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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]

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]