Nicotinic acetylcholine receptors (nAChRs) are ligand-gated ion channels that mediate the fast actions of the neurotransmitter, acetylcholine. Invertebrate nAChRs are of interest as they are targets of widely-selling insecticides and drugs that control nematode parasites. Here, we report the cloning of ShAR2β, a candidate nAChR subunit from the blood fluke, Schistosoma haematobium, which is the third trematode nAChR subunit to be characterized. While ShAR2β possesses key structural features common to all nAChRs, its amino acid sequence shares considerably low identity with those of insect, nematode and vertebrate nAChR subunits. In particular, the second transmembrane domain of ShAR2β, which lines the ion channel, bears unusual amino acid residues which will likely give rise to a receptor with distinct functional properties. Phylogenetic analysis shows that ShAR2β is a divergent nAChR subunit that may define a clade of trematode-specific subunits. We discuss our findings in the context of potentially exploiting this receptor as a target for controlling schistosome parasites.

The study was aimed at developing better orally active albendazole (ALB) formulations. Six formulations (ALB-1 to ALB-6) were prepared and tested against Brugia malayi in Mastomys coucha and jird (Meriones unguiculatus) at 200 mg/kg, orally, for 5 consecutive days. The anti-filarial efficacy was assessed against microfilariae (mf), adult worms and female reproductive potential. Three of the 6 ALB formulations showed greatly improved female worm sterilizing potential (ALB-1: 90%; ALB-3: 63%; ALB-4: 77% of untreated control) in B. malayi – M. coucha model. Sterilization efficacy of ALB-1 was also better than that shown by pure-ALB (P<0·001) or its marketed tablet formulation, Zentel (P<0·01), while that of ALB-4 was better than pure-ALB (P<0·05). The activity of ALB-3, pure-ALB and Zentel was, however, comparable. ALB-1 also showed late microfilaricidal activity with a maximum of 78% fall in microfilarial count. In contrast, neither the pure ALB nor Zentel showed any microfilaricidal activity. In the jird – B. malayi model, ALB-1 and ALB-4 showed marginal sterilizing efficacy whereas pure ALB or Zentel were ineffective. In conclusion the anti-filarial efficacy of ALB-1 was found to be superior to pure-ALB or Zentel.

The reservoir capacity of domestic cats and dogs for Trypanosoma cruzi infection and the host-feeding patterns of domestic Triatoma infestans were assessed longitudinally in 2 infested rural villages in north-western Argentina. A total of 86 dogs and 38 cats was repeatedly examined for T. cruzi infection by serology and/or xenodiagnosis. The composite prevalence of infection in dogs (60%), but not in cats, increased significantly with age and with the domiciliary density of infected T. infestans. Dogs and cats had similarly high forces of infection, prevalence of infectious hosts (41–42%), and infectiousness to bugs at a wide range of infected bug densities. The infectiousness to bugs of seropositive dogs declined significantly with increasing dog age and was highly aggregated. Individual dog infectiousness to bugs was significantly autocorrelated over time. Domestic T. infestans fed on dogs showed higher infection prevalence (49%) than those fed on cats (39%), humans (38%) or chickens (29%) among 1085 bugs examined. The basic reproduction number of T. cruzi in dogs was at least 8·2. Both cats and dogs are epidemiologically important sources of infection for bugs and householders, dogs nearly 3 times more than cats.

Leishmania infantum and Leishmania donovani both pertain to the L. (L.) donovani complex and are responsible for visceral leishmaniasis. To explore the L. donovani complex, we focused our study on cysteine protease B (cpb) and especially on 2 cpb copies: cpbE and cpbF. We selected cpb genes because of their phylogenetic interest and host–parasite interaction involvement. Sequencing these 2 copies revealed (i) that cpbE is specific to L. infantum and cpbF is specific to L. donovani and (ii) that these 2 copies are different in length and sequence.Nucleotide sequence data reported in this paper are available in the GenBank database under Accession numbers AY896776 AY896777, AY896778, AY896779, AY896780, AY896781, AY896782, AY896783, AY896784, AY896785, AY896786, AY896787, AY896788, AY896789, AY896790, AY896791. Phylogenetic analysis and protein predictions were carried out in order to compare these copies (i) with other trypanosomatid cpb, especially L. mexicana, and (ii) within the L. donovani complex. Our results revealed patterns specific to the L. donovani complex such as the COOH-terminal extension, potential epitopes and N-glycosylation sites. Moreover, phylogenetic analysis revealed different levels of polymorphism between L. infantum and L. donovani and confirmed the ancestral status of the latter. L. infantum has a shorter sequence and a deleted sequence responsible for modifications in protein conformation and catalytic triad. Considering the clinical aspect, L. infantum dermotropic strains appeared more polymorphic than L. infantum viscerotropic strains.

The protozoan parasite Neospora caninum is an important cause of reproductive disease in cattle worldwide. The dog is a definitive host for the parasite and the oocyst stage, shed in the faeces, is a source of infection for cattle through consumption of contaminated feed or water. In addition, transplacental transmission of N. caninum is a very efficient means of the parasite infecting a new host and this can occur in successive pregnancies and over several generations. Neospora parasites may cause disease during pregnancy resulting in death of the foetus or birth of live congenitally infected calves that may show some neurological clinical signs at birth. The stage of pregnancy at which infection/parasitaemia occurs is an important factor in determining disease severity. Neospora infection in the first trimester of pregnancy may have more severe consequences for the foetus compared with infection occurring in the final trimester. The host-parasite relationship during pregnancy is a fascinating interaction and research in this area will improve understanding of disease pathogenesis and the various consequences of the host immune response, being host-protective, parasite protective and contributing to disease pathology. Pregnancy poses an interesting problem for the immune system of the dam as she is essentially carrying a semi-allogeneic tissue graft (the foetus) without immunological rejection taking place. To facilitate the pregnancy the cytokine environment in the placenta favours the regulatory Th-2-type cytokines, whose role is to counteract the pro-inflammatory Th1-type immune responses. Protective immunity to N. caninum, similar to many other intracellular parasites, involves Th1-type immune responses, which may pose problems for the dam trying to control a Neospora infection during pregnancy.

This paper will discuss the host-parasite relationship at different stages of gestation in pregnant cattle and review the implications of this research for our understanding of disease pathogenesis, parasite transmission and host protection.

The mechanism of action of praziquantel (PZQ), the drug of choice against schistosomiasis, is still unclear. Since exposure of schistosomes to the drug is associated with calcium influx and muscular contraction, calcium channels have been suggested as the target, although direct combination of PZQ with their subunits was never demonstrated. We report a hitherto unknown effect of PZQ, namely the inhibition of nucleoside uptake, as observed in living worms using radio-isotope labelled adenosine and uridine. This effect is clearly seen in schistosomes but is absent in mammalian cells in culture. Moreover it is a specific pharmacological effect seen exclusively with the active levo-R(−)stereo isomer of the drug, and is shared by at least one benzodiazepine having antischistosomal activity. This novel effect acquires significance given that schistosomes cannot synthesize purine nucleosides de novo. A possible relationship between this novel effect and the known action of PZQ on calcium channels is discussed, since adenosine is known to bind to specific receptors and to behave as an indirect antagonist of calcium release in mammalian cells. If calcium channels were correlated with adenosine receptors also in schistosomes, as they are in mammals, this would support the hypothesis that PZQ-induced calcium influx may be correlated to adenosine receptor blockade.

We have recently identified 2 surface proteins in Entamoeba histolytica as intermediate subunits of galactose- and N-acetyl-D-galactosamine-inhibitable lectin (EhIgl1 and EhIgl2); these proteins both contain multiple CXXC motifs. Here, we report the molecular characterization of the corresponding proteins in Entamoeba dispar, which is neither pathogenic nor invasive. Two Igl genes encoding 1110 and 1106 amino acids (EdIgl1 and EdIgl2) were cloned from 2 strains of E. dispar. The amino acid sequence identities were 79% between EdIgl1 and EdIgl2, 75–76% between EdIgl1 and EhIgl1, and 73–74% between EdIgl2 and EhIgl2. However, all the CXXC motifs were conserved in the EdIgl proteins, suggesting that the fold conferred by this motif is important for function. Comparison of the expression level of the Igl genes by real-time RT-PCR showed 3–5 times higher expression of EdIgl1 compared to EdIgl2. Most EdIgl1 and EdIgl2 proteins were co-localized on the surface and in the cytoplasm of trophozoites, based on confocal microscopy. However, a different localization of EdIgl1 and EdIgl2 in intracellular vacuoles and a different level of phenotypic expression of the two Igls were also observed. These results demonstrate that Igls are important proteins even in non-pathogenic amoeba and that Igl1 and Igl2 may possess different functions.

This study investigated a new effective method for controlling the capsalid monogenean Neobenedenia girellae. We examined in vitro and in vivo the effect on the percentage survival of N. girellae in buffers containing different metallic ions. Decreased survival was observed in buffer solutions lacking two ions. In particular, the percentage survival of N. girellae was significantly decreased after 10 min exposure to buffer containing neither Ca2+ nor Mg2+. Transmission electron microscopic observations showed that treatment with this buffer disrupted intercellular junctions. This significant effect on percentage survival of N. girellae using Ca2+/Mg2+-free buffer was confirmed in an in vivo assay. Ca2+/Mg2+-free buffer had no effect on the condition of the host, spotted halibut Verasper variegates (Pleuronectidae). These results suggest that treatment with Ca2+/Mg2+-free buffer is a new effective control method, which could replace existing control methods.

Within the genus Cryptosporidium 2 lineages have evolved, one adapted to the acid environment of the stomach and abomasum, the other comprising parasites that multiply in the small intestine. We tested whether the release of sporozoites from oocysts, a process known as excystation, is triggered by conditions which mimic the site of infection. Specifically, we exposed oocysts from gastric and intestinal Cryptosporidium species to acid conditions or to a neutral solution of taurocholic acid, at 37°C. We found that oocysts from the gastric species C. muris and C. andersoni excysted in both conditions, whereas the intestinal species C. parvum and C. hominis did not respond to acid. When the effect of temperature alone was tested on C. muris and C. parvum, only oocysts from the former species excysted in significant numbers. Oocysts from intestinal species did not respond to temperature alone, nor to acidity. These observations are consistent with the need of gastric species to rapidly excyst and release the sporozoites upon ingestion, and indicate that Cryptosporidium oocysts have evolved to maximize delivery of sporozoites to the region of the gastro-intestinal tract where the parasite multiplies.

Two morphologically novel Kudoa species are characterized from brain tissue of fish, Kudoa chaetodoni n. sp. from Chaetodon unimaculatus (Chaetodontidae) and Kudoa lethrini n. sp. from Gymnocranius audleyi and Lethrinus harak (Lethrinidae). Additionally we characterized a 5-spore valve (SV) Kudoa species from the brain of Sillago ciliata (Sillaginidae). Intriguingly, its 18S rDNA sequence was identical to that of the 7 SV Kudoa yasunagai extracted from the brain of a paralichthyid halibut in Japan. These 2 species may either prove to be con-specific, even though morphology and distribution differ, or demonstrate the limit of specific resolution in the small subunit rDNA gene region. Small subunit rDNA sequences from these new species were used in molecular phylogenetic analyses of kudoids to examine congruence of phylogeny with tissue tropism, geographical distribution, and host specificity. There was significant correlation between tissue tropism in the form of well-supported brain and heart-infecting clades. Host specificity and geographical distribution showed some correlations with genotype.

Although Ixodes ricinus ticks are mainly associated with woodland, they are also present in open habitat such as pastures. The distribution of nymphal I. ricinus was monitored by drag sampling the vegetation in May–June 2003 on 61 grazed permanent pastures for cattle located in central France. After selecting explanatory variables from among a set of 155, tick abundance was modelled on the perimeter of the pasture using a negative binomial model that took into account data overdispersion. An abundant tree layer at the perimeter of the pasture associated with a high humidity before sampling greatly enhanced the average number of captured I. ricinus nymphs. The presence of apple or cherry trees around the pasture perimeter, the presence of trees or bushes at the pasture edge, woodland around the pasture and a high number of I. ricinus nymphs in the nearest woodland to the pasture were also favourable to nymph abundance in the pasture. The study highlighted that woodland vegetation associated with humidity and the presence of attractive foraging areas for tick hosts around the pasture played a key role in the abundance of I. ricinus. Finally, the results raised the question of whether and how transfer of ticks between woodland and grazed pastures occurs.

We studied the effect of density on feeding success of 2 fleas, Xenopsylla conformis and Xenopsylla ramesis, when exploiting rodents Meriones crassus and Gerbillus dasyurus. We tested 2 alternative hypotheses: (i) that intraspecific interference competition occurs and, thus, feeding success of a flea decreases with an increase in density and (ii) that facilitation via suppression of a host defence system occurs and thus, feeding success of a flea increases with an increase in density. The mean size of a bloodmeal and the proportion of highly engorged individuals in X. conformis feeding on both hosts were affected by density. When on G. dasyurus, both the size of a bloodmeal and the proportion of highly engorged individuals were lower at low (5–15 fleas per host) than at high (25–50 fleas per host) densities. The opposite was true when this flea fed on M. crassus. The mean bloodmeal size and proportions of highly engorged X. ramesis parasitizing either host were not affected by flea density. This study showed that the density dependence of feeding success of a flea (a) varied both between fleas and within-fleas between hosts and (b) indicated either intraspecific competition or facilitation via the host in a particular flea-host association.

A cDNA encoding the immunogen Pso o 1 from Psoroptes ovis was obtained by polymerase chain reaction (PCR) amplification. The amplicon contained the entire coding sequence for the prepro-enzyme in an open reading frame (ORF) of 966 bp. This gene encoded a predicted protein of 322 amino acids (aa) with 64% aa identity (80% similarity) to the major house dust mite faecal allergen Der f 1. The pro-enzyme form of Pso o 1 was expressed as a recombinant protein in the Pichia pastoris-eukaryotic expression system. Maturation of the recombinant pro-enzyme by autocatalytic activation was not observed, and such maturation could not be achieved using a number of techniques known to activate recombinant Der p 1 and Der f 1 expressed in the same system. Serum raised against recombinant Pso o 1 cross-reacted with mature Der p 1 and allowed Pso o 1 to be immunolocalized to the gut of P. ovis.

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.

Species of Protopolystoma are monogenean flukes that only infect allopolyploid hosts in the anuran genus Xenopus. Multivariate analyses of morphometric sclerite characters in the nominal species Protopolystoma simplicis suggest that morphologically distinguishable populations occur in the tetraploid host, Xenopus laevis victorianus, and in each of the octoploid hosts, X. vestitus and X. wittei. The species-level divergence of a lineage specific to X. laevis is supported by sequence variation in the mitochondrial cytochrome c oxidase subunit 1 (cox1) gene. Protopolystoma simplicis from X. laevis is redesignated P. microsclera n. sp., with P. simplicis being retained for populations in octoploid hosts. This division is consistent with large differences in egg hatching schedule, fixed differences at the mannose-6-phosphate isomerase and fumarate hydratase loci, and host-specificity in experimental analyses. Although the respective P. simplicis populations in X. vestitus and X. wittei also show significant diversity in allozyme expression, morphometrics and egg hatching schedule, they are retained in the same species because their level of mitochondrial DNA divergence is similar to that found within other Protopolystoma species. The consequences of splitting P. simplicis for a recent interpretation of the origin of Protopolystoma faunas in octoploid Xenopus spp. is discussed.

As a consequence of comprehensive transcriptome analysis followed by sequencing and draft assembly of the genome, the emphasis of schistosome research is shifting from the identification of genes to the characterization of their functions and interactions. Developmental biologists have long used whole mount in situ hybridization (WISH) to determine gene expression patterns, as a vital tool for formulating and testing hypotheses about function. This paper describes the application of WISH to the study of gene expression in larval and adult schistosomes. Fixed worms were permeablized by proteinase K treatment for hybridization with digoxygenin-labelled RNA probes, with binding being detected by alkaline phosphatase-coupled anti-digoxygenin antibodies, and BM Purple substrate. Discrete staining patterns for the transcripts of the molecules Sm29, cathepsin L, antigen 10.3 and chorion were observed in the tegument cell bodies, gut epithelium, oesophageal gland and vitelline lobules, respectively, of adult worms. Transcripts of the molecules SGTP4, GP18-22 and cathepsin L were localized to tegument cell bodies and embryonic gut, respectively, of lung schistosomula. We also showed that Fast Red TR fluorescent substrate can refine the pattern of localization permitting use of confocal microscopy. We believe that method of WISH will find broad application, in synergy with other emerging post-genomic techniques, such as RNA interference, to studies focused at increasing our molecular understanding of schistosomes.

Polistes dominulus are social wasps which are the host of the strepsipteran endoparasite Xenos vesparum. In the hibernating phase, unparasitized and parasitized wasps leave natal nests and aggregate together in sheltered quarters. In aggregations, wasps are socially active, and some individuals perform helping behaviour. Here we investigated if castrated parasitized wasps perform worker tasks in mixed aggregations. Moreover, by gas chromatography and mass spectrometry, we examined the cuticular hydrocarbons of unparasitized and parasitized wasps to evaluate if the infection alters the composition of cuticular waxes that are recognition cues in social insects. In clusters, infected females do not perform helping behaviour and they are less active than unparasitized wasps. Cuticular hydrocarbons are slightly differentiated between unparasitized and parasitized wasps but, generally, unparasitized wasps are more similar to wasps infected by Xenos females compared to wasps infected by Xenos males. Wasps infected by Xenos males do not usually survive the winter. This chemical similarity is probably a consequence of the similar physiological condition of unparasitized and female-affected Polistes wasps. At this stage, it is difficult to affirm whether these modifications are a true parasite manipulation or a consequence of infection.

This study begins to redress our lack of knowledge of the interactions between colonial hosts and their parasites by focusing on a novel host-parasite system. Investigations of freshwater bryozoan populations revealed that infection by myxozoan parasites is widespread. Covert infections were detected in all 5 populations studied and were often at high prevalence while overt infections were observed in only 1. Infections were persistent in populations subject to temporal sampling. Negative effects of infection were identified but virulence was low. Infection did not induce mortality in the environmental conditions studied. However, the production of statoblasts (dormant propagules) was greatly reduced in bryozoans with overt infections in comparison to uninfected bryozoans. Overtly-infected bryozoans also grew more slowly and had low fission rates relative to colonies lacking overt infection. Bryozoans with covert infections were smaller than uninfected bryozoans. High levels of vertical transmission were achieved through colony fission and the infection of statoblasts. Increased fission rates may be a strategy for hosts to escape from parasites but the parasite can also exploit the fragmentation of colonial hosts to gain vertical transmission and dispersal. Our study provides evidence that opportunities and constraints for host-parasite co-evolution can be highly dependent on organismal body plans and that low virulence may be associated with exploitation of colonial hosts by endoparasites.

The aim of the present study was to analyse the relation between parasitism and subsequent metallothioneins (MT) in the case of metal contamination. Experimental exposure of parasitized and unparasitized cockles (Cerastoderma edule) to cadmium (Cd) was performed, with the cockle as first or second intermediate host of 2 digenean species. After 7 days of Cd exposure in microcosms, cockles infected as first intermediate host by Labratrema minimus exhibited metal concentrations in tissues double that in uninfected cockles. Jointly, MT concentrations of parasitized cockles were not modified in comparison with uninfected individuals in which concentrations were increased 4·3-fold compared with controls. In cockles experimentally infected as the second intermediate host by Himasthla elongata, cadmium concentrations significantly increased again in parasitized cockles compared with uninfected individuals in contaminated conditions. Simultaneously, MT concentrations in healthy cockles increased, whereas they significantly decreased in parasitized individuals. Therefore, the presence of digenean parasites in Cd-exposed cockles leads to a maintenance or a decrease in MT concentrations compared with healthy individuals, whereas Cd accumulation in tissues is significantly increased. These experiments indicate a significant alteration of the protective effect of metallothioneins towards metals which could consequently enhance cockle vulnerability. Moreover, these results highlight the limit of the use of MT as a biomarker of metal pollution in field monitoring if parasitism is not taken into account.

We report 21 thelastomatoid species parasitizing 31 described and 5 undescribed geoscapheine and panesthiine cockroaches, representing all but 1 of the known species of these subfamilies in Australia. The nematodes have 3 distinct patterns of host distribution: dominant, moderate and rare. The 4 dominant species, Cordonicola gibsoni, Leidynemella fusiformis, Travassosinema jaidenae and Aoruroides queenslandensis, are highly prevalent, found in nearly all host species examined, and broadly distributed. The 8 moderate species have lower prevalences but are still widely distributed. Many of these species are more common in one host subfamily than the other. The remaining 9 rare species have highly restricted host and geographical distributions. Six of the 21 species are exclusive to geoscapheines, 5 to panesthiines and 10 are shared. These patterns suggest that most of the reported thelastomatoid species are generalists rather than specialists, that host-specificity within this group is low and that co-evolutionary speciation has had little, if any, impact on structuring the thelastomatoid fauna of Australian burrowing cockroaches. In a broader context, this study provides the first comprehensive examination of the role of coevolutionary speciation and host specificity in regulating the distribution of pinworms in arthropods.