Identification of conditionally essential genes for <i>Streptococcus suis</i> infection in pigs

<i>Streptococcus suis</i> is a Gram-positive bacterium and zoonotic pathogen that causes meningitis and sepsis in pigs and humans. The aim of this study was to identify genes required for <i>S. suis</i> infection. We created Tn-Seq libraries in a virulent <i>S. suis</i> strain 10, which was used to inoculate pigs in an intrathecal experimental infection. Comparative analysis of the relative abundance of mutants recovered from different sites of infection (blood, cerebrospinal fluid, and meninges of the brain) identified 361 conditionally essential genes, i.e. required for infection, which is about 18% of the genome. The conditionally essential genes were primarily involved in metabolic and transport processes, regulation, ribosomal structure and biogenesis, transcription, and cell wall membrane and envelope biogenesis, stress defenses, and immune evasion. Directed mutants were created in a set of 10 genes of different genetic ontologies and their role was determined in <i>ex vivo</i> models. Mutants showed different levels of sensitivity to survival in whole blood, serum, cerebrospinal fluid, thermic shock, and stress conditions, as compared to the wild type. Additionally, the role of three selected mutants was validated in co-infection experiments in which pigs were infected with both wild type and isogenic mutant strains. The genetic determinants of infection identified in this work contribute to novel insights in <i>S. suis</i> pathogenesis and could serve as targets for novel vaccines or antimicrobial drugs.猪链球菌（Streptococcus suis）是一种革兰氏阳性菌（Gram-positive），同时也是可引发猪与人类罹患脑膜炎、败血症的人畜共患病原体。本研究旨在鉴定猪链球菌感染所必需的基因。研究人员在强毒猪链球菌10菌株中构建了转座子测序（Tn-Seq）文库，并将该文库用于颅内途径的猪实验感染接种。对从不同感染部位（血液、脑脊液及脑膜）回收的突变体相对丰度开展比较分析后，共鉴定出361个感染必需的条件必需基因（conditionally essential genes），约占该菌株基因组的18%。此类条件必需基因主要参与代谢与转运过程、基因调控、核糖体结构与生物发生、转录过程、细胞壁膜及包膜生物发生、应激防御以及免疫逃逸等生物学功能。研究人员针对隶属于不同遗传本体类别的10个基因构建了定向突变株，并通过体外（ex vivo）模型验证了其功能。与野生型菌株相比，各突变株在全血、血清、脑脊液中的存活能力以及热激和应激条件下均呈现出不同程度的敏感性差异。此外，研究人员通过共感染实验验证了3个筛选得到的突变株的功能：该实验中猪同时感染野生型菌株与同基因突变株。本研究鉴定的猪链球菌感染遗传决定因子，为该菌的致病机制研究提供了全新见解，同时可作为新型疫苗或抗菌药物的开发靶点。