A deterministic mathematical model which predicts the probability of developing a new drug-resistant parasite population within the human host is reported. The model incorporates the host's specific antibody response to PfEMP1, and also investigates the influence of chemotherapy on the probability of developing a viable drug-resistant parasite population within the host. Results indicate that early treatment, and a high antibody threshold coupled with a long lag time between antibody stimulation and activity, are risk factors which increase the likelihood of developing a viable drug-resistant parasite population. High parasite mutation rates and fast PfEMP1 var gene switching are also identified as risk factors. The model output allows the relative importance of the various risk factors as well as the relationships between them to be established, thereby increasing the understanding of the conditions which favour the development of a new drug-resistant parasite population.

The taeniid tapeworm Echinococcus multilocularis is here reported for the first time at the Svalbard Archipelago in the Norwegian Arctic. This new finding is interesting because the establishment of E. multilocularis is due to a recent anthropogenic introduction of its intermediate host – the sibling vole Microtus rossiaemeridionalis at Svalbard. The parasite itself has probably become naturally transferred to Svalbard due to migratory movements of its final host – the arctic fox Alopex lagopus between source areas for E. multilocularis in Siberia and Svalbard. We report macroscopically determined prevalence of E. multilocularis from a sample of 224 voles trapped in August in 1999 and 2000. The prevalence was among the highest ever recorded in intermediate hosts and was dependent on age and sex of the hosts approaching 100% in overwintered males. The high prevalence and the simplicity of the vole – arctic fox – E. multilocularis system at Svalbard makes it an eminent model system for further epidemiological studies.

Tams1 is the polymorphic immunodominant merozoite-piroplasm surface protein of Theileria annulata. Evidence for selection of divergent forms of Tams1 has been obtained recently. This study was performed to address whether selection takes place during persistent infection of the bovine host or during passage through the Hyalomma tick vector. Four calves were infected with a T. annulata isolate representing multiple parasite genotypes. The development of the parasite population was analysed by denaturing gradient gel electrophoresis (DGGE) using the Tams1 gene as a marker. In addition, the parasitaemia was measured by a semi-quantitative reverse line blot hybridization assay in order to correlate Tams1 variation to changes in parasitaemia. It was found that both parasitaemia and parasite population displayed limited variation during persistent infection. Ticks were allowed to acquire T. annulata during 2 periods of the bovine infection. Tams1 alleles detected in ticks fed during acute infection were identical to the population in the bovine host. However, ticks fed during the carrier status acquired parasites showing a single Tams1 isotype that represented, in several cases, a minor population in the bovine host at the time of infestation. Although only a limited number of ticks were studied, these preliminary data suggest that specific parasite genotypes may be selected during tick transmission from a carrier animal.

The variability within schistosome populations was explored using mixed populations of cercariae from multimiracidial snail infections and individual clones of Schistosoma mansoni cercariae obtained from monomiracidial snail infections. We investigated the heterogeneity between different clones of S. mansoni with respect to infectivity and metabolism. One difference between clones of cercariae was found in the recovery of adult worms from Balb/C mice. Recovery of adult worms was greater after infections with a mixed population than with a clonal population. To investigate some biochemical features of individuals in clones or mixed populations, the uptake of [35S]methionine into individual parasites and their membrane proteins was measured. Isoelectric focusing of a soluble membrane fraction: the frozen–thawed supernatant extracted from individual clones, showed the presence of proteins of isoelectric point between 7.2 and 8.2 in all clones. These proteins were less labelled with [35S]methionine in the clones than in the mixed population. It was concluded that basic proteins are synthesized by all clones and in the mixed population but at different rates. Differences in the rate of incorporation of [35S]methionine into the surface membranes of schistosomula and adult worms derived from individual clones are reported. In addition, a direct correlation between the percentage of recovery of adult worms from mice infected with individual clones of S. mansoni and the rate of incorporation of [35S]methionine into schistosomula of these particular clones was observed. It is suggested that the high rate of metabolism shown by an individual clone may account for the enhanced survival of the cercariae derived from that clone during penetration of the skin and migration through the vertebrate host. In order to examine individuals in a population of schistosomula, from a clone or mixed population, the lysosome-specific fluorescent probe LysoTracker DND-99 was used to label the parasites and quantitative fluorescent measurements were made on individual parasites. There were significant differences between clones and a mixed population. Furthermore, the variation between individuals from a mixed population was greater than from that in any clone, just as was found in the infectivity studies. Freshly transformed schistosomula of individual clones labelled with the LysoTracker DND-99 showed less variations in the quantitative uptake of the dye within a single clone when compared to the mixed population. We conclude that for any biochemical and biological parameter, a population of cercariae consists of individuals showing a wide range of values, with a much greater range in a mixed population. This variability is likely to have great relevance for infectivity of the final host and the efficacy of drugs and the immune system.

The 14-3-3 protein is a key player in signal transduction processes in various species. We have previously cloned and expressed the 14-3-3 of Schistosoma mansoni. Using the purified protein we have now raised antibodies against it. A highly specific, affinity-purified antibody preparation was employed for the localization of the 14-3-3 protein in the parasite, by both immunohistochemistry and immunoelectron microscopy. The results demonstrate wide distribution of this protein. It was observed in the female excretory system, the nephridia as well as in the genital systems of both sexes, namely in the vitelline gland of female and in the testis of the male. It is also present in the parenchyma and muscle of both male and female worms. Immunoelectron microscopy demonstrated the presence of immunogold-labelled protein in the tegument, subtegument, muscle, parenchyma and in the female reproductive system, in both the cytoplasm and nucleus of vitelline cells, and oocytes. The possible role of the 14-3-3 protein in the genital organs is discussed.

The red poultry mite, Dermanyssus gallinae, is one of the most economically deleterious ecto-parasites of layer hens worldwide. D. gallinae is difficult to eliminate from infested poultry farms, and even to study, because it resides on the host only during the bloodmeal at night, and hides in the crevices of poultry houses during the day. Here, the life-cycle of D. gallinae was reproduced entirely in vitro. Mites were incubated in a glass pipette at 30 °C, 60–95% relative humidity and total darkness. A feeding apparatus, composed of a membrane, reservoir and blood was fitted on the pipette during bloodmeals. We tested feeding rates on blood mixed with 1 of 3 anti-coagulants (EDTA, heparin and trisodium citrate) at different concentrations, and biological and artificial membranes. The best engorgement and survival rates for all 3 haematophagous life-stages of the parasite were observed in 1-day-old chick membranes and heparinized (0.02 mmol/ml) blood. We then describe the steps in developing a complete self-sustaining in vitro life-cycle. A colony of mites was maintained in vitro for 7 generations. Losses in the first generation were heavy, but survival had multiplied 5-fold by the fifth generation. We hypothesize that heavy mortality rates during the first life-cycle correspond to selective pressure: only the mites which fed and survived in vitro were able to reproduce.

Thirty Holstein Friesian heifer calves, 3 months of age, were used to evaluate whether a low primary infection with Dictyocaulus viviparus would result in immunity against challenge infection 5 or 12 weeks later. Fifteen calves were experimentally infected with 30 larvae at day 0, while the other calves remained as uninfected controls. On day thirty-five 6 primary infected (G1) and 6 non-infected calves (G2) were challenged with 2000 larvae. These groups were necropsied on day 70. On day 84 the remaining 9 primary infected (G3) and 9 non-infected (G4) calves were similarly challenged and necropsied on day 119. Respiratory signs developed in most calves from approximately 2 weeks after challenge, but signs were more severe in G2 and G4 than in G1 and G3. The low primary infection resulted in significantly reduced faecal larval excretion (>80%) and worm burdens (>70%). In the primary infected groups the proportion of female worms in the worm burden was increased, suggesting that part of the developed immunity affected survival of the worms after their establishment in the host. Worm lengths and worm fecundity were also significantly reduced. All observed effects were stronger following challenge on day 84 than on day 35, but the effect of the duration of a primary infection (contrast between G1 and G3) was only statistically significant in the case of mean faecal larval excretion on the day of necropsy (P<0.05) and fecundity (P<0.05).

Parasite aggregation within the gill arches of 1 host species (Rutilus rutilus L.) was studied in the case of 9 congeneric monogeneans belonging to the genus Dactylogyrus. Both intra- and interspecific aggregation were followed by considering gill arches as independent patches. Parasite species were found to be aggregated both intra- and interspecifically. We showed that the intensity of parasite infection was positively related to intraspecific aggregation only for the more abundant species. No relationship was found between intensity of parasite infection and niche size when correcting for total parasite intensity. This may suggest that when parasite intensities are low, intraspecific competition may not have a strong effect on parasites. Conversely, when looking at the evolution of niche size and intraspecific aggregation for each species separately, intraspecific aggregation decreased for the most abundant species (D. crucifer) suggesting that this species is more competitive than others when total parasite intensity increases. When considering interspecific aggregation, Dactylogyrus species were found to be positively and negatively aggregated. Following the prediction of morphological and ecological similarity for congeneric species, Dactylogyrus species similar in morphometry of attachment parts tend to be more positively aggregated among their gill microhabitats than Dactylogyrus species with morphometric differences in attachment parts.

The complete nucleotide sequence of the mitochondrial (mt) DNA molecule of the liverfluke, Fasciola hepatica (phylum Platyhelminthes, class Trematoda, family Fasciolidae), was determined. It comprises 14462 bp, contains 12 protein-encoding, 2 ribosomal and 22 transfer RNA genes, and is the second complete flatworm (and the first trematode) mitochondrial sequence to be described in detail. All of the genes are transcribed from the same strand. Of the genes typically found in mitochondrial genomes of eumetazoans, only atp8 is absent. The nad4L and nad4 genes overlap by 40 nt. Most intergenic sequences are very short. Two larger non-coding regions are present. The longer one (817 nt) is located between trnG and cox3 and consists of 8 identical tandem repeats of 85 nt, rich in G and C, followed by 1 imperfect repeat. The shorter non-coding region (187 nt) exhibits no special features and is separated from the longer region by trnG. The gene arrangement resembles that of some other trematodes including the eastern Asian Schistosoma species (and cyclophyllidean cestode species) but it is strikingly different from that of the African schistosomes, represented by Schistosoma mansoni. The genetic code is as inferred previously for flatworms. Transfer RNA genes range in length from 58 to 70 nt, their products producing characteristic ‘clover leaf’ structures, except for tRNA(SUCN) and tRNA(SAGN) lacking the DHU arm.

The complexity of the life-cycle of many parasitic helminths has driven the evolution of several well-documented adaptations serving to facilitate the completion of each difficult step in the cycle. In some trematode taxa, selection has even favoured the truncation of the life-cycle: progenetic larval stages (metacercariae) are capable of producing eggs inside their usual intermediate hosts, eliminating the need to be transmitted by predation to a definitive host. In some species, progenesis is shown by all individuals, whereas in other species both the normal and truncated life-cycles coexist in the same populations. Here, the strategies chosen by normal and progenetic metacercariae of the trematode Coitocaecum parvum are investigated and compared. Normal metacercariae, i.e. metacercariae that only develop into adults following ingestion by a suitable fish definitive host, were not capable of manipulating the behaviour of their amphipod intermediate host in ways that could facilitate their capture by fish. These metacercariae were associated with increased mortality in amphipods. Progenetic metacercariae, on the other hand, showed lower virulence levels than normal metacercariae. At the time of their death, amphipods harbouring progenetic metacercariae contained close to the maximum number of eggs that can be produced by progenetic metacercariae, suggesting that the parasite lowers its virulence to keep its host alive long enough to maximize egg output. The maintenance of the 2 strategies in the C. parvum population suggests that they may have equal fitness payoffs: progenesis and low virulence guarantee the production of at least some eggs, whereas the normal life-cycle and its associated higher virulence in the intermediate host provides a low probability of much higher fecundity.

A polymorphic set of 14 kDa excretory–secretory (E–S) antigen-encoding cDNAs, with similarity to a previously characterized 15 kDa E–S antigen of Haemonchus contortus, was cloned from Cooperia punctata. Five cDNAs encoding predicted proteins of 70–80% identity were sequenced. Nucleotide sequence data reported in this manuscript are available in the GenBankTM databases under the accession numbers AF352718, AF352719, AF352720, AF352721, AF352722, AF352723, AF352724, AF352724, AF352726 and AF352727. Genomic analyses of individuals proved the existence of three 14 kDa E–S antigen-encoding genes, excluding that the differences reflected polymorphisms between individuals in a population. Southern blots indicated the presence of additional members of this gene family. Thus, despite the fact that heterologously expressed C. punctata 14 kDa E–S products are shown to be recognized by immune sera, potential pitfalls in the development of a recombinant vaccine are presented by this genetic diversity. Vaccine design could be further rationalized by knowledge of the function, and possible redundancy in function, of the E–S products which is presently lacking. The limitations encountered in assigning a function to the 14/15 kDa family of E–S proteins that is thus far unique to the trichostrongyloid nematodes are discussed.