Table_1_Targeted proteomics links virulence factor expression with clinical severity in staphylococcal pneumonia.xlsx

IntroductionThe bacterial pathogen Staphylococcus aureus harbors numerous virulence factors that impact infection severity. Beyond virulence gene presence or absence, the expression level of virulence proteins is known to vary across S. aureus lineages and isolates. However, the impact of expression level on severity is poorly understood due to the lack of high-throughput quantification methods of virulence proteins.
MethodsWe present a targeted proteomic approach able to monitor 42 staphylococcal proteins in a single experiment. Using this approach, we compared the quantitative virulomes of 136 S. aureus isolates from a nationwide cohort of French patients with severe community-acquired staphylococcal pneumonia, all requiring intensive care. We used multivariable regression models adjusted for patient baseline health (Charlson comorbidity score) to identify the virulence factors whose in vitro expression level predicted pneumonia severity markers, namely leukopenia and hemoptysis, as well as patient survival.
ResultsWe found that leukopenia was predicted by higher expression of HlgB, Nuc, and Tsst-1 and lower expression of BlaI and HlgC, while hemoptysis was predicted by higher expression of BlaZ and HlgB and lower expression of HlgC. Strikingly, mortality was independently predicted in a dose-dependent fashion by a single phage-encoded virulence factor, the Panton-Valentine leucocidin (PVL), both in logistic (OR 1.28; 95%CI[1.02;1.60]) and survival (HR 1.15; 95%CI[1.02;1.30]) regression models.
DiscussionThese findings demonstrate that the in vitro expression level of virulence factors can be correlated with infection severity using targeted proteomics, a method that may be adapted to other bacterial pathogens.

**引言**：作为一种细菌性致病菌，金黄色葡萄球菌（Staphylococcus aureus）携带诸多影响感染严重程度的毒力因子。除毒力基因的存在与否外，已有研究表明，毒力蛋白的表达水平在金黄色葡萄球菌的不同谱系及分离株中存在差异。然而，由于缺乏针对毒力蛋白的高通量定量方法，人们对表达水平对感染严重程度的影响仍知之甚少。

**方法**：本研究提出一种靶向蛋白质组学方法，可在单次实验中同时检测42种葡萄球菌蛋白。利用该方法，我们对136株金黄色葡萄球菌分离株的定量毒力组（virulome）进行了比较，这些分离株来自法国全国范围内的重症社区获得性葡萄球菌肺炎患者队列，所有患者均需接受重症监护。我们采用针对患者基线健康状况（查尔森合并症指数（Charlson comorbidity score））校正的多变量回归模型，以鉴定能够通过体外表达水平预测肺炎严重程度指标（即白细胞减少症与咯血）及患者存活率的毒力因子。

**结果**：研究发现，HlgB、Nuc及Tsst-1的高表达与BlaI、HlgC的低表达可预测白细胞减少症；而BlaZ与HlgB的高表达、HlgC的低表达则可预测咯血。值得注意的是，在逻辑回归（比值比OR=1.28；95%置信区间CI[1.02;1.60]）与生存回归（风险比HR=1.15；95%CI[1.02;1.30]）模型中，单一噬菌体编码的毒力因子——潘顿-瓦伦丁杀白细胞素（Panton-Valentine leucocidin，PVL）可通过剂量依赖性方式独立预测患者死亡率。

**讨论**：本研究结果表明，借助靶向蛋白质组学技术，可将毒力因子的体外表达水平与感染严重程度建立关联，该方法或可推广应用于其他细菌性致病菌的相关研究。