Two experiments are described in each of which three groups of calves received infective larvae of Ostertagia ostertagi daily, each group at a different rate. Changes in worm burdens were followed by the periodic slaughter of calves.

Numbers of adult worms quickly rose to and remained at levels which bore a positive relation to the infection rate. There was evidence that a turnover of worms was occurring and it appeared that burdens of adult worms were regulated by a loss of worms which depended on the number present. As the host's experience of infection increased, the adult worms present were smaller and an increasing proportion of females lacked a fully developed vulval flap. Evidence is presented which suggests that these were effects of resistance and that they depended on separate mechanisms.

Faecal egg counts of groups carrying different numbers of worms rose to the same peak and then declined according to the same logarithmic curve. This finding, together with data concerning the number of eggs in the uteri of the worms, suggested that the egg output of the entire population was restricted by the host to a limit which depended on previous egg-laying.

Infection did not have an adverse effect on the growth of the calves unless the infection rate exceeded a critical value. The resistance of calves to infection was impaired by clinical ostertagiasis.

Variant antigenic types (VATs) represented in a total of 47 first relapse populations of 6 clones of Trypanosoma brucei LUMP 227 were identified by immunofluorescent staining of living trypanosomes, using antiserum raised against purified surface antigens. The relative growth rates of these 6 clones were measured both individually and when grown together in a mixed population, and were found to be different under these two sets of conditions. A pattern emerged in the VATs represented in relapses of each clone, with some types being expressed more frequently than others and certain VATs being only very rarely expressed. It is suggested that new VATs are expressed according to a statistically definable order of priority which is different for each parent VAT, and that some VATs may be able to change to certain others only after passing through an intermediate VAT. The order of priority of appearance of VATs does not appear to correlate with growth rate measured either in individual clones or when clones are grown in a mixed population.

An apparatus for the artificial feeding of tsetse has been described. Fresh defibrinated blood at 36 °C is circulated with a peristaltic pump through feeding capsules. The efficiency of the method has been compared with that of the Cockings' method and with the live host feeding. The flies exposed to feeding on this apparatus showed significantly higher feeding frequency than those allowed to feed on the Cockings' apparatus. Compared with the live host feeding, the feeding frequency for circulating blood method was slightly lower but the difference was not significant. Once the fly is induced to feed, it would in most cases complete engorgement unless disturbed. The rate of crop emptying in flies fed by the Cockings' method was abnormally slower as compared to those fed on the circulating blood. The present study indicates that for the successful artificial feeding of tsetse, it is essential to circulate the blood and regulate the temperature of the medium and the feeding surface to approximately 36 °C.

I wish to thank Mr S. B. Kutuza for technical help, and the Director of E.A.T.R.O. for permission to publish this work.

The occurrence of multiple-clone Plasmodium falciparum haploid blood-stage infections is a pre-requisite for cross-fertilization and genetic exchange at the diploid stage in the mosquito. Using monoclonal antibodies against 3 polymorphic blood-stage antigens, a method of two-colour differential immunofluorescence allowed the resolution of between 1 and 4 clones/isolate. A mean of 2 P. falciparum clones was detected in the blood of malaria patients in The Gambia. The mean number of clones/patient showed no correlation with age, parasitaemia, or disease severity. There was a slight difference in mean number between sample periods, probably reflecting temporal differences in transmission intensity. A statistical analysis of 2-locus genetic diversity of clones within isolates concludes that not all multiple-clone infections result from superinfection, but that some are due to single multiple-clone inoculations.

The value of 2 PCR methods, targeting genomic and kinetoplast minicircle DNA respectively, was investigated for both diagnosis and prevalence studies of canine visceral leishmaniasis (CVL). The first method (R) was 5000-fold less sensitive than the second (method KRV). Both were tested for diagnosis of CVL in 44 sick dogs with confirmed disease using different biological samples. Method R was highly efficient when using invasive samples, but the use of method KRV proved necessary for a 100% sensitive diagnosis using peripheral blood. This method was applied to peripheral blood and skin samples in 263 dogs during a mass survey in the Cévennes focus. PCR was compared to serology and all results were analysed according to clinical status. The ‘CVL-infection’ prevalence was found to be 79.8% by PCR compared with 29.6% by serology: 89.4% of symptomatic and 65.2% of asymptomatic dogs harboured parasites in peripheral blood. This study confirms the high prevalence of asymptomatic carriers of Leishmania. In total, for the diagnosis of CVL in sick dogs, method R is recommended in view of its 100% positive predictive value (compared with 30% for method KRV). A strategy best adapted for prevalence surveys might combine serology and highly sensitive PCR on peripheral blood.

Neospora caninum is an apicomplexan parasite that is closely related to Toxoplasma gondii, the causative agent of toxoplasmosis in humans and domestic animals. However, in contrast to T. gondii, N. caninum represents a major cause of abortion in cattle, pointing towards distinct differences in the biology of these two species. There are 3 distinct key features that represent potential targets for prevention of infection or intervention against disease caused by N. caninum. Firstly, tachyzoites are capable of infecting a large variety of host cells in vitro and in vivo. Secondly, the parasite exploits its ability to respond to alterations in living conditions by converting into another stage (tachyzoite-to-bradyzoite or vice versa). Thirdly, by analogy with T. gondii, this parasite has evolved mechanisms that modulate its host cells according to its own requirements, and these must, especially in the case of the bradyzoite stage, involve mechanisms that ensure long-term survival of not only the parasite but also of the host cell. In order to elucidate the molecular and cellular bases of these important features of N. caninum, cell culture-based approaches and laboratory animal models are being exploited. In this review, we will summarize the current achievements related to host cell and parasite cell biology, and will discuss potential applications for prevention of infection and/or disease by reviewing corresponding work performed in murine laboratory infection models and in cattle.

Several species of malarial protozoans commonly parasitize the same host population and often the same individual host. This paper reviews the evidence for interactions among such host-sharing parasites. Field studies measuring the cross-sectional prevalence of malarial species often record fewer mixed infections than expected by chance, suggesting that one parasite has excluded another or suppressed its parasitaemia to undetectable levels. Prevalences may vary reciprocally between seasons, with one species increasing in prevalence while another decreases, despite parallel increases in the transmission rates of both, again suggesting suppression of one species by another. However, longitudinal studies of individual hosts indicate that malarial parasites may also favourably affect the host environment for each other, perhaps due to their depressive effect on the immune system: this is shown by the recrudescence of a latent malarial species immediately before or after the parasitic wave of another species. The suppression hypothesis is supported by data derived from the simultaneous inoculation of two Plasmodium species into laboratory animals; many studies have shown that one or both species are suppressed. This may be mediated by competition for host cells or nutrients, or by heterologous immunity. However, the suppressed species rebounds after the other species has abated, and may show a prolonged infection. Experimental evidence that one species can facilitate the recrudescence of another is minimal, but this may reflect the paucity of investigations of this phenomenon. Laboratory studies show only minor cross-resistance between host-sharing species, which is consistent with the hypothesis that their co-occurrence has led to antigenic divergence or that species showing strong heterologous resistance cannot co-exist in the same host population. Such complementarity occurs not only with the host immune response but also with many other life-history characteristics of host-sharing parasites, such as host cell preference. I conclude that malarial species have been important in each other's evolution, particularly in the tropics where multi-species complexes are common.

1. The egg-shell in many trematodes and pseudophyllidean cestodes is a quinone-tanned protein; and histochemical methods for polyphenol oxidase or for phenols may be used to demonstrate regions producing or containing shell material (i.e. the vitellaria, its ducts and the uterus) in whole mount preparations.

2. Specimens so treated may be further stained by routine whole mount methods. Gower's carmine is particularly recommended.

3. 70% alcohol was found to be the best general fixative for these methods.

4. These histochemical methods—with certain modifications—may also be used for demonstrating the egg-shell producing regions in sections.

5. Because of their affinity for basic proteins, bromo-phenol-blue and malachite green are suitable for staining these regions in sections. These stains are unsuitable for whole mounts.

6. There is some evidence to suggest that all trematodes and pseudophyllidean cestodes do not form and harden their egg-shell by quinone tanning; consequently, some forms may give negative or unsatisfactory results with these methods.

Sand fly saliva plays an important role in Leishmania transmission. We characterized the host antibody response to saliva from 3 sand fly species. Specific IgG was observed in sera from experimentally bitten mice as well as in sera from individuals living in the endemic area of Leishmania tropica in Sanliurfa, Turkey. Sera of Sanliurfa inhabitants showed high IgG levels against saliva of Phlebotomus sergenti and P. papatasi, the 2 most abundant sand fly species in this area, but did not react with saliva of the New World sand fly, Lutzomyia longipalpis. Patients with active Le. tropica lesions possessed significantly higher anti-P. sergenti IgG levels than the healthy individuals from the same place while anti-P. papatasi IgG levels were equal in both groups. Major protein bands in P. papatasi and P. sergenti saliva reacted with both, human and mice sera; in P. papatasi, however, mouse IgG recognized preferentially the 42 kDa protein band while the human IgG reacted strongly with the 30 kDa band. Our data suggest that the antibody response to sand fly saliva could be used for monitoring the exposure of humans and other hosts to sand flies and might be used as a marker of risks for Leishmania transmission in endemic areas.

Few studies have described the combined effect of age, gender, management and control programmes on helminth prevalence and egg shedding in grazing equines. Here, fecal samples collected from 1221 Thoroughbred horses, residing at 22 studs in the UK, were analysed. The distribution of strongyle eggs amongst individuals in relation to age, gender and management practices was investigated. Fecal worm egg counts (FWECs), described as the number of eggs per gramme (epg) of feces, were determined using a modification of the salt flotation method. The FWEC prevalence (mean%) of strongyles, Parascaris equorum, tapeworm spp. and Strongyloides westeri was 56, 9, 4 and 8%, respectively. Strongyle, P. equorum, tapeworm spp. and S. westeri infections were detected on 22 (100%), 11 (50%), 9 (41%) and 8 (36%) of studs, respectively. Within all age and gender categories, strongyle FWECs were highly over-dispersed (arithmetic mean = 95 epg, aggregation parameter k=0·111) amongst horses. Animal age, last anthelmintic type administered and management practices (for example, group rotation on grazing) most strongly influenced strongyle prevalence and level of egg shedding (P < 0·05). Overall, 11% of equines (range: 234–2565 epg) were responsible for excreting 80% of the strongyle eggs detected on FWEC analysis. The results confirm that the judicious application of targeted treatments has potential to control equine strongyle populations by protecting individual horses from high burdens, whilst promoting refugia for anthelmintic susceptible genotypes.

Mucus is a sticky visco-elastic material which coats all mucosal surfaces. Florey, in 1955, noted the following three functions for gastrointestinal mucus: protection of the underlying mucosa from chemical and physical injury, lubrication of the mucosal surface to facilitate passage of luminal contents, and removal of parasites by binding and entrapment. In the 31 years since Florey's review, detailed analyses of the composition of mucus and of the biochemistry of mucin glycoproteins, as well as measurements of the physical properties of mucus from different organs and sites have yielded information at the molecular level which provide additional support for his views on its function (Allen, 1981; Forstner, Wesley & Forstner, 1982).

From hundreds of independent transitions from a free-living existence to a parasitic mode of life, separate parasite lineages have converged over evolutionary time to share traits and exploit their hosts in similar ways. Here, we first summarize the evidence that, at a phenotypic level, eukaryotic parasite lineages have all converged toward only six general parasitic strategies: parasitoid, parasitic castrator, directly transmitted parasite, trophically transmitted parasite, vector-transmitted parasite or micropredator. We argue that these strategies represent adaptive peaks, with the similarities among unrelated taxa within any strategy extending to all basic aspects of host exploitation and transmission among hosts and transcending phylogenetic boundaries. Then, we extend our examination of convergent patterns by looking at the evolution of parasite genomes. Despite the limited taxonomic coverage of sequenced parasite genomes currently available, we find some evidence of parallel evolution among unrelated parasite taxa with respect to genome reduction or compaction, and gene losses or gains. Matching such changes in parasite genomes with the broad phenotypic traits that define the convergence of parasites toward only six strategies of host exploitation is not possible at present. Nevertheless, as more parasite genomes become available, we may be able to detect clear trends in the evolution of parasitic genome architectures representing true convergent adaptive peaks, the genomic equivalents of the phenotypic strategies used by all parasites.

Studies of patterns of reinfection with four species of intestinal nematodes (Ascaris, hookworm, Trichuris and Enterobius) in 174 individual patients following chemotherapeutic treatment revealed statistical evidence for predisposition to heavy or light infection (relative to the average level in the overall population). Analyses of associations between the abundances of the four species of nematodes within a combined sample of 525 worm burdens showed significant correlations between 5 out of the 6 possible pair-wise comparisons between species. The relevance of these results to the design of control programmes based on chemotherapeutic application is discussed.

The current trend of antimony (Sb) unresponsiveness in the Indian subcontinent is a major impediment to effective chemotherapy of visceral leishmaniasis (VL). Although contributory mechanisms studied in laboratory-raised Sb-R parasites include an up-regulation of drug efflux pumps and increased thiols, their role in clinical isolates is not yet substantiated. Accordingly, our objectives were to study the contributory role of thiols in the generation of Sb unresponsiveness in clinical isolates. Promastigotes were isolated from VL patients who were either Sb responsive (n=2) or unresponsive (n=3). Levels of thiols as measured by HPLC and flow cytometry showed higher basal levels of thiols and a faster rate of thiol regeneration in Sb unresponsive strains as compared with sensitive strains. The effects of antimony on generation of reactive oxygen species (ROS) in normal and thiol-depleted conditions as also their H2O2 scavenging activity indicated that in unresponsive parasites, Sb-mediated ROS generation was curtailed, which could be reversed by depletion of thiols and was accompanied by a higher H2O2 scavenging activity. Higher levels of thiols in Sb-unresponsive field isolates from patients with VL protect parasites from Sb-mediated oxidative stress, thereby contributing to the antimony resistance phenotype.

In parallel with massive research efforts in human schistosomiasis over the past 30 years, persistent efforts have been made to understand the basis for compatibility and incompatibility in molluscan schistosomiasis. Snail plasma contains molecules that are toxic to trematodes, but these seem to kill only species that never parasitize the mollusc used as the source of plasma. A sporocyst will be killed actively by haemocytes alone if they are from a snail that is resistant to the trematode. Oxygen-dependent killing mechanisms play a major role. Enzymes such as NADPH oxidase, superoxide dismutase, myeloperoxidase and nitric oxide synthase are critical components of the putative killing pathways. Metabolic intermediates such as hydrogen peroxide and nitric oxide appear to be more important against trematodes than the shorter-lived intermediates that are more important in anti-microbial defences. Products secreted by trematode larvae influence the physiology of snail haemocytes, implying active counter-defences mounted by the parasite, but these remain largely unexplored. A possible molecular basis for the susceptibility/resistance dichotomy in molluscan schistosomiasis is suggested to be deficient forms of enzymes in the respiratory burst pathway, and a selective disadvantage for schistosome resistance is an integral component of this model.

Polymerase chain reaction–ribosomal ITS-1 DNA (rDNA) restriction fragment length polymorphism (PCR–RFLP) analysis and sequencing of the mitochondrial cytochrome c oxidase subunit 1 (CO1) and NADH dehydrogenase 1 (ND1) genes were used to characterize 33 Echinococcus granulosus isolates collected from different regions and hosts in Argentina, and to determine which genotypes occurred in humans with cystic hydatid disease. The results of the study demonstrated the presence of at least 4 distinct genotypes; the common sheep strain (G1) in sheep from Chubut Province and in humans from Río Negro Province, the Tasmanian sheep strain (G2) in sheep and 1 human from Tucumán Province, the pig strain (G7) in pigs from Santa Fe Province and the camel strain (G6) in humans from Río Negro and Buenos Aires Provinces. The finding that pigs harboured the pig strain and the occurrence of the Tasmanian sheep strain has considerable implications for the implementation of hydatid control programmes due to the shorter maturation time of both strains in dogs compared with the common sheep strain. Furthermore, this is the first report of the presence of the G2 and G6 genotypes in humans which may also have important consequences for human health.

Recombinant vaccines have been developed which are highly effective in preventing infection with Taenia ovis in sheep, Taenia saginata in cattle, Taenia solium in pigs and Echinococcus granulosus in livestock animals. T. ovis and T. saginata are economically significant parasites and the commercial success or otherwise of vaccines against them will rely on their economic value. E. granulosus and T. solium are zoonotic parasites that cause cystic hydatid disease and neurocysticercosis, respectively, in humans. Vaccines against these parasites have been developed to assist with the control of transmission of the human diseases rather than for prevention of infections in livestock per se. Regions of high prevalence for cystic hydatid disease and neurocysticercosis occur primarily in the developing world. As a consequence, vaccines against them are of little or no commercially interest – they are Orphan Vaccines. Lack of commercial interest in these vaccines has made public sector support for their development necessary well beyond the research phase trough into completion of commercial scale-up and other more commercially-related assessments. Practical use of the vaccines will require commercial-scale production according to international manufacturing standards. Identifying partners and support in this endeavour is now of prime importance in efforts to achieve the potential of these vaccines as new tools for the control of cystic hydatid disease and neurocysticercosis.

Six murine monoclonal antibodies to Leishmania tropica major have been prepared and the properties of these antibodies studied. Two (WIC 79.3 and 79.7) were L. tropica major species-specific and bound to promastigote cell surfaces, to parasitized macrophages, but not isolated amastigotes. No evidence was found for the production of antibody to the antigenic determinants recognized by WIC 79.3 or 79.7 during L. tropica major infections in mice and hamsters. One antibody (WIC 79.1) bound to sub-cellular organelles of Leishmania species but to a different sub-cellular organelle of Trypanosoma cruzi. Two others bound to the flagellum, one (WIC 79.2) to all Leishmania species, T. cruzi and Trypanosoma brucei, the other (WIC 79.4) only of L. tropica major and L. donovani species. One antibody (WIC 79.5) was directed against an unknown internal antigen found in all Leishmania species and T. cruzi

Commitment to the production of female and male gametocytes was studied in the NF54 line of the human malaria parasite Plasmodium falciparum. The development of sibling parasites derived from individual schizonts was followed, and 2 antisera against the female gametocyte-specific protein Pfg377 and the male gametocyte-specific protein α-tubulin II were used to determine the sex of sibling gametocytes. The experiment showed that individual cohorts of sibling gametocytes were stained in a mutually exclusive fashion by only one or the other antiserum, indicating that individual schizonts committed to yield sexual parasite progeny produce gametocytes of the same sex. This work suggests that in P. falciparum commitment to sexual differentiation occurs prior to schizont maturation, at the same moment when the sex of the resulting gametocytes is determined.

Recent developments in parasite immune evasion and exploitation are reviewed with special reference to the papers presented in this volume. Parasites, broadly defined, of animals with good immune responses have evolved many strategies that adapt them to survive and reproduce. These strategies may be passive, or may involve active intervention with host immune regulation, and can be categorized as immune evasion, immune exploitation and molecular piracy. The concept of immune evasion began with Paul Ehrlich's demonstration of antigenic variation in African trypanosomes and was reinforced by later ideas on molecular mimicry. Molecular mimicry is updated in the light of recent discoveries about degeneracy and plasticity of TCR/MHC-peptide recognition. Possible connections between two of its postulated consequences, evasion and autoimmunity, are discussed. Another putative consequence of molecular mimicry, host antigenic polymorphism, is also updated. The concept of exploitation of host immune responses by parasites has been reinforced by new data on its first known examples, especially the immune dependence of schistosome egg excretion. Newer examples include use of host cytokines as parasite growth factors, virokines, viroreceptors and helminth pseudocytokines. Finally, questions of host gene capture by viruses and possible horizontal gene transfer between host and parasite mediated by retroviruses are examined. The latter is compared with molecular conservation as a source of molecular mimicry and other aspects of host–parasite coevolution.