High-frequency intragenomic heterogeneity of the ribosomal DNA intergenic spacer region in Trichophyton violaceum

The intergenic spacer (IGS) of the rRNA genes was analyzed from the dermatophyte Trichophyton violaceum isolated from cases of tinea capitis in Taiwan and Iran. T. violaceum strains were cultured from different colonies, from single conidial colonies derived by dilution plating, and from micromanipulation of single conidia from clinical samples. A ribosomal DNA probe hybridizing to multiple EcoRI fragments was used to compare restriction fragment length polymorphisms in different T. violaceum isolates. The arthroconidia of T. violaceum that form in vivo during infection were shown to contain a single nucleus by 4',6'-diamidino-2-phenylindole staining. IGS regions from an isolate cultured from a single conidium were amplified, cloned, and sequenced. The results identified that heterogeneity exists between IGS regions within a single T. violaceum genome due to different copy numbers of a 171-bp tandem repeat. This suggests that the IGS of T. violaceum is partially excluded from the concerted evolution of the rRNA gene locus. The heterogeneous character of the IGS regions in T. violaceum contrasts with the closely related dermatophyte Trichophyton rubrum, posing further questions on the phylogeny and the evolution of dermatophyte fungi.     

Direct Species Identification of Common Pathogenic Dermatophyte Fungi in Clinical Specimens by Semi-nested PCR and Restriction Fragment Length Polymorphism

OBJECTIVE: To seek a rapid and reliable molecular biology method to identify the common pathogenic dermatophyte fungi from clinical samples. METHOD: The genome DNA was extracted from cultured strains of seven common dermatophyte fungi species and part of each positive clinical specimen by microscopy. Intergenic spacer regions of ribosomal DNA (ITS) were amplified by semi-nested PCR (snPCR) with three universal primers (NS5, ITS1, and ITS4) for fungi. The amplified products were digested with two restriction endonucleases (BciT130 I, Dde I), the Restriction Fragment Length Polymorphism(RFLP). The rest of each clinical specimen was cultured in Sabouraud's Agar medium. Then the results of RFLP were compared with the traditional culture results. RESULTS: The digestion of seven common dermatophyte fungi produced seven different restriction profiles. Restriction profiles of 17 clinical specimens matched, respectively, to that of the cultured strains, and 14 profiles of the 17 ones matched the culture result completely. The coincidence was 100.0%. CONCLUSIONS: snPCR-RFLP analysis of intergenic spacer regions of ribosomal DNA is a valuable method of exactness and clarity for species identification of common dermatophyte fungi from clinical specimens.     

Survey of haircoat fungal flora for the presence of dermatophytes in a population of white-tailed deer (Odocoileus virginianus) in Virginia

Before releasing rehabilitated wildlife, patients should be cured of all infectious agents that pose a risk to free-roaming wildlife or humans after release. Dermatophyte fungi, commonly known as "ringworm," have zoonotic potential and may be carried as normal flora on the haircoats of certain species. Outbreaks of ringworm are anecdotally reported to occur in white-tailed deer (Odocoileus virginianus) fawns, but no prevalence surveys have been conducted on the haircoat flora of free-roaming individuals. In November 2008, we tested 60 legally hunted white-tailed deer for dermatophyte flora by using a modified MacKenzie technique. Results indicate it is unlikely that wild, mature white-tailed deer in Virginia, USA, carry dermato-phyte fungi as normal haircoat flora. Therefore, wildlife rehabilitators and hunters are at low risk for dermatophyte infection by direct contact with this species. In addition, the RapidVet-D 3 Day Test for Veterinary Dermatophytosis was determined to have poor specificity for presence of dermatophyte fungi on asymptomatic white-tailed deer in Virginia.     

Multicenter study of dermatophyte distribution in the metropolitan area of Barcelona (Catalonia,Spain)

The results of a multicenter study on the distribution of dermatophyte species in the metropolitan area of Barcelona (Spain) are presented.In the period 1977–1983, 638 dermatophyte strains were isolated from tineas by three different mycological laboratories. Zoophilic strains were prevalent comprising 64.4% of the isolates. T. mentagrophytes var. granulosum was isolated in 51% of the cases.A signigicant drop in the prevalence of T. tonsurans and T. violaceum was noted. Important variations were observed in dermatophyte distribution in relation to other geographical areas of Spain.     

An In Vitro Model for the Study of the Macrophage Response Upon Trichophyton rubrum Challenge

Diversity in the macrophage models currently employed in immunology studies may lead to opposed results and interpretations. In this study, we aimed to analyze the suitability of J774 macrophage-like cells as a model for the interaction between the dermatophyte Trichophyton rubrum and macrophages. J774 cells were competent in fungal phagocytosis, but succumbed to hyphal growth. Nevertheless, they could also secrete IL-1β in response to the dermatophyte. On the opposite direction, inflammatory, thioglycollate-induced peritoneal macrophages did not succumb to fungal growth and showed no significant IL-1β production. The proteomic profiling of these cells uncovered vimentin and plastin-2 as proteins whose abundance was altered by the fungal interaction. Our study indicates that this cell line could be an interesting tool in the investigation of T. rubrum infection biology.

     

Species Identification of Dermatophytes by MALDI-TOF MS

MALDI-TOF (matrix-assisted laser desorption/ionization time-of-flight) mass spectrometry (MS) is a new tool for the identification of microorganisms, inclusive of dermatophytes. The technique is faster, more straightforward, and powerful when compared to conventional dermatophyte identification methods (culture and microscopy). Accurate species identification in dermatophytes is not only essential to survey epidemiologic situations, but also for an appropriate medical treatment and to locate the source of infection (zoophilic or anthropophilic). Multiple platforms from a number of well-established commercial manufacturers (Andromas SAS, Bruker Daltonics, bioMérieux) have been used for dermatophyte identification with different success. Independent from the platform used, all of the studies reviewed here report on problems with the identification of phylogenetically closely related anthropophilic and zoophilic dermatophyte species. Thus, supplementation of the databases/libraries as well as standardized extraction protocols and cultivation methods are the precondition required for optimal species identification.     

An unusual lipid in the human pathogenic fungus Epidermophyton floccosum.

The dermatophyte Epidermophyton floccosum contains an unusual lipid characterized as 1(3),2-diacylglyceryl-3(1)-O-4'-(N,N,N-trimethyl)homoserine. Its concentration in E. floccosum is approx. 15% of the polar lipids. The role of this lipid in the biosynthesis of polar lipids and fatty acids is not yet known. However, it is interesting from a chemotaxonomic viewpoint that the lipids from two other genera of dermatophyte, Microsporum cookei and Trichophyton rubrum do not contain this novel lipid.     

The Clonal Population of Trichophyton equinum from Dermatophytoses of Japanese Racehorses

Mycopathologia - Trichophyton equinum is a zoophilic dermatophyte that is frequently isolated from horse dermatophytosis and rare infections in humans. In the present study, molecular and...     

MALDI-TOF-Based Dermatophyte Identification

MALDI-TOF MS has become increasingly popular for microorganism identification in the routine laboratory. Compared with conventional morphology-based techniques, MALDI-TOF is relatively inexpensive (per-unit identification), involves a rapid result turnaround time and yields more accurate results without the need for highly qualified staff. However, this technology has been technically difficult to implement for filamentous fungi identification. Identification of dermatophytes, a type of filamentous fungi, remains particularly challenging, partly due to the lack of clear species definition for some taxa or within some species complexes. Review of the ten studies published between 2008 and 2015 shows that the accuracy of MALDI-TOF MS-based identification varied between 13.5 and 100 % for dermatophytes. This variability was partly due to inconsistencies concerning critical steps of the routine clinical laboratory process. Use of both a complete formic acid-acetonitrile protein extraction step and a manufacturer library supplemented with homemade reference spectra is essential for an accurate species identification. This technique is conversely unaffected by variations in other routine clinical laboratory conditions such as culture medium type, incubation time and type of mass spectrometry instrument. Provided that a reference spectra library is adequate for dermatophyte identification, MALDI-TOF MS identification is more economical and offers an accuracy comparable to that of DNA sequencing. The technique also represents an advantageous alternative to the protracted and labor-intensive dermatophyte identification via macroscopic and microscopic morphology in the routine clinical laboratory.     

Fluorescent method for studying the morphogenesis and viability of dermatophyte cells

The dermatophyte Microsporum canis is commonly isolated from human and animal infection. The morphogenesis of this fungus was studied during its developmental stages through the fluorescent method Fluorescein Diacetate and Ethidium Bromide. To this end, 50 l dermatophyte suspension were transferred onto cellophane wrapping esterilized discs (2.5 cm of diameter) placed over the surface of Sabouraud dextrose agar on Petri dishes and incubated at 25 °C for 30 days. Every 60 minutes during the first 24 hours and every 12 hours for next 29 days, one disc was transferred onto glass slide, covered with equal volumes of freshly prepared fluorescein diacetate (FDA) and ethidium bromide (EB) solution, mounted with a coverslip and incubated in the dark for 30 minutes, at 25 °C. Each preparation was then examined on a fluorescent microscope. M. canis presented well defined growth stages: (1) tumescence of cells; (2) germination; (3) development of hyphae; (4) production of conidia and (5) tumescence and formation of arthroconidiae. Using the fluorescent method, non viable cells showed a light bright red coloration and viable cells presented green fluorescence. The principal morphological changes have occurred between the 3rd until the 18th day of culture. The method is very useful to demonstrate the dermatophyte growth stages as well as the perfect differentiation between viable and non viable cells.This revised version was published online in October 2005 with corrections to the Cover Date.     

Non-dermatophyte Dermatoses Mimicking Dermatophytoses in Animals

Dermatophytoses in animals are fungal diseases of the skin caused by dermatophyte fungi of the genus Microsporum or Trichophyton. Because the infection is generally follicular, the most common clinical sign is one or many circular areas of alopecia with variable erythema, scaling and crusting, and the primary differential diagnoses are follicular infections, such as bacterial folliculitis and demodicosis. Although dermatophyte folliculitis or ringworm is the most commonly observed lesion of dermatophytoses in animals, other presentations may be observed according to the host species and the dermatophyte involved: dermatophyte folliculitis or ringworm, scaling and crusting in dermatophytosis due to Microsporum persicolor, nodule in case of kerion or mycetoma, matted hairs, seborrheic dermatosis or miliary dermatitis in cats, generalized exfoliative dermatoses in dogs, cats and horses, superficial non-follicular pustules, papules and macules in the Devon Rex cat, pruritic dermatophytoses in dogs, cats and horses, and onychomycosis in dogs, cats and horses. Since manifestations of dermatophytosis are highly variable, particularly in the cat, dermatophytosis should be considered in case of any annular, papular, nodular or pustular dermatoses, alopecic or not, sometimes pruritic, and nodular dermatoses as well.     

濮阳市部分浅部真菌病中癣菌和念珠菌混合感染的分析

目的了解濮阳地区部分浅部真菌病中两种及两种以上真菌混合感染的情况。方法对临床确诊为浅部真菌病的患者456例,取标本行10%KOH直接镜检、真菌培养及菌种鉴定。登记确诊患者相关临床资料,分析其中混合感染的发病特点。结果确诊皮肤癣菌和念珠菌混合感染病例36例,分离率为7.89%,多见于股部(58.33%)和足部(27.78%);分离菌株72株,皮肤癣菌中以红色毛癣菌为主(75.00%),念珠菌中以非白念珠菌为主(72.22%)。结论濮阳地区部分浅部真菌病中混合感染主要为皮肤癣菌和念珠菌的混合感染,多见于股部和足部;致病菌以红色毛癣菌和非白念珠菌为主。     

Epidemiology of Dermatophytoses in an Area South of Tehran,Iran

Dermatophyte infections have been considered to be a major public health problem in many parts of the world. The aim of this study was to identify the etiological and epidemiological factors of dermatophyte infections in an area south of Tehran. A total of 1254 patients suspected to have dermatophytic lesions were examined over a period of three years (1999-2001). Material collected from skin, hair, and nails was submitted to direct microscopic examination using KOH, cultured in Sabouraud dextrose agar and microscopically examined for colony morphology, in order to the identify the 169 dermatophytes isolated. The prevalence of dermatophytoses was 13.5% (95% CI: 11.7-15.5%). Their incidence was 10.6 per 100,000 person-years (95% CI: 8.5-13.2). Epiderophyton floccosum was the most frequent dermatophyte isolated (31.4%) followed by Trichophyton rubrum (18.3%), T. mentegrophytes (17.2%), T. violaceum (16.6%), Microsporum canis (6.5%), T. verrucosum (4.7%) and M. gypseum (4.1%). Epidermophytes floccosum was found to be the most common isolated dermatophyte in age groups 20-29 (30.2%). Tinea corporis (31.4%) was the most common type of infection, followed by tinea cruris (20.7%), tinea manuum (15.4%), tinea capitis (12.4%), tinea pedis (10.6%), tinea faciei (7.1%), and tinea unguium (2.4%). The frequency rate of all of the types of tinea was higher in males than in females. The anthrophilic species E. floccosum was the most common dermatophyte as a causative agent of tinea. The most prevalent fungal infection was tinea corporis caused by E. floccosum.     

First Isolation of Pseudogymnoascus destructans,the Fungal Causative Agent of White-Nose Disease,in Bats from Italy

Mycopathologia - White-nose disease, caused by the dermatophyte Pseudogymnoascus destructans, is a devastating pathology that has caused a massive decline in the US bat populations. In Europe, this...     

Plant genetics predicts intra-annual variation in phytochemistry and arthropod community structure

With the emerging field of community genetics, it is important to quantify the key mechanisms that link genetics and community structure. We studied cottonwoods in common gardens and in natural stands and examined the potential for plant chemistry to be a primary mechanism linking plant genetics and arthropod communities. If plant chemistry drives the relationship between plant genetics and arthropod community structure, then several predictions followed. We would find (i) the strongest correlation between plant genetic composition and chemical composition; (ii) an intermediate correlation between plant chemical composition and arthropod community composition; and (iii) the weakest relationship between plant genetic composition and arthropod community composition. Our results supported our first prediction: plant genetics and chemistry had the strongest correlation in the common garden and the wild. Our results largely supported our second prediction, but varied across space, seasonally, and according to arthropod feeding group. Plant chemistry played a larger role in structuring common garden arthropod communities relative to wild communities, free-living arthropods relative to leaf and stem modifiers, and early-season relative to late-season arthropods. Our results did not support our last prediction, as host plant genetics was at least as tightly linked to arthropod community structure as plant chemistry, if not more so. Our results demonstrate the consistency of the relationship between plant genetics and biodiversity. Additionally, plant chemistry can be an important mechanism by which plant genetics affects arthropod community composition, but other genetic-based factors are likely involved that remain to be measured.     

Enhanced gene replacements in Ku80 disruption mutants of the dermatophyte, Trichophyton mentagrophytes

The frequency of targeted gene disruption via homologous recombination is low in the clinically important dermatophyte, Trichophyton mentagrophytes . The Ku genes, Ku70 and Ku80 , encode key components of the nonhomologous end-joining pathway involved in DNA double-strand break repair. Their deletion increases the homologous recombination frequency, facilitating targeted gene disruption. To improve the homologous recombination frequency in T. mentagrophytes , the Ku80 ortholog was inactivated. The nucleotide sequence of the Ku80 locus containing a 2788-bp ORF encoding a predicted product of 728 amino acids was identified, and designated as TmKu80 . The predicted TmKu80 product showed a high degree of amino acid sequence similarity to known fungal Ku80 proteins. Ku80 disruption mutant strains of T. mentagrophytes were constructed by Agrobacterium tumefaciens -mediated genetic transformation. The average homologous recombination frequency was 73.3 ± 25.2% for the areA/nit-2 -like nitrogen regulatory gene ( tnr ) in Ku80⁻ mutants, about 33-fold higher than that in wild-type controls. A high frequency ( c . 67%) was also obtained for the Tri m4 gene encoding a putative serine protease. Ku80 ⁻ mutant strains will be useful for large-scale reverse genetics studies of dermatophytes, including T. mentagrophytes , providing valuable information on the basic mechanisms of host invasion.     

Dermatophytosis caused by <Emphasis Type="Italic">Microsporum canis</Emphasis> in Eastern cottontail (<Emphasis Type="Italic">Sylvilagus floridanus</Emphasis>)

Eastern cottontail (Sylvilagus floridanus, fam. Leporidae) has previously been shown to be a potential healthy carrier of dermatophyte fungi both geophilic (Microsporum gypseum, M. cookei, Trichophyton ajelloi, T. terrestre) and zoophilic (M. canis, T. mentagrophytes). In this communication, the first case, to the best of our knowledge, of a symptomatic dermatophyte infection in S. floridanus is described.     

Eastern Cottontail (<Emphasis Type="Italic">Sylvilagus Floridanus)</Emphasis> as Carrier of Dermatophyte Fungi

Eastern cottontail (Sylvilagus floridanus, fam. Leporidae), introduced into Piedmont (Italy) in the 1960s, was studied as carrier of dermatophyte fungi. Of 216 hair samples collected from animals culled between September 1999 and July 2000 in the Province of Alessandria (Piedmont, Italy) during a pest control project, 57 (26.4%) yielded dermatophyte colonies. As two different species of dermatophytes grew from two samples, a total of 59 fungal isolates (26.5%) were obtained. Six dermatophyte species both geophilic (M. gypseum, M. cookei, Trichophyton ajelloi, T. terrestre) and zoophilic (M. canis, T. mentagrophytes) were identified. No sex-related differences were found but season-related differences were observed. The highest prevalence of dermatophyte-positive samples was recorded in May–September, due to the geophilic fungi whose prevalence decreased during colder and increased during warmer months ( p < 0.001). The presence of zoophilic dermatophytes, T. mentagrophytes, commonly associated with rodents, small mammals and lagomorphs and M. canis, usually correlated with domestic environment, did not change the whole year round. As Eastern cottontail has been showed to be a carrier of dermatophytes transmissible to man (M. canis, T. mentagrophytes and M. gypseum), it may represent a source of infection for gamekeepers, hunters and veterinarians.     

Isolations of dermatophyte from clinically normal scalps inM. Canis infections using the hairbrush method

The scalp hair of patients with dermatophytosis due toM. canis but without scalp lesions, and that of their family members without dermatophytosis were examined using the hairbrush method. The dermatophyte was detected in 93.8% of the scalps of those who lived in homes where cats were kept, and in 25% of those without cats. After the source of infection had been treated, the dermatophyte showed a gradually decreasing presence, finally disappearing altogether, so that no case of the disease on the scalp hair could be found.We inferred from the above findings inM. canis infections that, since the dermatophytes are seen in a high proportion of cases without scalp lesions, the dermatophytes, in many cases, exist only as saprophytes on the hair.