Polymerase chain reaction in the diagnosis of onychomycosis: a large, single-institute study

Litz, C.E.; Cavagnolo, Robert

doi:10.1111/j.1365-2133.2010.09852.x

Cited by 46 publications

(34 citation statements)

References 18 publications

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“…In the present study, a similar approach was adopted, and TRFLP analysis was used to identify infectious fungi based on differences in their 28S rDNA amplicons. Other DNA sequences, such as that of the chitin synthase 1 gene or small ribosomal subunit 18S rRNA, were successfully used for fungal species delineation and identification (7,27,28). The polymorphism of the internal transcribed spacers (ITS) of ribosomal DNA regions (ITS1 and ITS2) flanking the DNA sequence composing the 5.8S rDNA is the most discriminating tool for distinguishing different fungi (1).…”

Section: Discussionmentioning

confidence: 99%

Identification of Infectious Agents in Onychomycoses by PCR-Terminal Restriction Fragment Length Polymorphism

Verrier

Pronina²,

Peter³

et al. 2012

J Clin Microbiol

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A fast and reliable assay for the identification of dermatophyte fungi and nondermatophyte fungi (NDF) in onychomycosis is essential, since NDF are especially difficult to cure using standard treatment. Diagnosis is usually based on both direct microscopic examination of nail scrapings and macroscopic and microscopic identification of the infectious fungus in culture assays. In the last decade, PCR assays have been developed for the direct detection of fungi in nail samples. In this study, we describe a PCR-terminal restriction fragment length polymorphism (TRFLP) assay to directly and routinely identify the infecting fungi in nails. Fungal DNA was easily extracted using a commercial kit after dissolving nail fragments in an Na 2 S solution. Trichophyton spp., as well as 12 NDF, could be unambiguously identified by the specific restriction fragment size of 5=-end-labeled amplified 28S DNA. This assay enables the distinction of different fungal infectious agents and their identification in mixed infections. Infectious agents could be identified in 74% (162/219) of cases in which the culture results were negative. The PCR-TRFLP assay described here is simple and reliable. Furthermore, it has the possibility to be automated and thus routinely applied to the rapid diagnosis of a large number of clinical specimens in dermatology laboratories.

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Section: Discussionmentioning

confidence: 99%

Identification of Infectious Agents in Onychomycoses by PCR-Terminal Restriction Fragment Length Polymorphism

Verrier

Pronina²,

Peter³

et al. 2012

J Clin Microbiol

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“…Litz et al [11] employed nested PCR specifically detecting dermatophytes and achieved a positive rate (37%) comparable to that of traditional microscopy (40%). If both methods were applied, the positive rate was 52% and similar to that of periodic acid-Schiff staining of the nails (54%).…”

Section: Real-time Pcr-based Strategiesmentioning

confidence: 90%

“…In the recent literature, numerous differing approaches have been applied including manual homogenization of the clinical specimen [7,8], bead beating [9,10 && ], freeze/thaw processing [11] and traditional but toxic phenolchloroform extraction [12,13]. Yet, there have been significant improvements from laborious multistep procedures [14,15] .…”

Section: Dna Extraction Methodsmentioning

confidence: 99%

Molecular diagnosis of dermatophyte infections

Jensen

Arendrup

2012

Current Opinion in Infectious Diseases

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“…A technical difficulty of PCR is the determination of the appropriate quantity of sample that will reassure a positive result. Different recommended amounts varying from 1-2 to 30-100 mg are mentioned in the literature (Bontems et al, 2009;Litz & Cavagnolo, 2010). In our study, the use of the dermatophyte PCR kit was evaluated in a larger number of samples (418 specimens), with a fixed large amount of sample (30-33 mg per sample).…”

Section: Discussionmentioning

confidence: 99%

“…However, the low sensitivity of culture in the diagnosis of onychomycosis is commonly accepted (Litz & Cavagnolo, 2010;Mehlig et al, 2014). However, direct microscopy, especially with the addition of fluorochrome and periodic acid-Schiff staining (Gupta et al, 2008;Idriss et al, 2013;Litz & Cavagnolo, 2010), has higher sensitivity but lacks specificity as it cannot provide identification of species or even at the genus level and does not differentiate unquestionably between dermatophytes and other moulds (Bontems et al, 2009;Brillowska-Dabrowska et al, 2007;Garg et al, 2007;Jensen & Arendrup, 2012;Mehlig et al, 2014). Taking .…”

mentioning

confidence: 99%

Evaluation of a commercial PCR test for the diagnosis of dermatophyte nail infections

Spiliopoulou

Bartzavali

Jelastopulu

et al. 2015

Journal of Medical Microbiology

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Tinea unguium, known as onychomycosis, is a dermatophyte infection of nails with worldwide distribution. Conventional methods for detecting fungi in nail specimens are either non-specific (microscopy) or insensitive (culture). PCR has been used to improve sensitivity in detecting the causative fungi in nail specimens from patients with suspected onychomycosis. Results of a commercial multiplex PCR for the detection of dermatophytes, especially Trichophyton rubrum (the main dermatophyte implicated), as compared to conventional methods are presented. A total of 418 nail scrapings obtained from dermatological outpatients were handled in the Laboratory of Microbiology between May 2010 and May 2013. Among them, multiplex PCR detected 126 (30.1 %) dermatophyte-positive samples, whereas culture revealed 44 (10.5 %). Direct microscopy revealed 63 (15.1 %) positive specimens. T. rubrum was identified in 116 out of 126 (92 %) positive PCR samples and 40 out of 44 (91 %) dermatophyte-positive cultures. Implementation of PCR increased species-specific detection of dermatophytes by 21.1 %, leading to a threefold increase as compared to culture alone. Multiplex PCR offers a time-saving diagnostic tool for tinea unguium and augments laboratory assistance to clinical evaluation for proper treatment. INTRODUCTIONTinea refers to superficial infection of skin, hair and nails due to one of three fungal genera, Microsporum, Epidermophyton and Trichophyton, collectively known as dermatophytes (Clayton & Midgley, 1989;Hay, 1995;Moriarty et al., 2012). These associated infections are among the most common diseases worldwide and cause serious chronic morbidity. Tinea unguium, a dermatophyte infection of nails, also known as onychomycosis, has a prevalence of at least 12.4 % in the general population of Europe (Moriarty et al., 2012), whereas in older individuals it is as high as 50 % (Salgo et al., 2003). The disease is often atypical and aggressive in patients with untreated human immunodeficiency virus infection. Patients with psoriasis not only have an increased risk of onychomycosis but also require laboratory confirmation as the two conditions may be clinically similar (Moriarty et al., 2012). Trichophyton rubrum is the main pathogen implicated (Brillowska-Dabrowska et al., 2007;Tsoumani et al., 2011), followed by Trichophyton interdigitale, formerly Trichophyton mentagrophytes var. interdigitale (Cafarchia et al., 2013;Clayton & Midgley, 1989; Gräser et al., 1999;Nenoff et al., 2007). Less commonly associated species are Epidermophyton floccosum and Trichophyton verrucosum (Moriarty et al., 2012). In addition to dermatophytes, Candida and non-dermatophyte moulds may be recovered from clinically affected nails; however, their clinical significance is controversial (Brillowska-Dabrowska et al., 2007). Conventional diagnosis is based on detection of fungal elements by direct microscopy of clinical specimens, followed by culture and morphological identification of the fungus. The whole procedure is time-consuming, requiring 10 to 15 ...

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Polymerase chain reaction in the diagnosis of onychomycosis: a large, single-institute study

Abstract: PCR is a specific, relatively sensitive test for onychomycosis. When used in conjunction with KOH, PCR can produce positivity rates similar to those with PAS alone.

Cited by 46 publications

References 18 publications

Identification of Infectious Agents in Onychomycoses by PCR-Terminal Restriction Fragment Length Polymorphism

Identification of Infectious Agents in Onychomycoses by PCR-Terminal Restriction Fragment Length Polymorphism

Molecular diagnosis of dermatophyte infections

Evaluation of a commercial PCR test for the diagnosis of dermatophyte nail infections

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