1887

Abstract

Four primer systems, amplifying fragments of the gene coding for the small ribosomal subunit (18S rRNA) were characterised with pure cultures of 65 medically relevant fungal species plus two mushrooms. A primer cocktail (TR1/CA1-TR2/AF2) amplified 59 of 67 fungal species; the universal fungal primer 1 (UF1) in combination with the eukaryotic primers S3 or EU1 amplified 64 and 65 of 67 fungal species, respectively. The design of an additional primer (RZY1) enabled the amplification of the missing members of the zygomycetes. The primer systems amplified all the medically relevant fungi tested. These included eight spp. and seven other yeast species, 13 dermatophytes, 32 moulds (including six zygomycetes and five dimorphic fungi) and two mushrooms. Eleven controls including DNA from and man were not amplified. The oligonucleotide CA hybridised with and ; the oligonucleotide TR hybridised with the 13 dermatophytes; the oligonucleotide AF hybridised with , but not with or ; and the oligonucleotide HC hybridised with three varieties of . These oligonucleotides did not hybridise with the other fungi nor the controls. The specificity of the newly designed primer systems was confirmed by selective amplification of fungal DNA from human lung tissue spiked with fungal biomass and from vitrectomy fluid of a patient with candida endophthalmitis.

Loading

Article metrics loading...

/content/journal/jmm/10.1099/00222615-47-9-811
1998-09-01
2022-01-16
Loading full text...

Full text loading...

/deliver/fulltext/jmm/47/9/medmicro-47-9-811.html?itemId=/content/journal/jmm/10.1099/00222615-47-9-811&mimeType=html&fmt=ahah

References

  1. White T. J., Bruns T., Lee S., Taylor J. Amplification and direct sequencing of fungal ribosomal RNA genes for phylogenetics. In Innis M. A., Gelfand D. H., Sninsky J. J., White T. J. (eds) PCR protocols: a guide to methods and applications San Diego, CA: Academic Press; 1990315–322
    [Google Scholar]
  2. Polanco A. M., Rodríguez-Tudela J. L., Martínez-Suárez J. V. Detection of pathogenic fungi in human blood by the polymerase chain reaction. Eur J Clin Microbiol Infect Dis 1995; 14:618–621
    [Google Scholar]
  3. Bock M., Maiwald M., Kappe R., Nickel P., Näher H. Polymerase chain reaction-based detection of dermatophyte DNA with a fungus-specific primer system. Mycoses 1994; 37:79–84
    [Google Scholar]
  4. Makimura K., Murayama S. Y., Yamaguchi H. Detection of a wide range of medically important fungi by the polymerase chain reaction. J Med Microbiol 1994; 40:358–364
    [Google Scholar]
  5. Makimura K., Murayama S. Y., Yamaguchi H. Specific detection of Aspergillus and Penicillium species from respiratory specimens by polymerase chain reaction (PCR). Jpn J Med Sci Biol 1994; 47:141–156
    [Google Scholar]
  6. Kappe R., Fauser C., Okeke C. N., Maiwald M. Universal fungus specific primer systems and group-specific hybridization oligonucleotides for 18SrDNA. Mycoses 1996; 39:25–30
    [Google Scholar]
  7. Einsele H., Hebart H., Roller G. Detection and identification of fungal pathogens in blood by using molecular probes. J Clin Microbiol 1997; 35:1353–1360
    [Google Scholar]
  8. Mitchell T. G., Sandin R. L., Bowman B. H., Meyer W., Merz W. G. Molecular mycology: DNA probes and applications of PCR technology. J Med Vet Mycol 1994; 32: Suppl 1351–366
    [Google Scholar]
  9. Neefs J.-M., Van de Peer Y., De Rijk P., Chapelle S., De Wachter R. Compilation of small ribosomal subunit RNA structures. Nucleic Acids Res 1993; 21:3025–3049
    [Google Scholar]
  10. Van de Peer Y., Van den Broeck I., De Rijk P., De Wachter R. Database on the structure of small ribosomal subunit RNA. Nucleic Acids Res 1994; 22:3488–3494
    [Google Scholar]
  11. Van de Peer Nicolai S., De Rijk P., De Wachter R. Database on the structure of small ribosomal subunit RNA. Nucleic Acids Res 1996; 24:86–91
    [Google Scholar]
  12. Verweij P. E., Meis J. F. G. M., van den Hurk P., Zoll J., Samson R. A., Melchers W. J. G. Phylogenetic relationships of five species of Aspergillus and related taxa as deduced by comparison of sequences of small subunit ribosomal RNA. J Med Vet Mycol 1995; 33:185–190
    [Google Scholar]
  13. Sandhu G. S., Kline B. C., Stockman L., Roberts G. D. Molecular probes for diagnosis of fungal infections. J Clin Microbiol 1995; 33:2913–2919
    [Google Scholar]
  14. Krüger M., Osterburg G. On the alignment of two or more molecular sequences. Computer Programs in Biomedicine 1983; 16:68–69
    [Google Scholar]
  15. Maiwald M., Kappe R., Sonntag H.-G. Rapid presumptive identification of medically relevant yeasts to the species level by polymerase chain reaction and restriction enzyme analysis. J Med Vet Mycol 1994; 32:115–122
    [Google Scholar]
  16. Bowman B. H. A model PCR/probe system for the identification of fungal pathogens. In Persing D. H., Smith T. F., Tenover F. C., White T. J. (eds) Diagnostic molecular microbiology, principles and applications Washington, DC: American Society for Microbiology; 1993423–430
    [Google Scholar]
  17. Karlsen F., Kalantari M., Chitemerere M., Johansson B., Hagmar B. Modifications of human and viral deoxyribonucleic acid by formaldehyde fixation. Lab Invest 1994; 71:604–611
    [Google Scholar]
  18. Olsson M., Elvin K., Löfdahl S., Linder E. Detection of Pneumocystis carinii DNA in sputum and bronchoalveolar lavage samples by polymerase chain reaction. J Clin Microbiol 1993; 31:221–226
    [Google Scholar]
  19. Spreadbury C., Holden D., Aufauvre-Brown A., Bainbridge B., Cohen J. Detection of Aspergillus fumigatus by polymerase chain reaction. J Clin Microbiol 1993; 31:615–621
    [Google Scholar]
  20. Montone K., Litzky L. A. Rapid method for detection of Aspergillus 5S ribosomal RNA using a genus-specific oligonucleotide probe. Am J Clin Pathol 1995; 103:48–51
    [Google Scholar]
  21. Bretagne S., Costa J.-M., Marmorat-Khuong A. Detection of Aspergillus species DNA in bronchoalveolar lavage samples by competitive PCR. J Clin Microbiol 1995; 33:1164–1168
    [Google Scholar]
  22. Miyakawa Y., Mabuchi T., Kagaya K., Fukazawa Y. Isolation and characterization of a species-specific DNA fragment for detection of Candida albicans by polymerase chain reaction. J Clin Microbiol 1992; 30:894–900
    [Google Scholar]
  23. Chryssanthou E., Andersson B., Petrini B., Löfdahl S., Tollemar J. Detection of Candida albicans DNA in serum by polymerase chain reaction. Scand J Infect Dis 1994; 26:479–485
    [Google Scholar]
  24. Holmes A. R., Cannon R. D., Shepherd M. G., Jenkinson H. F. Detection of Candida albicans and other yeasts in blood by PCR. J Clin Microbiol 1994; 32:228–231
    [Google Scholar]
  25. Rand K. H., Houck H., Wolff M. Detection of candidemia by polymerase chain reaction. Mol Cell Probes 1994; 82:215–221
    [Google Scholar]
  26. Monteagudo C., Marcilla A., Mormeneo S., Llombart-Bosch A., Sentandreu R. Specific immunohistochemical identification of Candida albicans in paraffin-embedded tissue with a new monoclonal antibody (1B12). Am J Clin Pathol 1995; 103:130–135
    [Google Scholar]
  27. Lischewski A., Amann R. I., Harmsen D., Merkert H., Hacker J., Morschhauser J. Specific detection of Candida albicans and Candida tropicalis by fluorescent in situ hybridization with an 18S rRNA-targeted oligonucleotide probe. Microbiology 1996; 142:2731–2740
    [Google Scholar]
  28. Williams D. W., Wilson M. J., Lewis M. A. O., Potts A. J. C. Identification of Candida species in formalin fixed, paraffin wax embedded oral mucosa by sequencing of ribosomal DNA. J Clin Pathol: Mol Pathol 1996; 49:M23–M28
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/jmm/10.1099/00222615-47-9-811
Loading
/content/journal/jmm/10.1099/00222615-47-9-811
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