The viability of 4 human isolates of Giardia intestinalis cysts using either the fluorogenic vital dyes fluorescein diacetate (FDA) and propidium iodide (PI) or in vitro excystation was assessed. Whereas viable cysts, as defined by in vitro excystation were present in each of the 4 isolates, cysts from only 3 of the 4 isolates took up the vital dyes. FDA consistently over-estimated cyst viability whilst PI under-estimated non-viable cysts when compared with in vitro excystation. Following in vitro excystation, both FDA and PI stained a proportion of unexcysted cysts indicating that FDA stained cysts which were incapable of excystation, whereas PI did not stain all cysts which were incapable of excystation. One human cyst isolate, which underwent in vitro excystation, could not be stained with either FDA or PI. In the absence of currently more specific fluorescent indicators of viability, PI alone could be used to determine the lower limit of nonviability in positive water-related samples, where small numbers of cysts are to be expected.

To understand the relationship between mode of transmission and virulence, we investigated 2 microsporidian parasites in Daphnia magna laboratory populations. Pleistophora intestinalis is only transmitted horizontally, while Tuzetia sp. is transmitted vertically with high efficiency from mothers to parthenogenetic male and female offspring. We were not able to transmit Tuzetia horizontally in the laboratory. Tuzetia reduces host life-time reproductive success and host survival to a much greater extent than does P. intestinalis. Tuzetia-infected hosts were rapidly outcompeted by uninfected hosts. We suspect that Tuzetia infections may persist in natural populations by an as yet undiscovered horizontal transmission. It is possible that an alternate host species may be involved. We present a mathematical model to analyse the conditions for the persistence of a parasite with perfect vertical and an additional degree of horizontal transmission. We show that horizontal and vertical transmission contribute additively to a parasite's ability to invade and persist. Since the fitness contribution of horizontal transmission increases with population size, only very low transmission probabilities per host-to-host contact are necessary for the persistence of parasites occurring in large populations such as those commonly found for Daphnia. The detection of such low rates of horizontal transmission appears unlikely under laboratory conditions since the necessary number of host-to-host (or host-to-spore) contacts is not feasible. We review mechanisms that can maintain vertically transmitted parasites in nature.

We have investigated the effects of host age and sex on human antibody isotype responses to Schistosoma mansoni and Schistosoma japonicum adult worm (AW) and soluble egg (SEA) antigens, using sera from subjects in Kenya and the Philippines. Similar trends with age were observed between the two populations despite host, parasite and environmental differences between the two geographical locations. IgE to AW increased with age, whereas most isotype responses to SEA decreased with age. IgG1, IgG3 and IgG4 subclass responses to adult worm, however, did not show a broadly rising or falling pattern with age. Males were found to have higher IgG1, IgG4 and IgE to AW in both populations. This sex difference remained significant in the Kenyan population even after controlling statistically for confounding factors such as age and differences in intensity of infection. Analysis of S. mansoni and S. japonicum adult worm antigens reactive with IgE revealed a predominant 22 kDa band in both parasites. Only those individuals with relatively high IgE titres specifically reactive with S. mansoni or S. japonicum AW had detectable IgE against Sj22 or Sm22.

This study focuses on the effect of acidic water and aqueous aluminium on the monogenean ectoparasite Gyrodactylus salaris, infecting Atlantic salmon (Salmo salar) parr. G. salaris-infected salmon were exposed to various combinations of acidity and aluminium concentrations. The most pronounced effect was the elimination of parasites after 4 days when 202 μg Al/l was added to the water. The effect of aluminium was concentration dependent, but was relatively independent of pH (5·2, 5·6 and 5·9). At the lowest pH of 5·0 the effect of aluminium was enhanced. Acidic aluminium-poor water had no or minor effects on the G. salaris infections except at pH 5·0 where all parasites were eliminated within 9 days. The G. salaris populations increased exponentially in untreated control water. The results show for the first time that aqueous aluminium can, to a limited extent, have a positive effect on fish health. This study emphasizes that basic knowledge about abiotic environmental factors is of importance in order to understand the population dynamics, range extension and dispersal of ectoparasites such as G. salaris. Finally, our results suggest that aluminium treatment could form an effective disinfection method against ectoparasites in hatcheries and laboratories, as well as complementing the controversial rotenone treatments used against natural populations of G. salaris.

Schistosoma mansoni egg counts by faecal examination vary considerably and are not very sensitive, so prevalences are underestimated. The distribution of egg counts can adequately be described by a stochastic model which distinguishes variation in counts between persons and variation in repeated counts within a person. Based on this model a pocket chart has been developed which predicts the proportion of individuals harbouring at least 1 S. mansoni worm pair – the ‘true prevalence’ – from a simple single survey prevalence and geometric mean egg count (using common duplicate 25 mg Kato–Katz smears). The current paper describes the validation of this chart by comparing predicted true prevalences with prevalences observed after 5–7 repeated Kato–Katz faecal examinations (Burundi), by examination of a large quantity of stool using the Visser filter (Brazil) or a selective sedimentation–filtration method (Surinam). Because 5–7 repeated examinations do not suffice to measure all infections, predictions have been made of the cumulative proportion positives over 5–7 surveys – the ‘approximate true prevalence’ – as well. After dividing the data into age groups, 12 different subsets were considered for validation. In all 12 cases, predicted true prevalences (or approximate true prevalences for the Burundi data) agree well with those observed. The overall agreement depends only slightly on the assumed relationship between worm numbers and mean egg counts, with a good fit for a productivity between 0·8 and 4·4 eggs per gramme faeces (EPG) per worm pair (WP). This interval includes the most plausible value from the literature, i.e. 1·0 EPG/WP, which has been applied in the initial pocket chart. These findings support the validity of the chart to predict true prevalences for a wide range of productivity assumptions, and reinforces the applicability of its underlying stochastic model to describe egg count variation. However, as predictions appear to vary importantly when using only part of the data, it is also concluded that the pocket chart never compensates for limited validity of initial single survey prevalences and geometric means in consequence of small sample sizes.

The genetic make-up of an infecting Toxoplasma gondii strain may be important for the outcome of infection and the risk of reactivation of chronic disease. In order to survey the distribution of different genotypes within an area, free-range chickens act as a good model species. In this study 85 chickens were used to investigate the prevalence, genotype and mouse virulence of T. gondii in Kampala, Uganda. Antibodies were detected in 40 chickens, of which 20 had MAT-titres of 1:20 or higher and were also positive by PCR. Genotyping of 5 loci (SAG1, SAG2, SAG3, BTUB and GRA6) showed that 6 strains belonged to genotype I, 8 to Type II and 1 to Type III. Five chickens had multiple infections; 3 individuals with Type I plus Type II and a further 2 harbouring Types I, II and III. Isolates were obtained from 9 chickens via bioassay in mice, 6 were Type II strains and 3 were from animals with mixed infection. This is the first set of African T. gondii strains to be genotyped at multiple loci and in addition to the 3 predominant lineages we found a small number of new polymorphisms and a high frequency of multiple infections.

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.

Prevalence of Toxoplasma gondii within 6 UK farmstead wild rat populations, and 1 population maintained within a captivecat-free environment for 2·5 years, was examined. The mean prevalence found was 35% (n = 235), which is more than 3 times as high as all other wild rat prevalence levels previously reported in the UK. There were no significant age, sexor site differences in prevalence between or within populations irrespective of habitat type or presence of cats. Toxoplasma was also maintained within the captive rat population in the absence of cats at a prevalence, intensity, age and sexdistribution similar to that of the farmstead rat populations. These results suggest, firstly, that Toxoplasma can beperpetuated within wild rat populations without the sympatric presence of cats and secondly, that the congenital route isthe predominant route of transmission in wild rats. This study concludes that wild rats represent a significant and persistent wildlife intermediate host reservoir for toxoplasmosis.

Temperature-dependent survival and reproduction of Gyrodactylus bullatarudis Turnbull, 1956 were examined at 17, 21, 25·5, 27·5 and 30 °C on isolated guppies under conditions where parasite density dependence and host responses were minimized. Parasites had the longest life-span (5·5 days) at 21 °C, the highest average fecundity (1·73 offspring) at 25·5 °C and the highest instantaneous per capita birth rate (0·543/parasite/day) at 27·5 °C. The intrinsic rate of increase of the parasite population was maximum (0·230/parasite/day) at 27·5 °C. Data obtained from this study indicate that G. bullatarudis are not able to survive at 30 °C.

Soil from 403 hedgerow, roadside verge, woodland, heathland or field sites was assessed for presence of entomopathogenic rhabditid nematodes with aGalleria larva baiting technique. Steinernematids were recovered from 48 % of the sites but only one site yielded Heterorhabditis sp. The species recovered most frequently was Steinernema bibionis (Bovien). Another unidentified Steinernema sp. was also isolated. Prevalence of steinernematids in the different types of habitat formed continuum so that roadside verges harboured them most often while heathland sites yielded them least often. Mean soil temperatures at 5 cm were about 15 °C in fields and verges, 13 °C in hedgerows and heathland and 12 °C in woodland. Prevalence of the nematodes varied in different parts of Britain and was influenced by soil type. They were associated with calcareous soils, especially those with a calcareous subsoil horizon. Soils which are periodically or seasonally water-logged were suitable. Soils enriched by humus and hence high in organic matter frequently harboured the nematodes and some suitable soils had significant clay contents or subsoils high in clay. It appears that the two steinernematids are relatively unspecialized species, occurring in diverse habitats and soil types and are adapted to cooler temperatures.

The present paper describes how age–intensity profiles of macroparasite burdens are affected by processes underlying the distribution of the parasite numbers in host populations. In a comparative way, we consider the following 6 processes: (i) age-dependent exposure, (ii) parasite-induced host mortality, (iii) heterogeneity within the host population, (iv) clumped infection, (v) density-dependent parasite mortality and (vi) density-dependent parasite establishment. For each of these processes, we show typical patterns in the age–intensity profile and provide, if possible, explicit and simple solutions for the age-dependent mean parasite burden and the corresponding dispersion patterns. Emphasis is given to density-dependent parasite establishment and to age–intensity profiles resulting from the superposition of different processes. By means of 2 examples we show that the interpretation of observed patterns can be ambiguous if more than 1 process takes place. These findings underline that age–intensity profiles should be interpreted on the basis of available a priori knowledge about the processes assumed to be involved. For purposes of testing different hypotheses, a simulation program is provided with which discrepancies between model prediction and data can be explored.

The malaria parasite exists in an extracellular form at several stages in its life cycle. Within the vertebrate host, sporozoites and merozoites have to invade specific cell types. Proteins on the surface of the parasite or externalized from specialized organelles have been implicated as ligands for receptors on the host cell surface. Direct binding studies have identified parasite proteins that interact with the target cell surface. Examination of the deduced amino acid sequences has allowed the identification of primary structural motifs which may have roles in this process. On the sporozoite, the circum-sporozoite protein and sporozoite surface protein-2, a protein initially located within micronemes, have been found to contain an amino acid sequence thought to be involved in mediating recognition of sulphated polysaccharides on the surface of a liver cell. On the merozoite, merozoite surface protein-1 may be involved in the initial recognition of red blood cells; this protein undergoes a complex series of modifications in the time between its synthesis as a precursor molecule and successful erythrocyte invasion. Other merozoite proteins located at the apical end of the parasite have been identified as erythrocyte or reticulocyte binding proteins.

Fixation of carbon dioxide has been demonstrated for extracts from Crithidia fasciculata, Trypanosoma mega and Trypanosoma brucei brucei bloodstream and culture forms. The enzymes involved in this fixation were found to be ADP-stimulated phosphoenolpyruvate carboxykinase (E.C. 4. 1. 1. 32), ‘malic’ enzyme (E.C. 1. 1. 138–40) and pyruvate carboxylase (E. 0. 6. 4. 1. 1). The subcellular localization of these enzymes has been investigated in all three organisms. Products of short and long term fixation experiments were separated and identified.

The importance of carboxylation reactions is discussed in relation to the maintenance of oxidized and reduced coenzyme levels.

An enzyme-linked immunosorbent assay (ELISA) was used to detect antibodies to Encephalitozoon cuniculi in man, using spores derived from tissue culture as antigen. Negligible cross-reactions were found with other microsporidia of vertebrate or of invertebrate origin and there was no cross-reaction with Toxoplasma gondii, using ELISA, immunoperoxidase or immunofluorescence staining. A high prevalence of antibodies to E. cuniculi was found in patients suffering from schistosomiasis, malaria and neurological and psychiatric disorders, but not in healthy individuals.

The fact that no inquiry seems to have been made into a matter of some general interest, namely, the reasons which, apart from “hunger,” induce mosquitoes to bite, may serve as an excuse for presenting the preliminary results of an investigation which has had, for the time being, to be discontinued. The following observations on mosquitoes were made at the Pusa Research Institute.

Ecdysteroids (moulting hormones), juvenoids and neuropeptides in ticks are reviewed but, by far, the emphasis is on the former since this class of hormones has been the subject of most investigations. In immature stages of ticks, ecdysteroids have been shown to regulate moulting and to terminate larval diapause. Although there is a paucity of information on the molecular action of ecdysteroids in ticks, their action appears to be via a heterodimeric ecdysone/ultraspiracle receptor, as in insects. The role of ecdysteroids in sperm maturation in adult males is considered. In females, ecdysteroids function in the regulation of salivary glands, of production of sex pheromones and of oogenesis and oviposition. There is evidence for ecdysteroid production in the integument and pathways of hormone inactivation are similar to those in insects. Ecdysteroids also function in embryogenesis. Although evidence for the occurrence and functioning of juvenile hormones in ticks has been contradictory, in recent thorough work it has not been possible to detect known juvenile hormones in ticks, nor to demonstrate effects of extracts on insects. Factors (neuropeptides) from the synganglion affect physiological processes and limited immunocytochemical studies are reviewed. Sigificantly, a G-protein-coupled receptor has been cloned, expressed, and specifically responds to myokinins.

Plasmodium ookinetes are elongate, motile and invasive while inside the mosquito gut but promptly metamorphose into spherical immobile oocysts upon coming in contact with the basement membrane surrounding the midgut. There they begin a prolonged growth period characterized by massive DNA synthesis for the production of sporozoites. Living Plasmodium gallinaceum ookinetes attached avidly to the murine extracellular matrix proteins, laminin and collagen type IV. In ELISA-type assays, the main ookinete surface protein, Pgs28 was implicated as a mediator of parasite attachment to these basement membrane constituents. Laminin and collagen IV adhered to ookinete and oocyst lysates spotted onto nitrocellulose membranes. Receptor–ligand blot assays demonstrated that Pgs28 and an oocyst-specific antigen recognized by the mAb 10D6 interact with murine collagen IV and laminin. 10D6 antigen was also recognized by monospecific antiserum against the human epidermal growth factor receptor. Mosquito-derived laminin was incorporated into oocyst capsules of P. gallinaceum growing in Aedes aegypti. We hypothesize that contact with the mosquito basement membrane triggers the transformation of ookinetes into oocysts. Coalescence of basement membrane proteins onto the capsules masks developing oocysts from the mosquito's immune system and facilitates their prolonged extracellular development in the mosquito body cavity.

The parasitic mite, Varroa destructor, is the most important threat for apiculture in most bee-keeping areas of the world. The mite is carried to the bee brood cell, where it reproduces, by a nurse bee; therefore the selection of the bee stage by the parasite could influence its reproductive success. This study investigates the role of the cuticular hydrocarbons of the European honeybee (Apis mellifera) in host-selection by the mite. Preliminary laboratory bioassays confirmed the preference of the varroa mite for nurse bees over pollen foragers. GC-MS analysis of nurse and pollen bees revealed differences in the cuticular hydrocarbons of the two stages; in particular, it appeared that pollen bees have more (Z)-8-heptadecene than nurse bees. Laboratory experiments showed that treatment of nurse bees with 100 ng of the pure compound makes them repellent to the varroa mite. These results suggest that the mite can exploit the differences in the cuticular composition of its host for a refined selection that allows it to reach a brood cell and start reproduction. The biological activity of the alkene encourages further investigations for the development of novel control techniques based on this compound.

The environmental route of transmission is important for many protozoan and helminth parasites, with water, soil and food being particularly significant. Both the potential for producing large numbers of transmissive stages and their environmental robustness (with the ability to survive in moist microclimates for prolonged periods of time) pose persistent threats to public and veterinary health. Increased demands made on natural resources increase the likelihood of encountering environments and produce contaminated with parasites. In the last 30 years, endemic and epidemic waterborne and foodborne outbreaks in developed countries have led to a reappraisal of conventional isolation and detection methods. While these methods have proved invaluable in our understanding of environmental transmission routes for helminths, they have been less effective for the parasitic protozoa. Robust, efficient detection, viability and typing methods are required to assess risk and to further epidemiological understanding. Greater awareness of parasite contamination of our environment and its impact on health has precipitated the development of better detection methods. Currently, nowhere is this more apparent than with Cryptosporidium, with a broad range of immunological, microscopical and molecular methods available. The upsurge in molecular techniques, particularly the polymerase chain reaction, for determining occurrence and viability have brought with them the added benefits of increased sensitivity and specificity, yet many methods still have to be shown to address these issues consistently in the field. Rapid commercialization of reagents and standardization of methods provide consistency. The advances identified in non-destructive and destructive methods for the protozoa have application for helminths and emerging pathogens and should determine the importance of the matrices involved in the environmental transmission of parasites, further safeguarding public and veterinary health.

The relationship between varying intensities of Trichuris trichiura infection and iron status was examined in Jamaican schoolchildren, aged 7 to 11 years. A total of 409 children was identified with T. trichiura (epg > 1200). A control group comprised 207 uninfected children who were matched by school and class to every pair of infected subjects. Blood samples were obtained from 421 children: 264 infected and 157 controls. Compared to the rest of the children, those with heavy infections (epg > 10 000) had significantly lower (P < 0·05) Hb (11·5 ± 1·3 vs. 12·1 ± 1·1 g/dl), MCV (78·6 ± 6·3 vs. 81·2 ± 5·5 fl), MCH (26·2 ± 2·9 vs. 27·5 ± 2·5 pg) and MCHC (33·2 ± 1·5 vs. 33·9 ± 1·4 g/dl). Similarly, the prevalence of anaemia (Hb < 11·0 g/dl) amongst heavily infected children (33%) was significantly higher (P < 0·05) than the rest of the sample (11%). These differences remained significant after controlling for confounding variables including socio-economic status, age, gender, area of residence and the presence of Ascaris infections. Differences in red cell count, ferritin, and free erythrocyte protoporphyrin were not statistically significant and showed no association with the infectious load. These results suggest that in the Jamaican children studied, iron deficiency anaemia is associated with Trichuris infections over 10 000 epg, but not with less intense infections.