During the last 2 decades there have been numerous reports of the emergence of mefloquine resistance in Southeast Asia and nearly 50% resistance is reported in Thailand. A World Health Organization report (2001) considers mefloquine as an important component of ACT (artesunate+mefloquine) which is the first line of treatment for the control of uncomplicated/multi-drug resistant (MDR) Plasmodium falciparum malaria. In view of the emergence of resistance towards this drug, it is proposed to develop new drug combinations to prolong the protective life of this drug. Prior studies have suggested that mefloquine resistance can be overcome by a variety of agents such as ketoconazole, cyproheptadine, penfluridol, Icajine and NP30. The present investigation reports that clarithromycin (CLTR), a new macrolide, being a potent inhibitor of Cyt. P450 3A4, can exert significant resistance reversal action against mefloquine resistance of plasmodia. Experiments were carried out to find out the curative dose of CLTR against multi-drug resistant P. yoelii nigeriensis. Mefloquine (MFQ) and clarithromycin (CLTR) combinations have been used for the treatment of this MDR parasite. Different dose combinations of these two drugs were given to the infected mice on day 0 (prophylactic) and day 1 with established infection (therapeutic) to see the combined effect of these combinations against the MDR malaria infection. With a dose of 32 mg/kg MFQ and 225 mg/kg CLTR, 100% cure was observed, while in single drug groups, treated with MFQ or CLTR, the cure was zero and 40% respectively. Therapeutically, MFQ and CLTR combinations 32+300 mg/kg doses cleared the established parasitaemia on day 10. Single treatment with MFQ or CLTR showed considerable suppression of parasitaemia on day 14 but neither was curative. Follow-up of therapeutically treated mice showed enhanced anti-malarial action as reflected by their 100% clearance of parasitaemia. The present study reveals that CLTR is a useful antibiotic to be used as companion drug with mefloquine in order to overcome mefloquine resistance in plasmodia.

For Isospora (Protozoa: Eimeriidae) parasites of passerine birds, diurnal periodicity of oocyst output is a well-described phenomenon. From the temporal zone to the tropics, oocyst production is correlated with the light-dark cycle, peaking in the afternoon hours. However, nothing is known about the existence of diurnal periodicity of these parasites in the birds of High Arctic environments, under permanent light during summer. We sampled free-ranging Snow Bunting (Aves: Passeriformes), on Svalbard in summer and tested oocysts output of Isospora plectrophenaxia. Here we show that under the permanent light conditions of Arctic summer in the wild, Isospora plectrophenaxia, a parasite of the Snow Bunting, still keeps the 24-h rhythm of oocyst output with the peak in the post-meridiem hours, despite the absence of diurnal periodicity in host's activity. Our findings prove the ability of avian Isospora to invoke alternative cues for synchronizing the circadian rhythms. Possible cues and adaptive significance of diurnal periodicity of parasite output in High Arctic are discussed. The maintenance of synchronization and timing of the parasite life-cycle stages is under positive selection pressure even in permanent daylight in the Arctic.

Methionine adenosyltransferase (MAT) is an important enzyme for metabolic processes, to the extent that its product, S-adenosylmethionine (AdoMet), plays a key role in trans-methylation, trans-sulphuration and polyamine synthesis. Previous studies have shown that a MAT-overexpressing strain of Leishmania donovani controls AdoMet production, keeping the intracellular AdoMet concentration at levels that are compatible with cell survival. This unexpected result, together with the fact that MAT activity and abundance changed with time in culture, suggests that different regulatory mechanisms acting beyond the post-transcriptional level are controlling this protein. In order to gain an insight into these mechanisms, several experiments were carried out to explain the MAT abundance during promastigote cell growth. Determination of MAT turnover in cycloheximide (CHX)-treated cultures resulted in a surprising 5-fold increase in MAT turnover compared to CHX-untreated cultures. This increase agrees with a stabilization of the MAT protein, whose integrity was maintained during culture. The presence of proteasome inhibitors, namely MG-132, MG-115, epoxomycin and lactacystin in the culture medium prevented MAT degradation in both MAT-overexpressing and ‘mock-transfected’ leishmanial strains. The role of the ubiquitin (Ub) pathway in MAT down-regulation was supported using immunoprecipitation experiments. Immunoprecipitated MAT cross-reacted with anti-Ub antibodies, which provides evidence of a proteasome-mediated down-regulation of the leishmanial MAT abundance.

Histone H1 in Leishmania presents relevant differences compared to higher eukaryote counterparts, such as the lack of a DNA-binding central globular domain. Despite that, it is apparently fully functional since its differential expression levels have been related to changes in chromatin condensation and infectivity, among other features. The localization and the aggregation state of L. braziliensis H1 has been determined by immunolocalization, mass spectrometry, cross-linking and electrophoretic mobility shift assays. Analysis of H1 sequences from the Leishmania Genome Database revealed that our protein is included in a very divergent group of histones H1 that is present only in L. braziliensis. An antibody raised against recombinant L. braziliensis H1 recognized specifically that protein by immunoblot in L. braziliensis extracts, but not in other Leishmania species, a consequence of the sequence divergences observed among Leishmania species. Mass spectrometry analysis and in vitro DNA-binding experiments have also proven that L. braziliensis H1 is monomeric in solution, but oligomerizes upon binding to DNA. Finally, despite the lack of a globular domain, L. braziliensis H1 is able to form complexes with DNA in vitro, with higher affinity for supercoiled compared to linear DNA.

Life-cycle stages of the microsporidia Nosema bombycis, the pathogen causing silkworm pebrine, were separated and purified by an improved method of Percoll-gradient centrifugation. Soluble protein fractions of late sporoblasts (spore precursor cells) and mature spores were analysed by two-dimensional polyacrylamide gel electrophoresis (2D-PAGE). Protein spots were recovered from gels and analysed by mass spectrometry. The most abundant differential protein spot was identified by database search to be a hypothetical spore wall protein. Using immunoelectron microscopy, we demonstrated that HSWP5 is localized to the exospore of mature spores and renamed it as spore wall protein 5 (NbSWP5). Further spore phagocytosis assays indicated that NbSWP5 can protect spores from phagocytic uptake by cultured insect cells. This spore wall protein may function both for structural integrity and in modulating host cell invasion.

Following previous work on the anti-giardial effect of blueberry polyphenols, a range of polyphenol-rich extracts from berries and other fruits was screened for their ability to kill Giardia duodenalis, an intestinal parasite of humans. Polyphenol-rich extracts were prepared from berries using solid-phase extraction and applied to trophozoites of Giardia duodenalis grown in vitro. All berry extracts caused inhibition at 166 μg gallic acid equivalents (GAE)/ml phenol content but extracts from strawberry, arctic bramble, blackberry and cloudberry were as effective as the currently used drug, metronidazole, causing complete trophozoite mortality in vitro. Cloudberry extracts were found to be the most effective causing effectively complete trophozoite mortality at 66 μg GAE/ml. The polyphenol composition of the more effective berry extracts suggested that the presence of ellagitannins could be an important factor. However, the potency of cloudberry could be related to high ellagitannin content but also to the presence of substantial amounts of unconjugated p-coumaric acid and benzoic acid. These in vitro effects occur at concentrations easily achievable in the gut after berry ingestion and we discuss the likelihood that berry extracts could be effective anti-giardial agents in vivo.

The protozoan parasite Toxoplasma gondii is prevalent worldwide and can infect a remarkably wide range of hosts despite felids being the only definitive host. As cats play a major role in transmission to secondary mammalian hosts, the interaction between cats and these hosts should be a major factor determining final prevalence in the secondary host. This study investigates the prevalence of T. gondii in a natural population of Apodemus sylvaticus collected from an area with low cat density (<2·5 cats/km2). A surprisingly high prevalence of 40·78% (95% CI: 34·07%–47·79%) was observed despite this. A comparable level of prevalence was observed in a previously published study using the same approaches where a prevalence of 59% (95% CI: 50·13%–67·87%) was observed in a natural population of Mus domesticus from an area with high cat density (>500 cats/km2). Detection of infected foetuses from pregnant dams in both populations suggests that congenital transmission may enable persistence of infection in the absence of cats. The prevalences of the related parasite, Neospora caninum were found to be low in both populations (A. sylvaticus: 3·39% (95% CI: 0·12%–6·66%); M. domesticus: 3·08% (95% CI: 0·11%–6·05%)). These results suggest that cat density may have a lower than expected effect on final prevalence in these ecosystems.

Transposons of the Mutator superfamily have been widely described in plants, but only recently have metazoan organisms been shown to harbour them. In this work we describe novel Mutator superfamily transposons from the genomes of the human parasites Schistosoma mansoni and S. japonicum, which we name Curupira-1 and Curupira-2. Curupira elements do not have Terminal Inverted Repeats (TIRs) at their extremities and generate Target Site Duplications (TSDs) of 9 base pairs. Curupira-2 transposons code for a conserved transposase and SWIM zinc finger domains, while Curupira-1 elements comprise these same domains plus a WRKY zinc finger. Alignment of transcript sequences from both elements back to the genomes indicates that they are subject to splicing to produce mature transcripts. Phylogenetic analyses indicate that these transposons represent a new lineage of metazoan Mutator-like elements with characteristics that are distinct from the recently described Phantom elements. Description of these novel schistosome transposons provides new insights in the evolution of transposable elements in schistosomes.

Melarsomine hydrochloride can cure Trypanosoma evansi infection in camels at a dose of 0·25 mg/kg, but at that dose relapses occur in cattle. In our study, the efficacy of an intramuscular injection of melarsomine hydrochloride at 0·5 mg/kg was assessed in 3 normal and 3 splenectomized dairy cattle experimentally infected with a stock of T. evansi from Thailand. The animals were monitored for 5 months by haematocrit centrifugation, blood- or cerebrospinal fluid-mouse inoculation, polymerase chain reaction, the card agglutination test (CATT) for T. evansi, and the enzyme-linked immunosorbent assay‑T. evansi. Parasitological and DNA tests became and remained negative just after treatment. By the end of the experiment, CATT was negative and ELISA scores were below or very close to the cut-off value. One of the splenectomized cattle died from anaplasmosis during the experiment, but tested negative for surra. It was concluded that the parasites had been cleared from the cattle, and melarsomine hydrochloride at 0·5 mg/kg can be recommended for treatment against T. evansi infection in dairy cattle in Thailand. Further work is necessary to validate the efficacy of the treatment in the event of confirmed CSF-infection.

We identified 2 novel genes encoding different 2-Cys peroxiredoxins (PRxs), designated CsPRx2 and CsPRx3, in Clonorchis sinensis, which invades the human hepatobiliary tracts. The CsPRx2 gene expression was temporally increased along with the parasite's development and its protein product was detected in almost all parts of adult worms including subtegument, as well as excretory-secretory products. Conversely, CsPRx3 expression was temporally maintained at a basal level and largely restricted within interior parts of various tissues/organs. The recombinant forms of CsPRx proteins exhibited reducing activity against various hydroperoxides in the presence of either thioredoxin or glutathione (GSH) as a reducing equivalent, although they preferred H2O2 and GSH as a catalytic substrate and electron donor, respectively. A steady-state kinetic study demonstrated that the CsPRx proteins followed a saturable, Michaelis-Menten-type equation with the catalytic efficiencies (kcat/Km) ranging from 103 to 104 M−1 s−1, somewhat lower than those for other PRxs studied (104–105 M−1 s−1). The expression patterns and histological distributions specific to CsPRx2 and CsPRx3 might suggest different physiological functions of the antioxidant enzymes in protecting the worms against oxidative damage.

We asked whether the age of a rodent host affects the feeding performance of fleas. We predicted that fleas would perform better on young and old than on adult rodents. To test this prediction, we determined bloodmeal size, rate of digestion and time of survival after a single bloodmeal in Xenopsylla ramesis feeding on Meriones crassus of different ages. Fleas took less blood from subadult and adult than from juvenile and old animals. Fleas digested blood of old hosts at the highest rate and blood of juvenile hosts most slowly. After a bloodmeal, fleas survived the longest if fed on a juvenile host. The effect of host gender on bloodmeal size and survival after a bloodmeal was manifested only in (a) subadult and adult hosts and (b) subadult hosts, respectively. Host gender affected the rate of digestion differently among digestion stages and host age cohorts. We explain the observed patterns in flea performance in terms of changes in the host's immune defences and nutritional quality of blood during the host's individual life. Our observations suggested that a host of a particular age could not be unequivocally predicted to be more or less beneficial for a parasite than a younger or an older host.

Phylogenetic analyses of SSU rDNA sequences have previously revealed the existence of 2 Ichthyobodo species able to infect Atlantic salmon (Salmo salar L.). Ichthyobodo necator sensu stricto (s.s.) is assumed to be a freshwater parasite, while a genetically distinct but undescribed species, Ichthyobodo sp. II sensu Todal et al. (2004) have been detected on Atlantic salmon in both fresh- and seawater. In the present study a morphological description of Ichthyobodo sp. II from the gills of salmon reared in fresh-, brackish- and seawater is presented, using both light- and electron microscopy. Comparative morphometry show that Ichthyobodo sp. II from both freshwater and seawater displays a different cell shape, and is significantly smaller than I. necator s.s. Also, ultrastructural characteristics distinguish these two species, notably differences in the attachment region and the presence of spine-like surface projections in Ichthyobodo sp. II. Based on both unique SSU rDNA sequences and morphological characteristics, we conclude that Ichthyobodo sp. II. represents a novel species for which we propose the name Ichthyobodo salmonis sp. n.

There is a gap in our understanding of the relative and interactive effects of different parasite species on the same host population. Here we examine the effects of the acanthocephalan Acanthocephalus galaxii, an unidentified cyclophyllidean cestode, and the trematodes Coitocaecum parvum and Microphallus sp. on several fitness components of the amphipod Paracalliope fluviatilis, using a combination of infection surveys and both survival and behavioural trials. In addition to significant relationships between specific parasites and measures of amphipod survival, maturity, mating success and behaviour, interactions between parasite species with respect to amphipod photophilia were also significant. While infection by either A. galaxii or C. parvum was associated with increased photophilia, such increases were negated by co-infection with Microphallus sp. We hypothesize that this is due to the more subtle manipulative effect of A. galaxii and C. parvum being impaired by Microphallus sp. We conclude that the low frequency at which such double infections occur in our sampled population means that such interactions are unlikely to be important beyond the scale of the host individual. Whether or not this is generally true, implying that parasitological models and theory based on single parasite species studies do generally hold, requires cross-species meta-analytical studies.

Each transmission event in complex, multi-host life cycles create obstacles selecting for adaptations by trematodes. One such adaptation is life cycle abbreviation through progenesis, in which the trematode precociously matures and reproduces within the second intermediate host. Progenesis eliminates the need for the definitive host and increases the chance of life cycle completion. However, progenetic individuals face egg-dispersal challenges associated with reproducing within metacercarial cysts in the tissues or body cavity of the second intermediate host. Most progenetic species await host death for their eggs to be released into the environment. The present study investigated temporal variation of progenesis in Stegodexamene anguillae in one of its second intermediate fish hosts and the effect of the fish's reproductive cycle on progenesis. The study involved monthly sampling over 13 months at one locality. A greater proportion of individuals became progenetic in the gonads of female fish hosts. Additionally, progenesis of worms in the gonads was correlated with seasonal daylight and temperature changes, major factors controlling fish reproduction. Host spawning events are likely to be an avenue of egg dispersal for this progenetic species, with the adoption of progenesis being conditional on whether or not the parasite can benefit from fish spawning.

The cyclooctadepsipeptide PF1022A derived from the fungus, Mycelia sterilia, is characterized by a broad spectrum of activity against different parasitic gastrointestinal nematodes of livestock. In the present work the anthelmintic activity of PF1022A against Heligmosomoides bakeri, a widely used laboratory model was studied. Albendazole, ivermectin and levamisole served as reference. In vitro, PF1022A showed low activity on embryonation but significantly inhibited egg hatch (10 and 100 μg/ml), whereas albendazole (10 and 100 μg/ml) revealed statistically significant inhibitions of both embryonation and egg hatch. PF1022A (1–100 μg/ml) completely inhibited larval movement at most examination points. Comparable significant anthelmintic activity on the larval stages of H. bakeri was observed with levamisole (48–100%), while slightly lower activities were observed with ivermectin (20–92%) and albendazole (0–87%) at 1–100 μg/ml. PF1022A and levamisole significantly inhibited motility and egg release of adult worms, while albendazole and ivermectin failed to demonstrate activity. Significant worm burden reductions were achieved with PF1022A, levamisole and ivermectin in vivo. For example, at 0·125 mg/kg PF1022A a worm burden reduction of 91·8% was observed. The use of drug combinations did not further enhance the in vitro and in vivo activity of PF1022A. In conclusion, further investigations are warranted with PF1022A, as the drug is characterized by significant larvicidal and nematocidal activity in vitro and in vivo.