Identification of the causative dermatophyte of tinea capitis in children attending Mbarara Regional Referral Hospital in Uganda by PCR-ELISA and comparison with conventional mycological diagnostic methods.

Tinea capitis is a dermatophyte infection common among prepubertal children in sub-Saharan Africa and mainly caused by Trichophyton and Microsporum species. Accurate identification is challenging as conventional methods like culture and microscopy are slow and mostly based on morphological characteristics, which make them less sensitive and specific. Modern molecular methods, like polymerase chain reaction (PCR) assays, are gaining acceptance and are quick as well as accurate. The aim of this study was to investigate the clinical patterns of tinea capitis and to accurately identify the most common causative dermatophytes affecting the scalps of children aged 1 to 16 years attending the Skin Clinic at Mbarara University of Science and Technology (MUST), Mbarara, Uganda, East Africa, using both conventional mycological methods and PCR-ELISA for detection of dermatophyte DNA. One hundred fifteen clinical samples from children from Western Uganda attending the MUST Skin Clinic with a clinical diagnosis of tinea capitis were analyzed. T. violaceum was identified as the most common causative agent, followed by M. audouinii, T. soudanense, and T. rubrum. The early identification of the causative agent of tinea capitis is a prerequisite for the effective management of the disease, the identification of probable source and the prevention of spreading. Children with tinea capitis in Western Uganda should be treated by systemic therapy rather than topical preparations to ensure high cure rates as the most common causative dermatophytes T. violaceum exhibits an endothrix rather than ectothrix invasion of the hair follicle.

Unusual strains of Microsporum audouinii causing tinea in Europe.

We comment on an unusual strain of Microsporum (M.) audouinii. It was isolated from tinea corporis of a boy who lived in Germany and most likely had acquired his infection during a stay on a farm with animal husbandry in Poland. The strain showed features of M. canis (plenty of markedly rough-walled macroconidia, growth on rice, positive hair perforation) as well as of M. audouinii (white thallus, long macroconidia with central constriction) and in vitro it degraded hair of various mammals. Because its ribosomal internal transcribed spacer region showed 99.9% homology to a M. audouinii reference strain it was finally identified as M. audouinii. We relate these findings with recent observations of M. audouinii causing tinea in Europe. This appraisal suggests that irrespective of an identical ribosomal ITS region distinct M. audouinii strains can display a spectrum of morphological and physiological features that is broader than currently outlined in mycological textbooks. Certain unusual characteristics like an enhanced capacity to utilise keratins may even be associated with unexpected transmission routes. Above all sporadic M. audouinii infections in Europe that bear no relation to an endemic area should be analysed from this perspective.

Cat favus caused by Microsporum incurvatum comb. nov.: the clinical and histopathological features and molecular phylogeny.

Favus is a distinctive form of infection that is caused by exclusively dermatophytes. Its clinical presentation is characterized by scutula, which are concave, thick fungal crusts. The best-known examples of human scalp favus are caused by Trichophyton schoenleinii and those of mouse favus are caused by T. quinckeanum. However, other dermatophytes, such as T. violaceum, T. verrucosum, Microsporum audouinii, M. gallinae, M. gypseum, and M. canis, have been reported sporadically to cause favic lesions. Favus on cats has rarely been mentioned in the literature, and the pathogens with which it has been associated are, for the most part, unknown. Here, we examine four cat favus cases, focusing on clinical presentations and histopathological features. In all cases the etiologic agent was identified as M. incurvatum based on its morphological characteristics and sequences of internal transcribed spacers (ITS) of nuclear ribosomal DNA. Phylogenetic analysis using the neighbor-joining method, which is based on ITS, showed that these four isolates belonged to two strains of M. incurvatum; one strain was a new combination from the basionym Nannizzia incurvata.

Standardization of hyphal growth inhibition rate as a means of evaluating Microsporum spp. in vitro susceptibility to terbinafine, griseofulvin, and ciclopiroxolamine.

Reference methods for antifungal susceptibility tests recommend the use of conidia as inoculum. However, some isolates produce few conidia, while the invasive form of filamentous fungi in general is hyphae making susceptibility tests infeaseble. These facts suggest that other than conidia broth dilution method is required for susceptibility tests. The aim of this study was to clarify if the hyphal growth inhibition rate could be used as a method of determining the antifungal susceptibility of genus Microsporum. For this reason, a method which traces hyphal tips automatically and measures their growth rate was standardized for Microsporum spp. Control growth curves and test growth curves obtained by real-time observation of the hyphae groups responses to different concentrations of terbinafine, griseofulvin, and ciclopiroxolamine were used to compare with minimum inhibitory concentrations (MICs) obtained by conidia broth microdilution method. A visible reduction in the growth inhibition rate was observed when hyphal activity was evaluated using the third or fourth serial two-fold dilution below the MIC determined by broth microdilution for terbinafine and ciclopiroxolamine. For griseofulvin, this reduction occurred after the fifth dilution below the MIC. This study highlights the importance of the inoculum type used to determine the in vitro susceptibility of Microsporum strains. We conclude that measurement of hyphal growth inhibition, despite being time consuming, could be a suitable method for evaluating antifungal susceptibility, particularly for fungi as Microsporum spp. that produce a small (or not at all) number of conidia.

Tinea corporis caused by an unusual strain of Microsporum audouinii that perforates hair in vitro.

We report on a dermatophyte infection acquired by a young woman from Germany who had worked in Ghana. The strain isolated from her skin lesions showed morphological and physiological features compatible with Microsporum audouinii but a clearly positive hair perforation test made its definite identification by conventional methods equivocal. A genetic analysis finally unambiguously revealed Microsporum audouinii. This is the first observation of a Microsporum audouinii strain with a positive hair perforation test. The ability to perforate hair may be related to attributes favouring an inflammatory host response.

First case of Microsporum ferrugineum from Tunisia.

This is the first case of Microsporum ferrugineum isolated from a Tunisian patient. A 60-year-old man was admitted for tinea sycosis associated with circinate herpes of the hand. Examination disclosed diffuse erythematic and perifollicular papules and pustules in the beard area. Typical ringworm vesiculo-pustular lesions involved skin of the hand. Isolates were identified as Microsporum sp on the basis of macroscopic and microscopic colony characteristics. The diagnosis of M. ferrugineum was confirmed by PCR sequencing of Chitin Synthase1 gene. The patient was treated successfully with Griseofulvin, which was administered for 4 weeks.

Glucose improves the in vitro viability of Microsporum canis and Trichophyton mentagrophytes var. mentagrophytes.

We describe simple and cost-effective methods using carbohydrates to improve the in vitro viability of dermatophytes. Glucose and sucrose in different concentrations (3, 6, 9 and 12%) were used to maintain fifteen strains of M. canis and T. mentagrophytes var. mentagrophytes at 4 and -20 degrees C. The strains were phenotypically analyzed before storage and reevaluated at 1, 3, 6 and 9 months. At 1 and 3 months, any alterations in the viability or phenotype pattern of the stored strains were noted. At 6 months, both dermatophytes were 100% viable, when preserved in glucose (3, 6, 9 and 12%) at -20 degrees C. All T. mentagrophytes strains were also viable in sucrose (12%), at 4 degrees C and -20 degrees C. However, sucrose failed to improve the viability of M. canis at both temperatures. At 9 months, the higher viabilities without pleomorphism were seen for both dermatophytes preserved in glucose (9 and 12%) at -20 degrees C.

Evaluation of Microsporum canis in different methods of storage.

The main objective of this investigation was to evaluate different methods of storage for Microsporum canis based on materials and equipment that are readily available in developing countries. We tested 32 strains of M. canis at - 20 degrees C in potato dextrose agar (PDA) in its plain condition, or amended with 10% dimethyl sulfoxide or with 10% glycerol. In addition, we tested 25 degrees C storage of isolates in plain saline (0.9% NaCl) and in saline covered with a mineral-oil layer. After 9 months of storage, none of the M. canis strains frozen in PDA supplemented with glycerol survived, while only 16 and 6%, respectively, of the isolates in plain and DMSO medium lost viability. Nine month's storage in saline with or without mineral oil increased the amount of pleomorphic development of sterile hyphae; this phenomenon occurred at a significantly higher level than was seen in isolates stored at -20 degrees C. The physiological characteristics of M. canis were not affected by the different storage tests. The results suggest that, in order to ensure optimal viability, purity and pristine isolate condition, each M. canis isolate maintained should be held in at least two methods of storage, namely, PDA at -20 degrees C and saline with a mineral-oil layer at 25 degrees C.

Economical microscope configuration for direct mycological examination with fluorescence in dermatology.

BACKGROUND: The use of fluorochromes such as Blankophor or Calcofluor allows to detect immediately and without ambiguity fungal elements in dermatological preparations. Whereas fluorescence microscopy is widely practised in clinical laboratories, it is not generally used in private practice because of the high price of a epifluorescence microscope. OBJECTIVE: To propose an economical microscope configuration to visualize fungal elements using fluorescence. METHODS: The preparations were examined with a standard microscope for routine observations, equipped with only two supplementary filters. RESULTS: Because the fungal elements produce a particularly bright fluorescence, a 25-watt halogen light is sufficient to visualize them in dermatological preparations: CONCLUSIONS: The proposed microscope configuration for direct mycological examination is particularly economical since equipment for epifluorescence and a vapour mercury lamp are not necessary.

Microsporum nanum--a report from Malawi (Africa).

Microsporum nanum Fuentes 1956 has been identified in scrapings from a skin lesion in a patient in Karonga District, Malawi, Africa. We believe that this is the first such identification from the African continent. The patient had a single skin lesion in his popliteal fossa and was known to rear pigs.

Antifungal activity of some 2,2':5',2"-terthiophene derivatives.

The dermatophyte Microsporum cookei Ajello was treated with nine new natural and synthetic 2,2':5',2"-terthiophenes to determine their possible antifungal activity. In the dark the thiophenes were inactive, while when photoactivated with UV-A they induced a remarkable reduction in the growth rate of the fungus. The only exception was (E)-N-(2-methylpropyl)-3-(2,2':5',2"-terthien-5-yl)-propenamide , which was not fungistatic even at the highest dose tested (24 microM). The more active compounds were 3'-methoxy-2,2':5',2"-terthiophene and 3'-methylthio-2,2':5',2"-terthiophene, whose activity seems to be related to the presence of a substituent in the 3' position of the central ring of thiophenes. Transmission electron microscopic observations demonstrated the photoactive nature of the synthetic molecules to be similar to that of alpha-terthienyl, a natural thiophene present in some Asteraceae. The dark treatment caused only the accumulation of the compound in vacuoles, without other evident alterations. After UV-A irradiation the activated thiophene causes severe modifications to the endomembrane system, probably via oxygen-dependent mechanism.

Preliminary report on the isolation of a dysgonic variety of Microsporum canis together with the normal variety from a cattery.

The isolation of a dysgonic variety of Microsporum canis from a large number of cats and kittens in a cattery is described. The normal variety of this fungus was isolated at the same time from the same animals. Dysgonic varieties are thought to be mutants of normal strains, but this isolation of both forms together suggests that the relationship may be more complex.

Microsporum gypseum complex in man and animals.

Twenty-eight strains of the Microsporum gypseum complex isolated from humans and animals were studied. The perfect form was found for 25 of the isolates. Nannizzia incurvata was the species most frequently involved in human pathology, while Nannizzia gypsea was most frequently found on animal lesions. Nannizzia fulva was rarely involved pathologically and Nannizzia corniculata was not isolated during this study. It is surprising to note that this species was not found even though most of our strains (22/28) came from Africa. Reliable methods are not available for differentiating among the anamorphs, which are commonly called M. gypseum, Microsporum fulvum or Microsporum boullardii. The Sabouraud medium conventionally used for medical mycology makes almost no distinction among them. We found that the species could be easily distinguished by colonial and microscopic features when grown on Takashio medium. When strains are atypical, sexual reproduction remains the reference technique but, in most cases, Takashio medium makes it possible to avoid this long drawn-out procedure.

An atypical Microsporum canis isolate.

An atypical strain of Microsporum canis isolated from a two-year-old boy with tinea corporis is described. When cultured on Lactritmel agar the strain presented the typical pigment of M. canis without developing characteristic macroconidia. After 6 weeks, scarce, rudimentary, fusiform macroconidia 120-150 microns long developed on Lactritmel and Sablac microcultures. In vitro, the strain developed hair perforating organs. Experimental inoculation of a healthy volunteer produced a tinea with typical fluorescence and endoectothrix attack. After isolation the strain remained stable. These results differ from the results reported by English and Tucker. This is the first atypical strain of M. canis reported in Venezuela.

Mycological studies on Microsporum equinum isolated in Finland, Sweden and Norway.

Sixteen isolates of Microsporum equinum from cases of equine dermatophytosis in Finland, Sweden or Norway were studied mycologically. All the isolates produced typical macroconidia and were negative in the hair perforation test in vitro and were urease-positive. The growth pattern of M. equinum on polished rice greatly resembled that of M. audouinii during the first 2-3 weeks of incubation. All the isolates were incompatible with Nannizzia otae, the teleomorph of M. canis, but fresh M. equinum isolates showed growth stimulation against the (-) tester strain of N. otae. No cleistothecia or pseudocleistothecia were seen.

Correlation between morphological and physiological characteristics in species of microsporum.

The correlations between the physiological and morphological characteristics of various Microsporum strains were studied together with the inhibitory activities toward different microorganisms.

Isolation of saprophytic Microsporum praecox Rivalier from sites associated with horses.

Several M. praecox isolates of saprophytic origin were obtained in Belgium from horses and their surroundings. Visualization of macroconidia in dust collected in stables proved its saprophytic origin. A few strains were obtained from human cases of tinea corporis.