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(Referência obtida automaticamente do Web of Science, por meio da informação sobre o financiamento pela FAPESP e o número do processo correspondente, incluída na publicação pelos autores.)

Molecular Diagnosis of Pathogenic Sporothrix Species

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Autor(es):
Rodrigues, Anderson Messias [1] ; Sybren de Hoog, G. [2] ; de Camargo, Zoilo Pires [1]
Número total de Autores: 3
Afiliação do(s) autor(es):
[1] Univ Fed Sao Paulo, UNIFESP, Dept Microbiol Imunol & Parasitol, Disciplina Biol Celular, Sao Paulo, SP - Brazil
[2] CBS KNAW Fungal Biodivers Ctr, Utrecht - Netherlands
Número total de Afiliações: 2
Tipo de documento: Artigo Científico
Fonte: PLoS Neglected Tropical Diseases; v. 9, n. 12 DEC 2015.
Citações Web of Science: 21
Resumo

Background Sporotrichosis is a chronic (sub) cutaneous infection caused by thermodimorphic fungi in the order, Ophiostomatales. These fungi are characterized by major differences in routes of transmission, host predilections, species virulence, and susceptibilities to antifungals. Sporothrix species emerge in the form of outbreaks. Large zoonoses and sapronoses are ongoing in Brazil and China, respectively. Current diagnostic methods based on morphology and physiology are inaccurate due to closely related phenotypes with overlapping components between pathogenic and non-pathogenic Sporothrix. There is a critical need for new diagnostic tools that are specific, sensitive, and cost-effective. Methodology We developed a panel of novel markers, based on calmodulin (CAL) gene sequences, for the large-scale diagnosis and epidemiology of clinically relevant members of the Sporothrix genus, and its relative, Ophiostoma. We identified specific PCR-based markers for S. brasiliensis, S. schenckii, S. globosa, S. mexicana, S. pallida, and O. stenoceras. We employed a murine model of disseminated sporotrichosis to optimize a PCR assay for detecting Sporothrix in clinical specimens. Results Primer-BLAST searches revealed candidate sequences that were conserved within a single species. Species-specific primers showed no significant homology with human, mouse, or microorganisms outside the Sporothrix genus. The detection limit was 10-100 fg of DNA in a single round of PCR for identifying S. brasiliensis, S. schenckii, S. globosa, S. mexicana, and S. pallida. A simple, direct PCR assay, with conidia as a source of DNA, was effective for rapid, low-cost genotyping. Samples from a murine model of disseminated sporotrichosis confirmed the feasibility of detecting S. brasiliensis and S. schenckii DNA in spleen, liver, lungs, heart, brain, kidney, tail, and feces of infected animals. Conclusions This PCR-based method could successfully detect and identify a single species in samples from cultures and from clinical specimens. The method proved to be simple, high throughput, sensitive, and accurate for diagnosing sporotrichosis. (AU)

Processo FAPESP: 09/54024-2 - Biologia molecular e proteômica de fungos de interesse médico: Paracoccidioides brasiliensis e Sporothrix schenckii
Beneficiário:Zoilo Pires de Camargo
Linha de fomento: Auxílio à Pesquisa - Temático