Establishment of Multi-Site Infection Model in Zebrafish Larvae for Studying Staphylococcus aureus Infectious Disease
Establishment of Multi-Site Infection Model in Zebrafish Larvae for Studying Staphylococcus aureus Infectious Disease
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斑马鱼幼虫多部位感染模型的建立用于研究金黄色葡萄球菌传染病
DOI：
10.1016/j.jgg.2012.07.006
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发表时间：
2012-09-01
影响因子：
5.9
通讯作者：
Hu, Bing
中科院分区：
文献类型：
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作者：
Li, Ya-juan;Hu, Bing
斑马鱼(Danio Rerio)是研究传染病发病机制以及宿主与病原体相互作用的理想模型。作为硬骨鱼，斑马鱼已经形成了与哺乳动物相似的完整免疫系统。此外，大量透明胚胎的容易获得使其成为基因操作和药物筛选的良好候选者。在斑马鱼感染模型中，胚胎内细菌显微注射的所有部位、时机和剂量都是决定宿主细菌感染的重要因素。在这里，我们建立了一种斑马鱼受精后36小时幼虫的多部位感染模型，将野生型或表达GFP的金黄色葡萄球菌(S.aureus)以梯度负荷注射到不同的胚胎部位，包括心包腔(PC)、眼、第四后脑室(4V)、卵黄循环谷(YCV)、尾静脉(CV)、卵黄体(YB)和Cuvier管(DC)，以模拟人类传染病。将表达绿色荧光蛋白的金黄色葡萄球菌与转基因斑马鱼TG(coro1a：EGFP；Lyz：Dsred)和转基因斑马鱼TG(Lyz：Dsred)的巨噬细胞或中性粒细胞进行荧光标记，通过体内多色共聚焦荧光成像观察细菌感染的动态过程。对斑马鱼胚胎存活率、细菌增殖和骨髓细胞吞噬功能的分析表明，不同细菌进入途径的感染存在部位和剂量依赖的差异。这一工作为今后通过选择多点感染模型来研究致病机理和宿主抗性提供了考虑。通过斑马鱼多点感染模型，可以确定病原菌、毒力因子与斑马鱼免疫应答之间更多的相互作用机制。
Zebrafish (Danio rerio) is an ideal model for studying the mechanism of infectious disease and the interaction between host and pathogen. As a teleost, zebrafish has developed a complete immune system which is similar to mammals. Moreover, the easy acquirement of large amounts of transparent embryos makes it a good candidate for gene manipulation and drug screening. In a zebrafish infection model, all of the site, timing, and dose of the bacteria microinjection into the embryo are important factors that determine the bacterial infection of host. Here, we established a multi-site infection model in zebrafish larvae of 36 hours post-fertilization (hpf) by microinjecting wild-type or GFP-expressing Staphylococcus aereus (S. aureus) with gradient burdens into different embryo sites including the pericardial cavity (PC), eye, the fourth hindbrain ventricle (4V), yolk circulation valley (YCV), caudal vein (CV), yolk body (YB), and Duct of Cuvier (DC) to resemble human infectious disease. With the combination of GFP-expressing S. aureus and transgenic zebrafish Tg (coro1a: eGFP; lyz: Dsred) and Tg (lyz: Dsred) lines whose macrophages or neutrophils are fluorescent labeled, we observed the dynamic process of bacterial infection by in vivo multicolored confocal fluorescence imaging. Analyses of zebrafish embryo survival, bacterial proliferation and myeloid cells phagocytosis show that the site- and dose-dependent differences exist in infection of different bacterial entry routes. This work provides a consideration for the future study of pathogenesis and host resistance through selection of multi-site infection model. More interaction mechanisms between pathogenic bacteria virulence factors and the immune responses of zebrafish could be determined through zebrafish multi-site infection model.