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Clinico-Histopathological Findings of Buruli Ulcer Milanga

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Clinico-Histopathological Findings of Buruli Ulcer Milanga Jpn.J.Leprosy 69, 93-100(2000) Clinico-histopathological Findings of Buruli Ulcer Milanga Mwanatambwe'), Yukiko Fukunishi2), Mikihisa Yajima3), Keiji Suzuki3), Kingsley Asiedu4), Samuel Etuafel5)., Nobutaka Yamada') and Goro Asano') 1)Department of Pathology, Nippon Medical School,Japan 2)National Sanatorium, Matsuoka Hoyo-en, Aomori, Japan 3)National (Sanatorium, Tama-Zenshoen,) Higashi murayama, Japan 4)Global Tuberculosis Program, WHO, Geneva, Switzerland 5)St. Martin Hospital, Agroyesum, Ashanti Country, Ghana [Received:20 Apr 2000/ Accepted:6 June 2000] Keywords: BuruliUlcer, Histopathology, Mycobacterium ulcerans, Acid-fast bacilli (AFB) Here,we reportthe clinico-pathologicalfindings of Buruliulcer. The patientswere 2 females,9 and 23 yearsof age and one male,47 yearsof age fromthe Ashanti Country of Ghana.Clinically, cutaneous lesions wereclassified as nodular,ulcero-nodular and ulcerative.Histopathologically, lesions involved cutaneous and subcutaneoustissue, which showed lympho-epithelioid cell proliferationand panniculitiswith characteristic fat necroticchanges. Vascular inflammation, with the nervetissue involvement, are prominent features on the chronologicalspectrum of the 3 cases.In all butthe earlycase, Mycobacterium ulcerans could be visualized fromthe mid dermalarea to the subcutisby Fite-Faracoand Haradastain. The ulceratedlesions were also immunoreactiveto phenolicglycolipid-I(PGL-I). These findings suggest Mycobacterium ulcerans infection withlesions of differentages. Further,we also showthe needto identifydistinct caracteristics for differential diagnosiswith lesions caused by othermycobacteria. Introduction world, especially in western Africa (Fig. 1). The dis ease is caused by Mycobacterium ulcerans, which is Buruli ulcer is a skin ulcerating disease, affect closely related to the mycobacteria of tuberculosis ing children and adults of both sex, without distinc and leprosy. Buruli ulcer is the third most frequent tion of age, but particularly prevalent in children be mycobacterial disease after tuberculosis and leprosy. tween 2 and 14 years of age. The disease, described In endemic areas, only experienced medical practi a century ago in Ouganda Buruli county', is re tioners would be able to make accurate differential emerging at an alarming rate in some parts of the diagnosis between lesions of Buruli ulcer, and those caused by other microorganisms like, cutaneous lep 'Corresponding author : rosy, cutaneous tuberculosis, spirochetiosis, Department of Pathology Nippon Medical School, Bunkyo-ku, Sendagi 1-1-5 diphteriosis, actinomycosis, fusobacillosis, filariosis Tokyo 113 0086, Japan and leishmaniosisz). The culture of Mycobacterium 93 ulcerans or the confirmation of its genomic sequence by Buruli ulcer's patients. The 3 patients did not re by PCR will not be a practical diagnostic method in ceive any other medication prior to the surgical pro most endemic countries in the near future. Histopa cedure. From the surgically removed materials, the thology, if supported by well defined criteria, will specimen for histopathology were taken from the remain the gold standard diagnostic method. most active area of the lesions. Specimens from each patient were then divided in 2 separate portions. One Materials and Methods set of specimen was fixed in 10% neutral formalin and the other in 2.5% glutaraldehyde solution in 0.1M The skin specimen in this study, were obtained in cacodylate buffer at pH 7.4. The formalin fixed speci the course of skin reparative surgery, from 3 patients mens were routinely processed and stained with He admitted at the Agroyesum missionary hospital in matoxylin-eosin(H&E), Fite-Faraco and Harada re the Ashanti Country in Ghana. The clinical data of agents. For the phenolyc glycolipid-1(PGL-1) the 3 patients are summarized in Table 1. The surgi immunostaining, paraffin blocks of the formalin cal excisions were performed under local anesthesia fixed specimen were sectioned into 3 um sections according to a surgical procedure in use atAgroyesum and processed for PGL-1 immunostaining using the St. Martin Hospital. All the cases were admitted to ABC method. this hospital with the presumptive diagnosis of Buruli ulcer. These presumptive diagnoses were based on Results the endemicity of the area, the clinical history of the diseases, the physical examination of lesions and the The specimens from all 3 cases showed characteris experience of the 3 medical practitioners in charge tic histopathologic changes as a result of Mycobac of this 80 beds' hospital of which 1/3 are occupied terium ulcerans infection. In case 1, macroscopically Worlddistribution ofBuruli Ulcer (-), withthe highest prevalence inAfrica inthe last decade(). Fig.i. World distribution of Buruli Ulcer, showing the highest prevalence in Africa 94 Jpn.J.Leprosy 69, 93-100(2000) a single nodule with firm consistency was seen on served in perivascular spaces from the papillary der the posterior aspect of the left elbow (Table 1). Mi mis down the reticular dermis. In the subcutis, in croscopically, inflammatory cell infiltration was ob flammatory cells were scantly scattered in the adi Table I. Summary of patients' clinical data Table II. Summary of histopathological findings of the 3 cases 95 pose tissue (Fig.2a). Ultrastructurally, blood vessels able inflammatory cell infiltration was evident in fatty showed hypertrophic endothelial cells with vacuoliza tissue (Fig 3b). Some blood vessels showed the reca tion and perivascular increase of collagen fibrils (Fig. nalization of thrombi with collagenous degeneration 2b). In case 2, macroscopically an ulcerated nodule of their walls. In the perivascular fibrotic region, on the upper, anterior aspect of the left arm was noted. slight inflammatory cell infiltration was evident. Histologically, the reticular dermis showed coagula Scattered acid-fast bacilli (AFB) were also present tive necrosis of the connective tissue along with pan in these blood vessel walls (Fig.3c). Macroscopically, niculitis (Fig 3a). Perivascular edema with remark case 3 showed an ulcer on the lateral aspect of the Fig.2a Inflammatory cell infiltrationis observed in fatty tissue of Fig.3a.Coagulative necrosis of the connective tissue and the subcutis in case 1.(H&Estain x 200) panniculitis are shown in this picture of the reticular Fig.2b.Bloodvessels showing hypertrophic endothelial cells with dermis of case 2. (H&E stain x 200) vacuolization and increase perivascular collagen fibrils Fig.3b. Remarkable perivascular edema and lesions of vas in this ultrastructural picture of case1.(x 3600) culitis are demonstrated in the adipose tissue of this evolutive case 2.( H&E stain x 200) Fig.3c.A blood vessel shows slight inflammatory cell infiltra tion in perivascular fibrotic region in case 2.(Fite Faraco stain x 200) 96 Jpn.J.Leprosy 69, 93-100(2000) right upper arm. Microscopically, coagulative necro ters of AFB in the necrotic reticular dermis and the sis of all layers of the dermis in the ulcer bed was subcutis (Fig.4b). Ultrastructurally, various degen evident. Ghost adipocytes were scattered in the sub erative changes in the bundles of peripheral nerves cutis. Fibrotic granulomatous lesions, including were observed, associated with thickening of multinucleated giant cells, epithelioid cells, histio Schwann cell basement membrane and vacuolization cytes and lymphocytes were observed in the margins of axons. Mast cell infiltration was also visible of ulcer bed (Fig.4a). AFB stains (Fite-Faraco and (Fig.4c). Also, the lumen of some blood vessels Harada) demonstrated small globi and confluent clus showed blood stasis, endothelial degeneration and Fig.4a.The margins of the ulcerative lesion of case 3 show a Fig.4c. Ultrastructure of peripheral nerve in case 3: De granulomatous lesion comprising multinuclear giant generative change of peripheral nerve, consistent cells, hystiocytes, epithelioid cells and lymphocytes with thickening of Schwann cell basement mem (H&E stain x 200). brane and axons' vacuolization, associated with Fig.4b.ln the reticular dermis of case 3, the main confluent mast cell infiltration(x 4300). cluster of AFB are seen in foamy structures (Harada Fig.4d.Ultrastructure of blood vessel in case 3: The lumen stain x 200). of a blood vessel of case 3, shows blood stasis and endothelial degeneration and Mycobacterium ulcerans can be visualized as round-shape dense bodies (x 5800; arrow). 97 contained, Mycobacterium ulcerans, as round-shape of mycobacterial infection, are rarely visualized in dense bodies (Fig.4d, arrow). The histopathological lupus vulgaris like in early cases of Buruli ulcer'. In findings of the 3 cases are summarized in Table 11. advance cases of Buruli ulcer, most of the well ac Immunohistochemically, PGL-1 was positive in the tive looking AFB were organized in confluent globi subcutis in case 2 and case 3. However, it was nega like formations or what has been designated foamy tive in case 1. structures. Small tufts or individual AFB could be seen up to the reticular dermis by the Harada stain. Discussion They were not found by either of our AFB stains (Fite Faraco and Harada stains) in case I of our report. On a chronologic spectrum, lesions described in these The absence of AFB in early cases and, the re-epi 3 cases suggest a continuous degenerative process, thelialization stage of any form of Buruli ulcer, like advancing toward an extended necrosis of tissue, from in case 3, with fibrosis and granuloma, may render superficial to deep dermal layers down to the subcu the differential
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