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

总结 这一建议将测试过度假设，肠外致病性大肠杆菌 （ExPEC）通过依次激活相互连接的酸来克服泌尿生殖道乳酸杆菌的抑制作用 抵抗（AR）机制。我们进一步假设，进入阴道上皮细胞的ExPEC瞬时内化 细胞通过增强抗酸性和其他持久性机制的诱导来增加适应性， 使细菌能够进入并在其他恶劣的宿主环境中生存。待测试的假设 基于以下强有力的初步数据制定了：（1）我们发现了一种新的AR机制 在ExPEC中，其通过非规范信号传导系统BtsS-YpdB控制，并且其使用L-丝氨酸脱氨基 中和细菌胞质pH值。我们称这种新的AR机制为AR 6。(2)诱导BtsS-YpdB信号传导 在感染期间和对几种乳杆菌属物种的反应中。(3)btsS-ypdB的缺失显著 降低ExPEC酸耐受性和阴道定植。(4)缺乏L-丝氨酸脱氨酶或BtsS-的菌株 YpdB对代表性泌尿生殖L. gasseri和L.德氏 分离株(5)btsS-ypdB的缺失改变了已知酸敏感系统EvgSA的诱导和功能 它控制着最著名的AR机制AR 2。我们将使用最多的 流行的ExPEC致病型、尿路致病性E.杆菌肾盂肾炎大肠大肠杆菌是泌尿道感染的主要原因 感染（UTI），这是一种不成比例地折磨妇女的感染。同样，ExPEC菌株是主要的 导致感染相关的死产尽管主流观点认为阴道的低pH值可以保护 病原体，我们和其他人已经表明，阴道的定植由ExPEC可以作为一个病灶的感染 泌尿道、子宫颈、子宫角和妊娠子宫。因此阴道定植是关键的一步 在发病机理上。虽然已经确定了几种在肠道中活跃的抗酸（AR）机制， 在ExPEC感染期间，每种AR机制的相对作用仍然不确定。我们的目标是， 将：询问宿主中瞬时细菌扩增作为启动微生态位的意义， 增强抗酸性和其他持久性机制（目标1）。我们将对个人进行评估， AR机制对ExPEC在阴道、膀胱和肠道中定植潜力的联合作用 并将阐明我们发现的新的AR 6通路与AR 2的诱导和功能的联系 (Aim 2）。最后，基于令人兴奋的初步数据，我们将研究泌尿生殖道乳酸杆菌的潜力 菌株克服ExPEC酸抗性的能力，旨在确定有效的益生菌策略，以防止 阴道中的ExPEC储库形成（目的3）。完成这些目标将揭示ExPEC如何利用 他们的多个AR系统，以过境主机和逃避消除，并将告知我们的努力， 制定有效的益生菌策略，以预防阴道定植和对抗泌尿生殖道 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.