Role of TraJ in Neonatal E. coli Sepsis and Meningitis

描述（由申请方提供）：尽管使用了现代治疗和诊断措施，新生儿细菌性败血症和脑膜炎仍然是一种发病率和死亡率不可接受的疾病。这种疾病的死亡率为15- 75%;此外，大约50%的脑膜炎幸存者有明显的神经和发育异常。E.大肠杆菌K1（革兰氏阴性脑膜炎的最常见病原体）是口服的。肠道定植后，细菌通过胃肠道移位至肠系膜淋巴结（MLN）、肝脏、脾脏和血液的肠外部位。E.然后，大肠杆菌K1在血流中系统性繁殖，达到菌血症的必要阈值，以进入中枢神经系统（CNS）。最近，我们发现了一个E。coliK 1质粒编码的血脑屏障（BBB）侵袭基因traJ与多种F-样质粒tra操纵子的traJ具有同源性。我们的初步数据表明，traJ突变特异性减弱从MLN到肝，脾，血液和CNS的新生大鼠的传播。此外，尽管经口感染traJ突变体的动物在肝脏或脾脏中表现出减少或没有可恢复的细菌，但这些组织显示出显著的炎症反应。体外研究确定，traJ突变体被巨噬细胞摄取较少，并显示出55 kDa分泌蛋白的损失。本申请的中心假设是TraJ控制一组基因的表达，这些基因的产物（即，55 kDa分泌蛋白）参与E.大肠杆菌K1传播，全身感染和穿越血脑屏障，这些事件在疾病过程中发生的TraJ依赖性的相互作用与专业吞噬细胞。下面提出的实验旨在测试和证实我们的假设。本研究的目的是：1）阐明traJ基因和质粒的遗传和分子特征，评估内源质粒的自转移能力，并确定含traJ基因的质粒在大肠杆菌中的潜在作用; coli K1新生大鼠毒力，2）评价TraJ调节蛋白的功能（即，55 kDa分泌蛋白）。coli K1全身传播和脑膜炎，以及3）确定新生大鼠中TraJ依赖性宿主炎症反应的机制。提高对早期全身传播的分子机制的认识以及在这些事件中宿主炎症反应的确切相互作用将有助于实现我们的长期目标，即确定新的合理方法，以开发新的治疗方法和预防策略。大肠杆菌K1败血症和脑膜炎。

DESCRIPTION (provided by applicant): Despite the use of modern therapeutics and diagnostic measures, neonatal bacterial sepsis and meningitis continues to be a disease with unacceptable rates of morbidity and mortality. The fatality rate of this disease is 15-75 percent; furthermore, approximately 50 percent of meningitis survivors have significant neurological and developmental abnormalities. The primary, route of infection for E. coli K1 (the most common causative agent of gram-negative meningitis) is oral. Following intestinal colonization, bacteria translocate through the GI tract to extra-intestinal sites of mesenteric lymph nodes (MLN), liver, spleen, and the blood. E. coli K1 then multiply systemically within the bloodstream reaching a necessary threshold of bacteremia to gain access to the central nervous system (CNS). Recently we identified an E. coli K1 plasmid-encoded blood-brain barrier (BBB) invasion gene, traJ, with homology to traJ of various F-like plasmid tra operons. Our preliminary data indicate that the traJ mutation specifically attenuates dissemination from the MLN to the liver, spleen, blood, and the CNS in the neonatal rat. In addition, although animals orally infected with the traJ mutant demonstrated a decrease or no recoverable bacteria in the liver or spleen, these tissues showed a significant inflammatory response. In vitro studies determined that the traJ mutant is taken-up less by macrophages and shows a loss of a 55 kDa-secreted protein. The central hypothesis of this application is that TraJ controls the expression of a set of genes whose products (i.e., 55 kDa secreted protein) are involved in E. coli K1 dissemination, systemic infection and crossing the BBB, and these events within the disease process occur via TraJ-dependent interactions with professional phagocytes. The following proposed experiments are designed to test and substantiate our hypotheses. We aim to 1) Elucidate the genetic and molecular characteristics of the traJ region and plasmid, evaluate the ability of the endogenous plasmid to self-transfer, and determine the potential role of the traJ-containing plasmid in E. coli K1 neonatal rat virulence, 2) Evaluate the function of the TraJ-regulated proteins (i.e., 55 kDa-secreted protein) in E. coli K1 systemic dissemination and meningitis, and 3) Determine the mechanism of TraJ-dependent host inflammatory response in the neonatal rat. Improved knowledge of molecular mechanisms for early systemic dissemination and the exact interplay of the host inflammatory response during these events will assist in achieving our long-term goal of identifying novel rational approaches to development of new treatments and preventive strategies for E. coli K1 sepsis and meningitis.