Echinococcus granulosus and Echinococcus multilocularis are cestode parasites, of which the metacestode (larval) stages cause the diseases cystic echinococcosis (CE) and alveolar echinococcosis (AE), respectively. Albendazole and mebendazole are presently used for chemotherapeutical treatment. However, these benzimidazoles do not appear to be parasiticidal in vivo against AE. In addition, failures in drug treatments as well as the occurrence of side-effects have been reported. New drugs are needed to cure AE and CE, which are considered to be neglected diseases. Strategies currently being implemented to identify novel chemotherapeutical treatment options include (i) conventional primary in vitro testing of broad-spectrum anti-infective drugs, either in parallel with, or followed by, animal experimentation; (ii) studies of drugs which interfere with the proliferation of cancer cells and of Echinococcus metacestodes; (iii) exploitation of the similarities between the parasite and mammalian signalling machineries, with a special focus on targeting specific signalling receptors; (iv) in silico approaches, employing the current Echinococcus genomic database information to search for suitable targets for compounds with known modes of action. In the present article, we review the efforts toward obtaining better anti-parasitic compounds which have been undertaken to improve chemotherapeutical treatment of echinococcosis, and summarize the achievements in the field of host-parasite interactions which may also lead to new immuno-therapeutical options.

Serum-derived fatty acids are essential for the intraerythrocytic proliferation of Plasmodium falciparum in humans. We previously reported that only limited combinations of fatty acids can support long-term parasite culture, and palmitic acid (C16:0)/oleic acid (C18:1, n-9), palmitic acid (C16:0)/vaccenic acid (C18:1, n-7), or stearic acid (C18:0) are required in these combinations, implying that these fatty acids are key molecules for intraerythrocytic parasite growth (Mi-Ichi et al.2006). Here, we analysed profiles of parasitaemia changes as well as morphologies during the erythrocytic cycle and confirmed the importance of C16:0 and C18:1, n-9. We also provide evidence that C18:1, n-9 but not other C18 monoenoic or dienoic acids maintain the synchronicity of parasite development in serum-free medium when paired with C16:0, resulting in maintained exponential growth. Thus, C18:1, n-9 is indispensable for the intraerythrocytic proliferation of P. falciparum.

The current trend of antimony (Sb) unresponsiveness in the Indian subcontinent is a major impediment to effective chemotherapy of visceral leishmaniasis (VL). Although contributory mechanisms studied in laboratory-raised Sb-R parasites include an up-regulation of drug efflux pumps and increased thiols, their role in clinical isolates is not yet substantiated. Accordingly, our objectives were to study the contributory role of thiols in the generation of Sb unresponsiveness in clinical isolates. Promastigotes were isolated from VL patients who were either Sb responsive (n=2) or unresponsive (n=3). Levels of thiols as measured by HPLC and flow cytometry showed higher basal levels of thiols and a faster rate of thiol regeneration in Sb unresponsive strains as compared with sensitive strains. The effects of antimony on generation of reactive oxygen species (ROS) in normal and thiol-depleted conditions as also their H2O2 scavenging activity indicated that in unresponsive parasites, Sb-mediated ROS generation was curtailed, which could be reversed by depletion of thiols and was accompanied by a higher H2O2 scavenging activity. Higher levels of thiols in Sb-unresponsive field isolates from patients with VL protect parasites from Sb-mediated oxidative stress, thereby contributing to the antimony resistance phenotype.

Gene expression is known to vary significantly during the Leishmania life-cycle. Its monitoring might allow identification of molecular changes associated with the infective stages (metacyclics and amastigotes) and contribute to the understanding of the complex host-parasite relationships. So far, very few studies have been done on Leishmania (Viannia) braziliensis, one of the most pathogenic species. Such studies require, first of all, reference molecular markers. In the present work, we applied differential display analysis (DD analysis) in order to identify transcripts that might be (i) candidate markers of metacyclics and intracellular amastigotes of L. (V.) braziliensis or (ii) potential controls, i.e. constitutively expressed. In total, 48 DNA fragments gave reliable sequencing data, 29 of them being potential markers of infective stages and 12 potential controls. Eight sequences could be identified with reported genes. Validation of the results of DD analysis was done for 4 genes (2 differentially expressed and 2 controls) by quantitative real-time PCR. The infective insect stage-specific protein (meta 1) was more expressed in metacyclic-enriched preparations. The oligopeptidase b showed a higher expression in amastigotes. Two genes, glucose-6-phosphate dehydrogenase and a serine/threonine protein kinase, were found to be similarly expressed in the different biological samples.

Leishmania (Leishmania) amazonensis is a protozoan of the American Continent that causes localized cutaneous leishmaniasis and, rarely, the diffuse cutaneous form of disease in humans. It has become clear in recent years that the course of Leishmania major infection in the mouse model differs when low numbers of purified metacyclic forms are used as inocula in comparison with the traditionally hitherto studied infection models that used large numbers of stationary-phase (SP) promastigotes. The low-number metacyclic inocula are thought to reproduce more closely the natural infection transmitted by the vector. In the present study the course of L. amazonensis infection, its local and distant dissemination patterns, and parasite load were compared in susceptible BALB/c and relatively resistant C57BL/6 mice infected in the footpad with inocula of 107 SP-promastigotes or with 104 purified metacyclic forms. Longer lag-phases were observed for infection with purified metacyclics but the characteristic patterns of disease susceptibility and cytokine production for either mouse strain were similar to those observed for SP-promastigote inocula. An inoculation dose of the order of 104 metacyclics was required to obtain consistent infections; 10- or 100-fold lower doses resulted in variable infection rates. Characteristically, L. amazonensis infection spread to distant organs and persisted there also in the relatively resistant C57BL/6 mice examined after 6 months of infection.

The sex ratio may be influenced by many factors, such as stress and immunosuppression, age of parents, parity and sex of preceding siblings. In animal systems, parasitism often changes the sex ratio of infected hosts, which can increase the probability of their transmission. The most common human protozoan parasite in developed countries, Toxoplasma gondii (prevalence 20%−80%), is known to change the behaviour of its intermediate hosts, thereby increasing the probability of transmission to its definitive host (the cat) by predation. The intermediate hosts, which under natural conditions are rodents, serve as the vector for Toxoplasma. Therefore, we speculate that Toxoplasma can alter the secondary sex ratio (i.e. male to female ratio in the offspring) of infected females to increase the proportion of (congenitally infected) male offspring, which are the more migratory sex in most rodent species. Here we studied the sex ratio of experimentally infected laboratory mice, expressed here as the proportion of males in the litter. In accordance with our hypothesis and results of previous retrospective cohort studies on human subjects, mice with toxoplasmosis produced a higher sex ratio than controls, in the early phase of latent infection. In the later phase of infection, mice with congenital toxoplasmosis had a lower sex ratio than controls, which is in accord with the Trivers-Willard hypothesis of sex ratio manipulation, suggesting that females in poor physical condition give birth to more female offspring.

Parasite transmission dynamics are fundamental to explaining the evolutionary epidemiology of disease because transmission and virulence are tightly linked. Horizontal transmission of microsporidian parasites, e.g. Nosema bombi, may be influenced by numerous factors, including inoculation dose, host susceptibility and host population heterogeneity. Despite previous studies of N. bombi and its bumble bee hosts, neither the epidemiology nor impact of the parasite are as yet understood. Here we investigate the influence N. bombi spore dosage (1000 to 500 000 spores), spore source (Bombus terrestris and B. lucorum isolates) and host age (2- and 10-day-old bees) have on disease establishment and the presence of patent infections in adult bumble bees. Two-day-old bees were twice as susceptible as their 10-day-old sisters, and a 5-fold increase in dosage from 100 000 to 500 000 spores resulted in a 20-fold increase in the prevalence of patent infections. While intraspecific inoculations were 3 times more likely to result in non-patent infections there was no such effect on the development of patent infections. These results suggest that host-age and dose are likely to play a role in N. bombi's evolutionary epidemiology. The relatively low levels of horizontal transmission success are suggestive of low virulence in this system.

Using a combination of morphological, life-history and molecular data, we redescribe Sphaerospora ranae (Morelle, 1929) n. comb. (previously Leptotheca ranae) and emend its taxonomic status. Renal infection was recorded in 2 spp. of frogs (out of 5 amphibian spp. examined), Rana dalmatina (proposed type host) and Rana temporaria, suggesting restricted host specifity of S. ranae. We provide a description of sporogonic stages of S. ranae for the first time and suggest possible modes of its developmental cycle. Phylogenetic analysis inferred from the small-subunit ribosomal DNA revealed a close relationship of S. ranae with piscine Sphaerospora elegans (type species of the genus) and Sphaerospora truttae, forming together with distantly related Leptotheca fugu a ‘Sphaerosporid clade’, the basal branch to all myxosporean species. The close relationship of the 3 Sphaerospora spp. is further supported by the presence of 2 areas with extensive nucleotide insertions in the V4 region of the SSU rDNA (absent in L. fugu), morphology and life-history features. We conclude, that the spore morphology of Sphaerospora s.l., is very simple and probably represents a ‘primitive’, basal morphotype retained in most myxosporean lineages. Based on presented data, we propose emendation of the genus Sphaerospora using morpological, life-history and molecular features.

Various mechanisms that enable and improve transmission success of myxozoan actinospore stages towards fish hosts are described, based upon a combination of experimental data and functional analysis of morphological characters. For this purpose, laboratory-reared actinospores of Myxobolus cerebralis, Myxobolus parviformis, Henneguya nuesslini and Myxobolus pseudodispar were employed to exemplarily investigate aspects of host attachment and invasion. The process of polar filament discharge of M. cerebralis actinospores was analysed, showing that full discharge occurs in less than 10 msec. Additionally, a mechanism that rapidly contracts the discharged filament after discharge is described for the first time. Its purpose is most likely to bring the actinospore apex rapidly into intimate contact with the surface of the host. Unlike M. cerebralis, M. parviformis actinospores did not discharge polar filaments after mechanical and chemical stimulation, suggesting a different mode of triggering. For H. nuesslini actinospores, experimental results indicated that polar filament discharge is independent of external calcium-ion concentration but is influenced by osmolality. After attachment of an actinospore and prior to penetration into the host, an ensheathed unit (‘endospore’), containing the sporoplasm, was emitted from the valves in a manner which varied from species to species. Experimentally induced sporoplasm emission was time-dependent and was found to be independent of polar filament discharge in H. nuesslini. Remarkably, it could be concluded that the sporoplasm is able to recognize host-stimuli while still within the intact spore. An updated summary of the sequential course of events during host recognition and invasion by actinospores is given.

The aims of the study were (1) to describe the Dactylogyrus communities living on selected South European Barbus species, (2) to analyse morphometric variability of their attachment and reproductive organs, and (3) to perform molecular phylogenetic analyses, in order to investigate the mode of speciation in Dactylogyrus species parasitizing congeneric hosts. In Bulgaria, Dactylogyrus crivellius, D. dyki and D. petenyi were found on B. balcanicus, and D. dyki on B. cyclolepis. In Spain, Dactylogyrus carpathicus and D. dyki were detected on B. meridionalis. Morphometric analyses of D. dyki revealed significant differences in the attachment and reproductive organs when individuals from different Barbus species were compared. Two monophyletic groups were recognized from the molecular phylogenetic analyses: the first included D. carpathicus and D. crivellius which have large body size and anchors, with a weakly supported basal position of D. malleus from B. barbus; the second included D. dyki and D. petenyi which have small body and anchor sizes. The comparison of host and parasite phylogenies did not indicate the intrahost speciation. Intraspecific molecular variability was found between individuals of D. dyki and D. carpathicus from different Barbus species, suggesting the need for a taxonomic revision for these species.

We tested the hypothesis that host specificity in ectoparasites does not depend exclusively on the features of the host but also on surrounding habitats, using 2 fish ectoparasites, Argulus coregoni and A. foliaceus (Crustacea: Branchiura), occurring sympatrically in Finnish lakes. Although these parasites are considered to be of low specificity, we found that the larger of the 2 species, A. coregoni developed a pronounced preference for salmonid hosts at the beginning of maturation (defined by the presence of copulating specimens). Argulus foliaceus infects a much wider range of fish hosts. We showed that specialization of A. coregoni on salmonids does not necessarily result from incompatibility with other fishes, but could instead reflect higher sensitivity of oxygen depletion compared with A. foliaceus. Adult A. coregoni may meet these demands by attaching to salmonids, the typical inhabitants of well-aerated waters. Young parasites of both species showed little host specificity and attached mainly to fishes with higher body reflectivity. In host choice experiments, A. coregoni of 4–5 mm length preferred salmonids (rainbow trout) to cyprinids (roach) irrespective of the type of fish host, on which it had been previously grown in the laboratory. We suggest that such an innate ontogenetic shift in host preference maintains the major part of the parasite population on its principal host, ensuring successful reproduction within suitable habitats.

Developing and adult Schistosoma mansoni and S. haematobium intact worms do not bind specific antibodies, likely because of structural and biochemical modifications of the outer lipid bilayer. We have estimated the amount of cholesterol in the apical membrane of adult schistosomes via extraction with the membrane-impermeable, cholesterol-binding drug, methyl-β-cyclodextrin (MBCD), followed by filipin staining of the worms, and evaluation of the amount of cholesterol released in the medium by a commercially available, enzymatic colorimetric assay. Positive correlations between amount of released cholesterol, MBCD concentration, and worm number and age provided evidence for the sensitivity and validity of the newly developed method. Treatment with 40 mm MBCD for 2 h at 37°C led to total loss of cholesterol from the worm outer membrane, as assessed by filipin staining, and the released cholesterol values were used to estimate the amount of cholesterol per worm and per an approximate surface area unit. Additionally, total depletion of outer membrane cholesterol was associated with exposure of surface membrane antigens to specific antibody binding in 50% and 70% of S. haematobium and S. mansoni worms, respectively. These findings together suggest that cholesterol is an essential, but not the sole, factor in sequestration of surface membrane antigens in schistosomes.

Expansion of human activities frequently results in habitat fragmentation, a phenomenon that has been widely recognized in the last decades as one of the major threats to world's biodiversity. The transformation of a continuous forest into a fragmented area results in a hyper-dynamic landscape with unpredictable consequences to overall ecosystem health. The effect of the fragmentation process on Trypanosoma cruzi infection among small wild mammals was studied in an Atlantic Rain Forest landscape. Comparing continous forest to fragmented habitat, marsupials were less abundant than rodents in the continuous landscape. An overall decrease in small wild mammal richness was observed in the smaller fragments. An anti-T. cruzi seroprevalence of 18% (82/440) was deteced by immunofluorescence assay. Moreover, this seroprevalence was higher in the fragmented habitat than in the continuous forest. According to the collected data, 3 main factors seem to modulate infection by T. cruzi in small wild mammals: (i) habitat fragmentation; (ii) biodiversity loss; (iii) increase of marsupial abundance in mammal communities. Furthermore, an extremely mild controlled infection by T. cruzi was detected since no patent parasitaemia could be detected in fresh blood samples, and no parasites were isolated by haemoculture.

A review of the Giardia duodenalis sequences currently available on the GenBank database was completed to compare the different genotyping loci (small subunit ribosomal DNA, glutamate dehydrogenase, triose-phosphate isomerase and beta giardin) for their ability to discern assemblage and subassemblage groups and infer phylogenetic relationships. In total, 405 Giardia duodenalis sequences were sorted and aligned to examine the substitutions within and between the assemblages – A and B (zoonotic), C and D (dogs), E (livestock), F (cats) and G (rodents). It was found that all of the genes could reproducibly group isolates into their assemblages and that the AI/AII subassemblage groups were robust and identifiable at all loci. However, the assemblage B subgroups were not reproducible at half of the loci (small subunit ribosomal DNA and beta giardin), not due to their conserved nature, but because there was insufficient sequence data of reference isolates available for comparison. It is anticipated that further investigation of these loci may reveal the core subgroups of this medically important and zoonotic assemblage and also those of others. The closer, more recent, phylogenetic relationships amongst the assemblages appear to be resolved; however, more sequence data from the current loci, and possibly new loci, will be required to establish the remaining relationships.

Neobenedenia girellae, a capsalid monogenean, is a significant pathogen due to both its ability to cause high mortality in fishes and its low host specificity. Established control methods have both advantages and disadvantages. Biological control measures with no unfavourable effects on the environment should be incorporated into the control strategy. The response of N. girellae oncomiracidia to brightness and black-and-white contrast was investigated to search for an alternative approach of disease prevention or control. Japanese flounder, Paralichthys olivaceus (Paralichthyidae), were exposed to oncomiracidia in an aquarium divided into areas of different brightness (∼1·3, 41·3 and 138·0 lux). The number of parasites on the fish group reared in 138·0 lux was significantly higher than on those reared in the lower brightness levels. Thus, the fish tended to be more vulnerable to infection by N. girellae under brighter conditions. Challenge trials using host fish mucus and whole live fish were established to detect the response by oncomiracidia to black-and-white contrast on a white versus a black background. Markedly more N. girellae oncomiracidia attached to black-painted areas and dark-coloured fish (normal spotted halibut, Verasper variegatus (Pleuronectidae) compared with white-painted areas and light-coloured fish (mal-coloured V. variegatus) on a white-coloured background. On a black-coloured background, more N. girellae oncomiracidia tended to attach to white-painted areas and light-coloured fish. Thus, black-and-white contrast is considered important for host finding by N. girellae oncomiracidia. The simplicity of the positive phototactic behaviour and the response to black-and-white contrast may lead to the development of a simple, practical and inexpensive method to control N. girellae outbreaks.

Cysteine proteinases from the fruit and latex of plants, such as papaya, pineapple and fig, have previously been shown to have substantial anthelmintic efficacy, in vitro and in vivo, against a range of animal parasitic nematodes. In this paper, we describe the in vitro effects of these plant extracts against 2 sedentary plant parasitic nematodes of the genera Meloidogyne and Globodera. All the plant extracts examined caused digestion of the cuticle and decreased the activity of the tested nematodes. The specific inhibitor of cysteine proteinases, E-64, blocked this activity completely, indicating that it was essentially mediated by cysteine proteinases. In vitro, plant cysteine proteinases are active against second-stage juveniles of M. incognita and M. javanica, and some cysteine proteinases also affect the second-stage juveniles of Globodera rostochiensis. It is not known yet whether these plant extracts will interfere with, or prevent invasion of, host plants.

Some parasites with complex life-cycles are able to manipulate the behaviour of their intermediate hosts in a way that increases their transmission to the next host. Gammarids infected by the tapeworm Cyathocephalus truncatus (Cestoda: Spathebothriidea) are known to be more predated by fish than uninfected ones, but potential behavioural manipulation by the parasite has never been investigated. In this study, we tested the hypothesis that C. truncatus is able to manipulate the behaviour of one of its intermediate hosts, Gammarus pulex (Crustacea: Amphipoda). To assess if any behavioural change was linked to other phenotypic alterations, we also measured the immunity of infected and uninfected individuals and investigated the pathogenic effects of the parasite. Infected gammarids were significantly less photophobic than uninfected ones, but no effect of infection on the level of immune defence was found. The results on survival, swimming activity and oxygen consumption suggest that the parasite also has various pathogenic effects. However, the alteration in host phototaxis was not correlated to some of these pathogenic effects. Therefore, we propose that the modification in host reaction to light is a behavioural manipulation, explaining the previously observed increase of gammarid predation rate.