Evaluation of Protein Channel-Attenuated Salmonella Vaccines

蛋白质通道减毒沙门氏菌疫苗的评价

• 批准号: 7849975
• 负责人: Xinghong Yang
• 金额: $ 17.81万
• 依托单位: MONTANA STATE UNIVERSITY - BOZEMAN
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-06-01 至 2011-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7849975
• 关键词: Adherence; Affect; Animals; Antibiotic Resistance; Attenuated; Attenuated Vaccines; Bacteria; Bacterial Adhesins; Biogenesis; Biological; Brucella abortus; Cell surface; Cells; Cessation of life; Consumption; Country; Disease; Docking; Electrolytes; Enterobacteriaceae; Epidemic; Equilibrium; Erythromycin; Evaluation; Experimental Autoimmune Encephalomyelitis; Flagella; Food; Gastroenteritis; Gene Expression; Gene Mutation; Gene Silencing; Genes; Genomics; Gram-Negative Bacteria; Human; In Vitro; Infection; Intestines; Ions; Laboratories; Lead; Liquid substance; Mediating; Membrane; Methods; Molecular; Morphology; Mutation; Needles; Operon; Pathway interactions; Plasmid Cloning Vector; Protein Overexpression; Protein Secretion; Proteins; Recombinants; Regulator Genes; Research; SJL Mouse; Salmonella; Salmonella Vaccines; Salmonella enterica; Salmonella infections; Salmonella typhi; Salmonella typhimurium; Scanning Electron Microscopy; Scanning Probe Microscopes; Site-Directed Mutagenesis; Structural Genes; Surface; Techniques; Testing; Typhoid Fever; United States; Vaccines; Virulence; Water; appendage; attenuation; bacterial vector; base; colonization factor antigens; dalton; enterotoxigenic Escherichia coli; enterotoxin ST; fimbria; human disease; in vivo; macrophage; meetings; mouse model; mutant; overexpression; pathogen; prevent; protective efficacy; public health relevance; receptor; research study; small molecule; vaccine development; vector

DESCRIPTION (provided by applicant): The long-term objective of the project involves developing live vaccines through heterologous gene-mediated inactivation, as obtained using colonization factor antigen I (CFA/I) in Salmonella. We suspect this attenuation is because of the overexpression of channel CfaC and not due to the fimbriae CfaBE, although CfaBE may enhance the channel-mediated attenuation. Since this attenuation strategy was not previously described, we questioned whether it could be utilized for Salmonella vaccine development. Since conventional live vaccine development methods based on gene inactivation have been met with obstacles, this proposed study will generate Salmonella live vaccines for application in humans. As such, we set the following two Specific Aims: Specific Aim 1: Examine whether the deletion of fimbrial subunits, the secretion apparatus, or both are responsible for the observed attenuation of wild-type Salmonella. Specific Aim 2: Determine which of the deletion mutants of the cfa/I operon attenuates wild-type Salmonella and can therefore be used as a protective vaccine against salmonellosis. To achieve these two aims, we will install CfaC channel in the Salmonella cell outer membrane (OM). The channeled Salmonella will be evaluated for cell extension via atomic force microscope (AFM), viability in various conditions, morphology via Field Emission Scanning Electron Microscopy (FESEM), in vitro virulence via macrophages, and in vivo virulence by mouse model. Since the channel CFA/I in cell OM allows 741 dalton erythromycin to enter, it will also permit other smaller molecules, such as water, ions, and toxic molecules to pass. Thus, cell OM osmotic balance will be partially lost, which will result in the attenuation of bacterial pathogens. This study will possibly develop new live vaccines against Salmonella for human usage. PUBLIC HEALTH RELEVANCE: The proposed project entitled, "Evaluation of Protein Channel-Attenuated Salmonella Vaccines" will construct a live vaccine that aims at preventing human salmonellosis. Salmonella enterica is a disease that causes a variety of food and water-borne illnesses, such as gastroenteritis and typhoid fever. It can infect a wide range of animal hosts and also humans. Of the three common Typhoid fever disease, S. Typhi alone is estimated to cause approximately 16 million cases, with 600,000 deaths annually worldwide and 600 fatalities recently occurring each year in the United States. To make the situation even worse, this disease has acquired resistance to the antibiotics previously used to treat it. Recently, our laboratory invented a new strategy for inactivating the Salmonella. This new strategy is totally different from the conventionally used virulence gene inactivation method. If demonstrated effective in Salmonella, this strategy will be also applicable to other enterobacteria, as well. Therefore, this project will benefit not only the USA but also other countries epidemic with salmonellosis.

描述（由申请人提供）：该项目的长期目标涉及通过异源基因介导的失活而开发活疫苗，如使用定植因子抗原I（CFA/I）在沙门氏菌中获得的。我们怀疑这种衰减是因为CFAC通道的过度表达，而不是由于fimbriae CFABE引起的，尽管CFABE可能会增强通道介导的衰减。由于以前没有描述这种衰减策略，因此我们质疑它是否可以用于沙门氏菌疫苗的开发。由于基于基因失活的常规活疫苗开发方法已经存在障碍，因此这项拟议的研究将生成沙门氏菌活疫苗以供人类应用。因此，我们设定了以下两个特定目的：具体目标1：检查纤维亚基的缺失，分泌设备或两者是否负责观察到的野生型沙门氏菌的衰减。具体目标2：确定CFA/I操纵子的哪种缺失突变体会减弱野生型沙门氏菌，因此可以用作防御沙门氏菌病的防护疫苗。为了实现这两个目标，我们将在沙门氏菌细胞外膜（OM）中安装CFAC通道。将通过原子力显微镜（AFM）评估通道沙门氏菌的细胞延伸，在各种条件下的生存力，通过田间发射扫描电子显微镜（FESEM），通过巨噬细胞的体外毒力以及小鼠模型的体内毒力。由于细胞OM中的CFA/I通道允许741 Dalton红霉素进入，因此它还将允许其他较小的分子（例如水，离子和有毒分子）通过。因此，细胞OM渗透平衡将部分丢失，这将导致细菌病原体的衰减。这项研究可能会开发针对人类使用的沙门氏菌的新现场疫苗。公共卫生相关性：拟议的项目标题为“蛋白质通道衰减的沙门氏菌疫苗的评估”将建造一种旨在预防人类沙门氏菌病的现场疫苗。肠道沙门氏菌是一种疾病，会引起各种食物和水传播疾病，例如胃肠炎和伤寒。它可以感染各种动物宿主和人类。在三种常见的伤寒疾病中，据估计，单独的鼠伤寒链球菌将造成约1600万例病例，全球每年有60万例死亡，每年在美国每年发生600例死亡。为了使情况变得更糟，这种疾病已经获得了对先前用于治疗的抗生素的抗药性。最近，我们的实验室发明了一种使沙门氏菌失活的新策略。这种新策略与常规使用的毒力基因失活方法完全不同。如果证明在沙门氏菌中有效，此策略也将适用于其他肠杆菌。因此，该项目不仅将使美国，而且其他国家的杂种病流行。