1887

Abstract

The virulence plasmid of the pathogenic actinomycete is essential for proliferation of this pathogen in macrophages and the development of disease. The pathogenicity island of this plasmid encodes a family of virulence-associated proteins (Vap), one of which (VapA) is a virulence factor. This paper describes the operon ( o-expressed ene), located upstream of the operon. Transcription of the operon gave rise to transcripts with a half-life similar to those determined for other virulence plasmid genes (1.8 min). Transcription started at a promoter similar to the promoter, and proceeded through an inefficient terminator into the downstream gene. In addition, is also transcribed from a promoter downstream of The and operons were coordinately regulated by temperature and pH in a synergistic manner. The latter parameter only affected transcription at higher growth temperatures, indicating that temperature is the dominant regulatory signal. Transcription of the operon increased 10-fold during the late exponential and stationary growth phases. Transcription was also upregulated during the initial hours following phagocytosis by phagocytic cells. In contrast to and , the gene is conserved in the porcine VapB-encoding plasmid, as well as in pathogenic mycobacteria. The coordinated regulation of and , transcription of following phagocytosis and conservation of in pathogenic mycobacteria indicate a role for and the genes in the virulence of .

Funding
This study was supported by the:
  • Research Stimulus Fund of the Department of Agriculture, Fisheries and Food (Award RSF 06-379)
  • Science Foundation Ireland (Award 02/IN.1/B203)
Loading

Article metrics loading...

/content/journal/micro/10.1099/mic.0.049759-0
2011-08-01
2021-08-04
Loading full text...

Full text loading...

/deliver/fulltext/micro/157/8/2357.html?itemId=/content/journal/micro/10.1099/mic.0.049759-0&mimeType=html&fmt=ahah

References

  1. Butcher P. D., Mangan J. A., Monahan I. M. ( 1998). Intracellular gene expression. Analysis of RNA from mycobacteria in macrophages using RT-PCR. Methods Mol Biol 101:285–306[PubMed]
    [Google Scholar]
  2. Buttner M. J., Chater K. F., Bibb M. J. ( 1990). Cloning, disruption, and transcriptional analysis of three RNA polymerase sigma factor genes of Streptomyces coelicolor A3(2). J Bacteriol 172:3367–3378[PubMed]
    [Google Scholar]
  3. Byrne B. A., Prescott J. F., Palmer G. H., Takai S., Nicholson V. M., Alperin D. C., Hines S. A. ( 2001). Virulence plasmid of Rhodococcus equi contains inducible gene family encoding secreted proteins. Infect Immun 69:650–656 [View Article][PubMed]
    [Google Scholar]
  4. Byrne G. A., Russell D. A., Chen X., Meijer W. G. ( 2007). Transcriptional regulation of the virR operon of the intracellular pathogen Rhodococcus equi . J Bacteriol 189:5082–5089 [View Article][PubMed]
    [Google Scholar]
  5. Byrne G. A., Boland C. A., O’Connell E. P., Meijer W. G. ( 2008). Differential mRNA stability of the vapAICD operon of the facultative intracellular pathogen Rhodococcus equi . FEMS Microbiol Lett 280:89–94 [View Article][PubMed]
    [Google Scholar]
  6. Cole S. T., Brosch R., Parkhill J., Garnier T., Churcher C., Harris D., Gordon S. V., Eiglmeier K., Gas S. et al. ( 1998). Deciphering the biology of Mycobacterium tuberculosis from the complete genome sequence. Nature 393:537–544 [View Article][PubMed]
    [Google Scholar]
  7. De La Peña-Moctezuma A., Prescott J. F., Goodfellow M. ( 1996). Attempts to find phenotypic markers of the virulence plasmid of Rhodococcus equi . Can J Vet Res 60:29–33[PubMed]
    [Google Scholar]
  8. Demangel C., Stinear T. P., Cole S. T. ( 2009). Buruli ulcer: reductive evolution enhances pathogenicity of Mycobacterium ulcerans . Nat Rev Microbiol 7:50–60 [View Article][PubMed]
    [Google Scholar]
  9. Devulder G., Pérouse de Montclos M., Flandrois J. P. ( 2005). A multigene approach to phylogenetic analysis using the genus Mycobacterium as a model. Int J Syst Evol Microbiol 55:293–302 [View Article][PubMed]
    [Google Scholar]
  10. Felsenstein J. ( 1989). phylip – phylogeny inference package (version 3.2). Cladistics 5:164–166
    [Google Scholar]
  11. Fernandez-Mora E., Polidori M., Lührmann A., Schaible U. E., Haas A. ( 2005). Maturation of Rhodococcus equi-containing vacuoles is arrested after completion of the early endosome stage. Traffic 6:635–653 [View Article][PubMed]
    [Google Scholar]
  12. Fleischmann R. D., Alland D., Eisen J. A., Carpenter L., White O., Peterson J., DeBoy R., Dodson R., Gwinn M. et al. ( 2002). Whole-genome comparison of Mycobacterium tuberculosis clinical and laboratory strains. J Bacteriol 184:5479–5490 [View Article][PubMed]
    [Google Scholar]
  13. Giguère S., Hondalus M. K., Yager J. A., Darrah P., Mosser D. M., Prescott J. F. ( 1999). Role of the 85-kilobase plasmid and plasmid-encoded virulence-associated protein A in intracellular survival and virulence of Rhodococcus equi . Infect Immun 67:3548–3557[PubMed]
    [Google Scholar]
  14. Hondalus M. K., Mosser D. M. ( 1994). Survival and replication of Rhodococcus equi in macrophages. Infect Immun 62:4167–4175[PubMed]
    [Google Scholar]
  15. Hondalus M. K., Diamond M. S., Rosenthal L. A., Springer T. A., Mosser D. M. ( 1993). The intracellular bacterium Rhodococcus equi requires Mac-1 to bind to mammalian cells. Infect Immun 61:2919–2929[PubMed]
    [Google Scholar]
  16. Hovel-Miner G., Pampou S., Faucher S. P., Clarke M., Morozova I., Morozov P., Russo J. J., Shuman H. A., Kalachikov S. ( 2009). σS controls multiple pathways associated with intracellular multiplication of Legionella pneumophila . J Bacteriol 191:2461–2473 [View Article][PubMed]
    [Google Scholar]
  17. Hughes K. L., Sulaiman I. ( 1987). The ecology of Rhodococcus equi and physicochemical influences on growth. Vet Microbiol 14:241–250 [View Article][PubMed]
    [Google Scholar]
  18. Iriarte M., Stainier I., Cornelis G. R. ( 1995). The rpoS gene from Yersinia enterocolitica and its influence on expression of virulence factors. Infect Immun 63:1840–1847[PubMed]
    [Google Scholar]
  19. Jain S., Bloom B. R., Hondalus M. K. ( 2003). Deletion of vapA encoding virulence associated protein A attenuates the intracellular actinomycete Rhodococcus equi . Mol Microbiol 50:115–128 [View Article][PubMed]
    [Google Scholar]
  20. Jones D. T., Taylor W. R., Thornton J. M. ( 1992). The rapid generation of mutation data matrices from protein sequences. Comput Appl Biosci 8:275–282[PubMed]
    [Google Scholar]
  21. Kang J. G., Hahn M. Y., Ishihama A., Roe J. H. ( 1997). Identification of sigma factors for growth phase-related promoter selectivity of RNA polymerases from Streptomyces coelicolor A3(2). Nucleic Acids Res 25:2566–2573 [View Article][PubMed]
    [Google Scholar]
  22. Kelly B. G., Wall D. M., Boland C. A., Meijer W. G. ( 2002). Isocitrate lyase of the facultative intracellular pathogen Rhodococcus equi . Microbiology 148:793–798[PubMed]
    [Google Scholar]
  23. Klinkert B., Narberhaus F. ( 2009). Microbial thermosensors. Cell Mol Life Sci 66:2661–2676 [View Article][PubMed]
    [Google Scholar]
  24. Larkin M. A., Blackshields G., Brown N. P., Chenna R., McGettigan P. A., McWilliam H., Valentin F., Wallace I. M., Wilm A. et al. ( 2007). clustal_w and clustal_x version 2.0. Bioinformatics 23:2947–2948 [View Article][PubMed]
    [Google Scholar]
  25. Letek M., Ocampo-Sosa A. A., Sanders M., Fogarty U., Buckley T., Leadon D. P., González P., Scortti M., Meijer W. G. et al. ( 2008). Evolution of the Rhodococcus equi vap pathogenicity island seen through comparison of host-associated vapA and vapB virulence plasmids. J Bacteriol 190:5797–5805 [View Article][PubMed]
    [Google Scholar]
  26. Letek M., González P., Macarthur I., Rodríguez H., Freeman T. C., Valero-Rello A., Blanco M., Buckley T., Cherevach I. et al. ( 2010). The genome of a pathogenic Rhodococcus: cooptive virulence underpinned by key gene acquisitions. PLoS Genet 6:e1001145 [View Article][PubMed]
    [Google Scholar]
  27. Li L., Bannantine J. P., Zhang Q., Amonsin A., May B. J., Alt D., Banerji N., Kanjilal S., Kapur V. ( 2005). The complete genome sequence of Mycobacterium avium subspecies paratuberculosis . Proc Natl Acad Sci U S A 102:12344–12349 [View Article][PubMed]
    [Google Scholar]
  28. Lührmann A., Mauder N., Sydor T., Fernandez-Mora E., Schulze-Luehrmann J., Takai S., Haas A. ( 2004). Necrotic death of Rhodococcus equi-infected macrophages is regulated by virulence-associated plasmids. Infect Immun 72:853–862 [View Article][PubMed]
    [Google Scholar]
  29. Meijer W. G., Prescott J. F. ( 2004). Rhodococcus equi . Vet Res 35:383–396 [View Article][PubMed]
    [Google Scholar]
  30. Miranda-CasoLuengo R., Duffy P. S., O’Connell E. P., Graham B. J., Mangan M. W., Prescott J. F., Meijer W. G. ( 2005). The iron-regulated iupABC operon is required for saprophytic growth of the intracellular pathogen Rhodococcus equi at low iron concentrations. J Bacteriol 187:3438–3444 [View Article][PubMed]
    [Google Scholar]
  31. Muscatello G., Leadon D. P., Klayt M., Ocampo-Sosa A., Lewis D. A., Fogarty U., Buckley T., Gilkerson J. R., Meijer W. G., Vazquez-Boland J. A. ( 2007). Rhodococcus equi infection in foals: the science of ‘rattles’. Equine Vet J 39:470–478 [View Article][PubMed]
    [Google Scholar]
  32. Nielsen A. T., Dolganov N. A., Otto G., Miller M. C., Wu C. Y., Schoolnik G. K. ( 2006). RpoS controls the Vibrio cholerae mucosal escape response. PLoS Pathog 2:e109 [View Article][PubMed]
    [Google Scholar]
  33. Ocampo-Sosa A. A., Lewis D. A., Navas J., Quigley F., Callejo R., Scortti M., Leadon D. P., Fogarty U., Vázquez-Boland J. A. ( 2007). Molecular epidemiology of Rhodococcus equi based on traA, vapA, and vapB virulence plasmid markers. J Infect Dis 196:763–769 [View Article][PubMed]
    [Google Scholar]
  34. Pfaffl M. W. ( 2004). Quantification strategies in real-time PCR. A–Z of Quantitative PCR87–112 Bustin S. A. La Jolla, CA: International University Line;
    [Google Scholar]
  35. Platt T. ( 1986). Transcription termination and the regulation of gene expression. Annu Rev Biochem 55:339–372 [View Article][PubMed]
    [Google Scholar]
  36. Polidori M., Haas A. ( 2006). VapI, a new member of the Rhodococcus equi Vap family. Antonie van Leeuwenhoek 90:299–304 [View Article][PubMed]
    [Google Scholar]
  37. Ren J., Prescott J. F. ( 2003). Analysis of virulence plasmid gene expression of intra-macrophage and in vitro grown Rhodococcus equi ATCC 33701. Vet Microbiol 94:167–182 [View Article][PubMed]
    [Google Scholar]
  38. Ren J., Prescott J. F. ( 2004). The effect of mutation on Rhodococcus equi virulence plasmid gene expression and mouse virulence. Vet Microbiol 103:219–230 [View Article][PubMed]
    [Google Scholar]
  39. Ripoll F., Pasek S., Schenowitz C., Dossat C., Barbe V., Rottman M., Macheras E., Heym B., Herrmann J. L. et al. ( 2009). Non mycobacterial virulence genes in the genome of the emerging pathogen Mycobacterium abscessus . PLoS ONE 4:e5660 [View Article][PubMed]
    [Google Scholar]
  40. Rohde K. H., Abramovitch R. B., Russell D. G. ( 2007). Mycobacterium tuberculosis invasion of macrophages: linking bacterial gene expression to environmental cues. Cell Host Microbe 2:352–364 [View Article][PubMed]
    [Google Scholar]
  41. Russell D. A., Byrne G. A., O’Connell E. P., Boland C. A., Meijer W. G. ( 2004). The LysR-type transcriptional regulator VirR is required for expression of the virulence gene vapA of Rhodococcus equi ATCC 33701. J Bacteriol 186:5576–5584 [View Article][PubMed]
    [Google Scholar]
  42. Sambrook J., Russell D. W. ( 2001). Molecular Cloning: a Laboratory Manual Cold Spring Harbor, NY: Cold Spring Harbor Laboratory;
    [Google Scholar]
  43. Sekine M., Tanikawa S., Omata S., Saito M., Fujisawa T., Tsukatani N., Tajima T., Sekigawa T., Kosugi H. et al. ( 2006). Sequence analysis of three plasmids harboured in Rhodococcus erythropolis strain PR4. Environ Microbiol 8:334–346 [View Article][PubMed]
    [Google Scholar]
  44. Stinear T. P., Seemann T., Pidot S., Frigui W., Reysset G., Garnier T., Meurice G., Simon D., Bouchier C. et al. ( 2007). Reductive evolution and niche adaptation inferred from the genome of Mycobacterium ulcerans, the causative agent of Buruli ulcer. Genome Res 17:192–200 [View Article][PubMed]
    [Google Scholar]
  45. Stinear T. P., Seemann T., Harrison P. F., Jenkin G. A., Davies J. K., Johnson P. D., Abdellah Z., Arrowsmith C., Chillingworth T. et al. ( 2008). Insights from the complete genome sequence of Mycobacterium marinum on the evolution of Mycobacterium tuberculosis . Genome Res 18:729–741 [View Article][PubMed]
    [Google Scholar]
  46. Takai S., Fukunaga N., Kamisawa K., Imai Y., Sasaki Y., Tsubaki S. ( 1996). Expression of virulence-associated antigens of Rhodococcus equi is regulated by temperature and pH. Microbiol Immunol 40:591–594[PubMed] [CrossRef]
    [Google Scholar]
  47. Takai S., Hines S. A., Sekizaki T., Nicholson V. M., Alperin D. A., Osaki M., Takamatsu D., Nakamura M., Suzuki K. et al. ( 2000). DNA sequence and comparison of virulence plasmids from Rhodococcus equi ATCC 33701 and 103. Infect Immun 68:6840–6847 [View Article][PubMed]
    [Google Scholar]
  48. Toyooka K., Takai S., Kirikae T. ( 2005). Rhodococcus equi can survive a phagolysosomal environment in macrophages by suppressing acidification of the phagolysosome. J Med Microbiol 54:1007–1015 [View Article][PubMed]
    [Google Scholar]
  49. von Bargen K., Wohlmann J., Taylor G. A., Utermöhlen O., Haas A. ( 2011). Nitric oxide-mediated intracellular growth restriction of pathogenic Rhodococcus equi can be prevented by iron. Infect Immun 79:2098–2111 [View Article][PubMed]
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/micro/10.1099/mic.0.049759-0
Loading
/content/journal/micro/10.1099/mic.0.049759-0
Loading

Data & Media loading...

Supplements

Supplementary material 1

PDF

Supplementary material 2

PDF
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