1887

Abstract

Species identification of in routine laboratory testing is hampered by the lack of highly discriminatory phenotypic tests. Amplification polymorphism of the intergenic 16S-23S spacer regions (ISR) has been previously developed for identification of species within the [Hookey. J. V., Birtles, R. J. & Saunders, N. A. (1995). 33, 2377-2381], but it did not provide enough resolution to distinguish all members of the bluish-white autofluorescent species and the red autofluorescent group of the By choosing new primers that target regions 4 (positions 1521-1541 of 16S rRNA gene) and 6 (positions 114-132 of 23S rRNA gene) within the rDNA operon close to the 16S-23S intergenic spacer, 34 profiles were determined among the 79 type and reference strains representing 42 species that were tested. Analysis of the RFLP generated after I restriction digestion of the PCR products further improved the method, allowing complete discrimination among the species and subspecies of tested. Twenty-three well-identified strains from unrelated origins belonging to seven species gave amplification patterns identical to that of their type strain. The technique was also tested on 80 field isolates that could not be unequivocally assigned to groups by phenotypic methods. Seventy-two per cent (58/80) of these isolates had a profile identical to that of a type strain, while 27% (22/80) may correspond to new taxa since their ISR-PCR profiles did not match any of the known profiles.

Loading

Article metrics loading...

/content/journal/ijsem/10.1099/00207713-48-3-723
1998-07-01
2022-12-02
Loading full text...

Full text loading...

/deliver/fulltext/ijsem/48/3/ijs-48-3-723.html?itemId=/content/journal/ijsem/10.1099/00207713-48-3-723&mimeType=html&fmt=ahah

References

  1. Altschul S. F., Gish W., Miller W., Myers E. W., Lipman A. J. 1990; Basic local alignment search tool. J Mol Biol 215:404–410
    [Google Scholar]
  2. Benson R. F., Thacker W. L., Daneshvar M. I., Brenner D. J. 1996; Legionella waitersii sp. nov. and an unnamed Legionella genomospecies isolated from water in Australia. Int J Syst Bacteriol 46:631–634
    [Google Scholar]
  3. Brenner D. J. 1986; Classification of Legionellaceae. Current status and remaining questions. Isr J Med Sci 22:620–632
    [Google Scholar]
  4. Edelstein P. H. 1981; Improved semiselective medium for isolation of Legionella pneumophila from contaminated clinical and environmental specimens. J Clin Microbiol 14:294–303
    [Google Scholar]
  5. Grimont F., Lefé M., Ageron E., Grimont P. A. D. 1989; rRNA gene restriction patterns of Legionella species: a molecular identification system. Res Microbiol 140:615–626
    [Google Scholar]
  6. Urtler V., Barrie H. D. 1995; Typing of Staphylococcus aureus strains by PCR-amplification of variable-length 16S-23S rDNA spacer regions: characterization of spacer sequences. Microbiology 141:1255–1265
    [Google Scholar]
  7. Urtler V., Stanisich V. A. 1996; New approaches to typing and identification of bacteria using the 16S-23S rDNA spacer region. Microbiology 142:3–16
    [Google Scholar]
  8. Harrell L. J., Andersen G. L., Wilson K. H. 1995; Genetic variability of Bacillus anthracis and related species. J Clin Microbiol 33:1847–1850
    [Google Scholar]
  9. Hookey J. V., Birtles R. J., Saunders N. A. 1995; Inter- genic 16S rRNA gene (rDNA)-23S rDNA sequence length polymorphisms in members of the famil Legionellaceae.. J Clin Microbiol 33:2377–2381
    [Google Scholar]
  10. Hookey J. V., Saunders N. A., Fry N. K., Birtles R. J., Harrison T. G. 1996; Phylogeny of Legionellaceae based on small-subunit ribosomal DNA sequences and proposal of Legionella lytica comb. nov. for Legionella-like amoebal pathogens. Int J Syst Bacteriol 46:526–531
    [Google Scholar]
  11. Jaulhac B., Nowicki M., Bornstein N., Meunier O., Prevost G., Piemont Y., Fleurette J., Monteil H. 1992; Detection of Legionella spp in bronchoalveolar lavage fluids by DNA amplification. J Clin Microbiol 30:920–924
    [Google Scholar]
  12. Jensen M. A., Webster J. A., Straus N. 1993; Rapid identification of bacteria on the basis of polymerase chain reaction-amplified ribosomal DNA spacer polymorphisms. Appl Environ Microbiol 59:945–952
    [Google Scholar]
  13. Lo Presti F., Riffard S., Vandenesch F., Reyrolle M., Ronco F., Ichai P., Etienne J. 1997; The first clinical isolate of Legionella parisiensis, from a liver transplant patient with pneumonia. J Clin Microbiol 35:1706–1709
    [Google Scholar]
  14. Marmur J. 1961; A procedure for the isolation of deoxyribonucleic acid from microorganisms. J Mol Biol 3:208–218
    [Google Scholar]
  15. Meunier J. R., Grimont P. A. D. 1993; Factors affecting reproducibility of random amplified polymorphic DNA fingerprinting. Res Microbiol 144,373–379
    [Google Scholar]
  16. Navarro E., Simonet P., Normand P., Bardin R. 1992; Characterization of natural populations of Nitrobacter spp. using PCR/RFLP analysis of the ribosomal intergenic spacer. Arch Microbiol 157:107–115
    [Google Scholar]
  17. Normand P., Cournoyer B., Nazaret S., Simonet P. 1992; Analysis of a ribosomal RNA operon in the actinomycete Frankia . Gene 111:119–124
    [Google Scholar]
  18. Normand P., Ponsonnet C., Nesme X., Neyra M., Simonet P. 1996; ITS analysis of prokaryotes. In Molecular Microbial Ecology Manual pp 1–12 Edited by Akkermans A. D. L., van Elsas J. D., De Bruijn F. J. Dordrecht, The Netherlands: Kluwer;
    [Google Scholar]
  19. Rodgers F. G., Pasculle A. W. 1991 Legionella. In Manual of Clinical Microbiology, 5th edn. pp 442–453 Edited by Balows A., Hausler W. J. Jr, Herrmann K. L., Isenberg H. D., Shadomy H. J. Washington, DC: American Society for Microbiology;
    [Google Scholar]
  20. Verissimo A., Morais P. V., Diogo A., Gomes C., Costa M. S. D. 1996; Characterization of Legionella species by numerical analysis of whole-cell protein electrophoresis. Int J Syst Bacteriol 46:41–49
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/ijsem/10.1099/00207713-48-3-723
Loading
/content/journal/ijsem/10.1099/00207713-48-3-723
Loading

Data & Media loading...

Most cited this month Most Cited RSS feed

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