Complex Population Structure and Virulence Differences among Serotype 2 Streptococcus suis Strains Belonging to Sequence Type 28

Streptococcus suis is a major swine pathogen and a zoonotic agent. Serotype 2 strains are the most frequently associated with disease. However, not all serotype 2 lineages are considered virulent. Indeed, sequence type (ST) 28 serotype 2 S. suis strains have been described as a homogeneous group of low virulence. However, ST28 strains are often isolated from diseased swine in some countries, and at least four human ST28 cases have been reported. Here, we used whole-genome sequencing and animal infection models to test the hypothesis that the ST28 lineage comprises strains of different genetic backgrounds and different virulence. We used 50 S. suis ST28 strains isolated in Canada, the United States and Japan from diseased pigs, and one ST28 strain from a human case isolated in Thailand. We report a complex population structure among the 51 ST28 strains. Diversity resulted from variable gene content, recombination events and numerous genome-wide polymorphisms not attributable to recombination. Phylogenetic analysis using core genome single-nucleotide polymorphisms revealed four discrete clades with strong geographic structure, and a fifth clade formed by US, Thai and Japanese strains. When tested in experimental animal models, strains from this latter clade were significantly more virulent than a Canadian ST28 reference strain, and a closely related Canadian strain. Our results highlight the limitations of MLST for both phylogenetic analysis and virulence prediction and raise concerns about the possible emergence of ST28 strains in human clinical cases.猪链球菌（Streptococcus suis）是一种重要的猪源性致病菌，同时也是人畜共患病原。血清型2（serotype 2）菌株是与疾病关联最为频繁的菌株类型，但并非所有血清型2谱系均具有致病性。事实上，序列型（sequence type, ST）28型血清型2猪链球菌菌株曾被认为是一组均一的低毒力菌株。然而，在部分国家，ST28菌株常从发病猪只中分离得到，且目前已有至少4例人类ST28感染病例被报道。本研究采用全基因组测序（whole-genome sequencing）与动物感染模型，验证了“ST28谱系包含不同遗传背景与毒力水平菌株”这一假说。本次研究共纳入51株ST28菌株：其中50株分离自加拿大、美国与日本的发病猪只，剩余1株为从泰国人类感染病例中分离得到的ST28菌株。本研究揭示了51株ST28菌株复杂的种群结构：其多样性源于可变基因含量、重组事件以及大量非重组源性的全基因组多态性。基于核心基因组单核苷酸多态性（core genome single-nucleotide polymorphisms）的系统发育分析（phylogenetic analysis）显示，这些菌株可分为4个具有显著地理聚集特征的独立进化枝，以及第5个由美国、泰国与日本菌株组成的进化枝。在实验动物模型中检测发现，该第5进化枝的菌株毒力显著高于加拿大ST28参考菌株与一株亲缘关系相近的加拿大菌株。本研究结果凸显了多位点序列分型（Multi-Locus Sequence Typing, MLST）在系统发育分析与毒力预测方面的局限性，同时也引发了关于ST28菌株可能在人类临床病例中出现流行的担忧。