فصلنامه تازه های بیوتکنولوژی سلولی مولکولی
پیاپی 48 (پاییز 1401)

 Covid-19 disease with acute respiratory symptoms appeared in 2019. The disease was associated with fever, cough, and severe respiratory distress. The causative agent of the disease belonged to the beta-coronavirus family and was named SARS-CoV-2. The outbreak of Covid-19 and the creating of a global epidemic led to many attempts to understand the structure and function of the virus. The spike protein is a relatively heavy glycosylated protein consisting of three identical subunits that appear as a homo-trimmer on the virus's surface. After binding to the surface cell receptor (angiotensin-converting enzyme), the protein conformational changes and eventually causes the virus to enter the host cell. This process is necessary for viruses to enter human cells and infect them. Therefore, investigating the structural details of the spike protein through molecular dynamics simulation can play an influential role in understanding how the subunits connect and identifying the hotspot amino acids.

 First, the tertiary structure of spike proteins related to SARS-CoV-2, bat-SL-CoVZC21 (Bat Coronavirus), and PCoV_GX-P4L (Pangolin coronavirus) was constructed through modeling homology. After evaluation and validation, these structures were simulated using NAMD software. The structural properties of spike proteins were evaluated in the level of 3D structure, measurement of the interface area between monomers, thermodynamic parameters, trimmer structures' stability, and hotspot amino acids' identification in making connections between subunits.

 Evaluation of the trimeric structure of spike protein showed that this protein in SARS-CoV-2 has the highest ΔGdiss value (113.2 kcal/mol), which indicates the stability of the oligomeric protein structure among other spike proteins. The lowest ΔGdiss (100.6 kcal/mol) was obtained for the spike protein of PCoV_GX-P4L. The interface area between the subunits was almost the same in all spike proteins. Y369, D405, Y707 in subunit A, Y369, D574, Y707, Y837, D985 in subunit B, and Y707 in subunit C were identified as red hotspot amino acids for SARS-CoV-2 spike protein.

 The study of the structure of virus surface proteins at the atomic scale, and the recognition of hotspot residues that play an essential role in establishing monomeric connections, has created a window to further recognition and function of the virus. Thus, it can provide new information about neutralizing the virus.

The bacterium Streptococcus mutans is the main cause of dental caries. The formation of biofilms of this bacterium depends on the presence of the enzyme glucosyltransferase, which is encoded by the gtf gene. The aim of this study was evaluated the effect of polysaccharide nanocomposites of Chitosan(Cs)/Carboxymethyl Starch(CMS) /CQD.Mg /Trachyspermum on gtf gene expression in Streptococcus mutans using Real Time-PCR method in the formation of dental biofilm.

After preparation of Chitosan(Cs)/Carboxymethyl Starch(CMS) /CQD.Mg /Trachyspermum nanocomposite, its antimicrobial effect on Streptococcus mutans ATCC 35668 was investigated by microdilution method. The morphology and structural properties of the nanocomposite were investigated by FTIR and SEM analysis. The expression of gtfD, gtfC, gtfB genes was examined by Real Time-PCR technique. Data analysis was performed by SPSS22 software and the means were compared based on ANOVA at the level of 0.05.

The particle size of Chitosan(Cs)/Carboxymethyl Starch(CMS)/CQD.Mg/Trachyspermum nanocomposite was obtained in the range of 36-40 nm. The results of FTIR spectrum indicate the formation of a bond between carbon-quantum materials-magnesium and Trachyspermum essential oil in the nanocomposite. The antimicrobial effect of Cs/ CMS/CQD.Mg/Trachyspermum (0.168 mg/ ml) was obtained. RT-PCR results showed that gtfC gene expression was significantly reduced (P=0.049) in nanocomposite-treated cells, indicating the strong effect of Cs/CMS/CQD.Mg/Trachyspermum nanocomposite suppresses gtfC gene and reduces biofilm formation. No significant relationship was observed in the expression of gtfB and gtfD genes in nanocomposite-treated cells compared to the control group (P = 0.067, P = 0.187).

According to the results, the expression of gftC gene in cells treated with nanocomposite containing Trachyspermum has decreased and nanocomposite has a strong effect in suppressing genes and reducing the formation of microbial biofilm of teeth.
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Infections caused by Staphylococcus aureus are on the rise due to the spread of resistant strains, an increase in immunocompromised patients, and the overuse of medical equipment. The bacterium is also an opportunistic pathogen that provides conditions for invading the host by producing and secreting various toxins, including various enterotoxins.The aim of this study was to evaluate the antibiotic resistance and frequency of sea gene in Staphylococcus aureus isolated from clinical sources in Karaj.
 

This descriptive-observational study was performed on 100 Staphylococcus aureus isolates collected from clinical sources in Karaj. Collected samples were transferred to the laboratory under appropriate conditions by culture, microscopic and biochemical methods and then the susceptibility of isolated bacteria to various antibiotics was measured by disk diffusion agar method according to CLSI (2019) instructions and finally to test production ability. Enterotoxin A, the presence of sea gene was assessed by PCR

Based on the results of biochemical tests, 100 isolates of Staphylococcus aureus were identified. According to the antibiogram test, these bacteria had the highest resistance to penicillin with 92% and ceftazidime with 82% and also 100% of the isolates were sensitive to vancomycin. The results of PCR reaction also showed that out of 100 isolates, 79% contained sea genes.

Considering the importance and role of Staphylococcus aureus in the pathogenesis and development of nosocomial infections, drug resistance and the presence and expression of enterotoxin genes in clinical specimens of this bacterium should be considered in disease control management.

The use of creatininase is an enzymatic method for measuring amount of creatine and creatinine in body fluids to assess kidney health. The aim of this study was to investigate the effect of gold nanoparticles on the activity and stability of creatininase enzyme for use in creatine and creatinine assay kits.

In this study, E. coli BL21 containing pET28a amplification vector containing creatininase gene was first cultured in liquid LB medium and then induced at 28°C. After sonication, purification of enzyme was performed by affinity chromatography.  After confirmation of expression by SDS-PAGE, The expressed enzyme concentration was measured by Bradford method. Then the activity of the enzyme was examined and compared by colorimetric method in the presence and absence of gold nanoparticles. Finally, the effect of gold nanoparticles on the structure of creatininase enzyme was investigated by fluorescence spectroscopy.

Creatininase activity in the absence of gold nanoparticles 14.6 U/ml and in the presence of gold nanoparticles with concentrations of 1.97, 3.94, 7.88, 11.82, 15.76, 19.7 and 39.4 ngr/l 14/1, 12.4, 11.2, 10.1, 8.4, 7.5, 4.7 U/ml were calculated which indicates a decrease in creatininase activity in the presence of gold nanoparticles. On the other hand, fluorescence spectroscopy of creatininase in the presence and absence of gold nanoparticles at the excitation wavelength of 295 nm showed a decrease in fluorescence intensity in the presence of gold nanoparticles.

Gold nanoparticles reduced the activity of the creatininase enzyme. Gold nanoparticles also reduced the fluorescence intensity of creatininase enzyme, which could indicate a change in the microenvironment of the amino acid tryptophan in the structure of the creatininase enzyme, so using gold nanoparticles is not a good option for stable structure and activity of creatininase for use in commercial kits.

Sargassum ilicifolium is a multicellular brown algae that contains beta - carotene and fucoxanthin pigments, soluble acids of immune stimulants such as phycocyanin, polysaccharide, iron, zinc and essential amino acids, which promote safety. In this study, the effect of Sargassum iliisifolium algae extract on linear wound healing percentage and its effect on TGF β 1 and VEGFA gene expression in rats were investigated.

This research is an experimental study that was performed in the laboratory of Parand University of Medical Sciences. For this study, 30 male Wistar rats weighing 180 gr were used and 2 cm deep linear dermis was created from the neck to the waist of rats. Mice were divided into 3 groups of ten: The control group,The group received alpha ointment as a positive control drug and The group algae extract of Sargassum iliisifolium. The groups were examined for both wound size and expression of VEGFA and TGFB1 genes .

The alpha group had a 10.38 fold increase in TGFΒ1 gene expression compared to the control group, with a probability value of P <0.001. Compared with the algal group and the control group, it had a 5.39 increase in gene expression with a probability equal to P <0.001 . Also in assessing VGEFA gene expression :the alpha group significantly increased from the control group by 1.86 times the increase in gene expression with a probability value of p <0.044 , the sargassum algae group compared to the control group by 1.70 There is significant difference between the increase in expression and the probability value of p <0.040 . In terms of the percentage of wound healing, it shows that: the control group, reduced the wound length from 2 cm to 0.64 cm, the alpha group reduced the wound length from 2 cm to 0.5 cm and sargassum algae, the amount of wound has been reduced from 2 cm to 0.3 cm

Sargassum iliisifolium algae extract due to its antibacterial, anti -inflammatory, antioxidant and antimicrobial properties and due to the presence of alginate in it biometrically heals wounds. Available in the market, however, sargassum algae extract make a significant difference in increasing the expression of TGFβ1 and VEFGA gene .sargasum algae extract may use as a wound healer to heal skin damages

Assessing the genetic diversity of natural and protected germplasm is one of the most strategic global activities and as the national capital of each country. Over the years, various mutations and crosses have caused the accumulation of some useful genes in this germplasm and can be used as genetic resources in critical situations. Evaluation of diversity based on morphological and phenotypic traits is not stable due to the influence of the environment and the interaction of genotype in the environment. Therefore, the use of DNA-based markers that have high reproducibility. Codon-based DNA amplifier-based markers have recently been used to translate SCoT in plants due to heritability and reproducibility. The aim of this study was to evaluate the genetic diversity of hazelnut genetic germplasms in the forests of the northwest of the Iran by SCoT markers and it’s accordance to diversity estimates based on national markers of hazelnut differentiation tests.

In this study, 77 genotypes from hazelnut forests of Ardabil, Miyaneh and Arasbaran were selected by locating each genotype using GPS. Qualitative traits were evaluated using the national guidelines for uniform differentiation and hazelnut stability tests. after DNA extraction from leaf tissue, their polymorphism diversity was assessed using 15 SCoT-specific primers.

The results showed that the genotypes were classified according to the qualitative traits of differentiation of 17 clusters. Using 15 SCoT markers, 165 polymorphic bands were produced with an average of 11 bands per marker and the average percentage of polymorphisms with an average of 11 bands per marker and the average percentage of polymorphisms for markers 90.2% of genotypes based on these markers in 19 sub-clusters Were grouped.

Analysis of qualitative traits based on bands created in SCoT showed that markers 3, 5, 11, 12, 18 and 20 had more genetic diversity and that the degree of  accordance of genotype grouping based on the SCoT markers was 0.87 on The basis of the Mantel test.

Using two genetic markers, 75% (R2) of the variety of hazelnut differentiation descriptors can be accessed.

Enterococcus faecalis species are significant issue in severe nosocomial infections. In this study we aimed to investigate the effect of ionic liquids based on amino acids on pathogenic genes of this bacterium using Real time PCR.

In this study, 100 samples of oral infections were taken from patients and Enterococcus faecalis strains were identified using biochemical tests. The presence and frequency of cyl, hyl and esp genes and their expression under the influence of ionic fluid were evaluated using real time-PCR.

From the total samples taken, twelve Enterococcus faecalis bacteria were isolated and identified. The highest frequency of pathogenic genes was related to cyl gene which was present in 11 strains and the expression of target genes in the treated group under the influence of ionic fluid decreased compared to the control group (p <0.05). With decreasing ion-based amino acid concentration, the ability of bacteria to form biofilms increased (p <0.05).

Widespread antibiotic resistance is the most important concern about Enterococcus and cytolysin is a gelatinase enzyme and one of the most important invasive agents of Enterococcus faecalis. The results of this study provide a basis for promoting the use of new antibacterial agents, including ionic fluid, to prevent further pathogenicity of Enterococcus faecalis.

Probiotics are a solution of living microorganisms and if are administered in adequate quantities, will be beneficial to the host's health. Probiotics have positive effects on the gastrointestinal tract; hence, these microorganisms must be resistant to the process of producing and storage in order to survive in the commercial product at the completion of their useful life, higher than the threshold of 106 CFU / g. We have examined several ways in order to increase the stability and activity of probiotics in commercial products: Selecting species resistant to acid and bile acids, compatibility to stress, and absorbing micronutrients. Microencapsulation.

in this case, Probiotic cultures were isolated from different randomly cheese samples. the using garam stain and biochemichal test and Bergey's Manual of Determinative Bacteriology, we isolated probiotic lactobacilli. then we microencapsulated them.to reach a high density and high concentration network of low-fat milk solution as a gel forming solution.

Based on past internal and external research, we witnessed successful microencapsulation. In this research, we could produce spherical capsules and moreover saw very high microcapsulation effectiveness about 100%. we applied the special characteristics of milk and capsules produced in this approach gives very high characteristics to the product.

insoluble microcapsules in water with milk protein can be a very good alternative to encapsulation of probiotics from herbal gels by ionotropic or polysaccharides like alginate. In addition to the considerable functional features of milk protein, it can also be efficient to decrease the size of capsules, as the size of capsules produced due to sensory taste features is very significant not to touch under the tongue.