Evaluation of Pan-Dermatophyte Nested PCR in Diagnosis of Onychomycosis

Garg, Jaya; Tilak, Ragini; Singh, Sanjay; Gulati, Anil Kumar; Garg, Avneet; Prakash, Pradyot; Nath, Gopal

doi:10.1128/jcm.02367-06

Cited by 51 publications

(66 citation statements)

References 9 publications

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“…Thus, an alternative approach by use of a combination of PCR and culture dermatophyte-positive results as the gold standard stands for a more realistic assessment of true onychomycosis cases. Similar approaches have been implemented by other researchers (Garg et al, 2007;Rothmund et al, 2013). Low PCR diagnostic indices such as PPV, likelihood ratios and specificity, at a lesser extent, by the use of conventional methods as the gold standard can be explained by the lower number of onychomycosis cases identified by culture and/or direct microscopy A. Spiliopoulou and others as compared to culture.…”

Section: Discussionmentioning

confidence: 84%

“…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 .…”

Section: Discussionmentioning

confidence: 99%

“…Statistical significance was achieved if P,0.05. For an appropriate diagnostic test, desirable values for LR+ and LR2 should be ¢10 and ¡0.1, respectively (Garg et al, 2007). A DOR .1 is indicative of useful tests and higher DORs are indicative of better test performance.…”

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confidence: 99%

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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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Section: Discussionmentioning

confidence: 84%

Section: Discussionmentioning

confidence: 99%

See 1 more Smart Citation

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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“…mentagrophytes'e özgül nPCR arasında iyi düzeyde (κ= 0.78), pan-dermatofi t nPCR ile T.rubrum/T.mentagrophytes'e özgül nPCR arasında mükemmel düzeyde uyum bulunmuştur (κ= 0.93). Hat 4,5,[7][8][9][10]12,16,19,21: Negatif örnekler; Hat 6,11,[13][14][15]17,18,20, …”

Section: çAlışmaya Alınan 123 öRneğin 67'sinde (%55) Pan-dermatofi T unclassified

The Use of Polymerase Chain Reaction in Laboratory Diagnosis of Dermatophytosis

Tiryaki¹,

Korkmazgil

Eyigör³

et al. 2015

Mikrobiyol Bul

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* Bu çalışma, Adnan Menderes Üniversitesi Araştırma Fon Saymanlığı tarafından TPF-10010 sayılı proje olarak desteklenmiş ve XXXV. Türk Mikrobiyoloji Kongresi (3-7 Kasım 2011, Kuşadası)'nde sözel bildiri olarak sunulmuştur. ÖZDermatofi tler toplumda sık görülen enfeksiyonlara yol açan mantarlardır. Dermatofi tozların tanısında kullanılan klasik yöntemlerden direkt mikroskopik incelemenin tür ayrımını yapamaması, kültürün ise uzun süre gerektirmesi ve duyarlılığının düşük olması gibi dezavantajları bulunmaktadır. Bu çalışmada, klinik örneklerden dermatofi tlerin saptanmasında ve kültürden izole edilen suşların tanımlanmasında polimeraz zincir reaksiyonu (PCR) yönteminin performansının araştırılması amaçlanmıştır. Çalışmaya, dermatofi toz ön tanısı konulan 110 hastaya (69 kadın, 41 erkek; yaş aralığı: 4-82 yıl) ait 63'ü deri ve 60'ı tırnak kazıntısı olmak üzere toplam 123 örnek alınmıştır. Örnekler, rutin direkt mikroskopi ve mantar kül-türünün yanı sıra iki turlu (nested) PCR (nPCR) yöntemine dayalı iki ayrı protokol ile incelenmiştir. Birinci protokolde dermatofi tlerin kitin sentaz genini (CHS-1) hedefl eyen pan-dermatofi t nPCR; ikinci protokolde ise Trichophyton rubrum ve T. mentagrophytes'e özgül ITS-1 genlerini hedefl eyen primerlerin kullanıldığı nPCR uygulanmıştır. Kültürde üreyen suşlara da aynı şekilde PCR uygulanmıştır. Pan-dermatofi t nPCR'da pozitif, T.rubrum/T.mentagrophytes'e özgül nPCR ile negatif sonuç veren örnekler ayrıca dizi analizine alınmıştır. Çalışmamızda, örneklerin 62'sinde (%50) direkt mikroskopik incelemede hif ve/veya spor yapıları görülmüş, 30'unun (%24) kültüründe dermatofi t üremiştir. T.rubrum/T.mentagrophytes'e özgül nPCR ile kültürde üreyen izolatların 28'i T.rubrum, ikisi T.mentagrophytes olarak tanımlanmıştır. Direkt uygulamada klinik örneklerin 67'si (%55) pan-dermatofi t nPCR ile, 65'i (%53) ise T. rubrum/T. mentagrophytes'e özgül nPCR ile pozitif bulunmuştur. Direkt mikroskopide mantar elemanı görülmeyen örneklerin kültüründe de üreme olmamış, bu örneklerin dokuzu pan-dermatofi t nPCR, sekizi T. rubrum/T. mentagrophytes'e özgül nPCR ile pozitif sonuç vermiştir. Kültüründe dermatofi t üremesi olan 30 örneğin ikisi, direkt olarak uygulanan pan-dermatofi t nPCR ile, biri T.rubrum/T.mentagrophytes'e özgül nPCR ile Geliş Tarihi (Received): 05.11.2014 • Kabul Ediliş Tarihi (Accepted): 02.12.2014

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“…Molecular methods, such as restriction fragment length polymorphism analysis of mitochondrial DNA [11], sequencing of the internal transcribed spacer (ITS) region of the ribosomal DNA [12], sequencing of protein-encoding genes, arbitrarily primed PCR [AP-PCR] [13], pan-dermatophyte nested PCR and PCR fi ngerprinting [14], have brought important progress in distinguishing between species and strains.…”

Section: Introductionmentioning

confidence: 99%

Genotyping and Molecular Characterization of Dermatophytes Isolates Collected from Clinical Samples

Sheikhi¹,

Saeed²,

Mirzaahmadi³

et al. 2017

Arch Pulmonol Respir Care

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Background: Detection of dermatophytes by microbiological method is sometimes problematic and some atypical microscopic or macroscopic morphology are non-detectable. Due to morphological similarity and existing intermediate forms and variants, unequivocally separating these dermatophytes is not always straightforward, and sampling appropriate isolates for research is often troublesome. The aim of this study was to compare and evaluate use of sequencing chitin synthase 1 gene (CHS1) with conventional methods for identifi cation of dermatophytes species and we researched the genetic patterns of samples collected for general phylogenetic analysis. Material and Method:In the primary screening of 250 clinical samples by KOH microscopy method, 64 isolates has been detected as dermatophytes. All samples were cultured and amplifi ed by PCR Method and positive PCR samples have been sequenced. Clinical isolates (64/250) were analyzed by using sequencing gene CHS1 and genotyped by program DNAMAN and MEGA. Result:The all data were compared with the international database of national center for biotechnology information website. Based on reference sequences of different genotype strains, it was noted that most strains of Trichophyton mentagrophytes were misidentifi cations of Trichophyton interdigitale. Conclusion:This research demonstrated that nested PCR and sequencing can be considered as standard method for the diagnosis of dermatophytosis. Also research gives a fi rst result on genetic evolution of the Dermatophytes strains distributing in Iran. It may aid in the creation of a national database that will be a valuable support for further studies.

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Evaluation of Pan-Dermatophyte Nested PCR in Diagnosis of Onychomycosis

Cited by 51 publications

References 9 publications

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

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

The Use of Polymerase Chain Reaction in Laboratory Diagnosis of Dermatophytosis

Genotyping and Molecular Characterization of Dermatophytes Isolates Collected from Clinical Samples

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