Universal fungus‐specific primer systems and group‐specific hybridization oligonucleotides for 18S rDNA

Summary. We designed two primer systems that amplify a fragment of the gene coding for the small ribosomal subunit (18S rRNA). A broadly reactive, yet fungus‐specific, primer cocktail comprises two previously published primers, TR1 and TR2, which specifically amplify dermatophytes, and two newly designed primers, CA1 and AF2, which specifically amplify Candida and Aspergillus respectively. This primer cocktail amplifies a DNA fragment of approximately 578 basepairs (bp) in length (from position 838 to 1415), which contains variable, possibly species‐specific regions (V5, partly V7). Another newly designed primer, UF1 (universal fungal primer 1), along with the eukaryotic primer S3 amplifies a 926‐bp fragment (from position 263 to 1188) that includes the variable regions V3, V4 and V5. Both primer systems amplified DNA from Saccharomyces cerevisiae, Candida albicans, Cryptococcus neoformans, Aspergillus fumigatus, Penicillium marneffei, Fusarium oxysporum and Trichophyton mentagrophytes, but not the DNA from Prototheca zopfii, Escherichia coli or humans. The previously published oligonucleotides TR and HC, which are specific for dermatophytes and Histoplasma respectively, and the newly designed group‐specific oligonucleotides, CA and AF, hybridized with T. mentagrophytes, Histoplasma capsulatum, C. albicans and A. fumigatus respectively, but not with the other six fungi or with the three controls.

[1]  J. Meis,et al.  Phylogenetic relationships of five species of Aspergillus and related taxa as deduced by comparison of sequences of small subunit ribosomal RNA. , 1995, Journal of medical and veterinary mycology : bi-monthly publication of the International Society for Human and Animal Mycology.

[2]  M. Vidaud,et al.  Detection of Aspergillus species DNA in bronchoalveolar lavage samples by competitive PCR , 1995, Journal of clinical microbiology.

[3]  A. Llombart‐Bosch,et al.  Specific immunohistochemical identification of Candida albicans in paraffin-embedded tissue with a new monoclonal antibody (1B12). , 1995, American journal of clinical pathology.

[4]  T. G. Mitchell,et al.  Molecular mycology: DNA probes and applications of PCR technology. , 1994, Journal of medical and veterinary mycology : bi-monthly publication of the International Society for Human and Animal Mycology.

[5]  B. Johansson,et al.  Modifications of human and viral deoxyribonucleic acid by formaldehyde fixation. , 1994, Laboratory investigation; a journal of technical methods and pathology.

[6]  S. Murayama,et al.  Specific detection of Aspergillus and Penicillium species from respiratory specimens by polymerase chain reaction (PCR). , 1994, Japanese journal of medical science & biology.

[7]  K. Rand,et al.  Detection of candidemia by polymerase chain reaction. , 1994, Molecular and cellular probes.

[8]  K. Makimura,et al.  Detection of a wide range of medically important fungi by the polymerase chain reaction. , 1994, Journal of medical microbiology.

[9]  P. Nickel,et al.  Polymerase chain reaction‐based detection of dermatophyte DNA with a fungus‐specific primer system , 1994, Mycoses.

[10]  R. Kappe,et al.  Rapid presumptive identification of medically relevant yeasts to the species level by polymerase chain reaction and restriction enzyme analysis. , 1994, Journal of medical and veterinary mycology : bi-monthly publication of the International Society for Human and Animal Mycology.

[11]  R. Cannon,et al.  Detection of Candida albicans and other yeasts in blood by PCR , 1994, Journal of clinical microbiology.

[12]  Yves Van de Peer,et al.  Compilation of small ribosomal subunit RNA structures , 1993, Nucleic Acids Res..

[13]  C. Spreadbury,et al.  Detection of Aspergillus fumigatus by polymerase chain reaction , 1993 .

[14]  E. Linder,et al.  Detection of Pneumocystis carinii DNA in sputum and bronchoalveolar lavage samples by polymerase chain reaction , 1993, Journal of clinical microbiology.

[15]  Tadashi Mabuchi,et al.  Isolation and characterization of a species-specific DNA fragment for detection of Candida albicans by polymerase chain reaction , 1992, Journal of clinical microbiology.

[16]  Y. Peer,et al.  Database on the structure of small ribosomal subunit RNA , 1997, Nucleic Acids Res..

[17]  Y. Peer,et al.  Database on the structure of large ribosomal subunit RNA , 1996, Nucleic Acids Res..

[18]  L. Litzky,et al.  Rapid method for detection of Aspergillus 5S ribosomal RNA using a genus-specific oligonucleotide probe. , 1995, American journal of clinical pathology.

[19]  B. Petrini,et al.  Detection of Candida albicans DNA in serum by polymerase chain reaction. , 1994, Scandinavian journal of infectious diseases.