The serotoninergic and peptidergic components of the nervous system of the cercaria of Sanguinicola inermis (Digenea: Sanguinicolidae) were examined using whole-mount immunocytochemistry and a plan of the nervous system has been described. Antibodies to serotonin (5-hydroxytryptamine, 5-HT) and the neuropeptides, FMRFamide, GFNSALMFamide (S1) and SGPYSFNSGLTFamide (S2) were used in the study. Immunoreactivity (IR) was demonstrated to all but the S2 antisera and showed a similar fundamental distribution. IR was found in paired cerebral ganglia located anteriorly within the body and connected by a cerebral commissure. From the ganglia paired ventral and dorsal longitudinal nerve cords extend anteriorly into the cephalic organ and into the body. There is no apparent connection with the tail. Several transverse commissures connect the longitudinal nerve cords throughout the body and several associated cell bodies have been located. A double-stranded dorsal and ventral longitudinal nerve cord extends the length of the tail and six cell bodies are associated with these cords, uniquely demonstrating either FMRFamide and S1, or 5-HT-like IR. Only 5-HT-like IR was found to extend into the posterior tail furcae and there appears to be a lack of any peripheral tegumental innervation. Double-labelling experiments suggest that the serotoninergic and peptidergic components of the cercarial nervous system are distinct.

The interaction between Bothriocephalus acheilognathi Yamaguti, 1934 (Cestoda)and pronephric lymphocytes of carp, Cyprinus carpio L. was studied by examining proliferation of lymphocytes isolated from both naïve fish and fish injected intraperitoneally with cestode extract. Lymphocytes from naïve hosts were stimulated to proliferate in the presence of the extract depending upon the extract protein concentrations; lower concentrations (0.01–0.05μg/ml) induced the greatest response, and immunosuppression occurred at higher concentrations. Significant differences were noted in fish that received intraperitoneal injections of parasite extracts. Five days post-injection, lymphocyte proliferation was significantly greater in these individuals compared with sham injected or untreated controls. This difference was reduced at 10 days post-injection, although the response was dependent on the concentration of the parasite extract. The possible significance of the observed stimulation/suppression of lymphocyte activity to establishment of the parasite in the wild is discussed.

Metacercariae of Steganoderma formosum Stafford, 1904 infected 17 of 358 Acadian hermit crabs (Pagurus acadianus) on the Scotian Shelf east of Nova Scotia, Canada, in the Northwest Atlantic Ocean in 1989, 1990 and 1995. Overall prevalence and abundance in P. acadianus on the Scotian Shelf were 5% and 0.4 metacercariae per crab, with a maximum intensity of 56. The highest prevalence (34%) and abundance (2.9) were recorded in the winter of 1989. All infected hermit crabs were greater than 9 mm in cephalothorax length. The parasite was not found in Acadian hermit crabs (n = 74) from Georges Bank, nor in any hairy hermit crabs (P. arcuatus and P. pubescens) on the Scotian Shelf (n = 808), on Georges Bank (n = 14), or in the northern Gulf of St. Lawrence (n = 87). A single sand shrimp (Crangon septemspinosa) of 1254 collected from the Scotian Shelf in 1989 and 1990 was infected with a metacercaria of S. formosum. None of 421 pink shrimps (Pandalus spp.) collected from the Scotian Shelf in 1989 and 1995 was infected. Male and female reproductive systems were well developed and spermatozoa were observed in the seminal receptacles of most parasites, suggesting that these metacercariae are progenetic. The distribution of the parasite among potential intermediate hosts suggests that S. formosum demonstrates greater specificity for its second intermediate host in the Northwest Atlantic than it does in the Pacific or in the Northeast Atlantic oceans.

The successful colonization, establishment and spread of Pomphorhynchus laevis in a small, isolated, Devon river 128 km from the parasite's nearest focus in Dorset was followed over 11 years from 1985. The parasite was first detected in Anguilla anguilla and Platichthys flesus in 1988: by 1995 it had attained prevalence levels of 22.6% in A. anguilla and 43.6% in P. flesus and also occurred in 100% Salmo trutta, 50% Cottus gobio and Noemacheilus barbatulus. As judged by prevalence, abundance, proportion of females gravid and weight of gravid females, S. trutta was the preferred definitive host although C. gobio was a suitable host and may play a role in cycling the parasite: the other three species were unsuitable hosts. The intermediate host was the freshwater Gammarus pulex: the euryhaline G. zaddachi was not infected. On biological grounds, the P. laevis could be assigned to the English freshwater strain and was almost certainly introduced to the river by anthropochore stocking of S. trutta from a Dorset hatchery. The findings demonstrate conclusively that the English strain of P. laevis can colonize and establish in a new locality from which Leuciscus cephalus and Barbus barbus, its normal preferred hosts, are absent and use S. trutta instead. The results also confirm that P. laevis is a poor natural colonizer and appears always to be introduced to new localities by anthropochore transfers of fish. The implications of these conclusions for understanding the present distribution of P. laevis are discussed and it is considered that they provide direct evidence in support of the hypothesis that P. laevis was introduced to Ireland from England and subsequently adapted to use of S. trutta and G. duebeni there.

The ability of Hymenolepis nana oncosphere extract to induce eosinophil chemotactic response was examined in vitro and in vivo. The extract showed a chemotactic activity specific for eosinophils but not for neutrophils. Partially purified eosinophil chemotactic factors (ECFs) from the oncosphere extract showed apparent molecular mass from 5.5 to 9.6kDa and 30 to 40kDa. These were resistant to heating and proteinase K digestion but sensitive to periodate oxidation. Peritoneal injection of the crude extract or partially purified ECFs to mice resulted in a preferential eosinophil infiltration. The chemotactic activity for eosinophils was not separable from the adhesion molecule expression or oxygen radical-inducing activity by means of chromatography or chemical treatments. Furthermore, histological examination demonstrated a marked tissue eosinophilia around H. nana larvae in the intestinal lamina propria of both humoral and cell-mediated immunodeficiency mice. The present findings suggest that H. nana oncosphere-derived molecules facilitate in vivo the intestinal eosinophilia during the infection.

For the sensitive detection of eggs of Echinococcus multilocularis in fox faeces by PCR we have evaluated a method based on the previous concentration of helminth eggs by a combination of sequential sieving of faecal samples and flotation of the eggs in zinc chloride solution. The eggs were microscopically detected in the fractions retained in 40 and 20µm mesh sieves. DNA of the taeniid eggs retained in the 20 µm sieve was obtained after alkaline lysis and PCR was performed using E. multilocularis species-specific primers. Compared to the parasitological findings after examination of the small intestines of the foxes, the specificity of the PCR was 100% (no false-positive result with 20 foxes free of E. multilocularis) and the sensitivity was 94% (33 positive results from total 35 foxes proven to be infected with E. multilocularis). Both false-negative results were obtained with faeces from foxes harbouring immature worms. Using faecal volumes between 2 and 20 ml, no inhibition of PCR was observed as was demonstrated by the amplification of size-modified target in parallel reactions. The tests were undertaken with fresh faeces stored in 70% ethanol, but egg detection by PCR was also possible after inactivation of eggs by freezing the faeces at −80°C for one week or by incubation at +70°C for 2 h.

The presence of naturally occurring entomopathogenic nematodes (epns) was surveyed in the West-Flanders province in the north-west of Belgium. In 21 sites of different agronomical situations, 130 soil samples were taken. Using the Galleria larva bait technique, 16 soil samples were found positive for epns. Fifteen samples were found to contain Steinernema spp. (nine S.feltiae, five S. affinis, one Steinemema species B3). The remaining positive sample contained Heterorhabditis sp. (North West European strain). The morphometric characters of the isolates were highly variable and did not correspond precisely to the original descriptions; however, biochemical characterization confirmed their identity. The epns were isolated from 50%, 18.8% or 12.3% of the samples taken in sand dunes, grassland or woodlands, respectively. S. feltiae and S. affinis were isolated in these three habitats; Heterorhabditis sp. was found in a grassland habitat. Steinernema feltiae was prevalent in loamy sand soils with a wide range of organic matter content; S. affinis, Heterorhabditis sp. and Steinernema species B3 were isolated in sandy loam soils. All the positive sample sites were in the pH range of 4.0–8.1. This is the first report of naturally occurring entomopathogenic nematodes in Belgium.

Ten-day-old adults of Echinostoma caproni and E. paraensei, some of which had been exposed to 3-H-tyrosine to label sperm, were transplanted to mice in various combinations. Mating behaviour was followed on autoradiograms of worms recovered after 5 days by detecting the transfer of labelled sperm from labelled worms to themselves or to unlabelled worms. When single, labelled E. caproni adults were transplanted with unlabelled E. paraensei, they self-inseminated but did not show evidence of interspecies mating. No interspecies mating but self-insemination was observed when single, labelled E. paraensei and unlabelled E. caproni were transplanted together. When the labelled species had a choice of unlabelled adults of its own species or the opposite species, it mated in similar fashion to that seen in single species infections. The labelled species acted as if the opposite species was not present and showed an unrestricted mating pattern where it would both self- and cross-inseminate. Even though after transplant the E. paraensei adults were found in the duodenum and the E. caproni adults were found in the ileum, approximately 25% of the transplanted worms of both species were found within 1 cm of each other. Thus habitat separation was not a major cause of the lack of interspecies mating. Lack of reproductive recognition is another reason to separate E. caproni and E. paraensei into distinct species.

A total of 120 gastro-intestinal tracts and 960 faecal samples were examined to assess the prevalence and seasonal changes in the gastro-intestinal helminth parasites of Red Sokoto (maradi) goats slaughtered at Ibadan between May 1991 and April 1992. Egg types of strongyles, Strongyloides, Trichuris, Skrjabinema, Dicrocoelium and Moniezia were encountered in 93%, 83%, 44%, 0.9%, 2.3% and 31% of the faecal samples respectively. However, only strongyle, Strongyloides and Trichuris eggs occurred in large numbers and were more common during the rainy season than in the dry season. The parasites recorded and their prevalences were Haemonchus contortus (90.0%), H. ovis (5.0%), Strongyloides papillosus (80.8%), Trichostrongylus colubriformis (78.3%), T. axei (69.2%), Trichuris ovis (72.5%), T. globulosa (38.3%), Oesophagostomum columbianum (67.5%), Cooperia curticei (58.3%) Gaigeria pachyscelis (40.8%), Skrjabinema ovis (5.0%), Nematodirus battus (5.8%), Moniezia expansa (29.2%), M. benedeni (10.0%), Paramphistomum spp. (5.0%) and Cysticercus tenuicollis (33.3%). Haemonchus ovis is reported for the first time in Nigeria. Mixed infections were most prevalent. Young goats were more commonly infected and had higher worm counts than adult goats. Only Haemonchus, Trichostrongylus, Strongyloides and Cooperia spp. occurred in large numbers. Irrespective of the age of the goats, higher worm counts were generally encountered during the rainy season than in the dry season. The results are discussed in relation to the control of helminthiasis in grazing animals in Nigeria.

Adults of Echinostoma paraensei grown in mice were exposed to 3-H-tyrosine and transplanted to uninfected mice singly or with unexposed worms. The presence of sperm labelled with the radioactive tyrosine in the seminal receptacles of labelled and unlabelled worms recovered after 5 days indicated the mating pattern of this species. Of nine isolated, labelled worms recovered, four had self-inseminated (44%). Of the 39 unlabelled found with eight labelled worms, only eight exhibited labelled sperm for a cross-insemination rate of 21%. Six of these eight labelled worms had self-inseminated (75%) in addition to the cross-insemination. This pattern of mating is termed unrestricted, unlike the restricted mating seen in three species of eyeflukes, which will not self-inseminate in groups. In contrast to the other species of echinostomes investigated, very little sperm was found in the seminal receptacles of the serially sectioned E. paraensei. Adults were always found in the duodenum, even when transplanted in the ileum.

The surface tegument of G. tigrinum generally resembles that described for other digeneans. It contains surface tubercles and is covered with a glycocalyx. In the anterior region the tegument bears spines while tubular-like canals and occasional deep invaginations are present on the anteroventral surface. Two types of secretory body are present in the tegumental syncytium and produced in separate tegumental cells. Mitochondria are present in the surface syncytium, suggesting active transport occurs across the tegument in non-hibernating frogs.

The ultrastructural morphology of the alimentary tract of the third-stage larvae of Litomosoides carinii (Nematoda: Filarioidea) is described. The oesophagus possesses the characteristic tri-radiate lumen and is lined by cuticle. The lumen of the oesophagus is tightly occluded anteriorly by particulate material and posteriorly at the oesophagointestinal junction. Three glands are present in the glandular region, one dorsal and two subventral. These glands contain secretory granules with similar morphological and staining properties. The lumen of the intestine is ill-defined without microvilli and apparently non-functional. It is likely that the larvae feed via the transcuticular route.

The activities of selected enzymes of the respiratory chain system in Onchocerca fasciata (Filarioidea: Onchocercidae) have been investigated histochemically. Thus, the localization and distributions of NADH dehydrogenase (EC 1.6.99.3), succinate dehydrogenase (SDH) (EC 1.3.99.1) and cytochrome oxidase (EC 1.9.3.1) were investigated in various tissues of the adult female worm by employing MTT, Nitro BT (dehydrogenases) and DAB (cytochrome oxidase). Different tissues varied considerably in their enzymatic activities. The hypodermis and reproductive tissues showed strong and identical localization of NADH and SDH dehydrogenase activities reflecting high metabolic rates. Little or no dehydrogenase activities were observed in the intestine and cuticle. In contrast to the two dehyrogenases, no activity was observed for cytochrome oxidase in any of the tissues in adult or embryonic stages of the worm. The significance of these results with respect to the energy metabolism of the worm is discussed. It is suggested that O. fasciata lacks a classical, mammalian-type respiratory pathway and that oxidative phosphorylation is of no importance as an energy generating pathway in this essentially anaerobic parasite.

The random amplified polymorphic DNA (RAPD) assay was used to find molecular markers able to distinguish Trichinella spiralis from T. britovi, the two recognized Spanish Trichinella species. Fourteen Spanish Trichinella isolates, as well as reference Trichinella isolates representing the five species T. spiralis (T1), T. nativa (T2), T. britovi (T3), T. pseudospiralis (T4) and T. nelsoni (T7) and the three other taxa Trichinella T5, Trichinella T6 and Trichinella T8 of the genus, were characterized by RAPD using both purified and crude DNAs from infective muscle larvae (ML) and seven arbitrary primers. Three primers yielded diagnostic RAPD markers for the Spanish T. spiralis and T. britovi isolates as well as for the Trichinella reference isolates analysed, and in the case of crude DNAs the results were obtained in few hours. In addition, the species-specificity of the diagnostic RAPD markers from Spanish Trichinella isolates was studied by cross-hybridization assays. These assays confirmed that the selected diagnostic DNA fragments were not species-specific, but showed potential differences in the copy number among the examined Trichinella genetic clusters.

A serum IgG response to Strongyloides papillosus infection was analysed in rabbits. IgG production was induced against 30–200 kDa larval somatic antigens and 21–160 kDa adult excretory/secretory (ES) antigens at 4 weeks postinfection. No immunoreactivity was detected between sera of infected animals and any proteins of larval ES and adult somatic antigens. Protection against larval challenge infection was investigated in rabbits immunized by primary infection and by intradermal inoculations of larval somatic and adult ES antigens. Primarily infected animals had 90% fewer or less worms from day 2 onwards after challenge, and lower faecal egg outputs after challenge, when compared with those in susceptible animals. Immunization with larval somatic antigens induced effective protection, showing 52.6–81.5% reductions in worm recovery from day 5 onwards after challenge, and lower faecal egg outputs after challenge. Systemic immunization with adult ES antigens failed to protect the animals against challenge. The possibility that resistance to S. papillosus reinfection is performed on the killing of migrating larvae in the early stages of infection is discussed. Systemic immune responses to larval somatic antigens might play an important role in the resistance.

Computer-assisted microscopy and multivariate statistics were used to establish and evaluate a procedure for identification of bovine strongylid eggs. Ostertagia ostertagi, Cooperia oncophora, Haemonchus placei, Trichostrongylus axei, and Oesophagostomum radiatum eggs were obtained from faeces voided by monospecifically infected calves. Images of single eggs (400× magnification) were recorded by a CCD camera fitted onto a microscope and digitizied on a PC. After separation of eggs from the image background, the pixel (picture element) positions of the egg outline were analysed by algorithms to describe size and shape. A stepwise discriminant analysis was subsequently used to select and rank descriptive features of 4207 eggs according to discriminatory power. Classification criteria were developed by linear discrimination analysis on the basis of selected features, and the criteria evaluated by cross-validation. A maximum average percentage of correct classification of 85.8% resulted when nineteen features were employed in a linear classification criterion. The percentages correct classification for each species were: O. ostertagi 76.3%, C. oncophora 90.8%, O. radiatum 87.8%, H. placei 90.1%, and T. axei 83.8%. Classification based on the five most important features gave an overall correct classification of 81.5%. Images of ‘unknown’ eggs could be identified automatically by the classification criteria after procedural steps performed by PC were linked in a batch program.

Eosinophil counts, interleukin 5 (IL-5) concentrations in peripheral blood and larval recovery after infection with Toxocara canis were compared between IL-5 transgenic (Tg) and nontransgenic counterparts, C3H/HeN mice. In Tg mice characterized by a constant high level of peripheral eosinophils, eosinophils in peripheral blood fell to the lowest level on day 4 of a T. canis infection and then returned to the preinfection level on day 14. Serum IL-5 level fell to the lowest level on day 4 of infection, then recovered rapidly and peaked on day 14 postinfection. In contrast, the eosinophil and IL-5 levels in the peripheral blood peaked on days 11 and 7 of infection, respectively in C3H/HeN mice. The degree of eosinophil infiltration into the lung 4 days after infection was far more pronounced in Tg than C3H/HeN mice. The highest number of larvae recovered from the lungs of infected Tg and C3H/HeN mice occurred on day 4 postinfection. Strains of Tg and C3H/HeN mice vaccinated with excretory and secretory (ES) antigens of T. canis larvae were infected with T. canis and the recovery of larvae on day 21 analysed. There were no significant differences in the mean number of larvae recovered from nonvaccinated Tg and C3H/HeN mice or vaccinated Tg and C3H/HeN mice. However, significant differences were demonstrated in the mean total number of larvae recovered from vaccinated and nonvaccinated Tg or C3H/HeN mice. These results suggest that immunocompetent cells other than eosinophils may play a significant role in the expulsion and killing of T. canis larvae in infected mice.

Natural infection of the plankton Epischura baicalensis Sars (Calanoida: Temoridae) by procercoids of the genus Proteocephalus in Lake Baikal is analysed in detail. Over one year Epischura was infected by procercoids of proteocephalids at a prevalence of 0.13% (in Listvenichnyi Bay, South Baikal). Male E. baicalensis were infected more (0.39%). The number of procercoids of genus Proteocephalus was 2.7 specimens/m3. The largest number of procercoids was found in females (1.26), the least in the third copepodid stage (0.03 specimens/m3). Because of the close ecological contacts between E. baicalensis and Coregonus autumnalis migratorius it is probable that E. baicalensis is infected by P. exiguus, the species usually found in C. a. migratorius (Zaika, 1965). The parasite lives for about 8 months in the winter-spring Epischura generation and about 10 months in the summer generation.

The behaviour and infectivity of the infective juveniles (IJs) of two species of entomopathogenic nematodes, Steinernema carpocapsae Weiser and Steinernema feltiae Filipjev (Rhabditida: Steinernematidae), were determined after exposure to two neuroactive pesticides, oxamyl and fenamiphos, widely used against plant parasitic nematodes. Oxamyl (50 μg ml−1) initially stimulated non-sinusoidal movement in S. feltiae but had no appreciable effect on IJs of S. carpocapsae, while fenamiphos (10 and 50 μg ml−1) increased and sustained non-sinusoidal movement in both species over an observation period of 48 h. The two pesticides reduced the infectivity of both nematode species against late instar larvae of the wax moth, Galleria mellonella L. in a sand-tube bioassay when compared with controls. Infective juveniles of S. feltiae showed limited body waving behaviour on the sand surface whereas S. carpocapsae showed the full range of nictation movements, including body waving, stationary straight postures and leaping. Oxamyl had no significant (P>0.05) effect on the number of IJs of S. carpocapsae which nictated except when the IJs were pre-incubated with this compound for 24 h (P<0.05). In contrast, fenamiphos completely suppressed all nictation and body waving behaviour in both species. Nictating IJs of S. carpocapsae exposed to late instar G. mellonella for 12 h in a filter-paper bioassay were found to be more infective compared with non-nictating IJs. Treating IJs of S. carpocapsae with oxamyl (50 μg ml−1) increased the infectivity of non-nictating IJs compared with controls but reduced infectivity of nictating IJs when exposed to G. mellonella for 12 h. The work is discussed in relation to proposals to use these or other pesticides to enhance the activity and infectivity of IJs of entomopathogenic nematodes.

Enzyme electrophoresis has been used to examine genetic diversity in a population of Proteocephalus exiguus La Rue, 1911 (Cestoda: Proteocephalidae), parasitizing salmonid and coregonid fishes. Among 16 loci tested, three polymorphic loci (Ada, Got, Pgm-2) were found. Six different genotypes at the Got locus distributed in Hardy-Weinberg equilibrium suggest remarkable genetic flexibility of P. exiguus. Balancing selection is proposed as the mechanism maintaining genetic variation within the species. Data of genetic variability parameters (Ho = 0.064; He = 0.07; P = 0.19) and outcrossing rate (t = 0.842) of P. exiguus population have been provided. Possible factors affecting these data are discussed.