Expression of sap Operon in NTHI-induced Otitis Media

描述（由申请人提供）：细菌已经进化出抵抗宿主防御机制的策略，包括对抗菌肽（AP）的抗性。 sap（对抗菌肽的敏感性）操纵子在这种防御中起着重要作用，如动物模型中sap操纵子突变体的衰减所证明的。该提议的假设是，sap操纵子通过维持体内对AP的抵抗力和上调特定AP抵抗决定因素的表达，在中耳炎灰鼠模型中不可分型流感嗜血杆菌（NTHI）的生存中发挥着重要作用。将通过流式细胞术和组织学分析中耳炎病程期间或体外暴露于灰鼠嗜中性粒细胞或嗜中性粒细胞颗粒提取物时sap操纵子的表达。NTHI sap操纵子的六个基因中的突变将定义每个基因在体内的功能要求。最后，微阵列分析将被用来确定作为SAP操纵子表达的结果的决定对AP的抗性的基因。从这些研究中产生的数据将显着推进该领域的理解NTHI如何适应在疾病进展过程中遇到的微环境中的体内先天免疫反应，并将确定新的毒力机制，导致SAP操纵子表达，使NTHI生存和引起疾病。

DESCRIPTION (provided by applicant): Bacteria have evolved strategies to resist host defense mechanisms including resistance to antimicrobial peptides (APs). The sap (sensitivity to antimicrobial peptides) operon plays a significant role in this defense, as demonstrated by attenuation of sap operon mutants in animal models. The hypothesis of this proposal is that the sap operon plays a significant role in nontypeable Haemophilus influenzae (NTHI) survival in a chinchilla model of otitis media by maintaining resistance to APs in vivo and by up-regulating expression of specific AP resistance determinants. Expression of the sap operon during the disease course of otitis media or when exposed in vitro to chinchilla neutrophils or neutrophil granule extract will be analyzed by flow cytometry and histology. Mutations in the six genes of the NTHI sap operon will define the functional requirement for each gene in vivo. Finally, microarray analysis will be used to identify genes that determine resistance to APs as a result of sap operon expression. The data generated from these studies will significantly advance the field in understanding how NTHI adapt to the innate immune response in vivo in microenvironments encountered during disease progression and will identify novel virulence mechanisms resulting from sap operon expression that enable NTHI to survive and cause disease.