Susceptibility to oxidative stress is a well-established feature of the malarial parasite. Pharmacologists have taken advantage of this property to design highly effective pro-oxidant antimalarial drugs. Less well appreciated is the fact that nutritional manipulation of host oxidative stress status by dietary means can have a profound effect on the growth of the parasite. In particular, rapid induction of vitamin E deficiency in mice by feeding highly unsaturated fatty acids (fish oil) strongly suppresses plasmodial growth. Likewise, the status of other antioxidant nutrients (e.g., riboflavin or vitamin C) may also influence the course of malarial infection under certain conditions. A combined nutritional pharmacology approach may offer some promise in controlling malaria.

Vaccination with whole male worm extract (AMA), stichoseome extract (SA) and a short-term inculbation fluid (EXA) of adult Trichuris muris induced a high degree of protective immunity in mice as assessed by reduction in larval worm burden.

Immunodiffusion of AMA against rabbit anti-AMA serum revealed five precipitin lines, one of which was common to both EXA and SA with which only single lines were obtained.

Physico-chemical tests suggested that the antigen is a protein and that carbohydrate, lipid and DNA do not contribute to its antigenicity. Partial fractionation was obtained with ammonium sulphate. All preparations with immunogenic activity showed the common line of EXA on immunodiffusion.

It is concluded that one of the protective immunogens is a protein which can be associated with the precipitin line and originates in the stichosome.

Several approaches were adopted for the disruption and removal of the tegumental surface from protoscoleces of the horse strain of the hydatid organism, Echinococcus granulosus. The effectiveness of each method and the purity of subsequent microthrix-enriched fractions obtained by differential centrifugation were evaluated by electron microscopy, by the amount of protein released and by the degree of enrichment of surface plasma membrane marker enzymes. Incubation in saponin for 10 min produced the purest microtriche preparation, but in low yield; freeze/thawing, incubation in Triton X-100 for 10 mm or in saponin for 20 min produced fractions containing significant amounts of relatively pure microtriches, but mild homogenization was a poor method for surface disruption and subsequent isolation of microtriches. Phosphodiesterase, adenosine triphosphatase (total and ouabain-inhibited), leucine aminopeptidase and glutamyltransferase were active in the protoscoleces but none were enriched in any of the microthrix fractions. In contrast, alkaline phosphatase, acid phosphatase, 5′ nucleotidase and maltase were enriched significantly in all of the isolated microtriche preparations, which suggests that these enzymes are predominantly surface membrane bound. The protein profiles of the microthrix-enriched fractions, following SDS—PAGE, were basically similar, although there were some qualitative and quantitative differences in the proteins released by each isolation procedure. Three major PAS-staining components were present in all the preparations and these probably originated from the glycocalyx. One of these PAS-positive components, with an approximate molecular weight of 110 kDa, may be a glycoprotein specific to the horse strain of E. granulosus.

1. The spores of Eimeria (not the oocysts) are the real infective agents and, as in other Sporozoa, they readily set free their sporozoites in the duodenum or in vitro under suitable conditions.

2. The protective oocyst has double walls: the ectocyst is thick and impervious to fluids but easily fractures by mechanical means, such as drying. Even though cracked, it may continue to protect the thin endocyst and spores until they reach the specially muscular regions of the alimentary canal. Free spores are probably also sometimes ingested.

3. The ectocyst is a continuous envelope without a persistent micropyle. It resembles keratin in many ways.

The rapid elimination of the intestinal phase of Trichinella spiralis in NIH mice is associated with progressive inflammation of the intestinal tract. The non-specific effects of this inflammation were studied in. mice concurrently infected with an unrelated parasite, Hymenolepis diminuta, which does not stimulate a visible inflammatory response but is also immunologically rejected by this strain of mice.

It was demonstrated that the rejection phase of T. spiralis infection had a marked effect upon the growth and survival of H. diminuta,. The cestode either failed to establish or to grow; if the worms were already strobilate when inflammation developed then destrobilation occurred. There was no cross–immunity between the parasites, nor was the interaction a direct consequence of inter–specific competition.

The metacyclic form of African trypanosomes is the first to express genes for the Variant Surface Glycoprotein (VSG) and it uses an unusually predictable subset of the VSG gene repertoire. We have developed a model system for the analysis of metacyclic VSG (M-VSG) gene expression and have used this to demonstrate that, for two M-VSG genes, different modes of expression operate in the insect and mammalian phases of the life-cycle. In metacyclic-derived clones, these genes are expressed in situ, whereas they are routinely activated by duplication in bloodstream trypanosomes. The expression loci for both M-VSG genes studied are structurally simple and we present a model, based on this, for the maintenance of a separate M-VSG repertoire and expression system.

The tegument of Proteocephalus pollanicoli is organized at two levels. The external level is a syncytial layer of protoplasm composed of vesicles in a granular matrix. The surface of this level has microvillus-like structure, microtriches, with sharp electron dense tips and less dense bases. Pinocytotic vesicles, large vacuoles, and evaginations occur at the surface of the external level, whereas mitochondria and lipid droplets tend to be concentrated basally. The basal plasma membrane has in some places small invaginations and at others large evaginations forming protoplasmic tubes which connect the external and internal levels.

The internal level is composed of individual areas of dense, granular protoplasm, each of which has a nucleus. These areas, or cuticular cells, contain lipid droplets, mitochondria, short lengths of agranular endoplasmic reticulum and phospholipid bodies. Two cuticular cells lying close together may be joined by a protoplasmic bridge and thereby form a limited syncytium.

The morphology of the tegument of this proteocephalid is compared with that of the cyclophyllidean species and the function of the tegument in parasitic helminths is discussed.

I should like to acknowledge the technical assistance of Mr W. Ferguson and Mr A. Lyness. My thanks are also due to the Wellcome Trust for a grant to purchase the Akashi Transcope and ancillary equipment.

Parasites can exert strong effects on population to ecosystem level processes, but data on parasites are limited for many global regions, especially tropical marine systems. Characterizing parasite diversity and distributions are the first steps towards understanding the potential impacts of parasites. The Panama Canal serves as an interesting location to examine tropical parasite diversity and distribution, as it is a conduit between two oceans and a hub for international trade. We examined metazoan and protistan parasites associated with ten oyster species collected from both Panamanian coasts, including the Panama Canal and Bocas del Toro. We found multiple metazoan taxa (pea crabs, Stylochus spp., Urastoma cyrinae). Our molecular screening for protistan parasites detected four species of Perkinsus (Perkinsus marinus, Perkinsus chesapeaki, Perkinsus olseni, Perkinsus beihaiensis) and several haplosporidians, including two genera (Minchinia, Haplosporidium). Species richness was higher for the protistan parasites than for the metazoans, with haplosporidian richness being higher than Perkinsus richness. Perkinsus species were the most frequently detected and most geographically widespread among parasite groups. Parasite richness and overlap differed between regions, locations and oyster hosts. These results have important implications for tropical parasite richness and the dispersal of parasites due to shipping associated with the Panama Canal.

Two full-length beta-tubulin cDNAs, representing isotypes 1 and 2, were cloned from the cattle nematode Cooperia oncophora. The predicted protein sequences span 448 amino acids, and show a high degree of identity to beta-tubulins from other nematodes. While C. oncophora isotype 1 sequence had the highest identity to Haemonchus contortus isotype 1 and Teladorsagia circumcincta sequences (95% identity), the C. oncophora isotype 2 sequence was most similar to H. contortus isotype 2 and Trichostrongylus colubriformis (92% identity). Alignment of the two C. oncophora sequences with other trichostrongylid beta-tubulins deposited in GenBank showed a clear distinction between isotype 1 and 2 beta-tubulin classes. The two classes differed at 19 amino acid positions, most notably at the carboxy terminus. These isotype-defining residues were conserved among different trichostrongylid species within a class. Analysis of fragments of both genes revealed a high degree of genetic variability in coding and non-coding regions. However, all nucleotide differences detected in the coding region were silent, as they did not result in any amino acid substitution. Analysis of 2 groups of worms for the codon 200 polymorphism associated with benzimidazole resistance revealed a proportion of worms in 1 of the groups bearing a tyrosine at this position.

Gyrodactylus specimens infecting both anadromous Arctic charr (Salvelinus alpinus) from River Signaldalselva (northern Norway) and resident Arctic charr from Lake Pålsbufjorden (southern Norway) were identified as G. salaris using molecular markers and morphometrics. The infection in Pålsbufjorden represents the first record of a viable G. salaris population infecting a host in the wild in the absence of salmon (Salmo salar). G. salaris on charr from Signaldalselva and Pålsbufjorden bear different mitochondrial haplotypes. While parasites infecting charr in Signaldalselva carry the same mitochondrial haplotype as parasites from sympatric Atlantic salmon, G. salaris from charr in Pålsbufjorden bear a haplotype that has previously been found in parasites infecting rainbow trout (Oncorhynchus mykiss) and Atlantic salmon, and an IGS repeat arrangement that is very similar to those observed earlier in parasites infecting rainbow trout. Accordingly, the infection may result from 2 subsequent host-switches (from salmon via rainbow trout to charr). Morphometric analyses revealed significant differences between G. salaris infecting charr in the 2 localities, and between those on sympatric charr and salmon within Signaldalselva. These differences may reflect adaptations to a new host species, different environmental conditions, and/or inherited differences between the G. salaris strains. The discovery of G. salaris on populations of both anadromous and resident charr may have severe implications for Atlantic salmon stock-management as charr may represent a reservoir for infection of salmon.

This review describes some of the developments in helminth biochemistry that have taken place over the last 40 years. Since the early 1970s the main anabolic and catabolic pathways in parasitic helminths have been worked out. The mode of action of the majority of anthelmintics is now known, but in many cases the mechanisms of resistance remain elusive. Developments in helminth biochemistry have depended heavily on developments in other areas. High throughput methods such as proteomics, transcriptomics and genome sequencing are now generating vast amounts of new data. The challenge for the future is to interpret and understand the biological relevance of this new information.

Hosts living on islands carry few parasite species, and the prevalence and intensity of directly transmitted parasites are often higher in insular than in mainland populations. However, it is unclear whether density or other features of insular populations can be responsible for the pattern observed. We compared the parasite richness, prevalence and intensity of parasites between 2 feral populations of cats living either at low density on an island (Kerguelen) or at high density on the mainland (Lyon). Parasite richness was higher in Lyon than in Kerguelen, where only Toxocara cati was found. T. cati egg prevalence was higher in Kerguelen (71·1%) than in Lyon (58·0%). Because cat density cannot explain this pattern, we propose that the low number of parasite species, the diet and/or immunity of cats act to increase prevalence in Kerguelen. Moreover, prevalence, intensity and variance-to-mean ratio increased with age and body mass in Kerguelen whereas, in Lyon, prevalence decreased with age and body mass. We hypothesize that the pattern of exposure differs between populations, and that density-dependent parasite mortality is lower in Kerguelen than in Lyon. We discuss the consequences concerning the influence of parasites on insular host populations.

The antimalarial drug mefloquine has promising antischistosomal properties killing haematophagous adult schistosomes as well as schistosomula. The mode of action and involved drug targets of mefloquine in Schistosoma mansoni schistosomula are unknown. In order to identify mefloquine-binding proteins and thus potential drug targets, mefloquine affinity chromatography with S. mansoni schistosomula crude extracts was performed. We found one specific mefloquine-binding protein that was identified by mass spectrometry as the glycolytic enzyme enolase (Q27877). Enolase activity assays were performed on schistosomula crude extracts and on the recombinant enolase Q27877 expressed in Escherichia coli. In schistosomula crude extracts enolase activity was inhibited by mefloquine and by the enolase inhibitor sodium fluoride, while activity of the recombinant enolase was not affected. In contrast to enolase from crude extracts, recombinant Q27877 did not bind to mefloquine-agarose. Using isothermal microcalorimetry, we next investigated the metabolic inhibition of mefloquine and 3 known glycolytic inhibitors in Schistosoma spp., namely sodium fluoride, 3-bromopyruvate and menadione on schistosomula in the presence or absence of glucose. We found that in the presence of glucose, schistosomula were less affected by mefloquine, sodium fluoride and 3-bromopyruvate, whereas glucose had no protective effect when schistosomula had been exposed to menadione. These results suggest a potential role of mefloquine as an inhibitor of glycolysis, at least in stages where other targets like haem degradation are not relevant.

Within aquatic habitats, the hyper-abundant Order Crustacea appear to be the predominant host group for members of the Phylum Microsporidia. The musculature, a common site of infection, provides access to biochemical (carbohydrate-rich) and physiological (mitochondria-rich) conditions conducive to prolific parasite replication and maturation. The significant proportion of body plan devoted to skeletal musculature in Crustacea provides the location for a highly efficient intracellular parasite factory. In this study, we utilize histological, ultrastructural and phylogenetic evidence to describe a previously known (Inodosporus octospora) and novel (Ovipleistophora arlo n. sp.) microsporidian parasites infecting the musculature of the common prawn (Palaemon serratus) from the same site, at the same time of year. Despite similar clinical signs of infection, both parasites are otherwise distinct in terms of pathogenesis, morphology and phylogeny. Based upon partial subunit ribosomal RNA (SSU rDNA) sequence, we show that that I. octospora may be identical to a Kabatana sp. previously described infecting two-spot goby (Gobiusculus flavescens) in Europe, or at least that Inodosporus and Kabatana genera are synonyms. In addition, SSU rDNA sequence for O. arlo places it within a distinct clade containing Ovipleistophora mirandellae and Ovipleistophora ovariae, both infecting the oocytes of freshwater fish in Europe. Taken together, our data provide strong evidence for trophic-transfer between crustacean and fish hosts for two different microsporidians within clade 5 of the phylum. Furthermore, it demonstrates that morphologically and phylogenetically distinct microsporidians can infect the same tissues of the same host species to impart clinical signs which mimic infection with the other.

The present study aimed to search for and characterize parasite molecules, whose expression levels correlate with the viability and growth activity of Echinococcus multilocularis metacestodes. We focused on the expression profiles of 2 parasite-derived genes, 14-3-3 and II/3-10, as putative molecular markers for viability and growth activity of the larval parasite. In experiments in vivo, gene expression levels of 14-3-3 and II/3-10 were relatively quantified by real-time reverse transcription-PCR using a housekeeping gene, beta-actin, as a reference reaction. All three reactions were compared with growth activity of the parasite developing in permissive nu/nu and in non-permissive wild type BALB/c mice. At 2 months p.i., the transcription level of 14-3-3 was significantly higher in parasites actively proliferating in nu/nu mice compared to parasites moderately growing in wild type mice. Immunoblotting experiments confirmed at the protein level that 14-3-3 was over-expressed in parasites derived from nu/nu mice at 2 months p.i. In vitro treatment of E. multilocularis with an anti-echinococcal drug nitazoxanide resulted in a significant decrease of both 14-3-3 and II/3-10 transcription levels found after 8 days of treatment, which correlated with the kinetics of a housekeeping gene, beta-actin. The conclusion is that 14-3-3, combined with II/3-10, exhibits good potential as a molecular marker to assess viability and growth activity of the parasite.

A primary objective of schistosomiasis control programmes is to achieve, and hence also demonstrate, a quantifiable reduction in schistosome-associated morbidity as a consequence of chemotherapeutic intervention. Inherent within such an objective, it is necessary to define and validate direct and indirect indicators of schistosome-related morbidity. However, to define and thereby document such morbidity, and its reduction following treatment, may not be straightforward, particularly for intestinal schistosomiasis-induced morbidity, which is often not apparent in all but the most severe or chronic cases. Within all ‘Schistosomiasis Control Initiative’ activities, across selected sub-Saharan African countries since 2002, a range of standard and novel potential morbidity markers have been monitored and evaluated. Parasitological intensity measures, combined with haemoglobin/anaemia counts and ultrasonography, proved valuable schistosomiasis-related morbidity indicators, being both logistically practical and informative. Additional measures tested, such as albumin excretion profiles, were promising, and are subject to ongoing research, whilst some measures, such as distended stomach/umbilical circumference, anthropometrics and health questionnaires proved less reliable. These results serve to both illustrate the success of current control activities in reducing schistosome-induced morbidity, and to highlight key tools and techniques for continued application within ongoing and future mass drug administration programmes.

We examined for the presence of trypanosomes in blood samples from 259 anurans (47 species from 8 families), the majority of which were from the Brazilian Amazonia, Atlantic Forest and Pantanal biomes. Trypanosomes were detected by a combination of microhaematocrit and haemoculture methods in 45% of the anurans, and 87 cultures were obtained: 44 from Hylidae, 22 from Leptodactylidae, 15 from Bufonidae, 5 from Leiuperidae and 1 from an unidentified anuran. High morphological diversity (11 morphotypes) was observed among blood trypanosomes from anurans of different species and of the same species as well as among trypanosomes from the same individual. Conversely, morphologically similar trypanosomes were found in anurans from distinct species and biomes. ITS and SSU rDNA polymorphisms revealed high diversity among the 82 isolates examined.† Twenty-nine genotypes could be distinguished, the majority distributed in 11 groups. Phylogenetic relationships based on rDNA sequences indicated that isolates from more phylogenetically related anurans are more closely related. Comparison of anuran trypanosomes from Brazil and other countries revealed several new species among the isolates examined in this study. Phylogenetic relationships suggest that host restriction, host switching and overall ecogeographical structure may have played a role in the evolution of the anuran trypanosomes.

The 4 species of ruminants (dromedary, zebu cattle, sheep and goat) in arid areas of Mauritania harboured Haemonchus spp. as the most frequent internal parasite. This was a rare situation where the 3 putative species, H. longistipes (dromedary), H. placet (zebu cattle) and H. contortus (sheep and goat) occurred sympatrically. The study was undertaken on hosts slaughtered at the Nouakchott abattoir, on the basis of monthly collection of worms. The environment was very unfavourable to H. placei and unfavourable to H. contortus, as intensity of infection remained low throughout the year, whereas infection in the dromedary was 10 to 20-fold higher. The survival strategies during the long, dry season were different: the surviving stages were either 4th-stage larvae in digesta (dromedaries), 4th-stage larvae either in digesta or mucosae (cattle), or 4th-stage larvae in mucosae and few adults (sheep and goats). The prolificacy of female worms, indicative of the potential to contaminate pastures, was similar for all Haemonchus spp. in the rainy season. H. longistipes behave differently during the pre-rainy season as no increase of prolificacy could be demonstrated as observed in the other species. Traits of vulvar morphology are considered as markers of ecological adaptation and were studied. The knobbed and smooth female morphs (in equal proportions) were the most frequent in H. longistipes, the knobbed morph out-numbered the other morphs in H. placei, and all 3 morphs were present in sheep and goats with the linguiform form being predominant. Genetic characterization of the 3 species was performed by means of Random Amplified Polymorphic DNA (RAPD). Three groups were obtained from analysis of these data: 1 group with individuals of H. contortus, 1 group with individuals of H. placei, and 1 group with individuals of H. longistipes. This indicated that, although the 3 species were valid, H. contortus and H. placei were more similar. Intraspecific variability was 2-fold higher in H. contortus than in the 2 other species. The ecological, morphological and genetical studies showed that H. longistipes, H. placei and H. contortus could be arranged in increasing order of variability.

The melanization of Dirofilaria immitis microfilariae in Armigeres subalbatus haemolymph in vitro is a two-step process. Firstly, the microfilariae are encased in a transparent capsule, then the capsule material is melanized later. Benzamidine HCl and p-amidinophenylmethanesulfonyl fluoride, both serine protease inhibitors, inhibited the deposition of the transparent capsule material and melanization. Diethyldithiocarbamate, a phenoloxidase inhibitor, did not prevent the deposition of the transparent capsule material but it did inhibit melanization. m-Hydroxybenzylhydrazine, a dopa decarboxylase inhibitor, had no inhibitory action on the deposition of the transparent capsule material. However, the time for the transparent capsule to become melanized was delayed due to the presence of m-hydroxybenzylhydrazine in the test system. Immunocytochemical localization showed that prophenoloxidase and/or phenoloxidase was present in the transparent capsule material. These results suggest (1) that prophenoloxidase cascade is essential in the melanization of microfilariae in mosquitoes and its putative activation behaviour is similar to that of other insects and (2) that the deposition of transparent capsule material is a result of the activation of prophenoloxidase by serine proteases and the melanization of the transparent capsule material is due to the presence of active phenoloxidase in capsule.

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.