Epidemiological study of reservoir hosts in an endemic area of zoonotic cutaneous leishmaniasis in Iran.

Epidemiological study of reservoir hosts in an endemic area of zoonotic cutaneous leishmaniasis in Iran M.R. Yaghoobi-Ershadi1 & E. Javadian2 The study was carried out in the central part of the Islamic Republic of Iran over a 12-month period in 1991-92 in Borkhar, a rural district lying north of Isfahan city. The objective was to determine the ecology of natural reservoir hosts of leishmaniasis for possible future field trials of leishmania vaccine. The main reservoir host in this area is Rhombomys opimus, the great gerbil, followed by Meriones libycus, the Libyan jird, and Hemiechinus auritis, the long-eared hedgehog. Of the 179 small mammals examined in the Borkhar area, the great majority were R. opimus (82.1%), then M. libycus (15.7%), and last H. auritis (2.2%). The highest rate of infection of R. opimus was in September (90.5%), the rate varying between 22.2% and 80.4% in different villages. The average infection rate of M. libycus was 17.9%. These rodents probably play an important role as reservoir hosts in the epidemiology of zoonotic cutaneous leishmaniasis in this area. Sixteen domestic and stray dogs appeared to be uninfected because examination showed no active lesion or scar. Introduction Zoonotic cutaneous leishmaniasis (ZCL) is endemic and very common in many rural areas of the Islamic Republic of Iran. Rhombomys opimus, the great gerbil, is the main animal reservoir in foci in the north-east and central part of the country; in the south-west, beyond the geographical distribution of R. opimus in the province of Khuzistan, the main animal reservoir is Tatera indica. In the southern part of Iranian Baluchistan, bordering Pakistan, the animal reservoir has been identified as Meriones hurrianae (1-8). Phlebotomus papatasi, the most important spe- cies of the genus Phlebotomus, is the only known vector to humans and gerbils in these foci. The para- site has been identified as Leishmania major (6, 9, 10). Zoonotic cutaneous leishmaniasis is a serious and increasing public health problem in the rural areas that lie north and east of Isfahan city in central Islamic Republic of Iran. Enzyme characterization of numerous isolates from R. opimus and patients with cutaneous leishmaniasis has revealed that I Vice-Dean, Department of Medical Entomology and Vector Con- trol, School of Public Health, Teheran University of Medical Sci- ences, P.O. Box 6446-14155, Teheran, Islamic Republic of Iran. Requests for reprints should be sent to this author. 2 Vice-Dean, School of Public Health, Teheran University of Medi- cal Sciences, Teheran, Islamic Republic of Iran. Reprint No. 5741 L. major zymodeme LON-1 is widespread in this area (6, 10). In 1991, we isolated Leishmania (Kines- toplastida: family Trypanosomatidae) promastigotes from P. papatasi and P. caucasicus collected from rodent burrows in the villages of Borkhar, a rural district lying north-east of Isfahan city. They were typed at the Faculty of Medicine in Montpellier, France, by the examination of 15 isoenzymes and identified as L. major Yakimoff & Schockov zymodeme MON-26 (10, 11). Human prevalence of the disease and the inci- dence of new cases remain high despite passive case detection and treatment by well-trained staff in local clinics and public health services. House spraying with DDT, poisoning of gerbils, and application of DDT powder in rodent burrows have failed to con- trol ZCL in this area (12-14). The study was carried out in four villages in Borkhar district in 1991-92 to determine the ecology of natural reservoirs and their relative importance and to obtain up-to-date information on the local epidemiology prior to future field evaluations of killed Leishmania major (KLm) vaccinea in this hyperendemic focus. a Tropical diseases, progress in research 1989-1990. Tenth programme report. Geneva, UNDP/World Bank/WHO Special Programme for Research and Training in Tropical Diseases, 1991. Bulletin of the World Health Organization, 1996, 74 (6): 587-590 © World Health Organization 1996 587 M.R. Yaghoobi-Ershadi & E. Javadian Materials and methods Characteristics of the study area The investigation was carried out over a period of 12 months (from 21 March 1991 to 20 March 1992) in four villages (Komshecheh, Parvaneh, Aliabadchi and parts of Habibabad; total population, 6284) in the district of Borkhar, 30-48km north of Isfahan city. The selected villages are part of an endemic focus of leishmaniasis in Isfahan. The soil of the area is gen- erally a little salty, and in some parts sandy (1). How- ever, wheat, barley, cotton, vines, beetroot, pistachio, alfalfa, Indian corn, clover and summer crops are cultivated. The underground water table is about 80m deep in this area (data from General Office of Agri- culture, Isfahan Province), which has a desert climate, and is very hot in summer and cold in winter. In 1991, the maximum and minimum mean monthly tempera- tures were 27.9°C and -1.2°C in June and December, respectively. The total annual rainfall in 1991 was 104.8mm minimum of 3mm in March and maxi- mum of 27.4mm in December. The minimum mean monthly relative humidity was 24.5% (August) and the maximum was 68% (December and January). Methods The selected villages were located about 1-1.5km from the colonies of gerbils. These rodents were trapped from the colonies using 40 live traps once a month by two experienced field technicians, and transferred to the leishmaniasis laboratory of Isfahan Training and Research Station. The gerbils were held from the back of the neck with forceps and from the tail by hand. Their ears, regardless of any ob- vious lesions, were cleaned with alcohol and scraped using a clean piece (2cm x 2cm) of coarse sand- paper. The surface of a clean slide was then pressed onto the scraped part of the ear and moved along to produce a smear on the slide. The slides were fixed with methanol, stained with standard Giemsa, and examined carefully under high magnification for an hour or more to look for amastigotes (15). Six smears were prepared and examined from each rodent. Samples from infected rodents were injected at the base of the tail of outbred or inbred (Balb/c) mice to identify the parasite. Household and stray dogs were physically examined for the presence of any active lesion or scar in a house-to-house survey. small mammals found in the Borkhar area is R. opimus, the great gerbil (82.1%); the other two are Meriones libycus, the Libyan jird or gerbil (15.7%), and Hemiechinus auritis, the long-eared hedgehog (2.2%). R. opimus and M. libycus are rodents in the subfamily Gerbillinae and H. auritis is an insectivore of the subfamily Erinaceinae, family Erinaceidae. There are many active colonies of R. opimus in the area, especially around the eastern part of Habibabad city. The characteristic vegetation is Salsola and many small trees (Haloxylon) have been planted by the Organization of Forests and Pastures to prevent movement of the flowing sands. Both R. opimus and M. libycus were found to be infected with Leishmania; 80 out of 147 R. opimus (54.4%) and 5 out of 28 M. libycus (17.9%) were positive. Leishmanial infection of R. opimus was found in all months during the study except for March. The highest rate of infection was 90.5% in September and the lowest was 25% in April (Fig. 1), which was statistically significant (P < 0.01). Of the 80 infected animals, 41 showed leishmanial infection on one ear and 39 on both ears. Most of the R. opimus examined were infected on both ears during September to November. The infection rate varied between 22.2% and 80.4% for R. opimus and 16.7% and 20% for M. libycus in different villages. The parasites were almost always scanty. Parasites from infected great gerbils were injected subcutaneously at the base of the tail of 32 laboratory mice (souris) and 9 Balb/c mice. Nodules and ulcers appeared after 18 days in souris and 7 days in the Balb/c mice. Sixteen domestic and stray dogs were examined and none of them appeared to be infected. Fig. 1. Percentage prevalence of leishmanial infection among Rhombomys opimus gerbils in Borkhar dis- trict, Islamic Republic of Iran, by month, during 1991- 92. 100 - 80- 0-- a60- C 540- & 20 Results A total of 179 small mammals were captured and examined. Predominant among the three species of 0- Apr May Jun Jul Aug Sep Oct Nov Dec Jan Feb Mar 1991 1992 0i Ito WHO Bulletin OMS. Vol 74 1996 I 588 Reservoir hosts of zoonotic cutaneous leishmaniasis in Iran Discussion Based on this survey, the great gerbil (R. opimus) appears to be the main reservoir host of zoonotic cutaneous leishmaniasis in this part of the Islamic Republic of Iran (area, 150km x 35km). The results showed that over 54% of these animals had L. major infection, which agrees with studies carried out in 1968 (1). Close contact between infected R. opimus and the human population makes it easy for infected sandflies to feed on humans and transmit the infection. The main human-gerbil vector is P. papatasi; P. caucasicus, P. mongolensis and P. ansarii act as vec- tors between rodents. This finding confirms previous data obtained by our group (10, 11, 16) and others (1). The highest infection rate in sandflies appeared in late August and September, accompanying the second and largest peak of sandfly activity in the area (16). This increased rate of infectivity among sandflies during these months seems to be due to close contact with many infected hosts. Young R. opimus gerbils become adults after the winter, and 25-33.3% of them are infected at the beginning of the active season of sandflies. Breeding starts in early spring and the population grows for 3 months. Transmission in this area begins when the sandflies emerge in late May, but cannot keep pace with the rising rodent population so that the propor- tion of infected sandflies declines. From July to Sep- tember the increase in the numbers of Rhombomys declines and the proportion of infected animals rises; the highest rate of R. opimus infection is therefore in September (Fig. 1) and the chances of infection in sandflies that bite them is then greater. About 62% of the inhabitants showed a scar of leishmanial infec- tion. The high risk of contracting the disease from infected sandflies is due to the existence of numerous rodent burrows. In our study of M. libycus, 5 rodents out of 28 (17.9%) were infected, which is a much higher rate than that found in an earlier study in 1968 (only 1 infected out of 38 (2.6%)) (1). Our result may be related to the technique we used to prepare smears from small mammals for the detection of Leishmania parasites, and also due to an increase in the rate of infectivity. The present level of leishmanial infection in this rodent shows that this animal also may play an important role as a reservoir host in the epidemiol- ogy of zoonotic cutaneous leishmaniasis in this area. It seems that hyperendemic zoonotic cutaneous leishmaniasis will remain a major public health prob- lem until new methods of intervention, such as an effective vaccine (17),b can be implemented. b See footnote a on page 587. Acknowledgements We are grateful to the leishmaniasis laboratory staff in the Isfahan Training and Research Station for assistance in collecting the rodents. This work was supported by the UNDP/World Bank/WHO Special Programme for Re- search and Training in Tropical Diseases (TDR) through grant No. ID-880635 for the purchase of equipment. Resume Etude epidemiologique des h6tes reservoirs dans une zone d'endemie de la leishmaniose cutanee zoonosique en Iran L'etude a ete r6alisee sur une p6riode de douze mois en 1991-1992 a Borkhar, un district rural situe au nord d'ispahan, dans le centre de l'Iran. Elle avait pour objectif de d6terminer l'6cologie des h6tes r6servoirs naturels de la leishmaniose en vue d'eventuels essais sur le terrain d'un vaccin antileishmaniose. Les principaux h6tes reservoirs rencontres dans la r6gion sont Rhombomys opimus, une gerbille, Meriones libycus, un merion, et Hemiechinus auritis, un h6risson. Sur 179 petits mammiferes examines dans la region de Borkhar, la plupart 6taient des R. opimus (82,1%), suivis de loin par M. libycus (15,7%) et H. auritis (2,2%). Le plus fort taux d'infection de R. opimus a 6te observ6 en septembre (90,5%) et variait de 22,2% a 80,4% d'un village a l'autre. Le taux moyen d'infection chez M. libycus 6tait de 17,9%. Ces rongeurs jouent probablement un r6le important en tant qu'h6tes r6servoirs dans 1'6pid6miologie de la leishmaniose cutanee zoo- nosique dans la r6gion. Seize chiens domestiques et errants examin6s n'ont en revanche montr6 aucun signe d'infection (ni l6sion ni cicatrice). References 1. Nadim A, Faghih M. The epidemiology of cutaneous leishmaniasis in the Isfahan province of Iran. I. The reservoir. II. The human disease. Transactions of the Royal Society of Tropical Medicine and Hygiene, 1968, 61: 534-542. 2. Nadim A, Mesghali A, Seyedi-Rashti MA. Epidemi- ology of cutaneous leishmaniasis in Turkemen- Sahara, Iran. Joumal of tropical medicine and hy- giene, 1968, 71: 238-239. 3. Seyedi-Rashti MA, Nadim A. Epidemiology of cuta- neous leishmaniasis in Iran. B. Khorassan area. Part I. The reservoir. Bulletin de la Societe de Pathologie exotique, 1967, 60: 510-518. 4. Javadian E et al. Epidemiology of cutaneous leishmaniasis in Iran. B. Khorassan. Part V. Report on a focus of zoonotic cutaneous leishmaniasis in WHO Bulletin OMS. Vol 74 1996 589 M.R. Yaghoobi-Ershadi & E. Javadian Esferayen. Bulletin de la Societe de Pathologie exotique, 1976, 69: 140-143. 5. Seyedi-Rashti MA, Salehzadeh A. A new focus of zoonotic cutaneous leishmaniasis near Teheran, Iran. Abstracts of VIl International Congress of Parasitol- ogy, Paris, France, 20-24 August 1990. Bulletin de la Soci6t6 fran9aise de parasitologie, 1990, supplement 2:1145. 6. Nadim A. Current situation with regard to leishmaniasis in the countries of the Eastern Medi- terranean region: an overview. In: Walton BC, Wijeyaratne PM, Modabber F, eds. Proceedings of an International Workshop on Research on Control Strat- egies for Leishmaniasis, Ottawa, Canada, 1-4 June 1987. Ottawa, International Development Research Center, 1987: 162-167 (mimeographed document No. 184e). 7. Javadian E. Reservoir host of cutaneous leishmaniasis in Iran. In: Abstracts of XII International Congress for Tropical Medicine and Malaria, Amster- dam, 18-23 September 1988. Amsterdam, Elsevier, 1988: 35. 8. Seyedi-Rashti MA, Nadim A. Cutaneous leishmaniasis in Baluchistan, Iran. Xl Intemational Congress for Tropical Medicine and Malaria, Calgary, Canada, 16-22 September 1984, Abstract and poster volume. ICCA, Calgary, 1984: 124. 9. Nadim A, Seyedi-Rashti MA. A brief review of the epidemiology of various types of leishmaniasis in Iran. Acta medica iranica, 1971, 15: 99-106. 10. Yaghoobi-Ershadi MR, Javadian E, Tahvildare- Bidruni GH. Leishmania major MON-26 isolated from naturally infected Phlebotomus papatasi (Diptera: psychodidae) in Isfahan Province, Iran. Acta tropica, 1995, 59: 279-282. 11. Yaghoobi-Ershadi MR, Javadian E, Tahvildare- Bidruni GH. The isolation of Leishmania major from Phlebotomus (Paraphlebotomus) caucasicus in Isfahan Province, Islamic Republic of Iran. Transac- tions of the Royal Society of Tropical Medicine and Hygiene, 1994, 88: 518-519. 12. Nadim A, Amini H. The effect of antimalaria spray- ing on the transmission of zoonotic cutaneous leishmaniasis. Tropical and geographical medicine, 1970, 22: 479-481. 13. Seyedi-Rashti MA, Nadim A. Attempts to control zoonotic cutaneous leishmaniasis in the Isfahan area, Iran. Iranian journal of public health, 1974, 2: 199- 202. 14. Seyedi-Rashti MA, Nadim A. Re-establishment of cutaneous leishmaniasis after cessation of anti- malaria spraying. Tropical and geographical medi- cine, 1975, 27: 79-82. 15. Edrissian GH, Zovein Z, Nadim A. A simple tech- nique for preparation of smears from the ear of Rhombomys opimus for the detection of leishmanial infection. Transactions of the Royal Society of Tropical Medicine and Hygiene, 1982, 76: 706-707. 16. Yaghoobi-Ershadi MR, Javadian E. Seasonal varia- tion of Leishmania major infection-rates in sandflies from rodent burrows in Isfahan Province, Iran. Journal of medical and veterinary entomology, 1996, 10: 181- 184. 17. Modabber F. Development of vaccine against leishmaniasis. Scandinavian journal of infectious dis- ease, 1990, 76 (suppl.): 72-78. 590 WHO Bulletin OMS. Vol 74 1996