Evaluation of Gradient Concentration Strips for Detection of Terbinafine Resistance in Trichophyton spp.

The number of dermatophytosis cases resistant to terbinafine is increasing all over the world. Therefore, there is a need for antifungal susceptibility testing of dermatophytes for better management of the patients. In the present study, we have evaluated a gradient test (GT) method for testing the susceptibility of dermatophytes to terbinafine. MIC values to terbinafine determined by the EUCAST reference technique and by gradient test were compared for 79 Trichophyton spp. isolates. Overall, MICs were lower with gradient test (MIC50 of 0.002 µg/mL) than with EUCAST (MIC50 of 0.016 µg/mL). Good categorical agreement (>90%) between the 2 techniques was obtained but the essential agreement was variable depending on the batch of gradient test.

Reliability of a terbinafine agar containing method for the screening of dermatophyte resistance.

The increase in terbinafine resistance worldwide due to Trichophyton indotineae underlies the need for surveillance networks, deploying easy to perform methods to correctly identify resistant isolates and thereby reduce their spread. In the present study, we evaluated the performances of the terbinafine containing agar method (TCAM). Different technical parameters, such as culture medium (RPMI agar [RPMIA] or Sabouraud dextrose agar [SDA]) and inoculum size, were evaluated. Our study showed that terbinafine susceptibility determined using the TCAM was reliable and independent of the inoculum or medium used. We then performed a multicenter, blinded study. 5 isolates of T. indotineae and 15 of genotype I or II of T. interdigitale, including 5 terbinafine-resistant isolates (4 T. indotineae and 1 T. interdigitale), were sent to eight clinical microbiology laboratories. Each laboratory analyzed the 20 isolates' terbinafine susceptibility by the TCAM using both culture media. TCAM allowed all participants to correctly determine the terbinafine susceptibility of analyzed isolates without prior training. All participants agreed that the dermatophyte tested, regardless of species or genotype, grew better on SDA than on RPMIA medium but accumulated fungal growth after 14 days eventually minimized the effect of this difference. In conclusion, TCAM is a reliable, easy to perform screening method for assessing terbinafine resistance. However, despite good performances, TCAM is a qualitative method and minimal inhibitory concentrations must be determined by the European Committee for Antimicrobial Susceptibility Testing standardized method to follow the evolution of terbinafine resistance levels.

The increase in terbinafine resistance worldwide due to Trichophyton indotineae underlies the need for surveillance networks. The terbinafine containing agar method is a reliable and easy-to-use tool in clinical microbiology laboratories. It can be used to rapidly screening resistant isolates, thereby reducing their spread.

MALDI-TOF Mass Spectrometry Online Identification of Trichophyton indotineae Using the MSI-2 Application.

Trichophyton indotineae is an emerging pathogen which recently spread from India to Europe and that is more prone than other species of the Trichophyton mentagrophytes complex to show resistance to terbinafine, resulting in the necessity of rapid identification. Here, we improved the online MSI-2 MALDI-TOF identification tool in order to identify T. indotineae. By multiplying the culture conditions (2 culture media and 6 stages of growth) prior to protein extractions for both test isolates and reference strains, we added 142 references corresponding to 12 strains inside the T. mentagrophytes complex in the online MSI-2 database, of which 3 are T. indotineae strains. The resulting database was tested with 1566 spectra of 67 isolates from the T. mentagrophytes complex, including 16 T. indotineae isolates. Using the newly improved MSI-2 database, we increased the identification rate of T. indotineae from 5% to 96%, with a sensitivity of 99.6%. We also identified specific peaks (6834/6845 daltons and 10,634/10,680 daltons) allowing for the distinction of T. indotineae from the other species of the complex. Our improved version of the MSI-2 application allows for the identification of T. indotineae. This will improve the epidemiological knowledge of the spread of this species throughout the world and will help to improve patient care.

Terbinafine Resistance in Dermatophytes: A French Multicenter Prospective Study.

In recent years, we have moved from the sporadic description of terbinafine-resistant (TerR) Trichophyton spp. isolates to the Indian outbreak due to T. indotineae. Population flows have spread TerR worldwide, altering local epidemiology. We conducted a prospective multicentric study to determine the relative frequency of TerR isolates in France (Paris area) and of the newly introduced T. indotineae species. TerR isolates were screened by the terbinafine-containing-agar-medium (TCAM) method and confirmed by EUCAST. Sequencing methods were used to identify isolates to the species/genotype level and to analyze substitutions in the squalene epoxidase gene (SQLE). In total, 3 isolates out of 580 (T. rubrumn = 1; T. interdigitalen = 1; T. indotineaen = 1) grew on TCAM, showed terbinafine resistance by EUCAST and harbored the Phe397Leu (n = 2) or Leu393Ser (n = 1) substitution in the SQLE. ITS-sequencing of isolates of the T. mentagrophytes/interdigitale complex (n = 125) revealed a relative frequency of 4.8% for T. indotineae and the presence of T. mentagrophytes genotype VII. Despite the detection of terbinafine resistance, isolates from this complex remained susceptible to itraconazole, voriconazole and amorolfine. Terbinafine resistance is present in France and the dermatophyte epidemiology is changing. Efficient systems must be implemented to survey the evolution of newly introduced species and to identify TerR isolates.

Trichophyton bullosum: a new zoonotic dermatophyte species.

We report the first human case of dermatophytosis caused by Trichophyton bullosum in a 21-year-old male who had a skin lesion located on his forearm. The dermatophyte was isolated in culture and further identified by sequence analysis of internal transcripted spacer regions. The species T. bullosum is a zoophilic dermatophyte rarely isolated from the coat of horses in Africa and Asia. In the present case, it was probably transmitted by contact with an infected donkey in a rural area in France. Antifungal therapy led to remission of the lesion in the patient after 2 months of treatment. T. bullosum ITS region sequences were closely related to those of the African species of Arthroderma benhamiae and grouped in a zoophilic cluster with Trichophyton verrucosum, T. erinacei and the Trichophyton anamorph of A. benhamiae (zoophilic species of the T. mentagrophytes complex). Systematic molecular identification could contribute to an accurate identification of this unusual species.

[Epidemiology of tinea capitis. Five-year retrospective study in three hospitals in the Val de Marne]. / Epidémiologie des teignes du cuir chevelu. Etude rétrospective sur 5 ans dans 3 centres hospitaliers du Val-de-Marne.

OBJECTIVE: Describe the epidemiology of Tinea capitis in three hospitals in the Val de Marne (suburban district southeast of Paris) and evaluate the usefulness of screening the families of index cases. METHODS: The study included 3 hospitals: Henri Mondor Hospital, Creteil Intermunicipal Hospital Center and Bicêtre Hospital. Index patients had a positive culture for dermatophytes during visits to the mycology-dermatology or pediatric clinics from January 1998 through December 2002. The diagnostic procedure was identical in all centers. In two centers, scalp samples were taken routinely from family members, even in the absence of clinical lesions. Species were identified according to the phenotypic features of the cultures. We recorded the age, sex, and geographic origin of each patient, when available. RESULTS: Samples were obtained from 487 outpatients (including family members): 383 were positive (356 children under 16 years, 27 adults). Of the children with positive cultures, 214 were boys (60%); of the adults, 18 (66%) were women. Distribution of geographic origin was: sub-Saharan Africa (71%), Europe (10%), Caribbean (5%) and North Africa (4,7%). Species identification confirmed the predominance of anthropophilic species, with 46% of the isolates Trichophyton soudanense and 33% Microsporum langeronii. T. violaceum was reported in 5.5% of cases and T. tonsurans in 2.8%. M. canis (9%), found in 34 Europeans, was the main zoophilic species. Family screening accounted for 263 of the 487 outpatients. Of 242 children from 86 families, 153 (63%) had positive cultures. Of the 21 parents, 5 had a positive culture. Globally, the family screening identified 158 cases in 263 tests (60%). CONCLUSION: Tinea capitis in the Val de Marne is mainly due to anthropophilic species and involves children of African origin. The frequency of family contamination indicates that routine screening of family members of infected children is valuable.