Development and validation of a real-time multiplex PCR assay for the detection of dermatophytes and Fusarium spp.
J Med Microbiol
; 68(11): 1641-1648, 2019 Nov.
Article
em En
| MEDLINE
| ID: mdl-31526456
Introduction: Onychomycosis is a debilitating, difficult-to-treat nail fungal infection with increasing prevalence worldwide. The main etiological agents are dermatophytes, which are common causative pathogens in superficial fungal mycoses. Conventional detection methods such as fungal culture have low sensitivity and specificity and are time-consuming.Aim: The main objective of this study was to design, develop and validate a real-time probe-based multiplex qPCR assay for the detection of dermatophytes and Fusarium species.Methodology: The performance characteristics of the qPCR assays were evaluated. The multiplex qPCR assays targeted four genes (assay 1: pan-dermatophytes/Fusarium spp.; assay 2: Trichophyton rubrum/Microsporum spp.). Analytical validation was accomplished using 150 fungal isolates and clinical validation was done on 204 nail specimens. The performance parameters were compared against the gold standard (fungal culture) and expanded gold standard (culture in conjunction with sequencing).Results: Both the single-plex and multiplex qPCR assays performed well especially when compared against the expanded gold standard. Among the 204 tested nail specimens, the culture method showed that 125 (61.3 %) were infected with at least one organism, of which 40 yielded positive results for dermatophytes and Fusarium spp. These target organisms detected include 20 dermatophytes and 22 Fusarium spp. The developed qPCR assays demonstrated excellent limit of detection, efficiency, coefficient of determination, analytical and clinical sensitivity and specificity.Conclusion: The multiplex qPCR assays were reliable for the diagnosis of onychomycosis, with shorter turn-around time as compared to culture method. This aids in the planning of treatment strategies to achieve optimal therapeutic outcome.
Palavras-chave
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Dermatomicoses
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Arthrodermataceae
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Fusariose
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Reação em Cadeia da Polimerase Multiplex
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Reação em Cadeia da Polimerase em Tempo Real
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Fusarium
Tipo de estudo:
Diagnostic_studies
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Evaluation_studies
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Risk_factors_studies
Limite:
Humans
Idioma:
En
Revista:
J Med Microbiol
Ano de publicação:
2019
Tipo de documento:
Article
País de afiliação:
Singapura
País de publicação:
Reino Unido