Schistosomula of Schistosoma mansoni were incubated in RPMI 1640 medium containing 10% fetal calf serum, 10% human portal venous or 10% human peripheral venous sera in the presence of bromodeoxyuridine (BrdU) in order to measure differences in cell proliferation. The rates of cell proliferation as expressed by BrdU labelling indices (BLI) were determined as a function of time of incubation by immunohistochemistry using monoclonal antibody to BrdU. Compared to schistosomula cultured in the presence of RPMI plus 10% fetal calf serum, BLIs were increased by 60% in the presence of human portal, but not in peripheral serum. This stimulatory effect was substantially reproduced by a fraction of portal serum with a molecular weight range between 1 and 50 kDa. However, in the presence of human peripheral venous serum, either whole or fractionated, schistosomula showed no significant difference compared to RPMI plus 10% fetal calf serum alone. Furthermore, human portal serum fractions of molecular weight greater than 50 kDa also revealed no significant difference relative to control. The results indicate that portal venous serum component(s) of a molecular weight range higher than most simple nutrients can greatly stimulate the rate of cell proliferation of Schistosoma mansoni schistosomula.

A single exposure to radiation-attenuated cercariae of Schistosoma mansoni induces a high level of protective immunity in C57BL/6 mice, which is mediated by Th1 responses. Events in the skin and/or draining lymph nodes early after exposure are crucial for the induction of protection, and we have investigated the interactions of vaccinating parasites with host leucocytes in these 2 locations. We observed extensive lateral spreading of cercarial secretions along layers of the stratum corneum but not between keratinocytes. There was little direct contact with host leucocytes during the first 1–2 days when the parasites lay at the base of the epidermis, but cells accumulated in the underlying dermis. In contrast to normal parasites, attenuated larvae persisted in the dermis for >10 days, often surrounded by aggregates of macrophages/dendritic cells. Whilst cells bearing MHC II, CD11b or CD11c markers were present in the lymph nodes, particularly in the periphery and paracortical areas, no obvious redistribution was seen as a result of parasite residence there for 5–15 days. However, ultrastructural observations revealed numerous cells with macrophage/dendritic morphology in the vicinity of parasites, in some instances closely adherent to the tegument. The observations strongly suggest that the tegument is a potent source of the antigens which prime the immune system in the lymph nodes of vaccinated mice for a protective response.

An enzyme presenting kallikrein-like activity (designated sK1) was purified from the supernatant of Schistosoma mansoni adult worm homogenate. The enzyme cleaves bradykinin from purified rat plasma kininogen. Activity was optimal at pH 9·0 and the enzyme showed amidolytic activity, since it hydrolysed the kallikrein synthetic substrate d-Pro-Phe-Arg-p-nitroanilide. The activity of sK1 upon rat plasma kininogen was strongly inhibited by the serine proteinase inhibitors phenylmethanesulfonyl fluoride, aprotinin or soybean trypsin inhibitor, but not by ethylenediaminetetraacetic acid or sodium tetrathionate. The molecular mass of sK1, as determined by polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate, was 66 kDa and the pI value, estimated by analytical chromatofocusing, was 4·2. Physical and chemical properties suggest that sK1 is a serine proteinase of the kallikrein family. Evidence is presented which suggests that sK1 is a component of the tegumental surface of the parasite and the levels of its activity in the male adult worm are approximately 21 times higher than those in the female adult worm. The intravenous injection of 3 μg of sK1 into an anaesthetized rat induced a drastic reduction in the arterial blood pressure of the animal. This effect lasted for about 1 min, and was followed by a progressive recovery of the arterial pressure. Neither bradycardia nor cardiac arrhythmias were noticed, suggesting a peripheral vasodilation effect. The presence of sK1 on the surface of adult male worms could play an important role in the wandering capacity of coupled worms into the visceral vasculature of the host.

To identify the genes encoding novel immunodominant antigens of Theileria parva a λgt11 library of piroplasm genomic DNA was immunoscreened with bovine recovery serum and a gene encoding a 150 kDa antigen (p150) was identified. The predicted polypeptide contains an N-terminal secretory signal sequence and a proline-rich region of repeated amino acid motifs. The repeat region is polymorphic between stocks of T. parva in both copy number and sequence, and analysis of the repeat region from 10 stocks of T. parva revealed 5 p150 variants. A monoclonal antibody (mAb) against the T. parva polymorphic immunodominant molecule (PIM) cross-reacted with the recombinant p150. The p150 has sequence homology with a PIM peptide sequence containing the anti-PIM mAb epitope. Immunoelectron microscopy demonstrated that the p150 antigen, like PIM, is located in the microspheres of the sporozoites and is exocytosed following sporozoite invasion of the host lymphocyte. By immunoelectron microscopy p150 was subsequently transiently detectable on the sporozoite surface and in the lymphocyte cytosol. Immunoblotting showed that p150 is also expressed by the schizont stage, but at much lower levels compared to the sporozoite. These results suggest a major role for p150 in the early events of host–sporozoite interaction.

Current theories assume that adaptive parasite evolution explains variation in the level of virulence and parasite success. In particular, mixed-genotype infections by parasites should generally be more virulent, and faster multiplying strains more successful, either because fixed strategies have evolved or because parasites facultatively alter virulence in response to co-infecting competitors. We compared several measures of parasite success and virulence between single-clone and mixed-clone infections of 2 strains of the trypanosome Crithidia bombi in its bumblebee host, Bombus terrestris. Contrary to expectation, we could not find differences between single-clone and mixed-clone infections in parasite prevalence, infection success, duration and clearance rate. However, a clearly significant effect of colony on infection intensity was present, and the colony effect emerged in virtually all other measures. We thus conclude that host environment as defined by the family (colony) genotype and thus host heterogeneity are more important in determining parasite virulence than the parasite characteristics. This does not invalidate modern theories of parasite evolution but suggests that variation in both hosts and parasites must be taken into account in more detail.

Entomopathogenic nematodes (EPN) are useful biological control agents of insect pests. However, the infective juvenile (IJ) stage which is the only stage to occur outside the host is susceptible to environmental extremes such as desiccation. We have isolated desiccation-tolerant strains of the EPN Heterorhabditis megidis. In this paper we describe the surface properties of these desiccation-tolerant mutants. Heterorhabditid IJs retain the sheath of the previous larval stage. The mutant lines possess alterations in the surface properties of the sheath. Differences were observed in fluorescent lipid analogue insertion into the surface of the sheath. Furthermore, cationized ferritin-binding studies demonstrated that the mutant lines possessed an increase in net negative surface charge. Removal of the surface layer of the sheath resulted in the loss of the mutant phenotype and in a reduction in the desiccation tolerance of the parental strain. Therefore, the negatively charged ‘surface coat’ appears to play an important role in the desiccation tolerance of Heterorhabditis species.

The present study examines antigenic variability for the human whipworm Trichuris trichiura. Recognition by IgG of somatic antigens of individual worms collected from 3 intensely infected children from Jamaica, West Indies has been investigated by immunoblotting. When probed with 1 plasma sample, significant differences in recognition of 2 selected antigens among worm populations and between male and female worms was observed. In addition, there was evidence for antigenic variability within worm populations at the individual worm level. Such variation may have considerable implications for the development of immunity to parasitic nematodes.

It has recently been shown using genetic markers that Ascaris in humans and pigs in Central America comprise reproductively isolated populations. We present a similar analysis for a region of China in which close association between pigs and humans has been the norm for thousands of years, and agricultural practices will result in frequent exposure to eggs from both sources. DNA fragments from selected regions of mitochondrial and ribosomal DNA were amplified by PCR and allelic forms identified following digestion with a panel of restriction enzymes, using DNA from a total of 115 individual worms from both people and pigs from 2 neighbouring villages. Significant frequency differences in both mtDNA haplotypes and the rDNA spacer were found between the 2 host-associated populations, indicating that they represented reproductively isolated populations. Mitochondrial haplotype frequencies were different from those observed in Guatemala and also from other Asian Ascaris populations, suggesting low levels of gene flow between populations. However, we found no evidence for significant heterogeneity in the genetic composition of Ascaris infrapopulations in either humans or pigs, possibly indicative of agricultural practices in China which have resulted in a random distribution of alleles within the parasite populations.

The initiation of further development is fundamental to the infectious processes of parasitic nematodes. We have examined early developmental activation of Trichinella spiralis larvae during host invasion, with particular emphasis on the timing of events. Using a novel approach, we have observed changes in tissue-specific transcriptional activity in live larvae during the infectious process with the fluorescent nucleic acid dyes SYTO12 and acridine orange. Simultaneously, the metabolic switch from anaerobic metabolism, characteristic of the infective stage, to aerobic metabolism, as found in the enteral stages, was tracked by measuring activities of the key regulatory enzymes phosphoenolpyruvate carboxykinase and pyruvate kinase, as well as isocitrate dehydrogenase (NADP) activity, and used as a co-indicator for developmental activation. Both metabolic enzyme activities and transcription patterns were found to change in response to host death, liberation from the nurse cell, and exposure to components of the host stomach environment. The results give a clear indication that the activation processes of T. spiralis infective larvae occur at a much earlier time than previously thought, and are stimulated upon liberation of the larvae from the nurse cell inside the host stomach.

The cestode Schistocephalus solidus is a simultaneous hermaphrodite that grows in 2 intermediate hosts and reproduces rapidly within a few days in the gut of a bird. Reproduction takes place by self- or cross-fertilization. Here, it was tested whether egg production differs between S. solidus that reproduce alone and those that are allowed to reproduce in pairs. Egg production in an in vitro system was found to depend on the cestodes' social situation. When kept alone, larger cestodes produced larger eggs. This was not so when kept in pairs – the difference between these 2 reproductive modes being highly significant in this respect. Further experiments revealed that, within the first 3 days, these hermaphrodites produced a larger total egg mass when kept alone than when kept in pairs. This was also reflected by the energy contents of the cestodes after this time-span: selfers had used up more energy than paired worms. Furthermore, S. solidus appeared to adjust its investment per egg depending on whether the offspring will be the result of self- or cross-fertilization. Selfers produced larger numbers of eggs, but these eggs were smaller and contained even smaller embryos per given egg size than eggs of potentially outbreeding cestodes. Selfed eggs reached lower hatching rates. Although this is to be expected from inbreeding depression it may also be an effect of the reduced maternal investment per egg. The observed phenotypic plasticity in the reproduction of S. solidus is discussed within 4 evolutionary frameworks: local mate competition adjusted for hermaphrodites, the hermaphrodite's dilemma, bet-hedging, and sib-competition.

Traditionally, the family Campulidae has been associated either with the family Fasciolidae, parasites of ruminants, or the Acanthocolpidae, parasites of fishes, based on morphological similarities. Since morphology does not seem to resolve clearly the problem of the relationships of campulids, we have used the sequences of the 18S rRNA gene of the campulids Zalophotrema hepaticum, Campula oblonga and Nasitrema globicephalae, the fasciolid Fasciola hepatica, the acanthocolpid Stephanostomum baccatum and the outgroup Schistosoma mansoni to infer a phylogeny. Maximum parsimony and neighbour-joining methods were applied. Both methods indicated that campulids are closer to acanthocolpids than fasciolids. In order to confirm this relationship, we generated a second phylogeny using all the partial sequences of the 18S published for trematodes: Lobatostoma manteri, Echinostoma caproni, Calicophoron calicophorum, Tetracerasta blepta, Gyliauchen sp. and Opistorchis viverrini, plus those mentioned above, and Dicrocoelium dendriticum. The aspidogastrean L. manteri was used as the outgroup. Results were identical to the first analysis. According to this and the most recent Digenean phylogeny, which considers campulids and acanthocolpids as sister groups, we suggest that a common origin for these 2 groups would imply a host-switching process. The life-cycle of acanthocolpids includes marine gastropods as first intermediate hosts, and fishes as second intermediate and definitive hosts. In this context, the hypothesis would be that trematodes whose cycle ended in fishes were able to switch to mammalian hosts.

The epidemiology of the sucking lice Echinophthirius horridus was studied over a 4 year period in which their harbour seal hosts experienced marked inter-annual changes in food availability. Prevalence and intensity of infection varied in relation to host age but not sex. Burdens were highest on immature seals, but both prevalence and intensity of infection were significantly higher in years when food availability was low. Observed intra-population variations in ectoparasite dynamics suggested that reported geographical differences in the prevalence of E. horridus may have resulted from methodological differences. In contrast to previous studies of other pinnipeds, lice were also absent on weaned pups, suggesting that E. horridus is transferred horizontally. No significant differences were found in the haematological parameters of infected and uninfected hosts. Nevertheless, there was a significant negative correlation between intensity of infection and several erythrocyte parameters, suggesting that high burdens of lice may compromise diving ability.