in 40% Trichloroacetic acid and attached to Sepharose4B according to the Amersham-Biosciences instructions. The attachment of the ligand to the Sepharose 4B beads wasinvestigated by Fourier transform infrared (FTIR) spectroscopy. An acidic crude extract of nettle rhizome passes fromchromatographic columns in two sizes with dimensions: 24 x 0.51 cm and 8.44 x 0.86 cm. Quantity and quality of purifiedlectin were calculated by the Bradford microplate method: SDS-PAGE gel electrophoresis and human erythrocyte cell(RBC) hemagglutination, respectively.Results: The analysis of FTIR spectrograms showed that major changes were observed in the fingerprint regions. Besides,due to the dissolution of Sepharose 4B and chitin in the aqueous phase, this difference was not significant in the Imineand Nitrile regions. On the other hand, the comparative results of purification chromatograms showed that increasingthe column length causes a smaller half-width and increases the length of the purified peak. Also, it leads to high-qualitypurified UDA lectin, with a molecular weight of almost 12.5 kDa in gel electrophoresis. Hemagglutination activityon trypsinized red blood cells was displayed, and agglutination of purified UDA lectin started at least at 300 μg.mL-1concentration.Conclusion: According to our findings, we suggested that dissolving chitin in the polar solvent of Trichloroacetic acid,using Sepharose 4B as the beads of a matrix, and increasing the column length might lead to a decrease in the half-widthof the peak. These can increase the purity and concentration of purified UDA lectin, and speed up the purification process. These findings could be used by researchers to accelerate the purification of UDA lectin in other studies, dealing with drug delivery systems, ELISA techniques, and cell growth.
       
  • A Comprehensive Review of the Key Characteristics of the Genus Mentha,
           Natural Compounds and Biotechnological Approaches for the Production of
           Secondary Metabolites

    • Abstract: Context: The genus Mentha is one of the most aromatic and well-known members of the Lamiaceae family. A wide range of bioactive compounds has been reported in mints. Regarding the high economic importance of Mentha plants due to the presence of valuable metabolites, the demand for their products is growing exponentially. Therefore, to supply such demand, new strategies should be adopted to improve the yield and medicinal quality of the products.Evidence acquisition: The current review is written based on scientific literature obtained from online databases, including Google Scholar, PubMed, Scopus, and Web of Science regarding the characteristic features of some species of the genus Mentha, their distribution and cultivation, main uses and benefits, phytochemical composition, biotechnological approaches for the production of secondary metabolites, and strategies for enhanced production of mints secondary metabolites.Results & Conclusions: In this article, we offer an overview of the key characteristics, natural compounds, biological properties, and medicinal uses of the genus Mentha. Current research describes biotechnological techniques such as in vitro culture methods for the production of high-value secondary metabolites. This review also highlights the strategies such as elicitation, genetic, and metabolic engineering to improve the secondary compounds production level in mint plants. Overall, it can be concluded that identifying the biosynthetic pathways, leading to the accumulation of pharmaceutically important bioactive compounds, has paved the way for developing highly productive mint plants with improved phytochemical profiles.
       
  • Introns and their therapeutic applications in biomedical researches: A
           Review

    • Abstract: Context: Although for a long time, it was thought that intervening sequences (introns) were junk DNA without any function, their critical roles and the underlying molecular mechanisms in genome regulation have only recently come to light. Introns not only carry information for splicing, but they also play many supportive roles in gene regulation at different levels. They are supposed to function as useful tools in various biological processes, particularly in the diagnosis and treatment of diseases. Introns can contribute to numerous biological processes, including gene silencing, gene imprinting, transcription, mRNA metabolism, mRNA nuclear export, mRNA localization, mRNA surveillance, RNA editing, NMD, translation, protein stability, ribosome biogenesis, cell growth, embryonic development, apoptosis, molecular evolution, genome expansion, and proteome diversity through various mechanisms.Evidence acquisition: In order to fulfill the objectives of this study, the following databases were searched: Medline, Scopus, Web of Science, EBSCO, Open Access Journals, and Google Scholar. Only articles published in English were included.Results & Conclusions: The intervening sequences of eukaryotic genes have critical functions in genome regulation, as well as in molecular evolution. Here, we summarize recent advances in our understanding of how introns influence genome regulation, as well as their effects on molecular evolution. Moreover, therapeutic strategies based on intron sequences are discussed. According to the obtained results, a thorough understanding of intron functional mechanisms could lead to new opportunities in disease diagnosis and therapies, as well as in biotechnology applications.
       
  • CRISPR/Cas9-Induced Fam83h knock-out Leads to Impaired Wnt/β-Catenin
           Pathway and Altered Expression of Tooth Mineralization Genes in Mice

    • Abstract: Background: Dental enamel formation is a complex process that is regulated by various genes. One such gene, Family With Sequence Similarity 83 Member H (Fam83h), has been identified as an essential factor for dental enamel formation. Additionally, Fam83h has been found to be potentially linked to the Wnt/β-catenin pathway.Objectives: This study aimed to investigate the effects of the Fam83h knockout gene on mineralization and formation of teeth, along with mediators of the Wnt/β-catenin pathway as a development aspect in mice.Materials and Methods: To confirm the Fam83h-KnockOut mice, both Sanger sequencing and Western blot methods were used. then used qPCR to measure the expression levels of genes related to tooth mineralization and formation of dental root, including Fam20a, Dspp, Dmp1, Enam, Ambn, Sppl2a, Mmp20, and Wnt/β-catenin pathway mediators, in both the Fam83h-Knockout and wild-type mice at 5, 11 and 18 days of age. also the expression level of Fgf10 and mediators of the Wnt/β-catenin pathway was measured in the skin of both Knockout and wild-type mice using qPCR. A histological assessment was then performed to further investigate the results.Results: A significant reduction in the expression levels of Ambn, Mmp20, Dspp, and Fgf10 in the dental root of Fam83h-Knockout mice compared to their wild-type counterparts was demonstrated by our results, indicating potential disruptions in tooth development. Significant down-regulation of CK1a, CK1e, and β-catenin in the dental root of Fam83h-Knockout mice was associated with a reduction in mineralization and formation-related gene. Additionally, the skin analysis of Fam83h-Knockout mice revealed reduced levels of Fgf10, CK1a, CK1e, and β-catenin. Further histological assessment confirmed that the concurrent reduction of Fgf10 expression level and Wnt/β-catenin genes were associated with alterations in hair follicle maturation.Conclusions: The concurrent reduction in the expression level of both Wnt/β-catenin mediators and mineralization-related genes, resulting in the disruption of dental mineralization and formation, was caused by the deficiency of Fam83h. Our findings suggest a cumulative effect and multi-factorial interplay between Fam83h, Wnt/Β-Catenin signaling, and dental mineralization-related genes subsequently, during the dental formation process.
       
  • Research on Circular RNA Expression Profiles in the Photoaging Mouse Model

    • Abstract: Background: Nude mouse has been widely used to study photoaging induced by long-term chronic UV exposure. Circular RNAs (circRNAs) have been previously identified in several diseases. However, the roles of circRNAs in photoaging and potential regulatory mechanisms remain unclear.Objectives: To identify specific circRNAs differentially expressed in photoaged skin and investigate their potential role in aging.Materials and Methods: In this study, we screened out the microarray data to profile the expression of circRNAs. The circRNAs were analyzed by Gene Ontology(GO) and Kyoto Encyclopedia of Genes and Genomes(KEGG)biological pathway. Results: 36 circRNAs were identified to be differentially expressed between the UV group and control group (fold change > 1.5; P < 0.05), including 6 upregulated and 30 downregulated circRNAs. GO and KEGG biological pathway analyses indicated that the changes in circRNAs were associated with cancer, inflammation, oxidative stress, and metabolism.Conclusions: This present study revealed a circRNAs expression profiling in vivo. These findings not only provide a new possibility to prevent the occurrence of photoaging but also have therapeutic values for photoaging and associated skin diseases.
       
  • Gene Networks Analysis of Salmonella Typhimurium Reveals New Insights on
           Key Genes Involved in Response to Low Water Activity

    • Abstract: Background: When Salmonella enterica serovar Typhimurium, a foodborne bacterium, is exposed to osmotic stress, cellular adaptations increase virulence severity and cellular survival.Objectives: The aim of the gene network analysis of S. Typhimurium was to provide insights into the various interactions between the genes involved in cellular survival under low water activity (aw).Materials and Methods: We performed a gene network analysis to identify the gene clusters and hub genes of S. Typhimurium using Cytoscape in three food samples subjected to aw stress after 72 hours.Results: The identified hub genes of S. Typhimurium belonged to down-regulated genes and were related to translation, transcription, and ribosome structure in the food samples. The rpsB and Tig were identified as the most important of the hub genes. Enrichment analysis of the hub genes also revealed the importance of translation and cellular protein metabolic processes. Moreover, the biological process associated with organonitrogen metabolism in milk chocolate was identified. According to the KEGG pathway results of gene cluster analysis, cellular responses to stress were associated with RNA polymerase, ribosome, and oxidative phosphorylation. Genes encoding RNA polymerase activity, including rpoA, rpoB, and rpoZ, were also significantly identified in the KEGG pathways. The identified motifs of hub DEGs included EXPREG_00000850, EXPREG_00000b00, EXPREG_000008e0, and EXPREG_00000850.Conclusion: Based on the results of the gene network analysis, the identified hub genes may contribute to adaptation to food compositions and be responsible for the development of low water stress tolerance in Salmonella. Among the food samples, the milk chocolate matrix leads to more adaptation pathways for S. Typhimurium survival, as more hub genes were down-regulated and more motifs were detected. The identified motifs were involved in carbohydrate metabolism, carbohydrate transport, electron transfer, and oxygen transfer.
       
  • Green Synthesis of Polylactic acid/Fe3O4@β-Cyclodextrin Nanofibrous
           Nanocomposite Loaded with Ferulago Angulata Extract as a Novel
           Nano-biosorbent: Evaluation of Diazinon Removal and Antibacterial Activity
           

    • Abstract: Background: Organophosphate pesticides are one of the most extensively applied insecticides in agriculture. These insecticides persist in the environs and thereby cause severe pollution problems. Iron oxide polymer nanocomposites are wastewater remediation agents synthesized by various methods. When compared to chemical processes, green synthesis using plant extract is thought to be more cost- and environmentally-friendly. Objectives: This study aimed to evaluate the green synthesis of Fe3O4@β-Cyclodextrin (Fe3O4@β-CD) nanoparticles using Ferulago angulata (F. angulata) methanol extract. These nanoparticles are loaded on polylactic acid (PLA) nanofibrous nanocomposite along with Ferulago angulata extract (2, 4, and, 6wt %) to produce PLA/Fe3O4@β-CD/F. angulata extract nanofibrous nanocomposite as a new nano biosorbent. Furthermore, the antibacterial properties of this compound and its activity in diazinon removal have been evaluated.Materials and Methods: Fe3O4@β-CD nanoparticles synthesis was performed via co-precipitation method using FeCl3.6H2O and FeCl2.4H2O and β-cyclodextrin, and Ferulago angulata extract. Then polylactic acid/ Fe3O4@β-CD / F. angulate.extract nanofibrous nanocomposite was prepared by the electrospinning method. Energy-dispersive X-ray spectroscopy (EDS), X-ray diffraction analysis (XRD), vibrating sample magnetometer (VSM), and Fourier transform infrared spectroscopy (FTIR) were used to analyze the structure of the nanocomposite. The antibacterial activity of this nanocomposite against several fish and human bacterial pathogens, as well as its effectiveness in diazinon elimination, have been evaluated in the sections that follow. The nanocomposite structure demonstrated that Fe3O4 nanoparticles were produced and put into the polylactic acid matrix with an average particle size of 40 nm. Furthermore, the results showed that this nanocomposite exhibited removal efficiency of diazinon over 80% after 120 minutes under pH=7 and 2.5 gr.L-1 nanocomposite concentration. Also, this structure showed above 70% antibacterial ability against Bacillus cereus, Staphylococcus epidermidis and 60% antibacterial ability against Streptococcus iniae and Yersinia ruckeri.Conclusion: Fe3O4 nanocomposite synthesis may be accomplished in a delicate and efficient manner by using Ferulago angulata to produce Fe3O4@-CD nanoparticles. The stability of the nanoparticles was enhanced by combining Ferulago angulata extract with polylactic acid nanofibers to create an antibacterial homocomposition nanocomposite. This device may be used to remove and disinfect diazinon from aqueous media in an environmentally friendly manner.
       
  • Enhancement of Essential Oil Production and Expression of Some Menthol
           Biosynthesis-Related Genes in Mentha piperita Using Cyanobacteria

    • Abstract: Background: Mentha piperita L. is one of the most important aromatic crops and is cultivated worldwide for essential oils (EOs). Objectives: The aim of the present study was to investigate the potential of two cyanobacteria, Anabaena vaginicola ISB42 and Nostoc spongiaeforme var. tenue ISB65, as biological-elicitors to improve the growth and essential oil production of M. piperita.Materials and Methods: In this experiment, inoculation of M. piperita with cyanobacteria was performed by adding 1% cyanobacterial suspension to the soil of treated pots on the first time of planting and every 20 days thereafter. The experiment was performed in a randomized complete block design in an experimental greenhouse condition. After 90 days planting, the vegetative growth factors, the content of photosynthetic pigments, as well as the quantity and quality of EOs of treated and control plants were evaluated. Also, quantitative changes in the expression of some menthol biosynthesis-related genes were investigated.Results: Cyanobacterial application led to significant increases in M. piperita growth indices including root and shoot biomass, leaf number, leaf area, node number and ramification, as well as photosynthetic pigments content. The statistical analysis showed a 41-75 % increase in some of these growth indices, especially in Nostoc-treated plants. A. vaginicola and N. spongiaeforme var. tenue inoculation led to a 13% and 25% increase in the EOs content of M. piperita, respectively. The EOs components were also affected by cyanobacterial treatments. According to the statistical analysis, Nostoc-treated plants showed the highest amount of (-)-menthone and (-)-limonene, with a 2.36 and 1.87-fold increase compared to the control. A. vaginicola and N. spongiaeforme var. tenue inoculation also led to 40% and 98% increase in transcript level of (-)-limonene synthase gene, respectively. The expression of the (-)-menthone reductase gene, was also increased by 65% and 55% in response to A. vaginicola and N. spongiaeforme var. tenue application, respectively. Conclusions: Our data demonstrated that in addition to growth enhancement, these two heterocystous cyanobacteria improved the quantity and quality of EOs by up-regulating the key genes involved in the menthol biosynthetic pathway. Based on our results, these cyanobacteria can be considered valuable candidates in the formulation of low-cost and environmentally friendly biofertilizers in sustainable peppermint production.
       
  • Molecular identification of Juglans regia endophyte LTL-G3, its antifungal
           potential and bioactive substances

    • Abstract: Background: Endophyte is one of the potential biocontrol agents for inhibiting plant pathogens. However, the mechanisms and characteristics involved in the inhibition of different phytopathogenic fungi by endophytes, especially walnut endophytes, are still largely unknown.Objectives: The present study aimed to identify the walnut endophytic fungus LTL-G3 from a genetic point of view, assess the strain's antifungal activity, and determine the bioactivities of the substances it produces against plant pathogens.Materials and Methods: The homologous sequence of strain LTL-G3 was examined, and typical strains of the Trichoderma virens group were used to build NJ phylogenetic trees and analyze the taxonomic position of the strain. The biocontrol agent's antagonistic potential for many plant pathogenic fungi. By using silica gel G chromatography, the active components of the strain were separated and purified. The active components were identified using GC-MS and NMR.Results: The strain LTL-G3 was identified as Trichoderma virens. Its fermentation and secondary metabolite extracts had a broad spectrum and strong inhibitory effect on the spread of six plant pathogens (Botrytis cinerea, Fusarium graminearum, Gloeosporium fructigenum, Phytophthora capsici, Rhizoctonia solani, and Valsa mali) evaluated, Of which, its inhibition rate against Valsa mali reached 76.6% (fermentation extract) and 100% (ethyl acetate and n-butanol extracts). On silica gel G chromatography, bioactive compounds were divided into 6 fractions and 7 sub-fractions. Fr.2-2 was the sub-fraction that showed the greatest inhibitory against V. mali, as an inhibition percentage of 89.36% in 1 mg mL-1. Fifteen key inhibitory chemicals identified using GC-MS. By examining the NMR data, the chemical make-up of the precipitated white solid was identified. The inhibition rate against V. mali increased by over 95% at a dosage of 1 mg mL-1, indicating a significant linear association between compound A and that rate.Conclusions: The strain LTL-G3 can be applied as an efficient biological control agent against V. mali, and its highly inhibitive secondary metabolites provide the mechanism for this action.
       
  • Identification of a New Compound (4-Fluoro-2-Trifluoromethyl Imidazole)
           Extracted From a New Halophilic Bacillus aquimaris Strain Persiangulf TA2
           Isolated From the Northern Persian Gulf With Broad-Spectrum Antimicrobial
           Effect

    • Abstract: Background: The unique ecosystem of the Persian Gulf has made it a rich source of natural antimicrobial compounds produced by various microorganisms, especially bacteria, which can be used in the treatment of infectious diseases, especially those of drug-resistant microbes.Objectives: This study aimed to identify antimicrobial compounds in the bacteria isolated from the northern region of the Persian Gulf in Abadan (Choobdeh port), Iran, for the first time. Materials and Methods: Sampling was performed in the fall on November 15, 2019, from 10 different stations (water and sediment samples). The secondary metabolites of all isolates were extracted, and their antimicrobial effects were investigated. 16S ribosomal ribonucleic acid sequencing was used for the identification of the strains that showed the best inhibition against selected pathogens, and growth conditions were optimized for them. A fermentation medium in a volume of 5000 ml was prepared to produce the antimicrobial compound by the superior strain. The extracted antimicrobial compounds were identified using the gas chromatography-mass spectrometry technique. Minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) were determined for the superior strain. The effects of salinity, pH, and temperature on the production of antimicrobial compounds were determined by measuring the inhibitory region (mm) of methicillin-resistant Staphylococcus aureus (MRSA).Results: Four new strains with antimicrobial properties (i.e., Halomonas sp. strain Persiangulf TA1, Bacillus aquimaris strain Persiangulf TA2, Salinicoccus roseus strain Persiangulf TA4, and Exiguobacterium profundum strain Persiangulf TA9) were identified. The optimum growth temperatures were determined at 37-30, 37, and 40 °C for TA1 and TA2, TA4, and TA9 strains, respectively. The optimum pH values for the four strains were 7, 6-7, 7.5, and 6.5-7.5, respectively. The ethyl acetate extract of strain Persiangulf TA2 showed extensive antimicrobial activity against human pathogens (75%) and MRSA. The optimal salt concentrations for the four strains were 15%, 2.5-5%, 7.5%, and 5%, respectively. The most abundant compound identified in TA2 extract was the new compound 4-fluoro-2-trifluoromethyl imidazole. The MBC and MIC for the ethyl acetate extract of strain TA2 were 20 and 5 mg/ml (Staphylococcus aureus), 40 and 20 mg/ml (MRSA, Escherichia coli, and Enterococcus faecalis), 40 and 10 mg/ml (Acinetobacter baumannii), and 80 and 40 mg/ml (Staphylococcus epidermidis, Shigella sp., Bacillus cereus, and Klebsiella pneumoniae), respectively. The optimal conditions for antibiotic production by TA2 strain were 5% salt concentration, pH of 7, and temperature of 35 °C. Conclusion: Newly detected natural compounds in TA2 strain due to superior antimicrobial activity even against MRSA strain can be clinically valuable in pharmacy and treatment.
       
 
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