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Development and Testing of Species-specific Quantitative PCR Assays for Environmental DNA Applications
Published on: November 5, 2020
Genus- and Species-Specific PCR Detection Methods
Jan Springer1, Jürgen Löffler2
1Medizinische Klinik II, Universitätklinikum Würzburg, Josef-Schneider-Str. 2, Würzburg, 97080, Germany.
Abstract:
PCR-based detection of fungal pathogens offers a sensitive and specific tool for the diagnosis of invasive fungal infections. A large variety of different clinical specimen types can be used as original material. However, certain precautions, in addition to the published MIQE guidelines [1], need to be taken to prevent contaminations from airborne fungal spores and PCR reagents. In addition, the European Aspergillus PCR Initiative (EAPCRI) recently defined standards for Aspergillus PCR [2, 3], following these recommendations leads to superior sensitivity. The combination of fungal PCR with the galactomannan ELISA assay increases the sensitivity for the detection of Aspergillus DNA from blood, compared to a single assay only [4, 5].
Insights
Polymerase chain reaction (PCR) is a sensitive method for detecting fungal infections. Following specific guidelines and combining PCR with galactomannan ELISA assays enhances detection sensitivity, especially for Aspergillus species.
Area of Science:
- Clinical Microbiology
- Molecular Diagnostics
- Infectious Diseases
Background:
- Invasive fungal infections require sensitive and specific diagnostic tools.
- Polymerase chain reaction (PCR) is a key molecular technique for pathogen detection.
- Contamination control is crucial for accurate PCR-based fungal diagnostics.
Purpose of the Study:
- To highlight the utility of PCR for diagnosing invasive fungal infections.
- To emphasize the importance of contamination prevention and adherence to MIQE guidelines.
- To present the European Aspergillus PCR Initiative (EAPCRI) standards for Aspergillus PCR.
- To evaluate the combined sensitivity of fungal PCR and galactomannan ELISA for Aspergillus detection in blood.
Main Methods:
- Utilizing PCR for the detection of fungal pathogens in various clinical specimens.
- Implementing strict precautions against airborne fungal spores and reagent contamination.
- Adhering to MIQE (Minimum Information for the Detection of Quantitative DNA Experiments) guidelines.
- Applying EAPCRI-defined standards for Aspergillus PCR.
- Combining fungal PCR with galactomannan ELISA assays.
Main Results:
- PCR provides a sensitive and specific method for diagnosing invasive fungal infections.
- Adherence to EAPCRI recommendations significantly improves Aspergillus PCR sensitivity.
- Combining fungal PCR with galactomannan ELISA assay enhances the detection of Aspergillus DNA in blood samples compared to single assays.
Conclusions:
- PCR is a valuable tool for fungal infection diagnosis, requiring careful contamination control.
- Standardized protocols, such as those from EAPCRI, optimize PCR performance.
- The combined use of fungal PCR and galactomannan ELISA offers improved sensitivity for detecting Aspergillus in blood.
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