[1]马水燕,王楠楠,董雨豪,等.嗜水气单胞菌Ⅵ型分泌系统假定效应蛋白基因tle1AH的生物学功能[J].南京农业大学学报,2020,43(5):896-902.[doi:10.7685/jnau.201910004]
 MA Shuiyan,WANG Nannan,DONG Yuhao,et al.Biological functions of the type Ⅵ secretion system putative effector gene tle1AH in Aeromonas hydrophila[J].Journal of Nanjing Agricultural University,2020,43(5):896-902.[doi:10.7685/jnau.201910004]
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嗜水气单胞菌Ⅵ型分泌系统假定效应蛋白基因tle1AH的生物学功能()
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《南京农业大学学报》[ISSN:1000-2030/CN:32-1148/S]

卷:
43卷
期数:
2020年5期
页码:
896-902
栏目:
动物科学
出版日期:
2020-09-15

文章信息/Info

Title:
Biological functions of the type Ⅵ secretion system putative effector gene tle1AH in Aeromonas hydrophila
作者:
马水燕 王楠楠 董雨豪 刘锦 陆承平 刘永杰
南京农业大学动物医学院, 江苏 南京 210095
Author(s):
MA Shuiyan WANG Nannan DONG Yuhao LIU Jin LU Chengping LIU Yongjie
College of Veterinary Medicine, Nanjing Agricultural University, Nanjing 210095, China
关键词:
嗜水气单胞菌Ⅵ型分泌系统效应蛋白tle1AH基因生物学功能
Keywords:
Aeromonas hydrophilatype Ⅵ secretion systemeffector proteintle1AH genebiological functions
分类号:
S855.1+2
DOI:
10.7685/jnau.201910004
摘要:
[目的] 本文旨在鉴定和研究嗜水气单胞菌中国流行株NJ-35的Ⅵ型分泌系统(T6SS)新型效应蛋白及其功能,进一步阐明其致病机制。[方法] 基于已报道的保守结构域DUF4123,对NJ-35株全基因组序列进行潜在的T6SS效应蛋白基因预测,采用同源重组技术构建基因缺失株和相应互补株。对野生株、缺失株和互补株分别进行细菌竞争试验,并测定细菌生长曲线、运动性、生物被膜形成能力、抗巨噬细胞吞噬能力及其对斑马鱼的半数致死量(LD50)等。[结果] 成功预测到假定的效应蛋白基因tle1AH(U876-17550),并构建tle1AH基因缺失株(Δtle1AH)和相应互补株(CΔtle1AH)。特性分析发现,与野生株比较,缺失株抑制大肠杆菌生长和抗小鼠巨噬细胞吞噬的能力明显降低,生物被膜形成能力增强,对斑马鱼的LD50增加10余倍(P<0.01),互补株各项指标均恢复到野生株水平。[结论] 鉴定到嗜水气单胞菌T6SS的一个假定效应蛋白基因(tle1AH),并对其功能进行初步研究,为进一步探讨T6SS在该菌致病过程中的作用奠定基础。
Abstract:
[Objectives] The aim of the study was to identify the novel effectors of type Ⅵ secretion system (T6SS) and explore its function in Aeromonas hydrophila Chinese epidemic strain NJ-35,which was beneficial to further investigating the pathogenesis of pathogens.[Methods] The putative effector was screened in the whole genome of A.hydrophila NJ-35 based on the reported conservative domain DUF4123. Furthermore,the mutant and complementation strains were constructed via homologous recombination,and the competitiveness,growth,motility,biofilm formation,anti-phagocytosis and virulence in zebrafish were measured.[Results] A putative gene was obtained and named tle1AH (U876-17550),and its mutant (Δtle1AH) and complemented (CΔtle1AH) strains were constructed. The Δtle1AH showed a significant decrease in anti-bacterial efficacy against Escherichia coli and anti-phagocytosis activity. In addition,the biofilm formation ability of Δtle1AH increased significantly,and its virulence decreased in zebrafish,more than a 10-fold increase in LD50 over the wild-type strain (P<0.01). Meanwhile,the above characteristics of CΔtle1AH were restored to the same level of the wild type strain.[Conclusions] In this study,a functional analysis was conducted on a putative effector protein of T6SS in A.hydrophila,which provides a basis for further research on the role of T6SS in the pathogenic mechanism of this bacteria.

参考文献/References:

[1] Chopra A K,Houston C W. Enterotoxins in Aeromonas-associated gastroenteritis[J]. Microbes and Infection,1999,1(13):1129-1137.
[2] Citterio B,Biavasco F. Aeromonas hydrophila virulence[J]. Virulence,2015,6(5):417-418.
[3] Suarez G,Sierra J C,Kirtley M L,et al. Role of Hcp,a type 6 secretion system effector,of Aeromonas hydrophila in modulating activation of host immune cells[J]. Microbiology,2010,156(12):3678-3688.
[4] Ho B T,Dong T G,Mekalanos J J. A view to a kill:the bacterial type Ⅵ secretion system[J]. Cell Host & Microbe,2014,15(1):9-21.
[5] Renault M G,Zamarreno B J,Douzi B,et al. The gp27-like hub of VgrG serves as adaptor to promote hcp tube assembly[J]. Journal of Molecular Biology,2018,430(18):3143-3156.
[6] Shneider M M,Buth S A,Ho B T,et al. PAAR-repeat proteins sharpen and diversify the type Ⅵ secretion system spike[J]. Nature,2013,500(7462):350-353.
[7] Liang X Y,Moore R,Wilton M,et al. Identification of divergent type Ⅵ secretion effectors using a conserved chaperone domain[J]. Proc Natl Acad Sci USA,2015,112(29):9106-9111.
[8] Russell A B,LeRoux M,Hathazi K,et al. Diverse type Ⅵ secretion phospholipases are functionally plastic antibacterial effectors[J]. Nature,2013,496(7446):508-512.
[9] Wang N N,Wu Y F,Pang M D,et al. Protective efficacy of recombinant hemolysin co-regulated protein(Hcp)of Aeromonas hydrophila in common carp(Cyprinus carpio)[J]. Fish & Shellfish Immunology,2015,46(2):297-304.
[10] Abolghait S K. Suicide plasmid-dependent IS1-element untargeted integration into Aeromonas veronii bv.sobria generates brown pigment-producing and spontaneous pelleting mutant[J]. Current Microbiology,2013,67(1):91-99.
[11] Livak K J,Schmittgen T D. Analysis of relative gene expression data using real-time quantitative PCR and the 2ΔΔCT method[J]. Methods,2001,25(4):402-408.
[12] O’Toole G A,Pratt L A,Watnick P I,et al. Genetic approaches to study of biofilms[J]. Methods in Enzymology,1999,310(1):91-109.
[13] Macintyre D L,Miyata S T,Kitaoka M,et al. The Vibrio cholerae type Ⅵ secretion system displays antimicrobial properties[J]. Proc Natl Acad Sci USA,2010,107(45):19520-19524.
[14] Dong Y H,Geng J Z,Liu J,et al. Roles of three TonB systems in the iron utilization and virulence of the Aeromonas hydrophila Chinese epidemic strain NJ-35[J]. Applied Microbiology and Biotechnology,2019,103(10):4203-4215.
[15] Pang M D,Lin X Q,Hu M,et al. Tetrahymena:an alternative model host for evaluating virulence of Aeromonas strains[J]. PLoS One,2012,7(11):e48922.
[16] Dong Y H,Wang Y,Liu J,et al. Discovery of lahS as a global regulator of environmental adaptation and virulence in Aeromonas hydrophila[J]. International Journal of Molecular Sciences,2018,19(9):2709.
[17] Costa T R D,Felisberto-Rodrigues C,Meir A,et al. Secretion systems in Gram-negative bacteria:structural and mechanistic insights[J]. Nature Reviews Microbiology,2015,13(6):343-359.
[18] Russell A B,Peterson S B,Mougous J D. Type Ⅵ secretion system effectors:poisons with a purpose[J]. Nature Reviews Microbiology,2014,12(2):137-148.
[19] Ma J L,Pan Z H,Huang J H,et al. The Hcp proteins fused with diverse extended-toxin domains represent a novel pattern of antibacterial effectors in type Ⅵ secretion systems[J]. Virulence,2017,8(7):1189-1202.
[20] Sha J,Rosenzweig J A,Kozlova E V,et al. Evaluation of the roles played by Hcp and VgrG type 6 secretion system effectors in Aeromonas hydrophila SSU pathogenesis[J]. Microbiology,2013,159(6):1120-1135.
[21] Gallique M,Decoin V,Barbey C,et al. Contribution of the Pseudomonas fluorescens MFE01 type Ⅵ secretion system to biofilm formation[J]. PLoS One,2017,12(1):e0170770.
[22] Wang N N,Liu J,Pang M D,et al. Diverse roles of Hcp family proteins in the environmental fitness and pathogenicity of Aeromonas hydrophila Chinese epidemic strain NJ-35[J]. Applied Microbiology and Biotechnology,2018,102(16):7083-7095.
[23] Kirchberger P C,Unterweger D,Provenzano D,et al. Sequential displacement of type Ⅵ secretion system effector genes leads to evolution of diverse immunity gene arrays in Vibrio cholerae[J]. Scientific Reports,2017,7:45133.

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[10]严亚贤,孙建和,陈怀青,等.嗜水气单胞菌菌毛的提纯及特性分析[J].南京农业大学学报,1995,18(3):88.[doi:10.7685/j.issn.1000-2030.1995.03.015]
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备注/Memo

备注/Memo:
收稿日期:2019-10-09。
基金项目:国家自然科学基金项目(31372454);江苏省水产三新工程项目(D2017-3-1)
作者简介:马水燕,硕士研究生。
通信作者:刘永杰,教授,博导,研究方向为兽医微生物学,E-mail:liuyongjie@njau.edu.cn。
更新日期/Last Update: 1900-01-01