Prelinzirwry and Short Report

Transcription

1 Prelinzirwry and Short Report ELECTRON MICROSCOPIC STUDY OF INFECTED HAIRS IN TRICHOMYCOSIS AXILLARIS* LEOPOLDO F. MONTES, M.D., CESAR VASQUEZ, M.D.' AND MARTA S. CATALDI, PH.D.' The etiologic agent of trichomycosis axillaris was isolated by Crissey et at. (1) from all 28 cases of the disease seen in the Philadelphia area during the summer months of The cultural, microscopic and growth characteristics, mouse pathogenicity, and antibiotic sensitivities of this organism were also studied. It was then considered to be a diphtheroid, and the name Corynebacterium tenuis was proposed for its designation. In the present paper we report the results of our attempt to observe the morphology of C. tenuis in infected hairs studied under the electron microscope. Material and Methods Multiple infected hairs were obtained from both axillae of a 45-year-old Italian male having the red variety of trichomycosis axillaris. The infected hairs were readily distinguished under the Wood's light by their orange fluorescence. KOH and gram stained preparations of the hairs confirmed the clinical diagnosis. The concretions showed numerous gram positive organisms. Some of the hairs were placed in Petri dishes prepared with blood agar and incubated at 37 C. Colonies developed rapidly and smears obtained at 48 hours were stained with gram as suggested by Crissey (1). A gram positive organism of diphtheroid form was the only one isolated. This bacteria fermented dextrose and its antibiotic sensitivity was similar to that reported by the Philadelphia group. In addition, we tested the effect of erythromycin in vitro, and this was highly inhibitory. For electron microscopic study parts of the hairs including the concretions were carefully selected under a stereomicroscope. They were cut in small pieces, fixed in acetate veronal buffered osmium tetroxide (ph 7.6) (2) for one hour, and included in methacrylate. In order to obtain both longitudinal and transverse sections of the hairs, these were oriented accordingly in the gelatin capsules. A Porter-Blum microtome was used for * From the Instituto de Biologia y Medicina Experimental, Obligado 2490, and the Laboratorio de Micologia, SecciOn de Dermatologia, Clinica Marini, Buenos Aires, Argentina. 'From the Instituto de Investigaciones Hematológicas, Melo 3081, Buenos Aires. Professor of Microbiology, Facultad de Ingenieria Quimica, Universidad de Buenos Aires. This study was supported in part by a research grant from The Upjohn Company. Erythromycin was kindly supplied by Eli Lilly and Co. of Argentina. Received for publication February 15, ultrathin sectioning. A Siemens Elmiskop I electron microscope was used for observation and photography of the sections. Some of these were stained with lead using both Karnovsky's (3) and Millonig (4) procedures. Observations Under the electron microscope the concretions appeared to be made up of bacteria almost entirely. The organisms were packed together and well preserved in the deeper layers, whereas near the surface they were arranged in a more loose pattern and many of them undergoing lysis (Figs. 1 and 2). The length of the organisms ranged between 1.2 and 1.6 microns, and its maximum diameter, around the nuclear zone was about 0.5 micron. Remnants of cell walls as described by Slade (5), were frequently observed (Fig. 5). Some longitudinal sections revealed the presence of C. tenuis in the superficial horny layers of the hairs (Fig. 1). The different parts of the ultrastructure of C. tenuis could be identified under high magnifications (Figs. 3 to 6). A cell wall, and a cell membrane as seen by Chapman (6) in corynebacteria and in other types of bacteria were observed. The nuclear material was located in the central portion of the organism. It was represented by a zone of lower density than the cytoplasm, containing a filamentous network and more dense chromatinlike particles (Fig. 4). Discussion Our study supports Crissey's observations that trichomycosis axillaris is caused by a diphtheroid organism. The fact that the bacteria are more numerous in the deeper layers of the concretions, and also observed in the superficial horny layers of the hair, would indicate a keratinophylic tendency in C. tennis. Further studies are required to confirm this theory. It will also be important to learn whether this organism is in any way related to the erythrasma dip htheroid of Taplin, Sarkany and Blank (7). The fluoroscence of the concretions, already recorded by Montgomery (8), and the in vitro response to erythromycin observed by us, is also interesting in this regard. The systemic use of this or any other antibiotic may well result in a disappearance of the bacterial concretions from the axillary hairs. Summary The results of some electron microscopic observations of infected hairs obtained from a 45 year-old male having the red variety of tricho-

2 274 THE JOURNAL OF INVESTIGATIVE DERMATOLOGY :4 9!, - t, Pd I /,##.1 FIG. 1. Longitudinal section of a hair showing numerous C. tenuis oriented parallel to the axis of the hair. The micro-organisms seem to be lying between the keratin layers. X 4,500.

3 ELECTRON MICROSCOPIC STUDY OF INFECTED HAIRS 275 *1 S : _. r Fm. 2. Cross section of a concretion. Many of the bacteria show well preserved cell walls and cell membranes. Some of them (upper right side of the picture), located near the surface of the concretion, are undergoing early lysis. X 22, Fm. 3. C. tenuis. X 22,500.

4 276 THE JOURNAL OF INVESTIGATIVE DERMATOLOGY Cw N CM FJG. 4. The different parts of the ultrastructure of C. tenuis are illustrated. Cell wall (CW), cell membrane (CM), nuclear zone (N) are clearly seen. Many organisms are observed in cross section near their tips (T). X 60,000.

5 ELECTRON MICROSCOPIC STUDY OF INFECTED HAIRS FIG, 5. Almost longitudinal section of a C. tenuis. Remnant of a cell wall (CW) is a present. X 60, FIG. 6. Electron micrograph of a C. tenuis apparently undergoing early lysis. Rupture of the cell wall and cell membrane is taking place around an area of low density which probably represents the nuclear zone (N). X 107,000.

6 278 THE JOURNAL OF INVESTIGATIVE DERMATOLOGY mycosis axillaris, are reported. Our study confirms the existing data with reference to the bacterial origin of the disease. REFERENCES 1. CRJSSEY, J. T., RENaL, C. C. AND LASKAS, J. C.: Studies on the causative organism of trichomycosis axillaris. J. Invest. Derm., 19: 187, PALADE, G. E.: A study of fixation for electron microscopy. J. Exp. Med., 95: 285, KARNOvSKY, M. J.: Simple methods for "staining with lead" at high ph in electron microscopy. J. Biophys. Biochem. Cytol., 11: 729, MILLONIG, G.: A modified procedure for lead staining of thin sections. J. Biophys. Biochem. Cytol., 11: 736, SLADE, H. D. AND SLAMP, W. C.: Studies on Streptococcys pyogenes. V. Biochemical and microscopic aspects of cell lysis and digestion by enzymes from Streptomyces albus. J. Bact., 79: 103, CHAPMAN, C. B.: Electron microscopy of ultrathin sections of bacteria. J. Bact., 78: 96, TAPLIN, D., SARKANY, I. AND BLANK, H.: The erythrasma diphtheroid: a bacteriological study. J. Bact. (in press). 8. CRI55EY, J. T. AND MURRAY, P. F.: Trichomycosis axillaris. New York J. Med., 54: 2841, 1954.