Immune Evasion Mechanisms of Mycoplasma genitalium

项目摘要 生殖支原体（MG）是一种新出现的生殖道病原体，具有重大的公共卫生问题。 这种性传播细菌引起的疾病谱与淋病奈瑟菌和衣原体相似 沙眼，但在某些临床环境中可能更为普遍。令人担忧的是，MG正变得越来越 对抗生素耐药，有些感染在美国完全无法用推荐的方案治疗。的 最近FDA批准的MG诊断测试肯定会提高公众对MG的认识，并要求 来改善治疗。MG感染的一个标志是长期持续，尽管存在特异性 生殖道中的抗体。在过去的十年里，我们的实验室一直专注于 免疫显性MgpB和MgpC粘附蛋白，以及对这些蛋白的局部和全身抗体应答 感染期间的蛋白质我们证明了MgpB和MgpC通过一种特异性的免疫反应经历了相位和抗原性变异。 mgpBC表达位点和部分拷贝之间的同源重组的独特系统 在染色体上。我们建议扩展这些研究授权的三个最新进展：（1） 确定MgpC蛋白质结构，其中包括唾液酸结合口袋，（2）我们详细的 MG感染的纵向标本中抗体反应性和抗原变异的分析 包括MgpC的保守C-末端区域含有靶向的显性抗原的证据。 （3）发现了MG 281免疫球蛋白结合蛋白，其在 发病机制尚未探索。MgpC C-末端表位将使用工程化缺失和丙氨酸氨基转移酶来定义。 通过ELISA和表面等离子体共振（SPR）分析取代，以探索热力学和 抗体结合动力学。补体杀伤和调理吞噬测定将定位靶点， 杀菌抗体确定患者血清识别的MgpC表位将有助于改进血清学检测， 有必要探讨MG与严重的女性上生殖道后遗症的关系。作为唾液酸 酸结合口袋嵌入MgpC的可变区，我们将测量唾液酸结合的变化， 不同变体中的亲和力，以及变体特异性抗体对唾液酸结合和粘附的影响。 了解MG 281在避免抗MG抗体的生物活性中的作用是MG治疗的首要任务。 特别是MG与先天免疫效应物的相互作用的研究尚未确定。所以我们的 第三个目标将把目标1和目标2的结果联系起来，以确定MG 281是否能防止特定的 MG（+）男性血清和MG（+）女性宫颈阴道分泌物中的抗体。这些实验，沿着 我们的新方法开创了支原体研究这一困难领域的实验方法， 激发改进预防战略，以对抗这种日益具有抗药性的重要病原体。

PROJECT SUMMARY Mycoplasma genitalium (MG) is an emerging reproductive tract pathogen of significant public health concern. This sexually transmitted bacterium elicits a disease spectrum similar to Neisseria gonorrhoeae and Chlamydia trachomatis, yet may be more prevalent in certain clinical settings. Alarmingly, MG is becoming increasingly resistant to antibiotics with some infections totally untreatable with recommended regimens in the US. The recent FDA approval of an MG diagnostic test will certainly increase public awareness of MG, and demands for improved treatments. A hallmark of MG infection is long-term persistence despite the presence of specific antibodies in the genital tract. Over the past decade our laboratory has focused on the variability of the immunodominant MgpB and MgpC adherence proteins, and the local and systemic antibody response to these proteins during infection. We demonstrated that MgpB and MgpC undergo phase and antigenic variation via a unique system of homologous recombination between the mgpBC expression site and partial copies archived in the chromosome. We propose to extend these studies empowered by three recent advances: (1) the determination of the MgpC protein structure, which includes a sialic acid binding pocket, (2) our detailed analysis of antibody reactivity and antigenic variation in a collection of longitudinal specimens from MG-infected men, including evidence that the conserved C-terminal region of MgpC contains the dominant antigen targeted by patient antibodies, and (3) the discovery of the MG281 immunoglobulin binding protein whose role in pathogenesis is unexplored. MgpC C-terminal epitopes will be defined using engineered deletions and alanine substitutions analyzed by ELISA and surface plasmon resonance (SPR) to explore thermodynamics and kinetics of antibody binding. Complement killing and opsonophagocytosis assays will locate targets of bactericidal antibodies. Defining MgpC epitopes recognized by patient sera will inform improved serologic tests, essential to explore association of MG with serious upper reproductive tract sequelae in women. As the sialic acid binding pocket is embedded in the variable region of MgpC, we will measure changes in sialic acid binding affinity in different variants, and the effect of variant-specific antibodies on sialic acid binding and adherence. Understanding the role of MG281 in avoiding the biologic activity of anti-MG antibodies is a top priority in MG research especially as interactions of MG with innate immune effectors has not been defined. Therefore, our third aim will link the findings of Aims 1 and 2 to determine whether MG281 prevents killing by specific antibodies in sera of MG(+) men and in cervicovaginal exudates of MG(+) women. These experiments, along with our novel approaches pioneering experimental methods for the difficult field of mycoplasma research, will inspire improved prevention strategies to combat this increasingly antibiotic-resistant and important pathogen.