Conducting the RBD of SARS-CoV-2 Omicron Variant with Phytoconstituents from Euphorbia dendroides to Repudiate the Binding of Spike Glycoprotein Using Computational Molecular Search and Simulation Approach.

(1) Background: Natural constituents are still a preferred route for counteracting the outbreak of COVID-19. Essentially, flavonoids have been found to be among the most promising molecules identified as coronavirus inhibitors. Recently, a new SARS-CoV-2 B.1.1.529 variant has spread in many countries, which has raised awareness of the role of natural constituents in attempts to contribute to therapeutic protocols. (2) Methods: Using various chromatographic techniques, triterpenes (1-7), phenolics (8-11), and flavonoids (12-17) were isolated from Euphorbia dendroides and computationally screened against the receptor-binding domain (RBD) of the SARS-CoV-2 Omicron variant. As a first step, molecular docking calculations were performed for all investigated compounds. Promising compounds were subjected to molecular dynamics simulations (MD) for 200 ns, in addition to molecular mechanics Poisson-Boltzmann surface area calculations (MM/PBSA) to determine binding energy. (3) Results: MM/PBSA binding energy calculations showed that compound 14 (quercetin-3-O-ß-D-glucuronopyranoside) and compound 15 (quercetin-3-O-glucuronide 6″-O-methyl ester) exhibited strong inhibition of Omicron, with ΔGbinding of -41.0 and -32.4 kcal/mol, respectively. Finally, drug likeness evaluations based on Lipinski's rule of five also showed that the discovered compounds exhibited good oral bioavailability. (4) Conclusions: It is foreseeable that these results provide a novel intellectual contribution in light of the decreasing prevalence of SARS-CoV-2 B.1.1.529 and could be a good addition to the therapeutic protocol.

Immunoinformatics assisted profiling of West Nile virus proteome to determine immunodominant epitopes for the development of next-generation multi-peptide vaccine.

Emerging infectious diseases represent a significant threat to global health, with West Nile virus (WNV) being a prominent example due to its potential to cause severe neurological disorders alongside mild feverish conditions. Particularly prevalent in the continental United States, WNV has emerged as a global concern, with outbreaks indicating the urgent need for effective prophylactic measures. The current problem is that the absence of a commercial vaccine against WNV highlights a critical gap in preventive strategies against WNV. This study aims to address this gap by proposing a novel, multivalent vaccine designed using immunoinformatics approaches to elicit comprehensive humoral and cellular immune responses against WNV. The objective of the study is to provide a theoretical framework for experimental scientists to formulate of vaccine against WNV and tackle the current problem by generating an immune response inside the host. The research employs reverse vaccinology and subtractive proteomics methodologies to identify NP_041724.2 polyprotein and YP_009164950.1 truncated flavivirus polyprotein NS1 as the prime antigens. The selection process for epitopes focused on B and T-cell reactivity, antigenicity, water solubility, and non-allergenic properties, prioritizing candidates with the potential for broad immunogenicity and safety. The designed vaccine construct integrates these epitopes, connected via GPGPG linkers, and supplemented with an adjuvant with the help of another linker EAAAK, to enhance immunogenicity. Preliminary computational analyses suggest that the proposed vaccine could achieve near-universal coverage, effectively targeting approximately 99.74% of the global population, with perfect coverage in specific regions such as Sweden and Finland. Molecular docking and immune simulation studies further validate the potential efficacy of the vaccine, indicating strong binding affinity with toll-like receptor 3 (TLR-3) and promising immune response profiles, including significant antibody-mediated and cellular responses. These findings present the vaccine construct as a viable candidate for further development and testing. While the theoretical and computational results are promising, advancing from in-silico predictions to a tangible vaccine requires comprehensive laboratory validation. This next step is essential to confirm the vaccine's efficacy and safety in eliciting an immune response against WNV. Through this study, we propose a novel approach to vaccine development against WNV and contribute to the broader field of immunoinformatics, showcasing the potential to accelerate the design of effective vaccines against emerging viral threats. The journey from hypothesis to practical solution embodies the interdisciplinary collaboration essential for modern infectious disease management and prevention strategies.

Metagenomic next-generation sequencing of nasopharyngeal microbiota in COVID-19 patients with different disease severities.

Throughout the COVID-19 pandemic, extensive research has been conducted on SARS-COV-2 to elucidate its genome, prognosis, and possible treatments. However, few looked at the microbial markers that could be explored in infected patients and that could predict possible disease severity. The aim of this study is to compare the nasopharyngeal microbiota of healthy subjects, moderate, under medication, and recovered SARS-COV-2 patients. In 2020, 38 nasopharyngeal swabs were collected from 6 healthy subjects, 14 moderates, 10 under medication and 8 recovered SARS-COV-2 patients at the Prince Mohammed Bin Abdulaziz Hospital Riyadh. Metatranscriptomic sequencing was performed using Minion Oxford nanopore sequencing. No significant difference in alpha as well as beta diversity was observed among all four categories. Nevertheless, we have found that Streptococcus spp including Streptococcus pneumoniae and Streptococcus thermophilus were among the top 15 most abundant species detected in COVID-19 patients but not in healthy subjects. The genus Staphylococcus was found to be associated with COVID-19 patients compared to healthy subjects. Furthermore, the abundance of Leptotrichia was significantly higher in healthy subjects compared to recovered patients. Corynebacterium on the other hand, was associated with under-medication patients. Taken together, our study revealed no differences in the overall microbial composition between healthy subjects and COVID-19 patients. Significant differences were seen only at specific taxonomic level. Future studies should explore the nasopharyngeal microbiota between controls and COVID-19 patients while controlling for confounders including age, gender, and comorbidities; since these latter could affect the results and accordingly the interpretation.IMPORTANCEIn this work, no significant difference in the microbial diversity was seen between healthy subjects and COVID-19 patients. Changes in specific taxa including Leptotrichia, Staphylococcus, and Corynebacterium were only observed. Leptotrichia was significantly higher in healthy subjects, whereas Staphylococcus and Corynebacterium were mostly associated with COVID-19, and specifically with under-medication SARS-COV-2 patients, respectively. Although the COVID-19 pandemic has ended, the SARS-COV-2 virus is continuously evolving and the emergence of new variants causing more severe disease should be always kept in mind. Microbial markers in SARS-COV-2 infected patients can be useful in the early suspicion of the disease, predicting clinical outcomes, framing hospital and intensive care unit admission as well as, risk stratification. Data on which microbial marker to tackle is still controversial and more work is needed, hence the importance of this study.

Computational screening of natural products to identify potential inhibitors for human neuropilin-1 (NRP1) receptor to abrogate the binding of SARS-CoV-2 and host cell.

Recently, a new variant B.1.1.529 or Omicron variant and its sub-variants (BA2.75, BA.5) of SARS-CoV-2 (Severe acute respiratory virus 2) have been reported with a larger number of mutations in the spike protein and particularly in the RBD (receptor-binding domain). The omicron (B.1.1.529) variant has aggravated the pandemic situation further and needs more analysis for therapeutic development. Keeping in view the urgency of the required data, the current study used molecular modeling and simulation-based methods to target the NRP1 (Neuropilin 1) protein to halt the entry into the host cell. Employing a molecular screening approach to screen the North-East African natural compounds database (NEANCDB) revealed Subereamine B with a docking score of -8.44 kcal/mol, Zinolol with the docking score of -8.05 while Subereamine A with a docking score of -7.88 kcal/mol as the best hits against NRP1. Molecular simulation-based further validation revealed stable dynamics, good structural packing, and dynamic residues flexibility index. Moreover, hydrogen bonding fraction analysis demonstrated the interactions remained sustained during the simulation. Furthermore, the total binding free energy for Subereamine B was -44.24 ±0.91 kcal/mol, for Zinolol -34.32 ±0.40 kcal/mol while for Subereamine A the TBE was calculated to be -41.78 ± 0.36 kcal/mol respectively. This shows that the two arginine-based alkaloids, i.e. Subereamine B and Subereamine A could inhibit the NRP1 more strongly than Zinolol. In conclusion, this study provides a basis for the development of novel drugs against SARS-CoV-2.Communicated by Ramaswamy H. Sarma.

Multifunctional Lipophobic Polymer Dots from Cyclodextrin: Antimicrobial/Anticancer Laborers and Silver Ions Chemo-Sensor.

ß-Cyclodextrin (CD) is currently exploited for the implantation of lipophobic polymer dots (PDs) for antimicrobial and anticancer laborers. Moreover, the PDs were investigated to act as a chemo-sensor for metal detection. The data revealed that under basic conditions, photoluminescent PDs (5.1 nm) were successively clustered with a controllable size at 190 °C, whereas under acidic conditions, smaller-sized non-photoluminescent carbon nanoparticles (2.9 nm) were obtained. The fluorescence intensity of synthesized PDs under basic conditions was affected by pH, and such an intensity was significantly higher compared to that prepared under acidic conditions. The PDs were exploited as florescent detectors in estimation of Ag+ ions in aquatic streams. Treatment of Ag+ ion colloids with PDs resulted in fluorescence quenching attributing to the production of AgNPs that approved by spectral studies. The cell viability percent was estimated for Escherichia coli, Staphylococcus aureus, and Candida albicans after incubation with PDs implanted under basic conditions for 24 h. The cell mortality percent was estimated for breast cancer (MCF-7) after incubation with different concentrations of PDs that were implanted under acidic versus basic conditions to show that treatment of the tested cells with 1000 µg/mL PDs prepared under basic (IC50 232.5 µg/mL) and acidic (IC50 88.6 µg/mL) conditions resulted in cell mortality percentages of 70 and 90%, respectively.

Evaluation of the Levels of Peripheral CD3+, CD4+, and CD8+ T Cells and IgG and IgM Antibodies in COVID-19 Patients at Different Stages of Infection.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection affects the stimulatory levels of cellular-mediated immunity, which plays an essential role in controlling SARS-CoV-2 infection. In fact, several studies have shown the association of lymphopenia with severe COVID-19 in patients. The aim of this study is to investigate the response of the immune system, including cell-mediated immunity and antibody production, during different stages of SARS-CoV-2 infection. Peripheral blood and serum samples were collected from patients with moderate infection, patients under medication (hospitalized), patients who had recovered, and healthy individuals (n = 80). Flow cytometry analysis was performed on peripheral blood samples to determine the cellular immunity profile of each patient. The data showed a significant reduction in the levels of CD3+, CD4+, and CD8+ T cells and CD45+ cells in the moderate and under-medication groups, suggesting lymphopenia in those patients. Also, enzyme-linked immunosorbent assay (ELISA) was conducted on the serum samples to measure the levels of antibodies, including IgM and IgG, in each patient. The results revealed a significant increase in the levels of IgM in the moderate infection and under-medication patients, thus indicating the production of IgM during the first week of infection. Furthermore, changes in the levels of IgG were significantly detected among recovered patients, indicating therefore a remarkable increase during the recovery stage of SARS-CoV-2 infection and thus a strong humoral-mediated immunity. In summary, the results of this study may help us to understand the main role of the cellular immune responses, including CD3+, CD4+, and CD8+ T cells, against SARS-CoV-2 infection. This understanding might support the development of SARS-CoV-2 treatments and vaccines in the near future. IMPORTANCE Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) emerged in late 2019 in China. This virus is a serious threat to people not only in China but also worldwide, where it has been detected in over 222 countries. It has been reported that ∼3.4% of SARS-CoV-2-infected patients have died. The significance of our study relies on the fact that an enzyme-linked immunosorbent assay and flow cytometry were used to measure the levels of antibodies and cellular immune response, respectively, from clinical samples of patients infected with SARS-CoV-2.

Design of a Multi-Epitopes Based Chimeric Vaccine against Enterobacter cloacae Using Pan-Genome and Reverse Vaccinology Approaches.

Enterobacter cloacae (EC) is a significant emerging pathogen that is occasionally associated with lung infection, surgical site infection, urinary infection, sepsis, and outbreaks in neonatal intensive care units. In light of the fact that there is currently no approved vaccine or therapeutic option for the treatment of EC, the current study was developed to concentrate on applications based on modern computational approaches to design a multi-epitope-based E. cloacae peptide vaccine (MEBEPV) expressing the antigenic determinants prioritized from the EC genome. Integrated computational analyses identified two potential protein targets (phosphoporin protein-PhoE and putative outer-membrane porin protein) for further exploration on the basis of pangenome subtractive proteomics and immunoinformatic in-depth examination of the core proteomes. Then, a multi-epitope peptide vaccine was designed, which comprised shortlisted epitopes that were capable of eliciting both innate and adaptive immunity, as well as the cholera toxin's B-subunit, which was used as an adjuvant in the vaccine formulation. To ensure maximum expression, the vaccine's 3D structure was developed and the loop was refined, improving the stability by disulfide engineering, and the physicochemical characteristics of the recombinant vaccine sequence were found to be ideal for both in vitro and in vivo experimentation. Blind docking was then used for the prediction of the MEBEPV predominant blinding mode with MHCI, MHCII, and TLR3 innate immune receptors, with lowest global energy of -18.64 kJ/mol, -48.25 kJ/mol, and -5.20 kJ/mol for MHC-I, MHC-II, and TLR-4, respectively, with docked complexes considered for simulation. In MD and MMGBSA investigations, the docked models of MEBEPV-TLR3, MEBEPV-MHCI, and MEBEPV-MHCII were found to be stable during the course of the simulation. MM-GBSA analysis calculated -122.17 total net binding free energies for the TLR3-vaccine complex, -125.4 for the MHC I-vaccine complex, and -187.94 for the MHC II-vaccine complex. Next, MM-PBSA analysis calculated -115.63 binding free energy for the TLR3-vaccine complex, -118.19 for the MHC I-vaccine complex, and -184.61 for the MHC II-vaccine complex. When the vaccine was tested in silico, researchers discovered that it was capable of inducing both types of immune responses (cell mediated and humoral) at the same time. Even though the suggested MEBEPV has the potential to be a powerful contender against E. cloacae-associated illnesses, further testing in the laboratory will be required before it can be declared safe and immunogenic.

In silico screening of some compounds derived from the desert medicinal plant Rhazya stricta for the potential treatment of COVID-19.

The latest coronavirus pandemic (SARS-CoV-2) poses an exceptional threat to human health and society worldwide. The coronavirus (SARS-CoV-2) spike (S) protein, which is required for viral-host cell penetration, might be considered a promising and suitable target for treatment. In this study, we utilized the nonalkaloid fraction of the medicinal plant Rhazya stricta to computationally investigate its antiviral activity against SARS-CoV-2. Molecular docking and molecular dynamics simulations were the main tools used to examine the binding interactions of the compounds isolated by HPLC analysis. Ceftazidime was utilized as a reference control, which showed high potency against the SARS-CoV-2 receptor binding domain (RBD) in an in vitro study. The five compounds (CID:1, CID:2, CID:3, CID:4, and CID:5) exhibited remarkable binding affinities (CID:1, - 8.9; CID:2, - 8.7; and CID:3, 4, and 5, - 8.5 kcal/mol) compared to the control compound (- 6.2 kcal/mol). MD simulations over a period of 200 ns further corroborated that certain interactions occurred with the five compounds and the nonalkaloidal compounds retained their positions within the RBD active site. CID:2, CID:4, and CID:5 demonstrated high stability and less variance, while CID:1 and CID:3 were less stable than ceftazidime. The average number of hydrogen bonds formed per timeframe by CID:1, CID:2, CID:3, and CID:5 (0.914, 0.451, 1.566, and 1.755, respectively) were greater than that formed by ceftazidime (0.317). The total binding free energy calculations revealed that the five compounds interacted more strongly within RBD residues (CID:1 = - 68.8, CID:2 = - 71.6, CID:3 = - 74.9, CID:4 = - 75.4, CID:5 = - 60.9 kJ/mol) than ceftazidime (- 34.5 kJ/mol). The drug-like properties of the selected compounds were relatively similar to those of ceftazidime, and the toxicity predictions categorized these compounds into less toxic classes. Structural similarity and functional group analyses suggested that the presence of more H-acceptor atoms, electronegative atoms, acidic oxygen groups, and nitrogen atoms in amide or aromatic groups were common among the compounds with the lowest binding affinities. In conclusion, this in silico work predicts for the first time the potential of using five R. stricta nonalkaloid compounds as a treatment strategy to control SARS-CoV-2 viral entry.

Comparison of methodologies for enumerating and detecting the viability of Ascaris eggs in sewage sludge by standard incubation-microscopy, the BacLight Live/Dead viability assay and other vital dyes.

The aim of this study was to evaluate the Live/Dead BacLight viability kit as a method for enumerating viable eggs of Ascaris suum in sewage sludge as a surrogate for the human roundworm. The number and viability status of eggs of A. suum were accurately measured directly in sewage sludge samples by the BacLight method, compared to the conventional incubation-microscopy procedure. BacLight stains were not toxic to A. suum eggs, in contrast to some conventional vital dyes which disrupted viable eggs. The method was effective for the direct examination of eggs in heavily contaminated samples or seeded sludge containing ∼200 eggs/g DS in sludge with 5% DS content. However, a recovery method would be necessary to examine samples with small numbers of eggs, for instance in sludge from regions where the prevalence of infection with Ascaris lumbricoides is low. The BacLight technique may therefore be an effective alternative to conventional incubation-microscopy for enumerating Ascaris eggs in contaminated field samples or to validate sludge treatment processes by examining decay rates of inoculated A. suum eggs in laboratory simulations. Most field samples would require recovery from an appropriate number of composite samples prior to vital staining.