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Detection of pathogenic fungi in human blood by the polymerase chain reaction
A M Polanco1, J L Rodríguez-Tudela, J V Martínez-Suárez
1Unidad de Micología, Instituto de Salud Carlos III, Madrid, Spain.
Abstract:
The ability of the polymerase chain reaction (PCR) to detect pathogenic fungi in human blood was investigated. A DNA fragment of about 300 bp from the 18S rDNA, highly conserved in all fungi, was amplified with target DNA from 18 different species of fungi commonly isolated from clinical samples. The presence of PCR products was confirmed by hybridization with a fluorescein-labelled internal probe (21-mer). The PCR assay described is sensitive enough to detect 125 fg of purified Candida albicans DNA and 10 to 100 yeast cells per millilitre of blood.
Insights
This study demonstrates the polymerase chain reaction (PCR) can detect pathogenic fungi in human blood. The developed PCR assay is highly sensitive for identifying fungal DNA and yeast cells in blood samples.
Area of Science:
- Medical Mycology
- Molecular Biology
- Diagnostic Microbiology
Background:
- Fungal infections pose significant health risks, particularly in immunocompromised individuals.
- Accurate and rapid detection of pathogenic fungi in clinical samples is crucial for effective treatment.
- Current diagnostic methods may lack sensitivity or require prolonged incubation periods.
Purpose of the Study:
- To investigate the efficacy of the polymerase chain reaction (PCR) for detecting pathogenic fungi directly in human blood.
- To establish a sensitive and specific molecular assay for fungal identification.
- To evaluate the limit of detection for fungal DNA and cells in blood.
Main Methods:
- Amplification of a conserved 300 bp fragment from the 18S ribosomal DNA (rDNA) gene present in various fungal species.
- Utilized target DNA from 18 common fungal species isolated from clinical specimens.
- Confirmation of PCR products via hybridization with a fluorescein-labeled 21-mer internal probe.
Main Results:
- The PCR assay successfully amplified fungal DNA from 18 different species of pathogenic fungi.
- The assay demonstrated high sensitivity, detecting as little as 125 fg of purified Candida albicans DNA.
- The method could identify 10 to 100 yeast cells per milliliter of blood.
Conclusions:
- The developed PCR assay is a sensitive and specific method for detecting pathogenic fungi in human blood.
- This molecular technique offers a rapid alternative to conventional methods for diagnosing fungal infections.
- The assay's sensitivity supports its potential application in clinical diagnostics for bloodstream fungal infections.