Real-time nanoscale analysis of immune evasion molecules from Streptococcus pyogenes using bio-imaging technologies

中性粒细胞是最初招募到感染部位的免疫细胞的一部分。然而，在包括链球菌中毒性休克综合征在内的严重链球菌感染中，化脓性链球菌附近几乎没有发现中性粒细胞，它迅速感染STSS患者的各个器官。化脓性链球菌被认为配备了毒力因子来逃避中性粒细胞免疫系统。以前，我们发现补体C3b作为调理素在链球菌中毒性休克综合征患者的血清中被降解。此外，化脓性链球菌还能破坏中性粒细胞胞外陷阱(Net)。在本研究中，我们研究了化脓性链球菌逃逸中性粒细胞的机制。Biaccore分析表明，rSpeB和野生型化脓性链球菌能迅速降解C3b。在小鼠模型中，ΔSpeB突变体的细菌存活率低于野生型菌株。此外，组织病理学观察显示，ΔSpeB突变体可见中性粒细胞渗入感染部位，而注射野生型的小鼠则很少迁移至感染部位，化脓性链球菌继续沿筋膜扩散。时间推移显微镜分析表明，野生型菌株对Net有消化作用，而ΔSpeB突变株没有。我们的结果表明，SpeB通过降解初始感染部位的C3b和Net，帮助化脓性链球菌逃避中性粒细胞的免疫。

Neutrophils are part of the initial group of immune cells recruited to the site of infection. However, in severe streptococcal infection including streptococcal toxic shock syndrome, few neutrophils have been found near Streptococcus pyogenes, which rapidly infects various organs in STSS patients. S.pyogenes is thought to be equipped with virulence factors to evade the neutrophil immune system. Previously, we found that complement C3b functioned as an opsonin is degraded in sera from patients with streptococcal toxic shock syndrome. Furthermore, it has been revealed that S.pyogenes is able to fragment neutrophil extracellular traps (NETs). In the present study, we investigated the mechanism of S.pyogenes evasion from neutrophils. Biacore analysis demonstrated that rSpeB and wild-type S.pyogenes rapidly degrade C3b. In a mouse model, bacterial survival rate was lower for the ΔspeB mutant than the wild-type strain. Furthermore, histopathological observations showed that neutrophils infiltrated to the site of infection by the ΔspeB mutant, whereas few migrated to the infection site in mice injected with the wild type and S. pyogenes continued to spread along the fascia. Time-lapse microscopic analysis revealed that digestion of NETs was seen by the wild-type strain, but not ΔspeB mutant strain. Our results indicate that SpeB contributes to the escape of S. pyogenes from neutrophil immunity by degrading C3b and NETs at the site of initial infection.