We have shown that dermatophyte species can be easily identified on the basis of a DNA sequence encoding a part of the large-subunit (LSU) rRNA (28S rRNA) by using the MicroSeq D2 LSU rRNA Fungal Sequencing Kit. Two taxa causing distinct dermatophytoses were clearly distinguished among isolates of the Trichophyton mentagrophytes species complex.Dermatophytes are the main cause of superficial mycoses (9,15,16). These fungi have the capacity to invade keratinized tissue of humans or animals to produce infections that are generally restricted to the corneocytes of the skin, hair, and nails. Among the approximately 10 species isolated in Europe, Trichophyton rubrum and T. mentagrophytes are the most commonly observed, with frequencies varying from 27 to 74% and from 17 to 41%, respectively (13).Dermatophytes are usually identified on the basis of macroscopic appearance, together with microscopic examination of cultures. Important characteristics are the rate of growth, the shape and texture of the culture on solid media, color, diffusion of pigments into agar, and sporulation. However, identification of dermatophytes often remains difficult or uncertain because there are variations from one isolate to another. Recent advances in molecular biology and progress in technology have allowed the development of new techniques for species determination and strain typing in microbiology. The molecular approach used to identify fungi is often based on sequence analysis of the ribosomal DNA (rDNA) and in particular on the internal transcribed spacer (ITS). The polymorphism of the ITS1 and ITS2 regions flanking the DNA sequence encoding the 5.8S rRNA was previously shown to be suitable for the identification of clinically important yeasts (1), Aspergillus sp. (8),and dermatophyte species (3-5). In contrast, the gene coding for the small-subunit rRNA (18S rRNA) did not discriminate sufficiently between dermatophyte species (7).The MicroSeq D2 large-subunit (LSU) rRNA Fungal Sequencing Kit (Applied Biosystems, Rotkreuz, Switzerland) was recently developed to identify fungal species after amplification of a partial sequence of the DNA encoding the LSU rRNA (28S rDNA). The sequence of a given fungus can then be compared for identification with the rDNA sequences of the MicroSeq D2 Fungal database, which includes more than 500 validated sequences from different fungal species but not from dermatophytes. In the present study, we tested the MicroSeq D2 LSU rRNA Fungal Sequencing Kit by using it to identify dermatophyte species from patients referred to the mycological laboratory of the Department of Dermatology at the University Hospital in Lausanne, Switzerland (Table 1). Neotypes of different species and other reference strains (Table 2) were used for comparison of DNA sequences. This study allows the extension of the MicroSeq D2 Fungal database to the determination of dermatophytes.Isolates. Skin and nail scrapings and hair fragments were collected from patients with suspected mycoses. Routinely, one part of each sample was examine...

Background: Dermatophytes are usually identified on the basis of macroscopic characteristics and microscopic examination of the cultures. Identification of dermatophytes often remains difficult or uncertain because there are variations from one isolate to another and overlapping characteristics between species. Objective: To identify dermatophyte species producing numerous microconidia and resembling Trichophyton mentagrophytes by DNA sequence analysis. Methods: The complete ITS1 + 5.6s + ITS2 rDNA region of various dermatophytes isolated in culture was amplified by PCR and sequenced. Results: Nine isolates of a fast-growing dermatophyte species were identified as Arthroderma benhamiae by DNA sequencing. Retrospective investigations revealed that the isolates were from 8 children and 1 adult suffering from inflammatory dermatophytosis. Eight of the 9 patients had had previous contact with rodents, mostly guinea pigs. Conclusion: It is the first time that A. benhamiae is reported in Switzerland. In cases of dermatophytosis attributed to A. benhamiae, a rodent is the most likely cause of infection.

Background:Fusarium species are isolated from about 3% of onychomycoses in the Swiss native population. On the basis of macroscopic characters and microscopic examination of the cultures, identification of Fusarium often remains difficult or uncertain because of variations from one isolate to another and overlapping characteristics between species. Objective: To obtain information about the prevailing species of Fusarium collected from onychomycoses. Methods: An analysis of the Fusarium specimens isolated in the Department of Dermatology at the University Hospital of Lausanne was conducted during a 2-year period (71 isolates). A 311-bp fragment of the gene encoding 28S rRNA was amplified by PCR and sequenced. DNA sequences were compared to those available for reference strains. Results:Fusarium oxysporum was the most frequently isolated species, accounting for 54% of the isolates. F. proliferatum and 4 taxons belonging to the F. solani species complex were identified with an appreciable frequency ranging from 4 to 14%. Conclusion: The Fusarium species identified were the same as those known to cause disseminated fusariosis in immunocompromised patients. The presence of these Fusarium species in onychomycoses warrants that careful attention should be paid to abnormal nails before beginning immunosuppressive treatments in patients.

A new, slow-growing, scotochromogenic mycobacterium was isolated from a lymph node of an immunocompromised child and subsequently from tap water and from a respiratory specimen of a patient with chronic fibrosis. Alcohol-acid-fastness, lipid patterns and the G+C content clearly support the placement of this organism in the genus Mycobacterium. The isolates grew very slowly at temperatures ranging from 25 to 32 "C and showed activities of nitrate reductase, catalase, urease, arylsulfatase and Tween 80 hydrolysis. The organism was susceptible to all antimycobacterial drugs tested. The 16s rDNA sequence was unique and phylogenetic analysis placed the organism close to fast -g ro w i n g species such as Mycobacterium farcinogenes, Mycoba cterium komossense and Mycobacterium aichiense. These data support the conclusion that the isolates represent a new mycobacterial species, for which the name Mycobacterium tusciae sp. nov. is proposed. The type strain is strain FI-25796'; a culture of this strain has been deposited in the DSMZ as strain DSM 44338'.

Between August 1994 and September 1996, 28 glycopeptide-resistant enterococci (GRE) were isolated from 8 infected patients and 11 intestinal carriers hospitalized at the University Hospital of Geneva. Identification to the species was made by both phenotypic (API 20 STREP and Rapid ID 32 STREP systems, and Vitek Gram Positive Identification Card) and genotypic methods using a multiplex PCR assay developed also for the determination of the genotype of glycopeptide resistance (vanA, vanB, vanC1, andvanC2-C3 genes). Fifteen isolates were identified asEnterococcus faecium, 8 as E. gallinarum, 4 asE. faecalis, and 1 as E. hirae. All of the phenotypic identification methods failed to differentiate some isolates of E. gallinarum from E. faecium, or vice versa. Both vanA (n = 18) andvanB (n = 4) glycopeptide resistance genotypes were found. For the first time, the vanBdeterminant was found in two isolates of E. gallinarum. Two patients were colonized by two different species containing thevanA gene and one by two different species containing thevanB gene. All vanA isolates were highly resistant to both vancomycin and teicoplanin except for three isolates which were susceptible to teicoplanin. Molecular typing by pulsed-field gel electrophoresis showed identical or similar patterns among E. faecium isolates with thevanA gene in five patients for whom the epidemiological link could not be always elucidated. This study emphasizes the necessity of utilizing both phenotypic and genotypic methods to characterize GRE.