{"title":"[A simplified system for generation of germ-free zebrafish embryos and its application in Listeria monocytogenes infection].","authors":"Ying Shan, Yikai Zhang, Changyong Cheng, Chun Fang, Jinrong Pen, Weihuan Fang","doi":"","DOIUrl":null,"url":null,"abstract":"<p><strong>Objective: </strong>Under conventional cultivation conditions zebrafish harbors numerous microbes from the environment, leading to activation of its innate immune systems and interfering the results of relevant studies. We aimed to establish a germ-free zebrafish embryo model suitable for studies of host immune responses to infections.</p><p><strong>Methods: </strong>A germ-free cultivation process including simple disinfection of the fertilized eggs and growth in a positive-pressured thermostatic isolator. Sterility testing of germ-free zebrafish embryos and water samples was done according to the national standards. The transcriptional level of TLRs, the mark genes indicating activation of the innate immune system, was detected by qPCR. Listeria monocytogenes was used as an infection model.</p><p><strong>Results: </strong>The cultivation system and disinfection process could ensure germ-free status as shown by absence of microbes in zebrafish embryos and egg water. TLRs were barely detectable in zebrafish raised in the germ-free system, but highly induced in conventionally raised zebrafish or in germ-free zebrafish immersion-infected with pathogenic Listeria monocytogenes. The germ-free fish was sensitive to infection by L. monocytogene EGDe at a 100-CFU dose with 100% mortality in one week, while its isogenic mutants Δmpl and ΔplcB exhibited reduced death (70% and 40%, respectively). Macrophages were recruited around the intestine in EGDe immersion infected fish, but not in Δmpl and ΔplcB infected fish.</p><p><strong>Conclusion: </strong>Zebrafish embryos produced by this simple process were free of microbes and could be used to study the innate immune responses and the pathogenesis of microbial pathogens.</p>","PeriodicalId":7120,"journal":{"name":"微生物学报","volume":null,"pages":null},"PeriodicalIF":0.0000,"publicationDate":"2016-11-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"微生物学报","FirstCategoryId":"1089","ListUrlMain":"","RegionNum":0,"RegionCategory":null,"ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"","JCRName":"","Score":null,"Total":0}
引用次数: 0
Abstract
Objective: Under conventional cultivation conditions zebrafish harbors numerous microbes from the environment, leading to activation of its innate immune systems and interfering the results of relevant studies. We aimed to establish a germ-free zebrafish embryo model suitable for studies of host immune responses to infections.
Methods: A germ-free cultivation process including simple disinfection of the fertilized eggs and growth in a positive-pressured thermostatic isolator. Sterility testing of germ-free zebrafish embryos and water samples was done according to the national standards. The transcriptional level of TLRs, the mark genes indicating activation of the innate immune system, was detected by qPCR. Listeria monocytogenes was used as an infection model.
Results: The cultivation system and disinfection process could ensure germ-free status as shown by absence of microbes in zebrafish embryos and egg water. TLRs were barely detectable in zebrafish raised in the germ-free system, but highly induced in conventionally raised zebrafish or in germ-free zebrafish immersion-infected with pathogenic Listeria monocytogenes. The germ-free fish was sensitive to infection by L. monocytogene EGDe at a 100-CFU dose with 100% mortality in one week, while its isogenic mutants Δmpl and ΔplcB exhibited reduced death (70% and 40%, respectively). Macrophages were recruited around the intestine in EGDe immersion infected fish, but not in Δmpl and ΔplcB infected fish.
Conclusion: Zebrafish embryos produced by this simple process were free of microbes and could be used to study the innate immune responses and the pathogenesis of microbial pathogens.
期刊介绍:
Acta Microbiologica Sinica(AMS) is a peer-reviewed monthly (one volume per year)international journal,founded in 1953.It covers a wide range of topics in the areas of general and applied microbiology.The journal
publishes original papers,reviews in microbiological science,and short communications describing unusual observations.
Acta Microbiologica Sinica has been indexed in Index Copernicus (IC),Chemical Abstract (CA),Excerpt Medica Database (EMBASE),AJ of Viniti (Russia),Biological Abstracts (BA),Chinese Science Citation Database
(CSCD),China National Knowledge Infrastructure(CNKI),Institute of Scientific and Technical Information of China(ISTIC),Chinese Journal Citation Report(CJCR),Chinese Biological Abstracts,Chinese Pharmaceutical
Abstracts,Chinese Medical Abstracts and Chinese Science Abstracts.