1887

Abstract

, a Gram-negative bacterium, is an emerging human pathogen equipped with both a type 3 and a type 6 secretion system (T6SS). In this study, we evaluated the roles played by paralogous T6SS effector proteins, hemolysin co-regulated proteins (Hcp-1 and -2) and valine glycine repeat G (VgrG-1, -2 and -3) protein family members in SSU pathogenesis by generating various combinations of deletion mutants of the their genes. In addition to their predicted roles as structural components and effector proteins of the T6SS, our data clearly demonstrated that paralogues of Hcp and VgrG also influenced bacterial motility, protease production and biofilm formation. Surprisingly, there was limited to no observed functional redundancy among and/or between the aforementioned T6SS effector paralogues in multiple assays. Our data indicated that Hcp and VgrG paralogues located within the T6SS cluster were more involved in forming T6SS structures, while the primary roles of Hcp-1 and VgrG-1, located outside of the T6SS cluster, were as T6SS effectors. In terms of influence on bacterial physiology, Hcp-1, but not Hcp-2, influenced bacterial motility and protease production, and in its absence, increases in both of the aforementioned activities were observed. Likewise, VgrG-1 played a major role in regulating bacterial protease production, while VgrG-2 and VgrG-3 were critical in regulating bacterial motility and biofilm formation. In an intraperitoneal murine model of infection, all Hcp and VgrG paralogues were required for optimal bacterial virulence and dissemination to mouse peripheral organs. Importantly, the observed phenotypic alterations of the T6SS mutants could be fully complemented. Taking these results together, we have further established the roles played by the two known T6SS effectors of by defining their contributions to T6SS function and virulence in both and models of infection.

Loading

Article metrics loading...

/content/journal/micro/10.1099/mic.0.063495-0
2013-06-01
2024-12-05
Loading full text...

Full text loading...

/deliver/fulltext/micro/159/6/1120.html?itemId=/content/journal/micro/10.1099/mic.0.063495-0&mimeType=html&fmt=ahah

References

  1. Altwegg M., Martinetti Lucchini G., Lüthy-Hottenstein J., Rohrbach M. ( 1991). Aeromonas-associated gastroenteritis after consumption of contaminated shrimp. Eur J Clin Microbiol Infect Dis 10:44–45[PubMed] [CrossRef]
    [Google Scholar]
  2. Aschtgen M. S., Bernard C. S., De Bentzmann S., Lloubès R., Cascales E. ( 2008). SciN is an outer membrane lipoprotein required for type VI secretion in enteroaggregative Escherichia coli. . J Bacteriol 190:7523–7531 [View Article][PubMed]
    [Google Scholar]
  3. Ballister E. R., Lai A. H., Zuckermann R. N., Cheng Y., Mougous J. D. ( 2008). In vitro self-assembly of tailorable nanotubes from a simple protein building block. Proc Natl Acad Sci U S A 105:3733–3738 [View Article][PubMed]
    [Google Scholar]
  4. Basler M., Pilhofer M., Henderson G. P., Jensen G. J., Mekalanos J. J. ( 2012). Type VI secretion requires a dynamic contractile phage tail-like structure. Nature 483:182–186 [View Article][PubMed]
    [Google Scholar]
  5. Bröms J. E., Meyer L., Lavander M., Larsson P., Sjöstedt A. ( 2012). DotU and VgrG, core components of type VI secretion systems, are essential for Francisella LVS pathogenicity. PLoS ONE 7:e34639 [View Article][PubMed]
    [Google Scholar]
  6. Brouqui P., Raoult D. ( 2001). Endocarditis due to rare and fastidious bacteria. Clin Microbiol Rev 14:177–207 [View Article][PubMed]
    [Google Scholar]
  7. Cascales E. ( 2008). The type VI secretion toolkit. EMBO Rep 9:735–741 [View Article][PubMed]
    [Google Scholar]
  8. Castillo M. ( 2012). Flesh-eating infection victim Aimee Copeland recovering in rehab with positive outlook. http://www.cbsnews.com/8301-504763_162-57466495-10391704/flesh-eating-infection-victim-aimee-copeland-recovering-in-rehab-with-positive-outlook/
    [Google Scholar]
  9. Chopra A. K., Houston C. W. ( 1999). Enterotoxins in Aeromonas-associated gastroenteritis. Microbes Infect 1:1129–1137 [View Article][PubMed]
    [Google Scholar]
  10. Chopra A. K., Houston C. W., Peterson J. W., Jin G. F. ( 1993). Cloning, expression, and sequence analysis of a cytolytic enterotoxin gene from Aeromonas hydrophila. . Can J Microbiol 39:513–523 [View Article][PubMed]
    [Google Scholar]
  11. Chopra A. K., Peterson J. W., Xu X. J., Coppenhaver D. H., Houston C. W. ( 1996). Molecular and biochemical characterization of a heat-labile cytotonic enterotoxin from Aeromonas hydrophila. . Microb Pathog 21:357–377 [View Article][PubMed]
    [Google Scholar]
  12. Das S., Chakrabortty A., Banerjee R., Chaudhuri K. ( 2002). Involvement of in vivo induced icmF gene of Vibrio cholerae in motility, adherence to epithelial cells, and conjugation frequency. Biochem Biophys Res Commun 295:922–928 [View Article][PubMed]
    [Google Scholar]
  13. Edberg S. C., Browne F. A., Allen M. J. ( 2007). Issues for microbial regulation: Aeromonas as a model. Crit Rev Microbiol 33:89–100 [View Article][PubMed]
    [Google Scholar]
  14. Edwards R. A., Keller L. H., Schifferli D. M. ( 1998). Improved allelic exchange vectors and their use to analyze 987P fimbria gene expression. Gene 207:149–157 [View Article][PubMed]
    [Google Scholar]
  15. Enos-Berlage J. L., Guvener Z. T., Keenan C. E., McCarter L. L. ( 2005). Genetic determinants of biofilm development of opaque and translucent Vibrio parahaemolyticus. . Mol Microbiol 55:1160–1182 [View Article][PubMed]
    [Google Scholar]
  16. Erova T. E., Pillai L., Fadl A. A., Sha J., Wang S., Galindo C. L., Chopra A. K. ( 2006). DNA adenine methyltransferase influences the virulence of Aeromonas hydrophila. . Infect Immun 74:410–424 [View Article][PubMed]
    [Google Scholar]
  17. Galindo C. L., Sha J., Fadl A. A., Pillai L., Chopra A. K. ( 2006). Host immune responses to Aeromonas virulence factors. Curr Immunol Rev 2:13–26 [View Article]
    [Google Scholar]
  18. Hachani A., Lossi N. S., Hamilton A., Jones C., Bleves S., Albesa-Jové D., Filloux A. ( 2011). Type VI secretion system in Pseudomonas aeruginosa: secretion and multimerization of VgrG proteins. J Biol Chem 286:12317–12327 [View Article][PubMed]
    [Google Scholar]
  19. Hood R. D., Singh P., Hsu F., Güvener T., Carl M. A., Trinidad R. R., Silverman J. M., Ohlson B. B., Hicks K. G. & other authors ( 2010). A type VI secretion system of Pseudomonas aeruginosa targets a toxin to bacteria. Cell Host Microbe 7:25–37 [View Article][PubMed]
    [Google Scholar]
  20. Janda J. M., Guthertz L. S., Kokka R. P., Shimada T. ( 1994). Aeromonas species in septicemia: laboratory characteristics and clinical observations. Clin Infect Dis 19:77–83 [View Article][PubMed]
    [Google Scholar]
  21. Khajanchi B. K., Sha J., Kozlova E. V., Erova T. E., Suarez G., Sierra J. C., Popov V. L., Horneman A. J., Chopra A. K. ( 2009). N-acylhomoserine lactones involved in quorum sensing control the type VI secretion system, biofilm formation, protease production, and in vivo virulence in a clinical isolate of Aeromonas hydrophila. . Microbiology 155:3518–3531 [View Article][PubMed]
    [Google Scholar]
  22. Kirov S. M. ( 1993). The public health significance of Aeromonas spp. in foods. Int J Food Microbiol 20:179–198 [View Article][PubMed]
    [Google Scholar]
  23. Kozlova E. V., Popov V. L., Sha J., Foltz S. M., Erova T. E., Agar S. L., Horneman A. J., Chopra A. K. ( 2008). Mutation in the S-ribosylhomocysteinase (luxS) gene involved in quorum sensing affects biofilm formation and virulence in a clinical isolate of Aeromonas hydrophila. . Microb Pathog 45:343–354 [View Article][PubMed]
    [Google Scholar]
  24. Kühn I., Albert M. J., Ansaruzzaman M., Bhuiyan N. A., Alabi S. A., Islam M. S., Neogi P. K., Huys G., Janssen P. & other authors ( 1997). Characterization of Aeromonas spp. isolated from humans with diarrhea, from healthy controls, and from surface water in Bangladesh. J Clin Microbiol 35:369–373[PubMed]
    [Google Scholar]
  25. Leiman P. G., Basler M., Ramagopal U. A., Bonanno J. B., Sauder J. M., Pukatzki S., Burley S. K., Almo S. C., Mekalanos J. J. ( 2009). Type VI secretion apparatus and phage tail-associated protein complexes share a common evolutionary origin. Proc Natl Acad Sci U S A 106:4154–4159 [View Article][PubMed]
    [Google Scholar]
  26. Ma A. T., McAuley S., Pukatzki S., Mekalanos J. J. ( 2009). Translocation of a Vibrio cholerae type VI secretion effector requires bacterial endocytosis by host cells. Cell Host Microbe 5:234–243 [View Article][PubMed]
    [Google Scholar]
  27. Merino S., Rubires X., Knochel S., Tomas J. M. ( 1995). Emerging pathogens: Aeromonas spp. Int J Food Microbiol 28:157–168 [View Article][PubMed]
    [Google Scholar]
  28. Morohoshi T., Shiono T., Takidouchi K., Kato M., Kato N., Kato J., Ikeda T. ( 2007). Inhibition of quorum sensing in Serratia marcescens AS-1 by synthetic analogs of N-acylhomoserine lactone. Appl Environ Microbiol 73:6339–6344 [View Article][PubMed]
    [Google Scholar]
  29. Mougous J. D., Cuff M. E., Raunser S., Shen A., Zhou M., Gifford C. A., Goodman A. L., Joachimiak G., Ordoñez C. L. & other authors ( 2006). A virulence locus of Pseudomonas aeruginosa encodes a protein secretion apparatus. Science 312:1526–1530 [View Article][PubMed]
    [Google Scholar]
  30. Palú A. P., Gomes L. M., Miguel M. A., Balassiano I. T., Queiroz M. L., Freitas-Almeida A. C., de Oliveira S. S. ( 2006). Antimicrobial resistance in food and clinical Aeromonas isolates. Food Microbiol 23:504–509 [View Article][PubMed]
    [Google Scholar]
  31. Podladchikova O., Antonenka U., Heesemann J., Rakin A. ( 2011). Yersinia pestis autoagglutination factor is a component of the type six secretion system. Int J Med Microbiol 301:562–569 [View Article][PubMed]
    [Google Scholar]
  32. Pukatzki S., Ma A. T., Sturtevant D., Krastins B., Sarracino D., Nelson W. C., Heidelberg J. F., Mekalanos J. J. ( 2006). Identification of a conserved bacterial protein secretion system in Vibrio cholerae using the Dictyostelium host model system.. Proc Natl Acad Sci U S A 1031528–1533 [View Article]
    [Google Scholar]
  33. Pukatzki S., Ma A. T., Revel A. T., Sturtevant D., Mekalanos J. J. ( 2007). Type VI secretion system translocates a phage tail spike-like protein into target cells where it cross-links actin. Proc Natl Acad Sci U S A 104:15508–15513 [View Article][PubMed]
    [Google Scholar]
  34. Pukatzki S., McAuley S. B., Miyata S. T. ( 2009). The type VI secretion system: translocation of effectors and effector-domains. Curr Opin Microbiol 12:11–17 [View Article][PubMed]
    [Google Scholar]
  35. Rabaan A. A., Gryllos I., Tomás J. M., Shaw J. G. ( 2001). Motility and the polar flagellum are required for Aeromonas caviae adherence to HEp-2 cells. Infect Immun 69:4257–4267 [View Article][PubMed]
    [Google Scholar]
  36. Records A. R. ( 2011). The type VI secretion system: a multipurpose delivery system with a phage-like machinery. Mol Plant Microbe Interact 24:751–757 [View Article][PubMed]
    [Google Scholar]
  37. Seshadri R., Joseph S. W., Chopra A. K., Sha J., Shaw J., Graf J., Haft D., Wu M., Ren Q. & other authors ( 2006). Genome sequence of Aeromonas hydrophila ATCC 7966T: jack of all trades. J Bacteriol 188:8272–8282 [View Article][PubMed]
    [Google Scholar]
  38. Sha J., Agar S. L., Baze W. B., Olano J. P., Fadl A. A., Erova T. E., Wang S., Foltz S. M., Suarez G. & other authors ( 2008). Braun lipoprotein (Lpp) contributes to virulence of Yersiniae: potential role of Lpp in inducing bubonic and pneumonic plague. Infect Immun 76:1390–1409 [View Article][PubMed]
    [Google Scholar]
  39. Sierra J. C., Suarez G., Sha J., Foltz S. M., Popov V. L., Galindo C. L., Garner H. R., Chopra A. K. ( 2007). Biological characterization of a new type III secretion system effector from a clinical isolate of Aeromonas hydrophila–part II. Microb Pathog 43:147–160 [View Article][PubMed]
    [Google Scholar]
  40. Sierra J. C., Suarez G., Sha J., Baze W. B., Foltz S. M., Chopra A. K. ( 2010). Unraveling the mechanism of action of a new type III secretion system effector AexU from Aeromonas hydrophila. . Microb Pathog 49:122–134 [View Article][PubMed]
    [Google Scholar]
  41. Sierra J. C., Suarez G., Chopra A. K. ( 2011). An intriguing emerging human pathogen Aeromonas hydrophila: pathogenesis and vaccines. Vaccines Against Bacterial Biothreat Pathogens1–34 Feodorova V. A., Motin. V. L. Kerala, India: Research Signpost;
    [Google Scholar]
  42. Silver A. C., Rabinowitz N. M., Küffer S., Graf J. ( 2007). Identification of Aeromonas veronii genes required for colonization of the medicinal leech, Hirudo verbana . J Bacteriol 189:6763–6772 [View Article][PubMed]
    [Google Scholar]
  43. Silverman J. M., Brunet Y. R., Cascales E., Mougous J. D. ( 2012). Structure and regulation of the Type VI secretion system. Annu Rev Mcrobiol 66453–472 [View Article]
    [Google Scholar]
  44. Suarez G., Sierra J. C., Sha J., Wang S., Erova T. E., Fadl A. A., Foltz S. M., Horneman A. J., Chopra A. K. ( 2008). Molecular characterization of a functional type VI secretion system from a clinical isolate of Aeromonas hydrophila . Microb Pathog 44:344–361 [View Article][PubMed]
    [Google Scholar]
  45. Suarez G., Sierra J. C., Erova T. E., Sha J., Horneman A. J., Chopra A. K. ( 2010a). A type VI secretion system effector protein, VgrG1, from Aeromonas hydrophila that induces host cell toxicity by ADP ribosylation of actin. J Bacteriol 192:155–168 [View Article][PubMed]
    [Google Scholar]
  46. Suarez G., Sierra J. C., Kirtley M. L., Chopra A. K. ( 2010b). Role of Hcp, a type 6 secretion system effector, of Aeromonas hydrophila in modulating activation of host immune cells. Microbiology 156:3678–3688 [View Article][PubMed]
    [Google Scholar]
  47. Weber B., Hasic M., Chen C., Wai S. N., Milton D. L. ( 2009). Type VI secretion modulates quorum sensing and stress response in Vibrio anguillarum. . Environ Microbiol 11:3018–3028 [View Article][PubMed]
    [Google Scholar]
  48. Zhang L., Hinz A. J., Nadeau J. P., Mah T. F. ( 2011). Pseudomonas aeruginosa tssC1 links type VI secretion and biofilm-specific antibiotic resistance. J Bacteriol 193:5510–5513 [View Article][PubMed]
    [Google Scholar]
  49. Zheng J., Ho B., Mekalanos J. J. ( 2011). Genetic analysis of anti-amoebae and anti-bacterial activities of the type VI secretion system in Vibrio cholerae. . PLoS ONE 6:e23876 [View Article][PubMed]
    [Google Scholar]
/content/journal/micro/10.1099/mic.0.063495-0
Loading
/content/journal/micro/10.1099/mic.0.063495-0
Loading

Data & Media loading...

This is a required field
Please enter a valid email address
Approval was a Success
Invalid data
An Error Occurred
Approval was partially successful, following selected items could not be processed due to error