葡萄球菌黄素合成与菌膜形成的相关性及其分子调控机制

Staphylococcus aureus CC75 was newly established as a new species, Staphylococcus argenteus, which is naturally lack of one important carotenoid, staphyloxanthin. The applicant has got food isolates of S. argenteus for the first time. It is found that the biosynthesis of staphyloxanthin strengthens the bacterium's stress resistance, meanwhile biofilm is an important way to enhance stress resistance. Combining all the data we have, it is deduced that the biosynthesis of staphyloxanthin correlates with biofilm formation. How do they correlate? And what is the molecular mechanism for the regulation of staphyloxanthin to biofilm formation? To answer this scientific question, more S. argenteus isolates will be screened from our S. aureus library. The ability to synthesize staphyloxanthin and to form biofilm of the two staphylococcal species will be evaluated to demonstrate the external correlation between the pigment biosynthesis and biofilm formation. The S. aureus mutant of staphyloxanthin biosynthesis disruption and S. argenteus mutant of staphyloxanthin biosynthesis complementation will be constructed, and the ability to synthesize staphyloxanthin, to form biofilm, and the transcriptome profiles will be compared among these mutants and their wild types to further demonstrate the internal correlation between the pigment biosynthesis and biofilm formation, and to explore the corresponding regulators. Integrating with the genomic sequencing and comparison results among the S. aureus and S. argenteus isolates, the molecular mechanism of staphyloxanthin’s regulation to biofilm formation will be elucidated. This study will further interpret the function of staphyloxanthin, and provide theoretical basis for the new method via the inhibition of staphyloxanthin to prevent and control the formation of biofilm.

金黄色葡萄球菌的克隆系CC75刚被确立为新种即银色葡萄球菌，本项目组首次获得食品分离株，该菌天然缺失葡萄球菌黄素。该色素可增强菌体抗逆能力，而菌膜是重要抗逆形式，结合现有数据，可推断色素合成与菌膜形成相关。那么，二者如何相关？其分子调控机制是什么？为了回答这一问题，本项目将从金黄色葡萄球菌菌种库中发掘更多银色葡萄球菌，通过定性和定量评估两种菌的葡萄球菌黄素合成能力和菌膜形成能力，阐明色素合成与菌膜形成的表观相关性；构建该色素合成缺失的金黄色葡萄球菌突变株和色素合成回复的银色葡萄球菌突变株，通过比较突变株及其野生株的色素合成和菌膜形成能力差异及转录组差异，进一步阐明色素合成与菌膜形成之间的内在相关性，并挖掘重要的调控因子，再结合两种菌全基因组测序比对结果，阐明葡萄球菌黄素调控菌膜形成的分子机制。该项目的完成将更全面剖析葡萄球菌黄素的功能，并为通过抑制色素合成从而防控菌膜的新方法提供理论基础。

本项目从现有金黄色葡萄球菌菌种库中鉴定出6株银白色葡萄球菌，建立了一种快速区分银白色葡萄球菌和金黄色葡萄球菌的PCR检测方法。完成了19株金黄色葡萄球菌和5株银白色葡萄球菌的全基因组序列测定和相关基因组比较分析，鉴定出两种新型的葡萄球菌肠毒素超抗原SEl26和SEl27。从群体角度，未发现菌膜形成量和色素形成量之间的显著关联，但是根据野生型和色素合成能力缺失突变株的比较结果，色素合成在一定程度上会影响菌膜的形成；对金黄色葡萄球菌分离株进行了30种毒力因子编码基因筛查，并选取24株典型金黄色葡萄球菌和4株银色葡萄球菌构建小鼠脓血症模型，各类型菌株并未表现出明显的毒力差异；对金黄色葡萄球菌进行耐药谱筛查和菌膜形成能力测定，结果具有耐药表型的菌株菌膜形成能力较强，而菌膜形成受抑制时耐药性也随之减弱。发现色素合成阻断剂NTF及其衍生物均能抑制菌膜形成，且衍生物JX08806抑制效果最佳。通过转录组学和蛋白质组学对JX08806抑制菌膜形成进行关联分析，发现金黄色葡萄球菌菌膜相关粘附蛋白的表达下调；色素合成操纵子crtOPQMN基因及其蛋白表达显著下调。迄今已发表期刊论文10篇，其中SCI论文8篇；申请国家发明专利1项；培养研究生7名，其中博士生1名。