Bordetella Biofilms and Pathogenesis

描述(申请人提供)：百日咳由百日咳波尔德氏菌引起，尽管疫苗接种覆盖率很高，但在美国仍然是一个相当大的经济和健康负担。目前的百日咳疫苗虽然在预防严重形式的疾病方面有效，但不能防止携带者状态。成人和青少年携带者负责将细菌传播给婴儿和幼儿，他们的疾病很严重，有时甚至是致命的。迫切需要确定新的因素，这些因素将防止细菌携带，并将包括在无细胞疫苗中。据推测，人类鼻咽携带百日咳杆菌是因为它能够以一种被称为生物膜的固着形式存在。我们的初步结果表明，百日咳杆菌以多细胞群落的形式存在于小鼠的鼻腔上皮上，这使人联想到生物膜。本研究的总体目标是研究生物膜相关的BPS多糖在生物膜发育和发病机制中的作用，并深入了解控制BPS在生物膜和呼吸道中表达的调控电路。BPS与其他细菌的PGA/PIA/PNAG(聚-2-1，6-乙酰氨基葡萄糖)多糖相似，包括许多病原菌。在具体目标1中，将利用与生物学相关的体外模型系统来研究BPS在生物膜形成中作用的机制基础。BPS在提供抗微生物化合物保护方面的重要性将通过测试其在赋予对抗生素、抗菌肽和活性氧物种的抗药性方面的作用来检验。据推测，BPS通过发挥免疫调节作用和靶向免疫系统的一个或多个成分，介导百日咳杆菌在呼吸道的存活和持续。百日咳杆菌鼻腔感染的小鼠模型将被用来检验这些假设。在具体目标2中，将通过实时RT-PCR分析和GFP荧光测量来研究BPS在生物被膜形成过程和呼吸道中表达的时空梯度的建立。最后，为了深入了解控制BP表达的复杂网络，我们将研究调节蛋白BpsR在生物膜和小鼠呼吸道中控制BP基因表达的作用。与公共卫生的相关性。该项目将为更好地了解百日咳杆菌在哺乳动物宿主中的致病机制和感染循环提供依据。我们的研究将促进基于BPS的疫苗和试剂的开发，如用于治疗百日咳的抗体。由于BPS类多糖是由葡萄球菌、大肠杆菌和耶尔森氏菌等多种病原体产生的，我们的研究结果将有助于治疗和了解各种细菌感染。

DESCRIPTION (provided by applicant): Pertussis, caused by Bordetella pertussis remains a considerable economic and health burden in the USA, despite very high coverage of vaccination. Current pertussis vaccines although effective at preventing the severe form of disease do not prevent the carrier state. Adult and adolescent carriers are responsible for the transmission of the bacterium to infants and young children, in whom the disease is severe and sometimes lethal. There is an urgent need to identify new factors that will be protective against bacterial carriage and will be included in the acellular vaccine. It is hypothesized that nasopharyngeal carriage of B. pertussis in humans is due to its ability to exist in a sessile form known as biofilms. Our preliminary results demonstrate that B. pertussis exists in mice as multicellular community adherent to the nasal epithelium, which are reminiscent of biofilms. The overall objectives of this research are to study the role of the biofilm-associated Bps polysaccharide in biofilm development and pathogenesis and to gain insight into the regulatory circuitry that controls bps expression in biofilms and in the respiratory tract. Bps is similar to PGA/PIA/PNAG (poly-2-1,6-N- acetylglucosamine) polysaccharides of other bacteria including a number of pathogens. In Specific Aim 1, biologically relevant in vitro model systems will be utilized to study the mechanistic basis of the role of Bps in biofilm formation. The importance of Bps in providing protection from anti-microbial compounds will be examined by testing its role in conferring resistance against antibiotics, antimicrobial peptides and reactive oxygen species. It is hypothesized that Bps mediates survival and persistence of B. pertussis in the respiratory tract by playing an immunomodulatory role and by targeting one or more components of the immune system. Mouse models of B. pertussis intranasal infection will be employed to test these hypotheses. In Specific Aim 2, the establishment of temporal and spatial expression gradient of bps during biofilm formation and in the respiratory tract will be examined by real time RT PCR assays and measurement of GFP fluorescence. Finally, to provide insights into the complex networks that operate to control bps expression, the role of a regulatory protein BpsR in controlling the expression of the bps locus in biofilms and in the mouse respiratory tract will be examined. Relevance to public health. This project will provide a better understanding of the pathogenic mechanisms and the infectious cycle of B. pertussis in mammalian hosts. Our studies will stimulate the development of Bps-based vaccines and reagents like antibodies for treatment of pertussis. Since Bps-like polysaccharides are produced by a number of pathogens like Staphylococci, E. coli and Yersinia Spp, and possibly others, our results will contribute towards the treatment and the understanding of a wide variety of bacterial infections.