Neisseria gonorrhoeae central metabolism in the context of neutrophilic inflammation

摘要 淋球菌(GC)是性传播疾病淋病的病原体。GC是一种 由于出现了对所有当前病毒都有抵抗力的“超级细菌”菌株，对人类健康构成了突出的威胁 抗生素及其临床后遗症，包括不孕不育和慢性盆腔疼痛，具有严重的终身 效果。GC高度适应于在人类粘膜表面定植，尽管它在那里启动了强大的 炎症反应和中性粒细胞(PMN或中性粒细胞)的涌入，通常是透明的 细菌。我们最近使用了无偏见的全基因组方法，转座子测序和双重RNA- 对GC和PMN进行测序，以确定导致PMN成功感染GC的一组基因。气相色谱(GC) 代谢基因在这个队列中被过度表达，这表明GC利用不同的代谢途径 绕过人类的粘膜防御系统。我们已经将这些信息应用于系统生物学方法，以 为GC生成一个初步的代谢网络重建。这一分析导致预测GC 利用特定的宿主衍生的代谢物来为其代谢提供燃料--特别是乳酸。PMN是高度糖酵解的 从而分泌乳酸盐。乳酸可以被GC消耗，并被认为能刺激GC糖酵解。因此， 我们假设GC利用PMN代谢，特别是乳酸分泌，在粘膜表面生存。 在这项提案中，我们将确定GC在中性粒细胞存在时使用的中心碳代谢途径， 在GC代谢中使用代谢物分析、代谢建模和细菌基因敲除的组合 酵素。然后，我们将测试这些代谢途径在GC中提高细菌存活率的必要性 中性粒细胞及其基本机制，包括调节中性粒细胞的抗微生物反应，改变 GC对中性粒细胞抗微生物成分的敏感性，和/或规避宿主营养免疫，或 调节中性粒细胞的抗微生物反应。总而言之，这项建议将解决在以下方面的主要差距： 感染期间GC的基本代谢过程可能对预后有重大影响 淋病的症状。

ABSTRACT Neisseria gonorrhoeae (Gc) is the causative agent of the sexually transmitted disease gonorrhea. Gc is a prominent threat to human health due to emergence of “superbug” strains that are resistant to all current antibiotics and its clinical sequelae, including infertility and chronic pelvic pain, that have severe and lifelong effects. Gc is highly adapted to colonize human mucosal surfaces, where it survives despite initiating a robust inflammatory response and influx of polymorphonuclear leukocytes (PMNs or neutrophils) that typically clear bacteria. We recently used the unbiased, genome-wide approaches of transposon-sequencing and dual RNA- sequencing of Gc and PMNs to identify a cohort of genes contributing to successful Gc infection of PMNs. Gc metabolic genes were overrepresented in this cohort, suggesting that Gc utilizes distinct metabolic pathways to circumvent human mucosal defenses. We have applied this information to systems biology approaches to generate a preliminary metabolic network reconstruction for Gc. This analysis leads to the prediction that Gc utilizes specific host-derived metabolites to fuel its metabolism – in particular, lactate. PMNs are highly glycolytic and consequently secrete lactate. Lactate can be consumed by Gc and is known to stimulate Gc glycolysis. Thus we hypothesize that Gc exploits PMN metabolism, specifically lactate secretion, to survive at mucosal surfaces. In this proposal, we will identify the central carbon metabolic pathways used by Gc in the presence of neutrophils, using a combination of metabolite analysis, metabolic modeling, and bacterial genetic knockouts in Gc metabolic enzymes. We will then test the necessity of these metabolic pathways in Gc for enhancing bacterial survival from neutrophils and the underlying mechanisms, including modulating neutrophil antimicrobial responses, changing Gc sensitivity to neutrophil antimicrobial components, and/or circumventing host nutritional immunity, or modulating neutrophil antimicrobial responses. Together, this proposal will address key gaps in knowledge of the fundamental metabolic processes of Gc during infection that are likely to have major impacts on outcomes of gonorrhea.