1887

Abstract

Completion of the genome sequence of SK36 necessitates tools for further characterization of this species. It is often desirable to insert antibiotic resistance markers and other exogenous genes into the chromosome; therefore, we sought to identify a chromosomal site for ectopic expression of foreign genes, and to verify that insertion into this site did not affect important cellular phenotypes. We designed three plasmid constructs for insertion of , or resistance determinants into a genomic region encoding only a small (65 aa) hypothetical protein. To determine whether this insertion affected important cellular properties, SK36 and its erythromycin-resistant derivative, JFP36, were compared for: (i) growth , (ii) genetic competence, (iii) biofilm formation and (iv) virulence for endocarditis in the rabbit model of infective endocarditis (IE). The spectinomycin-resistant strain, JFP56, and tetracycline-resistant strain, JFP76, were also tested for virulence . Insertion of did not affect growth, competence or biofilm development of JFP36. Recovery of bacteria from heart valves of co-inoculated rabbits was similar to wild-type for JFP36, JFP56 and JFP76, indicating that IE virulence was not significantly affected. The capacity for mutant complementation was explored in an avirulent mutant background. Expression of from its predicted promoter in the target region restored IE virulence. Thus, the chromosomal site utilized is a good candidate for further manipulations of . In addition, the resistant strains developed may be further applied as controls to facilitate screening for virulence factors .

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2009-08-01
2019-12-14
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References

  1. Aas, J. A., Paster, B. J., Stokes, L. N., Olsen, I. & Dewhirst, F. E. ( 2005; ). Defining the normal bacterial flora of the oral cavity. J Clin Microbiol 43, 5721–5732.[CrossRef]
    [Google Scholar]
  2. Black, C., Allan, I., Ford, S. K., Wilson, M. & McNab, R. ( 2004; ). Biofilm-specific surface properties and protein expression in oral Streptococcus sanguis. Arch Oral Biol 49, 295–304.[CrossRef]
    [Google Scholar]
  3. Bochud, P. Y., Calandra, T. & Francioli, P. ( 1994; ). Bacteremia due to viridans streptococci in neutropenic patients: a review. Am J Med 97, 256–264.[CrossRef]
    [Google Scholar]
  4. Caufield, P. W., Dasanayake, A. P., Li, Y., Pan, Y., Hsu, J. & Hardin, J. M. ( 2000; ). Natural history of Streptococcus sanguinis in the oral cavity of infants: evidence for a discrete window of infectivity. Infect Immun 68, 4018–4023.[CrossRef]
    [Google Scholar]
  5. Claverys, J. P., Dintilhac, A., Pestova, E. V., Martin, B. & Morrison, D. A. ( 1995; ). Construction and evaluation of new drug-resistance cassettes for gene disruption mutagenesis in Streptococcus pneumoniae, using an ami test platform. Gene 164, 123–128.[CrossRef]
    [Google Scholar]
  6. Cremieux, A. C., Maziere, B., Vallois, J. M., Ottaviani, M., Azancot, A., Raffoul, H., Bouvet, A., Pocidalo, J. J. & Carbon, C. ( 1989; ). Evaluation of antibiotic diffusion into cardiac vegetations by quantitative autoradiography. J Infect Dis 159, 938–944.[CrossRef]
    [Google Scholar]
  7. Das, S., Kanamoto, T., Ge, X., Xu, P., Unoki, T., Munro, C. L. & Kitten, T. ( 2009; ). Contribution of lipoproteins and lipoprotein processing to endocarditis virulence in Streptococcus sanguinis. J Bacteriol 191, 4166–4179.[CrossRef]
    [Google Scholar]
  8. Delcher, A. L., Harmon, D., Kasif, S., White, O. & Salzberg, S. L. ( 1999; ). Improved microbial gene identification with GLIMMER. Nucleic Acids Res 27, 4636–4641.[CrossRef]
    [Google Scholar]
  9. Dintilhac, A., Alloing, G., Granadel, C. & Claverys, J. P. ( 1997; ). Competence and virulence of Streptococcus pneumoniae: Adc and PsaA mutants exhibit a requirement for Zn and Mn resulting from inactivation of putative ABC metal permeases. Mol Microbiol 25, 727–739.[CrossRef]
    [Google Scholar]
  10. Douglas, C. W., Heath, J., Hampton, K. K. & Preston, F. E. ( 1993; ). Identity of viridans streptococci isolated from cases of infective endocarditis. J Med Microbiol 39, 179–182.[CrossRef]
    [Google Scholar]
  11. Durack, D. T. ( 1975; ). Experimental bacterial endocarditis. IV. Structure and evolution of very early lesions. J Pathol 115, 81–89.[CrossRef]
    [Google Scholar]
  12. Durack, D. T., Beeson, P. B. & Petersdorf, R. G. ( 1973; ). Experimental bacterial endocarditis. 3. Production and progress of the disease in rabbits. Br J Exp Pathol 54, 142–151.
    [Google Scholar]
  13. Dyson, C., Barnes, R. A. & Harrison, G. A. J. ( 1999; ). Infective endocarditis: an epidemiological review of 128 episodes. J Infect 38, 87–93.[CrossRef]
    [Google Scholar]
  14. Froeliger, E. H. & Fives-Taylor, P. ( 2001; ). Streptococcus parasanguis fimbria-associated adhesin Fap1 is required for biofilm formation. Infect Immun 69, 2512–2519.[CrossRef]
    [Google Scholar]
  15. Gaustad, P. & Havardstein, L. S. ( 1997; ). Competence-pheromone in Streptococcus sanguis. Identification of the competence gene comC and the competence pheromone. Adv Exp Med Biol 418, 1019–1021.
    [Google Scholar]
  16. Ge, X., Kitten, T., Chen, Z., Lee, S. P., Munro, C. L. & Xu, P. ( 2008; ). Identification of Streptococcus sanguinis genes required for biofilm formation and examination of their role in endocarditis virulence. Infect Immun 76, 2551–2559.[CrossRef]
    [Google Scholar]
  17. Hancock, L. E. & Perego, M. ( 2004; ). Systematic inactivation and phenotypic characterization of two-component signal transduction systems of Enterococcus faecalis V583. J Bacteriol 186, 7951–7958.[CrossRef]
    [Google Scholar]
  18. Horton, R. M. ( 1995; ). PCR-mediated recombination and mutagenesis. SOEing together tailor-made genes. Mol Biotechnol 3, 93–99.[CrossRef]
    [Google Scholar]
  19. Kilian, M., Mikkelsen, L. & Henrichsen, J. ( 1989; ). Taxonomic study of viridans streptococci: description of Streptococcus gordonii sp. nov. and emended descriptions of Streptococcus sanguis (White and Niven 1946), Streptococcus oralis (Bridge and Sneath 1982), and Streptococcus mitis (Andrewes and Horder 1906). Int J Syst Bacteriol 39, 471–484.[CrossRef]
    [Google Scholar]
  20. Kitten, T., Munro, C. L., Michalek, S. M. & Macrina, F. L. ( 2000; ). Genetic characterization of a Streptococcus mutans LraI family operon and role in virulence. Infect Immun 68, 4441–4451.[CrossRef]
    [Google Scholar]
  21. Libus, J. & Storchova, H. ( 2006; ). Quantification of cDNA generated by reverse transcription of total RNA provides a simple alternative tool for quantitative RT-PCR normalization. Biotechniques 41, 156–164.[CrossRef]
    [Google Scholar]
  22. Loo, C. Y., Corliss, D. A. & Ganeshkumar, N. ( 2000; ). Streptococcus gordonii biofilm formation: identification of genes that code for biofilm phenotypes. J Bacteriol 182, 1374–1382.[CrossRef]
    [Google Scholar]
  23. Marsh, P. D. ( 2003; ). Are dental diseases examples of ecological catastrophes? Microbiology 149, 279–294.[CrossRef]
    [Google Scholar]
  24. Martin, B., Prudhomme, M., Alloing, G., Granadel, C. & Claverys, J. P. ( 2000; ). Cross-regulation of competence pheromone production and export in the early control of transformation in Streptococcus pneumoniae. Mol Microbiol 38, 867–878.[CrossRef]
    [Google Scholar]
  25. Moreillon, P. & Que, Y. A. ( 2004; ). Infective endocarditis. Lancet 363, 139–149.[CrossRef]
    [Google Scholar]
  26. Oggioni, M. R., Dowson, C. G., Smith, J. M., Provvedi, R. & Pozzi, G. ( 1996; ). The tetracycline resistance gene tet(M) exhibits mosaic structure. Plasmid 35, 156 [CrossRef]
    [Google Scholar]
  27. O'Toole, G. A. & Kolter, R. ( 1998; ). Initiation of biofilm formation in Pseudomonas fluorescens WCS365 proceeds via multiple, convergent signaling pathways: a genetic analysis. Mol Microbiol 28, 449–461.[CrossRef]
    [Google Scholar]
  28. Paik, S., Senty, L., Das, S., Noe, J. C., Munro, C. L. & Kitten, T. ( 2005; ). Identification of virulence determinants for endocarditis in Streptococcus sanguinis by signature-tagged mutagenesis. Infect Immun 73, 6064–6074.[CrossRef]
    [Google Scholar]
  29. Plummer, C., Wu, H., Kerrigan, S. W., Meade, G., Cox, D. & Douglas, C. W. I. ( 2005; ). A serine-rich glycoprotein of Streptococcus sanguis mediates adhesion to platelets via GPIb. Br J Haematol 129, 101–109.[CrossRef]
    [Google Scholar]
  30. Podbielski, A., Spellerberg, B., Woischnik, M., Pohl, B. & Lütticken, R. ( 1996; ). Novel series of plasmid vectors for gene inactivation and expression analysis in group A streptococci (GAS). Gene 177, 137–147.[CrossRef]
    [Google Scholar]
  31. Ramirez-Ronda, C. H. ( 1978; ). Adherence of glucan-positive and glucan-negative streptococcal strains to normal and damaged heart valves. J Clin Invest 62, 805–814.[CrossRef]
    [Google Scholar]
  32. Roberts, R. B., Krieger, A. G., Schiller, N. L. & Gross, K. C. ( 1979; ). Viridans streptococcal endocarditis: the role of various species, including pyridoxal-dependent streptococci. Rev Infect Dis 1, 955–966.[CrossRef]
    [Google Scholar]
  33. Senghas, E., Jones, J. M., Yamamoto, M., Gawron-Burke, C. & Clewell, D. B. ( 1988; ). Genetic organization of the bacterial conjugative transposon Tn916. J Bacteriol 170, 245–249.
    [Google Scholar]
  34. Tleyjeh, I. M., Abdel-Latif, A., Rahbi, H., Scott, C. G., Bailey, K. R., Steckelberg, J. M., Wilson, W. R. & Baddour, L. M. ( 2007; ). A systematic review of population-based studies of infective endocarditis. Chest 132, 1025–1035.[CrossRef]
    [Google Scholar]
  35. Vlessis, A. A., Hovaguimian, H., Jaggers, J., Ahmad, A. & Starr, A. ( 1996; ). Infective endocarditis: ten-year review of medical and surgical therapy. Ann Thorac Surg 61, 1217–1222.[CrossRef]
    [Google Scholar]
  36. Wilson, W., Taubert, K. A., Gewitz, M., Lockhart, P. B., Baddour, L. M., Levison, M., Bolger, A., Cabell, C. H., Takahashi, M. & other authors ( 2008; ). Prevention of infective endocarditis: guidelines from the American Heart Association: a guideline from the American Heart Association Rheumatic Fever, Endocarditis and Kawasaki Disease Committee, Council on Cardiovascular Disease in the Young, and the Council on Clinical Cardiology, Council on Cardiovascular Surgery and Anesthesia, and the Quality of Care and Outcomes Research Interdisciplinary Working Group. J Am Dent Assoc 139, 3S–24S.[CrossRef]
    [Google Scholar]
  37. Xu, P., Alves, J. M., Kitten, T., Brown, A., Chen, Z., Ozaki, L. S., Manque, P., Ge, X., Serrano, M. G. & other authors ( 2007; ). Genome of the opportunistic pathogen Streptococcus sanguinis. J Bacteriol 189, 3166–3175.[CrossRef]
    [Google Scholar]
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