Studies of the capsular-like antigen of F. tularensis

土拉热弗朗西丝菌是一种A类细菌选择剂，由于其极端的致病性和潜在的用途作为生物武器。卡莱尔和胡德的早期研究表明，F.土拉热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基因的同源物存在于弗朗西斯菌属基因组数据库中。初步研究，使用电子显微镜，表明并确认，胶囊状材料（CLM）包围F。土拉热。我们现在可以根据化学、色谱和免疫化学分析分离出这种不含弗朗西斯菌LPS的物质。该物质与细菌细胞松散结合，容易从细菌表面去除，并且由重复的四糖重复序列组成。利用转座子诱变技术，在F.在土拉热菌Schu S4中，我们已经鉴定了许多对我们的CLM特异性抗体XE 8没有反应性或具有有限反应性的突变体。我们目前已经确定了7个转座子突变体的插入位点，这些突变体改变了CLM的表达，将作为本提案中研究的无囊菌株。基于这些观察，我们提出以下假设：1）土拉热弗朗西丝菌表达一种在发病机制中很重要的荚膜样物质，我们认为它是第1组荚膜; 2）通过参与CLM生物合成和表达的基因突变改变这种荚膜样结构将改变土拉热弗朗西丝菌在细胞培养模型和呼吸道感染动物模型中的致病性; 3）靶向CLM的免疫应答的诱导将提供宿主免疫。以下具体目标将被用来解决这些假设：1）表征的F。在产生囊样物质方面有缺陷的土拉热杆菌Schu S4突变体和2）土拉热杆菌Schu S4囊样物质的生物学作用的表征和3）在弗朗西斯菌感染的鼠模型中研究被动和主动免疫的免疫原性和保护效力

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