Immune recognition of Klebsiella pneumoniae O2v1 and O2v2 O-antigen subtypes

肺炎克雷伯菌 O2v1 和 O2v2 O 抗原亚型的免疫识别

• 批准号: 10739041
• 负责人: David A. Rosen
• 金额: $ 23.33万
• 依托单位: WASHINGTON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-08-01 至 2025-07-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10739041
• 关键词: Accounting; Acute; Affect; Antibiotic Resistance; Antibiotic-resistant organism; Antibiotics; Antibodies; Antibody Response; Antigens; Bacteria; Bacteriology; Biological Assay; Body Weight decreased; Cessation of life; Clinical; Complement; Conjugate Vaccines; Development; Disease; Enzyme-Linked Immunosorbent Assay; Europe; Exhibits; Flow Cytometry; Future; Galactans; Galactose; Glycosyltransferase Gene; Goals; Gram-Negative Bacteria; Hospitalization; Hospitals; Human; Humoral Immunities; Immune; Immune response; Immunization; Immunoglobulin G; Immunologic Techniques; In Vitro; Infection; Infection prevention; Innate Immune Response; Klebsiella pneumoniae; Knowledge; Lipopolysaccharides; Literature; Lung; Measures; Mediating; Microbial Biofilms; Monitor; Mus; Natural Immunity; O Antigens; Organism; Ozone; Pathogenesis; Pathogenicity; Patients; Phagocytosis; Phase I Clinical Trials; Phenotype; Pneumonia; Polysaccharides; Predisposition; Production; Research Project Grants; Research Project Summaries; Resistance; Respiratory System; Respiratory Tract Infections; Sepsis; Serum; Structure; Surface; Testing; Urinary tract infection; Vaccinated; Vaccination; Vaccine Design; Vaccines; Vertebral column; Virulence; Virulence Factors; Western Blotting; adaptive immune response; bacterial fitness; bacterial resistance; bactericide; capsule; carbapenem resistance; carbapenemase; combat; cross immunity; cross reactivity; cytokine; experimental study; fitness; healthcare-associated infections; immunogenic; immunogenicity; in vivo; innate immune function; interest; mortality; mouse model; mutant; neutrophil; novel; pathogen; pneumonia model; prevent; resistant Klebsiella pneumoniae; response; vaccine development

PROJECT SUMMARY This research project will enhance our understanding of the immune responses elicited by the O-antigen of Klebsiella pneumoniae (Kp), an important target in putative vaccine development. The long-term goal of this proposal is to further our knowledge of the innate and adaptive immune responses elicited by two subtypes of the common serogroup O2 of Kp, O2v1 and O2v2. We seek to understand how the addition of the single branched galactose of O2v2 changes its bacterial fitness and host immune susceptibility relative to O2v1. Further, we will determine cross-reactivity between these two subtypes and whether infection or vaccination with one subtype confers cross-protection against the other, information critical to the design of vaccines that can broadly target this increasingly antibiotic-resistant organism. Kp infections, including pneumonia, urinary tract infection, and bloodstream infection, are sharply on the rise among hospitalized patients; CDC has declared that infections with Kp and other carbapenem-resistant Enterobacteriales demand a threat level of urgent. This project builds on the PI's background in bacteriology, murine models of infection, and pathogenesis studies of Gram-negative bacteria, by focusing specifically on the Kp virulence factor O-antigen and the host immunogenic response. We have previously demonstrated that use of bioconjugate vaccines targeting Kp are beneficial in preventing disease and seek to increase our understanding and breadth of coverage of future vaccines. Kp historically has eleven known serogroups of O-antigen. Recently, additional O-antigen subtypes within these serogroups have been identified. Yet, differences in pathogenic fitness, immunogenicity, functional antibody response, and cross-protection between related subtypes are not well understood. We have constructed a mutant of a classical Kp strain expressing O-antigen subtype O2v2 to generate an otherwise isogenic strain expressing O2v1. We will test if these bacteria exhibit similar phenotypic expression of virulence factors by measuring capsule, hypermucoviscosity, fimbriae and biofilm. A well-established murine model of pneumonia will be leveraged to determine pulmonary fitness and delineate host innate immune response to each pathogen. Neutrophil and complement-mediated bacterial killing assays will be employed to further explore bacterial resistance to innate immune attack, which the literature indicates may be enhanced in O2v2 strains. Next, we will perform a range of experiments testing if O2v1 and O2v2 are cross-protective. Using classically immunological techniques including ELISA and flow cytometry, we will determine the IgG subclasses and effector cytokines elicited by respiratory tract infection with these pathogens. Lastly, using novel O2v1 or O2v2 bioconjugate vaccines followed by challenge with O2v1 or O2v2 strains, we will assess cross-protection against weight loss and mortality from pneumonia. We will further characterize antibody functionality using serum bactericidal and opsonophagocytic killing assays. These studies will significantly advance our understanding of immune response to Kp O-antigen and aid in vaccine design to combat this pathogen.

项目摘要 这项研究项目将加强我们对O抗原引起的免疫反应的理解， 肺炎克雷伯氏菌（Kp）是公认疫苗开发的重要靶标。长期目标是 我们的建议是进一步了解先天性和适应性免疫反应引起的两个亚型， Kp、O2 v1和O2 v2的共同血清群O2。我们试图了解如何增加单一的 相对于O2 v1，O2 v2的支链半乳糖改变了其细菌适应性和宿主免疫易感性。 此外，我们将确定这两种亚型之间的交叉反应性，以及是否感染或接种了 一种亚型赋予了对另一种亚型的交叉保护，这是设计疫苗的关键信息， 广泛地针对这种越来越具有抗药性的生物体。Kp感染，包括肺炎、尿路感染 感染和血液感染，在住院患者中急剧上升; CDC宣布， 肺炎克雷伯氏菌和其他碳青霉烯类耐药肠杆菌的感染需要紧急的威胁水平。 这个项目建立在PI在细菌学、小鼠感染模型和发病机制研究方面的背景之上 革兰氏阴性菌，通过专门关注Kp毒力因子O-抗原和宿主 免疫原性反应。我们先前已经证明，使用靶向Kp的生物缀合物疫苗是有效的。 有利于预防疾病，并寻求增加我们的理解和覆盖面的未来 疫苗。Kp历史上有11个已知的O抗原血清群。最近，其他O抗原亚型 在这些血清群中已经被鉴定。然而，在致病适应性、免疫原性、功能性 抗体应答和相关亚型之间的交叉保护还不清楚。 我们构建了表达O-抗原亚型O2 v2的经典Kp菌株的突变体，以产生一个表达O-抗原亚型O2 v2的突变体。 另外表达O2 v1的同基因菌株。我们将测试这些细菌是否表现出类似的表型表达， 毒力因子通过测量荚膜、高粘滞性、菌毛和生物被膜。一种成熟的小鼠 将利用肺炎模型来确定肺适应性并描绘宿主先天免疫 对每一种病原体的反应。将采用中性粒细胞和补体介导的细菌杀伤试验， 进一步探索细菌对先天免疫攻击的抵抗力，文献表明， O2 v2菌株。接下来，我们将进行一系列实验，测试O2 v1和O2 v2是否具有交叉保护作用。使用 经典的免疫学技术包括ELISA和流式细胞术，我们将确定IgG亚类 以及由这些病原体的呼吸道感染引起的效应细胞因子。最后，使用新的O2 v1或 O2 v2生物结合疫苗，然后用O2 v1或O2 v2毒株攻毒，我们将评估交叉保护 对抗体重减轻和肺炎死亡率。我们将使用血清进一步表征抗体功能 杀菌和调理吞噬杀伤测定。这些研究将大大促进我们对 对Kp O-抗原免疫应答并有助于设计对抗该病原体的疫苗。