Immunoinformatics Approach to Design Novel Subunit Vaccine against the Epstein-Barr Virus.

Epstein-Barr virus (EBV) is a lymphotropic virus responsible for numerous epithelial and lymphoid cell malignancies, including gastric carcinoma, Hodgkin's lymphoma, nasopharyngeal carcinoma, and Burkitt lymphoma. Hundreds of thousands of people worldwide get infected with this virus, and in most cases, this viral infection leads to cancer. Although researchers are trying to develop potential vaccines and drug therapeutics, there is still no effective vaccine to combat this virus. In this study, the immunoinformatics approach was utilized to develop a potential multiepitope subunit vaccine against the two most common subtypes of EBV, targeting three of their virulent envelope glycoproteins. Eleven cytotoxic T lymphocyte (CTL) epitopes, 11 helper T lymphocyte (HTL) epitopes, and 10 B-cell lymphocyte (BCL) epitopes were predicted to be antigenic, nonallergenic, nontoxic, and fully conserved among the two subtypes, and nonhuman homologs were used for constructing the vaccine after much analysis. Later, further validation experiments, including molecular docking with different immune receptors (e.g., Toll-like receptors [TLRs]), molecular dynamics simulation analyses (including root means square deviation [RMSD], root mean square fluctuation [RMSF], radius of gyration [Rg], principal-component analysis [PCA], dynamic cross-correlation [DCC], definition of the secondary structure of proteins [DSSP], and Molecular Mechanics Poisson-Boltzmann Surface Area [MM-PBSA]), and immune simulation analyses generated promising results, ensuring the safe and stable response of the vaccine with specific immune receptors after potential administration within the human body. The vaccine's high binding affinity with TLRs was revealed in the docking study, and a very stable interaction throughout the simulation proved the potential high efficacy of the proposed vaccine. Further, in silico cloning was also conducted to design an efficient mass production strategy for future bulk industrial vaccine production. IMPORTANCE Epstein-Barr virus (EBV) vaccines have been developing for over 30 years, but polyphyletic and therapeutic vaccines have failed to get licensed. Our vaccine surpasses the limitations of many such vaccines and remains very promising, which is crucial because the infection rate is higher than most viral infections, affecting a whopping 90% of the adult population. One of the major identifications covers a holistic analysis of populations worldwide, giving us crucial information about its effectiveness for everyone's unique immunological system. We targeted three glycoproteins that enhance the virulence of the virus to design an epitope-based polyvalent vaccine against two different strains of EBV, type 1 and 2. Our methodology in this study is nonconventional yet swift to show effective results while designing vaccines.

Bioactive potentiality of secondary metabolites from endophytic bacteria against SARS-COV-2: An in-silico approach.

Five endophytic bacterial isolates were studied to identify morphologically and biochemically, according to established protocols and further confirmed by 16S rDNA Sanger sequencing, as Priestia megaterium, Staphylococcus caprae, Neobacillus drentensis, Micrococcus yunnanensis, and Sphingomonas paucimobiliz, which were then tested for phytohormone, ammonia, and hydrolytic enzyme production. Antioxidant compounds total phenolic content (TPC), and total flavonoid content (TFC) were assessed by using bacterial crude extracts obtained from 24-hour shake-flask culture. Phylogenetic tree analysis of those identified isolates shared sequence similarities with the members of Bacillus, Micrococcus, Staphylococcus, and Pseudomonas species, and after GenBank submission, accession numbers for the nucleotide sequences were found to be MW494406, MW494408, MW494401, MW494402, and MZ021340, respectively. In silico analysis was performed to identify their bioactive genes and compounds in the context of bioactive secondary metabolite production with medicinal value, where nine significant bioactive compounds according to six different types of bioactive secondary metabolites were identified, and their structures, gene associations, and protein-protein networks were analyzed by different computational tools and servers, which were reported earlier with their antimicrobial, anti-infective, antioxidant, and anti-cancer capabilities. These compounds were then docked to the 3-chymotrypsin-like protease (3CLpro) of the novel SARS-COV-2. Docking scores were then compared with 3CLpro reference inhibitor (lopinavir), and docked compounds were further subjected to ADMET and drug-likeness analyses. Ligand-protein interactions showed that two compounds (microansamycin and aureusimine) interacted favorably with coronavirus 3CLpro. Besides, in silico analysis, we also performed NMR for metabolite detection whereas three metabolites (microansamycin, aureusimine, and stenothricin) were confirmed from the 1H NMR profiles. As a consequence, the metabolites found from NMR data aligned with our in-silico analysis that carries a significant outcome of this research. Finally, Endophytic bacteria collected from medicinal plants can provide new leading bioactive compounds against target proteins of SARS-COV-2, which could be an effective approach to accelerate drug innovation and development.

SHANK3 genetic polymorphism and susceptibility to ASD: evidence from molecular, in silico, and meta-analysis approaches.

BACKGROUND: The SHANK3 gene encodes a master synaptic scaffolding protein at the excitatory synapse's postsynaptic density, which is predominantly responsible for constructing a synapse, maintaining synaptic structure, and functions. Recently, evidence from rare mutations and copy number variation provided an important clue about SHANK3 which acts as a strong candidate gene in the pathogenesis of Autism Spectrum Disorder (ASD). MATERIALS AND METHODS: To investigate potential allelic variants for the SHANK3 (rs9616915) gene as a genetic risk factor, we performed PCR-RFLP analysis and Sanger sequencing for 90 ASD and 90 healthy subjects. Moreover, to understand the functional and structural impacts of our selected non-synonymous SHANK3 SNP rs9616915, we have performed an in silico analysis. Subsequently, a meta-analysis of rs9616915 with a total of 6 eligible studies (including the present study) containing a total of 795 cases and 12,947 controls was obtained from a comprehensive online database search to evaluate the overall association with ASD. RESULTS: Our retrieved data, such as Pearson's chi-square test (p = 0.081) as well as logistic regression analysis of co-dominant (p = 0.1131), dominant (p = 0.3656) and recessive models (p = 0.0569) speculated no significant association between rs9616915 and our studied sample. Interestingly, by in silico analysis, we have observed two hydrogen bonds between amino acids instead of one hydrogen bond in the protein structure of rs9616915, which indicates this mutant structure could affect the proteins' stability. The findings of the meta-analysis revealed that four genetic association models were associated with ASD susceptibility. CONCLUSIONS: Our study suggested that targeted SHANK3 SNP of interest rs9616915 might not be associated with ASD in the southern part of the Bangladeshi population.

Exploring the Structural and Functional Effects of Nonsynonymous SNPs in the Human Serotonin Transporter Gene Through In Silico Approaches.

The sodium-dependent serotonin transporter SLC6A4 (solute carrier family 6 member 4) gene encodes an intrinsic membrane protein that transmits the serotonin neurotransmitter from synaptic clefts into presynaptic neurons. The product of the SLC6A4 gene is related to the regulation of mood and social behavior, sleep, appetite, memory, digestion, and sexual desire. This protein is a target for antidepressant and psychostimulant drugs, thus prolonged neurotransmitter signaling remains blocked. In this study, the functional consequences of nsSNPs in the human SLC6A4 gene were explored through computational tools: PhD-SNP, SIFT, Align GVGD, PROVEAN, PMut, nsSNP Analyzer, SNPs&GO, SNAP2, PolyPhen2, and PANTHER to identify the most deleterious and damaging nsSNPs. Then the mutant protein stabilities were assessed using I-Mutant, MUpro, and MutPred2; amino acid conservation using ConSurf, and posttranslational modification analysis using MusiteDEEP and PROSPER. Furthermore, the 3-dimensional (3D) model of the mutated proteins was predicted and validated using SPARKS-X, Verify3D, and PROCHECK. The protein-ligand binding sites were analyzed using the COACH meta-server. Results from this study predicted that T192M, G342E, R607C, W282S, R104C, P131L, P156L, and N351S were the most structurally and functionally significant nsSNPs in the human SLC6A4 gene. Arg607 and Pro156 were the predicted sites for posttranslational modifications, and Thr192 and Try282 were the ligand-binding sites in the human SLC6A4 gene. The analyzed data also suggested that R104C, P131L, P156L, T192M, G342E, and W282S mutants might affect the binding of sodium ions with this protein. Taken together, this study provided important information on structurally and functionally important nsSNPs of the human SLC6A4 gene for further experimental validation. In the future, these damaging nsSNPs of the SLC6A4 gene have the potential to be evaluated as prognostic biomarkers for SLC6A4-related disorder diagnosis and research.

Prevalence and impact of the use of electronic gadgets on the health of children in secondary schools in Bangladesh: A cross-sectional study.

BACKGROUND AND AIMS: Use of technological gadgets has rapidly been increasing among adolescents, which may result in health issues and technology addiction. This study focuses on the prevalence of usage of technological gadgets and health-related complications among secondary school-going children of Bangladesh. METHODS: A total of 1803 secondary school students from 21 different districts of Bangladesh participated in the study. The children were asked questions relating to their access to electronic gadgets, time spent on outdoor activities, and whether they experienced any health-complications as an after-effect of the usage. A binary logistic regression model was adapted considering time spent on gadgets as an independent variable and health problems (physical and mental) as the dependent variable. RESULTS: Among all the gadgets, 67.11% of the participants were reported to use mobile phones on a daily basis. Due to the ongoing COVID-19 pandemic, 24.48% of respondents used electronic gadgets for attending online classes. The participants were reported to use gadgets significantly more (P < .05) in 2020 as compared to 2019. Children showed less tendency to spend time in outdoor activities. More than 50% of the participants spend time doing outdoor activities for less than 1 hour daily. An association between gadget use and health problems like headache, backache, visual disturbance, and sleeping disturbance has been observed in our study. CONCLUSION: This study demonstrates that different socio-demographic factors have influence on the use of gadgets by children, and this use has greatly been affecting both the physical and mental health of the secondary school-going students of Bangladesh.

Microbial Bioremediation of Feather Waste for Keratinase Production: An Outstanding Solution for Leather Dehairing in Tanneries.

In leather industries and tanneries, large amount of wastes has been disposed; which polluting water, soil, and atmosphere and causing serious human health problems. In particular, chemical dehairing process of leather industries produces fair amount of toxic wastes. It is, thus, urgently needed to use alternative processes free from pollution. As more than 90% of keratin is contained in feather, it is desirable to develop bioremediation process using keratinolytic microorganisms. In the present investigation, therefore, we first identified Bacillus cereus and Pseudomonas sp. to be able to produce keratinase. Then, the optimization was performed to maximize the keratinase activity with respect to cultivation temperature, pH, and incubation time. Moreover, the effects of metal ions and various substrates on keratinase activity were also investigated. The result indicates that keratinase activity became maximum at 50°C for both strains, whereas the optimal pH was 10.0 for B. cereus and 7.0 for Pseudomonas sp. The highest keratinase activity of 74.66 ± 1.52 U/mL was attained by B. cereus, whereas 57.66 ± 2.52 U/mL was attained by Pseudomonas sp. Enzymatic dehairing efficiency of leathers was also compared with chemical dehairing (Na2S and CaO), where complete dehairing was achieved by treating them with crude keratinase. Partial enzyme purification was performed by acetone precipitation. Batch cultivation of B. cereus using 1 L fermentor indicates a potential candidate for large-scale keratinase production. Thus, keratinase enzyme by degrading poultry wastes (feather) can be an alternative approach to chemical dehairing in leather industries, thus preventing environmental pollution through bioremediation.

A single metabolite production by Escherichia coli BW25113 and its pflA.cra mutant cultivated under microaerobic conditions using glycerol or glucose as a carbon source.

Abundant, low prices and a highly reduced nature make glycerol to be an ideal feedstock for the production of reduced biochemicals and biofuels. Escherichia coli has been paid much attention as the platform of microbial cell factories due to its high growth rate (giving higher metabolite production rate) and the capability of utilizing a wide range of carbon sources. However, one of the drawbacks of using E. coli as a platform is its mixed metabolite formation under anaerobic conditions. In the present study, it was shown that ethanol could be exclusively produced from glycerol by the wild type E. coli, while d-lactic acid could be exclusively produced from glucose by pflA.cra mutant, where the glucose uptake rate could be increased by this mutant as compared to the wild type strain. It was also shown that the growth rate is significantly reduced in pflA.cra mutant for the case of using glycerol as a carbon source due to redox imbalance. The metabolic regulation mechanisms behind the fermentation characteristic were clarified to some extent.