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Detection and differentiation of fungi in clinical specimens using polymerase chain reaction (PCR) amplification and restriction enzyme analysis. 1993

R L Hopfer, and P Walden, and S Setterquist, and W E Highsmith
UNC Hospitals, Department of Clinical Microbiology-Immunology, University of North Carolina, Chapel Hill 27514.

We have developed a method for processing and detecting fungi in clinical specimens using the polymerase chain reaction (PCR) methodology. This PCR amplification of a segment of a ribosomal DNA gene results in a 310 bp product. The gene sequences used as amplimers have been highly conserved throughout the fungal kingdom and positive PCR results have been obtained for all genera and species of fungi tested (n = 42). Neither human nor a variety of pathogenic bacteria (n = 24) gave an amplified product. This PCR method can detect as few as 15 cells of Candida albicans in clinical specimens following a simple processing procedure. The sensitivity of the PCR for detecting other fungi remains to be determined. Analysis of the 310 bp amplified product, following digestion with the restriction enzyme HaeIII, can be used to further characterize the identity of the fungus involved into the following five groups: (i) Candida species and closely related yeasts; (ii) Cryptococci and Trichosporon species; (iii) Aspergilli and clinically related septate molds; (iv) the Zygomycetes; and (v) the dimorphic fungi.

UI MeSH Term Description Entries
D008969 Molecular Sequence Data Descriptions of specific amino acid, carbohydrate, or nucleotide sequences which have appeared in the published literature and/or are deposited in and maintained by databanks such as GENBANK, European Molecular Biology Laboratory (EMBL), National Biomedical Research Foundation (NBRF), or other sequence repositories. Sequence Data, Molecular,Molecular Sequencing Data,Data, Molecular Sequence,Data, Molecular Sequencing,Sequencing Data, Molecular
D009181 Mycoses Diseases caused by FUNGI. Fungus Diseases,Fungal Diseases,Fungal Infections,Fungus Infections,Disease, Fungal,Disease, Fungus,Diseases, Fungal,Diseases, Fungus,Fungal Disease,Fungal Infection,Fungus Disease,Fungus Infection,Infection, Fungal,Infection, Fungus,Infections, Fungal,Infections, Fungus
D009841 Oligonucleotides Polymers made up of a few (2-20) nucleotides. In molecular genetics, they refer to a short sequence synthesized to match a region where a mutation is known to occur, and then used as a probe (OLIGONUCLEOTIDE PROBES). (Dorland, 28th ed) Oligonucleotide
D012150 Polymorphism, Restriction Fragment Length Variation occurring within a species in the presence or length of DNA fragment generated by a specific endonuclease at a specific site in the genome. Such variations are generated by mutations that create or abolish recognition sites for these enzymes or change the length of the fragment. RFLP,Restriction Fragment Length Polymorphism,RFLPs,Restriction Fragment Length Polymorphisms
D004271 DNA, Fungal Deoxyribonucleic acid that makes up the genetic material of fungi. Fungal DNA
D004587 Electrophoresis, Agar Gel Electrophoresis in which agar or agarose gel is used as the diffusion medium. Electrophoresis, Agarose Gel,Agar Gel Electrophoresis,Agarose Gel Electrophoresis,Gel Electrophoresis, Agar,Gel Electrophoresis, Agarose
D005658 Fungi A kingdom of eukaryotic, heterotrophic organisms that live parasitically as saprobes, including MUSHROOMS; YEASTS; smuts, molds, etc. They reproduce either sexually or asexually, and have life cycles that range from simple to complex. Filamentous fungi, commonly known as molds, refer to those that grow as multicellular colonies. Fungi, Filamentous,Molds,Filamentous Fungi,Filamentous Fungus,Fungus,Fungus, Filamentous,Mold
D006801 Humans Members of the species Homo sapiens. Homo sapiens,Man (Taxonomy),Human,Man, Modern,Modern Man
D001483 Base Sequence The sequence of PURINES and PYRIMIDINES in nucleic acids and polynucleotides. It is also called nucleotide sequence. DNA Sequence,Nucleotide Sequence,RNA Sequence,DNA Sequences,Base Sequences,Nucleotide Sequences,RNA Sequences,Sequence, Base,Sequence, DNA,Sequence, Nucleotide,Sequence, RNA,Sequences, Base,Sequences, DNA,Sequences, Nucleotide,Sequences, RNA
D012680 Sensitivity and Specificity Binary classification measures to assess test results. Sensitivity or recall rate is the proportion of true positives. Specificity is the probability of correctly determining the absence of a condition. (From Last, Dictionary of Epidemiology, 2d ed) Specificity,Sensitivity,Specificity and Sensitivity

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