Babesia vesperuginis is described from blood of two species of British bat: Pipistrellus pipistrellus and Myotis myslacinus. Reticulocytes appeared significantly elevated in blood films of P. pipistrellus infected with B. vesperuginis compared with uninfected laboratory-maintained bats or apparently uninfected wild-caught bats. Infected captive bats had significantly enlarged spleens. B. vesperuginis was transmitted by inoculation of infected blood to 5 uninfected captive P. pipistrellus. The course of infection followed a pattern of a rising parasitaemia accompanied by a rise in reticulocytes, followed by a fall in parasitaemia to low (<0·1%) or undetectable levels. When sacrificed, the Babesia-infected bats had significantly lowered blood haemoglobin, significantly raised white blood cell counts and enlarged spleens compared to uninfected bats. Attempts to transmit the parasite to irradiated and athymic ‘nude’ mice by inoculation of infected blood were unsuccessful. The experimental results and observations of infected wild bats indicate the potential pathogenicity of B. vesperuginis to bats. It is likely that the vector of B. vesperuginis is Argas vespertilionis because no Ixodid ticks were found on P. pistrellus.

Plasmodium falciparum-infected erythrocytes (IRBC) synthesize 3 histidine-rich proteins: HRP-I or the knob-associated HRP, HRP-II and HRP-III or SHARP. In order to distinguish these proteins immunochemically we prepared monoclonal antibodies which react with HRP-I, HRP-II and HRP-III, and rabbit antisera against synthetic peptides derived from the HRP-II and HRP-III sequences. A comparative analysis of diverse P. falciparum parasites was made using these antibodies and immunoprecipitation or Western blotting. HRP-I (Mr 80000–115000) was identified in all knob-positive P. falciparum parasites including isolates examined directly from Gambian patients. However, this protein was of lower abundance in these isolates and in 6 knob-positive, culture-adapted parasites compared to Aotus monkey-adapted parasites or culture-adapted parasites studied previously. HRP-II (Mr 60000–105000) was identified in all P. falciparum parasites regardless of knob-phenotype, and was recovered from culture supernatants as a secreted water-soluble protein. Within IRBC, HRP-II was found as a complex of several closely spaced bands. Cell surface radio-iodination of IRBC from several isolates and immunoprecipitation with a rabbit antiserum against the HRP-II repeat sequence identified HRP-II as a surface-exposed protein. Like HRP-I, the abundance of HRP-II was lower in the Gambian isolates than with Aotus monkey-adapted parasites studied earlier. Neither HRP-I nor HRP-II were identified in a knob-positive isolate of P. malariae collected from a Gambian patient. Analogues of these HRP were also absent from asexual parasites of diverse primate and murine malaria species screened with this panel of antibodies. HRP-III (Mr 40000–55000) was distinguished by its lower apparent size and by specific reaction with rabbit antibody against its 5-mer repeat sequence. HRP-III was of lowest abundance compared with the other two HRP. These antibody reagents and distinguishing properties should prove useful in studies on the separate functions of the 3 P. falciparum HRP.

Species- and subspecies-specific trypanosome DNA hybridization probes have been employed in the detection and identification of trypanosome infections in Glossina morsitans centralis. Several ways of sample preparation including the use of tsetse organ suspensions, proboscides and dissected midguts, as well as tsetse abdominal content touch-blots were explored. The results of hybridization of radio-isotope-labelled species-specific DNA probes to tsetse samples indicated that it was possible to detect trypanosomes in the organs where parasite development is known to characteristically occur for each subgenus. Duplicate slot-blots of samples prepared from midguts of tsetse infected with 2 strains of T. congolense and from non-infected fly controls show that it is not only possible to detect infection in tsetse but also to identify the strain of parasite present in a sample after hybridization with the DNA probes specific for each strain. The results, obtained after hybridization of sequential abdominal touch-blots from the same fly with the DNA probe specific for one strain of T. congolense, indicated that at least 8 positive signals can be observed after an overnight exposure. Because of their simplicity and potentially low cost, the techniques described here would be appealing for screening large numbers of tsetse samples from the field for the presence of any trypanosome residing in the guts or proboscis of the vector. In addition, the possibility of doing multiple touch-blots from the same fly gives the opportunity of detecting mixed trypanosome infections in the vector.

A range of monoclonal antibodies specific for Plasmodium falciparum were tested in vitro for their abilities to inhibit the multiplication of a partially synchronized culture of P. falciparum, to augment the phagocytosis of the parasites by macrophages, and to enhance the killing of parasites by peritoneal cells depleted of adherent cells. Seven of 17 monoclonal antibodies, ranging from culture supernatant fluid and ascitic fluid to purified IgG, showed dose- and time-dependent inhibition of parasite growth in vitro. At a concentration of 0·6 mg/ml, the inhibitory capacity of these monoclonal IgGs was above 94% over a 3-day culture period, much higher than that of the relevant polyclonal IgG. Four of 6 monoclonal antibodies tested augmented the phagocytosis of the parasites by macrophages, which occurred as a result of opsonization of the parasites. Four of 7 monoclonal antibodies examined showed cytotoxic activity on malaria parasites. Peritoneal cells depleted of adherent cells were capable of killing the parasites in the presence of monoclonal antibodies. These results indicate that there may be ‘monofunction’, ‘bifunction’, and ‘multifunction’ types of monoclonal antibodies against P. falciparum. The putative protective antigen of malaria parasites purified by ‘multifunctional monoclonal antibody’ affinity chromatography may have potential interest as a vaccine against the parasite or as an immunodiagnostic reagent for human malaria.

In the experiments reported here treatment (with a single dose or daily for 4 days) was delayed until 3 days after inoculation. Various combinations of M&B 35769, 2:4-diamino-5-[3(4–4′-chlorophenylphenoxy)propyl-l-oxy]-6-methylpyrimidine HC1, plus sulphadoxine, and of M&B 35769 plus dapsone, were examined and it was concluded that no universally ideal ratio of constituents in a combination is possible because the optimum ratio depends upon the activity of the constituents on their own and therefore varies from strain to strain. Activity was assessed on the 24th day after infection. M&B 35769 was markedly superior to pyrimethamine against all strains when the compounds were given on their own but, although it showed good potentiation with sulphadoxine and dapsone, much of its lead over pyrimethamine was lost in combination. M&B 36821 2–4-diamino-5-[3-(3,4′-dibromobiphenyl-4-oxy)-propyl-l-oxy]-6-methyl pyrimidine HC1, was more active than M&B 35769 against pyrimethamine- and cycloguanil-resistant strains but not against sensitive strains.

A comparative isoenzyme study between a natural population of Echinostoma caproni and 2 strains of E. caproni and E. togoensis, maintained for a long time in the laboratory led to the following conclusions. (1) The natural E. caproni population showed polymorphism of associated PGM and GPI genotypes. Some zymograms were identical to those of experimental strains. Allelic frequencies show that the natural population is panmictic according to the Hardy-Weinberg equilibrium. (2) The laboratory strains were monomorphic, and interbreeding produced double-heterozygous F1 progeny, identical to those of the natural population.

We have examined the interaction between Schistosoma mansoni and a cytotoxin from Pseudomonas aeruginosa known to affect a variety of cell types through the formation of membrane pores. The killing effect of the cytotoxin on S. mansoni in vitro was strongly dependent on the parasite developmental stage. Skin schistosomula were most sensitive, while 4-week-old or older parasites were less so. In contrast, lung schistosomula were relatively resistant and juvenile mesenteric forms younger than 28 days were almost completely refractory to the action of the toxin. A sharp increase in sensitivity to the toxin was observed between 27 and 28 days of parasite age with a parasite length of about 1·80 mm appearing as the threshold. Thus, the schistosome sensitivity to the cytotoxin changed during parasite development in a way similar to the known sensitivity of the worms to in vitro immune attack. Killing of the parasites by the cytotoxin was preceded by the formation of vesicles which, in the case of adult worms, detached easily from the parasites. Evidence obtained by electron microscopy and by immunofluorescent assays with living worms demonstrated that the outer parasite membrane participated in the formation of the vesicle membrane. Thus, the cytotoxin may be a useful tracer for changes in the properties of the schistosome surface membrane during parasite development.

Eimerian sporozoites can be recovered from intestinal washings after oral administration of oocysts to chickens but suspensions of sporozoites are usually prepared in the laboratory by incubation of sporocysts or fractured oocysts in vitro, at body temperatures, with relatively high concentrations of trypsin and bile salts. Since these agents affect membrane structure, the surface membrane of proteins of Eimeria tenella sporozoites excysted in vivo and in vitro have been compared. Surface radio-iodination followed by sodium dodecyl sulphate–polyacrylamide gel electrophoresis (SDS–PAGE) showed that more 125I was incorporated into polypeptides on sporozoites excysted in vivo than on sporozoites excysted in vitro. The 125I-polypeptide profile of sporozoites excysted in vivo was more resistant to subsequent incubation with pure trypsin than that of sporozoites excysted in vitro, but incubation with bile salts resulted in the loss of some iodinated polypeptides from both preparations of iodinated sporozoites. Reaction with combinations of crude trypsin and bile salts led to the lysis of sporozoites. The method of excystation had no effect on the reaction of convalescent chicken serum with Western blots of sporozoites but the results of immunofluorescent staining carried out with mouse monoclonal antibodies indicated that the structure of the cell surface was altered and some antigenic determinants were lost from sporozoites excysted in vitro. In contrast, neither the infectivity of sporozoites determined in vivo, nor their invasion of cultured cells was changed by the method of excystation.

Employing a simple method of growing ookinetes of Plasmodium falciparum in culture, 40% of mature gametocytes convert to macrogametes, 4% reach the retort-form ookinete stage and 0·45% become mature ookinetes. A single-step gradient centrifugation method on 12·5% Nycodenz® enriches both gametocytes and ookinetes.

The tegumental surface of Schistosoma margrebowiei as viewed by scanning electron microscopy is described. Both spined and unspined tubercles were found on the dorsal and dorso-lateral surfaces of sexually mature (in copula) male worms. Unpaired males lacked spined tubercles and the development of the spines is considered to occur only when worms are in copula. The females lack tubercles. The importance of tubercle structure in the identification of schistosome species and the effect of sexual maturity on their development is discussed.

l-Amino acid esters, such as leucine methyl ester (Leu-OMe) can destroy intracellular as well as isolated amastigotes of Leishmania mexicana amazonensis by a mechanism which may involve ester hydrolysis by parasite enzymes. We show here that several other esters prevented the killing of the amastigotes by Leu-OMe. Destruction of Leishmania within macrophages in culture was assessed microscopically and viability of isolated parasites was monitored by reduction of the tetrazolium MTT. The main features of the protective effect were similar for intracellular and for isolated amastigotes. Thus, (i) effective prevention of parasite killing required that the protective ester be present in the medium prior to and during exposure of infected cells or parasites to Leu-OMe; (ii) the same esters protected intracellular and isolated Leishmania against damage by Leu-OMe. Ranks of protective activity, as determined on isolated amastigotes were: Gly-OBz > Tyr-OMe > Ile-OMe > Met-OMe > Val-OMe > Ala-OMe > Gly-OMe > D-Leu-OMe; (iii) several esters were inactive in both systems (Leu-OBz, Trp-OMe and Phe-OMe). Protective activity was associated with leishmanicidal (e.g. Gly-OBz, Tyr-OMe) as well as with non-leishmanicidal (e.g. Ile-OMe, Val-OMe) esters. The results are compatible with the hypothesis that protective esters inhibit the activity of parasite enzyme(s) which hydrolyse Leu-OMe.

Of the surface antigens identified by radio-iodination, two-dimensional gel analyses showed no similarities between those of Schistosoma haematobium and Schistosoma mansoni, thus providing a basis for the species specificity of these antigens described previously (Simpson, Knight, Hagan, Hodgson, Wilkins & Smithers (1985) Parasitology 90, 499–508). The surface antigens of S. haematobium were glycosylated and comprised an acidic polypeptide of Mr 17000 as well as a complex set of polypeptides of approximate pI 6–7, which resolved in the Mr range 20000–30000. At least one of the lower Mr forms of this complex is also present in the adult worm. Limited cross-reaction was observed with S. mansoni infection sera and this may be due to a shared carbohydrate epitope. In contrast, extensive cross-reaction was observed using sera from mice immunized with S. bovis. This pattern parallels the species-specificity of vaccine-induced immunity. Extensive cross-reaction was also observed within cell-free translation products of m-RNA from adult worms of S. haematobium and S. mansoni by use of heterologous human infection sera. The few antigens which were species-specific may represent surface antigens.

Guinea-pigs immunized with equivalent numbers of normal or radiation-attenuated cercariae of Schistosomamansoni are known todevelop close to complete resistance to reinfection at weeks 12 and 4·5 respectively. We here analyse and compare the immune responses induced by the two populations of cercariae. Both radiation-attenuated and normal parasites of S. mansoni elicited an extensive germinal centre response in guinea-pigs by week 4·5 post-immunization. The anti-parasite antibody titre and cytotoxic activity of serum from 4·5-week-vaccinated, or 4·5-week-infected guinea-pigs were approximately equal, but sera from 12-week-infected individuals had high titres of anti-parasite antibody, which promoted significant larvicidal activity in vitro. In all cases, larvicidal activity was mediated by the IgG2 fraction of the immune serum. Lymphocyte transformation tests conducted on splenic lymphocytes from 4–5-week vaccinated guinea-pigs revealed maximal stimulation against cercarial, 2-week and 3-week worm antigens, whereas spleen cells from 4–5-week-infected guinea-pigs were maximally stimulated by cercarial and 6-week worm antigens. The splenic lymphocyte responses of 12-week infected animals were dramatic against antigens prepared from all life-stages of the parasite.

Larval Schistosoma mansoni have been shown to induce morphological changes to the internal calcium reserves (in particular the calcareous inclusions in Type A calcium cells and to the inner, nacreous layer of the shell) of Biomphalaria glabrata within 48 h of miracidial penetration. Control experiments have shown that these changes were not due to either the experimental procedures used, mechanical damage or to starvation effects. The effects were, however, analogous to experimentally induced acidosis, suggesting that the rapidly transforming miracidium-sporocyst quickly induces changes in the host's metabolism, presumably by the production and release of CO2 and waste metabolites into the haemolymph.

In some Biomphalaria glabrata–Schistosoma mansoni combinations snails are susceptible to infection as juveniles, but have variable susceptibility as adults. These snails become non-susceptible at the onset of egg-laying and typically revert to susceptibility in old age. Certain stocks of B. glabrata have the capacity to form amoebocytic accumulations in the atrium, and this ability is under genetic control. The atrial amoebocytic accumulations are transitory, typically appearing at onset of egg-laying and disappearing after a few months. A snail stock which has genetic tendencies for both adult variable susceptibility and atrial amoebocytic accumulations was studied. An association between the time of occurrence of adult non-susceptibility and atrial accumulation is revealed as snails never became infected with S. mansoni when amoebocytic accumulations were present. Developing parasites, however, were not necessarily encapsulated and destroyed by amoebocytes. Some sporocysts were able to delay development until the amoebocytic accumulations disappeared. The timing of atrial amoebocytic accumulations and resulting transient non-susceptibility in this host-parasite combination could influence snail population dynamics.

In an attempt to find possible targets for benzimidazole action in muscle-stage larvae of Trichinella spiralis, the effects of mebendazole and thiabendazole were tested in vivo by oral treatment of infested mice and in vitro by including these anthelmintics in an adequate maintenance medium containing decapsulated larvae. The effects of the anthelmintics on succinate dehydrogenase and fumarate reductase activities, measured in the mitochondrial fraction obtained from the in vivo- or in vitro-treated larvae showed that only thiabendazole causes significant inhibition of fumarate reductase activity. On the other hand, measurements of free glucose, glycogen reserves and soluble protein in the treated larvae indicate that in vivo, mebendazole and thiabendazole clearly diminish free glucose levels, although in vitro only mebendazole produces the same diminution. Both the glycogen and protein contents of the larvae remained unchanged after treatment in vivo or in vitro. The importance of these findings with regard to a possible site of action for mebendazole and thiabendazole is discussed.

A double antibody sandwich ELISA technique has been developed for detection of antigens of African trypanosomes present in the sera of infected mammals. The assay uses a high titre, high affinity rabbit antiserum made to purified membranes of procyclic trypanosomes as ‘capture’ reagent and a mixture of three biotin-labelled trypanosome-specific monoclonal antibodies as detecting reagent. The monoclonal antibodies were chosen on the basis of their specificity for surface membrane antigens of Trypanosoma brucei spp., the relative abundance and solubility of their specific antigen in aqueous solvents (including serum), and the fact that each monoclonal antibody binds to distinct epitopes on the same antigen molecule. Thus, antigen capture from serum and subsequent detection was achieved using as little as 10 ng/ml of whole trypanosome lysate, or the equivalent of 5000 trypanosomes/ml when solubilized material was added to normal serum in an artificial system. Using the optimized assay, antigen was detected in the sera of trypanosome-infected mice as early as 2 days after infection with T. b. rhodesiense. The results indicate that the assay allows detection of low concentrations of specific membrane antigens of T. brucei spp. of African trypanosomes and thus may have immunodiagnostic utility.

A laboratory life-cycle of Schistosoma bovis was established in order to study the host-parasite relationship in immunologically intact and T-cell deprived mice. Normal mice were found to have ‘self-cured’ their S. bovis infections almost completely by 10 weeks after cercarial administration, and there was no evidence of self-cure by day 79 in T-cell deprived animals, Thus, groups of deprived mice autopsied between 9 and 11 weeks after infection were invariably found to have greater worm burdens and a greater total number of eggs in the liver than comparably-infected normal mice. However, liver egg counts/worm pair were similar in the two types of host, and differences between normal and deprived mice with respect to total S. bovis egg counts in the intestine were also not consistently in the same direction in all experiments. Faecal egg counts were always less in deprived mice than in normal mice, even in an experiment in which the deprived mice had a significantly higher intestinal tissue egg count than the normal control group. The results are discussed in relation to the better known S. mansoni/mouse host-parasite relationship.

Protoscoleces of Echinococcus granulosus absorb the l-amino acids proline, methionine, leucine, alanine, serine, phenylalanine, lysine and glutamic acid by a combination of mediated transport and diffusion. All eight amino acids were accumulated against a concentration gradient. Comparison of Kt and Vmax values suggests that a low affinity for a particular compound is compensated for by a relatively larger number of transport sites for that compound. Four systems serve for the transport of the eight substrates studied: 2 for neutral (EgNl, EgN2) and 1 each for acidic (EgA) and basic (EgB) amino acids. All eight amino acids are incorporated into protein to varying degrees and substantial portions of absorbed l-alanine and l-methionine are metabolized into other compounds.

The plasma of mice infected with pleomorphic Trypanosoma brucei brucei contains a peptidase which has the same electrophoretic mobility on starch gels as a parasite peptidase. An enzyme with this electrophoretic mobility was not detected in the plasma of uninfected mice. The molecular weight of this enzyme in either parasite lysate or plasma from infected mice was approximately 40000 Da when assayed on a size exclusion column using high-performance liquid chromatography. The enzyme can cleave the dipeptides leu-ala, val-leu and pro-leu, but not the dipeptide phe-ala. The enzyme also cleaved the tripeptides tyr-tyr-tyr and leu-gly-gly. Another parasite peptidase which migrates on starch gels to a different position than the above-mentioned peptidase cleaved the dipeptides leu-ala, val-leu and pro-leu but could not cleave the tripeptides tyr-tyr-tyr or leu-gly-gly. Furthermore, incubation of this parasite peptidase with normal mouse plasma at 37 °C resulted in an apparent loss of detectable activity. It is postulated that the plasma of mice modifies either the charge or enzymic activity of this peptidase. We speculate that the parasite peptidase present in the plasma of mice infected with T. brucei could contribute to pathogenesis.