279. Comparison of molecular techniques for identification of Fusarium species from clinical samples
Session: Poster Abstract Session: Diagnostic Microbiology; Novel Molecular Methods
Thursday, October 3, 2013
Room: The Moscone Center: Poster Hall C
Posters
  • Poster ID Week_ Marcela de Souza.pdf (460.4 kB)
  • Background: The genus Fusarium emerged over the past three decades as an opportunistic pathogen of immunocompromised hosts, mainly those with persistent and severe neutropenia. The early diagnosis of infection may optimize the antifungal therapy and be crucial for patient recovery. Nucleic acid based methods are often used to identify Fusarium sp. This project aimed to compare the techniques of loop - mediated isothermal amplification (LAMP), Real Time and DNA sequencing for the identification of Fusarium sp. isolated from blood culture of patients with onco-hematologic disease from University Hospital at Unicamp.

    Methods: Sixteen Fusarium sp. strains were selected from 2006 to 2012, belonging to the Mycology Laboratory. All strains have been previously identified by classical methods (macroscopic and micro morphological characteristics). The LAMP technique consisted of three primers pairs that identified six distinct regions of the gene RBP2. The Real Time PCR was performed in order to identify Fusarium solani species and Fusarium non-solani species complex using EF1α gene. The sequencing was performed in an ABI Prism Genetic Analyzer 3100 (Applied Biosystems) using EF1α gene amplification and BigDye® (Applied Biosystems) reagent, according to manufacturer instructions. The sequences were assembled using ATSQ version 6.0.1 (Genetix) and compared with database information available at NCBI bank (National center for biotechnology information).

    Results: Fourteen of the strains were identified as belonging to Fusarium solani species complex and two as Fusarium non-solani species complex using LAMP and Real Time PCR methodologies. The sequencing analysis identified fourteen strains as Fusarium solani species complex and two as Fusarium napiforme (non-solani). The tree methodology showed 100% of specificity and sensitivity and 100% of concordance among them.

    Conclusion: Considering the high performance of the three methodologies we conclude that LAMP is more accessible when compared with sequencing and real time PCR techniques, since it shows a lower cost and quickness performance. These results indicate a promising methodology that can be used for filamentous fungi routine diagnostic, assisting in therapy and appropriate treatment of patients.

    Marcela Souza, BS1, Tetsuhiro Matsuzawa, PhD2, Tohru Gonoi, PhD2, Katsuhiko Kamei, PhD2, Yasunori Muraosa, PhD2, Angélica Schreiber, PhD1, Luzia Lyra1, Maria Luiza Moretti, PhD1 and Plínio Trabasso, PhD1, (1)Unicamp, Campinas, Brazil, (2)Chiva University, Chuo-ku, Japan

    Disclosures:

    M. Souza, None

    T. Matsuzawa, None

    T. Gonoi, None

    K. Kamei, None

    Y. Muraosa, None

    A. Schreiber, None

    L. Lyra, None

    M. L. Moretti, None

    P. Trabasso, None

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