Annals of RSCB Vol.

XVII, Issue 1/2012

MORPHOLOGY AND ULTRASTRUCTURE ASPECTS IN SPECIES

BELONGS TO TRICHOPHYTON GENUS USING LIGHT AND
SCANNING ELECTRON MICROSCOPY
C. V. Mihali1, A. Buruiana2, Violeta Turcus1,2, Aurelia Covaci1,
A. Ardelean1,2
1 – INSTITUTE OF LIFE SCIENCES, UVVG ARAD
2 – THE FACULTY OF NATURAL SCIENCES, ENGINEERING AND INFORMATICS,
UVVG ARAD

Summary
The present paper presents the morphology features of the macroconidia, microconidia,
thalus and general aspect of mycelium in Trichophyton genus. The species studied were
Trichophyton mentagrophytes ssp. mentagrophytes and Trichophyton mentagrophytes
ssp. quinckeanum. These species belongs to Arthrodermataceae family, order
Onygenales of Ascomycota division from Fungi. Observations were made on fungal
fragments from culture media containing strains isolated from patients with clinical
suspicious of tinea corporis and tinea capitis. Trichophyton species are causative agents
of dermatophytosis, onychomycosis, tinea barbae, tinea capitis, tinea corporis, tinea
cruris, tinea faciei, and tinea pedis. In species belonging to Trichophyton genus, the
morphological aspects can be analysed using an scanning electron microscope with
environmental mode of observation (ESEM) for ultrastructural details in
micro/macroconidia and talus aspects and general overview images using light
microscopy.
Keywords: ESEM, ultrastructure, light microscopy, morphology, Trichophyton genus.

mihaliciprian@yahoo.com

Introduction tissues, it is considered as one of the main

Trichophyton is a dermatophyte fungus causes of hair, skin, and nail infections in
which can be isolated from the soil, humans
humans, or animals. The Trichophyton (Weitzman et al., 1988). They have several
genus includes anthropophilic, geophilic, enzymes, for instance – folic acid,
and zoophilic species (De George et al., proteinases, elastase, keratinases. Other
2001; Georg, 1959). Some of the enzymes such as proteinases are the major
Trichophyton species are cosmopolitan and virulence factors of Trichophyton species.
others have a limited geographic Trichophyton mentagrophytes a zoophilic
distribution. There are 22 species under dermatophyte has 5 five different ssp., T.
genus Trichophyton. Most common are mentagrophytes ssp. interdigitale
Trichophyton mentagrophytes, anthropophilic, T. mentagrophytes var
Trichophyton concentricum, Trichophyton nodulare a rare anthropophilic form
rubrum, Trichophyton schoenleinii, isolated from cases of tinea pedis, T.
Trichophyton tonsurans, Trichophyton mentagrophytes ssp. mentagrophytes, T.
verrucosum, and Trichophyton violaceum. mentagrophytes ssp. quinckeanum and T.
From the total number of species, 11 are mentagrophytes var erinacei are zoophilic
often associated with tinea of the scalp, the dermatophytes. All these ssp. shaping
nails, and the skin in humans and 4 are together the T. mentagrophytes complex.
often isolated from animals. Trichophyton Differentiation between these ssp. not be
genus is a keratinophilic filamentous fungus achieved on a single culture medium
which has the ability to invade keratinized

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(Ajello and Cheng, 1967; Houck et al. support the European dominance of A.
1996). benhamiae. Numerous variations of colony
The Trichophyton species have morphology are seen among the
teleomorphic forms and these teleomorphs Trichophyton mentagrophytes complex
are classified under genus Arthroderma, (Beneke and Rogers, 1971; Rippon, 1982).
two perfect states have been described for Macroscopic appearance of the colony -
T. mentagrophytes: Arthroderma growth rate may range from being slow to
benhamiae and Arthroderma moderately rapid, colonies are waxy,
vanbreuseghemii (Ajello and Cheng, 1967; glabrous, downy to cottony. The surface
Takashio, 1973). Takashio (Takashio, colony color ranges from white to bright
1972) divided A. benhamiae in 2 ssp., an yellowish beige or red violet and the
American-European ssp. and an African reverse may be pale, yellowish, brown, or
ssp. He observed that intra-ssp. matings red – brown. Macroconidia are multicellular
were very fertile but inter-ssp. matings were (with 2 or more cells), in some cases can be
only low or poorly fertile or sterile. He smooth, thin or thick walled, cylindrical,
observed that the ssp. erinacei which club shaped, or cigar shaped, and are often
belongs to the African ssp. and ssp. caviae absent. Microconidia are unicellular, round
which belongs to the American-European to pyriform shaped, numerously present,
ssp. were sexually degenerated by may be solitary or arranged in grape like
comparing with the normal strain of A. clusters.
benhamiae. The Finnish and Swedish
strains of T. mentagrophytes were
compatible with A. benhamiae (+ type) and

Material and methods

Were examinated 2 strains of one drop of lactophenol cotton blue
Trichophyton mentagrophytes ssp. between the glas slide and adhesive-tape.
mentagrophytes and 1 of Trichophyton ESEM with cooling stage - colony sample
mentagrophytes ssp. quinckeanum from of Microsporum gypseum and
patients with isolated cutaneous lesions of Microsporum canis were excised and
tinea corporis and tinea capitis, pedis, placed in Eppendorf tubes with
barbae. The fragments of fungal Glutaraldehyde 2,7% for 6 hr at 4ºC. After
dermatophytes culture media containing they were mounted into the Cooling stage
strains isolated from patients with clinical (which is a temperature controlled
suspicious of tinea corporis and tinea specimen holder of ESEM microscope)
capitis, barbae were examinated using which has 3°C and 100% RH . The
optical microscopy and (ESEM) morphology studies was made using Fei
environmental scanning electron Quanta 250 ESEM, the parameters of
microscopy (Ludwig, 1998; Patrick, 2009), ESEM examination were represented by
for morphological and structural the following items: the image mode, the
particularities studies (Glauert, 1974). working distance (WD), the beam
Light microscopy - native samples was conditions, chamber pressure and the
made using adhesive-tape method of relative humidity into sample. In the same
sampling, the morphology studies was order they were: the gaseous secondary
made using a BX 43 Olympus optical electron detector detector (GSED); the
microscope with semi plan apochromate working distance: 5 mm; the beam
objectives 10X, 20X, 100X mages were conditions: 12,5 – 15 kV, spot 4,5; the
captured using a digital CCD camera, CX3 pressure range: 910 to 1400 Pa and the
Olympus. The stain method used was with relative humidity of 90 - 100%.

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Results and discussions

Trichophyton mentagrophytes ssp. white on obverse side and beige with a
mentagrophytes showed a flat rapidly central brown spot on reverse side.
growing granular whyte colony. Mycelium In Trichophyton mentagrophytes ssp.
was sparse and the powdery or granular quinckeanum, the colony was well defined,
appearance is due to the production of The colony has a white higher central area
numerous microconidia. Numerous while the active growing area is superficial
variations of shape colony morphology at the surface of the medium. In the
were seen among the Trichophyton examination with light microscopy, we
mentagrophytes complex (Fig. 1 A-C). observed the following morphology
Macroscopic appearance of the colony aspects: a general overview showed the
colours ranges from white to bright presence of numerous spiral hyphae in
yellowish beige. The colony colours were Trichophyton

Fig.1 – T. mentagrophytes ssp. mentagrophytes, macroscopic appearance of the colony (A,C -

reverse; B,D – obverse), colours ranges from white to bright yellowish beige.

mentagrophytes ssp. mentagrophytes, many smooth walls. In some macroconidia the

microconidia and macroconidia (Fig.2). In apex were pointed tip and others with
Trichophyton mentagrophytes ssp. rounded terminal cell. Large and elongate
quinckeanum, the presence of numerous macroconidia were seen. The microconidia
microconidia and macroconidia were were globular. Macroconidia in T.m. ssp.
observed (Fig.3) The numbers of quinckeanum, were not so elongate as in
macroconidia in T.m. ssp. mentagrophytes anterior ssp. and they have multilocular
were lower compared with T.m. ssp. chambers with smooth walls. The branched
quinckeanum. In examined sample from thalus has terminal microconidia,
T.m. ssp. quinckeanum were not met spiral elongated/pyriform in shape. Microconidia
hyphae. Macroconidia in T.m. ssp. number is smaller than in the T.m. ssp.
mentagrophytes are multilocular with mentagrophytes.
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Fig. 2 – T. mentagrophytes ssp. mentagrophytes, macroconidia (A-100X), spiral hiphae (B- 40X),
spiral hiphae / macroconidia – general overview (C – 10X), microconidia/macroconidia (D – 100X).

Fig. 3 - T. mentagrophytes ssp. quinckeanum, macroscopic appearance of the colony (A - reverse; B –

obverse); macro/microconidia (C-100X).

In ESEM analysis could notice the mentagrophytes . In T. mentagrophytes ssp.

difference in colony density between the quinckeanum thalus is highly branched,
analyzed subspecies, the T. mentagrophytes branches are opening wide to thalus (Fig. 4
ssp. mentagrophytes colony has a uniform B-E). In T. mentagrophytes ssp.
appearance with a dense thalus, without quinckeanum the were observed spiral
spaces while the T. mentagrophytes ssp. hyphae, also (Fig. 5).
quinckeanum present discontinuity in the Microconidia in T. mentagrophytes ssp.
colony with a thalus multi-branched which mentagrophytes were arranged in grape
has many cordage (Fig. 4 A-D). In ESEM clusters, they were globular in shape while
investigations weren’t observed in the sp T. mentagrophytes ssp.
macroconidia. The thalus is without quinckeanum were elongated/pyriform and
branches with microconidia arranged in arranged in terminal ramifications.
clusters in T. mentagrophytes ssp.
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Fig. 4 – T. mentagrophytes ssp. mentagrophytes (A- mycelium general aspect; B – thalus with
microconidia; C- microconidia). T. mentagrophytes ssp. quinckeanum (D- mycelium general aspect;
E- branched thalus with terminal microconidia; F- microconidia detail).

Fig. 5 - T. mentagrophytes ssp. quinckeanum, thalus, spiral hiphae, microconidia.

Conclusions
By comparing the two ssp., they have In T. mentagrophytes ssp. mentagrophytes
the following differences: T. species were a greater number of
mentagrophytes ssp. mentagrophytes microconidia, spiral hyphae were observed
presents a less branched thalus with in greater numbers in T. mentagrophytes
abundant microconidia arranged in a cluster ssp. quinckeanum in ESEM investigations .
along the entire tall while in T. Colony aspect in T. mentagrophytes ssp.
mentagrophytes ssp. quinckeanum thalus is quinckeanum presents cordage and small
highly branched, branches are opening wide spaces between the mycelium hyphae while
to thalus. Microconidia are arranged on the in T. mentagrophytes ssp. mentagrophytes
terminal branches of thalus. the colony has a dense uniform appearance.
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Acknowledgements Houck H.E., Cooley J.E., Lowitt M.H, Kao

This work was cofinanced from the G.F., Tinea caput medusa; an unusual
European Social Fund through Sectoral presentation of Trichophyton mentagrophytes
Operational Programme Human Resources on the scalp. Cutis; 58:48-52. 1996.
Development 2007-2013, project number Ludwig R., Scanning Electron Microscopy:
POSDRU/89/1.5/S/63258 ”Postdoctoral school Physics of Image Formation and
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biotehnology based on the eco-economy and the 08372-3, 1998.
bio-economy required by eco-san-genesys”. Patrick E., Handbook of Sample Preparation for
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