Search Results
Confirmation of Dermatophytes in Nail Specimens Using In-Office Dermatophyte Test Medium Cultures
Insights from a Multispecialty Survey
Using data from a multicenter nationwide multispecialty survey, the authors investigated the efficacy of in-office dermatophyte test medium (DTM) and central laboratory cultures used to confirm onychomycosis across samples collected by podiatric, dermatologic, and primary-care physicians. The samples collected by podiatric physicians were both positive or both negative in 43% and 27% of patients, respectively. Samples harvested by dermatologists were both positive in 37% of patients and both negative in 32%, while the samples collected by primary-care physicians were both positive in 28% of patients and both negative in 38%. The accuracy of DTM and central laboratory tests is dependent on the proper collection of nail samples, and the accuracy of mycologic test results varied significantly across nail specimens harvested by podiatric, dermatologic, and primary-care physicians. DTM culture was found to be an effective and convenient method of confirming dermatophyte infections in patients with signs of onychomycosis. The data presented here indicate that the special expertise of podiatric physicians in treating foot-related illnesses translates into more accurate mycologic testing. (J Am Podiatr Med Assoc 93(3): 195-202, 2003)
Treatment of Dermatophyte Toenail Onychomycosis in the United States
A Pharmacoeconomic Analysis
This study attempted to determine the cost-effectiveness of therapies for dermatophyte toenail onychomycosis in the United States in 2001. The antimycotic agents evaluated were ciclopirox 8% nail lacquer and the oral agents terbinafine, itraconazole (pulse), itraconazole (continuous), fluconazole, and griseofulvin. A treatment algorithm for the management of onychomycosis was developed, and a meta-analysis was carried out to determine the average mycologic and clinical response rates for the various agents. The cost of the regimen was figured as the sum of the costs of drug acquisition, medical management, and management of adverse effects. The expected cost of management and disease-free days were determined, and a sensitivity analysis was conducted. It was concluded that ciclopirox 8% nail lacquer, which has recently become available in the larger size of 6.6 mL, is a cost-effective agent for the management of toenail onychomycosis. (J Am Podiatr Med Assoc 92(5): 272-286, 2002)
Background
Podiatric physicians routinely use electric drills for the treatment of nail and skin conditions. The grinding process produces human nail and skin dust that is generally vacuumed into bags in the grinding unit. Many of the nails are thought to be mycotic, particularly because they are obtained from patients with symptoms of dermatophyte infections. Currently, there is limited information available on the detection of fungi from nail dust samples. Herein, we attempt to address this situation and outline some of the difficulties that pathology laboratories face in isolating and identifying dermatophytes from nail samples.
Methods
Fifty nail dust bags from podiatric medical clinics across all of the states and territories of Australia were collected and analyzed. Samples from the bags were inoculated onto primary isolation media. Fungal colonies that grew were then inoculated onto potato dextrose agar for identification using standard morphological (macroscopic and microscopic) features.
Results
One hundred fifty-one colonies of dermatophytes were identified from 43 of the 50 samples. In addition 471 nondermatophyte molds were isolated, along with some yeasts and bacteria.
Conclusions
The most common dermatophytes isolated were from the Trichophyton mentagrophytes/interdigitale complexes. Trichophyton rubrum, Trichophyton tonsurans, Trichophyton soudanense, and Epidermophyton floccosum were also isolated. An unidentified group of dermatophytes was also present. The three most common genera of nondermatophyte molds were Aspergillus, Penicillium, and Scopulariopsis, all of which have been implicated in onychomycosis and more general disease. The presence of viable fungal pathogens in the dust could potentially pose a health problem to podiatric physicians.
Background
Reports of mixed infections with nondermatophyte molds (NDMs) and dermatophytes in onychomycosis are rare, possibly owing to the inhibition of NDM growth during traditional culture. We sought to determine the prevalence of mixed infections in onychomycosis using molecular identification.
Methods
Molecular analyses were used to identify infecting organisms directly from at least two serial great toenail samples from each of the 44 patients.
Results
Mixed infections were present in 41% of the patients (18 of 44). A single coinfecting NDM was the most common mixed infection and was detected in 34% of patients with onychomycosis (15 of 44), with Fusarium oxysporum present in 14% (6 of 44), Scopulariopsis brevicaulis in 9% (4 of 44), Acremonium spp in 2% (1 of 44), Aspergillus spp in 4.5% (2 of 44), and Scytalidium spp in 4.5% (2 of 44). Mixed infections with two NDMs were found in 7% of patients (3 of 44).
Conclusions
Mixed onychomycosis infections may be more prevalent than previously reported.
Background
Dermatomycoses are a group of pathologic abnormalities frequently seen in clinical practice, and their prevalence has increased in recent decades. Diagnostic confirmation of mycotic infection in nails is essential because there are several pathologic conditions with similar clinical manifestations. The classical method for confirming the presence of fungus in nail is microbiological culture and the identification of morphological structures by microscopy.
Methods
We devised a nested polymerase chain reaction (PCR) that amplifies specific DNA sequences of dermatophyte fungus that is notably faster than the 3 to 4 weeks that the traditional procedure takes. We compared this new technique and the conventional plate culture method in 225 nail samples. The results were subjected to statistical analysis.
Results
We found concordance in 78.2% of the samples analyzed by the two methods and increased sensitivity when simultaneously using the two methods to analyze clinical samples. Now we can confirm the presence of dermatophyte fungus in most of the positive samples in just 24 hours, and we have to wait for the result of culture only in negative PCR cases.
Conclusions
Although this PCR cannot, at present, substitute for the traditional culture method in the detection of dermatophyte infection of the nails, it can be used as a complementary technique because its main advantage lies in the significant reduction of time used for diagnosis, in addition to higher sensitivity.
Onychomycosis Infections
Do Polymerase Chain Reaction and Culture Reports Agree?
Background:
Mycological culture is the traditional method for identifying infecting agents of onychomycosis despite high false-negative results, slower processing, and complications surrounding nondermatophyte mold (NDM) infections. Molecular polymerase chain reaction (PCR) methods are faster and suited for ascertaining NDM infections.
Methods:
To measure agreement between culture and PCR methods for identification of infecting species of suspected onychomycosis, single toenail samples from 167 patients and repeated serial samples from 43 patients with suspected onychomycosis were processed by culture and PCR for identification of 16 dermatophytes and five NDMs. Agreement between methods was quantified using the kappa statistic (κ).
Results:
The methods exhibited fair agreement for the identification of all infecting organisms (single samples: κ = 0.32; repeated samples: κ = 0.38). For dermatophytes, agreement was moderate (single samples: κ = 0.44; repeated samples: κ = 0.42). For NDMs, agreement was poor with single samples (κ = 0.16) but fair with repeated samples (κ = 0.25). Excluding false-negative reports from analyses improved agreement between methods in all cases except the identification of NDMs from single samples.
Conclusions:
Culture was three or four times more likely to report a false-negative result compared with PCR. The increased agreement between methods observed by excluding false-negative reports statistically clarifies and highlights the major discord caused by false-negative cultures. The increased agreement of NDM identification from poor to fair with repeated sampling along with their poor agreement in the single samples, with and without false-negatives, affirms the complications of NDM identification and supports the recommendation that serial samples help confirm the diagnosis of NDM infections.
Background: A high rate of false-negative dermatophyte detection is observed when the most common laboratory methods are used. These methods include microscopic observation of potassium hydroxide–digested nail clippings and culture methods using agar-based media supplemented with cycloheximide, chloramphenicol, and gentamicin to isolate dermatophytes. Microscopic detection methods that use calcofluor white staining or periodic acid–Schiff staining may also be substituted for and have previously been reported to be more sensitive than potassium hydroxide–digested nail clippings.
Methods: Trichophyton rubrum infections were detected directly from nails in a double-round polymerase chain reaction assay that uses actin gene–based primers. This method was compared with detection of fungal hyphae by using calcofluor white fluorescence microscopy of nail samples collected from 83 patients with onychomycosis who were undergoing antifungal drug therapy.
Results: Twenty-six of 83 samples (31.3%) were found to be positive by calcofluor white fluorescence microscopy, and 21 of 83 samples (25.3%) yielded positive results for T rubrum when actin gene–based primers in a double-round polymerase chain reaction assay were used. When calcofluor white fluorescence microscopy and polymerase chain reaction assay were used, the combined detection was 46.9% compared with 31.3% when calcofluor microscopy and culture of nail samples on Sabouraud’s dextrose agar supplemented with cycloheximide, chloramphenicol, and gentamicin were used.
Conclusions: These results suggest that the use of a direct DNA protocol is an alternative method for detecting Trichophyton infections. When this protocol is used, the presence of T rubrum DNA is directly detected. However, the viability of the dermatophyte is not addressed, and further methods need to be developed for the detection of viable T rubrum directly from nail samples. (J Am Podiatr Med Assoc 98(3): 224–228, 2008)
Fungal Diversity and Onychomycosis
An Analysis of 8,816 Toenail Samples Using Quantitative PCR and Next-Generation Sequencing
Background:
Onychomycosis is a fungal infection of the nail that is often recalcitrant to treatment and prone to relapse. Traditional potassium hydroxide and culture diagnosis is costly and time-consuming. Therefore, molecular methods were investigated to demonstrate effectiveness in diagnosis and to quantify the microbial flora present that may be contributing to disease.
Methods:
A total of 8,816 clinically suspicious toenail samples were collected by podiatric physicians across the United States from patients aged 0 to 103 years and compared with a control population (N = 20). Next-generation sequencing and quantitative polymerase chain reaction were used to identify and quantify dermatophytes, nondermatophyte molds, and bacteria.
Results:
Approximately 50% of suspicious toenails contained both fungi and bacteria, with the dermatophyte Trichophyton rubrum contributing the highest relative abundance and presence in 40% of these samples. Of the remaining 50% of samples, 34% had bacterial species present and 16% had neither. Fungi only were present in less than 1% of samples. Nondermatophyte molds contributed to 11.0% of occurrences in fungus-positive samples. All of the control samples were negative for fungi, with commensal bacterial species composing most of the flora population.
Conclusions:
Molecular methods were successful in efficiently quantifying microbial and mycologic presence in the nail. Contributions from dermatophytes were lower than expected, whereas the opposite was true for nondermatophyte molds. The clinical significance of these results is currently unknown.
Onychomycosis is a common problem seen in clinical practice. Given the differential diagnosis of dystrophic nails, it is helpful to obtain a definitive diagnosis of dermatophyte infection before initiation of antifungal therapy. Potassium hydroxide preparation and fungal culture, which are typically used in the diagnosis of these infections, often yield false-negative results. Recent studies have suggested that nail plate biopsy with periodic acid–Schiff stain may be a very sensitive technique for the diagnosis of onychomycosis. In this article, we review the literature on the utility of histopathologic analysis in the evaluation of onychomycosis. Many of these studies indicate that biopsy with periodic acid–Schiff is the most sensitive method for diagnosing onychomycosis. We propose that histopathologic examination is indicated if the results of other methods are negative and clinical suspicion is high; therefore, it is a useful complementary technique in the diagnosis of onychomycosis. (J Am Podiatr Med Assoc 95(3): 258–263, 2005)
Background:
Onychomycosis is a chronic nail infection caused by dermatophytes, Candida, nondermatophyte molds, and Trichosporon. The purpose of this study was to identify the underlying pathogen in patients with onychomycosis in our region.
Methods:
A retrospective analysis of 225 cases with onychomycosis, diagnosed over a 27-month period at the Department of Dermatoveneorology, Bezmialem Vakif University, Istanbul, Turkey, and confirmed with culture, was performed.
Results:
Patient age ranged from 2 to 87 years (mean ± SD, 41.59 ± 17.61), and female patients were more commonly affected (120 cases, 53.3%) than male patients. Lateral and distal subungual onychomycosis was detected in 180 cases (80%). Etiologic agents were as follows: Trichophyton rubrum, 77 cases (34.2%); Trichophyton mentagrophytes, 30 cases (13.3%), Candida albicans, 28 cases (12.4%); Candida parapsilosis, 25 cases (11.1%); Acremonium species, one case (0.4%); Aspergillus species, two cases (0.9%); Fusarium species, four cases (1.3%); and Trichosporon species, three cases (1.3%).
Conclusions:
The most frequent isolated etiologic agents were T rubrum for toenails and C albicans for fingernails.
