Gardnerella vaginalis: toxin production and pathogenesis

描述(由申请人提供)：细菌性阴道病(BV)是一种非常常见的阴道微生物疾病，影响所有女性的30%，在怀孕和非裔美国人中发病率较高。携带BV病毒的妇女发生早产的风险大大增加，这是新生儿发病率和死亡率以及获得性传播疾病(包括人类免疫缺陷病毒)的主要原因。尽管BV对公共卫生具有重要意义，但其发病机制尚不清楚。我们最近鉴定了阴道溶血素(VLY)，这是一种来自阴道加德纳菌的胆固醇依赖性细胞溶血素(一种存在于BV环境中的阴道黏膜上的细菌物种，被认为与疾病的发病机制有关)，显示出精致的人类特异性。我们推测，这种物种特有的毒素可能是阴道毛滴虫的一个重要毒力因子，与BV的发病相关。在我们的初步数据中，我们已经鉴定了生殖道上皮细胞上的VLY受体(人CD59)。这种受体被引入不敏感的细胞，使它们对VLY敏感。我们设计了一只表达hCD59受体的转基因小鼠，并构建了一个不依赖hCD59的VLY嵌合体。这些代表了BV的体内候选模型。此外，我们还开发了对阴道毛滴虫进行遗传操作的技术，包括转座子突变。在目标1中，我们将使用新的诱变技术和毒素产生的检测来确定阴道毛滴虫毒力的遗传决定因素。在目标2中，我们将确定VLY在体外和体内宿主-病原体界面上的作用，重点是VLY-hCD59相互作用的独特方面。在这些研究的结论中，我们将扩大我们对阴道毛滴虫发病机制的了解，评估新的BV体内模型，确定抑制毒素-宿主相互作用的候选策略，并开发新的工具来继续研究这种重要疾病的发病机制。

DESCRIPTION (provided by applicant): Bacterial vaginosis (BV) is an exceedingly common disorder of the vaginal microflora affecting >30% of all women, with higher rates in pregnancy and among African- American populations. Women with BV are at substantially increased risk of preterm birth, which is a major cause of neonatal morbidity and mortality, as well as acquisition of sexually transmitted diseases including human immunodeficiency virus. Despite its public health importance, the pathogenesis of BV is not well understood. We have recently characterized vaginolysin (VLY), a cholesterol-dependent cytolysin from Gardnerella vaginalis (a bacterial species present on the vaginal mucosa in the setting of BV and thought to contribute to the pathogenesis of disease) that exhibits exquisite human specificity. We hypothesize that this species-specific toxin may be an important virulence factor of G. vaginalis with relevance to the pathogenesis of BV. In our preliminary data, we have characterized the receptor for VLY (human CD59) on genital tract epithelial cells. Introduction of this receptor into non-susceptible cells renders them sensitive to VLY. We have engineered a transgenic mouse expressing the hCD59 receptor and also constructed a VLY chimera that is hCD59-independent. These represent candidate in vivo models for BV. In addition, we have developed techniques for genetic manipulation of G. vaginalis, including transposon mutagenesis. In Aim 1, we will define genetic determinants of G. vaginalis virulence using new techniques for mutagenesis and assays of toxin production. In Aim 2, we will determine the role of VLY at the host-pathogen interface both in vitro and in vivo with a focus on unique aspects of the VLY-hCD59 interaction. At the conclusion of these studies, we will have expanded our knowledge of G. vaginalis pathogenesis, evaluated new in vivo models of BV, identified candidate strategies to inhibit toxin-host interaction, and developed new tools for continued investigation into the pathogenesis of an important disorder.