Accurate Identification of Candida parapsilosis (Sensu Lato) by Use of Mitochondrial DNA and Real-Time PCR

Autor(es)

Souza, Ana Carolina R. ^[1] ; Ferreira, Renata C. ^{[2, 1, 3]} ; Goncalves, Sarah S. ^[1] ; Quindos, Guillermo ^[4] ; Eraso, Elena ^[4] ; Bizerra, Fernando C. ^[1] ; Briones, Marcelo R. S. ^{[2, 3]} ; Colombo, Arnaldo L. ^[1]

Afiliação do(s) autor(es)

^[1] Univ Fed Sao Paulo, Lab Especial Micol, Disciplina Infectol, Sao Paulo - Brazil

^[2] Univ Fed Sao Paulo, Dept Microbiol Imunol & Parasitol, Sao Paulo - Brazil

^[3] Univ Fed Sao Paulo, Lab Genom Evolut & Biocomplexidade, Sao Paulo - Brazil

^[4] Univ Pais Vasco Euskal Herriko Unibertsitatea, Lab Micol Med, Dept Inmunol Microbiol & Parasitol, Fac Med & Odontol, Bilbao - Spain

• Tipo de documento: Artigo Científico
• Fonte: Journal of Clinical Microbiology; v. 50, n. 7, p. 2310-2314, JUL 2012.
• Citações Web of Science: 19

Resumo

Candida parapsilosis is the Candida species isolated the second most frequently from blood cultures in South America and some European countries, such as Spain. Since 2005, this species has been considered a complex of 3 closely related species: C. parapsilosis, Candida metapsilosis, and Candida orthopsilosis. Here, we describe a real-time TaqMan-MGB PCR assay, using mitochondrial DNA (mtDNA) as the target, which readily distinguishes these 3 species. We first used comparative genomics to locate syntenic regions between these 3 mitochondrial genomes and then selected NADH5 as the target for the real-time PCR assay. Probes were designed to include a combination of different single-nucleotide polymorphisms that are able to differentiate each species within the C. parapsilosis complex. This new methodology was first tested using mtDNA and then genomic DNA from 4 reference and 5 clinical strains. For assay validation, a total of 96 clinical isolates and 4 American Type Culture Collection (ATCC) isolates previously identified by internal transcribed spacer (ITS) ribosomal DNA (rDNA) sequencing were tested. Real-time PCR using genomic DNA was able to differentiate the 3 species with 100% accuracy. No amplification was observed when DNA from other species was used as the template. We observed 100% congruence with ITS rDNA sequencing identification, including for 30 strains used in blind testing. This novel method allows a quick and accurate intracomplex identification of C. parapsilosis and saves time compared with sequencing, which so far has been considered the ``gold standard{''} for Candida yeast identification. In addition, this assay provides a useful tool for epidemiological and clinical studies of these emergent species. (AU)

• Processo FAPESP: 10/02752-1 - Identificação de espécies do complexo Candida "para-meta-ortho-psilosis" por PCR-tempo real
• Beneficiário: Ana Carolina Remondi Souza
• Modalidade de apoio: Bolsas no Brasil - Mestrado
• Processo FAPESP: 07/08575-1 - Candidíase hematogênica: uma abordagem multidisciplinar para o aprimoramento de ferramentas diagnósticas e caracterização de biomarcadores relacionados ao seu prognóstico
• Beneficiário: Arnaldo Lopes Colombo
• Modalidade de apoio: Auxílio à Pesquisa - Temático
• Processo FAPESP: 10/17179-5 - Estudo de proteômica como estratégia para identificação de biomarcadores para diagnóstico precoce de candidemia
• Beneficiário: Fernando César Bizerra
• Modalidade de apoio: Bolsas no Brasil - Pós-Doutorado
• Processo FAPESP: 09/01230-4 - Aplicação clínica da sistemática molecular de Candida spp. na perspectiva da genômica comparativa por genealogias em rede e análise bayesiana
• Beneficiário: Renata Carmona e Ferreira
• Modalidade de apoio: Bolsas no Brasil - Pós-Doutorado