Department of Microbiology, University of Washington, Seattle, WA, USA.
Nat Protoc. 2013 Jun;8(6):1114-24. doi: 10.1038/nprot.2013.068. Epub 2013 May 16.
Mycobacterium marinum-infected zebrafish are used to study tuberculosis pathogenesis, as well as for antitubercular drug discovery. The small size of zebrafish larvae coupled with their optical transparency allows for rapid analysis of bacterial burdens and host survival in response to genetic and pharmacological manipulations of both mycobacteria and host. Automated fluorescence microscopy and automated plate fluorimetry (APF) are coupled with facile husbandry to facilitate large-scale, repeated analysis of individual infected fish. Both methods allow for in vivo screening of chemical libraries, requiring only 0.1 μmol of drug per fish to assess efficacy; they also permit a more detailed evaluation of the individual stages of tuberculosis pathogenesis. Here we describe a 16-h protocol spanning 22 d, in which zebrafish larvae are infected via the two primary injection sites, the hindbrain ventricle and caudal vein; this is followed by the high-throughput evaluation of pathogenesis and antimicrobial efficacy.
感染海洋分枝杆菌的斑马鱼被用于研究结核病的发病机制，以及用于抗结核药物的发现。斑马鱼幼虫体积小，光学透明，因此可以快速分析细菌负荷和宿主在遗传和药理学操纵细菌和宿主时的存活情况。自动化荧光显微镜和自动平板荧光计 (APF) 与简便的饲养相结合，可方便地对单个感染鱼进行大规模、重复分析。这两种方法都允许对化学文库进行体内筛选，每个鱼只需 0.1μmol 的药物即可评估疗效；它们还允许更详细地评估结核病发病机制的各个阶段。这里我们描述了一个跨越 22 天的 16 小时方案，在此期间，通过后脑室和尾静脉这两个主要注射部位将斑马鱼幼虫感染；随后对发病机制和抗菌功效进行高通量评估。
