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Analysis of aetiology of invasive fungal infections using a panfungal PCR-based method in tissue biopsies with proven infection

Abstract number: O151

Cuenca-Estrella M., Buitrago M.J., Bernal-Martinez L., Gomez-Lopez A., Rodriguez-Tudela J.L.

Objectives: The etiological cause of many IFIs is never revealed. PCR-based methods can detect specific DNA of fungal species in tissue biopsies, in order to know the prevalence of fungal species. Those data will help us to choose the most efficient therapeutic alternative for first line therapies of IFI.

Methods: A total of 116 paraffined and fresh tissue biopsies were sent to Spanish Reference Mycology Laboratory between 2006 and 2010 from 36 different hospitals. Biopsies came from 93 patients suffering from proven infection as microscopical examination of tissues showed invasion by fungal structures. Cultures were negative for all samples. A total of 36 (30%) biopsies came from lungs, 23 (19%) were subcutaneous or skin biopsies, 19 (16%) from gastrointestinal tract, 10 brain biopsies, 9 from nasal sinus, 7 from liver, and 18 from other tissues. Procedures of DNA extraction and amplification followed routine methods. The tissue samples were analyzed using a panfungal Real-Time PCR-based assay. It was designed to amplify the ITS regions 1 and 2 from fungal rDNA gene complex. Amplified DNA was identified by sequencing. Controls were used in each set of experiments which were done in duplicate.

Results: A total of 80% of biopsies were positive by the PCR-based technique (93/116). When data were analyzed by patient, fungal DNA was detected and identified in 77 out of 93 patients (83%). Results by patients of species identification were: 37/77 (48%) Aspergillus fumigatus, 4 (5%) Aspergillus flavus, 7 (9%) Zygomycetes, 7 (9%) other phaeohyphomycosis with 3 scedosporiosis, 6 (7.7%) hyalohyphomycosis with 3 fusariosis, and 6 (7.7%) Candida. A total of 10 (13%) cases of histoplasmosis were detected as well. Other 4 cases were mixed infection. By samples, main data were: Aspergillus DNA was detected in 76% of positive lung biopsies and 43% of brain tissues, and Candida DNA in 50% of gastrointestinal biopsies.

Conclusion: (i) PCR-based methods can be used for species identification in tissue biopsies as a complementary technique of cultures. (ii) Emerging fungal pathogens could be underestimated as microbiological conventional methods have a limited accuracy for species detection and identification. (iii) Further studies are warranted.

Session Details

Date: 07/05/2011
Time: 00:00-00:00
Session name: Abstracts of 21st ECCMID / 27th ICC
Subject:
Location: Milan, Italy, 7 - 10 May 2011
Presentation type:
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