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A matrix-assisted laser desorption/ionization time of flight mass spectrometry reference database for the
Clara Valero1, María J Buitrago1, Sara Gago1
1Servicio de Micología, Centro Nacional de Microbiología, Instituto de Salud Carlos III, Majadahonda, Madrid, Spain.
Abstract:
The isolation of the pathogenic fungus Histoplasma capsulatum from cultures together with the visualization of typical intracellular yeast in tissues are the gold standard methods for diagnosis of histoplasmosis. However, cultures are time-consuming, require level 3 containment and experienced personnel, and usually call for an additional confirmation test. Matrix-Assisted Laser Desorption Ionization Time of Flight Mass Spectrometry (MALDI-ToF MS) has been established as a suitable tool for microbial identification in several clinical laboratories. A reference database has been constructed for the identification of H. capsulatum by MALDI-ToF MS by using six H. capsulatum strains previously identified by molecular methods. For validation, 63 fungal strains belonging to the Collection of the Spanish National Centre for Microbiology were tested against the new reference database combined with other commercial and in-house databases. In a blind assay, all H. capsulatum strains (n = 30) were correctly identified by the database and 86.6% had scores above 1.7. Considering both phases of the fungus for the same strain, the most reliable results were obtained with the mycelial phase, with only 13.3% of isolates having scores below 1.7. The new database was able to identify both morphological phases of the fungus. MALDI-ToF technology yields a prompt and simple identification from H. capsulatum yeast forms and early mycelial cultures. It allows for reducing response time and decreasing risk in fungus manipulation.
Insights
Matrix-Assisted Laser Desorption Ionization Time of Flight Mass Spectrometry (MALDI-ToF MS) offers a rapid and accurate method for identifying Histoplasma capsulatum. This new database improves diagnostic speed and safety for histoplasmosis detection.
Area of Science:
- Medical Mycology
- Clinical Microbiology
- Mass Spectrometry
Background:
- Histoplasmosis diagnosis relies on culture and tissue visualization, which are slow and require specialized facilities.
- Current diagnostic methods for Histoplasma capsulatum are time-consuming and necessitate additional confirmation steps.
Purpose of the Study:
- To develop and validate a Matrix-Assisted Laser Desorption Ionization Time of Flight Mass Spectrometry (MALDI-ToF MS) reference database for rapid Histoplasma capsulatum identification.
- To evaluate the efficacy of MALDI-ToF MS in identifying both yeast and mycelial phases of Histoplasma capsulatum.
Main Methods:
- A reference database was created using six molecularly identified Histoplasma capsulatum strains.
- Sixty-three fungal strains were tested against the new database, alongside commercial and in-house databases.
- A blind assay was conducted to assess identification accuracy and scoring.
Main Results:
- All 30 Histoplasma capsulatum strains were correctly identified using the developed database.
- 86.6% of isolates achieved identification scores above 1.7, with the mycelial phase yielding more reliable results.
- The database successfully identified both morphological phases of the fungus.
Conclusions:
- MALDI-ToF MS provides a prompt, simple, and reliable method for identifying Histoplasma capsulatum.
- This technology reduces diagnostic response times and minimizes risks associated with handling pathogenic fungi.
- The developed database enhances the utility of MALDI-ToF MS for clinical diagnostics of histoplasmosis.