Skip to main content

Advertisement

SpringerLink
Log in

Cytolysin gene expression in Enterococcus faecalis is regulated in response to aerobiosis conditions

  • Original Paper
  • Published:
Molecular Genetics and Genomics Aims and scope Submit manuscript

Abstract

Here we investigate the expression of cylL L and cylL S , the genes that encode the structural subunits of the cytolysin/haemolysin of Enterococcus faecalis, in response to aerobiosis conditions. Haemolysis assays of E. faecalis strains cultured under aerobic and anaerobic conditions revealed three different haemolytic phenotypes, one of which exhibited greater haemolysis under anaerobic conditions than under aerobic conditions, and was shown to be associated with the presence of the cyl genes. Reporter gene studies revealed that cylL L L S promoter activity was significantly greater (up to 8.6-fold) under anaerobic compared to aerobic conditions throughout batch growth, demonstrating that these genes are regulated in response to the degree of aerobiosis. Band shift assays confirmed the binding of a protein factor to the region between 202 and 37 bp upstream of the cylL L start codon, and a higher level of binding was observed with anaerobically derived cell-free extracts than with extracts of aerobically grown cells. This is the first report of an oxygen-regulated virulence factor in E. faecalis (that is distinct from the quorum-sensing regulatory system reported previously), and may be of in vivo relevance for the bacterium in biofilms and other environments characterised by oxygen gradients.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1A–C.
Fig. 2A, B.
Fig. 3A–F
Fig. 4A–C.
Fig. 5A–F.

Similar content being viewed by others

References

  • Anderson DG, McKay LL (1983) Simple and rapid method for isolating large plasmid DNA from lactic streptococci. Appl Environ Microbiol 46:549–552

    CAS  PubMed  Google Scholar 

  • Basinger SF, Jackson RW (1968) Bacteriocine (hemolysin) of Streptococcus zymogenes. J Bacteriol 96:1895–1902

    PubMed  Google Scholar 

  • Booth MC, Hatter KL, Miller D, Davis J, Kowalski R, Parke DW, Chodosh J, Jett BD, Callegan MC, Penland R, Gilmore MS (1998) Molecular epidemiology of Staphylococcus aureus and Enterococcus faecalis in endophthalmitis. Infect Immun 66:356–360

    CAS  PubMed  Google Scholar 

  • Coburn PS, Hancock LE, Booth MC, Gilmore MS (1999) A novel means of self-protection, unrelated to toxin activation, confers immunity to the bactericidal effects of the Enterococcus faecalis cytolysin. Infect Immun 67:3339–3347

    CAS  PubMed  Google Scholar 

  • Costerton JW, Lewandowski Z, Caldwell DE, Korber DR, Lappin-Scott HM (1995) Microbial biofilms. Annu Rev Microbiol 49:711–745

    CAS  PubMed  Google Scholar 

  • Cruz-Rodz AL, Gilmore MS (1990) High efficiency introduction of plasmid DNA into glycine treated Enterococcus faecalis by electroporation. Mol Gen Genet 224:152–154

    CAS  PubMed  Google Scholar 

  • Day AM, Sandoe JAT, Cove JH, Phillips-Jones MK (2001) Evaluation of a biochemical test scheme for identifying clinical isolates of Enterococcus faecalis and Enterococcus faecium. Lett Appl Microbiol 33:392–396

    Article  CAS  PubMed  Google Scholar 

  • Dunny GM, Lee LN, LeBlanc DJ (1991) Improved electroporation and cloning vector system for Gram-positive bacteria. Appl Environ Microbiol 57:1194–1201

    CAS  PubMed  Google Scholar 

  • Endophthalmitis Vitrectomy Study Group (1996) Microbiologic factors and visual outcome in the endophthalmitis vitrectomy study. Am J Ophthalmol 122:830–846

    PubMed  Google Scholar 

  • Facklam RR, Collins MD (1989) Identification of Enterococcus species isolated from human infections by a conventional test scheme. J Clin Microbiol 27:731–734

    CAS  PubMed  Google Scholar 

  • Facklam RR, Sahm DF (1995) Enterococcus. In: Murray PR, Baron EJ, Pfaller MA, Tenover FC, Yolken RH (eds) Manual of clinical microbiology (6th edn). American Society for Microbiology, Washington DC, pp 308–314

  • Facklam RR, Teixeira LM (1998) Enterococcus. In: Collier L, Balows A, Sussman M (eds) Topley and Wilson's Microbiology and microbial infections. Arnold, London, pp 669–682

  • Gilmore MS, Segarra RA, Booth MC, Bogie C, Hall LR, Clewell DB (1994) Genetic structure of the Enterococcus faecalis cytolytic toxin system and its relationship to lantibiotic determinants. J Bacteriol 176:7335–7344

    CAS  PubMed  Google Scholar 

  • Haas W, Gilmore MS (1999) Molecular nature of a novel bacterial toxin: the cytolysin of Enterococcus faecalis. Med Microbiol Immunol 187:183–190

    Article  CAS  PubMed  Google Scholar 

  • Haas W, Shepard BD, Gilmore MS (2002) Two-component regulator of Enterococcus faecalis cytolysin responds to quorum-sensing autoinduction. Nature 415:84–87

    Article  CAS  PubMed  Google Scholar 

  • Heim R, Prasher DC, Tsien RY (1994) Wavelength mutations and posttranslational autoxidation of green fluorescent protein. Proc Natl Acad Sci USA 91:12501–12504

    CAS  PubMed  Google Scholar 

  • Huycke MM, Spiegel CA, Gilmore MS (1991) Bacteremia caused by hemolytic, high-level gentamicin-resistant Enterococcus faecalis. Antimicrob Agents Chemother 35:1626–1634

    CAS  PubMed  Google Scholar 

  • Ike Y, Hashimoto H, Clewell DB (1984) Hemolysis of Streptococcus faecalis subspecies zymogenes contributes to virulence in mice. Infect Immun 45:528–530

    CAS  PubMed  Google Scholar 

  • Ike Y, Hashimoto H, Clewell DB (1987) High incidence of hemolysin production by Enterococcus (Streptococcus) faecalis strains associated with human parental infections. J Clin Microbiol 25:1524–1528

    CAS  PubMed  Google Scholar 

  • Ike Y, Clewell DB, Segarra RA, Gilmore MS (1990) Genetic analysis of the pAD1 hemolysin/bacteriocine determinant in Enterococcus faecalis: Tn917 insertional mutagenesis and cloning. J Bacteriol 172:155–163

    CAS  PubMed  Google Scholar 

  • Joyanes PA, Pascual A, Martínez-Martínez L, Hevia A, Perea EJ (2000) In vitro adherence of Enterococcus faecalis and Enterococcus faecium to urinary catheters. Eur J Clin Microbiol Infect Dis 19:124–127

    Article  CAS  PubMed  Google Scholar 

  • Keane PF, Bonner MC, Johnston SR, Zafar A, Gorman SP (1994) Characterization of biofilm and encrustation on ureteric stents in vivo. Br J Urol 73:687–691

    Google Scholar 

  • Leclerq R (1997) Enterococci acquire new kinds of resistance. Clin Infect Dis 24 (Suppl 1):S80–S84

    PubMed  Google Scholar 

  • MacInnes JI, Kim JE, Lian CJ-, Soltes GA (1990) Actinobacillus pleuropneumoniae hlyX gene homology with the fnr gene of Escherichia coli. J Bacteriol 172:4587–4592

    CAS  PubMed  Google Scholar 

  • Macrina FL, Ash Tobian J, Jones KR, Evans RP, Clewell DB (1982) A cloning vector able to replicate in Escherichia coli and Streptococcus sanguis. Gene 19:345–353

    Article  CAS  PubMed  Google Scholar 

  • McGlynn P, Hunter CN (1992) Isolation and characterisation of a putative transcription factor involved in the regulation of the Rhodobacter sphaeroides pucBA operon. J Biol Chem 267:11098–11103

    CAS  PubMed  Google Scholar 

  • Megran DW (1992) Enterococcal endocarditis. Clin Infect Dis 15:63–71

    CAS  PubMed  Google Scholar 

  • Miyazaki S, Ohno A, Kobayashi I, Uji T, Yamaguchi K, Goto S (1993) Cytotoxic effect of hemolytic culture supernatant from Enterococcus faecalis on mouse polymorphonuclear neutrophils and macrophages. Microbiol Immunol 37:265–270

    Article  CAS  PubMed  Google Scholar 

  • Moellering RC Jr (1992) Emergence of Enterococcus as a significant pathogen. Clin Infect Dis 14:1173–1178

    PubMed  Google Scholar 

  • Phillips-Jones MK (2000) Use of a lux reporter system for monitoring rapid changes in α-toxin gene expression in Clostridium perfringens during growth. FEMS Microbiol Lett 188:29–33

    Article  CAS  PubMed  Google Scholar 

  • Ralph ET, Guest JR, Green J (1998) Altering the anaerobic transcription factor FNR confers a haemolytic phenotype on Escherichia coli K12. Proc Natl Acad Sci USA 95:10449–10452

    Article  CAS  PubMed  Google Scholar 

  • Ritchey TW, Seeley HW Jr (1974) Cytochromes in Streptococcus faecalis var. zymogenes grown in a haematin-containing medium. J Gen Microbiol 85:220–228

    CAS  PubMed  Google Scholar 

  • Ross RP, Claiborne A (1997) Evidence for regulation of the NADH peroxidase gene (npr) from Enterococcus faecalis by OxyR. FEMS Microbiol Lett 151:177–183

    CAS  PubMed  Google Scholar 

  • Sambrook J, Fritsch EF, Maniatis T (1989) Molecular cloning: a laboratory manual (2nd edn). Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y.

    Google Scholar 

  • Scott KP, Mercer DK, Glover LA, Flint HJ (1998) The green fluorescent protein as a visible marker for lactic acid bacteria in complex ecosystems. FEMS Microbiol Ecol 26:219–230

    Article  CAS  Google Scholar 

  • Shephard BD, Gilmore MS (1999) Identification of aerobically and anaerobically induced genes in Enterococcus faecalis by random arbitrarily primed PCR. Appl Environ Microbiol 65:1470–1476

    PubMed  Google Scholar 

  • Spiro S, Guest JR (1990) FNR and its role in oxygen-related gene expression in Escherichia coli. FEMS Microbiol Rev 75:399–428

    CAS  Google Scholar 

  • Terzaghi BE, Sandine WE (1975) Improved medium for lactic streptococci and their bacteriophages. Appl Microbiol 29:807–813

    CAS  Google Scholar 

  • Todd EW (1934) A comparative serological study of streptolysins derived from human and from animal infections, with notes on pneumococcal haemolysin tetanolysin and staphylococcus toxin. J Pathol Bacteriol 39:299–321

    CAS  Google Scholar 

  • Whitehead TR, Flint HJ (1995) Heterologous expression of an endoglucanase gene (endA) from the ruminal anaerobe Ruminococcus flavefaciens 17 in Streptococcus bovis and Streptococcus sanguis. FEMS Microbiol Lett 126:165–170

    Article  CAS  PubMed  Google Scholar 

  • Winstedt LL, Frankenberg L, Hederstedt L, von Wachenfeldt C (2000) Enterococcus faecalis V583 contains a cytochrome bd-type respiratory oxidase. J Bacteriol 182:3863–3866

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgements

We thank Dr. P. Duggan for provision of plasmid pVACMC1 and Dr. J. A. T. Sandoe for provision of previously described E. faecalis clinical isolates. AMD was supported by an Emma and Leslie Reid Scholarship from the University of Leeds

Author information

Authors and Affiliations

  1. Division of Microbiology, School of Biochemistry and Molecular Biology, University of Leeds, Leeds LS2 9JT, UK

    A. M. Day, J. H. Cove & M. K. Phillips-Jones

Authors
  1. A. M. Day
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. J. H. Cove
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. M. K. Phillips-Jones
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to M. K. Phillips-Jones.

Additional information

Communicated by W. Goebel

Rights and permissions

Reprints and permissions

About this article

Cite this article

Day, A.M., Cove, J.H. & Phillips-Jones, M.K. Cytolysin gene expression in Enterococcus faecalis is regulated in response to aerobiosis conditions. Mol Gen Genomics 269, 31–39 (2003). https://doi.org/10.1007/s00438-003-0819-1

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00438-003-0819-1

Keywords

Search

Navigation