Raquel Campo

Contributions to and review of dicrocoeliosis, with special reference to the intermediate hosts of Dicrocoelium dendriticum

An epidemiological study on dicrocoeliosis caused by Dicrocoelium dendriticum was carried out on sheep, molluscs and ants in the mountains of León 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 2.98%, of the 2084 Helicella itala examined and in 1.06% 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, 0.69% infection prevalence, 2-56 metacercariae per ant); F. sanguinea (234, 1.28%, 2-63); F. nigricans (1770, 4.97%, 1-186); F. rufibarbis (288, 6.59%, 2-107). In a flat area close to León town, 95.39% 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.

IgG antibody response to ES or somatic antigens of Dicrocoelium dendriticum (Trematoda) in experimentally infected sheep

Abstract The ELISA technique was used to study the kinetics of IgG antibodies against excretory–secretory (ES) and somatic (So) antigens of Dicrocoelium dendriticum in the sera of 32 lambs: 12 experimentally infected with 1,000 metacercariae, 12 with 3,000 and 8 controls. Both antigen types were used at a 2 μg/ml concentration. Dilutions of sera and the anti-sheep IgG peroxidase conjugate were: 1:200 and 1:6,000, respectively. Optical density values for each type of antigen in the two infected groups were higher than the cut-off point from day 30 post infection (p.i.), showing positive infection. Maximum antibody levels were observed 60 days p.i. and remained high until the experiment ended 180 days p.i. This pattern was similar for both ES and So antigens, although with slightly lower figures in the latter. Antibody kinetics were very similar for each of the two doses, except on day 30 p.i. No correlation between the antibody level and parasite burden could be established.

Preliminary study on genetic variability of Dicrocoelium dendriticum determined by random amplified polymorphic DNA

Genetic variability of adult specimens of Dicrocoelium dendriticum has been studied using random amplified polymorphic DNA (RAPD). The worms were collected from the infected livers of different sheep from several localities in León province (NW Spain). DNA fragments were amplified by means of decamer primer oligonucleotides of arbitrary sequence. Some primers produce complex and highly variable patterns of amplified DNA in D. dendriticum. Intra- and inter-population genetic variability of adult parasites were analyzed, scoring polymorphic and monomorphic reproducible bands by means of the Jaccard similarity, and dendrograms showing genetic relationships between individuals were obtained using the FITCH method. Genetic variability seems to be high in this parasite and genetic similarity within a population (worms infecting a single animal) is similar to the average similarity between worms from different sheep. These results suggest that each sheep is infected by numerous genetically different parasites from one or more populations of infected ants.

Characterization of adult Dicrocoelium dendriticum by isoelectric focusing

Water soluble extracts of 3131 adult specimens of Dicrocoelium dendriticum from cattle, sheep and goats, mainly from León province, were analysed by isoelectric focusing in thin-layer polyacrylamide gels. Activity of the following enzymes was studied: lactate dehydrogenase (LDH, EC 1.1.1.27), glucose phosphate isomerase (GPI, EC 5.3.1.9), phosphoglucomutase (PGM, EC 2.7.5.1), acid phosphatase (AcP, EC 3.1.3.2), alpha-glycerophosphate dehydrogenase (alpha-GPDH, EC 1.1.1.8), hydroxybutyrate dehydrogenase (HBDH, EC 1.1.1.30) and malate dehydrogenase (MDH, 1.1.1.37). Five distinct enzyme types were recognized for LDH (pH range 6.30-7.13), GPI (pH 6.13-6.80) and PGM (pH 6.20-6.60) whereas AcP showed three different patterns (pH 5.70-5.92). Weak and diffuse activity was detected for MDH (pH 4.8-6.2) and no activity was observed for alpha-GPDH and HBDH. In general, little phenotypic variation was observed between worms recovered from a single host, between those from hosts of the same species and between those from hosts of different species, although some enzyme types were found in some animals but not others. Nevertheless, it must be taken into account that most parasites came from sheep and also from a relatively small area in north-west Spain.

Larval development of Dicrocoelium dendriticum in Cernuella (Xeromagna) cespitum arigonis under controlled laboratory conditions

The larval development of Dicrocoelium dendriticum (Digenea: Dicrocoeliidae) in experimentally infected Cernuella (Xeromagna) cespitum arigonis (Schmidt, 1853), a species of mollusc important in the epidemiology of dicrocoeliosis in Spain, has been studied. A total of 948 specimens of this mollusc, distributed in five batches, were tested with individual doses of 50 to 150 parasite eggs, obtained from sheep, after 4 days without food. After infection these molluscs and control specimens were kept in an environmental simulation chamber at 20 degrees C, 50% relative humidity and 7 h of light per day. To detect the parasite, a minimum of six molluscs were examined every 20 days from day 1 post-infection (p.i.). The eggs of D. dendriticum were eliminated in the molluscan faeces 48 h post infection. The percentages of molluscs harbouring the parasite ranged between 17.53% and 75%. Daughter sporocysts with undifferentiated germinal masses and occupying very reduced areas of the hepatopancreas were observed 50 days p.i. and in the period immediately following. After 110 days p.i. sporocysts with cercariae at different stages of development were found although slimeball emission was never observed.