Type VII secretion system promotes Streptococcus agalactiae virulence through magnesium acquisition and capsule maintenance

Streptococcus agalactiae, also known as Group B Streptococcus (GBS), is a major pathogen causing substantial economic losses in global tilapia aquaculture. The type VII secretion systems (T7SS), present in Actinobacteria and Firmicutes, secrete effector proteins implicated in bacterial virulence, yet its functional mechanisms remain poorly defined. Here, we constructed an essC deletion mutant (∆essC) in S. agalactiae strain HN016 to investigate the role of T7SS in virulence. The ∆essC exhibited an impaired growth and declined intracellular magnesium ion concentration in THB or magnesium-limited chemically defined medium (1 mM Mg2+-CDM), these defect that were complemented by addition of wild-type culture supernatant. Transcriptomic and qRT-PCR analyses revealed altered sugar metabolism and significant downregulation of capsule biosynthesis genes (cpsA, cpsB, cpsD) in ∆essC. Consistent with this, the mutant produced markedly less capsular polysaccharide and displayed impaired capsule integrity. Furthermore, cellular assays confirmed that compared to HN016, the ∆essC mutant exhibited significantly reduced adhesion capacity, immune evasion ability and cytotoxicity. More importantly, the ∆essC mutant showed attenuated virulence in vivo, with reduced bacterial loads in host tissues and diminished ability to cross the blood-brain barrier (BBB). Our findings provide the first evidence that the T7SS influences magnesium homeostasis and is essential for maintaining capsule integrity, both of which contribute critically to pathogenicity. This study identifies T7SS as a potential target for novel therapeutic strategies against streptococcal disease in aquaculture.

无乳链球菌（Streptococcus agalactiae），又称B族链球菌（Group B Streptococcus, GBS），是造成全球罗非鱼养殖业重大经济损失的主要致病菌。VII型分泌系统（type VII secretion systems, T7SS）广泛存在于放线菌门与厚壁菌门中，其分泌的效应蛋白与细菌毒力密切相关，但该系统的具体功能机制仍有待阐明。本研究在无乳链球菌HN016菌株中构建了essC基因缺失突变株（ΔessC），以探究T7SS在细菌毒力中的作用。实验结果显示，ΔessC在THB培养基或镁限制型化学成分限定培养基（1 mM Mg²+-CDM）中生长受损，胞内镁离子浓度下降，上述缺陷可通过添加野生型菌株的培养上清得以修复。转录组分析与qRT-PCR实验表明，ΔessC中糖代谢通路发生改变，荚膜生物合成基因（cpsA、cpsB、cpsD）显著下调。与之相符的是，该突变株的荚膜多糖产量显著降低，荚膜完整性受损。进一步的细胞实验证实，相较于野生株HN016，ΔessC突变株的黏附能力、免疫逃逸能力与细胞毒性均显著下降。更为关键的是，ΔessC突变株在体内毒力显著减弱，宿主组织内的细菌载量降低，且穿过血脑屏障（blood-brain barrier, BBB）的能力大幅下降。本研究首次证实，T7SS可影响镁离子稳态，且对维持荚膜完整性至关重要，二者均对致病菌的致病性发挥关键作用。本研究明确T7SS可作为水产养殖中链球菌病新型防治策略的潜在靶点。