1887

Abstract

Salmonellosis is a major health problem worldwide. serovar Enteritidis (. Enteritidis) has been a primary cause of outbreaks in many countries. AvrA is an SPI-1 effector protein involved in the enteritis pathway, with critical roles in inhibiting inflammation and apoptosis. In this work, we constructed an AvrA-FLAG-tagged strain of Enteritidis to analyse the expression profile of AvrA , in cell culture and . AvrA expression and secretion were observed under culture conditions that mimicked intestinal and intracellular environments. In agreement, bacteria isolated from infected cell monolayers expressed and translocated AvrA for at least 24 h post-inoculation. For experiments, BALB/c mice were inoculated by the natural route of infection with the AvrA-FLAG strain. Infecting bacteria and infected cells were recovered from mesenteric lymph nodes (MLN). Our results showed that AvrA continues to be synthesized up to day 8 post-inoculation. Moreover, AvrA translocation was detected in the cytosol of cells isolated from MLN 8 days after infection. Interestingly, we observed that AvrA is secreted by both type three secretion system (T3SS)-1 and T3SS-2. In summary, these findings indicate that AvrA expression is not constrained to the initial host–bacteria encounter in the intestinal environment as defined previously. The AvrA effector may participate also in systemic Enteritidis infection.

Funding
This study was supported by the:
  • Consejo Nacional de Investigaciones Científicas y Técnicas, Argentina (Award PIP 2012-2014 GI 11220110100911)
  • Secretaría de Ciencia y Técnica de la Universidad de Buenos Aires, Argentina (Award UBACyT 20020110200087, 20020100100541 y 20020120200021BA)
Loading

Article metrics loading...

/content/journal/micro/10.1099/mic.0.077115-0
2014-06-01
2021-07-30
Loading full text...

Full text loading...

/deliver/fulltext/micro/160/6/1191.html?itemId=/content/journal/micro/10.1099/mic.0.077115-0&mimeType=html&fmt=ahah

References

  1. Bäumler A. J., Hargis B. M., Tsolis R. M. ( 2000). Tracing the origins of Salmonella outbreaks. Science 287:50–52 [View Article][PubMed]
    [Google Scholar]
  2. Ben-Barak Z., Streckel W., Yaron S., Cohen S., Prager R., Tschäpe H. ( 2006). The expression of the virulence-associated effector protein gene avrA is dependent on a Salmonella enterica-specific regulatory function. Int J Med Microbiol 296:25–38 [View Article][PubMed]
    [Google Scholar]
  3. Bollaerts K., Aerts M., Faes C., Grijspeerdt K., Dewulf J., Mintiens K. ( 2008). Human salmonellosis: estimation of dose-illness from outbreak data. Risk Anal 28:427–440 [View Article][PubMed]
    [Google Scholar]
  4. Boyle E. C., Bishop J. L., Grassl G. A., Finlay B. B. ( 2007). Salmonella: from pathogenesis to therapeutics. J Bacteriol 189:1489–1495 [View Article][PubMed]
    [Google Scholar]
  5. Broberg C. A., Orth K. ( 2010). Tipping the balance by manipulating post-translational modifications. Curr Opin Microbiol 13:34–40 [View Article][PubMed]
    [Google Scholar]
  6. Brumell J. H., Kujat-Choy S., Brown N. F., Vallance B. A., Knodler L. A., Finlay B. B. ( 2003). SopD2 is a novel type III secreted effector of Salmonella typhimurium that targets late endocytic compartments upon delivery into host cells. Traffic 4:36–48 [View Article][PubMed]
    [Google Scholar]
  7. Bruno V. M., Hannemann S., Lara-Tejero M., Flavell R. A., Kleinstein S. H., Galán J. E. ( 2009). Salmonella Typhimurium type III secretion effectors stimulate innate immune responses in cultured epithelial cells. PLoS Pathog 5:e1000538 [View Article][PubMed]
    [Google Scholar]
  8. Collazo C. M., Galán J. E. ( 1997). The invasion-associated type-III protein secretion system in Salmonella – a review. Gene 192:51–59 [View Article][PubMed]
    [Google Scholar]
  9. Collier-Hyams L. S., Zeng H., Sun J., Tomlinson A. D., Bao Z. Q., Chen H., Madara J. L., Orth K., Neish A. S. ( 2002). Cutting edge: Salmonella AvrA effector inhibits the key proinflammatory, anti-apoptotic NF-κB pathway. J Immunol 169:2846–2850[PubMed] [CrossRef]
    [Google Scholar]
  10. Datsenko K. A., Wanner B. L. ( 2000). One-step inactivation of chromosomal genes in Escherichia coli K-12 using PCR products. Proc Natl Acad Sci U S A 97:6640–6645 [View Article][PubMed]
    [Google Scholar]
  11. Drecktrah D., Knodler L. A., Galbraith K., Steele-Mortimer O. ( 2005). The Salmonella SPI1 effector SopB stimulates nitric oxide production long after invasion. Cell Microbiol 7:105–113 [View Article][PubMed]
    [Google Scholar]
  12. Du F., Galán J. E. ( 2009). Selective inhibition of type III secretion activated signaling by the Salmonella effector AvrA. PLoS Pathog 5:e1000595 [View Article][PubMed]
    [Google Scholar]
  13. Ehrbar K., Friebel A., Miller S. I., Hardt W. D. ( 2003). Role of the Salmonella pathogenicity island 1 (SPI-1) protein InvB in type III secretion of SopE and SopE2, two Salmonella effector proteins encoded outside of SPI-1. J Bacteriol 185:6950–6967 [View Article][PubMed]
    [Google Scholar]
  14. Ellermeier J. R., Slauch J. M. ( 2007). Adaptation to the host environment: regulation of the SPI1 type III secretion system in Salmonella enterica serovar Typhimurium. Curr Opin Microbiol 10:24–29 [View Article][PubMed]
    [Google Scholar]
  15. Galyov E. E., Wood M. W., Rosqvist R., Mullan P. B., Watson P. R., Hedges S., Wallis T. S. ( 1997). A secreted effector protein of Salmonella dublin is translocated into eukaryotic cells and mediates inflammation and fluid secretion in infected ileal mucosa. Mol Microbiol 25:903–912 [View Article][PubMed]
    [Google Scholar]
  16. Geddes K., Worley M., Niemann G., Heffron F. ( 2005). Identification of new secreted effectors in Salmonella enterica serovar Typhimurium. Infect Immun 73:6260–6271 [View Article][PubMed]
    [Google Scholar]
  17. Giacomodonato M. N., Uzzau S., Bacciu D., Caccuri R., Sarnacki S. H., Rubino S., Cerquetti M. C. ( 2007). SipA, SopA, SopB, SopD and SopE2 effector proteins of Salmonella enterica serovar Typhimurium are synthesized at late stages of infection in mice. Microbiology 153:1221–1228 [View Article][PubMed]
    [Google Scholar]
  18. Giacomodonato M. N., Sarnacki S. H., Llana M. N., García Cattaneo A. S., Uzzau S., Rubino S., Cerquetti M. C. ( 2009). Impaired synthesis and secretion of SopA in Salmonella Typhimurium dam mutants. FEMS Microbiol Lett 292:71–77 [View Article][PubMed]
    [Google Scholar]
  19. Giacomodonato M. N., Sarnacki S. H., Llana M. N., Cerquetti M. C. ( 2011). SopB effector protein of Salmonella Typhimurium is translocated in mesenteric lymph nodes during murine salmonellosis. FEMS Microbiol Lett 317:100–106 [View Article][PubMed]
    [Google Scholar]
  20. Gong H., Vu G.-P., Bai Y., Yang E., Liu F., Lu S. ( 2010). Differential expression of Salmonella type III secretion system factors InvJ, PrgJ, SipC, SipD, SopA and SopB in cultures and in mice. Microbiology 156:116–127 [View Article][PubMed]
    [Google Scholar]
  21. Hald T., Vose D., Wegener H. C., Koupeev T. ( 2004). A Bayesian approach to quantify the contribution of animal-food sources to human salmonellosis. Risk Anal 24:255–269 [View Article][PubMed]
    [Google Scholar]
  22. Hapfelmeier S., Ehrbar K., Stecher B., Barthel M., Kremer M., Hardt W.-D. ( 2004). Role of the Salmonella pathogenicity island 1 effector proteins SipA, SopB, SopE, and SopE2 in Salmonella enterica subspecies 1 serovar Typhimurium colitis in streptomycin-pretreated mice. Infect Immun 72:795–809 [View Article][PubMed]
    [Google Scholar]
  23. Hardt W. D., Galán J. E. ( 1997). A secreted Salmonella protein with homology to an avirulence determinant of plant pathogenic bacteria. Proc Natl Acad Sci U S A 94:9887–9892 [View Article][PubMed]
    [Google Scholar]
  24. Hendriksen R. S., Vieira A. R., Karlsmose S., Lo Fo Wong D. M. A., Jensen A. B., Wegener H. C., Aarestrup F. M. ( 2011). Global monitoring of Salmonella serovar distribution from the World Health Organization Global Foodborne Infections Network Country Data Bank: results of quality assured laboratories from 2001 to 2007. Foodborne Pathog Dis 8:887–900 [View Article][PubMed]
    [Google Scholar]
  25. Hogue A., White P., Guard-Petter J., Schlosser W., Gast R., Ebel E., Farrar J., Gomez T., Madden J. & other authors ( 1997). Epidemiology and control of egg-associated Salmonella Enteritidis in the United States of America. Rev Sci Tech 16:542–553[PubMed]
    [Google Scholar]
  26. Jones R. M., Wu H., Wentworth C., Luo L., Collier-Hyams L., Neish A. S. ( 2008). Salmonella AvrA coordinates suppression of host immune and apoptotic defenses via JNK pathway blockade. Cell Host Microbe 3:233–244 [View Article][PubMed]
    [Google Scholar]
  27. Kerrinnes T., Zelas Z. B., Streckel W., Faber F., Tietze E., Tschäpe H., Yaron S. ( 2009). CsrA and CsrB are required for the post-transcriptional control of the virulence-associated effector protein AvrA of Salmonella enterica . Int J Med Microbiol 299:333–341 [View Article][PubMed]
    [Google Scholar]
  28. Knodler L. A., Finlay B. B., Steele-Mortimer O. ( 2005). The Salmonella effector protein SopB protects epithelial cells from apoptosis by sustained activation of Akt. J Biol Chem 280:9058–9064 [View Article][PubMed]
    [Google Scholar]
  29. Kubori T., Galán J. E. ( 2003). Temporal regulation of Salmonella virulence effector function by proteasome-dependent protein degradation. Cell 115:333–342 [View Article][PubMed]
    [Google Scholar]
  30. Kubori T., Sukhan A., Aizawa S. I., Galán J. E. ( 2000). Molecular characterization and assembly of the needle complex of the Salmonella typhimurium type III protein secretion system. Proc Natl Acad Sci U S A 97:10225–10230 [View Article][PubMed]
    [Google Scholar]
  31. Lawley T. D., Chan K., Thompson L. J., Kim C. C., Govoni G. R., Monack D. M. ( 2006). Genome-wide screen for Salmonella genes required for long-term systemic infection of the mouse. PLoS Pathog 2:e11 [View Article][PubMed]
    [Google Scholar]
  32. Liao A. P., Petrof E. O., Kuppireddi S., Zhao Y., Xia Y., Claud E. C., Sun J. ( 2008). Salmonella type III effector AvrA stabilizes cell tight junctions to inhibit inflammation in intestinal epithelial cells. PLoS ONE 3:e2369 [View Article][PubMed]
    [Google Scholar]
  33. Livak K. J., Schmittgen T. D. ( 2001). Analysis of relative gene expression data using real-time quantitative PCR and the 2−ΔΔ C T method. Methods 25:402–408 [View Article][PubMed]
    [Google Scholar]
  34. Löber S., Jäckel D., Kaiser N., Hensel M. ( 2006). Regulation of Salmonella pathogenicity island 2 genes by independent environmental signals. Int J Med Microbiol 296:435–447 [View Article][PubMed]
    [Google Scholar]
  35. Lu R., Liu X., Wu S., Xia Y., Zhang Y. G., Petrof E. O., Claud E. C., Sun J. ( 2012). Consistent activation of the β-catenin pathway by Salmonella type-three secretion effector protein AvrA in chronically infected intestine. Am J Physiol Gastrointest Liver Physiol 303:G1113–G1125 [View Article][PubMed]
    [Google Scholar]
  36. Majowicz S. E., Musto J., Scallan E., Angulo F. J., Kirk M., O’Brien S. J., Jones T. F., Fazil A., Hoekstra R. M. International Collaboration on Enteric Disease ‘Burden of Illness’ Studies ( 2010). The global burden of nontyphoidal Salmonella gastroenteritis. Clin Infect Dis 50:882–889 [View Article][PubMed]
    [Google Scholar]
  37. Marcus S. L., Knodler L. A., Finlay B. B. ( 2002). Salmonella enterica serovar Typhimurium effector SigD/SopB is membrane-associated and ubiquitinated inside host cells. Cell Microbiol 4:435–446 [View Article][PubMed]
    [Google Scholar]
  38. McGhie E. J., Brawn L. C., Hume P. J., Humphreys D., Koronakis V. ( 2009). Salmonella takes control: effector-driven manipulation of the host. Curr Opin Microbiol 12:117–124 [View Article][PubMed]
    [Google Scholar]
  39. Miki T., Okada N., Danbara H. ( 2004). Two periplasmic disulfide oxidoreductases, DsbA and SrgA, target outer membrane protein SpiA, a component of the Salmonella pathogenicity island 2 type III secretion system. J Biol Chem 279:34631–34642 [View Article][PubMed]
    [Google Scholar]
  40. Moore G., Blair I. S., McDowell D. A. ( 2007). Recovery and transfer of Salmonella Typhimurium from four different domestic food contact surfaces. J Food Prot 70:2273–2280[PubMed]
    [Google Scholar]
  41. Morales R. A., McDowell R. M. ( 1999). Economic consequences of Salmonella enterica serovar Enteritidis infection in humans and the U. S. egg industry. Salmonella Enterica Serovar Enteritidis in Humans and Animals271–290 Saeed A. M., Gast R. K., Potter M. E., Wall P. G. Ames, IA: Iowa State University Press;
    [Google Scholar]
  42. Niemann G. S., Brown R. N., Gustin J. K., Stufkens A., Shaikh-Kidwai A. S., Li J., McDermott J. E., Brewer H. M., Schepmoes A. & other authors ( 2011). Discovery of novel secreted virulence factors from Salmonella enterica serovar Typhimurium by proteomic analysis of culture supernatants. Infect Immun 79:33–43 [View Article][PubMed]
    [Google Scholar]
  43. Oliveira K., Oliveira T., Teixeira P., Azeredo J., Henriques M., Oliveira R. ( 2006). Comparison of the adhesion ability of different Salmonella Enteritidis serotypes to materials used in kitchens. J Food Prot 69:2352–2356[PubMed]
    [Google Scholar]
  44. Pang J.-C., Chiu T.-H., Helmuth R., Schroeter A., Guerra B., Tsen H.-Y. ( 2007). A pulsed field gel electrophoresis (PFGE) study that suggests a major world-wide clone of Salmonella enterica serovar Enteritidis. Int J Food Microbiol 116:305–312 [View Article][PubMed]
    [Google Scholar]
  45. Patel J. C., Hueffer K., Lam T. T., Galán J. E. ( 2009). Diversification of a Salmonella virulence protein function by ubiquitin-dependent differential localization. Cell 137:283–294 [View Article][PubMed]
    [Google Scholar]
  46. Perales I., Audicana A. ( 1988). Salmonella Enteritidis and eggs. Lancet 332:1133 [View Article][PubMed]
    [Google Scholar]
  47. Prager R., Mirold S., Tietze E., Strutz U., Knüppel B., Rabsch W., Hardt W. D., Tschäpe H. ( 2000). Prevalence and polymorphism of genes encoding translocated effector proteins among clinical isolates of Salmonella enterica . Int J Med Microbiol 290:605–617 [View Article][PubMed]
    [Google Scholar]
  48. Prager R., Rabsch W., Streckel W., Voigt W., Tietze E., Tschäpe H. ( 2003). Molecular properties of Salmonella enterica serotype Paratyphi B distinguish between its systemic and its enteric pathovars. J Clin Microbiol 41:4270–4278 [View Article][PubMed]
    [Google Scholar]
  49. Raffatellu M., Wilson R. P., Chessa D., Andrews-Polymenis H., Tran Q. T., Lawhon S., Khare S., Adams L. G., Bäumler A. J. ( 2005). SipA, SopA, SopB, SopD, and SopE2 contribute to Salmonella enterica serotype Typhimurium invasion of epithelial cells. Infect Immun 73:146–154 [View Article][PubMed]
    [Google Scholar]
  50. Rodrigue D. C., Tauxe R. V., Rowe B. ( 1990). International increase in Salmonella enteritidis: a new pandemic?. Epidemiol Infect 105:21–27 [View Article][PubMed]
    [Google Scholar]
  51. Stecher B., Robbiani R., Walker A. W., Westendorf A. M., Barthel M., Kremer M., Chaffron S., Macpherson A. J., Buer J. & other authors ( 2007). Salmonella enterica serovar Typhimurium exploits inflammation to compete with the intestinal microbiota. PLoS Biol 5:2177–2189 [View Article][PubMed]
    [Google Scholar]
  52. Streckel W., Wolff A.-C., Prager R., Tietze E., Tschäpe H. ( 2004). Expression profiles of effector proteins SopB, SopD1, SopE1, and AvrA differ with systemic, enteric, and epidemic strains of Salmonella enterica . Mol Nutr Food Res 48:496–503 [View Article][PubMed]
    [Google Scholar]
  53. Sun J. ( 2009). Pathogenic bacterial proteins and their anti-inflammatory effects in the eukaryotic host. Antiinflamm Antiallergy Agents Med Chem 8:214–227 [View Article][PubMed]
    [Google Scholar]
  54. Uzzau S., Figueroa-Bossi N., Rubino S., Bossi L. ( 2001). Epitope tagging of chromosomal genes in Salmonella . Proc Natl Acad Sci U S A 98:15264–15269 [View Article][PubMed]
    [Google Scholar]
  55. Waterman S. R., Holden D. W. ( 2003). Functions and effectors of the Salmonella pathogenicity island 2 type III secretion system. Cell Microbiol 5:501–511 [View Article][PubMed]
    [Google Scholar]
  56. Wu S., Ye Z., Liu X., Zhao Y., Xia Y., Steiner A., Petrof E. O., Claud E. C., Sun J. ( 2010). Salmonella typhimurium infection increases p53 acetylation in intestinal epithelial cells. Am J Physiol Gastrointest Liver Physiol 298:G784–G794 [View Article][PubMed]
    [Google Scholar]
  57. Wu H., Jones R. M., Neish A. S. ( 2012). The Salmonella effector AvrA mediates bacterial intracellular survival during infection in vivo. Cell Microbiol 14:28–39 [View Article][PubMed]
    [Google Scholar]
  58. Ye Z., Petrof E. O., Boone D., Claud E. C., Sun J. ( 2007). Salmonella effector AvrA regulation of colonic epithelial cell inflammation by deubiquitination. Am J Pathol 171:882–892 [View Article][PubMed]
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/micro/10.1099/mic.0.077115-0
Loading
/content/journal/micro/10.1099/mic.0.077115-0
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