Evaluation of a Liquid Media MALDI-TOF MS Protocol for the Identification of Dermatophytes Isolated from Tinea capitis Infections.

One of the most common types of tinea is the superficial infection of the hair and scalp area known as tinea capitis. It is responsible for frequent outbreaks in nurseries and schools and represents a global health problem. Correct identification of the infection agent is essential in the determination of the infection source, epidemiological course, and treatment initiation. The conventional identification methods (direct exam, culture, DNA sequencing) are time-consuming, require experienced staff, are time-consuming, and the latter is expensive for routine identifications. Matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) is gaining new ground for routine identification of filamentous fungi. The main advantages of MALDI-TOF MS are its rapid and accurate identification capability, relatively low cost, and easy integration into the laboratory routine. Its accuracy heavily depends on the quality of the reference spectra database. Identification of clinical isolates with MALDI-TOF MS protocol requires a sub-culturing step to ensure reliable identification. It can take days to weeks before fungal growth appears on solid medium. In this study, a unique MALDI-TOF MS protocol using liquid cultures of dermatophyte species was developed in order to shorten the turnaround time for the culture and identification of clinical isolates. Material and Method A standard MALDI-TOF MS protocol was adapted for liquid instead of solid cultures. Three different databases were tested. Results Using the liquid media MALDI-TOF MS protocol, a global rate of 62% correct identification (RCI) was obtained, compared with 87% for the protocol based on solid cultures. Trichophyton tonsurans was not correctly identified in all isolates using liquid cultures, with 88% of the isolates misidentified as Trichophyton interdigitale. The turnaround time for primary isolates for the solid and liquid protocols were respectively 11.7 and 11.6 days (no significant difference between both methods (p = 0.96)). Conclusions The newly designed liquid MALDI-TOF MS protocol did not lead to a significantly shorter turnaround time for the identification of dermatophytes isolated from tinea capitis infections. The turnaround time for the method with primary isolates was not significantly lower, and the rate of correct identification decreased remarkably, which emphasizes the need for a sub-culturing step. Using different database did not lead to improvement in turnaround time or rate of correct identification. This study highlights the importance of the medium and the reference database when performing MALDI-TOF MS.

Asymptomatic Scalp Carriage among Household Contacts of Children Affected by Tinea Capitis: A Prospective Study in the Metropolitan Area of Brussels, Belgium.

Introduction: Tinea capitis (TC) is a superficial fungal infection affecting the scalp. The existence of asymptomatic carriers (ACs) could represent a potential reservoir responsible of (re)contamination and failure of treatment. No prospective studies on ACs in household contacts of TC patients in Europe have been published to date. Objectives: The aim of this study was to assess the prevalence of ACs in a cohort of household contacts of children who were diagnosed with TC in the metropolitan area of Bruxelles, Belgium. Methods: This prospective observational study was conducted from October 2015 to April 2016 at the Dermatology Department of the University Hospitals Brugmann, Saint-Pierre, Queen Fabiola Children Hospital. Results: Ninety-nine cases of TC from 95 different family circles were included. The main infectious agent identified was Microsporum audouinii in 53 cases. The mean age of TC patients was 5.8 years. Male/female ratio was 2.8. Eighty-one household contacts of TC patients were enrolled in the study. Two cases of ACs (5%) were identified. Conclusions: M. audouinii was the most common pathogen identified. The prevalence of ACs we report is on average higher compared to other European large cities. Larger prospective studies including all close contacts of affected patients are required in order to establish guidelines regarding identification and management of ACs.

Direct examination, histopathology and fungal culture for the diagnosis of onychomycosis: A retrospective, comparative study on 2245 specimens.

BACKGROUND: Onychomycosis affects 5.5% of the general population and represents up to 50% of all nail diseases. Diagnosis and pathogen identification are essential in order to plan an adequate treatment. Many diagnostic techniques are available, and however, no solid data regarding comparison between different techniques over a large number of specimens are available to date. OBJECTIVES: To compare sensitivity and specificity of direct examination, histopathology and fungal culture in our referral mycology laboratory. METHODS: Nail specimens received at the cutaneous pathology and mycology laboratory of the University Hospital Saint-Pierre (Brussels, Belgium) between 1 January and 15 May 2018 were retrospectively analysed. All specimens were submitted to direct examination and culture. In cases of adequate specimen size, histopathology was performed. Fungal culture was considered the gold standard for diagnosis. RESULTS: A total of 2245 nail samples were included in the study. Onychomycosis was diagnosed in 1266 specimens. Sensitivity and positive predictive value were found to be higher for direct examination compared to histopathology, while sensitivity of direct examination was found to be lower. Combined approach with all the three techniques showed the highest rate of positivity, followed by the association of direct examination and histopathology. CONCLUSIONS: To our knowledge, this study included the largest number of nail specimens to date, allowing a comparison between direct examination, culture and histopathology. Direct examination showed to be the most performing technique in routine practice. Histopathology represents the most effective option in cases where both specimen size and laboratory resources are adequate. Our paper adds to the literature the 'real-life' experience of the mycology laboratory of a referral centre for nail diseases.

Superficial mycoses in Belgium: Burden, costs and antifungal drugs consumption.

BACKGROUND: Monitoring of superficial mycoses requires more attention due to their important incidence, health costs and antifungal drugs consumption. OBJECTIVES: The objectives were to estimate the burden of superficial mycoses in Belgium and to assess trends in associated antifungal consumption. METHODS: The burden of dermatophytoses (including onychomycosis), as well as skin and genital candidiasis, was estimated using disability-adjusted life years (DALY). Moreover, trends in systemic and topical antifungal consumption in ambulatory care were examined for the period 2010-2017, together with their associated costs. RESULTS: Due to their high incidence and long treatment duration, dermatophytoses represented the bulk of the burden, accounting for 92.2% of the total DALYs of superficial mycoses. Terbinafine was the most prescribed antifungal in terms of doses (35.4% of the total doses) while fluconazole was the most delivered drug in terms of packages (29.1% of the total packages). More than 70% of the prescriptions were made by general practitioners while consumption varied according to age and gender of the patients. A global 12% decrease in antifungal prescriptions was observed between 2011 and 2017. However, this reduction would result mainly from packaging changes and increased self-medication. A significant decrease in itraconazole treatments was notably compensated by an increased prescription of fluconazole packages. CONCLUSION: This study emphasises that dermatological presentations of superficial mycoses are the most important in terms of both burden and antifungal consumption in Belgium. Further reduction in antifungals use can be achieved by applying the adequate treatment after identification of the causative agent.

An international survey about nail histology processing techniques.

BACKGROUND: There are limited data on nail histopathology techniques. The objective of this study was to examine nail histopathology techniques currently in use internationally. METHODS: An online survey was sent to the European Nail Society and Council for Nail Disorders during 2015-2016. RESULTS: There were 57 respondents, from twenty countries comprising dermatologists, podiatrists and pathologists. Specimens were unmarked or marked using ink or a suture and fixed in 10% formalin, from 6 to 48 hours before embedding in paraffin wax (90% [17/19]), liquid nitrogen (frozen section, 1/19) and 2-hydroxyethylmethacrylate (plastic, 1/19). Nail softening was undertaken by 71% (17/24) of respondents for 6 to 48 hours using Mollifex Gurr (12.5%, 3/24), 10% potassium hydroxide solution (12.5%, 3/24) or 10% potassium thioglycolate cream (12.5%, 3/24). Section thickness was 4 to 9 µm (62.5%), using a steel microtome (92%,12/13) on glass slides (91.6%, 11/12). Hematoxylin and eosin (H&E) was routine for all biopsies and Periodic acid Schiff (PAS) for fungus. The favored stain for differentiating melanin and hemoglobin was Fontana-Masson (60%, 6/10). For pigmented lesions, Melan-A was always employed by all respondents (9/9). CONCLUSION: Nail histopathology processing has some small variations from normal skin processing.

Is Pica under-reported in children with sickle cell disease? A pilot study in a Belgian cohort.

BACKGROUND: For centuries, writers have recorded their observations on pica. Nevertheless the association of pica with sickle cell disease (SCD) was poorly documented. METHODS: Cross-sectional evaluation performed on SCD children and caregivers attending the outpatient clinic who were invited to complete questionnaires assessing behavior of pica. RESULTS: Out of 55 sickle cell children, 31(56.4%) reported practicing pica regularly. Substances ingested by patients covered a broad spectrum. Compared with the non-pica group, subjects who reported pica were younger and had lower hemoglobin (8.3 g/dl (7.6-9.7) vs. 9.1 g/dl (7.9-10.5): P < 0.01). The level of ferritin, zinc, copper, and lead was similar between the pica and non-pica groups (P > 0.05). Discussion In this series, there are many substances consumed by SCD children and adolescents, and we did not find an occurrence of similar substances among this select group. Pica children were younger and more anemic than non-pica patients. CONCLUSION: This study suggests that pica remains an unknown and under-reported clinical problem in children with SCD and seems to be related to the severity of anemia. The next step of this project aims to clarify causal mechanisms for pica and its association with SCD in a larger population.

[Management of onychomycosis]. / Prise en charge des onychomycoses.

Onychomycosis accounts for half of all nail pathologies. Never prescribe a local or a systemic antifungal without confirmation of the diagnosis. The quality of the sampling is the cornerstone of mycological analysis. If the results of the mycology are negative, do not hesitate to harvest a new specimen if the clinical features are highly suggestive. Otherwise, think to another cause. Identify the clinical presentations at high risk of failure to classical treatments (thick hyperkeratosis, onycholysis, lateral disease, yellow spikes, moulds). Always eradicate as much as possible of the infected keratin (mechanically, chemically or surgically). A mycologic cure means a return to a complete normal nail with a negative culture. Prevent recurrences.

Nail tumors.

Most neoplasms of the nail apparatus have different clinical appearances, courses, and biological behaviors as compared with similar tumors located elsewhere on the skin. Some of these tumors are unique to the nail, such as onychomatricoma. As a general rule, benign lesions respect the general architecture of the nail apparatus, whereas malignant ones are destructive. Our review covers the most common nail tumors, from benign ones to the most frequent nail malignancy, the squamous cell carcinoma, which actually is the greatest simulator. We will also discuss new approaches to the diagnosis and treatment of melanoma of the nail apparatus. Physicians should be aware of these conditions and their management.

A retrospective study of squamous cell carcinoma of the nail unit diagnosed in a Belgian general hospital over a 15-year period.

BACKGROUND: Squamous cell carcinoma (SCC) is the most common malignant tumor at the nail unit. It mainly affects middle-aged men, with a peak incidence between 50 and 69 years of age. Diagnosis is often delayed because of the slow evolution of the lesion and multiple clinical features. OBJECTIVE: We sought to characterize the different clinical and histopathological patterns of SCC of the nail unit and evaluate their therapeutic outcome. METHODS: Records for 58 patients were retrieved from our department's dermatopathology database over a period of 15 years (1995-2011) and the patients recontacted. RESULTS: Of the 58 patients, 51 were eligible for follow-up. There was a male predominance (72.5%). The fingers were most commonly affected (98%), the right index and long fingers being most commonly affected (20.8% each). The nail bed was mainly affected. The commonest clinical signs were, in decreasing order, subungual hyperkeratosis, onycholysis, oozing, and nail plate destruction. The majority of SCC of the nail unit was in situ (63%). The recurrence rate of all treatments taken together was 30.6%. LIMITATIONS: Retrospective study design is a limitation. CONCLUSIONS: SCC of the nail unit mostly affects men aged 50 to 69 years. Most cases were the warty type, with oozing being an underrecognized clinical sign. Contrary to prior studies, most lesions were in situ, and bone involvement was uncommon. Conservative surgical resection should be the first-line treatment when the bone is not involved. Recurrence rate is high when a procedure other than Mohs micrographic surgery is performed.