Please wait a minute...
浙江大学学报（农业与生命科学版）
Journal of Zhejiang University (Agriculture and Life Sciences)  2022, Vol. 48 Issue (3): 303-309    DOI: 10.3785/j.issn.1008-9209.2021.02.152
Biological sciences & biotechnologies     
Evaluation of Aeromonas hydrophila outer membrane protein deletion strains in response to environmental stresses
Zeqi LI(),Yuqian WANG,Xiaoyan LI,Xiangmin LIN,Jun YUAN()
Fujian Provincial Key Laboratory of Agroecological Processing and Safety Monitoring, College of Life Sciences, Fujian Agriculture and Forestry University, Fuzhou 350002, China
Download: HTML   HTML ( )    PDF(3274KB)
Export: BibTeX | EndNote (RIS)      

Abstract  

In order to evaluate systematically the biological functions of outer membrane proteins (OMPs) under environmental stresses, we selected Aeromonas hydrophila ATCC 7966 as the research object, and constructed 33 OMP deletion strains to determine their growth status under the stress conditions of osmotic pressure, metal ions, H2O2 and pH. The results showed that the AHA_0461 and AHA_4275 (encoding TonB protein family) knockout strains grew better than the wild type strain under the osmotic pressure stress. The AHA_2282 (encoding an unknown functional protein) knockout strain grew weakly under the metal ion stresses, while ΔAHA_0904 strain grew better under the osmotic pressure stress. The AHA_2145 (encoding long-chain fatty acid transporter) knockout strain grew weakly under the H2O2 and metal ion stresses. The growth of AHA_1279, AHA_1281 (encoding OmpA family protein)and pilQ, AHA_0569 (encoding secretin family protein) knockout strains was weaker than the wild type strain under the H2O2 stress. It was suggested that these OMPs may play important roles in response to environmental stresses. These results may highlight a comprehensive understanding of the physiological functions of bacterial OMPs under different environmental stresses in the future, and provide possible targets for the prevention and control of this pathogen.

Key words： Aeromonas hydrophila      outer membrane protein      gene deletion      response to environmental stresses     
Received: 15 February 2021      Published: 07 July 2022
CLC:  Q 93-3  
Corresponding Authors: Jun YUAN     E-mail: lzq_943317937@sina.com;yjmail2008@126.com
Service
E-mail this article
Add to my bookshelf
Add to citation manager
E-mail Alert
RSS
Articles by authors
Zeqi LI
Yuqian WANG
Xiaoyan LI
Xiangmin LIN
Jun YUAN
Cite this article:

Zeqi LI,Yuqian WANG,Xiaoyan LI,Xiangmin LIN,Jun YUAN. Evaluation of Aeromonas hydrophila outer membrane protein deletion strains in response to environmental stresses. Journal of Zhejiang University (Agriculture and Life Sciences), 2022, 48(3): 303-309.

URL:

https://www.zjujournals.com/agr/10.3785/j.issn.1008-9209.2021.02.152     OR     https://www.zjujournals.com/agr/Y2022/V48/I3/303


嗜水气单胞菌外膜蛋白缺失株逆境响应功能评价

为系统评价外膜蛋白在环境胁迫条件下的生物学功能，以嗜水气单胞菌（Aeromonas hydrophila）ATCC 7966为研究对象，选取33株外膜蛋白缺失菌株，对其贴太紧在渗透压、金属离子、H2O2及pH等胁迫条件下的生长状态进行测定。结果发现：编码TonB家族蛋白基因AHA_0461、AHA_4275被敲除的菌株在渗透压胁迫条件下较野生型菌株生长更好；编码未知功能蛋白基因AHA_2282被敲除的菌株在金属离子胁迫条件下生长较弱，而ΔAHA_0904菌株在渗透压胁迫条件下生长状况较好；编码长链脂肪酸转运蛋白基因AHA_2145被敲除的菌株在H2O2和金属离子胁迫条件下生长状态较弱；而编码OmpA家族蛋白基因AHA_1279、AHA_1281和分泌素家族蛋白基因pilQ、AHA_0569被敲除的菌株在H2O2胁迫条件下的生长均弱于野生型菌株。提示这些蛋白可能在细菌响应环境胁迫方面起重要作用。本研究结果有望为今后全面地了解嗜水气单胞菌外膜蛋白响应不同环境胁迫的机制提供思路，为该病原菌的防治提供可能的靶点。


关键词： 嗜水气单胞菌,  外膜蛋白,  基因缺失,  逆境响应 
Fig. 1 PCR amplification results of wild type (WT) strain and partial OMP gene knockout strainsM: DL2000 marker; 1: WT; 31: ΔAHA_2991; 32: ΔAHA_0569; 33: ΔAHA_3509; 34: ΔAHA_0521.
Fig. 2 Determination results of WT strain and OMP gene knockout strains under different concentrations of NaCl
Fig. 3 Growth status of WT strain and OMP gene knockout strains under different pH conditions
Fig. 4 Growth status of WT strain and OMP gene knockout strains under different concentrations of H2O2CK: LB liquid medium.
Fig. 5 Growth status of WT strain and OMP gene knockout strains under different metal ion stresses
Fig. 6 Prediction results of interaction networks of 33 OMPs and summary of the phenotypes responding to environmental stressesDifferent solid line colors indicate the changes of strain phenotypes under environmental stress‍es (1: Increase in assay; －1: Decrease in assay). The OMPs of AHA_0389, AHA_2202 and AHA_2699 are not found to be involved in protein-protein interaction networks.
[1]   赵香汝.细菌外膜蛋白的结构与功能[J].中国兽医科技,1999,29(10):20-23.
ZHAO X R. Structure and function of bacterial outer membrane protein[J]. Chinese Journal of Veterinary Science and Technology, 1999, 29(10): 20-23. (in Chinese)
[2]   YANG D C H, ABEYKOON A H, CHOI B E, et al. Outer membrane protein OmpB methylation may mediate bacterial virulence[J]. Trends in Biochemical Sciences, 2017, 42(12): 936-945. DOI:10.1016/j.tibs.2017.09.005
doi: 10.1016/j.tibs.2017.09.005
[3]   林向民,王风平,王三英.不同盐度对Halomonas aquamarina外膜蛋白表达的影响[J].厦门大学学报(自然科学版),2005,44():128-131.
LIN X M, WANG F P, WANG S Y. The effect of different NaCl concentrations on OMP expression of Halomonas aquamarina [J]. Journal of Xiamen University (Natural Science), 2005, 44(): ‍128-131. (in Chinese with English abstract)
[4]   DUPONT M, JAMES C E, CHEVALIER J, et al. An early response to environmental stress involves regulation of OmpX and OmpF, two enterobacterial outer membrane pore-forming proteins[J]. Antimicrobial Agents and Chemotherapy, 2007, 51(9): 3190-3198. DOI:10.1128/AAC.01481-06
doi: 10.1128/AAC.01481-06
[5]   PRATT L A, HSING W H, GIBSON K E, et al. From acids to osmZ: multiple factors influence synthesis of the OmpF and OmpC porins in Escherichia coli [J]. Molecular Microbiology, 1996, 20(5): 911-917. DOI:10.1111/j.1365-2958.1996.tb02532.x
doi: 10.1111/j.1365-2958.1996.tb02532.x
[6]   王璐,梁利国,谢骏,等.嗜水气单胞菌双重PCR检测方法的建立及应用[J].水产科学,2013,32(4):219-223. DOI:10.16378/j.cnki.1003-1111.2013.04.010
WANG L, LIANG L G, XIE J, et al. Development and application of duplex PCR for detection of bacterium Aeromonas hydrophila [J]. Fisheries Science, 2013, 32(4): 219-223. (in Chinese with English abstract)
doi: 10.16378/j.cnki.1003-1111.2013.04.010
[7]   LI Z Q, WANG Y Q, LI X Y, et al. The characteristics of antibiotic resistance and phenotypes in 29 outer-membrane protein mutant strains in Aeromonas hydrophila [J]. Environmental Microbiology, 2019, 21(12): 4614-4628. DOI:10.1111/1462-2920.14761
doi: 10.1111/1462-2920.14761
[8]   ZGURSKAYA H I, LÓPEZ C A, GNANAKARAN S. Permeability barrier of Gram-negative cell envelopes and approaches to bypass it[J]. ACS Infectious Diseases, 2015, 1(11): 512-522. DOI:10.1021/acsinfecdis.5b00097
doi: 10.1021/acsinfecdis.5b00097
[9]   RAMAKRISHNAN G. Iron and virulence in Francisella tularensis [J]. Frontiers in Cellular and Infection Microbiology, 2017, 7: 107. DOI:10.3389/fcimb.2017.00107
doi: 10.3389/fcimb.2017.00107
[10]   BALIBAR C J, GRABOWICZ M. Mutant alleles of lptD increase the permeability of Pseudomonas aeruginosa and define determinants of intrinsic resistance to antibiotics[J]. Antimicrobial Agents and Chemotherapy, 2015, 60(2): 845-854. DOI:10.1128/AAC.01747-15
doi: 10.1128/AAC.01747-15
[11]   BOURASSA D V, KANNENBERG E L, SHERRIER D J, et al. The lipopolysaccharide lipid—a long-chain fatty acid is important for Rhizobium leguminosarum growth and stress adaptation in free-living and nodule environments[J]. Molecular Plant-Microbe Interactions, 2017, 30(2): 161-175. DOI:10.1094/MPMI-11-16-0230-R
doi: 10.1094/MPMI-11-16-0230-R
[12]   姜倩雯,张丽珊,李小艳,等.单壁碳纳米管载嗜水气单胞菌外膜蛋白亚单位疫苗的研制及免疫效果评价[J].水产学报,2020,44(9):1424-1434. DOI:10.11964/jfc.20190411755
JIANG Q W, ZHANG L S, LI X Y, et al. Development of single-walled carbon nanotubes carrying Aeromonas hydrophila outer membrane protein subunit vaccine and evaluation of immune effect[J]. Journal of Fisheries of China, 2020, 44(9): 1424-1434. (in Chinese with English abstract)
doi: 10.11964/jfc.20190411755
[13]   MORI N, ISHII Y, TATEDA K, et al. A peptide based on homologous sequences of the β‍-barrel assembly machinery component BamD potentiates antibiotic susceptibility of Pseudomonas aeruginosa [J]. Journal of Antimicrobial Chemotherapy, 2012, 67(9): 2173-2181. DOI:‍10.1093/jac/dks174
doi: ?10.1093/jac/dks174
[14]   ARTS I S, GENNARIS A, COLLET J F. Reducing systems protecting the bacterial cell envelope from oxidative damage[J]. FEBS Letters, 2015, 589(14): 1559-1568. DOI:10.1016/j.febslet.2015.04.057
doi: 10.1016/j.febslet.2015.04.057
[15]   TAMBOSI R, LIOTENBERG S, BOURBON M L, et al. Silver and copper acute effects on membrane proteins and impact on photosynthetic and respiratory complexes in bacteria[J]. mBio, 2018, 9(6): e01535-18. DOI:‍10.1128/mBio.01535-18
doi: ?10.1128/mBio.01535-18
[16]   NIE D, HU Y, CHEN Z, et al. Outer membrane protein A (OmpA) as a potential therapeutic target for Acinetobacter baumannii infection[J]. Journal of Biomedical Science, 2020, 27(1): 26. DOI:10.1186/s12929-020-0617-7
doi: 10.1186/s12929-020-0617-7
[17]   SHI L, RICHARDSON D J, WANG Z M, et al. The roles of outer membrane cytochromes of Shewanella and Geobacter in extracellular electron transfer[J]. Environmental Microbiology Reports, 2009, 1(4): 220-227. DOI:10.1111/j.1758-2229.2009.00035.x
doi: 10.1111/j.1758-2229.2009.00035.x
[1] Lu Lianming, Du Danchao, Cheng Baoping, Hu Xiurong, Zhang Liping, Chen Guoqing. Genetic variation analysis of an outer membrane protein gene of Candidatus Liberibacter asiaticus.[J]. Journal of Zhejiang University (Agriculture and Life Sciences), 2014, 40(2): 125-132.