Antigenic variation of Mycoplasma genitalium during persistent genital tract infection of pig-tailed macaques

项目总结 生殖支原体(MG)是一种性传播的细菌病原体，越来越被认为是一种 严重的公共卫生问题。在基于人群的研究中，MG的患病率从1-4%到更高 20%以上的患者是获得性传播感染的高危人群。MG的疾病谱是 与淋球菌和沙眼衣原体相似，包括男性尿道炎和 女人。尤其值得关注的是，MG感染与严重的上生殖道后遗症有关。 妇女包括盆腔炎、不孕症、早产和自然流产。令人震惊的是， MG的耐药性正在增加：40%-100%的菌株对阿奇霉素和 有些感染是完全无法用美国批准的疗法治疗的。FDA最近批准了两种MG 诊断性测试肯定会提高公众对MG的认识，并要求改善治疗。一个 迫切需要动物模型来了解MG感染的自然历史，包括MG感染的机制 持续性和免疫逃避，这项研究在患者中是困难的，因为必须治疗有症状的患者 感染。我们已经优化了猪尾猴持续性生殖道感染的模型，现在 建议使用该模型来研究抗原变异在免疫回避中的作用。延续我们之前的 免疫显性粘附素mgpB和mgpC的抗原性和时相变化机制的研究 蛋白质，我们将确定是否需要变异才能在生殖道中持续存在。首先，使用全基因组 我们将在感染18周期间将变异的出现与出现的 三种MG感染的灵长类动物和以前存档的标本中的特异性mgpB和mgc抗体 灵长类动物实验。第二，相同的与变异的菌株再次感染清除生殖器的动物的能力 将对肠道感染进行评估，以了解菌株的特异性免疫。第三，一个非变量的“锁定” 将构建不能进行抗原变异的MG株，并在体外进行鉴定。三 将对灵长类动物进行宫颈接种野生型和锁定MG的混合接种，以比较 在我们的模型的18周内，这两个菌株的持久性和上呼吸道上升同时发生。全 受感染的灵长类动物将接受尸检，以检查宫颈、子宫和输卵管的肉眼病理。 和组织学，并评估MG在上生殖道的存在。这些实验不会 只确定基因变异是否是持久性所必需的，但也将提供额外的观察 灵长类个体对下尿路的自然史持久性、上尿路上升和免疫力 应答(包括细胞因子、细胞浸润物和针对保守和可变MG的抗体 抗原)。这些拟议的实验具有非常重要的意义，因为它们填补了优先考虑的研究空白 NIAID赞助的小组，即发展动物模型，探索MG在 女性严重的上生殖道疾病，以及MG血清学测试的发展。

PROJECT SUMMARY Mycoplasma genitalium (MG), a sexually transmitted bacterial pathogen, is increasingly recognized as a significant public health concern. The prevalence of MG ranges from 1-4% in population-based studies to more than 20% in patients at high risk of acquiring sexually transmitted infections. The disease spectrum of MG is similar to Neisseria gonorrhoeae and Chlamydia trachomatis, and includes urethritis in men and cervicitis in women. Of particular concern, MG infection is associated with serious upper reproductive tract sequelae in women including pelvic inflammatory disease, infertility, preterm birth, and spontaneous abortion. Alarmingly, antimicrobial resistance in MG is increasing: 40-100% of strains are completely resistant to azithromycin and some infections are totally untreatable with US approved therapies. The recent FDA approval of two MG diagnostic tests will certainly increase public awareness of MG and demands for improved treatment. An animal model is urgently needed to understand the naturally history of MG infection including mechanisms of persistence and immune evasion, studies that are difficult in patients given the imperative to treat symptomatic infection. We have optimized our pig-tailed macaque model of persistent genital tract infection and now propose to use this model to study the role of antigenic variation in immune avoidance. Extending our previous work defining mechanisms of antigenic and phase variation of the immunodominant MgpB and MgpC adhesin proteins, we will determine if variation is required for persistence in the genital tract. First, using whole genome sequencing we will correlate the appearance of variants during 18 weeks of infection with the appearance of antibodies specific to MgpB and MgpC in three MG-infected primates and archived specimens from prior primate experiments. Second, the ability of the identical vs variant strain to re-infect animals that clear genital tract infection will be assessed in order to understand strain specific immunity. Third, a non-variable, “locked” MG strain that is unable to undergo antigenic variation will be constructed and characterized in vitro. Three primates will be inoculated cervically with a mixed inoculum of wild type and “locked” MG to compare the persistence and upper tract ascension of the two strains simultaneously over the 18 weeks of our model. All infected primates will undergo necropsy to examine the cervix, uterus, and Fallopian tubes for gross pathology and histology, and to assess the presence of MG in the upper reproductive tract. These experiments will not only determine if gene variation is required for persistence but will also provide additional observations in individual primates of the natural history of lower tract persistence, upper tract ascension, and immune response (including cytokines, cellular infiltrates and antibodies specific for conserved and variable MG antigens). These proposed experiments are highly significant in that they fill research gaps prioritized by NIAID-sponsored panels, namely the development of an animal model, the exploration of the role of MG in serious upper reproductive tract disease in women, and the development of an MG serologic test.