Identification of fungal pathogens in Formalin-fixed, Paraffin-embedded tissue samples by molecular methods

Area of Science:

• Medical Mycology
• Molecular Diagnostics
• Infectious Diseases

Background:

• Invasive fungal infections (IFI) etiology is poorly understood due to limitations in traditional diagnostic methods like cultures and histopathology.
• Diagnostic gaps hinder optimal antifungal use and contribute to adverse patient outcomes.

Purpose of the Study:

• To evaluate molecular methods, specifically PCR and in situ hybridization, for identifying fungal pathogens in Formalin-fixed, Paraffin-embedded (FFPE) tissue.
• To highlight the challenges and limitations in current molecular diagnostic approaches for IFI.

Main Methods:

• Utilized PCR assays (species-specific and broad-range) on FFPE samples from patients with proven IFI.
• Reviewed studies employing in situ hybridization for fungal identification in clinical samples.
• Assessed the impact of various factors on diagnostic sensitivity, including sample types, target sequences, and probe chemistry.

Main Results:

• Fungal species identification rates using PCR on FFPE samples ranged from 15-90%, with significant heterogeneity.
• Comparison across studies is difficult due to inconsistent reporting of controls for false positives, false negatives, and PCR inhibition.
• In situ hybridization studies also showed variability in sample types, targeted fungi, and detection methods.

Conclusions:

• Molecular methods offer an alternative for fungal pathogen identification from FFPE tissues, but diagnostic sensitivity varies.
• Standardized reporting of controls is crucial for comparing and validating molecular diagnostic studies for IFI.
• Parallel use of PCR and in situ hybridization may provide additive insights and a better understanding of fungal etiology, while acknowledging the limitations of each approach.