Intraerythrocytic malaria parasites ingest the cytosol of their host cell and digest it inside their acid food vacuoles. Acidified (pH 4–5·5, 37 °C) human red blood cell lysates were used to simulate this process, measuring the denaturation of haemoglobin (Hb) and the release of iron, in the absence or presence of exogenous protease. Spontaneous Hb denaturation and appearance of non-heme iron were observed upon lysate acidification, their rates decreasing with increasing pH, and increasing in the presence of protease. Both processes were inhibited by the quinoline-containing anti-malarial drugs (QCDs) chloroquine, quinine, mefloquine and amodiaquine at concentrations well below those expected in the acidic food vacuole of the parasite. Spectrophotometric analysis indicated that chloroquine complexes with heme in acid-denatured haemoglobin. Other weak bases as well as verapamil and diltiazem, known to reverse the resistance of malarial parasites to chloroquine, were without efifect indicating that the action of QCDs is specific. Based on our previous results and the present report, we suggest that iron release in acidified lysates is mediated through the formation of ferryl (Fe(IV)) radicals. QCDs possibly complex with this radical, as they do with heme, and prevent its contact with an adjacent heme molecule which is required for ring opening and iron release. These results may suggest that one of the anti-malarial effects of QCDs is to deprive the parasite of an adequate iron supply. Addition of iron to cultures of Plasmodium falciparum was expected to circumvent the deprivation of iron and reduce the anti-malarial eflfect of QCDs. However, adding iron as penetrating fructose or nitrilotriacetate complexes did not alter the parasite's susceptibility to chloroquine. Ascorbate markedly increased the release of iron in acidified lysates, and this effect was not reduced by chloroquine. Ascorbate was found to decrease parasite susceptibility to chloroquine, suggesting that iron deprivation may be an important factor in the anti-malarial action of QCDs.

The activity of atovaquone against Plasmodium berghei ANKA during sporogonic development has been examined. Anopheles stephensi mosquitoes were fed on gametocyte infected mice which had been treated 8 h previously with atovaquone or diluent alone. Mosquito midguts were examined for oocysts, and salivary gland infections were estimated using an ELISA for the circumsporozoite protein (CSP). The number of oocysts per midgut fell by at least 97% when mosquitoes were fed on mice dosed with 0·1–10 mg atovaquone/kg body weight. This was paralleled by a decrease in the prevalence of oocyst-infected mosquitoes from 70–90% in controls to 40% or 10% respectively. No oocysts were observed at a dose of 100 mg/kg. CSP ELISA results indicated that mosquitoes fed on atovaquone failed to produce sporozoites. Mosquitoes which fed on gametocytaemic, atovaquone-treated mice (0·1–100 mg/kg) did not transmit malaria to naive mice. These results demonstrate that atovaquone has a highly potent inhibitory activity against the mosquito stages of P. berghei.

Rabbit polyclonal and mouse monoclonal antibodies (Mabs) prepared against synthetic peptides patterned on exofacialloops 3 (amino acids 546–555) and 7 (821–834) of the human anion transport protein band 3 inhibited the cytoadherence of Plasmodium falciparum-infected erythrocytes to C32 amelanotic melanoma cells. Mabs directed against exofacial loop4 (amino acids 628–642) did not inhibit adherence to a significant degree. The murine Mabs recognized only P. falciparum- infected erythrocytes suggesting that the epitopes of loops 3, 4 and 7 are normally cryptic in uninfected erythrocytes.

Glycosylation variants of the virulent Leishmania major clone VI21 were generated by mutagenesis with N-methyl-N-nitroso-N-nitroguanidine and selected using the galactose-specific lectin Ricinus communis II (RCA II). Three mutants, 4B9, 1D1 and 1C12, which failed to bind RCA II, were found to have an altered expression of lipophosphoglycan (LPG), a molecule implicated in the attachment to host macrophages and survival within the phagolysosome. There were differences in the antigenicity, molecular weight and localization of LPG from mutant parasites as compared to V121. Expression of gp63, a surface molecule also implicated in attachment to macrophages, was unaltered. All 3 mutants caused disease when injected into genetically susceptible BALB/c mice but lesions developed at a much slower rate than those caused by the virulent V121 clone. This slow rate of lesion development did not correlate with promastigotes' ability to invade macrophages in vitro. Karyotype analysis showed that there was a reduction in the size of chromosome band number 2 in all 3 mutants. The differences in LPG and chromosome band 2 were retained by mutant clones following passage through mice, suggesting that these phenotypes are stable. Although the mutant parasites were infective and caused lesions, the changed structure of the LPG appeared to influence the virulence of the parasites.

Prevalence of Toxoplasma gondii within 6 UK farmstead wild rat populations, and 1 population maintained within a captivecat-free environment for 2·5 years, was examined. The mean prevalence found was 35% (n = 235), which is more than 3 times as high as all other wild rat prevalence levels previously reported in the UK. There were no significant age, sexor site differences in prevalence between or within populations irrespective of habitat type or presence of cats. Toxoplasma was also maintained within the captive rat population in the absence of cats at a prevalence, intensity, age and sexdistribution similar to that of the farmstead rat populations. These results suggest, firstly, that Toxoplasma can beperpetuated within wild rat populations without the sympatric presence of cats and secondly, that the congenital route isthe predominant route of transmission in wild rats. This study concludes that wild rats represent a significant and persistent wildlife intermediate host reservoir for toxoplasmosis.

The question of whether the known immunosuppressive effects of Babesia microti may disrupt the development of acquired resistance to its tick vector, Ixodes trianguliceps, in natural rodent hosts (Clethrionomys glareolus), and thus enhance thedisease transmission potential, is addressed experimentally. The results show for the first time that natural hosts can acquire resistance to ticks; that this acquired resistance is manifested chiefly by a strongly density-dependent reduction in the percentage of attached larvae that engorge; that the density dependence is quantitatively similar whether the host receives occasional large tick challenges or frequent low infestations; but that infection with B. microti does not disrupt this pattern of acquired resistance. Of two important natural host species, Apodemus sylvaticus can support repeated infestations of I. trianguliceps, but is a poor host to B. microti, while C. glareolus develops acquired resistance to the tickvector, but supports much higher-level, longer-lasting B. microti infections.

Sarcocysts from the tongue muscle of a European badger (Meles meles) are reported for the first time and described by light and transmission electron microscopy. Judging from the ultrastructure of the cyst wall, the parasite is similar to the species Sarcocystis gracilis Rátz, 1909 sensu Erber, Boch & Barth (1978) from roe deer and possibly identical with it. This is noteworthy regarding the intermediate host specificity.

Muscle fibres isolated from adult Schistosoma mansoni contracted in a dose-dependent manner when exposed to elevated K+ with a maximum response obtained with 25 mM K+. These contractions were dependent on extracellular Ca2+ since Co2+ (5 mM) or nicardipine (1 μM) blocked the high K+ contractions. Serotonin (300 nM or higher) was required for maintenance of high K+ contractions. With concentrations of serotonin less than 300 nM the response was dose dependent. 5-Methoxytryptamine or α-methylserotonin at 1 μM as well as 10 μM tryptamine were able to substitute for serotonin, but 1 μM 5-carboxyamidotryptamine was ineffective. The order of potency for antagonists (10 μM) was: methiothepin > metergoline > Ly-278,584 = ketanserin. This pattern of responsiveness does not fit well with any known mammalian serotonin receptor subtype. Since forskolin, an adenylate cyclase activator, is able to mimic the action of serotonin and H89, a protein kinase inhibitor, is able to block the effect of serotonin, the effect of serotonin on contractility of the muscle may be via a cAMP-dependent pathway.

We have used two monoclonal antibodies, 128C3/3 and 504B1, to immunolocalize their carbohydrate epitopes in different developmental stages of Schistosoma mansoni. Both epitopes contain fucose: mAb 128C3/3, as we have shown previously, recognizes fucose in a novel, possibly internal linkage (Levery et al. 1992) while mAb 504B1, as we show here, bound to the Lex epitope, which contains fucose α1 → 3 linked to N-acetyl-glucosamine. The tissue expression of these epitopes was strikingly different and both elicit an immune response in infected hosts. The mAb 128C3/3-defined epitope was exposed on the surface of all larval stages but not on adult worms; however, it was found in the excretory system of adult worms of both sexes. In contrast, surface expression of the Lex epitope was initiated after the transformation of cercariae to schistosomula and was maintained throughout the adult life in both sexes.

Gyrdicotylus gallieni is a viviparous gyrodactylid monogenean that infects the buccal cavity and pharynx oi Xenopus laevis laevis. Offspring directly recolonize the same host as their parent and transmission is carried out exclusively by adult worms which are capable of leech-like stepping locomotion. The infrapopulation dynamics (at 20 °C) of this species were studied by experimental exposure to single worms of previously uninfected hosts (which were dissected at various time intervals post-infection). Infrapopulations increased exponentially until 50 days post-infection (p.i.), during which time the intrinsic rate of increase was estimated by regression as 0·065/parasite/day. After this, infrapopulation sizes showed greater variability, with established infections becoming extinct 2–5 months p.i.: elimination was probably due to someform of host reaction, as parasite lineages were maintained for up to 10 months by transfers to successive uninfected hosts.The development of infrapopulations is similar to that reported for other viviparous gyrodactylids parasitic on the bodysurface of teleosts. However, the intrinsic rate of increase was comparatively slow in G. gallieni and the duration of infection in isolated hosts prolonged. Slow-growing infrapopulations may elicit a host response more slowly and extend the period during which dispersal to a new host or host population can occur. This might be important in G. gallieni for which the internal site of infection could limit the transmission rate, as worm migration from the oral cavity or accidental detachment and expulsion is necessary for host-host transfer to occur. Exposure of wild caught X. l. laevis of unknown infectionhistory to 1 (n = 33) or 10 (n = 10) worms produced only 2 established infrapopulations (in both cases hosts initially infected by single worms), compared with establishment of over 60% in naive hosts infected with single worms: this suggests that host resistance may be an important factor in the population dynamics of G. gallieni.

Whole worm homogenates and excretory/secretory (E/S) products of adult Nippostrongylus brasiliensis significantly decreased the amplitude of contractions of segments of uninfected rat intestine maintained in an organ bath, with significantly larger volumes of E/S products being required to bring about a similar decrease to that caused by the whole worm homogenate. Boiled samples of homogenate and E/S products significantly decreased the amplitude of contractions of uninfected rat intestine, but larger volumes were needed than with unboiled samples. Frequency of contraction was unaltered by homogenates or E/S products. When electric eel AChE was injected into the lumen of segments of uninfected rat intestine maintained in an organ bath there was no significant decrease in the amplitude of contractions. These results suggest that substances present in the E/S products of N. brasiliensis significantly reduce the amplitude of contractions of uninfected rat intestine in vitro and that the biochemical holdfast responsible for this phenomenon may not be AChE.

The primary immune response to infection with Heligmosomoides polygyrus was studied in mice differing in response phenotype (fast-SWR, intermediate-NIH, slow-CBA). Marked IgGl and IgE but not IgG2a antibody responses were detected in infected mice and the former were more intense in fast compared with slow responder strains. Mastocytosis, MMCP-1, and the secretion of cytokines by mesenteric lymph node cells, following stimulation in vitro by Con A, were also more intense initially in SWR mice. Secretion of IL-4 declined in all strains by the 4th week of infection, irrespective of response phenotype. IL-10 was only produced briefly by SWR mice. However, the temporal patterns of secretion of IL-3 and IL-9 clearly distinguished fast from slow responder phenotypes. Following initial intense secretion of IL-3, production declined in all strains but in the 5–6th weeks enhanced secretion was evident in SWR and NIH mice and was sustained until week 10 p.i. In contrast, CBA mice never recovered from the initial down-regulation in weeks 3–4 and secretion declined to background levels by week 6 p.i. despite the continued presence of adult worms. Temporal changes in the secretion of IL-9 were very similar: secretion declined in CBA mice by week 6 p.i., whilst SWR and NIH mice continued to secrete high amounts. We suggest that fast and slow responder mice differ not only in their initial responsiveness to parasite antigens but also in their ability to sustain a Th2 response to the parasite and we propose that the latter is in part determined by their different susceptibilities to parasite-mediated immunomodulation. Only the fast responder strains can sustain a Th2 response of sufficient intensity to facilitate expulsion of adult worms.

The metamorphosis of oncospheres of the rat tapeworm, Hymenolepis diminuta, to mature metacestodes induces several pathophysiological effects in the intermediate host, Tenebrio molitor (Coleoptera). Previous investigations have failed to elucidate the mechanism responsible for changes in the host reproductive physiology and behaviour. This work forms part of an assessment of the degree to which nutrient resource management may be involved in these interactions. We report that developing metacestodes exert a pronounced effect upon host carbohydrate reserves and also alter circulating carbohydrate titres at some stages post-infection. Biochemical studies of fat body glycogen demonstrated a significant depletion of reserves as early as 3 days post-infection in males and 5 days post-infection in females. Gas chromatography revealed trehalose to be the predominant haemolymph carbohydrate, titres being significantly elevated in male beetles 5 and 9 days post-infection and in females 12 days post-infection. Metacestodes had no effect on glucose concentrations in male or female beetles.

Evidence for the presence of adipokinetic hormone/hypertrehalosaemic hormone (AKH/HrTH)-like peptides in the parasitic nematodes Ascaridia galli and Ascaris suum has been obtained using insect bioassays which measure hyperglycaemic responses to peptides belonging to the AKH/HrTH family of insect hormones. A peptide fraction extracted from heads and tails of Ascaridia galli evoked a dose-dependent hyperglycaemic response when injected into the cockroach, Periplaneta americana. Maximal bioactivity was obtained with material that was equivalent to 38 mg (wet weight) of nematode. Bioactivity appeared to be highest in extracts from heads and tails of both male and female worms and could be fractionated into at least three peaks of hyperglycaemic activity by reversed-phase high-performance liquid chromatography. An extract from heads and tails of A. suum also evoked a hyperglycaemic response when injected into the cockroach, Blaberus discoidalis. The bioactivity was inactivated on incubation with pure endopeptidase 24.11, confirming the peptidic nature of the bioactive material. These results provide evidence for the existence of peptides related to the insect AKH/HrTH family of peptides in parasitic nematodes.