The free-living amoeba Naegleria fowleri is the aetiological agent of primary amoebic meningoencephalitis (PAM), a disease leading to death in the vast majority of cases. In patients suffering from PAM, and in corresponding animal models, the brain undergoes a massive inflammatory response, followed by haemorrhage and severe tissue necrosis. Both, in vivo and in vitro models are currently being used to study PAM infection. However, animal models may pose ethical issues, are dependent upon availability of specific infrastructural facilities, and are time-consuming and costly. Conversely, cell cultures lack the complex organ-specific morphology found in vivo, and thus, findings obtained in vitro do not necessarily reflect the situation in vivo. The present study reports infection of organotypic slice cultures from rat brain with N. fowleri and compares the findings in this culture system with in vivo infection in a rat model of PAM, that proved complementary to that of mice. We found that brain morphology, as present in vivo, is well retained in organotypic slice cultures, and that infection time-course including tissue damage parallels the observations in vivo in the rat. Therefore, organotypic slice cultures from rat brain offer a new in vitro approach to study N. fowleri infection in the context of PAM.

The authors of the above article regret that there are errors in the names of two of the authors on the title page. The authors' names should read:

The humoral innate immune response of gilthead seabream (Sparus aurata L.) against the myxozoan Enteromyxum leei has been studied. At 10, 22, 38, 52 and 108 days of cohabitation fish were sampled to examine gut histology and to determine serum innate immune parameters and the mRNA expression of pro-inflammatory cytokines (IL-1β and TNFα) in head-kidney. The parasite was successfully transmitted to 45% of the recipient fish and prevalence reached a maximum (62·5%) at the last sampling time (108 days). Recipient fish started to die after 74 days of cohabitation. In general, alternative complement activity was higher whereas the peroxidase level was lower in recipient fish than in controls. Moreover, IL-1β mRNA expression increased while the TNFα gene expression decreased in recipient fish. These data demonstrate the involvement of complement activity in the defence mechanisms of the gilthead seabream against the myxosporean E. leei. Within the recipient fish group, few differences were observed in the studied immune parameters between E. leei-parasitized and non-parasitized recipient fish. Parasitological and immunological implications of E. leei infections in Mediterranean fish farms are discussed.

This paper reviews the immunomodulatory effects (immunosuppression or immunoactivation) of various anthelmintics including levamisole, fenvalerate, dieldrin, carbofuran, aminocarb, thiabendazole, fenbendazole, oxfendazole and ivermectin. The induced modulation of immune function may occur via direct and/or indirect mechanisms. The immunomodulatory effects of these anti-parasitics have been studied in a variety of bacterial (e.g. brucellosis, salmonellosis, paratuberculosis, mastitis), viral (e.g. infectious bovine rhinotracheitis, Herpes, foot and mouth disease), parasitic (e.g. onchocerciasis, coccidiosis, ascariasis, schistosomiasis) and neoplastic diseases. Some antiparasitics have also been used to boost immunity in a number of human diseases including leprosy, Hodgkin's disease, rheumatoid arthritis, and in adjuvanted therapy of colorectal cancer. The ability to stimulate the immune response of animals offers a new means of disease intervention. Future research on immunomodulatory effects of anti-parasitics, for humans and domestic farm animals, will provide additional methods of treating immunosuppressed subjects. The immunopotentiating or immunosuppressing activity of anti-parasitics will dictate whether co-administration of vaccines and anthelmintics or administration of vaccines during the window of immunoactivation is justified or not.

Host manipulation by trophically transmitted parasites may predispose infected hosts to predation and in this way enhance parasite transmission. In most study systems, however, the evidence comes from laboratory studies, and therefore knowledge of the effect of manipulation on parasite transmission efficiency in the wild is still limited. Here we examined the effect of Diplostomum spathaceum (Trematoda) eye flukes on the susceptibility of fish intermediate hosts to predation by bird definitive hosts. Our earlier studies have shown that the parasite alters fish phenotype and increases their susceptibility to artificial predation under laboratory conditions. In the present field study, we allowed wild birds to feed on fish from cages placed into a lake, and found that predation vulnerability of infected fish did not differ from that of controls. However, we suggest that the experimental set-up likely affected the result because the cages allowed gulls, which caught most of the fish in the study, to feed on fish in an easy, unnatural manner by standing on the edges of the cages. Nevertheless, considerable predation was observed, which provides important initial evidence of how this question should be addressed in the wild.

The D1-D2 domains of LSU rDNA were used to reconstruct the phylogenetic relationships within the Ancyrocephalinae (Monogenea: Dactylogyridae) utilizing maximum-parsimony (MP), maximum-likelihood (ML), minimum evolution (ME) and neighbour-joining (NJ) methods. A total of 32 monogenean taxa were examined in the present study, including 9 Haliotrema species and 13 other species representing the Ancyrocephalinae, 4 Thaparocleidus species representing the Ancylodiscoididae, and 6 species representing the Diplectanidae which were used as multiple outgroups. All 4 analyses (i.e. MP, ML, ME and NJ) inferred the same interrelationship pattern: (Diplectanidae, (Ancylodiscoididae, Dactylogyridae)) with high bootstrap support. However, 9 Haliotrema species were dispersed to form 4 clades together with species from other genera, indicating the apparent non-monophyly of Haliotrema. Three major groups were defined based on reconstructed phylogenetic trees to explain the radiation of Haliotrema species. The morphology of the reproductive organ, particularly the male copulatory organ (MCO), was discussed to further understand the formation of each group. (1) Results of the present study indicated an intimate relationship among Metahaliotrema (2 species), Protogyrodactylus (4 species) and Haliotrema (2 of 9 species), and notably, all these species share vagina-absence. (2) Based on the present molecular analyses and the morphological characters of the MCO, we propose to transfer H. spirotubiforum and the undetermined Haliotrema sp. ZHDDb to Euryhaliotrema as new combinations. (3) We propose to erect a new genus to accommodate the Haliotrema species with horn-like shaped MCO. Taxonomic implications of the present molecular phylogenetic analyses are discussed. A wider range of taxa and more DNA markers displaying various evolutionary rates should be used to estimate phylogenetic relationships among species within the Ancyrocephalinae and Ancylodiscoididae in further studies.

Human death has been recognised as a significant personal and social event for many thousands of years, and classical archaeologists have revealed the changing complexity of rituals associated with it. The study of cell death, however, is a much more recent event, although many of the molecular pathways involved have now been identified, at least in mammalian systems. In studying the loss of cells, the use of the term ‘death’ is, perhaps, not altogether appropriate both since it carries the cultural resonance associated with bodily death, and because we do not study cell death itself, but rather the processes that lead up to it. Mammalian cell death processes are complex and involve a dynamic equilibrium between death promoting and death inhibiting factors, suggesting that some components of death pathways may have a paradoxical survival function. Since parasites must survive an often hostile environment, they may be a useful model to study whether component molecules of mammalian death pathways originally formed modules of parasite survival strategies, and whether survival and death pathways coevolved.

Toxoplasma gondii is an obligate intracellular protozoan parasite. The objective of the present study was to examine the behaviour of Toxoplasma gondii RH Ankara strain tachyzoites in a cell culture environment. The study represents the first step in determining whether T. gondii RH Ankara strain tachyzoites, grown in cell culture, are of sufficient quality to allow cessation of in vivo tachyzoite production for diagnostic assays. In the present study, T. gondii RH Ankara strain tachyzoites were continuously produced in myeloma X63.Ag8.653, HeLa, Hep-2, and Vero cell cultures for 2 months. The average size of the tachyzoites was 3×5·7 μm prior to the first inoculation but after continuous production, a marked decrease was noted in average tachyzoite size. The smallest tachyzoite size, was 1×2·1 μm after 2 months, in myeloma cell cultures even though the yield of tachyzoites increased. With other cell cultures, tachyzoite yields were not as high as myeloma cell culture although decrease in size was less. The smallest decrease in tachyzoite size, averaging 2×3·8 μm after 2 months, was observed in tachyzoites produced in HeLa cell cultures. A virulence assay in small groups of BALB/c mice, using tachyzoites derived from cell cultures, was also conducted. The preliminary results of the virulence assay suggest that as the size of the tachyzoites decreased, the virulence in mice decreased. Future research will focus on the effect of the size of cell culture-derived T. gondii RH Ankara strain tachyzoites on the virulence, protein expression, and the reliability of diagnostic assays. Ultimately, the behaviour of tachyzoites from various T. gondii strains will be observed in cell culture to determine if size is altered.

We have examined metabolic rate, lipid and carbohydrate of female Aedes aegypti during 10 days following a malaria-infected bloodmeal. In parallel, we determined bloodmeal size, portions retained and diuresed, and subsequent fecundity. We found that mosquitoes obtained identical masses of blood when feeding on an infected or control host. However, infected mosquitoes lost more mass during diuresis and retained a smaller mass. Infection led to a significant reduction in fecundity, the extent of which could not be explained by the difference in post-diuresis bloodmeal mass alone. We found no differences in lipid or carbohydrate content between infected and control mosquitoes during the 10 days post-infection, although infected mosquitoes had a lower body mass than controls. Metabolic rates were not different between groups, except during blood digestion, where the metabolic rate was lower in infected mosquitoes. These results suggest that infection by malaria does not lead to an increase in metabolic rate during the phases of midgut invasion and sporogony. However, infection does have a measurable effect on fecundity and subsequent body mass of the infected females.

Two experiments (Exps 1 and 2) were carried out to study the effect of dietary carbohydrates on the establishment of Trichuris suis in pigs. Two experimental diets based on barley flour were used; Diet 1 was supplemented with non-fermentable carbohydrates from oat hull meal, while Diet 2 was supplemented with fermentable carbohydrates from sugar beet fibre and inulin. In Exp. 1, thirty-two pigs were allocated randomly into 4 groups. Two groups were fed Diet 1 and 2 groups were fed Diet 2. Pigs from one of each diet group were inoculated with 2000 infective T. suis eggs each and the other two groups were uninfected controls. All pigs were slaughtered 8 weeks post-inoculation (p.i.). In Exp. 2, twenty-four pigs were allocated randomly into 2 groups and fed Diet 1 or Diet 2, respectively. All the pigs were inoculated with 2000 infective T. suis eggs. Six pigs from each group were slaughtered 8 weeks p.i. and the remaining 6 pigs from each group were slaughtered 12 weeks p.i. Infections were followed by faecal egg counts and worm burdens were assessed at necropsy. Pigs fed Diet 2 had lower egg counts in both experiments; in Exp. 2 the difference was significant (P<0·05). No differences were found in worm burdens 8 weeks p.i. in both experiments, however, worms from pigs on Diet 2 were significantly shorter (P<0·0001). Pigs fed Diet 2 and slaughtered 12 weeks p.i. had significantly lower worm counts (P<0·01) compared to pigs fed Diet 1. The results indicate that fermentable carbohydrates do not affect the establishment of T. suis in naïve pigs, but result in earlier expulsion and reduced growth of the established worms. Thus, diets with highly fermentable carbohydrates may be used in the control of T. suis.

There is unequivocal evidence that parasites influence the immune activity of their hosts, and many of the classical examples of this are drawn from assessment of helminth infections of their mammalian hosts. Thus, helminth infections can impact on the induction or course of other diseases that the host might be subjected to. Epidemiological studies demonstrate that world regions with high rates of helminth infections consistently have reduced incidences of autoimmune and other allergic/inflammatory-type conditions. Here I review and assess the possible ways by which helminth infections can block or modulate concomitant disease processes. There is much to be learned from careful analysis of immuno-regulation in helminth-infected rodents and from an understanding of the immune status of acutely and chronically infected humans. The ultimate reward from this type of investigation will likely be a more comprehensive knowledge of immunity, novel ways to intervene in the immune response to alleviate autoimmune and allergic diseases (growing concerns in economically developed areas), and perhaps the development of helminth therapy for patients suffering from specific inflammatory, autoimmune or allergic disorders.

We have examined Plasmodium falciparum gametocyte prevalence, density and their genetic complexity among children of 2 sympatric ethnic groups (Mossi and Fulani) in villages in Burkina Faso. The 2 groups are known to have distinct differences in their susceptibility and immune responses to malaria. We used RT-PCR and sequence-specific probes to detect and type RNA of the gametocyte-specific protein Pfs48/45. There were no differences in detection rates of asexual forms and gametocytes among the 2 groups, using PCR and RT-PCR, respectively. However, there were significant differences in densities of asexual forms and gametocytes, which were both higher among Mossi than Fulani. Both asexual forms and gametocyte densities were influenced by age and ethnicity. Multiple-clone infections with more than 1 gametocyte genotype were equally prevalent among Fulani and Mossi. These differences can most probably be attributed to genetic differences in malaria susceptibility in the 2 ethnic groups.

Human Oesophagostomum infections are locally common in northern Ghana. The present study describes the results of a cross-sectional survey involving 1011 subjects, selected by a compound-based random sampling method from 1227 compounds in 24 villages. Selected persons were examined by both Kato and coproculture methods. Hookworm-like eggs, representing ova of Oesophagostomum bifurcum and hookworm were detected in 87·5% of the Kato smears. The geometric mean egg count of the infected subjects was 1018. Upon coproculture, third-stage larvae of O. bifurcum and hookworm were detected in 53·0% and 86·9% of subjects respectively. Oesophagostomum infections were clustered but no clear explanation for aggregation of infections could be found as yet. Subjects infected with hookworm had a 5-fold higher risk of being infected with O. bifurcum. Infection rates in adult women were higher than in adult men. No association was found with family size, level of hygiene or with the presence of animals in the compounds. Representatives of the Bimoba-tribe were significantly more infected than those of the other tribes. It appears, however, that this tribal association is a geographical phenomenon: Bimoba are mostly living in villages with the highest infection rates.

A succession of antimalarial drugs has been deployed to treat human falciparum malaria but each has, in turn, been nullified by the spread of drug resistance. The consensus view has always been that, once present, resistance will inevitably rapidly increase to 100%. However, recent field evidence has shown this is not inevitable, and that drug resistance may initially spread and then stabilize at relatively low frequencies. It is proposed that intense competition between separate malaria clones co-infecting the same human can generate complex dynamics capable of explaining this observation. Standard population genetic analysis confirms this assertion. The dynamics underlying the evolution of antimalarial resistance may therefore be much more complex than previously realized, and can resolve the apparent paradox between field data and the underlying theory of the evolution of resistance. This explanation is novel and the results are equally applicable to other parasitic species where multiple infections of the same host are common.

The longevity of filarial nematodes is dependent on secreted immunomodulatory products. Previous investigation of one such product, ES-62, has suggested a critical role for post-translationally attached phosphorylcholine (PC) moieties. In order to further investigate this, ES-62 lacking PC was produced, using the Pichia pastoris recombinant gene expression system. Unlike parasite-derived ES-62, which is tetrameric the recombinant material was found to consist of a mixture of apparently stable tetramers, dimers and monomers. Nevertheless, the recombinant protein was considered to be an adequate PC-free ES-62 as it was recognized by existing antisera against the parasite-derived protein. However, subsequent to this, recognition of parasite-derived ES-62 by antibodies produced against the recombinant protein was found to be absent. In an attempt to explain this, recombinant ES-62 was subjected to structural analysis and was found to (i) contain 3 changes in amino acid composition; (ii) demonstrate significant alterations in glycosylation; (iii) show major differences in protein secondary structure. The effects of these alterations in relation to the observed change in immunogenicity were investigated and are discussed. The data presented clearly show that recognition by existing antibodies is insufficient proof that recombinant proteins can be used to mimic parasite-derived material in studies on nematode immunology and vaccination.

The nodule worm Oesophagostomum bifurcum (Nematoda: Strongylida) is a parasite of major human health importance predominantly in northern Togo and Ghana. Currently, it is estimated that 0·25 million people are infected with this nematode, and at least 1 million people are at risk of infection. Infection with this parasite causes significant disease as a consequence of encysted larvae in the wall of the large intestine. In spite of the health problems caused by O. bifurcum, there have been significant gaps in the knowledge of the biology, transmission and population genetics of the parasite. This review provides an account of some recent insights into the epidemiology and genetics of the parasite from human and non-human primate hosts in specific regions of Africa using molecular tools. Recent research findings are discussed mainly in relation to non-human primates being reservoirs of infection, and the consequences for the prevention and control of oesophagostomiasis in humans are briefly discussed.

Damage to the surface membrane of adult Schistosoma mansoni, and the activity of the excretory system, as shown by resorufin fluorescence, was observed following treatment with praziquantel and incubation with other molecules. Praziquantel treatment induced damage to the surface membrane as measured by the use of a variety of fluorescent compounds. The excretory system of the male worm was inhibited immediately after praziquantel treatment, but fully recovered after culture for 2 h following removal of praziquantel. The excretory system of the female, observed to be minimally active in untreated worm pairs, was often greatly activated in paired females, as shown by intense resorufin labelling, after praziquantel treatment, and this continued during recovery of the male excretory system. In experiments with normal worm pairs, the female could be activated by inhibiting the metabolic rate of the pair by a cooling procedure. The effects on the excretory system of changes in culture conditions (such as changes in pH, concentrations of bacterial lipopolysaccharide, cytokines, reactive oxygen species, compounds which remove cholesterol, such as beta-methyl cyclodextrin, and damaging basic poly-L-lysine) were also assessed. It is concluded that the extensive excretory system of the adult worm is responsive to drug treatment and to certain changes in environmental conditions. Its activity seems to be strongly linked to the integrity of the surface membrane.

The effect of acquired immunity on the duration of Plasmodium falciparum infections is unclear, although this is an important term in models of malaria transmission. It is problematical to determine the duration of infections because of the difficulty of distinguishing persisting infections from new ones, and because parasite densities are often transiently below the limit of detection. We recently developed a dynamic model for infection incidence, clearance and detection of multiple genotype P. falciparum infections and fitted it to a panel dataset from a longitudinal study in Northern Ghana. We now extend this model to allow for seasonal and age variation in infection rates and also age dependence in clearance and in detectability of infections. These models indicate that there is seasonal variation in the infection rate, and age dependence in detectability. The best fitting models had no age dependence in infection or clearance rates, suggesting that acquired immunity mainly affects detectability.

Until recently it had generally been assumed that apoptosis and other forms of programmed cell death evolved during evolution of the metazoans to regulate growth and development in these multicellular organisms. However, recent research is adding strength to the original phenotypic observations described almost a decade ago which indicated that some parasitic protozoa may have evolved a cell death pathway analogous to the process described as apoptosis in metazoa. Here we explore the implications of a programmed cell death pathway in the African tsetse-transmitted trypanosomes.