In vitro hair cultures for differentiating between atypical isolates ofTrichophyton mentagrophytes andTrichophyton rubrum

Summary Isolates ofT. mentagrophytes andT. rubrum are encountered that cannot be distinguished from each other solely on the basis of morphological criteria.Since these two species fundamentally differ in the manner in which they attack hairin vitro, this property can be used as a diagnostic aid, when correlated with their morphologic characteristics as developed on a variety of media. T. mentagrophytes radially penetrates hair segments immersed in water forming wedge-shaped perforations.T. rubrum does not perforate hair.The method of determining the ability or inability ofT. mentagrophytes orT. rubrum to perforate hair and the use of this test in identifying those species are described.     

Differentiation of trichophyton rubrum from trichophyton mentagrophytes

Conclusions Of the several morphological and physiological criteria that have been proposed to identifyTrichophyton rubrum andTrichophyton mentagrophytes differentially, only the hair invasion test agrees with the immunological results reported here.A typical strain ofT. rubrum andT. mentagrophytes was obtained from the Centraalbureau voor Schimmelcultures (Netherlands) and was used as reference strain. All isolates that did not invade hair shafts (in vitro) and that produced three precipitate lines in agargel diffusion tests against anti-T. rubrum serum (refence strain, Tr-1) were placed in one group. These isolates all also produced a red undersurface pigment and macroconidia with long parallel sides. They were labledT. rubrum.Other isolates that did invade hair shafts, and produced only two precipitate lines against anti-T. rubrum serum were placed in another group and were labeledT. mentagrophytes. These isolates had variable characteristics of pigment production, colony and conidial morphology.The antigenic studies thus corroborate the validity of the hair invasion test. The hair invasion test will probably remain in wider use because of its simplicity.     

Lipolytic activity of dermatophytes

Lipolytic activity of dermatophytes was tested by the method generally used forCandida lipolytica. Most of the freshly isolated strains ofMicrosporum canis, M. gypseum, Epidermophyton floccosum andTrichophyton mentagrophytes gave positive reactions, whereas, only few strains ofT. schoenleini, T. violaceum, T. megnini, T. rubrum andT. tonsurans yielded such reactions. Among subcultured strains, it was found a decreased lipolytic activity inMicrosporum species and an increased lipolytic activity inEpidermophyton and someTrichophyton species.Part of this work was performed at Serviço de Doenças Infecto-Contagiosas (Secção de Dermatologia), Hospital de Curry Cabral - Lisboa.     

Molecular confirmation of a Trichophyton violaceum isolate from human black-dot ringworm

A clinical isolate from a black-dot ringworm lesion of a 28-year-old female Japanese was investigated by morphological and biochemical analyses as well as molecular analyses. The isolate grew well onthiamine enriched agar and did not produce violetpigment, macroconidia or microconidia on Sabouraud's dextrose agar. Approximately 620-bp genomic DNA fragments of the CHS1 gene were amplified from Trichophyton mentagrophytes, T. rubrum, T. tonsurans and T. violaceum by polymerase chain reaction (PCR) and sequenced. The chitin synthase 1 (CHS1) nucleotide sequences of the clinical isolate showed more than 97% similarity to that of T. violaceum and less than 96% similarity to that of T. mentagrophytes, T. rubrum and T. tonsurans. The phylogenetic analysis of their sequences revealed that the clinical isolate was genetically close to T. violaceum and distinct from T. mentagrophytes, T. rubrum and T. tonsurans. Therefore, the isolate was confirmed as T. violaceum by mycological examination and molecular analyses. This revised version was published online in June 2006 with corrections to the Cover Date.     

Mating patterns of dermatophytes of diverse origin in India

The mating patterns of Trichophyton mentagrophytes var. interdigitale (74 isolates) and the Microsporum gypseum complex (17 isolates) of diverse origin and T. rubrum (25 isolates) and T. tonsurans (10 isolates) of clinical origin were studied. The results of the study showed that the teleomorph of the Indian isolates of T. mentagrophytes belong to Arthroderma vanbreuseghemii, undetermined teleomorphs of T. mentagrophytes var. interdigitale (+) mating types, and undetermined teleomorphs of T. mentagrophytes var. interdigitale indeterminate mating types. All the isolates of T. rubrum and T. tonsurans were found to be of the (-) mating type.     

Effective photodynamic treatment of Trichophyton species with Rose Bengal

Photodynamic therapy (PDT) with Rose Bengal has previously achieved eradication of Trichophyton rubrum infections causing toenail onychomycosis; however, its antifungal activity against other clinically relevant dermatophytes has yet to be studied. Here, we test the efficacy of PDT using Rose Bengal (140 μM) and 532 nm irradiation (101 J/cm²) against Trichophyton mentagrophytes and Trichophyton interdigitale spores, in comparison to T. rubrum. A significant reduction (>99%) of T. mentagrophytes and T. interdigitale was observed, while actual eradication of viable T. rubrum was achieved (99.99%). Laser irradiation alone inhibited growth of T. rubrum (55.2%) and T. mentagrophytes (45.2%) significantly more than T. interdigitale (25.5%) (P = .0086), which may indicate an increased presence of fungal pigments, xanthomegnin and melanin. The findings suggest that Rose Bengal-PDT can act against a broader spectrum of fungal pathogens, and with continued development may be employed in a wider range of clinical antifungal applications.     

Antifungal potential of essential oil and various organic extracts of <Emphasis Type="Italic">Nandina domestica</Emphasis> Thunb. against skin infectious fungal pathogens

This study was undertaken to assess the in vitro antifungal potential of the essential oil and n-hexane, chloroform, ethyl acetate, and methanol extracts of Nandina domestica Thunb. against dermatophytes, the casual agents of superficial infections in animals and human beings. The oil (1,000 μg/disc) and extracts (1,500 μg/disc) revealed 31.1–68.6% and 19.2–55.1% antidermatophytic effect against Trichophyton rubrum KCTC 6345, T. rubrum KCTC 6375, T. rubrum KCTC 6352, Trichophyton mentagrophytes KCTC 6085, T. mentagrophytes KCTC 6077, T. mentagrophytes KCTC 6316, Microsporum canis KCTC 6591, M. canis KCTC 6348, and M. canis KCTC 6349, respectively, along with their respective minimum inhibitory concentration values ranging from 62.5 to 500 and 125 to 2,000 μg/ml. Also, the oil had strong detrimental effect on spore germination of all the tested dermatophytic fungi as well as concentration and time-dependent kinetic inhibition of T. rubrum KCTC 6375. The present results demonstrated that N. domestica mediated oil and extracts could be potential sources of natural fungicides to control certain important dermatophytic fungi.     

Studies on the phospholipase A of dermatophytes

The presence of phospholipase A activity was detected in three dermatophytes:Microsporum cookei, Trichophyton mentagrophytes, andEpidermophyton floccosum. The activity was always higher inT. mentagrophytes than inM. cookei andE. floccosum. All exhibited phospholipase A₁ and A₂ activities, but the activity was largely A₂ inM. cookei and A₁ inE. floccosum. T. mentagrophytes possessed almost equal activities of phospholipase A₁ and A₂.     

Urease activity inTrichophyton mentagrophytes

Urease activity was detected in the dermatophyteTrichophyton mentagrophytes cells at early exponential phase of growth. Specific activity of urease decreased with culture age. At exogenous urea concentrations above 2 mm formation of urease was inhibited. The pH optimum lay at 7–7.5, the K_m being 14 mm. No urease activity could be detected in cell-free culture fluid ofT. mentagrophytes. No endoor exocellular urease activity could be detected in aT. rubrum strain grown with or without urea.     

Review of dermatophytoses in Galicia from 1951 to 1987, and comparison with other areas of Spain

We have reviewed all the dermatophytoses diagnosed in Galicia during four consecutive 9-year periods 1951–86 and 1987. From 4571 patients, we isolated 3351 fungal strains belonging, in decreasing order of frequency, to the following dermatophyte species: Microsporum canis (25.5%), Trichophyton rubrum (24.6%), T. mentagrophytes (21.4%), Epidermophyton floccosum (11.8%), M. gypseum (5.2%), T. tonsurans (3.9%), T. verrucosum (3.1%), T. schoenleinii (2.5%), T. violaceum (1.2%), T. mengninii (0.3%), M. audouinii (0.2%), T. equinum (0.1%) and T. soudanense (0.1%). Tinea capitis has diminished in frequency since 1951, though there was been a slight increase since 1978; M. canis has always been the most common agent, and between 1951 and 1959 T. schoenleinii was also very frequent but is no longer found. The frequency of tinea corporis, on the other hand, has experienced a considerable increase. Its most common causal agents in the last few years have been T. mentagrophytes, M. canis and T. rubrum. Until 1977 the most common tinea cruris dermatophyte was E. floccosum, but since then it has been T. rubrum. The commonest tinea pedis dermatophytes have been T. rubrum and T. mentagrophytes. Tinea unguium and tinea barbae have been the most frequent dermatophytoses, and their commonest causal agents T. rubrum and T. mentagrophytes respectively. We have documented the distribution of the various causal agents by location of the lesions, age and source of the patients (private or National Health Service patients), and we have compared the results with those obtained in other regions of Spain.     

Dermatophytes isolated from patients in University Hospital,Kuala Lumpur,Malaysia

A total of 576 dermatophytes were isolated from patients with a variety of skin infections from January 1993 to May 2000. Ten species of dermatophytes were identified: Epidermophyton floccosum (0.7%), Microsporum audouinii (1.1%), M. canis (3.1%), M. gypseum (0.3%), Trichophyton concentricum(3.5%), T. equinum (0.2%), T. mentagrophytes (36.%), T . rubrum (53.8%), T. verrucosum (0.2) and T. violaceum (1.0%). The body sites most frequently affected by dermatophytes were the buttocks, nails and trunk. Anthropophilic dermatophytes made up 60.1% of the isolates; the most common species was T. rubrum, T. mentagrophytes and M. canis were the two main zoophilic dermatophytes. T. mentagrophytes was isolated from all body sites except the scalp. M. canis was found to be associated with domestic dogs and wasnot isolated from ethnic Malays. The only geophilic dermatophyte was M. gypseum, an uncommon dermatophyte associated with tinea pedis.This revised version was published online in October 2005 with corrections to the Cover Date.     

A short review on dermatophytoses of animals in Romania

Dermatophytoses of animals in Romania are reviewed. Ringworm infections have been found in cattle (whereT. verrucosum was the only isolated agent), horses (the causative organisms beingT. verrucosum, T. mentagrophytes andM. equinum), cats (dermatophytes isolated:M. canis andT. quinckeanum), dogs (M. canis andM. gypseum), fowlsT. gallinae). From the laboratory animals: guinea pigs (T. mentagrophytes, M. audouinii, T. rubrum), mice (T. quickeanum andT. mentagrophytes), rats (T. mentagrophytes), rabbits (T. mentagrophytes andM. canis) and hamster)T. mentagrophytes). Among the wild animals, only the chamois had a ringworm infection byM. gypseum, other large or smaller wild animals harboring only saprophytically some poozhilic and geophilic dermatophytes.     

Molecular cloning ofTrichophyton mentagrophytes DNA sequences with promoter activity inEscherichia coli

A promoter probe library from the dermatophyte fungusTrichophyton mentagrophytes has been constructed in the pVB32 plasmid vector. Using this library, a set ofT. mentagrophytes DNA sequences with promoter activity inEscherichia coli has been cloned. The size and the resistance phenotype conferred by these DNA fragments varied. Southern blot analysis confirmed that they were derived fromT. mentagrophytes genomic DNA.     

In vitro characterization of Trichophyton rubrum and T. mentagrophytes biofilms

Dermatophytes are fungi responsible for a disease known as dermatophytosis. Biofilms are sessile microbial communities surrounded by extracellular polymeric substances (EPS) with increased resistance to antimicrobial agents and host defenses. This paper describes, for the first time, the characteristics of Trichophyton rubrum and T. mentagrophytes biofilms. Biofilm formation was analyzed by light microscopy, scanning electron microscopy (SEM) and confocal laser scanning microscopy (CLSM) as well as by staining with crystal violet and safranin. Metabolic activity was determined using the XTT reduction assay. Both species were able to form mature biofilms in 72?h. T. rubrum biofilm produced more biomass and EPS and was denser than T. mentagrophytes biofilm. The SEM results demonstrated a coordinated network of hyphae in all directions, embedded within EPS in some areas. Research and characterization of biofilms formed by dermatophytes may contribute to the search of new drugs for the treatment of these mycoses and might inform future revisions with respect to the dose and duration of treatment of currently available antifungals.     

Enzymes of Trichophyton rubrum

Jensen, Altschuler &Bard (1957) have surveyed the respiratory enzymes ofTrichophyton mentagrophytes. Enzymes of the hexosediphosphate, hexosemonophosphate, and tricarboxylic acid pathways were found plus a group of miscellaneous enzymes. In view of the close morphological, immunological, and nutritional similarities ofT. mentagrophytes toTrichophyton rubrum, we felt that a study of the enzymology of the latter organism had potential value in forming a basis for the comparison of the two dermatophytes. Our results showed that cell-free extracts ofT. rubrum contain glucokinase, TPN-dependent glucose and fructose dehydrogenases and separate TPN- and DPN-linked glycerol dehydrogenases. These results are in accord with findings of other investigators. ATPase activity was minimal, and shikimic acid kinase, and shikimic and quinic acid dehydrohydrogenases were not detected.     

Nutritional and environmental factors affecting pigmentation inTrichophyton rubrum

Summary 1. Pigmentation inTrichophyton rubrum appears to be dependent upon tyrosine and phenylalanine (as precursors), carbohydrate (primarily as an energy source), and oxygen.2. Cysteine and methionine inhibit the growth ofT. rubrum in a minimal medium. L-sorbose also inhibits the growth of this organism, even in a complete medium.3.T. rubrum apparently synthesizes all of the vitamins required for growth and pigmentation.4. At certain concentrations of ascorbic acid, it is possible to distinguish between the pigmentogenic and growth processes ofT. rubrum.5. Sunlight suppresses growth and pigmentation inT. rubrum.     

The detection of contamination in Trichophyton rubrum and trichophyton mentagrophytes

This report concerns the usefulness of two media, brain heart infusion agar (BHIA) and BCP milk dextrose agar in the detection of contamination inT. rubrum andT. mentagrophytes and provides cultural information in the identification of these species.Department of Microbiology, Public Health Laboratory, Ontario Department of Health, Box 9000, Postal Terminal A, Toronto 1, Ontario, Canada.Head, Medical Mycology Section.Mycologist, Medical Mycology Section.     

Rylux BSU stimulates spore germination in Trichophyton mentagrophytes and Aspergillus fumigatus and increases the survival rate after UV-irradiation

Calcofluor-allied optical brightener Rylux BSU stimulated spore germination rate inTrichophyton mentagrophytes andAspergillus fumigatus both if supplemented into Sabouraud glucose agar and if used for pretreatment of spore suspension prior to inoculation at low concentrations. Maximum stimulation of germination was obtained if 0.2% Rylux BSU was used for pretreatment in aqueous solution for 1 d prior to inoculation (130% inT. mentagrophytes and 150% inA. fumigatus, respectively). Pretreatment with 1% Rylux BSU provided strong protection against UV-irradiation and resulted in increased yields of cultural variants after UV-irradiation.     

Dermatophyte species,microscopic and cultural examination

The dermatophytes Epidermophyton floccosum, Microsporum canis, Trichophyton mentagrophytes var. interdigitale, T. rubrum, and T. verrucosum were compared with respect to the direct microscopic examination of a clinical material and the number of colonies obtained by culture.It was found that the results of microscopy as well as of culture depended to a marked extent upon which species were the cause of the mycosis.The extremes were E. floccosum and T. mentagrophytes var. interdigitale which showed 7.5 % and 32.5 % isolates with negative microscopic findings and 45.5 % and 5.0 % isolates with 10 colonies.     

Investigation of the dark metabolism of acetate in photoheterotrophically grown cells ofRhodospirillum rubrum

The mechanism of the aerobic dark assimilation of acetate in the photoheterotrophically grown purple nonsulfur bacteriumRhodospirillum rubrum was studied. Both in the light and in the dark, acetate assimilation inRsp. rubrum cells, which lack the glyoxylate pathway, was accompanied by the excretion of glyoxylate into the growth medium. The assimilation of propionate was accompanied by the excretion of pyruvate. Acetate assimilation was found to be stimulated by bicarbonate, pyruvate, the C₄-dicarboxylic acids of the Krebs cycle, and glyoxylate, but not by propionate. These data implied that the citramalate (CM) cycle inRsp. rubrum cells can function as an anaplerotic pathway under aerobic dark conditions. This supposition was confirmed by respiration measurements. The respiration of cells oxidizing acetate depended on the presence of CO₂ in the medium. The fact that the intermediates of the CM cycle (citramalate and mesaconate) markedly inhibited acetate assimilation but had almost no effect on cell respiration indicated that citramalate and mesaconate were intermediates of the acetate assimilation pathway. The inhibition of acetate assimilation and cell respiration by itaconate was due to its inhibitory effect on propionyl-CoA carboxylase, an enzyme of the CM cycle. The addition of 5 mM itaconate to extracts ofRsp. rubrum cells inhibited the activity of this enzyme by 85%. The data obtained suggest that the CM cycle continues to function inRsp. rubrum cells that have been grown anaerobically in the light and then transferred to the dark and incubated aerobically.