An Attenuated E.coli Vaccine for Enterotoxigenic E.coli (ETEC)

针对产肠毒素大肠杆菌 (ETEC) 的减毒大肠杆菌疫苗

• 批准号: 7843474
• 负责人: EDGAR C. BOEDEKER
• 金额: $ 22.5万
• 依托单位: UNIVERSITY OF NEW MEXICO
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-05-15 至 2012-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7843474
• 关键词: Adjuvant; Adverse effects; Age; Agglutination; Animal Model; Animals; Antibodies; Antigens; Area; Attenuated; Attenuated Live Virus Vaccine; Attenuated Vaccines; Autopsy; Bacteria; Bacterial Adhesins; Biological Assay; Blood Cell Count; Blood Chemical Analysis; Body Weight decreased; Breathing; CD4 Positive T Lymphocytes; Cattle; Cause of Death; Cells; Child; Clinical; DNA; Developing Countries; Development; Diarrhea; Dose; Drainage procedure; Eating; Electron Microscopy; Enterotoxins; Enzyme-Linked Immunosorbent Assay; Escherichia coli EHEC; Escherichia coli Infections; Escherichia coli Vaccines; Eye; Future; Genetic Techniques; Health Benefit; Heating; Helper-Inducer T-Lymphocyte; IgG1; IgG3; Immune response; Immune system; Immunization; Immunoblotting; Immunoglobulin A; Immunoglobulin G; Individual; Infant; International; Intestines; Irrigation; Laboratories; Licensing; Life; Lung; Measures; Metabolic Clearance Rate; Microspheres; Modeling; Molecular Genetics; Monitor; Morbidity - disease rate; Mucosal Immune Responses; Mus; Nose; Operon; Organ; Plasmids; Posture; Property; Public Health; Research Personnel; Risk; Safety; Sampling; Serum; Severities; Shiga Toxin; Shivering; Structure of mucous membrane of nose; Tissues; Toxin; Traveler' s diarrhea; Vaccinated; Vaccination; Vaccine Antigen; Vaccines; Virulence; Virulence Factors; Work; bactericide; base; clinical effect; cytokine; drinking; enterotoxigenic Escherichia coli; field study; high risk; immunogenicity; indexing; infancy; meetings; mortality; mouse model; mutant; oral vaccine; pathogen; prevent; protective efficacy; prototype; public health relevance; response; safety testing; vaccine efficacy; vaccine evaluation; vaccine safety; vector; vector vaccine; volunteer

DESCRIPTION (provided by applicant): Enterotoxigenic Escherichia coli (ETEC) are among the primary causes of infantile and traveler's diarrhea. Despite the number and severity of ETEC infections worldwide, at present no licensed vaccine is available for at-risk individuals. We have developed a live attenuated E. coli vaccine strain by deleting or modifying major virulence determinants of a WT O157:H7 Enterohemorrhagic E.coli (EHEC) isolate. We now propose to use this strain as a vector to express ETEC antigens. We hypothesize that by delivering critical ETEC virulence determinants (adhesins and toxin components) to the mucosal immune system in an attenuated, non-invasive live vaccine strain, effective systemic and local immune responses can be generated which will protect against ETEC challenge. We will use an established mouse intranasal model of immunization and pathogen challenge to determine safety and immunogenicity and protective efficacy of our vaccine strains. Our first aim is to prepare derivatives of our attenuated attaching/effacing enterohemorrhagic E. coli (EHEC) vaccine that efficiently express components of two critical ETEC antigens, the CFA/I colonization factor and the heat-labile enterotoxin (LT). Our second aim is to determine a safe immunizing dose of the attenuated vaccine strains by monitoring immunized mice for adverse effects. Our third aim is to measure serum and mucosal immune responses following intranasal immunization with the vaccine constructs in order to determine the correlates of protection, and also of reactogenicity. Our fourth aim is to determine the level of protective efficacy resulting from immunization with the vaccine constructs using an established intra-pulmonary challenge model which we have previously used to determine the efficacy of other ETEC vaccine constructs. To achieve these ends, the expression of ETEC antigens by the vaccine constructs will be demonstrated by SDS-PAGE, immunoblotting, and bacterial agglutination. We will assess the safety of the vaccine constructs by noting any adverse clinical signs, and gross and histological changes at necropsy and by performing complete blood cell counts and blood chemistry in the vaccinated animals. We will assess the level of both systemic and local immune responses in the vaccinated mice by ELISA, cytokine responses and functional assays. We will determine protective efficacy by intranasal challenge with a lethal dose of the wild-type ETEC strain, as well as by measuring ETEC bacterial clearance from the lungs of the vaccinated mice. Development of safe and effective vaccines directed against ETEC strains should have significant public health benefit for infants in developing countries and for international travelers. PUBLIC HEALTH RELEVANCE: Enterotoxigenic E. coli (ETEC) are important bacterial pathogens causing worldwide morbidity and mortality. ETEC infections are important causes of death in infants and children under the age of five years in developing countries. ETEC are also the leading cause of diarrhea in travelers to high-risk areas of the world. Despite the fact that virulence factors of ETEC are well understood, and although in the last few decades there have been several potential ETEC vaccines tested in volunteer trials and field studies, no safe and effective vaccine is yet available for at-risk individuals. The development of safe and effective, live attenuated ETEC vaccines should have great public health significance for infants in developing countries and for international travelers.

描述（由申请人提供）：肠毒素大肠杆菌（ETEC）是婴儿和旅行者腹泻的主要原因之一。尽管全世界ETEC感染的数量和严重程度，但目前尚无授权疫苗可用于高危个人。我们通过删除或修改WT O157的主要毒力决定因素：H7 enthohemorrhagic e.coli（EHEC）分离株，开发了一种活跃的大肠杆菌疫苗菌株。现在，我们建议将此菌株用作表达ETEC抗原的载体。我们假设通过在粘膜免疫系统中传递关键的ETEC毒力决定因素（粘附素和毒素成分），可以产生可保护ETEC挑战的衰减，无创的活疫苗菌株，有效的全身和局部免疫反应。我们将使用已建立的小鼠免疫和病原体挑战的鼻内模型来确定疫苗菌株的安全性和免疫原性和保护作用。我们的第一个目的是制备我们减毒/剥落的肠肠肠肠肠球菌（EHEC）疫苗的衍生物，该疫苗有效地表达了两种关键ETEC抗原的成分，CFA/I cFA/I定量因子和热固定型肠毒素（LT）。我们的第二个目的是通过监测免疫小鼠的不良反应来确定衰减疫苗菌株的安全免疫剂量。我们的第三个目的是测量与疫苗构建体鼻内免疫后血清和粘膜免疫反应，以确定保护的相关性以及反应生成性的相关性。我们的第四个目的是使用已建立的肺内挑战模型来确定通过疫苗构建体免疫造成的保护功效水平，我们以前用来确定其他ETEC疫苗构建体的功效。为了达到这些目的，通过SDS-PAGE，免疫印迹和细菌凝集来证明ETEC抗原的表达。我们将通过注意任何不良临床体征以及在尸检时的严重和组织学变化来评估疫苗构建体的安全性，并在疫苗接种的动物中进行全血细胞计数和血液化学。我们将通过ELISA，细胞因子反应和功能测定法评估接种小鼠的全身和局部免疫反应水平。我们将用野生型ETEC菌株的致命剂量以及测量接种小鼠肺的ETEC细菌清除率来确定鼻内挑战的保护效果。开发针对ETEC菌株的安全有效疫苗应为发展中国家和国际旅客的婴儿带来重大的公共卫生益处。公共卫生相关性：肠毒素大肠杆菌（ETEC）是重要的细菌病原体，导致全球发病率和死亡率。 ETEC感染是发展中国家五岁以下的婴儿和儿童死亡的重要原因。 ETEC也是旅行者到世界高风险地区的腹泻的主要原因。尽管ETEC的毒力因子已经充分了解了，尽管在过去几十年中，在志愿者试验和现场研究中已经测试过几种潜在的ETEC疫苗，但尚无安全有效的疫苗可用于处于危险中。安全有效，活跃的ETEC疫苗的开发对于发展中国家和国际旅行者的婴儿应具有很高的公共健康意义。