Naveed Muhammad, Waseem Muhammad, Aziz Tariq, Hassan Jawad Ul, Makhdoom Syeda Izma, Ali Urooj, Alharbi Metab, Alsahammari Abdulrahman
Department of Biotechnology, Faculty of Science and Technology, University of Central Punjab, Lahore 54590, Pakistan.
Department of Agriculture, University of Ioannina, 47100 Arta, Greece.
Biomedicines. 2023 Mar 28;11(4):1039. doi: 10.3390/biomedicines11041039.
The emergence of antibiotic-resistant microorganisms is a significant concern in global health. Antibiotic resistance is attributed to various virulent factors and genetic elements. This study investigated the virulence factors of to create an mRNA-based vaccine that could help prevent antibiotic resistance. Distinct strains of the bacteria were selected for molecular identification of virulence genes, such as , , , and , which were performed utilizing PCR techniques. DNA extraction from samples of was conducted using the Cetyl Trimethyl Ammonium Bromide (CTAB) method, which was confirmed and visualized using a gel doc; 16S rRNA was utilized to identify the bacterial strains, and primers of , , , and genes were employed to identify the specific genes. Sequencing was carried out at Applied Bioscience International (ABI) in Malaysia. Phylogenetic analysis and alignment of the strains were subsequently constructed. We also performed an in silico analysis of the , , , and genes to generate an antigen-specific vaccine. The virulence genes were translated into proteins, and a chimera was created using various linkers. The mRNA vaccine candidate was produced utilizing 18 epitopes, linkers, and an adjuvant, known as RpfE, to target the immune system. Testing determined that this design covered 90% of the population conservancy. An in silico immunological vaccine simulation was conducted to verify the hypothesis, including validating and predicting secondary and tertiary structures and molecular dynamics simulations to evaluate the vaccine's long-term viability. This vaccine design may be further evaluated through in vivo and in vitro testing to assess its efficacy.
抗生素耐药性微生物的出现是全球健康领域的一个重大问题。抗生素耐药性归因于多种毒力因子和遗传元件。本研究调查了[细菌名称]的毒力因子，以创建一种基于mRNA的疫苗，该疫苗有助于预防抗生素耐药性。选择了该细菌的不同菌株进行毒力基因的分子鉴定，如[具体基因名称1]、[具体基因名称2]、[具体基因名称3]和[具体基因名称4]，这些鉴定是利用PCR技术进行的。使用十六烷基三甲基溴化铵（CTAB）法从[细菌名称]样本中提取DNA，通过凝胶成像仪进行确认和可视化；利用16S rRNA鉴定细菌菌株，并使用[具体基因名称1]、[具体基因名称2]、[具体基因名称3]和[具体基因名称4]基因的引物来鉴定特定基因。测序在马来西亚的应用生物系统国际公司（ABI）进行。随后构建了菌株的系统发育分析和比对。我们还对[具体基因名称1]、[具体基因名称2]、[具体基因名称3]和[具体基因名称4]基因进行了计算机模拟分析，以生成一种抗原特异性疫苗。将毒力基因翻译成蛋白质，并使用各种连接子创建嵌合体。利用18个表位、连接子和一种名为RpfE的佐剂生产了mRNA候选疫苗，以靶向免疫系统。测试确定该设计覆盖了90%的群体保守性。进行了计算机免疫疫苗模拟以验证该假设，包括验证和预测二级和三级结构以及分子动力学模拟，以评估疫苗的长期生存能力。这种疫苗设计可通过体内和体外测试进一步评估其疗效。
