Nanobodies for the Detection and Neutralization of Listeria monocytogenes

描述（由申请人提供）：李斯特菌病是一种由单核细胞增生李斯特菌引起的严重食源性疾病，是一个重大的公共卫生问题。据估计，美国每年有 1600 例李斯特菌病病例，导致 250 人死亡。除了这种高死亡率之外，怀孕期间感染还可能导致流产或死产。及早检测食品供应中的病原体是保护公众健康的关键。近年来李斯特菌病的反复爆发凸显了开发快速、灵敏且廉价的生物传感器的必要性。此外，感染孕妇的治疗需要一个疗程的高侵入性静脉注射抗生素。一种廉价且有效的预防方法可以防止李斯特菌定植，这将是产前护理的有力补充。我们建议使用纳米抗体来解决防御李斯特菌病的这些关键漏洞。纳米抗体是源自骆驼科物种（骆驼、羊驼、美洲驼）的独特免疫系统库的重链单域抗体。纳米抗体生产成本低廉、高度稳定且具有独特的结合模式，使其特别适合开发新的李斯特菌生物传感器和疗法。该项目的目标是了解源自原始骆驼科噬菌体展示文库的纳米抗体如何与李斯特菌表面抗原结合并相互作用，为开发新型李斯特菌生物传感器和疗法奠定基础。我们的初步研究已经确定了四种纳米抗体，它们与表面暴露且李斯特菌毒力因子所需的靶标结合。通过结构生物学和结合研究，我们将评估源自组合的幼稚骆驼科噬菌体展示文库与李斯特菌毒力因子 InlB (AIM 1) 的纳米抗体识别的表位范围。我们还将利用微生物学和结构研究来检验骆驼科纳米抗体检测和中和李斯特菌（AIM 2）的潜力。

DESCRIPTION (provided by applicant): Listeriosis is a severe food-borne disease caused by the bacterium Listeria monocytogenes and is a significant public health concern. There are an estimated 1600 cases of Listeriosis per year in the US resulting in 250 deaths. In addition to this high mortality rate infection during pregnancy can lead to miscarriage or stillbirth. Early detection of the pathogen in the food supply is key to protecting public health. Repeated outbreaks of Listeriosis in recent years highlight the need for development of a rapid, sensitive and inexpensive biosensor. In addition, treatment of infected pregnant women requires a course of highly invasive intravenously administered antibiotics. An inexpensive and effective prophylactic that could prevent Listeria colonization would be a powerful addition to prenatal care. We propose using nanobodies to address these critical holes in the defense against Listeriosis. Nanobodies are heavy chain, single domain antibodies derived from the unique immune system repertoires found in Camelidae species (camels, alpacas, llamas). Nanobodies are inexpensive to produce, highly stable, and have distinctive binding modes making them uniquely suited to developing new Listeria biosensors and therapeutics. The goal of this project is to understand how nanobodies derived from a naïve Camelidae phage display library bind and interact with Listeria surface antigens, to lay the foundation for development of novel Listeria biosensors and therapeutics. Our preliminary studies have identified four nanobodies that bind to a target that is both surface exposed and required for Listeria virulence factor. Usin structural biology and binding studies we will assess the range of epitopes recognized by nanobodies derived from a combined naïve Camelidae phage display library to Listeria virulence factor InlB (AIM 1). We will also employ microbiological and structural studies to examine the potential of Camelidae nanobodies to detect and neutralize Listeria (AIM 2).