How E. coli Acid Response Mechanisms Breach Colonization Resistance in the Vagina

摘要 这项提议将检验肠外致病性大肠埃希菌 (Exc)通过相互连接的酸的顺序激活克服泌尿生殖乳杆菌的抑制 抗性(AR)机制。我们进一步假设ExPEC瞬间内化到阴道上皮细胞 细胞通过增强耐酸性的诱导和其他持久的机制来增强适应性 使细菌能够进入并在其他恶劣的宿主环境中生存。需要检验的假设 (1)我们发现了一种新的AR机制 在通过非规范信号系统控制的ExPEC中，BTSS-YpdB使用L丝氨酸脱氨 中和细菌胞液pH值。我们称这种新的AR机制为AR6。(2)BTSS-YpdB信令 在感染期间和对几种乳杆菌的反应。(3)BTSS-ypdB显著缺失 降低ExPEC的耐酸性和阴道定植。(4)缺乏L丝氨酸脱氨酶的菌株- YpdB对具有代表性的加瑟氏乳杆菌和德氏乳杆菌的抑制作用有差异。 分离株。(5)BTSS-ypDB的缺失改变了已知的酸敏系统EvgSA的诱导和功能 它控制着最突出的、已知的AR机制AR2。我们将使用最多的 流行的ExPEC病原型，尿路致病性大肠埃希菌。致尿性大肠埃希氏菌是尿路的主要原因 感染(尿路感染)，这是一种不成比例地困扰女性的感染。同样，ExPEC菌株是领先的 感染相关死产的原因。尽管主导模式是阴道的低pH值对 病原体，我们和其他人已经表明，ExPEC对阴道的定植可以作为感染的病灶。 尿路、宫颈、子宫角和妊娠子宫。因此，阴道定植是关键的一步 在发病机制上。虽然已经确定了几种在肠道中活跃的耐酸(AR)机制， 每种AR机制在ExPEC感染过程中的相对贡献尚不清楚。有了我们的目标，我们 威尔：询问寄主中瞬时细菌扩张作为启动利基的意义 增强耐酸性和其他持久性机制(目标1)。我们将评估个人和 AR机制对ExPEC在阴道、膀胱和肠道的定植潜能的联合作用 并将阐明我们发现的新的AR6通路与AR2的诱导和功能的关系 (目标2)。最后，在令人兴奋的初步数据的基础上，我们将调查泌尿生殖系统乳杆菌的潜力 菌株超越ExPEC耐酸性的能力，旨在确定有效的益生菌策略以防止 排除阴道内的蓄积物(目标3)。完成这些目标将揭示ExPEC如何利用 他们的多个AR系统，以过境宿主和逃避消除，并将通知我们的努力 制定有效的益生菌策略以预防、阴道定植和防治生殖道 感染。

SUMMARY This proposal will test the over-arching hypothesis that extra-intestinal pathogenic Escherichia coli (ExPEC) overcomes inhibition by urogenital Lactobacilli via sequential activation of inter-connected acid resistance (AR) mechanisms. We further postulate that transient internalization of ExPEC into vaginal epithelial cells increases fitness by enhancing induction of acid resistance and other mechanisms of persistence that enable bacteria to gain access and survive in otherwise harsh host environments. The hypothesis to be tested has been formulated based on the following strong preliminary data: (1) We discovered a novel AR mechanism in ExPEC that is controlled via a non-canonical signaling system, BtsS-YpdB and it uses L-serine deamination to neutralize bacterial cytosolic pH. We call this new AR mechanism, AR6. (2) BtsS-YpdB signaling is induced during infection and in response to several Lactobacillus species. (3) Deletion of btsS-ypdB significantly decreases ExPEC acid tolerance and vaginal colonization. (4) Strains lacking L-serine deaminases, or BtsS- YpdB differentially react to the inhibitory actions of representative urogenital L. gasseri and L. delbrueckii isolates. (5) Deletion of btsS-ypdB alters the induction and function of the known acid-sensing system EvgSA that controls the most prominent, known AR mechanism, AR2. We will test these hypotheses using the most prevalent ExPEC pathotype, uropathogenic E. coli. Uropathogenic E. coli is the main cause of urinary tract infections (UTIs), an infection that disproportionately afflicts women. Similarly, ExPEC strains are the leading cause of infection-related stillbirths. Despite the dominant paradigm that the low pH of vagina is protective against pathogens, we and others have shown that colonization of the vagina by ExPEC can serve as a nidus for infection of the urinary tract, the cervix, uterine horns and the gravid uterus. Vaginal colonization is therefore a key step in pathogenesis. While several acid resistance (AR) mechanisms have been identified that are active in the gut, the relative contribution of each AR mechanism during ExPEC infection remains undefined. With our aims, we will: Interrogate the significance of transient bacterial expansion in the host as a priming niche for the amplification of acid resistance and other persistence mechanisms (Aim 1). We will evaluate the individual and combined contributions of AR mechanisms to the colonization potential of ExPEC in the vagina, bladder and gut and will elucidate the connection of the novel AR6 pathway we discovered to the induction and function of AR2 (Aim 2). Finally, building on exciting preliminary data we will investigate the potential of urogenital Lactobacilli strains in their ability to override ExPEC acid resistance, aiming to identify effective probiotic strategies to prevent ExPEC reservoir formation in the vagina (Aim 3). Completion these aims will uncover how ExPEC leverage their multiple AR systems to transit the host and evade elimination and will inform our efforts towards developing effective probiotic strategies to prevent, vaginal colonization and combat genitourinary tract infections.