In order to elucidate mechanisms underlying snail/echinostome compatibility, numerous molecular studies comparing transcripts and proteins of Biomphalaria glabrata susceptible or resistant to Echinostoma caproni were undertaken. These studies focused on plasma and haemocytes of the two strains and revealed that some transcripts and/or proteins were differentially expressed between strains. The aim of the present study was to develop a complementary transcriptomic approach by constructing subtractive libraries. This work revealed some candidate transcripts already identified in previous studies (calcium-binding proteins and glycolytic enzymes) as well as novel candidate transcripts that were differentially represented between strains of B. glabrata. Among these newly identified genes, we revealed several genes potentially involved in immune processes encoding proteases, protease inhibitors, a lectin, an aplysianin-like molecule, and cell adhesion molecules.

Polysporoplasma sparis infection was studied in gilthead sea bream from different mariculture systems of the Spanish coasts. Culture conditions influenced the infection dynamics, as the parasite appeared only in semi-intensive cultures and was not found in intensive closed systems nor in open ones. No clear seasonal pattern was observed. No fish weighing less than 51 g was found parasitized in any group. A statistically significant dependence between infection prevalence and host weight was observed in some growing stocks. Light and transmission electron microscope observations revealed serious damage in the trunk kidney. Glomerular disease was provoked by the progressive occupation of the glomerular capillaries by P. sparis spores. Tubular epithelial cells were also affected. Inflammatory responses appeared towards the end of the infection, and consisted mainly of melanomacrophages and eosinophils. Rodlet cells were common close to infected capillaries and debris of rodlet sacs formed a belt encircling capillary vessels. Cytochemistry demonstrated the lipidic nature of these sacs and the glycogen and glycoprotein composition of the cytoplasmic granules of rodlet cells.

Many hermaphroditic parasites reproduce by both cross-fertilization and self-fertilization. To understand the maintenance of such mixed mating systems it is necessary to compare the fitness consequences of the two reproductive modes. This has, however, almost never been done in the context of host–parasite coevolution. Here we show the consequences of outcrossing and selfing in an advanced life-stage of the cestode Schistocephalus solidus, i.e. in its second intermediate host, the three-spined stickleback (Gasterosteus aculeatus). Each juvenile stickleback was simultaneously exposed to 2 experimentally infected copepods, one harbouring outcrossed the other selfed parasites. At 60 days p.i. parasites were removed from the fish's body cavity and, with microsatellite markers, assigned to either outcrossed or selfed origin. Prevalence was not significantly higher in outcrossed parasites. However, those fish that were infected contained significantly more outcrossed than selfed parasites. Thus the probability of a selfed parasite to progress in the life-cycle is reduced in the second intermediate host. Furthermore, we found that even the multiply infected fish increased in weight during the experiment. Nevertheless, total worm weight in multiply infected fish was significantly lower than in singly infected ones, which thus might be a parasite life-history strategy.

The ability of certain species of parasitic nematodes to survive desiccation for considerable periods is a fascinating example of adaptation to the demands of fluctuating environments that occasionally can become extreme and life threatening. Behavioural and morphological adaptations associated with desiccation survival serve primarily to reduce the rate of drying, either to prolong the time taken for the nematode's water content to reach lethal low levels or, in true anhydrobiotes, to enable the structural and biochemical changes required for long-term survival to take place. Examples of these adaptations are reviewed, together with information on the factors involved in rehydration that ensure successful exit from the dormant state. Information on desiccation survival is central to effective management and control options for parasitic nematodes. It is also required to assess the feasibility of enhancing the longevity of commercial formulations of entomopathogenic nematodes, both before and after application; current research and future prospects for enhancing survival of these bio-insecticides are discussed.

Varroa destructor is considered the most damaging parasite affecting honeybees (Apis mellifera L.). However, some honeybee populations such as the savannah honeybee (Apis mellifera scutellata) can survive mite infestation without treatment. It is unclear if survival is due to resistance mechanisms decreasing parasite reproduction or to tolerance mechanisms decreasing the detrimental effects of mites on the host. This study investigates both aspects by quantifying the reproductive output of V. destructor and its physiological costs at the individual host level. Costs measured were not consistently lower when compared with susceptible honeybee populations, indicating a lack of tolerance. In contrast, reproduction of V. destructor mites was distinctly lower than in susceptible populations. There was higher proportion of infertile individuals and the reproductive success of fertile mites was lower than measured to date, even in surviving populations. Our results suggest that survival of savannah honeybees is based on resistance rather than tolerance to this parasite. We identified traits that may be useful for breeding programmes aimed at increasing the survival of susceptible populations. African honeybees may have benefited from a lack of human interference, allowing natural selection to shape a population of honeybees that is more resistant to Varroa mite infestation.

Whole blood collected from koalas admitted to the Australian Zoo Wildlife Hospital (AZWH), Beerwah, QLd, Australia, during late 2006–2009 was tested using trypanosome species-specific 18S rDNA PCRs designed to amplify DNA from Trypanosoma irwini, T. gilletti and T. copemani. Clinical records for each koala sampled were reviewed and age, sex, blood packed cell volume (PCV), body condition, signs of illness, blood loss, trauma, chlamydiosis, bone marrow disease, koala AIDS and hospital admission outcome (‘survival’ / ‘non-survival’) were correlated with PCR results. Overall 73·8% (439/595) of the koalas were infected with at least 1 species of trypanosome. Trypanosoma irwini was detected in 423/595 (71·1%), T. gilletti in 128/595 (21·5%) and T. copemani in 26/595 (4·4%) of koalas. Mixed infections were detected in 125/595 (21%) with co-infections of T. irwini and T. gilletti (101/595, 17%) being most common. There was a statistical association between infection with T. gilletti with lower PCV values and body condition scores in koalas with signs of chlamydiosis, bone marrow disease or koala AIDS. No association between T. gilletti infection and any indicator of health was observed in koalas without signs of concurrent disease. This raises the possibility that T. gilletti may be potentiating other disease syndromes affecting koalas.

Periodontal diseases (gingivitis and periodontitis), result from a disruption of the host–oral microbiome homoeostasis. Whereas the pathological role of some specific bacterial strains during periodontal diseases is well documented, the impact of parasites in periodontium pathophysiology is still under debate. This review aims to collect data about the prevalence and the potential role of Trichomonas tenax during periodontal diseases. Data from 47 studies revealed that T. tenax prevalence in diseased periodontium ranged from 0 to 94·1%. The prevalence of oral protozoan infections was found to be largely greater in patients with periodontal diseases than with healthy periodontium. The parasite detection was mainly performed by direct microscopy. Trichomonas tenax presence was clearly correlated with periodontal disease. The high heterogeneity of its periodontal prevalence may be correlated with the diversity of the population screened (age, sex, systemic diseases), and the methods used for diagnosis. This protozoan seems to have the capacity to be involved in the inflammatory process of gum disease. Animal experimentation, using relevant physiopathological models of periodontitis, needs to be performed to investigate the ability of T. tenax to cause and/or worsen the disease. Further investigations using standardized experimental designs of epidemiologic studies are also needed.

We addressed the question whether diaplacental transmission of Neospora caninum can be controlled by metaphylactic chemotherapy using toltrazuril or enrofloxacin. Female C57/BL6 mice, infected on day 10 of pregnancy, were medicated for 6 consecutive days p.i. with 52·5 mg toltrazuril or – as an out-group control medication – 16·7 mg enrofloxacin per kg body weight per day. Other control groups received either infection but no medication or vice versa. Toltrazuril treatment significantly reduced pre- and perinatal losses (10 deliveries of healthy newborns, versus 1 abortion and 4 failures) when compared to control-enrofloxacin (2 deliveries, versus 1 abortion, 7 failures and 2 pre-parturient deaths of dams) and non-treated animals (3 deliveries, versus 6 abortions, 8 failures and 4 pre-parturient deaths). Simultaneously, PCR-based parasite detection in the brain of mothers, histopathological findings as well as clinical fatality were significantly less frequent in toltrazuril-treated dams. The overall toltrazuril treatment efficacy was determined as 87%, that of enrofloxacin-treatment as 17%. The progenies of toltrazuril-treated dams also exhibited a very low rate of PCR-positivity in their brain (3 out of 39), whereas untreated dams delivered litters with mostly PCR-positive brains (12 out of 14) and a relatively high death rate post-partum (5 out of 19 newborns died). Mice subjected to a second mating delivered newborns all negative by N. caninum-PCR, indicating that diaplacental tachyzoite passage does not occur in a later, repeated pregnancy. Overall, our experiments showed that toltrazuril-treatment of an acute N. caninum-infection – induced during pregnancy – results in a clear reduction of fetal losses and a marked reduction of diaplacental passage of the parasite to the fetal brain, whereas enrofloxacin, as an out-group control substance, failed to show the same effect.

Promastigotes of Leishmania braziliensis panamensis absorbed the purines adenine, hypoxanthine, adenosine and inosine by a combination of diffusion and mediated components. When the uptake rates for these substrates were corrected for diffusion and compared, the purine bases adenine and hypoxanthine were transported at a significantly slower rate than the purine nucleosides adenosine and inosine. Competitive interactions among those purines tested confirmed the presence of mediated and diffusion components and suggested that three transport loci may be operating (Fig. 6). The first transport locus, designated Locus 1, transported inosine, Locus 2, the purine bases hypoxanthine and adenine and Locus 3, adenosine. In addition, adenine and hypoxanthine inhibited the uptake of one another competitively. A comparison of Ki values derived from double reciprocal plots of labelled hypoxanthine and adenine uptake in the presence of the unlabelled substrates as inhibitors suggested that adenine has a greater affinity for the transport locus.

The central (CNS) and peripheral (PNS) nervous systems of the cyclophyllidean tapeworm, Moniezia expansa, were examined for the presence of cholinergic, serotoninergic and peptidergic elements using enzyme cytochemical and immunocytochemical techniques in conjunction with light and confocal scanning laser microscopy. Cholinesterase activity and 5-hydroxytryptamine- and regulatory peptide-immunoreactivities (IRs) were localized to the nerve fibres and cell bodies of all of the major neuronal components in the CNS of the worm, including the cerebral ganglia and connecting commissure, the 10 longitudinal nerve cords and associated transverse ring commissures. Although each of the 3 systems appeared well developed and comprised a significant portion of the nervous system, the serotoninergic constituent was the most highly developed, consisting of a vast array of nerve fibres and cell bodies distributed throughout the strobila of the worm. A close association of cholinesterase reactivity and peptide-IRs was evident throughout the CNS, indicating the possible co-localization of acetylcholine and neuropeptides. Within the PNS, cholinergic activity and serotoninergic- and peptidergic-IRs occurred in the subtegumental network of nerve fibres and somatic musculature. Although all 3 neuro-chemical elements were present in the acetabula, they were found in different nerve fibres; only cholinergic and peptidergic cell bodies were found. The common genital opening, vagina and ootype regions of the reproductive system displayed a rich innervation of all 3 types of neuronal populations. Within the peptidergic system, immunostaining with antiseraraised to the C-terminus of the neuropeptide Y superfamily of peptides and the invertebrate peptides, neuropeptide F (M. expansa) and FMRFamide was the most prevalent. Limited positive-IR for substance P and neurokinin A were also recorded in the CNS of the worm.

The ultrastructure of immature exo-erythrocytic forms of Plasmodium berghei in rat hepatocytes was studied at stages between 25 and 51 h of development. A new method was successfully applied to localize the parasites in a small portion of the liver by temporary ligature of blood vessels to the majority of the liver, and from the spleen and the pancreas. Nuclear profiles appeared to be part of a highly lobed nuclear reticulum. Peripheral vesiculation and vacuolization of the cytoplasm was increasingly prominent and ushered in the formation of pseudocytomeres. Spacing between host- and parasite-derived membranes could first be observed after 43 h of development. In general, organellar development was found to follow closely that described for plasmodial oocysts.

The surface antigens of Trypanosoma cruzi trypomastigotes were identified by immunoprecipitation and were compared with metabolically labelled excretory—secretory products (ES) released by the parasites in vitro. A series of major immunogenic components in the ES antigens were revealed (160 kDa, 130 kDa and 80–110 kDa). The trypomastigote surface also bears the 130 kDa band and the 80–110 kDa complex. Competition experiments demonstrated the common antigenic structure of the ES and the surface antigens. Two-dimensional analysis of ES antigens immunoprecipitated by human Chagasic serum revealed several spots in the 80–110 kDa region with a wide range of isoelectric points (PI between 5·4 and 6·7). This reflects a charge heterogeneity of these polypeptides. The trypomastigote 85 kDa polypeptide was also identified in the ES antigens by using a monoclonal antibody against this antigen. Two-dimensional analysis of the 85 kDa proteins shed from the surface of trypomastigotes and immunoprecipitated by the monoclonal antibody 155D3 showed 2 major spots: a major part of the 85 kDa polypeptide was found at pH 6·5–6·6, whereas a substantial amount of the antigen was found at pH 5·7. An additional component with molecular weight of approximately 58 kDa and isoelectric points of 6·5 and 6·6, was also visualized. Detection of the 85 kDa polypeptide circulating in serum from patients with acute and chronic Chagas' disease was achieved using an enzyme-linked immunosorbent assay. In addition, the data obtained showed that a polyclonal antibody to the 85 kDa polypeptide could be used to passively induce a partial protection of Fischer rats against acute lethal infection. Thus, the antigens recognized by polyclonal antibody appear to play a role in the development of protective immunity against T. cruzi.

In earlier studies on the supposedly anaerobic metabolism of Leishmania promastigotes it was suggested that the reduction of fumarate to succinate functions as the main electron sink during anoxia. Interestingly, however, our preliminary results demonstrated that rhodoquinone, an essential component for efficient fumarate reduction in eukaryotes, was absent in L. infantum promastigotes. Therefore, we re-investigated the energy metabolism and succinate production of these promastigotes. Our studies demonstrated that L. infantum promastigotes could, to a certain extent, survive periods without respiration but had a low capacity for anaerobic metabolism. When oxygen could not be used as terminal electron acceptor, the degradation of glucose was severely inhibited, forcing the parasite to reduce its energy expenditure, which resulted in inhibited motility and proliferation. In addition, we studied the mechanism of succinate production under aerobic conditions and showed that in L. infantum promastigotes this succinate was mainly produced via an oxidative pathway, the Krebs cycle, and not significantly via fumarate reduction, which correlated with the absence of rhodoquinone. Taken collectively our studies show that L. infantum promastigotes depend mainly on respiratory chain activity for energy generation, have a poor capacity for anaerobic functioning, and go into metabolic arrest during anoxic conditions.

A novel 28 kDa cysteine protease (Cs28CF) secreted by the hepatobiliary trematode, Clonorchis sinensis was identified. The protease was purified from the excretory-secretory products (ESP) of the adult worm using DEAE-ion exchange and Arginine-Sepharose 4B chromatography. It showed a high activity between pH 6·5 and 7·5 in a dithiothreitol (DTT)-dependent manner. Inhibitors specific to cysteine proteases down-regulated the activity. Addition of Cs28CF to monkey cholangiocyte cultures resulted in approximately 95% cell death after 7 days. The full-length cDNA (1078 bp) encoded a single peptide of 328 amino acids (aa) with an N-terminal hydrophobic sequence, an ERFNAQ motif in the propeptide and a mature domain. Expression of mRNA transcripts of Cs28CF was observed in both the metacercaria and adult stages. Bacterially expressed recombinant protein exhibited a specific antibody reaction with clonorchiasis sera. Deduced aa exhibited 52–76% sequence identity with the cathepsin F analogues from other organisms. A novel E/DXGTA motif was recognized in the propeptide region. Phylogenetic analysis of 63 papain family members revealed that the trematode cysteine proteases formed 2 major clades of cathepsins F and L. The trematode cysteine proteases classified as cathepsin F shared higher homology among themselves than those classified as cathepsin L. Cathepsin F is phylogenetically conserved in the trematode parasites as well as in mammals.

The spore wall of Nosema bombycis plays an important role in microsporidian pathogenesis. Protein fractions from germinated spore coats were analysed by two-dimensional polyacrylamide gel electrophoresis and MALDI-TOF/TOF mass spectrometry. Three protein spots were identified as the hypothetical spore wall protein NbHSWP12. A BAR-2 domain (e-value: 1.35e-03) was identified in the protein, and an N-terminal protein-heparin interaction motif, a potential N-glycosylation site, and 16 phosphorylation sites primarily activated by protein kinase C were also predicted. The sequence analysis suggested that Nbhswp12 and its homologous genes are widely distributed among microsporidia. Additionally, Nbhswp12 gene homologues share similar sequence features. An indirect immunofluorescence analysis showed that NbHSWP12 localized to the spore wall, and thus we renamed it spore wall protein 12 (NbSWP12). Moreover, NbSWP12 could adhere to deproteinized N. bombycis chitin coats that were obtained by hot alkaline treatment. This novel N. bombycis spore wall protein may function in a structural capacity to facilitate microsporidial spore maintenance.

Schistosome cercariae of the genus Trichobilharzia are the causative agent of swimmers' itch. In order to characterize the changes in parasites during and after the penetration of the host skin, in vitro and in vivo (in ducks and mice) transformations of T. szidati cercariae to schistosomula were performed. Ultrastructural observation revealed that cercariae possess a simple outer tegumental membrane with a thick glycocalyx. As with human schistosomes, the latter structure disappears during transformation and a new double membrane with putative protective function is formed. Our biochemical and immunological observations showed that the carbohydrate-rich glycocalyx of cercariae is readily bound by lectins and antibodies. The in vitro transformation to schistosomula can be detected by enhanced reactivity of 2 lectin probes (PNA and ConA) with the surface. The in vivo-transformed (skin and lung) schistosomula appear to have few surface ligands for the 12 lectin probes being tested. Similarly, the cercarial surface and its remnants on the in vitro-produced schistosomula is recognized by sera from immunized mice and humans with cercarial dermatitis; the tissue schistosomula fail to react with these antibodies. The loss of surface targets as a part of parasite immune evasion within the host is discussed.

IgA and IgE activity against Teladorsagia circumcincta was investigated in a flock of Texel lambs following natural, mixed nematode infection among lambs. The distribution of IgA activity was similar to a gamma distribution whereas IgE activity was different. Box-Cox analysis demonstrated that X0·25 was a suitable transformation to normalise IgE responses. The transformed IgE activity was under moderate to strong genetic control. Nine different allergens were identified by proteomic analysis. Tropomyosin was selected for further analysis. IgE activity against tropomyosin was moderately heritable and associated with decreased egg counts and with reduced body weight at the time of sampling.

Stochastic processes play a vital role in the early stages of the evolution of drug-resistant malaria. We present a simple and flexible method for investigating these processes and understanding how they affect the emergence of drug-resistant malaria. Qualitatively different predictions can be made depending on the biological and epidemiological factors which prevail in the field. Intense intra-host competition between co-infecting clones, low numbers of genes required to encode resistance, and high drug usage all encourage the emergence of drug resistance. Drug-resistant forms present at the time drug application starts are less likely to survive than those which arise subsequently; survival of the former largely depends on how rapidly malaria population size stabilizes after drug application. In particular, whether resistance is more likely to emerge in areas of high or low transmission depends on malaria intra-host dynamics, the level of drug usage, the population regulation of malaria, and the number of genes required to encode resistance. These factors are discussed in relation to the practical implementation of drug control programmes.

Ascaris is a large parasitic roundworm (nematode) of the small intestine of humans and pigs, which causes the socio-economically important disease, ascariasis. To better understand the relationship of Ascaris between the 2 host species, recent studies in China have focused on investigating the genetics and epidemiology of Ascaris from humans and pigs using a mutation scanning-based approach. Findings provided support for a low level of gene flow between the human and porcine Ascaris populations. Extending the studies of genotypic variability within Ascaris from humans and pigs, experimental infections of mice and pigs with selected genotypes of Ascaris were carried out. Initial results indicate that there is a significant difference in the ability of Ascaris eggs of genotype G1 (derived from human) and G3 (derived from pig) to infect and establish as adults in pigs, supporting the difference in the frequencies of these genotypes in natural Ascaris populations between pigs and humans in China. Taken together, current information supports that there is limited cross-infection of Ascaris between humans and pigs in endemic regions and that pigs are not a significant reservoir of human infection with the adult nematode in such areas.