Clinical evaluation of a<i>T. rubrum</i>-specific polymerase chain reaction and pandermatophyte polymerase chain reaction in the diagnosis of suspected onychomycosis in 183 Serbian patients

Dubljanin, Eleonora; Čalovski, Ivana Čolović; Vujčić, Isidora S.; Džamić, Aleksandar; Arendrup, Maiken Cavling; Petersen, Randi Føns; Jensen, Rasmus H.

doi:10.1111/bjd.13168

Cited by 6 publications

(7 citation statements)

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“…Recently, molecular techniques have shown promise in the detection of the causative organisms of tinea unguium [10][11][12][13]. These techniques include conventional PCR [10,11,14,15], nested PCR [3,8], and real-time PCR [13,16,17].…”

Section: Introductionmentioning

confidence: 99%

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Development and Evaluation of a Novel Real-Time PCR for Pan-Dermatophyte Detection in Nail Specimens

et al. 2015

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An accurate diagnosis of tinea unguium is necessary for the selection of antimycotics and successful treatment. To rapidly and accurately identify the aetiological agents causing tinea unguium, we improved upon the conventional boiling method for DNA extraction and developed a novel real-time PCR detection system that includes two assays. The two assays, based on the amplification of ribosomal internal transcribed spacer regions and 28S rDNA, were designed to detect pan-dermatophyte and Trichophyton rubrum, respectively. The analytical sensitivities of both assays permitted the detection of ten copies of plasmid DNA template. The analytical specificity of the detection system was confirmed using 11 dermatophyte strains and 25 non-dermatophyte strains. In total, 165 nail specimens were examined by microscopy, culture, conventional PCR, and the novel real-time PCR method. Real-time PCR gave positive results in 47.3 % of the specimens (78), a rate exceeding those obtained using microscopy (72, 43.6 %), conventional PCR (69, 41.8 %), and culture (49, 29.7 %). All conventional PCR-positive specimens were detected by real-time PCR, and real-time PCR detected nine specimens that were missed by conventional PCR. The results from latent class analysis, and further calculations, showed that real-time PCR was the most sensitive method, but the diagnostic specificity of the four approaches was equivalent. In particular, molecular approaches may be more effective than microscopy and culture when the clinical symptoms of tinea unguium are not evident.

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Section: Introductionmentioning

confidence: 99%

“…These techniques include conventional PCR [10,11,14,15], nested PCR [3,8], and real-time PCR [13,16,17]. A limited number of studies have compared molecular diagnoses and conventional methods, but comparisons between molecular methods are infrequent.…”

Section: Introductionmentioning

confidence: 99%

Development and Evaluation of a Novel Real-Time PCR for Pan-Dermatophyte Detection in Nail Specimens

et al. 2015

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“…), whereas Li et al (2011) distinguished the groups of pathogens implicated in onychomycosis (dermatophytes, yeasts and moulds). In a recent work, the primers adopted by the commercial PCR kit that is evaluated in the present study were used in two separate PCRs for the detection of pandermatophytes and specific detection of T. rubrum, supplemented with a third PCR for T. interdigitale (Dubljanin et al, 2014). However, real-time PCR is less laborious, may enable a broader spectrum of simultaneous species detection and as a closed system reduces contamination risk (Alexander et al, 2011;Bergman et al, 2013;Bergmans et al, 2010;Miyajima et al, 2013;Paugam et al, 2013;Wisselink et al, 2011).…”

Section: Discussionmentioning

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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“…In addition, most of the available RT‐PCR methods target the ribosomal RNA region which contains repetitive elements, leading to higher sensitivity . Molecular methods also have short processing times of as little as 4 hours . However, they are still not widely used in microbiology laboratories, and only a few have been standardised for routine clinical work …”

Section: Introductionmentioning

confidence: 99%

“…A strategy based on real-time polymerase chain reaction (RT-PCR) would have a twofold advantage: increased differentiation capacity among species and limited risk of contamination due to the automated sample preparation and closed PCR format, making subsequent diagnostic methods unnecessary. 12,13 In addition, most of the available RT-PCR methods target the ribosomal RNA region which contains repetitive elements, leading to higher sensitivity. 4,5,14 Molecular methods also have short processing times of as little as 4 hours.…”

Section: Introductionmentioning

confidence: 99%

Evaluation of multiplex real‐time PCR for identifying dermatophytes in clinical samples—A multicentre study

Sherman

Goshen²,

Treigerman

et al. 2017

Mycoses

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The gold-standard method for dermatophyte identification involves direct microscopy and culture, which have inherent shortcomings. Only few molecular methods have been standardised for routine clinical work. This study aimed to develop and test a platform for identifying the most common dermatophytes in Israel using multiplex real-time polymerase chain reaction (RT-PCR). Specific primers were designed for the multiplex system (LightCycler 480) according to known cultures and validated by reference isolates. The dermatophyte detection rate was compared to smear and culture in 223 clinical samples obtained from a tertiary medical centre. Inconsistencies between methods were evaluated by sequencing. The RT-PCR was further evaluated in 200 community-based samples obtained from a health maintenance organisation and 103 military-personnel-based samples analysed at a central laboratory. In hospital-based clinical samples, complete concordance between methods was observed in 190 samples (85%; Kappa = 0.69). In most cases of non-concordance, sequencing was consistent with RT-PCR results. RT-PCR correctly identified all smear- and culture-positive cases in community and military-personnel samples. The results were available within 4 hours. The multiplex RT-PCR platform is a rapid and efficient method for identifying dermatophyte species in clinical samples and may serve as a first step in the diagnostic algorithm of superficial fungal infections.

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Clinical evaluation of aT. rubrum-specific polymerase chain reaction and pandermatophyte polymerase chain reaction in the diagnosis of suspected onychomycosis in 183 Serbian patients

Cited by 6 publications

References 10 publications

Development and Evaluation of a Novel Real-Time PCR for Pan-Dermatophyte Detection in Nail Specimens

Development and Evaluation of a Novel Real-Time PCR for Pan-Dermatophyte Detection in Nail Specimens

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

Evaluation of multiplex real‐time PCR for identifying dermatophytes in clinical samples—A multicentre study

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