Function of Longus Type IV pili in enterotoxigenic E coli

IV型长菌毛在产肠毒素大肠杆菌中的功能

• 批准号: 7900470
• 负责人: Oscar G GOMEZ
• 金额: $ 12.74万
• 依托单位: UNIVERSITY OF IOWA
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-09-03 至 2011-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7900470
• 关键词: Adherence; Amino Acid Sequence Homology; Animal Model; Animals; Antibodies; Antigens; Attenuated; Binding; Biological Assay; Biological Models; Cell surface; Cells; Childhood; Developing Countries; Development; Diarrhea; Disease; Escherichia coli Vaccines; Evaluation; Exhibits; Fimbriae Proteins; Genes; Geographic Locations; Goals; Histology; Human; Immune response; Infection; Intestines; Knowledge; Life; Mediating; Microbial Biofilms; Molecular; Mus; Organelles; Pathogenesis; Phenotype; Pilum; Plasmids; Prevention; Process; Proteins; Pseudomonas aeruginosa; Role; SECTM1 gene; Salmonella; Serum; Severe Combined Immunodeficiency; Shapes; Site-Directed Mutagenesis; Structural Protein; Structure; Surface; Testing; Time; Traveler' s diarrhea; Vaccines; Variant; Vibrio cholerae; Virulence; Work; base; cell motility; cross reactivity; enteropathogenic Escherichia coli; enterotoxigenic Escherichia coli; gene cloning; immunogenic; immunogenicity; in vivo; interest; mouse model; mutant; novel; pathogen; polymerization; prevent; response; vaccine candidate; vector

DESCRIPTION (provided by applicant): Enterotoxigenic Escherichia coli (ETEC) is a leading cause of childhood diarrhea in developing countries and the most important cause of traveler's diarrhea. Effective vaccines for ETEC are not currently available. ETEC intestinal colonization is a crucial first step in the infection process, and it is believed to be mediated by species-specific pili. Longus (long pilus), a recently identified type IV pili (T4P) and one of the most prevalent pili described in human ETEC strains, has structural similarity and protein sequence homology to the well known virulence-associated T4Ps expressed by important human bacterial pathogens. The long-term goals of this project are to identify the role of Longus in ETEC colonization of the human gut and to develop vaccine candidates to prevent ETEC diarrhea. I hypothesize that Longus contributes to colonization of human intestine by binding to cells or non-cell surfaces, and that Longus-based vaccines, capable of inducing anti-Longus antibodies, will block ETEC colonization and prevent disease. To test this hypothesis I propose the following specific aims: 1) Identify the genes necessary for Longus expression and evaluate Longus-gene variation, 2) Evaluate Longus-mediated adherence and colonization of to human intestinal surfaces, and 3) Test immunogenicity of Longus in a mouse model. I will first clone the genes necessary for Longus expression, evaluate gene variation among worldwide isolates, and construct a Longus structural subunit mutant essential for the evaluation of Longus phenotypes. Wild type and mutants will be compared for cell adherence to human intestinal cells, and in biofilm formation assays. ETEC colonization will be evaluated by using a novel human intestine explant-Severe combined immunodeficiency (SCID) mouse model. The explant will be infected with ETEC and evaluated for colonization by colony count and histology. Longus immunogenicity and cross-reactivity with longus variants will be tested on sera from immunized mice with Longus-expressing attenuated Salmonella. Demonstration that Longus mediates binding to cells and biofilm formation will greatly contribute to our knowledge on Longus function. The development of a human explant/SCID mouse model will have crucial implications for the understanding ETEC colonization at the molecular level. Information that Longus is immunogenic across variants and that anti-Longus antibodies may block colonization and prevent disease will facilitate the construction of effective ETEC vaccines.

描述（由申请方提供）：产肠毒素大肠杆菌（ETEC）是发展中国家儿童腹泻的主要原因，也是旅行者腹泻的最重要原因。目前还没有有效的ETEC疫苗。ETEC肠道定植是感染过程中至关重要的第一步，据信由种属特异性皮利介导。Longus（长菌毛）是最近鉴定的IV型皮利（T4 P），也是人类ETEC菌株中描述的最普遍的皮利之一，与重要人类细菌病原体表达的众所周知的毒力相关T4 P具有结构相似性和蛋白质序列同源性。该项目的长期目标是确定Longus在人类肠道ETEC定植中的作用，并开发预防ETEC腹泻的候选疫苗。我假设Longus通过与细胞或非细胞表面结合促进人类肠道定植，并且能够诱导抗Longus抗体的基于Longus的疫苗将阻断ETEC定植并预防疾病。为了验证这一假设，我提出了以下具体目标：1）鉴定Longus表达所必需的基因并评价Longus基因变异，2）评价Longus介导的对人肠道表面的粘附和定殖，以及3）在小鼠模型中测试Longus的免疫原性。我将首先克隆Longus表达所需的基因，评估全球分离株之间的基因变异，并构建Longus表型评估所必需的Longus结构亚基突变体。将比较野生型和突变体对人肠细胞的细胞粘附，以及生物膜形成测定。将使用新型人肠道免疫缺陷-重度联合免疫缺陷（SCID）小鼠模型评价ETEC定植。将用ETEC感染外植体，并通过菌落计数和组织学评价定植。将在来自用表达Longus的减毒沙门氏菌免疫的小鼠的血清上测试Longus免疫原性和与Longus变体的交叉反应性。证明Longus介导与细胞的结合和生物膜形成将极大地有助于我们对Longus功能的了解。人外植体/SCID小鼠模型的开发将对在分子水平上理解ETEC定植具有重要意义。Longus在变体中具有免疫原性，抗Longus抗体可阻断定植并预防疾病，这一信息将有助于构建有效的ETEC疫苗。

项目成果

Murine immunization with CS21 pili or LngA major subunit of enterotoxigenic Escherichia coli (ETEC) elicits systemic and mucosal immune responses and inhibits ETEC gut colonization.

使用产肠毒素大肠杆菌 (ETEC) 的 CS21 pili 或 LngA 主要亚基对小鼠进行免疫接种，可引发全身和粘膜免疫反应，并抑制 ETEC 肠道定植。

• DOI: 10.1016/j.vetmic.2016.02.001
• 发表时间: 2017-04
• 期刊: Veterinary microbiology
• 影响因子: 3.3
• 作者: Zhang C;Iqbal J;Gómez-Duarte OG
• 通讯作者: Gómez-Duarte OG

Detection of Escherichia coli enteropathogens by multiplex polymerase chain reaction from children's diarrheal stools in two Caribbean-Colombian cities.

• DOI: 10.1089/fpd.2009.0355
• 发表时间: 2010-02
• 期刊: Foodborne pathogens and disease
• 影响因子: 2.8
• 作者: Gómez-Duarte OG;Arzuza O;Urbina D;Bai J;Guerra J;Montes O;Puello M;Mendoza K;Castro GY
• 通讯作者: Castro GY