Two domestic guinea-pigs (Cavia porcellus), bought in Pinheiros, São Paulo State, Brazil, were taken by their owners a farm in the rural district of Capão Bonito, close to the Atlantic Forest, São Paulo, where they both developed tumour-like and ulcerating lesions on the ears. The causative agent was identified as Leishmania (L.) enriettii, based on biological characters and isoenzyme profiles. Sources of the parasite in wild mammals, and the possible sandfly vector species discussed.

Receptors interacting with terminal sugars as ligands are involved in the binding of Leishmania donovani promastigotes to the macrophage surface and their subsequent internalization. Mannose and glucose are specifically involved in the binding process. Decreased binding occurs to macrophages already infected with L. donovani either in vivo or in vitro. When mannose- or glucose-bearing liposomes are used as ligands the binding shows similar trends and the percentage inhibition of binding with mannose-bearing liposomes increases when compared to that for the glucose-bearing ones. The decreased binding of the ligand seems to be due to a decrease in the number of receptors after infection. The affinity of the ligands for the binding sites either on the normal macrophages or on the infected macrophages apparently remains the same. The results based on the incorporation of [3H]phenyl alanine and supported by the binding of glycosylated liposomes to both infected and non-infected macrophages suggest that protein synthesis, in general, is suppressed in L. donovani infected macrophages thus affecting also mannose/glucose receptor protein synthesis, resulting in fewer receptors on the macrophage surface.

The severity of the trypanosomiasis problem in a particular location is traditionally assessed in terms of a challenge index – the product of some measure of tsetse abundance and infection rate – which is assumed to be proportional to the force of infection. However, this index masks variation in the force of infection between herds and among individuals within herds. It is also not comparable between sites since the relative abundance of tsetse to hosts may vary. We have studied spatial distribution of herds of cattle in relation to tsetse in The Gambia and calculated an index of challenge based on the ratio of vectors to hosts over the livestock ranging area. This index is strongly correlated with estimates of the force of infection calculated from the incidence of infection in susceptible zebu; and it provides information on heterogeneity in exposure of different herds to tsetse.

An immunodominant antigen in Trypanosoma congolense-infected cattle is a 69 kDa protein which is conserved among species and developmental stages of African trypanosomes. Immunoscreening of a cDNA expression library identified a 2·35 kbp clone which contains a complete open reading frame encoding a protein of 653 amino acids with a predicted molecular mass of 71 kDa. Protein sequence analyses revealed 45–65% identity with hsp70s from a broad range of organisms, the highest homology being with the mammalian BiP (immunoglobulin heavy chain binding protein). The 69 kDa trypanosome protein shares with other BiP-related molecules two characteristics that are associated with their localization in the endoplasmic reticulum and their function as chaperonins, i.e. a hydrophobic N-terminal signal sequence and a conserved C-terminal tetrapeptide (X)DEL. Divergence between the 69 kDa antigen and other BiP-homologues occurs in the C-terminal region. This may be responsible for the high immunogenicity of the trypanosome protein. The gene for the 69 kDa antigen appears to be present as a cluster of several copies which are not organized in tandem repeats. It is expressed in all developmental stages of T. congolense, but the specific mRNA levels are higher in metacyclics than in other stages.

The relationships between regional species richness and local species richness were examined in respect of helminth parasite communities in 32 species of freshwater fish in the British Isles. Fish were divided into 5 categories, for each of which the goodness of fit of the relationship to linear, exponential and power function models was tested. For all categories of fish combined, there was a significant, positive curvilinear relationship. Nested within this were two other patterns. For introduced fish, a linear model provided the best fit; for euryhaline and relict species it was impossible to determine the best model, but for the other categories the relationship was curvilinear and was best fitted by a power function model. The linear relationship found for introduced fish was interpreted as a temporary situation, reflecting the shortage of time for the communities to become saturated. It corresponded to the linear part of the curvilinear relationship of the other categories, which is believed to represent the fundamental form of the relationship for parasite communities. The communities reached a saturation level of richness, corresponding to the asymptote of the curve, which fell well below regional species richness. Explanations for local saturation are discussed, but neither community structure nor supply-side ecology can yet be preferred. It is concluded that local patterns in helminth community richness, in contrast to those in fish assemblages, are not significantly influenced by patterns on a larger, regional spatial scale and so regional species richness is not a key determinant of local species richness, nor does a knowledge of regional patterns improve predictability of local patterns.

Problems of pattern and scale are considered in relation to helminth communities of freshwater fish by examining them at different hierarchical taxonomic and spatial scales, with a view to seeking generalizations of heuristic value, assessing the importance of phylogenetic and ecological determinants of community structure and improving understanding of unpredictable communities. Initially, communities were analysed at the level of salmonid genera, focusing on Oncorhynchus, in its heartland in Canada: then in O. mykiss throughout its global range and finally in individual localities to which it has been introduced in Britain. In the heartland, communities are dominated by salmonid specialist helminths, forming a phylogenetic element: the minority ecological element comprises broad generalists and non-salmonid specialists. Most species except generic specialists are shared between host genera. As the distance to which O. mykiss was translocated from its heartland increases, so generic specialists disappear first and then salmonid specialists decline. The community is thus increasingly composed of generalists and it also becomes increasingly poor. Helminths may be acquired from native salmonids and/or unrelated hosts, depending on availability. This same pattern is paralleled in individual localities in a restricted region: the phylogenetic element reflects the native salmonid species present and the ecological element the presence of other genera of fish, i.e. a supply-side situation. The change of scale in analysis has thus enabled the recognition of generalizations and patterns of heuristic value and improved the understanding of unpredictable communities by interpreting local variation as ecological ‘noise’ that often obscures fundamental patterns. In this and other taxa of fish, phylogenetic elements dominate helminth communities in the heartlands, but ecological elements dominate as the host increasingly becomes a stranger in a strange land.

The present study describes Trichuris trichiura infections in a cohort of children over a period of approximately 5 years, and examines the relationships between changes in infection intensity and changes in parasite-specific isotype responses (measured by ELISA). The decrease in mean infection intensity with time was mirrored by time-dependent changes in IgG subclass responses, with IgA and IgE levels remaining relatively constant in the cohort. At the individual level, changes in infection intensity between the initial and final time-points correlated positively and significantly with changes in all isotype levels with the exception of IgA.

After i.v. injection of 305 × 103 microfilariae (mf) per animal (50 g) into naive jirds, 50·8% of them could be recovered at autopsy 15 min later. Of these, 65·8% were calculated to be in the peripheral circulating blood (PCB) and were completely intact; 18·6% were recovered by perfusion of the lungs and 13·6% from the liver. In both organs about half the mf were associated with adherent lymphocytes and neutrophils but a few were partly disintegrated. Only 2·6% were recovered from the kidneys and the spleen. In long-term injection experiments using the same inoculum size the autopsy was done 15 min and 1, 3 and 6 weeks post-injection (p.i.) of mf into naive jirds. Throughout the experimental period the density of mf remained more or less constant in the PCB, but 3 weeks p.i. the density in the lungs increased up to 14 times to that in the PCB, whereas in the liver it decreased at the same time to a density similar to that in the PCB. In patent animals with adult worms delivering mf these were distributed as follows: 34·7% were calculated to be in the PCB; 24·4% were obtained by perfusion from the lungs and 22·0% from the liver; the rest were found in the kidneys (16·6%) and spleen (2·3%). In the lungs and the liver about 5/6 were associated with adherent cells, partly disintegrated or as fragments. In view of the fact that very few mf become disintegrated immediately after i.v. injection and also from their extremely long sojourn in the PCB, a low turnover rate of mf is presumed. The limited fecundity of the female adults combined with the 20 weeks median circulation time of injected mf in the PCB supports this view.

Host invasion and tissue migration of several helminths have been linked to expression and release of parasite-derived proteinases. The plerocercoid of the cestode Proteocephalus ambloplitis can migrate into the visceral organs or, in the case of bass, from them into the intestinal tract of the same individual fish. It does this within a few hours, aided by secretion of a substance from its apical gland. Proteinase activity in this plerocercoid, obtained from the host liver, was defined by pH optimum, by substrate and inhibitor specificity, and by electrophoretic and chromatographic techniques. Homogenates of plerocercoid contained a metalloproteinase exhibiting a molecular weight of 30000 determined by gelatin substrate gel electrophoresis. Peak activity of this proteolytic enzyme in gel filtration fractions when azocoll was used as substrate then corresponded to a molecular weight of 31500. The proteinase showed collagenolytic, haemoglobinolytic and slight elastinolytic activity, and it had a pH optimum at 9·0. Enzyme activity could be inhibited by various chelating agents. The metalloproteinase identified in this study constitutes the only enzyme class present in this larval stage of P. ambloplitis. We suggest that the plerocercoid's metalloproteinase is the substance secreted from the apical organ, necessary for the previously recognized tissue migration phase. This enzyme might also have a nutritional function.

We have used a number of molecular genetic approaches to characterize the cervid strain (‘northern form’) of Echinococcus granulosus. PCR–RFLP analysis of the nuclear ITS1 region of the rDNA repeat can readily distinguish the cervid form from other strains of E. granulosus. The complexity of the RFLP patterns obtained suggests that a number of distinct ITS1 types are present in this strain which may represent an inter-strain E. granulosus hybrid. Mitochondrial CO1 sequence of the cervid genotype was ambiguous at 18 positions and closely resembles a cluster of previously characterized E. granulosus genotypes, G1 (common, domestic sheep)/G2 (Tasmanian sheep)/G3 (buffalo). In contrast, mitochondrial ND1 sequence, although unique, suggests that the cervid form is most similar to strains represented by the G6 (camel)/G7 (pig) genotypes. We assume that the CO1 and ND1 sequences obtained for the cervid genotype are linked in a single mitochondrial genome although this is difficult to explain if conventional molecular genetics of mitochondrial DNA are assumed. Based on its unique ND1 sequence and ITS1 PCR–RFLP pattern, the cervid strain appears to represent a distinct genotype (designated G8) of E. granulosus.

The recombinant Echinococcus multilocularis antigen ll/3–10 is one of the most promising tools for immunodiagnosis of alveolar echinococcosis in human patients. Its nucleic acid sequence represents a part of the E. multilocularis gene encoding the metacestode antigen II/3, the former being basically present and expressed in both E. multilocularis and E. granulosus. Most (94%) patients with alveolar echinococcosis respond to infection with a marked anti-II/3–10 IgG synthesis; in contrast, most of the cystic echinococcosis patients do not, for some reason, recognize the recombinant antigen. We tackled this problem by generating cDNA derived from both E. granulosus and E. multilocularis full length II/3 genes, performed by reverse transcription and PCR amplification. Sequence analysis revealed a very high degree of conservation of the primary sequence of the antigen II/3 in both Echinococcus species. cDNA fragments were subcloned and expressed in E. coli as fusion proteins with Schistosoma japonicum glutathione S-transferase. Recombinant proteins were affinity purified and comparatively assessed by ELISA with respect to antibody-binding characteristics. Sera from patients suffering from cystic echinococcosis showed no significant differences in reactivity with the antigens derived from either E. multilocularis or E. granulosus. Therefore, parameters other than some minor differences in the primary sequence seem to be responsible for the lack of antigen II/3 recognition in cystic echinococcosis.

Note

Nucleotide sequence data reported in this paper have been submitted to the GenBank® data base with the accession numbers U05573 and U05574.

The localization and distribution of the serotoninergic components of the nervous system in the hydatid organism, Echinococcus granulosus, were determined by immunocytochemical techniques in conjunction with confocal scanning laser microscopy (CSLM). The distribution of serotonin immunoreactivity (IR) paralleled that previously described for cholinesterase activity, although it was more widespread. Nerve cell bodies and nerve fibres immunoreactive for 5-HT were present throughout the central nervous system (CNS), occurring in the paired lateral, posterior lateral and rostellar ganglia, their connecting commissures and nerve rings in the scolex and in the ten longitudinal nerve cords that run posteriorly throughout the body of the worm. A large population of nerve cell bodies was associated with the lateral nerve cords. In the peripheral nervous system (PNS), immunoreactive nerve fibres occurred in well-developed nerve plexuses innervating the somatic musculature and the musculature of the rostellum and suckers. The genital atrium and associated reproductive ducts were richly innervated with serotoninergic nerve cell bodies and nerve fibres.

Four-day-old worms of the tapeworms Hymenolepis microstoma, H. diminuta and H. nana and newly excysted H. microstoma were exposed in vitro at 37 °C to immune serum from mice infected for 4–1 2 weeks with H. microstoma. Worms were fixed for electron microscopy after intervals of 5 min to 96 h. Within 10–15 min an homogeneous precipitate occurred between the microtriches of 4-day-old H. microstoma and H. nana, while on some areas of H. microstoma the precipitate extended distal to the microthrix border and contained small vesicles (30 nm in diameter) and shed microtriches. In H. diminuta precipitates were not found until 2 h post-incubation. The thickness of the precipitate and the number of small vesicles and shed microtriches increased with time after incubation. Since a similar precipitate occurred on worms kept in complement-depleted immune serum, antibodies alone may induce immune damage. The precipitate on newly excysted H. microstoma lacked microthrix fragments. After 48 h an extensive precipitate was found protruding from the rostellar glands on some H. microstoma, and within the culture vessel. Antibodies may therefore be complexing with tapeworm secretory products.

A method was developed, which enables substances to be injected into the feeding structure (syncytium) established by Heterodera schachtii in roots of Arabidopsis thaliana. The technique was used to study the uptake of nutrients by the feeding nematode. The fluorescent dye lucifer yellow CH (LYCH) and fluorescence-labelled dextrans of different molecular weights were injected into the thin and translucent roots of A. thaliana. Such roots are a feature of this plant and they provide optimal conditions for microinjection. Injected LYCH was taken up by feeding juveniles and adults, indicated by the staining of the alimentary duct and the digestive system. Fluorescent dextrans of 3, 10 and 20 kDa but not of 40 and 70 kDa were ingested, suggesting that molecules of a maximum Stokes radius of 3·2 to 4·4 nm are taken up. It is likely that the feeding tube, forming the interface between the plant cytosol and the nematode's digestive system, is responsible for this size exclusion effect. The injected fluorescent substances were not detected in plant cells adjacent to the syncytium or in the root vascular elements. Injections into parts of roots which were infested by several nematodes revealed that feeding H. schachtii individuals may share one syncytium.

The presence of superoxide dismutase (SOD) activity in the bovine lungworm Dictyocaulus viviparus was examined using the xanthine–xanthine oxidase assay system and by non-denaturing PAGE followed by specific enzyme staining. High levels of activity were detected in excretory–secretory (ES) products of adult worms and in soluble extracts of both the L3 and adult stages of the parasite. Stage-specific and ES-specific activities were indicated by differences in SOD isoenzyme profiles between adult and larval parasite extracts and between adult extract and ES products, with a fast migrating activity being specific to ES products. All isoenzymes were sensitive to cyanide, indicating copper/zinc dependency. The antigenicity of ES SOD was demonstrated by a reduction in SOD activity in both the chemical assay and non-denaturing PAGE following incubation of parasite ES products with IgG antibody purified from serum of infected or vaccinated bovine hosts. The high level of SOD activity released by adult D. viviparus may be a reflection of the problems faced by a parasite occupying an oxygen-rich environment. Antibody inhibition of SOD may, therefore, be an important target of protective immunity.