Avirulent Phenotype promotes Bordetella pertussis Adaptation to the Intramacrophage Environment. Avirulent Phenotype promotes Bordetella pertussis Adaptation to the Intramacrophage Environment

Bordetella pertussis, the causative agent of whooping cough, is an extracellular, strictly human pathogen. However, it has been shown that B. pertussis cells can escape phagocytic killing and survive in macrophages upon internalization. Our time-resolved RNA-seq data suggest that B. pertussis efficiently adapts to the intramacrophage environment and responds to host bactericidal activities. We show that this adaptive response is multifaceted and, surprisingly, related to the BvgAS two-component system, a master regulator of virulence. Our results show that expression of this regulatory circuit is downregulated upon internalization. Moreover, we demonstrate that the switch to avirulent Bvg- phase triggers a very complex process based on the adjustment of central and energy metabolism, cell wall reinforcement, maintenance of appropriate redox and metal homeostasis, and repair of damaged macromolecules. Nevertheless, not all observed effects could be simply attributed to the transition to Bvg- phase, suggesting that additional regulators are involved in the adaptation to the intramacrophage environment. Interestingly, a large number of genes required for metabolism of sulfur were strongly modulated within macrophages. In particular, the mutant lacking two genes encoding cysteine dioxygenases displayed strongly attenuated cytotoxicity toward THP-1 cells. Collectively, our results suggest that intracellular B. pertussis cells have adopted the Bvg- mode to acclimate to the intramacrophage environment and respond to antimicrobial activities elicited by THP-1 cells. Therefore, we hypothesize that the avirulent phase represents an authentic phenotype of internalized B. pertussis cells.

百日咳博德特菌（Bordetella pertussis）是百日咳的致病菌，属于胞外专性人类病原体。然而已有研究证实，百日咳博德特菌可逃逸吞噬杀伤，并在被巨噬细胞内化后于胞内存活。本研究的时间分辨RNA测序（time-resolved RNA-seq）数据表明，百日咳博德特菌可高效适应巨噬细胞内环境，并对宿主的杀菌活性产生应答。我们证实，该适应性应答具有多面性，且出乎意料地与毒力主调控因子——BvgAS双组分系统（BvgAS two-component system）相关。研究结果显示，该调控回路的表达在细菌内化后发生下调。此外，我们证明，向无毒力Bvg-相的转变会触发极为复杂的过程，包括中枢与能量代谢的调控、细胞壁加固、维持适宜的氧化还原与金属稳态，以及受损大分子的修复。然而，并非所有观测到的效应都可简单归因于向Bvg-相的转变，这提示存在额外的调控因子参与巨噬细胞内环境的适应过程。值得注意的是，大量参与硫代谢的基因在巨噬细胞内被显著调控。具体而言，缺失两个编码半胱氨酸双加氧酶（cysteine dioxygenases）的突变株，其对THP-1细胞（THP-1 cells）的细胞毒性显著减弱。综上，我们的研究结果表明，胞内的百日咳博德特菌会采用Bvg-模式以适应巨噬细胞内环境，并应对THP-1细胞所引发的抗菌活性。因此，我们提出假说：无毒力相是内化的百日咳博德特菌的真实表型。