WhiB4 is required for the reactivation of latent infection of Mycobacterium marinum in zebrafish [zebrafish RNAseq]. WhiB4 is required for the reactivation of latent infection of Mycobacterium marinum in zebrafish [zebrafish RNAseq]

Granulomas are the pathological hallmark of tuberculosis (TB). In individuals latently infected with Mycobacterium tuberculosis (M. tb), the bacteria are thought to reside within the granulomas in a nonreplicating dormant state due to the lack of oxygen and nutrients. However, a portion of these individuals will develop active TB and little is known on the bacterial mechanisms/factors involved in this process. In this study, we found that WhiB4, an oxygen sensor and a transcription factor, plays a critical role in disease progression and reactivation of Mycobacterium marinum (M. marinum) infection in zebrafish. We show that the whiB4 mutant of M. marinum caused latent infection in adult zebrafish, which is characterized by the stable bacterial loads, constant number of non-necrotized granulomas in fewer organs, and reduced immune responses compared to zebrafish infected with the wild type bacteria or the complemented strain. The mutant bacteria in zebrafish were also less responsive to antibiotic treatments. Moreover, the whiB4 mutant was defective in resuscitation from hypoxia-induced dormancy and that the DosR regulon was dysregulated in the mutant. Taken together, our results suggest that WhiB4 is a major driver of reactivation from latent infection and that WhiB4 is an attractive target for the development of novel therapeutics, which may help to prevent the reactivation of latent infection thereby reducing the incidences of active TB. Overall design: mRNA profiles of zebrafish kidney infected with whiB4 mutant strain and wild type;a total of 6 samples were analyzed, 3 relicates were included for control and experiment samples.

肉芽肿（granulomas）是结核病（tuberculosis, TB）的病理标志性特征。对于潜伏感染结核分枝杆菌（Mycobacterium tuberculosis, M. tb）的个体而言，由于缺氧与营养匮乏，细菌被认为以非复制休眠状态驻留于肉芽肿内。然而，其中部分个体将进展为活动性结核病，而该过程中涉及的细菌机制与因子仍鲜为人知。本研究发现，作为氧感应转录因子的WhiB4，在斑马鱼感染海分枝杆菌（Mycobacterium marinum, M. marinum）的疾病进展与再激活过程中发挥关键作用。研究证实，海分枝杆菌ΔwhiB4缺失突变株可在成年斑马鱼中引发潜伏感染，其特征为细菌载量稳定、少数器官内非坏死性肉芽肿数量恒定，且相较于野生型菌株或互补菌株感染的斑马鱼，宿主免疫应答水平显著降低。该突变株在斑马鱼体内对抗生素治疗的响应也更为迟钝。此外，ΔwhiB4突变株在缺氧诱导的休眠状态复苏方面存在缺陷，且突变株内的DosR调节子表达失调。综合上述结果，本研究表明WhiB4是潜伏感染再激活的核心驱动因子，同时也是新型治疗药物开发的极具潜力的靶点，或可助力预防潜伏感染再激活，从而降低活动性结核病的发病率。整体实验设计：感染whiB4突变株与野生型菌株的斑马鱼肾脏组织的mRNA转录组谱；共分析6个样本，对照组与实验组各包含3个生物学重复。