A preliminary purification has been carried out by continuous elution electrophoresis of a 49.5 kDa protease of crude extracts from Dicrocoelium dendriticum eggs. The enzyme showed a high capacity to degrade the collagen derivative azocoll at acidic pH. Although it is necessary to carry out further experiments to confirm any physiological role, this protease could be implicated in penetration mechanisms.

Effects of the anthelmintics, pyrantel and levamisole, on egg development of Angiostrongylus costaricensis were studied in vitro. After 7 days, about 80% of eggs developed to first-stage larvae in Ham's F-12 medium with 10% foetal calf serum under 5% CO2. Significant inhibition of development was caused by pyrantel (10-9–10-8 g ml-1) and levamisole (10-9–10-8 g ml-1) (Mann-Whitney U-test; P<0.05), and none of the eggs developed to first-stage larvae in higher concentrations of these anthelmintics (10-7 g ml-1). Furthermore, incubation with these drugs at 10-8 g ml-1 for at least 3 h or at 10-4 g ml-1 for 1 h caused irreversible effects on egg development.

The host-protective immune response to infection with gastrointestinal (GI) nematodes involves a range of interacting processes that begin with recognition of the parasite's antigens and culminate in an inflammatory reaction in the intestinal mucosa. Precisely which immune effectors are responsible for the loss of specific worms is still not known although many candidate effectors have been proposed. However, it is now clear that many different genes regulate the response and that differences between hosts (fast or strong versus slow or weak responses) can be explained by allelic variation in crucial genes associated with the gene cascade that accompanies the immune response and/or genes encoding constitutively expressed receptor/signalling molecules. Major histocompatibility complex (MHC) genes have been recognized for some time as decisive in controlling immunity, and evidence that non-MHC genes are equally, if not more important in this respect has also been available for two decades. Nevertheless, whilst the former have been mapped in mice, only two candidate loci have been proposed for non-MHC genes and relatively little is known about their roles. Now, with the availability of microsatellite markers, it is possible to exploit linkage mapping techniques to identify quantitative trait loci (QTL) responsible for resistance to GI nematodes. Four QTL for resistance to Heligmosomoides polygyrus, and additional QTL affecting faecal egg production by the worms and the accompanying immune responses, have been identified. Fine mapping and eventually the identification of the genes (and their alleles) underlying QTL for resistance/susceptibility will permit informed searches for homologues in domestic animals, and human beings, through comparative genomic maps. This information in turn will facilitate targeted breeding to improve resistance in domestic animals and, in human beings, focused application of treatment and control strategies for GI nematodes.

Mature specimens of Cucullanus heterochrous Rudolphi, 1802 (Nematoda: Cucullanidae) were obtained from the intestine of the flounder, Platichthys flesus, from Danish waters. Eggs embryonate in seawater but do not hatch. Fully developed larvae pressed out of eggs are 430 μm long with amphids and dereids and enclosed within the cuticle of a previous larval stage. Infective larvae are believed to be in their third stage. Experimental studies showed that the polychaetes, Nereis spp., Scoloplos armiger, Brada villosa and Capitella sp., may act as intermediate hosts. In N. diversicolor the larvae increase their length to 1 mm within four weeks (15°C) without moulting. Experimental infections showed that larvated eggs are not infective to fish, whereas >550 μm long larvae from polychaetes survived in 4–24 cm long flounders and plaice, Pleuronectes platessa. Third-stage larvae 550 μm to 1.1 mm long were found in the submucosa of the intestine one week post infection. At a length of about 800 μm to 1.4 mm they moult to fourth-stage larvae. Fourth-stage larvae, immature and mature worms occur in the intestine and rectum. Fourth-stage larvae and adults survived experimental transfer from one flounder to another. Similar developmental stages survived for two weeks in the intestine of experimentally infected cod, Gadus morhua.

Two groups of Galba truncatula and two groups of Lymnaea natalensis were experimentally infected with Fasciola gigantica to determine if snail species had an influence on the redial burden and cercarial shedding of this trematode when snails of both species were infected with the same isolate of miracidia. In the two groups used for the study of redial burden, the total number of free rediae was significantly higher at day 49 post-exposure in L. natalensis than in G. truncatula. In the groups used for cercarial shedding, the life-span of cercaria-shedding snails and those of infected snails which died without cercarial emission, and the duration of the prepatent period were significantly longer in L. natalensis than those noted in G. truncatula. However, the mean numbers of shed cercariae did not significantly differ and showed no differences in their daily distribution throughout the shedding period. These results demonstrate that G. truncatula might be the principal intermediate host of F. gigantica in Egypt, at least in the areas where this lymnaeid species lives.

Tetrathyridia of Mesocestoides corti were cultured in vitro in a diphasic medium consisting of a liquid medium (CMRL Sigma) and a thixotropic nutrient gel (Oxoid). Tests demonstrated that a 50% medium/gel mixture produced optimum conditions for the survival and development of tetrathyridia. Established anthelminthic drugs were inoculated into the gel which demonstrated that this system can be used for preliminary anthelminthic drug screening. The development and survival of the tetrathyridia were influenced by the addition of pepsin, trypsin and liver peptone to the culture media. The development and maturation of proglottids were observed in addition to asexual reproduction by the process of budding. Tetrathyridia maintained in vitro and reinfected into both mouse and rat hosts retained their viability.

Aminopeptidase-like activities in crude whole body extracts of the free-living nematode Caenorhabditis elegans and the plant parasitic soybean cyst nematode Heterodera glycines were examined. General characteristics including pH optima, heat lability, and inactivation of enzyme by organic solvent were the same for the two species. All developmental stages of H. glycines exhibited activity. In older females, activity was present primarily in the eggs. Affinity for the substrate L-alanine-4-nitroanilide was the same regardless of the stage examined, and was similar for the two species (Km=2.3±0.3 mM FOR C. ELEGANS AND 2.9±0.2 Mm for H. glycines). Nearly all (>95%) of C. elegans aminopeptidase-like activity was present in the soluble fraction of the extract, while H. glycines activity was distributed between the soluble and membrane fractions. Specific activities of the soluble enzymes were highest in C. elegans and H. glycines juveniles. The C. elegans enzyme was susceptible to a number of aminopeptidase inhibitors, particularly to amastatin and leuhistin, each of which inhibited aminopeptidase-like activity more than 90% at 90 μm. In H. glycines, aminopeptidase-like activity was inhibited 39% by amastatin at 900 μm. The apparent molecular weight of the soluble C. elegans enzyme is 70–80 kDa. Some activity in H. glycines is present in the 70–80 kDa range, but most activity (80–90%) is associated with a very high molecular weight (>240 kDa) component.

The ultrastructure of the spinous body tegument of the metacercaria of Timoniella imbutiforme (Molin, 1859) has recently been described. Other regions of the metacercarial tegument, including those of the oral sucker, pharynx, and nephridiopore, demonstrate considerable specializations. The oral sucker tegument had an aspinous outer syncytial layer that possessed a pimpled apical surface as well as enclosing two types of secretory bodies. The pharyngeal tegument likewise lacked spines, but possessed only one type of secretory body, and a smooth but folded outer surface. The nephridiopore tegument, however, showed the greatest degree of specialization possessing a single type of secretory body specific only to this region of the tegument. Also associated with the syncytium here was a prominent long filamentous glycocalyx, and microtubules which were observed for the first time in this region of the tegument.

Seasonal fluctuations in the prevalence and abundance of infection with intestinal helminths were studied in Apodemus sylvaticus (wood mouse, n = 399), from three contrasting habitats in southern England, to test the hypothesis that both intrinsic (host sex, age) and extrinsic (season, site) factors influence parasite species richness and abundance. Five species of helminths were recovered but only one of these (Capillaria murissylvatici) was site-specific (Dungeness). Total species richness was therefore 5 at Dungeness and 4 at the other two sites. Mean species richness was 1.4, but in adult mice there was a pronounced difference between the sites, and an independent highly significant effect of season. Syphacia stroma and Corrigia vitta both showed marked differences between sites in respect of prevalence and abundance of infection. Capillaria murissylvatici was encountered at Dungeness mostly in the spring whereas seasonal changes in abundance ofS. stroma were consistent across all three sites. Seasonal fluctuations in the abundance of Catenotaenia pusilla were compounded by differences between sites. Host sex was not a significant factor in any species, although a posteriorianalysis of S. stromaworm burdens for the Isle of Wight site revealed a moderate local sex effect. Overall the principal determinants of variation in helminth burdens were the extrinsic factors, site and season.

The effects of glucose in artificial spring water (ASW) on the survival, infectivity, and linear movement of Echinostoma caproni cercariae were studied. Cercariae maintained at 23°C in 1% glucose in ASW (ASWG) or ASW alone, reached 50% survival at 26 and 23 h, respectively. All cercariae in ASWG and ASW were dead by 50 and 32 h, respectively. Infectivity to juvenile Helisoma trivolvis (Colorado strain) snails was significantly less for cercariae aged 16 h in ASWG compared to cercariae aged 16 h in ASW. Linear movement, i.e. the ability of cercariae to traverse a 1-cm radius, ceased at 16 and 20 h for cercariae maintained in ASWG and ASW, respectively. Glucose added to ASW extended the survival time of E. caproni cercariae but decreased their ability to infect snails or move in a linear direction.

Of all the laboratory models of intestinal nematode infection, Trichuris muris in the mouse is arguably the most powerful. This is largely due to the fact that the ability to expel this parasite is strain dependent. Thus, most mouse strains readily expel T. muris. However certain mouse strains, and indeed some individuals within particular mouse strains, are unable to mount a protective immune response and harbour long term chronic infections. This unique model thus presents an opportunity to examine the immune events underlying both resistance to infection and persistent infection within the same host species, and in some cases, the same host strain.

Peroxynitrite (ONOO-) is a cytotoxic anion, produced by interaction between nitric oxide and superoxide in vivo in some inflammatory cells. This study investigated its effects on Fasciola hepatica and Dicrocoelium dendriticum isolated from ovine livers and kept in bile at room temperature. Peroxynitrite was synthesized using a quenched flow reactor and assayed spectrophotometrically. It was applied at different concentrations (10-3.5 to 10-2.3 M) to the flukes kept in bile. The viability of the peroxynitrite-treated flukes was compared with a control group (n=5–7 per group). Control F. hepatica and D. dendriticum lived for 226±11 and 208±14 min, respectively. Life times were decreased by peroxynitrite at all concentrations used (P<0.001). At the highest concentration of peroxynitrite, F. hepatica and D. dendriticum lived only for 6.1±0.4 and 4.1±4.1±0.2 min, respectively. Correlation between peroxynitrite concentration and parasite viability was significant in the case of F. hepatica (r=-0.842; P=0.0035). A single application of peroxynitrite can decrease the life span of ovine liver flukes. A failure in the activation of hepatic macrophages in infected animals may lead to a decreased production of free radicals and, thus, peroxynitrite. Such a failure is likely to deprive the body of a defence tool against multicellular parasites.

Anisakis third stage larvae utilize a variety of fish as intermediate hosts. Uncooked fish are rendered safe for human consumption by freezing. Larvae freeze by inoculative freezing from the surrounding medium but can survive freezing at temperatures down to -10°C. This ability may be aided by the production of trehalose, which can act as a cryoprotectant, but does not involve recrystallization inhibition. Monitoring of fish freezing in commercial blast freezers and under conditions which simulate those of a domestic freezer, indicate that it can take a long time for all parts of the fish to reach a temperature that will kill the larvae. This, and the moderate freezing tolerance of larvae, emphasizes the need for fish to be frozen at a low enough temperature and for a sufficient time to ensure that fish are safe for consumption.

A literature survey was undertaken in order to draw up a definitive list of helminth parasites of the wolf, Canis lupus. From 27 papers a total of 72 helminth species from 40 genera were recorded that infect wolves, of which 93% were identified from the gastrointestinal tract at necropsy. They comprised 28 species of nematode, 27 species of cestode, 16 species of trematode and one acanthocephalan. Of these, 46 species were able to be included in further meta-analysis of prevalence data derived from 25 publications for which the total number of wolves examined was 1282 (1066 from Nearctic populations, and 216 from the Palaearctic region). These two populations were further subdivided into three relevent ecosystems or biomes, i.e. temperate/montane (n=216), boreal (n=805) or tundra (n=261). The meta-analysis of relative prevalence indicated the most common helminth species to be the tapeworm Taenia hydatigena, which occurred at relative rates of >30% for either zoogeographic region as well as in each of the three biomes. The related tapeworm, Echinococcus granulosus also exhibited high meta-prevalence (>19%) in all host biomes. The hookworm Uncinaria stenocephala was the most prevalent nematode species by meta-analysis (meta-prevalence 44.9%) in the temperate/montane biome, while the ascarid Toxascaris leonina was the dominant helminth species (meta-prevalence 73.9%) in the tundra wolf populations. Trematodes in the genus Alaria were the dominant fluke (meta-prevalence 3–5%) in all biomes. Analysis of published studies for helminth biodiversity using the Shannon-Wiener index based on species number and meta-prevalence by region or biome, indicated that highest helminth diversity occurred in wolf populations of the temperate/ montane biome (Palaearctic), and was lowest in tundra wolf populations of the Nearctic (P<0.05). Helminth species assemblage in European wolf populations was therefore at least as great or more varied than was recorded for the larger less disturbed wolf populations of North America.

The karyotype of glirid tapeworm Rodentolepis myoxi (Rudolphi, 1819) (Cestoda: Hymenolepididae) comprises six pairs of small bi-armed chromosomes (2n = 12). All pairs of chromosomes possess uniform morphology, i.e. metacentric, submetacentric or meta-submetacentric types of structures. The formula of the karyotype structure is n = 2m + 1m-sm + 3sm. The absolute chromosome length ranges from 3.78 to 2.00 μm. The mean total length of the haploid complement is 15.98 μm. The first pair (group A) is the largest, pairs 2 and 3 can be grouped into group B while pairs 4–6 are smaller and can be classified as group C. The number of chromosomes of R. myoxi is the same for the congeneric species, however, karyological characteristics differ from all recently known karyotypes of rodent hymenolepidids.

The nurse cell in the cyst of Trichinella spiralis comprises at least two kinds of cytoplasm, derived from muscle or satellite cells, as indicated by the pattern of staining using regular dye (haematoxylin and eosin, or toluidine blue), alkaline phosphatase (ALP) expression, acid phosphatase (ACP) expression and immunostaining with an anti-intermediate filament protein (desmin or keratin). Muscle cells undergo basophilic changes following a T. spiralis infection and transform to the nurse cells, accompanied by an increase in ACP activity and the disappearance of desmin. Satellite cells are activated, transformed and joined to the nurse cells but remain eosinophilic. The eosinophilic cytoplasm is accompanied by an increase in desmin and ALP expression but not an increase in ACP activity. Differences in the staining results for ALP or ACP suggest that the two kinds of cytoplasm have different functions. Trichinella pseudospiralis infection results in an increase of ACP activity at a later stage than T. spiralis. There is also a difference in the location pattern of ACP in the cyst of T. spiralis compared with T. pseudospiralis. In T. spiralis, ACP is diffused within the cell, but in T. pseudospiralis, ACP distribution is spotty corresponding to the location of the nucleus. Trichinella pseudospiralis infection is accompanied by a slight increase in ALP activity. Activated satellite cells following a T. pseudospiralis infection exhibit an increase in desmin expression. The present study therefore reveals that nurse cell cytoplasm differs between the two Trichinella species and between the two origins of cytoplasm in the cyst of T. spiralis.

Four parameters of the intestinal inflammatory response (numbers of mucosal mast cells (MMC) and Paneth cells, villus:crypt ratios and mitotic figures) were measured in mice exposed to varying doses of infective larvae of Trichinella spiralis.The aim of the experiments was to determine whether generation of these components of inflammation required a threshold level of infection and whether, once triggered, inflammation became pan-mucosal. Near maximal MMC and Paneth cell responses were elicited even with infections as low as 35 larvae; changes in villus:crypt ratios and in mitotic indices also occurred at this level of infection, but were progressively greater with increasing levels of infection. In all infected mice, including those infected with 35 larvae, MMC and Paneth cell responses extended over most of the small intestine. These data are interpreted as showing: (i) that the intestinal mucosa is highly responsive to T. spiralis infection; (ii) that once triggered, components of the inflammatory response are amplified by T cell-dependent mechanisms, becoming pan-mucosal; and (iii) that MMC and Paneth cell responses, which require cell division and differentiation, become maximal at a lower infection threshold than changes in the villus:crypt ratio or in mitotic indices, which directly reflect increased rates of division in crypt cells.

Inequality in body sizes is a common feature in populations of helminth parasites, with potential consequences for egg production and population genetics. Inequalities in body lengths and the effects of intraspecific competition on worm length were studied in a species of mermithid nematode parasitic in the crustacean Talorchestia quoyana (Amphipoda: Talitridae). The majority of the 753 worms recovered were relatively small, and an analysis using a Lorenz curve and Gini coefficient suggested that there were no marked inequalities in body lengths among the worms. Total worm length in the 356 infected amphipods (i.e. the sum of the lengths of all the worms in a host) increased steadily as a function of the number of worms per amphipod, whereas the length of the longest worm per amphipod peaked in amphipods harbouring intermediate numbers of worms. This last result was not significantly accounted for by the observed increase in host size with increasing intensity of infection, but resulted from a correlation between worm length and host size. As the number of worms per amphipod increased, the relative sizes of the second-, third-, and fourth-longest worms per host increased markedly. This means that relative inequalities in sizes become less pronounced, i.e. subordinate worms get closer in size to the longest worm, as the number of worms per host increases. The main consequence of this phenomenon is that worm sizes in the mermithid population are more homogeneous than they would be if intraspecific competition had stronger effects on worm growth.

The biogenic elements zinc, manganese and cobalt are essential for metabolic processes in animals. Compounds of nGly.Me2+A. mH2O (Me2+=Zn2+, Mn2+, Co2+; A=Cl−, SO42−, n=1, 2; m=2, 5), as supplements in the diet, were used separately on different experimental groups of male Hisex chickens to correct the mineral deficiency caused by Ascaridia galli infections. An amelioration of body weight gain, reduction of mortality and restoration of trace element levels were estimated in infected chickens. A mathematical model has been proposed for A. galli population kinetics in chickens, taking into account the stimulating effect of these elements on the nematodes. The model parameters are considered as phenomenological constants of the host–parasite system. An agreement with experimental data is observed using, for the parameters ψ, α, μ and μs, values equal to those calculated in previously investigated A. galli–chicken systems. For parameter ν (immunological constant) the same value was obtained as in a previous experiment with high infection. This model is likely to be suitable for a range of host–nematode systems, including varying degrees of infection and treatment with different trace elements.

Anisakid nematodes belonging to the Anisakis simplex complex are highly prevalent in several fish species off the coast of Portugal and are an important zoonotic problem in the Iberian Peninsula. Two reproductively isolated sibling species of the Anisakis simplex complex were identified from Pleuronectiformes inhabiting the Portuguese coast using restriction fragment length polymorphism (RFLP). Recombinant genotypes corresponding to presumptive Anisakis simplex sensu stricto and Anisakis pegreffii hybrids were also detected by this technique, as well as the species Anisakis typica. Although 25 species of Pleuronectiformes were investigated, Anisakis spp. larvae were only found in seven: Arnoglossus imperialis, Arnoglossus laterna, Lepidorhombus boscii, Citharus linguatula, Platichthys flesus, Dicologlossa cuneata and Solea senegalensis. The occurrence of hybrids in relatively sedentary fishes such as the Pleuronectiformes suggests that the Portuguese coast may constitute an area of hybridization and, therefore, is of particular interest for the study of the process of hybridization and speciation for these anisakids.