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The Middle Eastern prevalence of sickle cell anemia, a genetic disorder that affects red blood cells, necessitates additional research. On a molecular level, we sought to identify and sort the oral microbiota of healthy individuals and those with sickle cell anemia. Furthermore, it is crucial to comprehend how changes in the genetic makeup of the oral microbiota impact the state of sickle cell anemia. Using next-generation sequencing, the 16S rRNA amplicon was examined using saliva samples from 36 individuals with sickle cell anemia and healthy individuals. These samples were obtained from sickle cell anemia patients (18 samples) and healthy control participants (controls, 18 samples). Various analyses are conducted using bioinformatic techniques to identify distinct species and their relative abundance. Streptococcus, followed by Fusobacterium nucleatum, Prevotella, and Veillonella were the most prevalent genera of bacteria in the saliva of the SCA and non-SCA individuals according to our findings. Rothia mucilaginosa, Prevotella scoposa, and Veillonella dispar species were the dominant species in both sickle cell anemia and non-sickle cell anemia subjects. Streptococcus salivarius, Actinomyces graevenitzii, Actinomyces odontolyticus, and Actinomyces georgiae spp. were the most prevalent bacterial spp. in the studied SCA cases. The sequencing of the 16S rRNA gene yielded relative abundance values that were visualized through a heatmap analysis. Alterations in the oral microflora's constitution can significantly affect the susceptibility of sickle cell anemia patients to develop more severe health complications. Salivary diagnosis is a potential tool for predicting and preventing oral microbiome-related diseases in the future. [ABSTRACT FROM AUTHOR]

STAPHYLOCOCCUS aureus can enter the bloodstream, leading to health complications such as sepsis, arthritis, endocarditis, and pneumonia. This study screened for the impact of six replication genes of cos phage 12 (φ12) on the evolution and transfer of cos S. aureus pathogenicity islands (SaPIbov5). An overnight culture of the S. aureus strain RN4220 diluted with fresh TSB was used in the bacteriophage φ12 titering assay. Phage-point mutagenesis was achieved using the pMAD vector, which facilitates homologous recombination in a two-step process. Finally, the transduction SaPI titering assays also utilized the S. aureus strain RN4220, which was then mixed with CaCl2 and fresh TSB. The study showed that ORF11, ORF12, and VirE mutants did not lyse or produce phage particles after titering into the recipient RN4220 strains. While ORF26, ORF10, and ORF04 mutants generated detectable plaques, the mutations may have an effect on phage replication or packaging after tittering. For instance, complemented φ12 ORF26, ORF10, and ORF04 mutant strains had fully restored phage titers. The experiment found that cos phages facilitate inter and intra-generically transfer of cosSaPIs. It was also established that φ12 transduces SaPIbov5. However, φ12 mutants in VirE, ORF12, and ORF11 did not show SaPI mobilization because of their effects on infective phage particles and lack of replication. The complemented φ12 VirE, ORF12, and ORF11 mutants had partly restored phage titers when they were expressed in the recipient and donor strains. Mutations in ORF26, 0RF10, and ORFO4 decrease the φ12's ability to transfer SaPIbov5. Generally, this study found a new mechanism that facilitates the transfer of SaPI and cos genes. While the VirE, ORF12, and ORF11 affect packaging and are necessary for replication and phage biology [ABSTRACT FROM AUTHOR]

Coronaviruses emerged three times in the last two decades and became a source of concern globally. Humulus lupulus plant has been used widely in medical science. Our objective in this study was to determine the effects of the crude extract of H. lupulus in inhibiting MERS-CoV and SARS-CoV-2 viruses' replication in vitro using Vero E6 cell lines and predict the antiviral activity of its identified compounds against the receptor binding (RBD) protein of both viruses in silico. We determined the concentration of the extract that induced less than 50% cell toxicity (CC50), and the antiviral activity based on IC50 and plaque reduction assay. We used molecular docking simulation to predict the potential of known active compounds in H. lupulus to inhibit the RBD protein. H. lupulus extract showed very low toxicity on Vero E6 cells with CC50= 23.25 µg/µL and antiviral activity toward MERS-CoV and SARS-CoV-2 with IC50= 0.18 and 0.9 µg/µL, respectively. The crude extract showed inhibition rate of 84.6% with MERS-CoV and 80% with SARS-CoV-2. In silico analysis predicted the compounds 5′-prenylxanthohumo, xanthogalenol, dehydrocycloxanthohumol hydrate, 6-prenylnaringenin, isoxanthohumol, catechin gallate, epicatechin gallate, 8-prenylnaringenin and xanthohumol to inhibit MERS-CoV and SARS-CoV-2 invasion of host cells by interfering with viral spike protein and the host cell receptor recognition process. Drug likeness and toxicity risk prediction analysis confirmed their capability as potential drugs. Based on our findings, isolation, purification and testing of the suggested active compounds could lead to novel anti-coronavirus drugs. [ABSTRACT FROM AUTHOR]

The emergence of superbugs and resistant pathogens poses a challenge in scientific and medical research as they threaten public health worldwide. Many herbal natural products currently used in therapies have been suggested to exert antimicrobial, antiviral and even virucidal activities against a vast majority of impervious pathogens. Rhazya stricta, a folk medicinal desert plant from Saudi Arabia was recently revealed to exhibit bactericidal activity against multidrug-resistant (MDR) microorganisms. The pandemic caused by the severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) is a threat to public health worldwide. Hence, we examined the antiviral activity of R. stricta against the virus. The R. stricta water extract was prepared at the traditional dose. The antagonistic effects of this extract against pathogens have been proven in previous studies, and those against SARS-CoV-2 were shown in the present study. Therefore, we explored the effects of the plant extracts and fractions against the virus for future drug development. All plant extracts showed antiviral effects against SARS-CoV-2 in the Vero E6 cell lines. Non-alkaloids showed the strongest effect against the virus, followed by weak base alkaloids and finally strong base alkaloids. A cytotoxicity assay was performed to explore the safest dose with the strongest antiviral effects. The non-alkaloid extract derived from R. stricta leaves is a promising antiviral candidate for the development of potential drugs with appropriate activity against COVID-19 and other life-threatening diseases. [ABSTRACT FROM AUTHOR]

Increase in the incidence of Insulin Dependent Diabetes Mellitus (IDDM) among people from developed and developing countries has created a large global market for insulin. Moreover, exploration of new methods for insulin delivery including oral or inhalation route which require very high doses would further increase the demand of cost-effective recombinant insulin. Various bacterial and yeast strains have been optimized to overproduce important biopharmaceuticals. One of the approaches we have taken is the production of recombinant human insulin along with C-peptide in yeast Pichia pastoris. We procured a cDNA clone of insulin from Origene Inc., USA. Insulin cDNA was PCR amplified and cloned into yeast vector pPICZ-α. Cloned insulin cDNA was confirmed by restriction analysis and DNA sequencing. pPICZ-α-insulin clone was transformed into Pichia pastoris SuperMan5 strain. Several Zeocin resistant clones were obtained and integration of insulin cDNA in Pichia genome was confirmed by PCR using insulin specific primers. Expression of insulin in Pichia clones was confirmed by ELISA, SDS-PAGE, and Western blot analysis. In vivo efficacy studies in streptozotocin induced diabetic mice confirmed the activity of recombinant insulin. In conclusion, a biologically active human proinsulin along with C-peptide was expressed at high level using Pichia pastoris expression system. [ABSTRACT FROM AUTHOR]

Influenza viruses have developed resistance to the current classes of drugs, which means they could eventually become more virulent and cause more mortality and hospitalization. Our study aims to investigate the antiviral activity of Rhazya stricta Decne leaves extract in vitro and search for new promising drugs from R. stricta identified compounds in silico. The study was performed in vitro by utilizing Madin-Darby Canine Kidney cell line (MDCK) as a substrate for the influenza virus and estimating the inhibition performance of the plant leaves extract. Additionally, in silico screening was conducted to explore the antiviral activity of R. stricta phytochemicals. We investigated the cytotoxicity of R. stricta leaves extract and its antiviral activity against influenza virus (A/Puerto Rico/8/34 (H1N1)) using the MTT assay. The mode of action of the plant leaves extract during the viral life cycle was tested using time-of-addition assay. In silico analyses were performed, including molecular docking, drug-likeness analysis, and toxicity risk assessment, to state the leading compounds to be developed into an anti-influenza virus drug. The 50% cytotoxicity concentration of the leaves extract was CC50: 184.6 µg/mL, and the 50% inhibition concentration was CI50: 19.71 µg\mL. The time of addition assay revealed that R. stricta leaves extract exerted its activity in the late step of the influenza virus replication cycle. In comparison to Oseltamivir, the leading compounds showed better binding affinity and can be developed into oral drugs with low toxicity risk. Isolation and purification of the leading compounds and testing their antiviral activity in vitro and in vivo are required. [ABSTRACT FROM AUTHOR]

The local medicinal plant Rhazya stricta Decne is reviewed for its folkloric medicinal, phytochemical, pharmacological, biological, and toxicological features. R. stricta has been used widely in different cultures for various medical disorders. The phytochemical studies performed on the R. stricta extract revealed many alkaloidal and fatty acid compounds. Moreover, several flavonoid and terpenoid compounds were also detected. Pharmacological activates of R. stricta extracts are approved to possess antimicrobial, antioxidant, anticancer, antidiabetic, and antihypertensive activities. Additionally, R. stricta extract was found to hold biological activates such as larvicidal and phytoremediation activates R. stricta extract was found to be toxic, genotoxic, and mutagenic. R. stricta contains novel phytochemical compounds that have not been investigated pharmacologically. Further research is needed through in vitro and in vivo experiments to pave the road for these compounds for medical, veterinary, and ecological uses. [ABSTRACT FROM AUTHOR]

Middle East respiratory syndrome coronavirus (MERS-CoV) is a highly infectious zoonotic virus first reported into the human population in September 2012 on the Arabian Peninsula. The virus causes severe and often lethal respiratory illness in humans with an unusually high fatality rate. The N-terminal domain (NTD) of receptor-binding S1 subunit of coronavirus spike (S) proteins can recognize a variety of host protein and mediates entry into human host cells. Blocking the entry by targeting the S1-NTD of the virus can facilitate the development of effective antiviral drug candidates against the pathogen. Therefore, the study has been designed to identify effective antiviral drug candidates against the MERS-CoV by targeting S1-NTD. Initially, a structure-based pharmacophore model (SBPM) to the active site (AS) cavity of the S1-NTD has been generated, followed by pharmacophore-based virtual screening of 11,295 natural compounds. Hits generated through the pharmacophore-based virtual screening have re-ranked by molecular docking and further evaluated through the ADMET properties. The compounds with the best ADME and toxicity properties have been retrieved, and a quantum mechanical (QM) based density-functional theory (DFT) has been performed to optimize the geometry of the selected compounds. Three optimized natural compounds, namely Taiwanhomoflavone B (Amb23604132), 2,3-Dihydrohinokiflavone (Amb23604659), and Sophoricoside (Amb1153724), have exhibited substantial docking energy >−9.00 kcal/mol, where analysis of frontier molecular orbital (FMO) theory found the low chemical reactivity correspondence to the bioactivity of the compounds. Molecular dynamics (MD) simulation confirmed the stability of the selected natural compound to the binding site of the protein. Additionally, molecular mechanics generalized born surface area (MM/GBSA) predicted the good value of binding free energies (ΔG bind) of the compounds to the desired protein. Convincingly, all the results support the potentiality of the selected compounds as natural antiviral candidates against the MERS-CoV S1-NTD. [ABSTRACT FROM AUTHOR]