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

中性粒细胞是被募集到感染部位的初始免疫细胞群的一部分。然而，在严重的链球菌感染，包括链球菌中毒性休克综合征，少数中性粒细胞已被发现附近的化脓性链球菌，迅速感染各种器官的STSS患者。化脓性链球菌被认为具有毒力因子以逃避中性粒细胞免疫系统。先前，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血清中降解。此外，已经揭示化脓性链球菌能够使中性粒细胞胞外陷阱（NET）片段化。在本研究中，我们探讨了化脓性链球菌逃避中性粒细胞的机制。Biacore分析表明，rSpeB和野生型化脓性链球菌快速降解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中性粒细胞浸润到ΔspeB突变体的感染部位，而在注射野生型和S.化脓继续沿沿着扩散。延时显微镜分析显示，NETs的消化被野生型菌株所观察到，但ΔspeB突变菌株没有。我们的结果表明SpeB对S的逃逸有贡献。化脓性链球菌通过降解初始感染部位的C3b和NET而从中性粒细胞免疫中获得。

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.