Studies of the capsular-like antigen of F. tularensis

土拉弗拉菌荚膜样抗原的研究

• 批准号: 7920674
• 负责人: Michael A. Apicella
• 金额: $ 41.56万
• 依托单位: UNIVERSITY OF IOWA
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-08-01 至 2015-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7920674
• 关键词: Active Immunization; Acylation; Address; Affect; Alveolar; Anabolism; Animal Model; Antibodies; Antigens; Bacteria; Bacterial Capsules; Binding; Biological; Biological Warfare; Cell Culture Techniques; Cell Wall; Cells; Chemicals; Classification; DNA; Data; Detergents; Electron Microscopy; Epithelial Cells; Epitopes; Francisella; Francisella tularensis; Galactose; Gene Mutation; Genes; Glucose; Heptoses; Homologous Gene; Human; Immune response; Immunity; Immunization; Immunofluorescence Immunologic; Infection; Investigation; Lead; Length; Lipid A; Lipoprotein (a); Mannose; Methods; Microscopic; Modeling; Monoclonal Antibodies; Morphology; Mus; Mutagenesis; Mutate; Mutation; O Antigens; Organism; Passive Immunity; Pathogenesis; Pathogenicity; Polymers; Population; Preparation; Production; Regulation; Regulator Genes; Respiratory Tract Infections; Rhamnose; Role; Ruthenium Red; Site; Solid; Staining method; Stains; Structure; Surface; System; T-Lymphocyte; Time; Tularemia; Virulence; Virulence Factors; Western Blotting; Work; bactericide; base; capsule; carbohydrate structure; genome database; immunogenicity; killings; macrophage; monocyte; mouse model; mutant; protective efficacy; protein transport; sugar; therapeutic development; uptake; vaccine candidate; weapons of mass destruction

Francisella tularensis is a class A bacterial select agent due to its extreme pathogenicity and potential use as a bioweapon. Early studies by Carlisle and Hood have indicated that F. tularensis produces a capsule-like material. Many bacteria produce capsules and the genes required to produce them encode proteins for transport, biosynthesis and regulation. Homologs of genes implicated in capsular biosynthesis are present in the Francisella genome database. Preliminary studies, using electron microscopy, indicate and confirm that a capsule-like material (CLM) surrounds F. tularensis. We can now isolate this material, free from Francisella LPS, based on chemical, chromatographic and immunochemical analysis. This material is loosely associated with the bacterial cell, is easily removed from the bacterial surface and is composed of a repeating tetrasaccharide repeat. Using transposon mutagenesis in F. tularensis Schu S4, we have identified a number of mutants with no or limited reactivity to our CLM specific antibody XE8. We have currently identified the site of insertion of 7 of these transposon mutants that have altered CLM expression that will serve as acapsular strains for the studies in this proposal. Based on these observations, we would pose the following hypotheses 1) Francisella tularensis expresses a capsular-like material that is important in pathogenesis and that we believe is a group 1 capsule; 2) Alteration of this capsule-like structure by mutations of the genes involved in biosynthesis and expression of CLM will alter pathogenicity of Francisella tularensis in cell culture models and in an animal model of respiratory infection; 3) Induction of an immune response targeted to the CLM will provide host immunity. The following specific aims will be used to resolve these hypotheses: 1) Characterization of the F. tularensis Schu S4 mutants that are defective in production of capsule-like material and 2) Characterization of the biological role of the P. tularensis Schu S4 capsule like material and 3) Study the immunogenicity and protective efficacy of passive and active immunization in murine models of Francisella infection

土拉菌因其极强的致病性和潜在的生物武器用途而被列为a类细菌选择剂。卡莱尔和胡德的早期研究表明，土拉菌会产生一种胶囊状物质。许多细菌产生胶囊，而产生胶囊所需的基因编码用于运输、生物合成和调节的蛋白质。与荚膜生物合成有关的同源基因存在于弗朗西斯菌基因组数据库中。使用电子显微镜的初步研究表明并证实了土拉菌周围有一种胶囊状物质（CLM）。我们现在可以通过化学，色谱和免疫化学分析，从Francisella LPS中分离出这种物质。这种物质与细菌细胞松散结合，很容易从细菌表面去除，由重复的四糖重复组成。利用转座子诱变法，我们在土拉菌Schu S4中发现了许多对CLM特异性抗体XE8没有或有限反应的突变体。我们目前已经确定了7个这些转座子突变体的插入位点，这些突变体改变了CLM的表达，将作为本提案研究的囊性菌株。基于这些观察结果，我们将提出以下假设：1)土拉菌表达一种囊状物质，这在发病机制中很重要，我们认为这是1组胶囊；2)在细胞培养模型和呼吸道感染动物模型中，参与CLM生物合成和表达的基因突变改变这种胶囊状结构将改变土拉弗朗西斯菌的致病性；3)诱导针对CLM的免疫应答将提供宿主免疫。为了解决这些假设，我们将进行以下具体的研究：1)鉴定在生产囊状物质方面存在缺陷的土拉菌Schu S4突变体；2)鉴定土拉菌Schu S4囊状物质的生物学作用；3)研究弗朗西斯菌感染小鼠模型中被动免疫和主动免疫的免疫原性和保护作用