S91 Contributions to and review of dicrocoeliosis, with special reference to the intermediate hosts of Dicrocoelium dendriticum M.Y.MANGA-GONZA; LEZ*, C.GONZA; LEZ-LANZA, E.CABANAS and R.CAMPO Consejo Superior de Investigaciones CientıTficas (CSIC), EstacioT n AgrıTcola Experimental, Apdo. 788, 24080 LeoT n, Spain An epidemiological study on dicrocoeliosis caused by Dicrocoelium dendriticum was carried out on sheep, molluscs and ants in the mountains of Leo! n province (NW Spain) between 1987–1991. The results concerning the intermediate hosts and a review of some aspects of dicrocoeliosis are summarized. Mollusc collection for the helminthological study was random throughout the study area at fortnightly intervals. Twenty-nine Gastropoda species were identified. D. dendriticum infection was only detected in 2n98% of the 2084 Helicella itala examined and in 1n06% of 852 H. corderoi. The highest infection prevalence was detected in H. itala in September and in H. corderoi in February. Daughter sporocysts with well- developed cercariae predominated in spring and autumn. Infection prevalence increased with mollusc age and size. Ants were collected from anthills or plants to which they were attached. The behaviour of ants in tetania was followed. Twenty- one Formicidae species were identified, but only the following harboured D. dendriticum: Formica cunicularia (1158 examined specimens, 0n69% infection prevalence, 2–56 metacercariae per ant); F. sanguinea (234, 1n28%, 2–63); F. nigricans (1770, 4n97%, 1–186); F. rufibarbis (288, 6n59%, 2–107). In a flat area close to Leo! n town, 95n39% of the 2085 F. rufibarbis specimens collected in tetania contained metacercariae (1–240) in the abdomen. These were used for parasite characterization by isoelectric focusing and to infect lambs and hamsters. Only one brainworm per ant was found. Key words: Dicrocoelium dendriticum, mollusc and ant intermediate hosts. Sudarikov et Ryjikov, 1951). D. dendriticum is found in America, Asia, North Africa and Europe (Malek, Dicrocoelioses are parasite infections caused by the 1980) including practically all of the Iberian Pen- species of the genus Dicrocoelium Dujardin, 1845 insula (Cordero et al. 1994). The second, the third (Trematoda, Digenea), although mainly by Dicro- and the fourth species have been found in Africa coelium dendriticum (Rudolphi, 1819) Looss, 1899. (Lucius, 1981), Asia (Tang et al. 1983) and the old This parasite, which cycles in land molluscs and Soviet Union and Austria (Hinaidy, 1983), respect- ants, is located in the bile ducts and gall bladders of ively. numerous species of domestic and wild mammals, This paper mainly concerns dicrocoeliosis mainly ruminants, which act as definitive hosts in produced by D. dendriticum as it is the most several countries in Europe, Asia, America and widespread amongst the ruminants of several North Africa (Malek, 1980). D. dendriticum is a very countries. For a long time this parasite was confused common species in ruminants in the Iberian pen- with an immature form of Fasciola hepatica, as both insula (Cordero, Castan4 o! n & Reguera, 1994). More- trematodes are frequently found together in the liver over, this parasite can also occasionally infect of ruminants. Its description thus came late. The humans (Mohamed & Mummery, 1990). Infection synonymy of this parasite is complex, due to the of the definitive hosts occurs by ingesting the ants different generic and specific denominations received which harbour infective metacercariae. (Mapes, 1951; Schuster, 1987). The genus Dicrocoelium is included in the sub- Dicrocoeliosis causes irritation of the mucosa of family Dicrocoeliinae Looss, 1899, of the Dicro- the large bile ducts in definitive hosts. This explains coeliidae Family Odhner, 1911. According to La the proliferation and increase in the secretion of the Rue (1957), this family belongs to the superfamily glandular cells as well as cholangitis and Plagiorchioidea, suborder Plagiorchiata, order cholangiectasis of the septal and hepatic bile ducts, Plagiorchiida and superorder Epitheliocystida. The granulomatous type portal hepatitis associated with most important species of this genus which infect portal, septal and perisinusoidal fibrosis and vacuolar ruminants are: Dicrocoelium dendriticum; D. hospes degeneration of the hepatocytes (Dhar & Singh, Looss, 1907, D. chinensis Tang & Tang, 1978 and D. 1963; Wolff, Hauser & Wild, 1984; Sanchez- suppereri Hinaiday, 1983 (syn. D. orientalis Campos et al. 1996; Ferreras et al. 1997). This disease is generally chronic and the immune response * Corresponding author: Tel: j34 987 317156. Fax: does not protect from reinfection. Experimental j34 987 317161. E-mail: y.manga!eae.csic.es studies carried out in lambs infected with 1000 and Parasitology (2001), 123, S91–S114. " 2001 Cambridge University Press DOI: 10.1017\0031182001008204 Printed in the United Kingdom M. Y. Manga-GonzaT lez and others S92 3000 D. dendriticum metacercariae showed that the (sometimes 2 or 3), called the ‘‘brainworm’’, settles first egg elimination took place between days 49 and in the suboesophageal ganglion of the ant and the 79 post-infection (Campo, Manga-Gonza! lez & rest become metacercariae in the abdomen. When Gonza! lez-Lanza, 2000), while the first detection of the temperature falls, the brainworm alters the IgG antibodies by ELISA technique was observed behaviour of the ant by causing tetania of its on day 30 p.i. Maximum antibody levels were mandibular muscles. Due to this the ant remains obtained 60 days p.i. and remained high until the temporarily attached to grass and this promotes experiment ended 180 day p.i. (Gonza! lez-Lanza et ingestion by the definitive host. The mature ab- al. 2000). Moreover the studies of the liver and dominal metacercariae excyst in the intestine, the hepatic lymph nodes (from the lambs slaughtered 2 young flukes migrate to the liver through the opening months p.i.) immunolabelled by avidin-biotin com- of the common bile duct (sometimes the portal plex system showed that the parasite induced a circulation) and become adult worms in the bile humoral and cell-mediated local immune response duct. When these are mature, they lay eggs which that contributed to the inflammation observed but exit in the faeces of the host and this allows the life did not seem effective for the destruction of the cycle to begin again. A more detailed description of parasite (Ferreras et al. 2000). the adult worm and of the different stages of the The economic and health significance of parasite can be found in Manga-Gonza! lez (1999) and dicrocoeliosis is partly due to the direct losses in Manga-Gonza! lez & Quiroz-Romero (1999). occasioned by the confiscation of altered livers (Del The role played by molluscs in the epidemiology Rio, 1967; Lukin, 1980; Karanfilovski, 1983) and of dicrocoeliosis is very important as D. dendriticum also the indirect ones caused by the digestive egg hatching and miracidium liberation only occur disorders derived from the hepatobiliary alterations in the intestine of the molluscs that act as in- caused by these parasites, such as decreased animal termediate hosts. Moreover, the parasite multiplies weight (Boray, 1985), growth delay (Hohorst & enormously by asexual reproduction inside them La$ mmler, 1962), reduced milk production (Cavani et (numerous cercariae can be formed from one al. 1982), amongst others. Moreover, the additional ingested egg). This increases the possibilities of costs incurred by the application of anthelminthic parasite transmission. Since Piana (1882) first treatments, to which the animals must be subjected, encountered the long-tailed cercaria in Helix have to be considered. carthusiana (l Monacha (M.) cartusiana), which The life cycle of Dicrocoelium dendriticum is was later described by Von Linstow (1887) as extremely complex because land molluscs and ants Cercaria vitrina (from Zebrina detrita) and associated are required as first and second intermediate hosts, with Dicrocoelium dendriticum, many studies have respectively. Until Krull & Mapes (1952, 1953) been carried out to discover the mollusc species managed to complete the life cycle for the first time, which act as first intermediate hosts for this parasite. numerous studies were carried out over more than a More than 100 mollusc species (Gastropoda, century to try elucidate it (reviewed by Mapes, 1951 Pulmonata, Stylommatophora) have been found and Del Rı!o, 1967). The adults of the genus D. receptive to D. dendriticum under natural and dendriticum live in the liver and bile ducts of the laboratory conditions (some of them are mentioned definitive hosts where they lay their embryonated in the Discussion below). It can be deduced from eggs which pass through the intestine to be this that D. dendriticum shows markedly little eliminated in the faeces. Egg hatching and specificity as regards its first intermediate host. In miracidium liberation only occur in the intestine of addition the parasite can develop in various mollusc numerous species of land molluscs that act as first species in the same area: Manga-Gonza! lez (1987, intermediate hosts. The miracidium penetrates the 1992) found 11 species of Helicidae infected with the intestinal wall of the mollusc and settles in the parasite in the province of Leo! n (Spain). The life hepatopancreas, where it becomes a mother spo- history of the mollusc intermediate hosts is of great rocyst, which takes the shape of the spaces between epidemiological interest, as regards both the in- hepatopancreatic lobules because it has