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See detailA three year survey of dermatophytoses in Belgium
SACHELI, Rosalie ULg; DARFOUF, Rajae ULg; GRAIDE, Hélène ULg et al

in Mycoses (2015, October), 58(Supplement 4), 135

Objectives Dermatophytosis refers to superficial fungal infections of keratinized tissues caused by keratinophilic dermatophytes. They are the most common cause of superficial fungal infections worldwide ... [more ▼]

Objectives Dermatophytosis refers to superficial fungal infections of keratinized tissues caused by keratinophilic dermatophytes. They are the most common cause of superficial fungal infections worldwide. Epidemiological studies regarding dermatophyte infections have been conducted in several countries and differences in the incidence and in etiological agents have been reported for different geographical areas. That is why national surveillance of circulating strains causing dermatophytosis is crucial. The Belgian National Reference Center (NRC) for Mycoses conducted a survey on dermatophytes strains circulating from 2012 to 2014. The present study was performed to assess the profile of dermatophytosis and to identify the species involved. Methods The Belgian NRC for Mycosis collected 9138 strains between January 2012 and December 2014. The isolates were cultured from patients clinically suspected for fungal infections of skin, hair and nails. Isolates were sent by Belgian laboratories to the two labs of the Belgian NRC (UZ Leuven and CHU of Liège) in order to identify the fungus or to confirm the identification. All isolates cultured from patients of UZ Leuven and CHU of Liège were also included. Fungal identification was performed by microscopy after subculture and in case of doubtful identifications by ITS sequencing. Results .Among the 9138 samples (results of UZ Leuven and CHU of Liège combined), 3587 were identified as dermatophytes. Trichophyton. rubrum (T. rubrum) was the most prevalent species accounting for 56,17% (n=2015) of the infections from all sources, followed by T. mentagrophytes complex (21,83%, n=783). The other main etiological agents of dermatophytosis recorded in this study in descending order of prevalence were M. audouinii (n=303), M. canis (n=120), T. violaceum (n=112), T. tonsurans (n= 95), T. soudanense (n=66), M. praecox (n=59), E. floccosum (n=14) Our data reveal the predominance of anthropophilic species causing tinea capitis especially M. audouinii responsible for 36,49% (n=163/448) of hair/scalp infection. Trichophyton violaceum rarely observed in our country is frequently found as 12,8% (n=57) of the reported cases of tinea capitis are due to this species. The retrospective evaluation of data collected also shows that zoophilic strains as M. canis well represented in the past epidemiology of tinea capitis, is decreasing in frequency accounting for only 7,2% (n=32) of clinical cases. Finally, our data confirm the high prevalence of T. rubrum commonly observed in Europe as causal agent of onychomycosis (70,9%, n=1603) followed by T. mentagrophytes complex (20,9%, n=455). T. rubrum and T. mentagrophytes complex are also responsible for the majority of skin infections as they represent respectively 40% (n=386) and 24,75% (n=239) of skin dermatophytosis during the study period. Conclusions The present work has provided recent data on the prevalence of several dermatophytes species circulating in Belgium. Such data is critical for the establishment of therapeutic strategies and measures for prevention and control of dermatophytes infections. Our study confirms the predominance of T. rubrum followed by T. mentagrophytes in the Belgian population but also highlights the emergence of new anthropophilic species such as M. audouinii and T. violaceum as causative agents of tinea capitis in children in relation with African immigration. [less ▲]

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See detailEpidemiological aspects and genotypic characterization of T.violaceum strains collected during a Belgian National survey on anthropophilic tinea
SACHELI, Rosalie ULg; Dekkers, Charlotte; GRAIDE, Hélène ULg et al

in Mycoses (2015, October), 58(Supplement 4), 189

Objectives The last two years, clinical cases of tinea capitis caused by Trichophyton violaceum (T. violaceum), have been identified in Belgium. To better understand the emergence of this species in the ... [more ▼]

Objectives The last two years, clinical cases of tinea capitis caused by Trichophyton violaceum (T. violaceum), have been identified in Belgium. To better understand the emergence of this species in the population, the Belgian National Reference Center (NRC Liège) launched a one-year national survey in 2013. Epidemiological aspects and genotypic characterization of the strains were included. Methods The study was conducted from March 2013 up to February 2014. All Belgian laboratories were asked to send M. audouinii and T. violaceum strains isolated from hair to the NRC with a form to fill in including epidemiological data. The fungal strains were identified by microscopy or ITS sequencing in case of doubtful identification. The genotypic analysis was performed by the DiversiLab® system (bioMérieux) for DNA fingerprinting and analysis. Epidemiological data were analyzed with the help of a biostatistician. Results Amongst the collected isolates, 23 strains were confirmed as T.violaceum (results concerning the 116 M. audouinii strains have already been reported). Analysis of the epidemiological characteristics of the infected population shows that the main age category concerns 0-4 year-old children (n=9, 39,1%) with a sex-ratio M/F of 1.875. Data concerning the geographic origin of the family were present in 82,6% of the cases and reveal that patients were mainly of Ethiopian origin (n=8, 57,9% of known cases). One patient was also from Burundi showing that T. violaceum strains probably circulate mainly in East Africa. The genotypic analysis led to the distinction of 2 variants of T. violaceum. The major group was composed of 17 strains which were mainly collected in the North of Belgium and included also the reference strain (18/23, 83,3%). The other group (6 strains) was close to the major group but the analysis of the spectral superposition showed some differences between these two groups, defining two distinct variants of T. violaceum in the Belgian population. This second variant was mainly recovered from South Belgium (5/6, 83,3%). No correlation could be made between the genotypic group and a particular ethnical origin as Ethiopian subjects were found in both groups. Conclusion The DiversiLab® system proved to be an efficient method to investigate the molecular epidemiology of dermatophytes infections as reported previously for M. audouinii. These results show that two distinct isolates co-exist in Belgium providing evidence of genetic heterogeneity and a possible spread of one genotypic variant in a restricted geographic area or the co-existence of two variants circulating in different African communities. However, no clear correlation could be established between the appartenance to a group and epidemiological factors, such as age or ethnical origin. [less ▲]

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See detailOnychomycosis: is it possible to increase the cure rate?
Hayette, Marie-Pierre ULg

in Mycoses (2015, October), 58(suppl 4), 35

Onychomycosis represent about 50% of nails disorders in the world with a very variable prevalence depending of the countries considered. Dermatophytes, non-dermatophyte molds and Candida sp. are the main ... [more ▼]

Onychomycosis represent about 50% of nails disorders in the world with a very variable prevalence depending of the countries considered. Dermatophytes, non-dermatophyte molds and Candida sp. are the main causing agents. A rapid and accurate diagnosis is essential in order to give an adequate treatment to the patient. Generally, a combination of microscopy and culture is used for laboratory diagnosis. However, microscopy does not always allow the distinction between yeasts or filamentous fungi, culture takes generally about a week before identification and the result is compromised if there is contamination by not relevant fungi. Therefore commercially available PCR-based methods have been developed in order to provide a rapid and accurate identification of dermatophytes and yeasts in nails samples. Combination of microscopy and PCR may provide a rapid and specific diagnosis in 2 working days. However this methodology is still not widely used by laboratories because of the high cost. Furthermore, this technology can detect DNA from dead fungi and therefore is not suitable for assessment of treatment efficacy. Onychomycosis therapy depends on different factors such as the causative agent, the number of nails and degree of nail involvement, the type of onychomycosis, potential drug interactions or drug intolerance and a failure to previous treatments. Oral and topical antifungals are mostly used separately or in combination. Oral therapy includes azoles (itraconazole, fluconazole) and/or allylamine (terbinafin), this latter being the most frequently prescribed antifungal for treatment of onychomycosis in North America and Europe. Topical amorolfine and ciclopirox formulations can be used alone in mild cases or in case of intolerance to oral antifungals. However, one of the biggest problems of therapy for onychomycosis is the high frequency of relapse which concerns about 20 to 40% of the patients treated by oral antifungals. Different strategies have been developed to overcome this problem amongst which are: optimization of the dosing regimens (continuous vs pulse therapy) or therapy duration, combination therapy (nail debridement + antifungals, oral + topical drugs, 2 oral drugs), improving drug delivery (use of physical or chemical enhancers, and modification of the pharmacological formulation for increasing drug uptake). Some strategies such as combination therapy (oral + topical) have demonstrated enhanced efficacy and should be recommended in case of poor efficacy of the initial treatment or in case of extended infection. Prophylactic topical therapy implemented after completion of oral treatment has been shown to delay relapse. Preventive measures such as treatment of concurrent tinea pedis and/or infected family members and regular cleaning of bathroom and shower floors can help to reduce the risk of reinfection particularly when a dermatophyte is the causative agent. In conclusion, treatment for onychomycosis is associated with frequent relapse. Consequently, follow-up is mandatory and combination therapy can be necessary in case of relapse or resistance to treatment. Patients should also be aware of the preventive hygiene measures to apply in order to decrease the risk of reinfection. New strategies improving treatment efficacy are promising but their efficacy have still to be demonstrated in comparative clinical trials before their implementation in therapy. [less ▲]

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See detailValidation of the DermaGenius Nail plus multiplex assay, a new commercial PCR assay developed for the detection and identification of dermatophyte and Candida in nails
Hayette, Marie-Pierre ULg; GRAIDE, Hélène ULg; ADJETEY BAHUN, Akolé ULg et al

in Mycoses (2015, October), 58(suppl 4), 223

Objectives Superficial dermatophytosis is the most common fungal infection in humans. Diagnosis of dermatophytosis is currently based on microscopy or histology associated with culture on specific agar ... [more ▼]

Objectives Superficial dermatophytosis is the most common fungal infection in humans. Diagnosis of dermatophytosis is currently based on microscopy or histology associated with culture on specific agar media. However, direct microscopy lacks specificity and culturing has a long turn-around-time of 2-4 weeks. These limitations can be prevented by the use of molecular diagnostics. The DermaGenius (DG) multiplex kit (PathoNostics, The Netherlands) is a new commercial realtime-PCR kit, which can differentiate various dermatophytes species including the nail pathogens T. rubrum, T. mentagrophytes, T. interdigitale and 2 Candida species (C. albicans and C. parapsilosis). This study aimed at the validation of the kit on nails clippings. Results were compared with histology and culture data. Methods A set of 76 nail clippings was collected from 76 patients attending the dermatology consultation at the University Hospital of Liege on suspicion of onychomycosis. All nails were divided in three pieces for histology, culture and the PCR multiplex assay. Histologic preparations were stained with PAS staining. Cultures were performed on 2 different Sabouraud agar medium slants (bioMérieux). The DNA extraction protocol used a proteinase K pre-treatment followed by an automated DNA-extraction (EasyMag, bioMérieux). An Internal Control (IC) was included to monitor for PCR inhibition or manual errors. The realtime-PCR amplification was performed with the DG kit on a Rotor-Gene Q instrument (Qiagen) by using quantitative amplification and melting curve analysis. Results In total, 35 of 76 cases (46%) were classified as confirmed onychomycosis based on positive microscopy (M+) with or without positive culture (C+) or just by positive culture of a confirmed pathogen. Based on negative microscopy (M-) and negative culture of a confirmed pathogen, 41 cases (54%) were reported as non-fungal onychodystrophy. Agreement between DermaGenius (DG) and culture was found in 52% of the cases while 86% agreement was reported when comparing positive DG with confirmed onychomycosis. Three positive cultures (microscopy negative) were not detected by DG (2 T. rubrum, 1 C. albicans). However, DG could detect 7 additional infections (9%). Eleven discrepancies DG+/C+ were determined which could be positively confirmed in favour of DG result by ITS sequence analysis. Most discrepancies could be explained by fungal/yeast species overgrowing the agar slant, including species of Candida, Fusarium, Trichosporon, which were not considered as the source of infection. Conclusion The DermaGenius Nail plus multiplex was able to detect the most prevalent pathogenic dermatophytes species in clinical nail specimens and proved to be more sensitive and specific than culture and direct microscopy. The DNA extraction procedure has been shown to work efficiently in diagnostics which enables the physician in charge of the patient to start a dedicated treatment rapidly. [less ▲]

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See detailDNA fingerprinting using Diversilab system for genotyping characterization of Microsporum audouinii and Trichophyton violaceum
SACHELI, Rosalie ULg; DIMO, Lauryl; GRAIDE, Hélène ULg et al

in Mycoses (2013, October 01), 56(Supplement S3), 99

Objectives: To investigate the epidemiological determinants responsible for the high number of anthropophilic dermatophytes received by the National Reference Center for Mycosis of Liege (NRCL) during the ... [more ▼]

Objectives: To investigate the epidemiological determinants responsible for the high number of anthropophilic dermatophytes received by the National Reference Center for Mycosis of Liege (NRCL) during the year 2012. To perform a genotypic characterization by the Diversilab® system focusing on the two main isolated species, Microsporum audouinii and Trichophyton violaceum. To present a preliminary study preceding the national survey launched in 2013. Methods: A total of 51 strains of M. audouinii (50 clinical + 1 reference (ref.) strains) and 15 strains of T. violaceum (14 clinical + 1 ref. strain) originating from different locations through Belgium were included in the study. The fungal strains were first cultivated on Malt agar, then sub-cultured in Sabouraud liquid medium (Fluka). The grown mycelium was processed for DNA extraction following recommendations of the manufacturer (Ultra Clean® DNA Microbial isolation kit, MoBio laboratories). Genotypic analysis was performed using the DiversiLab® system (BioMérieux) for DNA fingerprinting and analysis. Results: Regarding M. audouinii, four different genotypic groups of strains were separated. Group 1 includes 11 strains and is only found in the Liège surroundings. Group 2 includes only one strain with little differences compared to group 1 and collected from the Liège area. These two groups may be related to each other. Group 3 contains 36 strains and the reference strain. This genotype is distributed in different Belgium locations. The last group, group 4, contains only 3 isolates sharing low similarities in comparison with the 3 other groups. Concerning T. violaceum, 6 different genotypic groups with a mixed geographical distribution were determined. Group 1 includes 8 clinical isolates and the ref. strain. The other five isolates are all different and seem not to be related to each other. Conclusion: The automated typing DiversiLab® system proved to be an easy and efficient method to investigate the molecular epidemiology of dermatophytes infections. Preliminary results of the study show that, through Belgium, several groups of isolates co-exist for M. audouinii and T. violaceum providing evidence of genetic heterogeneity. This variation can be related to acquired mutations due to environmental adaptation. Further investigations are necessary to better understand the impact of this genotypic variation. [less ▲]

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See detailMechanisms of skin adherence and invasion by dermatophytes
Baldo, Aline ULg; Monod, M.; Mathy, Anne ULg et al

in Mycoses (2012), 55(3), 218-223

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See detailVentriculitis caused by Aspergillus fumigatus in a child with central nervous system tuberculosis
Antachopoulos, Charalampos; STERGIOPOULOU, Theodouli ULg; Simitsopoulou, Maria et al

in Mycoses (2011), 54(5), 627-30

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See detailCharacterization of a new potential virulence factor of Microsporum canis, the secreted subtilisin Sub6
Mathy, Anne ULg; Baldo, Aline ULg; Salamin, K. et al

in Mycoses (2011), 54(suppl 2), 112-113

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See detailThe nail under fungal siege in patients with type II diabetes mellitus.
Pierard, Gérald ULg; Pierard, Claudine ULg

in Mycoses (2005), 48(5), 339-42

Few studies have examined the prevalence of onychomycosis among diabetic patients. Given the morbidity linked to onychomycosis, and the ever-growing size of the diabetic population, a better recognition ... [more ▼]

Few studies have examined the prevalence of onychomycosis among diabetic patients. Given the morbidity linked to onychomycosis, and the ever-growing size of the diabetic population, a better recognition of this nail infection is welcome. To revisit the relative prevalence of dermatophyte, yeast and non-dermatophytic mould onychomycoses in diabetic adults in a prospective study using combined histomycology and cultures. Toenail clippings were collected for 3 years in 190 type II diabetic patients (136 men and 54 women) and from an age- and gender-matched group of non-diabetic subjects. All sampled nails showed clinical alterations reminiscent of onychomycosis. Histomycology and cultures were performed on each sample to distinguish onychomycosis from non-infectious onychodystrophy. Compared to non-diabetic subjects with nail alterations, diabetics showed a higher proportion of onychomycosis relative to non-fungal onychodystrophy. Diabetic men suffered more frequently from onychomycosis and onychodystrophy than diabetic women. When considering the nature of the fungal pathogens, dermatophytes predominated largely over yeast and non-dermatophytic moulds, both in diabetic and non-diabetic patients. Diabetic patients, particularly men, are at increased risk of developing onychomycosis. The morbidity linked to this disorder, and its impact on the foot status in diabetic subjects merit to be better appreciated by clinicians. [less ▲]

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See detailNew insights into the effect of amorolfine nail lacquer.
Flagothier, Caroline ULg; Pierard, Claudine ULg; Pierard, Gérald ULg

in Mycoses (2005), 48(2), 91-4

Despite improvements in antifungal strategies, the outcome of treating onychomycoses often remains uncertain. Several factors account for treatment failure, of which the pharmacokinetics and ... [more ▼]

Despite improvements in antifungal strategies, the outcome of treating onychomycoses often remains uncertain. Several factors account for treatment failure, of which the pharmacokinetics and pharmacodynamics of the antifungal are of importance. The taxonomic nature and ungual location of the fungus cannot be neglected, besides the type of nail and its growth rate. In addition, the biological cycle of the fungus and the metabolic activity of the pathogen likely play a marked influence in drug response. The presence of natural antimicrobial peptides in the nail is also probably a key feature controlling the cure rates. There are many outstanding publications that cover the full spectrum of the field. The purpose of this review is to put in perspective some facets of activity of the topical treatment using amorolfine nail laquer. The antifungal activity of the drug is likely less pronounced in onychomycosis than that expected from conventional in vitro studies. However, the nail laquer formulation should reduce the propensity to form antifungal-resistant spores and limit the risk of reinfection. [less ▲]

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See detailItraconazole corneofungimetry bioassay on Malassezia species.
Pierard, Gérald ULg; Arrese Estrada, Jorge ULg; Pierard, Claudine ULg

in Mycoses (2004), 47(9-10), 418-21

Yeasts of the genus Malassezia are part of the normal skin biocenosis and are involved in a series of distinct skin disorders and specific dermatomycoses in man and animals. Several species are currently ... [more ▼]

Yeasts of the genus Malassezia are part of the normal skin biocenosis and are involved in a series of distinct skin disorders and specific dermatomycoses in man and animals. Several species are currently distinguished. Their relative in vitro susceptibility to antifungals appears different according to the species and to the nature and route of administration of the drug. Corneofungimetry is an ex vivo bioassay allowing to test the fungal response on human stratum corneum following oral intake of a given antifungal by volunteers. Two series of cyanoacrylate skin surface strippings (CSSS) were harvested from the volar forearm of 30 volunteers before and after a 2-week treatment with itraconazole 200 mg daily. They were coated by olive oil and inoculated with suspensions of seven different Malassezia spp. After a 1-week culture on CSSS, the amount of viable yeasts was assessed using neutral red staining assisted by computerized image analysis. Growth of the seven species was not similar on the CSSS from untreated stratum corneum. The ranking order from the most proliferative to the least was M. restricta, M. sympodialis, M. globosa, M. furfur, M. obtusa, M. slooffiae and M. pachydermatis. Their growth was abated to almost the same level after itraconazole treatment. It is concluded that in vivo treatment with itraconazole is highly active against all Malassezia spp. colonizing the human stratum corneum. [less ▲]

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