Non-pathogenic Escherichia coli acquires virulence by mutating a growth-essential LPS transporter
Autoři:
Chikara Kaito aff001; Hirono Yoshikai aff002; Ai Wakamatsu aff003; Atsushi Miyashita aff002; Yasuhiko Matsumoto aff004; Tomoko Fujiyuki aff005; Masaru Kato aff006; Yoshitoshi Ogura aff007; Tetsuya Hayashi aff007; Takao Isogai aff008; Kazuhisa Sekimizu aff009
Působiště autorů:
Graduate School of Medicine, Dentistry, and Pharmaceutical Sciences, Okayama University, Kita-ku, Okayama, Japan
aff001; Graduate School of Pharmaceutical Sciences, The University of Tokyo, Bunkyo-ku, Tokyo, Japan
aff002; Japan Biological Informatics Consortium (JBIC), Koto-ku, Tokyo, Japan
aff003; Department of Microbiology, Meiji Pharmaceutical University, Kiyose, Tokyo, Japan
aff004; The Institute of Medical Science, The University of Tokyo, Minato-ku, Tokyo, Japan
aff005; Devision of Bioanalytical Chemistry, School of Pharmacy, Showa University, Shinagawa-ku, Tokyo, Japan
aff006; Department of Bacteriology, Faculty of Medical Sciences, Kyushu University, Fukuoka, Japan
aff007; Translational Research Center, Fukushima Medical University, Fukushima, Japan
aff008; Institute of Medical Mycology, Teikyo University, Hachioji, Tokyo, Japan
aff009
Vyšlo v časopise:
Non-pathogenic Escherichia coli acquires virulence by mutating a growth-essential LPS transporter. PLoS Pathog 16(4): e32767. doi:10.1371/journal.ppat.1008469
Kategorie:
Research Article
doi:
https://doi.org/10.1371/journal.ppat.1008469
Souhrn
The molecular mechanisms that allow pathogenic bacteria to infect animals have been intensively studied. On the other hand, the molecular mechanisms by which bacteria acquire virulence functions are not fully understood. In the present study, we experimentally evaluated the evolution of a non-pathogenic strain of Escherichia coli in a silkworm infection model and obtained pathogenic mutant strains. As one cause of the high virulence properties of E. coli mutants, we identified amino acid substitutions in LptD (G580S) and LptE (T95I) constituting the lipopolysaccharide (LPS) transporter, which translocates LPS from the inner to the outer membrane and is essential for E. coli growth. The growth of the LptD and LptE mutants obtained in this study was indistinguishable from that of the parent strain. The LptD and LptE mutants exhibited increased secretion of outer membrane vesicles containing LPS and resistance against various antibiotics, antimicrobial peptides, and host complement. In vivo cross-linking studies revealed that the conformation of the LptD-LptE complex was altered in the LptD and LptE mutants. Furthermore, several clinical isolates of E. coli carried amino acid substitutions of LptD and LptE that conferred resistance against antimicrobial substances. This study demonstrated an experimental evolution of bacterial virulence properties in an animal infection model and identified functional alterations of the growth-essential LPS transporter that led to high bacterial virulence by conferring resistance against antimicrobial substances. These findings suggest that non-pathogenic bacteria can gain virulence traits by changing the functions of essential genes, and provide new insight to bacterial evolution in a host environment.
Klíčová slova:
Amino acid substitution – Antimicrobial resistance – Bacterial evolution – Mutagenesis – Mutant strains – Silkworms – Substitution mutation – Vancomycin
Zdroje
1. Deatherage DE, Kepner JL, Bennett AF, Lenski RE, Barrick JE. Specificity of genome evolution in experimental populations of Escherichia coli evolved at different temperatures. Proc Natl Acad Sci U S A. 2017;114(10):E1904–E12. Epub 2017/02/17. doi: 10.1073/pnas.1616132114 28202733; PubMed Central PMCID: PMC5347587.
2. Lazar V, Nagy I, Spohn R, Csorgo B, Gyorkei A, Nyerges A, et al. Genome-wide analysis captures the determinants of the antibiotic cross-resistance interaction network. Nat Commun. 2014;5:4352. Epub 2014/07/09. doi: 10.1038/ncomms5352 25000950; PubMed Central PMCID: PMC4102323.
3. Poole DS, Yu S, Cai Y, Dinis JM, Muller MA, Jordan I, et al. Influenza A virus polymerase is a site for adaptive changes during experimental evolution in bat cells. J Virol. 2014;88(21):12572–85. Epub 2014/08/22. doi: 10.1128/JVI.01857-14 25142579; PubMed Central PMCID: PMC4248895.
4. Bull JJ, Molineux IJ. Predicting evolution from genomics: experimental evolution of bacteriophage T7. Heredity (Edinb). 2008;100(5):453–63. Epub 2008/01/24. doi: 10.1038/sj.hdy.6801087 18212807.
5. Wichman HA, Badgett MR, Scott LA, Boulianne CM, Bull JJ. Different trajectories of parallel evolution during viral adaptation. Science. 1999;285(5426):422–4. Epub 1999/07/20. doi: 10.1126/science.285.5426.422 10411508.
6. Ensminger AW, Yassin Y, Miron A, Isberg RR. Experimental evolution of Legionella pneumophila in mouse macrophages leads to strains with altered determinants of environmental survival. PLoS Pathog. 2012;8(5):e1002731. Epub 2012/06/14. doi: 10.1371/journal.ppat.1002731 22693450; PubMed Central PMCID: PMC3364954.
7. Kaito C, Kurokawa K, Matsumoto Y, Terao Y, Kawabata S, Hamada S, et al. Silkworm pathogenic bacteria infection model for identification of novel virulence genes. Mol Microbiol. 2005;56(4):934–44. Epub 2005/04/28. doi: 10.1111/j.1365-2958.2005.04596.x 15853881.
8. Kaito C, Akimitsu N, Watanabe H, Sekimizu K. Silkworm larvae as an animal model of bacterial infection pathogenic to humans. Microb Pathog. 2002;32(4):183–90. Epub 2002/06/25. doi: 10.1006/mpat.2002.0494 12079408.
9. Nikaido H. Molecular basis of bacterial outer membrane permeability revisited. Microbiol Mol Biol Rev. 2003;67(4):593–656. Epub 2003/12/11. doi: 10.1128/MMBR.67.4.593-656.2003 14665678; PubMed Central PMCID: PMC309051.
10. Zhang G, Meredith TC, Kahne D. On the essentiality of lipopolysaccharide to Gram-negative bacteria. Curr Opin Microbiol. 2013;16(6):779–85. Epub 2013/10/24. doi: 10.1016/j.mib.2013.09.007 24148302; PubMed Central PMCID: PMC3974409.
11. Schwechheimer C, Kuehn MJ. Outer-membrane vesicles from Gram-negative bacteria: biogenesis and functions. Nat Rev Microbiol. 2015;13(10):605–19. Epub 2015/09/17. doi: 10.1038/nrmicro3525 26373371; PubMed Central PMCID: PMC5308417.
12. Kuehn MJ, Kesty NC. Bacterial outer membrane vesicles and the host-pathogen interaction. Genes Dev. 2005;19(22):2645–55. Epub 2005/11/18. doi: 10.1101/gad.1299905 16291643.
13. Klieve AV, Yokoyama MT, Forster RJ, Ouwerkerk D, Bain PA, Mawhinney EL. Naturally occurring DNA transfer system associated with membrane vesicles in cellulolytic Ruminococcus spp. of ruminal origin. Appl Environ Microbiol. 2005;71(8):4248–53. Epub 2005/08/09. doi: 10.1128/AEM.71.8.4248-4253.2005 16085810; PubMed Central PMCID: PMC1183309.
14. Ruiz N, Kahne D, Silhavy TJ. Transport of lipopolysaccharide across the cell envelope: the long road of discovery. Nat Rev Microbiol. 2009;7(9):677–83. Epub 2009/07/28. doi: 10.1038/nrmicro2184 19633680; PubMed Central PMCID: PMC2790178.
15. Braun M, Silhavy TJ. Imp/OstA is required for cell envelope biogenesis in Escherichia coli. Mol Microbiol. 2002;45(5):1289–302. Epub 2002/09/05. doi: 10.1046/j.1365-2958.2002.03091.x 12207697.
16. Wu T, McCandlish AC, Gronenberg LS, Chng SS, Silhavy TJ, Kahne D. Identification of a protein complex that assembles lipopolysaccharide in the outer membrane of Escherichia coli. Proc Natl Acad Sci U S A. 2006;103(31):11754–9. Epub 2006/07/25. doi: 10.1073/pnas.0604744103 16861298; PubMed Central PMCID: PMC1544242.
17. Ruiz N, Falcone B, Kahne D, Silhavy TJ. Chemical conditionality: a genetic strategy to probe organelle assembly. Cell. 2005;121(2):307–17. Epub 2005/04/27. doi: 10.1016/j.cell.2005.02.014 15851036.
18. Abe S, Okutsu T, Nakajima H, Kakuda N, Ohtsu I, Aono R. n-Hexane sensitivity of Escherichia coli due to low expression of imp/ostA encoding an 87 kDa minor protein associated with the outer membrane. Microbiology. 2003;149(Pt 5):1265–73. Epub 2003/05/02. doi: 10.1099/mic.0.25927-0 12724388.
19. Manning AJ, Kuehn MJ. Contribution of bacterial outer membrane vesicles to innate bacterial defense. BMC Microbiol. 2011;11:258. Epub 2011/12/03. doi: 10.1186/1471-2180-11-258 22133164; PubMed Central PMCID: PMC3248377.
20. Doshi R, Nguyen T, Chang G. Transporter-mediated biofuel secretion. Proc Natl Acad Sci U S A. 2013;110(19):7642–7. Epub 2013/04/25. doi: 10.1073/pnas.1301358110 23613592; PubMed Central PMCID: PMC3651508.
21. Ruiz N, Chng SS, Hiniker A, Kahne D, Silhavy TJ. Nonconsecutive disulfide bond formation in an essential integral outer membrane protein. Proc Natl Acad Sci U S A. 2010;107(27):12245–50. Epub 2010/06/23. doi: 10.1073/pnas.1007319107 20566849; PubMed Central PMCID: PMC2901483.
22. Freinkman E, Chng SS, Kahne D. The complex that inserts lipopolysaccharide into the bacterial outer membrane forms a two-protein plug-and-barrel. Proc Natl Acad Sci U S A. 2011;108(6):2486–91. Epub 2011/01/25. doi: 10.1073/pnas.1015617108 21257904; PubMed Central PMCID: PMC3038725.
23. Dong H, Xiang Q, Gu Y, Wang Z, Paterson NG, Stansfeld PJ, et al. Structural basis for outer membrane lipopolysaccharide insertion. Nature. 2014;511(7507):52–6. Epub 2014/07/06. doi: 10.1038/nature13464 24990744.
24. Schwechheimer C, Kuehn MJ. Synthetic effect between envelope stress and lack of outer membrane vesicle production in Escherichia coli. J Bacteriol. 2013;195(18):4161–73. Epub 2013/07/16. doi: 10.1128/JB.02192-12 23852867; PubMed Central PMCID: PMC3754735.
25. Qiao S, Luo Q, Zhao Y, Zhang XC, Huang Y. Structural basis for lipopolysaccharide insertion in the bacterial outer membrane. Nature. 2014;511(7507):108–11. Epub 2014/07/06. doi: 10.1038/nature13484 24990751.
26. Stokes JM, French S, Ovchinnikova OG, Bouwman C, Whitfield C, Brown ED. Cold Stress Makes Escherichia coli Susceptible to Glycopeptide Antibiotics by Altering Outer Membrane Integrity. Cell Chem Biol. 2016;23(2):267–77. Epub 2016/02/09. doi: 10.1016/j.chembiol.2015.12.011 26853624.
27. Botos I, Majdalani N, Mayclin SJ, McCarthy JG, Lundquist K, Wojtowicz D, et al. Structural and Functional Characterization of the LPS Transporter LptDE from Gram-Negative Pathogens. Structure. 2016;24(6):965–76. Epub 2016/05/11. doi: 10.1016/j.str.2016.03.026 27161977; PubMed Central PMCID: PMC4899211.
28. Shimada T, Makinoshima H, Ogawa Y, Miki T, Maeda M, Ishihama A. Classification and strength measurement of stationary-phase promoters by use of a newly developed promoter cloning vector. J Bacteriol. 2004;186(21):7112–22. Epub 2004/10/19. doi: 10.1128/JB.186.21.7112-7122.2004 15489422; PubMed Central PMCID: PMC523215.
29. Jones SA, Gibson T, Maltby RC, Chowdhury FZ, Stewart V, Cohen PS, et al. Anaerobic respiration of Escherichia coli in the mouse intestine. Infect Immun. 2011;79(10):4218–26. Epub 2011/08/10. doi: 10.1128/IAI.05395-11 21825069; PubMed Central PMCID: PMC3187261.
30. Miyashita A, Kizaki H, Kawasaki K, Sekimizu K, Kaito C. Primed immune responses to gram-negative peptidoglycans confer infection resistance in silkworms. J Biol Chem. 2014;289(20):14412–21. Epub 2014/04/08. doi: 10.1074/jbc.M113.525139 24706746; PubMed Central PMCID: PMC4022907.
31. Miyashita A, Iyoda S, Ishii K, Hamamoto H, Sekimizu K, Kaito C. Lipopolysaccharide O-antigen of enterohemorrhagic Escherichia coli O157:H7 is required for killing both insects and mammals. FEMS Microbiol Lett. 2012;333(1):59–68. Epub 2012/05/23. doi: 10.1111/j.1574-6968.2012.02599.x 22612664.
32. Miki T, Yamamoto Y, Matsuda H. A novel, simple, high-throughput method for isolation of genome-wide transposon insertion mutants of Escherichia coli K-12. Methods Mol Biol. 2008;416:195–204. Epub 2008/04/09. doi: 10.1007/978-1-59745-321-9_13 18392969.
33. Miller JH. Experiments in molecular genetics. Cold Spring Harbor, N.Y.: Cold Spring Harbor Laboratory; 1972. xvi, 466 p. p.
34. Datsenko KA, Wanner BL. One-step inactivation of chromosomal genes in Escherichia coli K-12 using PCR products. Proc Natl Acad Sci U S A. 2000;97(12):6640–5. Epub 2000/06/01. doi: 10.1073/pnas.120163297 10829079; PubMed Central PMCID: PMC18686.
35. Ellis HM, Yu D, DiTizio T, Court DL. High efficiency mutagenesis, repair, and engineering of chromosomal DNA using single-stranded oligonucleotides. Proc Natl Acad Sci U S A. 2001;98(12):6742–6. Epub 2001/06/07. doi: 10.1073/pnas.121164898 11381128; PubMed Central PMCID: PMC34423.
36. Diner EJ, Garza-Sanchez F, Hayes CS. Genome engineering using targeted oligonucleotide libraries and functional selection. Methods Mol Biol. 2011;765:71–82. Epub 2011/08/05. doi: 10.1007/978-1-61779-197-0_5 21815087; PubMed Central PMCID: PMC3167224.
37. Ellis TN, Leiman SA, Kuehn MJ. Naturally produced outer membrane vesicles from Pseudomonas aeruginosa elicit a potent innate immune response via combined sensing of both lipopolysaccharide and protein components. Infect Immun. 2010;78(9):3822–31. Epub 2010/07/08. doi: 10.1128/IAI.00433-10 20605984; PubMed Central PMCID: PMC2937433.
38. Cunningham FX Jr., Gantt E. A portfolio of plasmids for identification and analysis of carotenoid pathway enzymes: Adonis aestivalis as a case study. Photosynth Res. 2007;92(2):245–59. Epub 2007/07/20. doi: 10.1007/s11120-007-9210-0 17634749.
39. Cunningham FX Jr., Gantt E. A study in scarlet: enzymes of ketocarotenoid biosynthesis in the flowers of Adonis aestivalis. Plant J. 2005;41(3):478–92. Epub 2005/01/22. doi: 10.1111/j.1365-313X.2004.02309.x 15659105.
40. Bligh EG, Dyer WJ. A rapid method of total lipid extraction and purification. Can J Biochem Physiol. 1959;37(8):911–7. Epub 1959/08/01. doi: 10.1139/o59-099 13671378.
41. Chng SS, Ruiz N, Chimalakonda G, Silhavy TJ, Kahne D. Characterization of the two-protein complex in Escherichia coli responsible for lipopolysaccharide assembly at the outer membrane. Proc Natl Acad Sci U S A. 2010;107(12):5363–8. Epub 2010/03/06. doi: 10.1073/pnas.0912872107 20203010; PubMed Central PMCID: PMC2851745.
Článek vyšel v časopise
PLOS Pathogens
2020 Číslo 4
- S diagnostikou Parkinsonovy nemoci může nově pomoci AI nástroj pro hodnocení mrkacího reflexu
- Proč při poslechu některé muziky prostě musíme tančit?
- Chůze do schodů pomáhá prodloužit život a vyhnout se srdečním chorobám
- Metamizol jako analgetikum první volby: kdy, pro koho, jak a proč?
- „Jednohubky“ z klinického výzkumu – 2024/44
Nejčtenější v tomto čísle
- Battling brain-eating amoeba: Enigmas surrounding immunity to Naegleria fowleri
- Drosophila as a model for the gut microbiome
- Genetic interaction analysis comes to the diploid human pathogen Candida albicans
- Human cytomegalovirus IE2 drives transcription initiation from a select subset of late infection viral promoters by host RNA polymerase II
