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Rapid identification of <i>Histoplasma capsulatum</i> in culture lysates

Rapid identification of Histoplasma capsulatum in culture lysates

Abstract

Background

Histoplasma capsulatum is the agent of histoplasmosis, a deep mycosis mainly afflicting immunocompromised patients. Rapid identification of the fungus isolated from clinical specimens allows timely administration of specific treatment.

Aim

To assess the ability of a dual PCR system targeting specific H. capsulatum DNA sites to identify fungal species in simple aqueous lysates from cultured fungi.

Methods

We analysed the performance of two independent PCR reactions designed to amplify fragments of 111 and 279bp included in H. capsulatum-specific gene AgM. We used 248 H. capsulatum strains and 68 isolates of other fungal species. Reaction templates consisted of aqueous lysates of cultured fungi (either in mycelial or yeast phase) obtained after three cycles of boiling and immediate cooling at 0°C. Selected strains were submitted to conventional DNA extraction and/or sequencing.

Results

Both PCR systems performed identically. Amplification from aqueous lysates was achieved from 239 H. capsulatum strains; the remaining 9 strains only showed specific bands when purified DNA was used as template. Of all other fungal species tested, only 2 Emmonsia crescens strains amplified H. capsulatum-specific bands and sequences of their amplified PCR products matched > 97% with H. capsulatum sequences. Total test time did not exceed 7h with 96% sensitivity, 97% specificity and 99% positive predictive value.

Conclusions

The assay is fast, accurate and economical, and can be an alternative method for presumptive identification of H. capsulatum in simple culture lysates.

Keywords: Histoplasma capsulatum. Molecular identification. PCR.


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Belén Ibarra-Camou a, Adriana Inés Toranzo a, William Lee a, Graciela Davel a, Cristina Elena Canteros a, ,

a Departamento de Micología, INEI-ANLIS Dr. Carlos G. Malbrán, Buenos Aires, Argentina
Journal: Rapid identification of <i>Histoplasma capsulatum</i> in culture lysates

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