Two sets of oligonucleotide primers, one derived from a repetitive sequence and the other from the gene encoding a 67 kDa sporozoite antigen of Theileria parva, were used to amplify parasite DNA from the blood of T. parva-infected carrier cattle using the polymerase chain reaction (PCR). PCR amplification products were obtained from 15 carrier cattle infected with one of 4 different T. parva stocks. Successful amplifications were performed using DNA from 2 cattle infected with T. p. parva Pemba Mnarani, 10 cattle infected with T. p. parva Marikebuni, 2 cattle infected with T. p. bovis Boleni and 1 animal infected with T. p. lawrencei 7014. No amplification products were obtained from any of 7 cattle which had been infected with the T. p. parva Muguga stock. A synthetic oligonucleotide, which hybridized specifically to T. p. parva Marikebuni DNA among 6 T. parva stocks tested, was designed using sequence data from within the region of the T. parva genome amplified by the repetitive sequence primers. The oligonucleotide was used to probe PCR products and to increase the sensitivity and specificity of carrier animal detection. Southern blot analysis using a T. parva repetitive sequence probe demonstrated the existence of restriction fragment length polymorphisms between parasites isolated from T. p. parva Marikebuni-infected carrier cattle. The use of the PCR and other methods of carrier animal detection are discussed.

Gyrodactylus salaris, an important pathogen of Atlantic salmon Salmo salar, has been shown to be highly sensitive to factors in host serum and mucus, being killed rapidly (50% within 1h) by serum at a dilution of 1[ratio ]200. The time needed for killing was inversely proportional to serum concentration. Similar effects were noted using host mucus, which contained approximately 1/20th of the anti-Gyrodactylus activity of serum. Serum activity was abolished completely by heating at 45°C for 30 min, and by addition of EDTA, but not by EGTA+1 mm magnesium ions. Activity was not dependent on whether the serum was from infected or naive fishes, nor was it species specific. Attempts to pre-coat parasites in salmon anti-Gyrodactylus antibodies also failed to enhance the activity of fresh serum. These observations suggest that killing is due to the complement system of the host, acting via the alternate pathway. G. salaris appears to be exceptionally sensitive to complement, being killed at concentrations which could be experienced in vivo. The role of complement in the protection of fishes against gyrodactylid infection therefore deserves further investigation.

Some factors controlling differentiation of the protoscoleces of Echinococcus into strobilate or cystic stages, in vitro, have been identified. Development in a strobilate direction requires (a) evagination of the scolex, (b) contact of the evaginated scolex with a suitable nutritive substrate. These conditions are satisfied by treating fresh protoscoleces with pepsin and by evaginating them with trypsin + pancreatin + dog bile and subsequently culturing in diphasic media.

The most satisfactory diphasic system so far developed consisted of a base of bovine serum (coagulated by heating) with a liquid phase of Parker 199 medium +20% hydatid fluid and a gas phase of 8% O2+5% CO2 in N2. In this diphasic system, protoscoleces have been cultured to the 3-proglottid strobilate stage with the anlagen of the male and female genitalia formed: the first proglottid appeared after 30 days culture in vitro compared with 16 days in vivo; three proglottids were formed after 60 days. Development in vitro stopped just short of mature egg-producing worms.

That an intimate contact of a ‘placental’ nature takes place between the parasite and the solid proteinaceous base is shown by the fact that strobilization never took place in cultures lacking bases or in those in which the serum base was separated from the protoscoleces by a cellulose membrane. Cytological and ultra-structure studies of an evaginated protoscolex revealed that fully developed microtriches were present only in the rostellar region and not in the post-sucker region. It is concluded that it is this anterior region which makes initial contact with the nutritive substrate—a contact which appears to provide the ‘strobilization stimulus’. It is not known if the ‘strobilization stimulus’ is stimulatory (i.e. nervous) or nutritive or both. Contact with a non-nutritive base (i.e. agar) does not induce strobilization.

In any one sample of protoscoleces, after a high initial evagination, only about 30–40% finally remained evaginated and continued to show sucker activity. The degree of evagination appears to be characteristic for any sample of protoscoleces and may be related to the source of the hydatid cysts. It may prove to be a genetically determined character.

Evaginated protoscoleces became inactive under certain conditions and the degree of activity seems to be related to stimulatory and nutritive factors. The activity of evaginated protoscoleces was used as a criterion for assessing the effectiveness of a culture medium or condition. Evaginated protoscoleces which lost their activity developed small posterior bladders as did the majority of unevaginated protoscoleces. In both cases, posterior bladders enlarged and secreted a PAS-positive layer which formed the laminated membrane of a miniature hydatid cyst.

The well-known appearance of vesicular forms in cultures appears to be related to unfavourable culture conditions in vitro, such as a low or high pH or a low or high O2 tension.

It is pointed out that since the morphogenesis of Echinococcus protoscoleces can now be controlled to a considerable extent in vitro and since the contents of any one cyst have the same genotype, this organism provides exceptionally interesting material for studies on morphogenesis.

Acknowledgement is made to the Australian Wool Board, the National Institutes of Health, U.S. Public Health Services Grant No. AI-04707–03 and the World Health Organizati on for generous financial assistance in support of this work.

The skilful technical assistance of Mr A. B. Howkins and Mrs Mary Barton is also acknowledged.

The diversity and abundance of parasites vary widely among populations of the same host species. These infection parameters are, to some extent, determined by characteristics of the host population or of its habitat. Recent studies have supported predictions derived from epidemiological models regarding the influence of host population density: parasite abundance and parasite species richness are expected to increase with increasing host population density, at least for directly transmitted parasites. Here, we test this prediction using a natural system in which populations of the crucian carp, Carassius carassius (L.), occur alone, with no other fish species, in a series of 9 isolated ponds in Finland. The ectoparasite communities in these fish populations consist of only 4 species of monogeneans (Dactylogyrus formosus, D. wegeneri, D. intermedius and Gyrodactylus carassii); the total and relative abundance of these 4 species varies among ponds, with one or two of the species missing from certain ponds. Across ponds, only one factor, total fish population size, explained a significant portion of the variance in both the mean number of monogenean species per fish and the mean total abundance of monogenean individual per fish. In contrast, fish population density did not influence either monogenean abundance or species richness, and neither did any of the other variables investigated (mean fish length per pond, number of fish examined per pond, distance to the nearest lake, and several water quality measures). In our system, proximity among fish individuals (i.e. host population density) may not be relevant to the proliferation of monogeneans; instead, the overall availability of host individuals in the host population appeared to be the main constraint limiting parasite population growth.

Additional ultrastructural features of the tegument of Fasciola hepatica, after glutaraldehyde fixation and high resolution, have been observed.

Two distinct types of tegumental cells are shown to be present, the type 1 cell having been previously described. Details of the fine structure of the nucleus, mitochondria, endoplasmic reticulum and Golgi complex of the type 2 cell are given. The type 2 cell is characterized by biconcave secretory bodies derived from the Golgi complex and the latter is intimately related to the granular endoplasmic reticulum, from which it is derived by blebbing. The type 2 cells are connected by protoplasmic tubes to the same surface syncytium as type 1 cells.

These new findings of the structure of the tegument of Fasciola are compared with previous reports on the ultrastructure of the tegument in other flukes.

I should like to express my sincere thanks to the following: the Wellcome Trust and S.R.C. for grants to purchase an Akashi TRS 50 and an A.E.I. E.M. 6B electron microscope respectively; the Royal Society for a personal grant to purchase a Reichert Ultramicrotome and a vacuum coating unit.

My thanks are also due to Mr W. Ferguson for photographic skill and to Mr A. Lyness for technical assistance and microscope maintenance, and my Research Associate, Mrs S. S. E. Dermott, for expert electron-microscopy.

The form of the egg, the structure of the oncomiracidium and the subsequent development of Entobdella soleae, a monogenean skin parasite of the common sole, are described. The reproductive organs develop in a constant sequence and the parasite is protandrous. Glands of Goto were observed in post-oncomiracidia of all ages.

The early embryo of E. soleae possesses eight pairs of marginal hooklets and two pairs of hamuli. One pair of hooklets, centrally situated on the adhesive organ, undergoes metamorphosis and becomes the pair of accessory sclerites associated with the hamuli in the adult.

My thanks are due to Dr J. Llewellyn for continued advice and criticism. The work was conducted during the tenure of a Fishery Research Training Grant from the Development Commission.

The 18S nuclear small subunit ribosomal RNA gene of piroplasms from wildlife and human cases of babesiosis in the western USA were isolated by PCR and sequenced. Phylogenetic analyses of these sequences and comparisons with sequences from other Babesia and Theileria species revealed that piroplasm isolates from the human cases were indistinguishable from some of the isolates from the western wildlife species, most notably the isolates from mule deer (Odocoileus hemionus). These results suggest that large ungulates may serve as reservoirs for human piroplasm infection. The western piroplasm isolates from humans and wildlife formed a distinct clade, separate from other piroplasms found worldwide.

Relatively simple protocols employing non-radioactive DNA probes have been used for the detection of African trypanosomes in the blood of mammalian hosts and the saliva of live tsetse flies. In combination with the polymerase chain reaction (PCR), the protocols revealed trypanosomes in buffy-coat samples from antigenaemic but aparasitaemic cattle and in the saliva of live, infected tsetse flies. Furthermore, the protocols were used to demonstrate concurrent natural infections of single tsetse flies with different species of African trypanosomes.

Patterns of genetic subdivision in parasite populations can provide important insights into transmission processes and complement information obtained using traditional epidemiological techniques. We describe mitochondrial sequence variation in 265 Ascaris collected from 62 individual hosts (humans and pigs) from 35 households in 3 Guatemalan locations. Restriction mapping of individual worms revealed 42 distinct mitochondrial genotypes. We ask whether the mitochondrial genotypes found in worms from individual hosts, from families of hosts and from villages represent random samples from the total Ascaris population. Patterns of genetic subdivision were quantified using F-statistics, while deviations from the null hypothesis of randomness were evaluated by a simple resampling procedure. The analysis revealed significant deviations from panmixia. Parasite populations were strongly structured at the level of the individual host in both humans and pigs: parasites bearing the same mitochondrial genotype were found more frequently than would be expected by chance within hosts. Significant heterogeneity was also observed among populations from different villages, but not from different families within a village. The clustering of related parasites within hosts suggests a similar clustering of related infective stages in the environment and may explain why sex ratios in Ascaris are female-biased. We discuss aspects of Ascaris biology which may lead to the observed patterns.

We investigated the genotypic composition of the digenetic parasite Schistosoma mansoni for its adult stages within the definitive host (the wild rat, Rattus rattus) and for the larval stages within the intermediate host (the snail, Biomphalaria glabrata) both collected at the same transmission site. Our analyses are based upon the recognition and distribution of 200 different multilocus genotypes generated by RAPD markers. While intramolluscan larval infrapopulations are characterized by a low infection rate (0·6% on average) and low intra-host genetic diversity (1·1 genotype on average per infected snail), adult infrapopulations within rats showed a high infection rate (94%) and a substantial intra-host genetic diversity (34 genotypes on average) linked to high intensities (160 worms per host on average). A single definitive host bearing 105 different genotypes harboured 52% of the total genetic diversity detected within the whole parasite population. Analysis of the genetic data allowed the identification of various ecological, behavioural and immunological factors which are likely to enhance transmission of multiple parasite genotypes towards the vertebrate hosts. From the distribution of repeated identical multilocus genotypes within the parasite population and among the hosts, we have inferred different parameters of the cercarial transmission efficiency as well as patterns and processes by which vertebrate hosts acquire infection in the field.

Non-availability of adult worms from living hosts remains a key problem in population genetic studies of schistosomes. Indirect sampling involving passage through laboratory animals presents significant ethical and practical drawbacks, and may result in sampling biases such as bottlenecking processes and/or host-induced selection pressures. The novel techniques reported here for sampling, storage and multi-locus microsatellite analysis of larval Schistosoma mansoni, allowing genotyping of up to 7 microsatellite loci from a single larva, circumvent these problems. The utility of these assays and the potential problems of laboratory passage, were evaluated using 7 S. mansoni population isolates collected from school-children in the Hoima district of Uganda, by comparing the associated field-collected miracidia with adult worms and miracidia obtained from a single generation in laboratory mice. Analyses of laboratory-passaged material erroneously indicated the presence of geographical structuring in the population, emphasizing the dangers of indirect sampling for population genetic studies. Bottlenecking and/or other sampling effects were demonstrated by reduced variability of adult worms compared to their parent field-collected larval samples. Patterns of heterozygote deficiency were apparent in the field-collected samples, which were not evident in laboratory-derived samples, potentially indicative of heterozygote advantage in establishment within laboratory hosts. Genetic distance between life-cycle stages in the majority of isolates revealed that adult worms and laboratory-passaged miracidia clustered together whilst segregating from field miracidia, thereby further highlighting the utility of this assay.

Existing models of the basic case reproduction number (R0) for vector-borne diseases assume (i) that the distribution of vectors over the susceptible host species is homogenous and (ii) that the biting preference for the susceptible host species rather than other potential hosts is a constant. Empirical evidence contradicts both assumptions, with important consequences for disease transmission. In this paper we develop an Ideal Free Distribution (IFD) model of host choice by blood-sucking insects, predicated on the argument that vectors must have evolved to choose the least defensive hosts in order to maximize their feeding success. From a re-analysis of existing data, we demonstrate that the interference constant, m, of the IFD can vary between host species. As a result, the predicted distribution of insects over hosts has 2 desirable and intuitively plausible behaviours: that it is heterogeneous both within and between host species; and that the intensity of heterogeneity varies with host and vector density. When the IFD model is incorporated into R0, the relationship with the vector: host ratio becomes non-linear. If correct, the IFD could add considerable realism to models which seek to predict the effect of these ecological parameters on disease transmission as they vary naturally (e.g. through seasonality in vector density or host population movement) or as a consequence of artificial manipulation (e.g. zooprophylaxis, vector control). It raises the possibility of targeting transmission hot spots with greater accuracy and concomitant reduction in control effort. The robustness of the model to simplifying assumptions is discussed.

Using the filamentous actin marker, FITC-conjugated phalloidin, the major muscle systems of adult male and female schistosomes have been examined. The body wall musculature comprises an outer sheath of circular fibres, within which there is a compact layer of short, spindle-shaped longitudinal fibres and a lattice-like arrangement of inner diagonal fibres. Within the oral sucker and acetabulum 3 fibre types, circular, radial and longitudinal can be distinguished. The wall of the oesophagus is lined by a grid-like array of circular and longitudinal fibres, whereas the walls of the intestinal caeca contain only comparably broad circular fibres. Within the female reproductive system, only circular fibres are present in the oviduct, vitelline duct and uterus. In contrast, the wall of the ootype displays closely arranged circular and longitudinal muscle fibres. Antisera to previously identified myoactive compounds (serotonin [5-hydroxytryptamine, 5-HT], neuropeptide F [Moniezia expansa] and GYIRFamide [Bdelloura candida, Dugesia tigrina]) were used as neuronal markers in a preliminary study of the spatial inter-relationships of specific nerve fibres and various muscle systems. Serotoninergic fibres innervate both suckers and also constitute a subtegumental nerve net. In males they provide innervation to the dorso-ventral muscle fibres of the gynaecophoric canal, and in females they innervate the circular and longitudinal muscle fibres of the ootype. Neuropeptide F and the FMRFamide-related peptide, GYIRFamide are both localized within nerve plexuses associated with the dorso-ventral fibres of the gynaecophoric canal, and are evident in the innervation of the ventral and oral sucker.

Canine leishmaniosis (CanL) is caused by the parasite Leishmania infantum and is a systemic disease, which can present with variable clinical signs, and clinicopathological abnormalities. Clinical manifestations can range from subclinical infection to very severe systemic disease. Leishmaniosis is categorized as a neglected tropical disease and the complex immune responses associated with Leishmania species makes therapeutic treatments and vaccine development challenging for both dogs and humans. In this review, we summarize innate and adaptive immune responses associated with L. infantum infection in dogs, and we discuss the problems associated with the disease as well as potential solutions and the future direction of required research to help control the parasite.

Since 2004 there has been a remarkable increment in our knowledge of the proteins and glycans that reside at, or are released from the surfaces of schistosomes in the mammalian host. Initial characterization of the soluble proteome permits distinctions to be made between the parasite secretome and its necrotome. The principal proteins secreted by the cercaria to gain access to the skin have been described as well as those released by migrating schistosomula. An inventory of transporters, enzymes and structural proteins has been shown to reside the tegument surface, but also immunoglobulins, complement factors and host CD44. The secreted membranocalyx that overlies the plasma membrane may contain a small number of proteins, not simply acting as physical barrier, but its lipid composition remains elusive. Analysis of worm vomitus has provided insights into blood feeding, increasing the number of known lysosomal hydrolases, and identifying a series of carrier proteins potentially involved in uptake of lipids and inorganic ions by the gut epithelium. The egg secretions that aid escape from the tissues include a mixture of MEG-2 and MEG-3 family variant proteins. The utility of identified proteins for the development of new diagnostics, and their potential as vaccines candidates is evaluated.

Several other journal supplements have documented progress made in the control of schistosomiasis in Egypt, China and Brazil, however, with more than 97% of the schistosome infections now estimated to occur in Africa, the relevance of this special issue in Parasitology cannot be overemphasized. In total, 18 articles are presented, inclusive of a lead-editorial from the WHO highlighting a seminal resolution at the 54th World Health Assembly in 2001 that advocated de-worming. Facilitated by a US$ 30 million grant from the Bill and Melinda Gates Foundation in 2002, the Schistosomiasis Control Initiative subsequently fostered implementation of large-scale schistosomiasis (and soil-transmitted helminthiasis) control programmes in six selected African countries. From 2005, CONTRAST, a European union-funded consortium, was formed to conduct multi-disciplinary research pertaining to optimisation of schistosomiasis control. Progress made in schistosomiasis control across sub-Saharan Africa since the turn of the new millennium is reviewed, shedding light on the latest findings stemming from clinical, epidemiological, molecular and social sciences research, inclusive of public health interventions with monitoring and evaluation activities. New opportunities for integrating the control of schistosomiasis and other so-called neglected tropical diseases are highlighted, but more importantly, several opportune questions that arise from it frame the remaining challenges ahead for an enduring solution.

In this review, we explore the state-of-the-art of sand fly relationships with microbiota, viruses and Leishmania, with particular emphasis on the vector immune responses. Insect-borne diseases are a major public health problem in the world. Phlebotomine sand flies are proven vectors of several aetiological agents including viruses, bacteria and the trypanosomatid Leishmania, which are responsible for diseases such as viral encephalitis, bartonellosis and leishmaniasis, respectively. All metazoans in nature coexist intimately with a community of commensal microorganisms known as microbiota. The microbiota has a fundamental role in the induction, maturation and function of the host immune system, which can modulate host protection from pathogens and infectious diseases. We briefly review viruses of public health importance present in sand flies and revisit studies done on bacterial and fungal gut contents of these vectors. We bring this information into the context of sand fly development and immune responses. We highlight the immunity mechanisms that the insect utilizes to survive the potential threats involved in these interactions and discuss the recently discovered complex interactions among microbiota, sand fly, Leishmania and virus. Additionally, some of the alternative control strategies that could benefit from the current knowledge are considered.

In the following account of the biology of Pediculus humanus, I have sought to bring together all that could be gathered from the publications of the many authors who have occupied themselves more or less with these insects, adding here and there my own observations2. I have sought throughout to give credit where it is due and have not withheld criticism where it appeared necessary.

The Xinjiang Uygur Autonomous Region, multi-ethnic province in northwestern China, is one of the most important foci of human cystic echinococcosis (CE) in the world. Two Echinococcus granulosus genotypes (G1 and G6) are known to infect the intermediate hosts in this area but, to date, the source of the human infection remains unclear. The current study aimed to genetically analyse 67 hydatid cysts removed from 47 CE patients for which epidemiological, clinical and serological data were also recorded. Mitochondrial cox 1 gene sequencing suggested that the E. granulosus G1 genotype is the major source of infection (45/47 CE patients). Nevertheless, for the first time in China, 2 patients were found with hydatid cysts of the G6 genotype. In addition, 45 E. granulosus gravid tapeworms, isolated from 13 dogs, were genotyped. The majority of adult worms (42/45) exhibited the G1 genotype, whereas 3 adult tapeworms with the G6 genotype were found in one dog, that also harboured E. granulosus tapeworms of the G1 genotype. This sympatric occurrence of G1 and G6 genotypes of E. granulosus, not only in the same area but also in the same definitive host, raises the interesting question of putative genetic recombination between these E. granulosus genotypes.

Disruption of the endocrine system has been shown to occur in wild freshwater fish populations across the globe. Effects range from subtle changes in the physiology and sexual behaviour of fish to permanently altered sexual differentiation, impairment of gonad development and/or altered fertility. A wide variety of adverse environmental conditions may induce endocrine disruption, including sub-optimal temperatures, restricted food supply, low pH, environmental pollutants, and/or parasites. Furthermore, it is conceivable that any/all of these factors could act simultaneously to cause a range of disparate or inter-related effects. Some of the strongest evidence for a link between an adverse health effect, as a consequence of endocrine disruption, and a causative agent(s) is between the condition of intersex in wild roach (Rutlius rutilus) in UK rivers and exposure to effluents from sewage treatment works. The evidence to indicate that intersex in roach (and other cyprinid fish living in these rivers) is caused by chemicals that mimic and/or disrupt hormone function/balance in treated sewage effluent is substantial. There are a few parasites that affect the endocrine system directly in fish, including the tape worm Ligula intestinalis and a few parasites from the micropsora phylum. L. intestinalis acts at the level of the hypothalamus restricting GnRH secretion (resulting in poorly developed gonads) and is one of the very few examples where an endocrine disrupting event has been shown to result in a population-level effect (reducing it). It is well established that many parasites affect the immune system and thus the most common effect of parasites on the endocrine system in fish is likely to be an indirect one.