Bacterial membrane vesicles as modulators of adverse pregnancy outcomes

B族链球菌（GBS）存在于生殖泌尿道，并在高达35%的孕妇中定菌，导致绒毛膜羊膜炎、早产、死胎、新生儿败血症和脑膜炎。GBS感染发生在子宫内，在胎盘膜被侵入后或在分娩时因阴道内的吸入而发生。我们之前的研究表明，特定的GBS基因型在新生儿疾病病例中比孕妇更常见，孕妇在母胎界面感染细胞的能力更强。然而，我们对不同GBS菌株持续定植和/或侵入胎盘膜的机制的理解存在重大差距。我们期望表型变异及其对宿主反应的影响是一个重要但尚未得到充分研究的因素，并代表了本提案的重点。我们的初步数据表明，不同的GBS菌株分泌的胞外膜囊泡（MVs）在数量、大小和蛋白质含量上都有所不同，这些MVs在体外引起巨噬细胞的炎症细胞因子反应。我们还发现，来自两种不同GBS菌株的mv含有不同丰度的蛋白质，这些蛋白质被归类为已知和新型GBS毒力因子，这表明mv在GBS发病机制中发挥了作用。虽然最近在GBS中描述了mv，但只研究了一种罕见基因型的对照菌株。因此，我们建议：1)确定GBS MVs对人妊娠组织宿主反应的影响；2)确定MVs对孕鼠宿主反应和胎儿结局的影响；3)测定GBS病毒的蛋白质含量。我们将对不同基因型的侵袭性和非侵袭性菌株的mv含量进行表征，并对从母胎界面恢复的原代细胞以及感染妊娠小鼠后的宿主反应进行分类。我们预测，来自侵袭性GBS菌株而非共生菌株的mv将引发炎症反应，从而导致胎儿或新生儿损伤。在短期内，我们将确定MVs在炎症反应、不良妊娠结局和毒性中的作用。我们的长期目标是发现与感染、疫苗靶点和递送系统相关的新型生物标志物，这些生物标志物可能会影响由GBS和其他具有类似生活方式的病原体引起的疾病。

Group B Streptococcus (GBS) resides in the genitourinary tract and colonizes up to 35% of pregnant women contributing to chorioamnionitis, preterm birth, stillbirth, and neonatal sepsis and meningitis. GBS infections occur in utero following invasion of the placental membranes or during childbirth upon aspiration within the vaginal canal. Our prior studies have shown that specific GBS genotypes are more common among neonatal disease cases than pregnant mothers with a superior ability to infect cells at the maternal-fetal interface. However, there are critical gaps in our understanding of the mechanisms that different GBS strains use to persistently colonize and/or invade the placental membranes. We expect that phenotypic variation and its impact on host responses is an important yet understudied contributor to consider and represents the focus of this proposal. Our preliminary data show that distinct GBS strains secrete extracellular membrane vesicles (MVs) that vary in quantity, size and protein content and that these MVs elicit inflammatory cytokine responses from macrophages in vitro. We also found that MVs from two different GBS strains contained differentially abundant proteins that were classified as known and novel GBS virulence factors, suggesting that MVs play a role in GBS pathogenesis. Although MVs were recently described in GBS, only one control strain of a rare genotype was studied. Consequently, we propose to: 1) Determine the influence of GBS MVs on host response in human gestational tissues; 2) Determine the influence of MVs on host response and fetal outcome in pregnant mice; and 3) Ascertain the protein content of GBS MVs. We will characterize the content of MVs from both invasive and non-invasive strains of varying genotypes and classify host responses from primary cells recovered from the maternal-fetal interface as well as following infection of pregnant mice. We predict that MVs from the invasive GBS strains and not the commensal strains will trigger inflammatory responses that contribute to fetal or neonatal injury. In the short term, we will determine the role that MVs play in inflammatory responses, adverse pregnancy outcomes and virulence. Our long-term goal is to discover novel biomarkers associated with infection, vaccine targets and delivery systems that can potentially impact diseases caused by GBS and other pathogens with similar lifestyles.