Characterization of Leishmania major causing cutaneous leishmaniasis in northern Cameroon

TRANSACTIONS OFTHE ROYAL SOCIETY OFTROPICAL MEDICINE AND HYGIENE (1998)92,677-678 Six weeks after successful chemotherapy, as defined by zero CCA levels and egg counts, CSEA could still be detected in the urine samples of 7 cases. In one of these patients, CSEA could even be demonstrated until 12 weeks after treatment (data not shown). This indicates that not all S. mansoni tissue eggs are killed by praziquante1 (not even by the highest dose) and secretion of egg antigens proceeds for a considerable period. However, to be ableto interpret these findings fully, more research is needed on the life snan of eees, the duration of CSEA secretion by the eggs, and thYeY mechanisms regulating clearance of CSEA. Faecal egg counts and/or detection of adult worm antigens remain more suitable indicators of the efficacy of chemotherapy. However, we have demonstrated that assessment of the levels of antigens secreted by eggs provides more information about the number of eggs retained in tissues before and after chemotherapy. As such, measurement of egg antigen might be used as a marker for morbidity. Acknowledgements We thank the ward staff of US Naval Medical Research Unit no. 3 for collecting urine samples. This investigation was financially supported by the UNDP/World Bank/WHO Special Programme for Research and Training in Tropical Diseases (TDR) and by the Research and Development Programme ‘Life Science and Technology for Developing Countries (STD-3)’ of the European Communities. References Giboda, M. & Smith, J. M. (1994). Schistosoma munsoni eggs as a target for praziquantel: efficacy of oral application in mice. %urnal oiTro&al Medicine and Hwiene. 97,98-102. Nibbeling, H. A.“M.,kan Lieshout, L., I’~&an,~I<.,~Stelma, F.

/ Short Report

Characterization of Leishmania major causing cutaneous leishmaniasis in northern Cameroon Blaise DondjiLW, Jacques Dereurel, Francine Pratlongl, Dotchka D. Duhlinskaz, Albert SameEkobo3 and Jean-Pierre Dedetl* JLaboratoire d’Ecologie Midicale et de Pathologic Parasitaire, Fact& de M.& decine, Universite’ de Montpellier I, 163 rue Auguste Broussonet, 34090 Montpellier, France; ZApplied Entomology and Parasitology Unit, University of Jos, Jos, Nigeria; 3Laboratoly of Parasitology, Mycology and Parasite Immunology, Yaoundi University Teaching Hospital, l? 0. Box 3266, Yaoundi, Cameroon Leishma-

Introduction Cutaneous leishmaniasis (CL) occurs in various parts of the world, mainly in tropical and subtropical regions. In the African continent, CL due to Leishmania maior, L. tropica and L. aethiopiia is unevenly distributed fr% the northern to the southern areas of the continent. Northern

Cameroon

is one of the West African

en-

demic CL foci, especially Mokolo region (DESJEUX et aZ., 198 1). The first cases in Cameroon were reported in the 1930s (HERvB, 1937). Subsequently, DJIBRILLA et *Author for correspondence: [email protected]

F., Polderman, A. M. & Deelder, A. M. (1998). Serum circulating egg antigen levels in two areas endemic for Schistosoma munsoni. Transactions of the Royal Society of TropicaL Medicine and Hygiene, 92,350-354. Nourel Din, M. S. A., Kornelis, D., Van Zeyl, R. J. M. & Deelder, A. M. (1994a). Immunologic characterization of two monoclonal antibodies reactive with repetitive carbohydrate epitopes of circulating Schistosoma mansoni egg antigen. American Journal of Tropical Medicine and Hygiene, 50, 487-498. Nourel Din, M. S. A., Nibbeling, R., Rotmans, J. I?, Polderman, A. M., Krijger, F. W. & Deelder, A. M. (1994b). Quantitative determination of circulating soluble egg antigen in urine and serum of Schistosoma mansoni-infected individuals using a combined two-site enzyme-linked immunosorbent assay. American Journal of Tropical Medicine and Hygiene, 50, 585-594. Richards, F., jr, Sullivan, J., Ruiz-Tiben, E., Eberhard, M. & Bishop, H. (1989). Effect of praziquantel on the eggs of Schistosoma mansoni, with a note on the implications for managing central n&vous system schistosomi&is. Annals of Tropical Medicine and Parasitology, 83,465-472. Sabah, A. A., Fletcher, C., Webbe, G. & Doenhoff, M. J. (1986). Schistosoma mansoni: chemotherapy of infections of different ages. Experimental Parasitology, 61,294-303. Van Lieshout, L., De Jonge, N., El Masry, N. A., Mansour, M. M., Bassily, S., Krijger, F. W. & Deelder, A. M. (1994). Monitoring the efficacy of different doses of praziquantel by quantification of circulating antigens in serum and urine of schistosomiasis patients. Parasitology, 108, 519-526. Xiao, S. H., Catto, B. A. & Webster, L. T. (1985). Effects of praziquantel on different developmental stages of Schistosoma munsoni in vitro and in viva. Journal of Infectious Diseases, 151,1130-1137. Received 9 June 1998; revised 24 August 1998; accepted for publication 25 August 1998

al. (1979) made a prospective study between December 1974 and Februarv 1975 and renorted 58 cases of CL in Mokolo city and its environs. In an earlier survey, we reported 14 cases with active lesions and 162 with characteristic leishmanial scars (DONDJI et al., in press). We report here the first characterization of the parasite responsible for CL in the Mokolo endemic focus of northern Cameroon.

1

Keywords: leishmaniasis, cutaneous leishmaniasis, niu major, zymodeme characterization, Cameroon

677

fax +33 (0)4 67 63 00 49, e-mail

Materials and Methods Clinico-parasitological screening for CL was conducted in the Mokolo region throughout 1997. All the patients were interviewed and information on sex, age, residential area and travelling history was recorded. The clinical diagnosis of the disease was confirmed by the presence of parasites in smears after May-Gtinwald-Giemsa (MGG) staining. Parasite isolation was attempted but was successful with only one patient, an 8 years old girl from Zileng village in Mokolo Subdivision (100-200 m altitude). She had been living in the village since birth. She had 3 lesions on her left leg (1.5 cm, 2 cm and 4 cm in diameter). The lesions appeared within one week and had lasted for one month before our visit. Tissue specimens were collected, sliced in physiological saline and inoculated into the footpad and tail junction of 2 B.&B/c mice, which both developed a lesion 23 and 34 d, respectively, after inoculations. One mouse was transported to Laboratoire d’Ecologie MCdicale et Pathologie Parasitaire in Montpellier, France, where an aspirate of the lesion was successfully cultureh in NNN medium. Isoenzymatic typing of the isolate obtained (MHOMICMI97IMOKl) was nerformed by starch gel electrophoresis using i5 enzymes (FCIOUXet al., 1990) (Table). The isolate was compared with the following reference strains of L. major: MHOMUvlA43l/LEM265 (MON-25) and L. major MHOMNDI76ILm62 (MON-26).

678

BLAISE DONDJI ETAL.

Table. Isoenzymatic Cameroon Strain

profiles

MDH

MHOMIMAI81ILEM 265b MHOMNDI76ILEM62b MHOMICMI97IMOKl

of the reference

strains

of Leishmania

major

and

Enzyme mobilitya ME ICD PGD G6l’D GLUD DIA NPl NP2 GOT1 GOT2 PGM

150

88

160

88

160

88

100 100 100

122 122 122

94 94 94

200 200 200

100 400 90 100 400 90 100 400 90

110 110 110

120 110 110

118 118 118

of the new

isolate

from

FH

Ml’1

GPI

Zymodeme

87 79 79

150 150 150

77 77 77

MON-25 MON-26 IMON-26

aMDH=malate dehydrogenase (EC 1.1.1.37), ME=malic enzyme (EC 1.1.1.40), ICD=isocitrate dehydrogenase (EC 1.1.1.42), PGD=phosphogluconate dehydrogenase (EC 1.1.1.44), G6PD=glucose 6-phosphate dehydrogenase (EC 1.1 .1.49), GLUD= glutamic dehydrogenase (EC 1.4.1.3), DIA=diaphorase (EC 1.6.2.2), NPl, NP2=purine nucleoside phosphorylases (EC 2.4.2.1), GOTl, GOT2=glutamate oxaloacetate transaminases (=aspartate aminotransferase, EC 2.6.1. I), PGM=phosphoglucomutase (EC 5.4.2.2), FH=fumarate hydratase (EC 4.2.1.2), MPI=mannose phosphate isomerase (EC 5.3.1.8), GPI=glucose phosphate isomerase (EC 5.3.1.9). bReference strains. Results

and Comments

We recorded a total of 47 individuals with active lesions and 4 with characteristic leishmanial scars in the Mokolo region throughout 1997. Their ages ranged from 8 months to 70 years; 22 were males and 29 were females. The number of lesions varied from one to 10 and the mean was 3, as observed in earlier studies (DONDJI et al., in press). The lesions included ulcerative and nodular types, and sometimes satellite nodules appeared around the main lesion. These are common clinical signs of CL infection in this focus (DJIBRILIA et al., 1979). The diameter of the lesions varied from few millimetres to 7 cm. Most patients came from suburban and rural areas between 100 m and 700 m altitude. Isoenzymatic characterization showed that the isolate belonged to L. major zymodeme MON-26 (Table). L. major is the causative agent ofzoonotic cutaneous leishmaniasis in Asia and Africa, and zymodeme MON-26 has been reported from Senegal (DEDET et al., 1986), Mali (IZRI et al., 1989), the Arabian peninsula (MAAZOUN et al., 1986; PETERS et al., 1985), Israel and Iraq (MAAZOUN et al., 1986), Jordan (SALIBA et al., 1988), Iran (YAGHOOBI-ERSHADI et al., 1995) and Syria (IWIAMI et al., 199 1). Diffuse cutaneous leishmaniasis due to L. major MON-26 has also been recorded (DEVELOUX et al., 1996). Eco-epidemiological surveys, including biotic factors (flora, reservoir host, humans, parasite) and abiotic factors (geology, pedology, climatology), should be conducted in northern Cameroon in order to define more fully the transmission cycle of leishmaniasis and the factors involved. Such information is a basic requirement for the design of a successful control programme. Acknowledgements We thank Dr I.Yimagou and Dr E. Nogbe (District Hospital, Mokolo, Cameroon), Dr P. Martin (Centre Pasteur, Yaoundt, Cameroon), and Mr I’. Lami, Mrs C. Rouquairol and G. Serres (Laboratoire d’Ecologie Medicale et de Pathologie Parasitaire, Montpellier, France) for their valuable technical assistance, and Dr P. Desjeux (WHO, Geneva) for his advice. The research visit of B. D. to Montpellier was supported by Mission Francaise de Cooperation et d’Action Culturelle au Cameroun. The work described here formed part of a thesis approved by the University of Jos, Nigeria for a PhD degree awarded to B. D. References Dedet, J. P., Saf’janova, J. M., Le Blancq, S. M., Desjeux, I’., Schnur, L. F., Emelyanova, L. I? & Chance, M. L. (1986). Caracterisation de souches de Leishmania ma.orYakimoff et Schokkor 1914 isolees au Senegal (Afrique de I’Ouest). In:

Leishmania: Taxanomie et Phylogenbe. Applications lko@d& miolgiques, Rioux, J. A. (editor). Montpellier: IMEEE, pp. 463-469. Desjeux, I’., Waroquy, L. & Dedet, J. l? (1981). La leishmaniose cutanee en Afrique de I’Ouest. Bulletin de la Socie’te de Pathologic Exotique, 74,414425. Develoux, M., Diallo, S., Dieng, Y., Mane, I., Huerre, M., Pratlong, F., Dedet, J. I? & Ndiaye, B. (1996). Diffuse cutaneous leishmaniasis due to Leishmania major in Senegal. Transactions of the Royal Society ojFTropica1 Medicine and Hygiene, 90,396-397. Djibrilla, K. B., Ripert, C., Ravisse, l?, Durand, D. & Carrie, J. (1979). Etude epidtmiologique du foyer de leishmaniose cutanee de Mokolo (Nerd-Cameroun). Bulletin de la So&e de Pathologic Exotique, 72, 442-450. Dondji, B., Duhlinska, D. D. & Agwale, S. M. (in press). Leishmaniasis in Cameroon. 1. A preliminary survey in Mokolo Subdivision, Far North Province. AJricanJournaZ of NaturaE Sciences. Her+, (1937). Note sur la leishmaniose cutanee au Cameroun. Am-tales de Medecine et de Pharmacie Coloniales, 35, 928-934. Izri, M. A., Doumbo, O., Belazzoug, S. & Pratlong, E (1989). Presence de Leishmania major MON-26 au Mali. Annales de Parasitologic Humaine et Comparee, 64, 5 1O-5 11. Khiami, A., Dereure, J., Pratlong, F., Martini, A. & Rioux, J. A. (1991). La leishmaniose cutanee humaine a Leishmania major MON-26 aux environs de Damas (Syrie). Bulletin de la Societe’ de l’athologie Exotique, 84, 340-344. Maazoun, R., Pratlong, F., Lanotte, G. & Rioux, J. A. (1986). Le complexe Leishmania major. A propos de l’analyse numerique de 35 souches identifiees par la methode ezymatique. In: Leishmania. Taxonomie et Phylogendse. Applications Eco-epidimiologiques, Rioux, J. A. (editor). Montpellier: IMEEE, pp. 119-128. Peters, W., Elbihari, S., Ching Liu, Le Blancq, S. M., Evans, D. A., Killick-Kendrick, R., Smith, V. & Baldwin, C. I. (1985). Leishmania infecting man and wild animals in Saudi Arabia 1. General survey. Transactions of the Royal Society of Tropical Medicine and Hygiene, 79, 831-839. Rioux, J. A., Lanotte, G., Serres, E., Pratlong, F., Bastien, P. & Per&e, J. (1990). Taxonomy of Leishmania: use of isoenzymes. Suggestions for a new classification. Annales de Parasitologie Humaine et Comp&e, 65, 111-125. Saliba, E. K., Highashi, G. I., Yates, J. A. & Oumeish, 0. Y. (1988). Cutaneous leishmaniasis in Tordan: biochemical identification of human and Psammo&s obesus isolates as Leishmania major. Annals of Tropical Medicine and Parasitology, 82, 21-25. Yaghoobi-Ershadi, M. R., Javadian, E. & Tahvildare-Bidruni, G. H. (1995). Leishmania major MON-26 isolated from naturally infected Phlebotomuspapatasi (Diptera: Psychodidae) in Isfahan Province, Iran. Acta Tropica, 59, 279-282. Received 30 June 1998; revised 17 August for publication 18 August 1998

1998; accepted