The literature contains many examples of changes induced by parasites in the behaviour and/or other phenotypic traits of insects. From an evolutionary perspective, the nature of these changes is usually difficult to assess. Parasite-induced changes in host behaviour can be adaptations of either host or parasite, or they can be mere pathological consequences of infection. Of the many criteria and experimental tests necessary to distinguish between adaptations and non-selected consequences, two are particularly important: the demonstration of fitness benefits for either host or parasite associated with the behavioural change, and the elucidation of the proximate mechanism responsible for the behavioural change. Another approach can serve to identify adaptive changes in behaviour: mapping specific behavioural alterations on a phylogeny of either hosts or parasites. The usefulness of this approach is illustrated with two examples, acanthocephalan- cockroach associations and insect-fungus associations. The adaptive nature of parasite-induced behavioural changes will always be difficult to evaluate because they are the product of two distinct but interacting genotypes. However, experimental and phylogenetic approaches can provide valuable insights into the evolutionary history of insect-parasite interactions.

A female-specifically expressed cDNA clone was obtained by screening of a subtractive cDNA library enriched for RNA from Schistosoma mansoni females. The deduced protein shows significant homology to a class of enzymes functioning as amidases. Northern blots revealed a transcript of 4·0 kb which is absent in larval stages, but is expressed in adult female worms. By in situ hybridization, the expression site of the gene was exclusively localized in the gastrodermis of female schistosomes. This is the first report of a female-specifically transcribed sequence of S. mansoni that is not expressed in the reproductive organs.

Twenty-eight isolates of E. granulosus, collected from humans at surgery, and a range of intermediate hosts, including sheep, cattle and camels from abattoirs in North and South Xinjiang Uygur Autonomous Region, People's Republic of China, were analysed for DNA sequence variation within regions of the mitochondrial cytochrome c oxidase I (COI) and NADH dehydrogenase subunit I (NDI) genes. The isolates were categorized into 2 distinct and uniform genotypic groupings, based on the sequences obtained, and the data clearly indicated that the camel/dog strain (G6 genotype) of E. granulosus as well as the cosmopolitan, common sheep strain (G1 genotype) occur in north Xinjiang. The presence of the camel strain has thus been confirmed in Xinjiang but it is evident from this and a previous molecular genetic survey of E. granulosus isolates from north-western China that the common sheep strain is the most predominant in the region. From the public health perspective, the majority of infected livestock will act as reservoirs of human infection there. During the course of the study, a polymerase chain reaction (PCR)-restriction fragment length polymorphism (RFLP) assay, based on the NDI sequence variation, was developed that allows rapid discrimination of the G1 and G6 genotypes.

Theileria annulata (Ta)-infected leucocytes are able to disseminate in scid mice. The dose of virulent parasites of the Ta-Ode line required to achieve quantifiable dissemination was found to be 2×10 cells given i.p. Dissemination was higher on day 11 post-inoculation than on day 18. The attenuated Ta-Ode cells were found to disseminate very poorly compared to their virulent progenitors, which correlates with a marked reduction in matrix metalloproteinase (MMP) expression. A daily i.p. injection of mice with BB94, a synthetic inhibitor of MMPs, almost completely ablated dissemination compared to controls. This provides strong evidence that metastasis of Theileria annulata macroschizont-infected host cells is mediated by host MMPs induced by the parasite. This has important implications for explaining a number of pathological features of tropical theileriosis in cattle.

Host fertility is often curtailed as a result of parasitic infection. The hypothesis that this may confer an adaptive advantage upon the symbionts if nutrients are directed from reproduction and made available for host/parasite maintenance is explored. The suggestion is made that an understanding of the mechanisms underlying the pathophysiology of fecundity reduction may shed light upon the evolutionary implications of this strategy for both parasite and host. To illustrate this the down-regulation of egg production is explored with reference to a particular model system, the association between metacestodes of the rat tapeworm, Hymenolepis diminuta and the mealworm beetle, Tenebrio molitor. Several aspects of host reproductive behaviour and physiology are affected by infection in this association, including vitellogenesis. Metacestodes directly inhibit the fat body synthesis of vitellogenin in a stage-specific, density-dependent manner. This inhibition is likely to be orchestrated by a modulator molecule, produced by the parasite. In the ovarian follicles, juvenile hormone III binding to a specific follicular membrane-binding protein is inhibited in infected beetles, resulting in the down-regulation of a cascade of events which enables vitellogenin to pass into the developing oocyte. Data to support the proposed existence of a parasite-induced antigonadotrophin, of host origin, are discussed. Evidence that similar mechanisms operate in Plasmodium-iniected anopheline mosquitoes and Onchocerca-infected blackflies is presented in support of the possibility that a parasite-induced reduction in host reproductive fitness is an adaptive strategy and an assessment of who is manipulating whom is made.

The Zn2+-chelating metalloprotease inhibitor 1,10-phenanthroline (phenanthroline, 5–150 μM) elicited dose-dependent contraction of the longitudinal and circular (transverse) musculature of adult male schistosomes. At the same concentrations, phenanthroline did not cause contraction of dispersed individual muscle fibres. The phenanthroline-induced contractions were reduced by the inclusion of 100 or 300 μM Zn2+ in the extracellular medium. Phenanthroline (0·5–150 μM) also inhibited the egg production of adult worm pairs in vitro, with a 98% reduction at 50 μM. When worm pairs were exposed to phenanthroline, the males detached from the dish and released the females, resulting in unpaired worms. At the higher concentrations (50 and 150 μM), the worms were killed in vitro. Worm burdens were reduced by over 50% in infected mice injected with phenanthroline (20 mg/kg/day for 4 days), but twice the dose resulted in only a 25% reduction. Phenanthroline injections also induced an hepatic shift and an unpairing of adult worms in infected mice, and the female worms appeared degenerate and lacked gut pigmentation. Mice fed a diet containing 0·3% phenanthroline received significant protection from infection when challenged with schistosome cercaria, where phenanthroline-fed mice had 94% fewer adult worms than control mice. The broad range of phenanthroline effects on schistosomes suggests broad and important functions for metalloproteases in these worms.

An aspartic proteinase precursor, herein named BYC (Boophilus Yolk pro-Cathepsin) was isolated from eggs of the hard tick, Boophilus microplus. As judged by electrophoresis on sodium dodecyl sulfate polyacrylamide slab gel (SDS–PAGE), purified BYC presented 2 bands of 54 and 49 kDa, bearing the same NH2-terminal amino acid sequence. By Western blot analysis, BYC was also found in the haemolymph, indicating an extraovarian site of synthesis. Several organs were incubated in culture medium with [35S] methionine, and only the gut and fat body showed synthesis of BYC polypeptides. Protein sequencing of both the NH2-terminal and an internal sequence obtained after cyanogen bromide (CNBr) cleavage of BYC revealed homology with several aspartic proteinase precursors. Incubation at pH 3·5 resulted in autoproteolysis of BYC, which produced the mature form of the enzyme, that displayed pepstatin-sensitive hydrolytic activity against haemoglobin. Western blot analysis using anti-BYC monoclonal antibodies showed proteolytic processing of BYC during embryogenesis and suggested activation of the enzyme during development. A role of BYC in degradation of vitellin, the major yolk protein of tick eggs, is discussed.

We investigated the influence of infection by the trematode Curtuteria australis on the burrowing behaviour of its intermediate host, the bivalve Austrovenus stutchburyi. Laboratory experiments and field observations revealed that cockles, unable to bury completely or even partially under the sediment, have a reduced foot length compared with buried individuals. The ability to bury proved to be highly repeatable in field experiments: cockles found at the surface and transplanted to an experimental area did not bury themselves, and cockles found buried stayed buried when relocated. All metacercariae of C. australis were found strictly in the foot and for each of 3 samples collected in different sites, there was a negative and significant relationship between the relative length of the foot and the parasite load. A predation test conducted under natural conditions indicated that cockles with the stunted foot and the altered behaviour are significantly more susceptible to predation by aquatic birds than other cockles. Given that the definitive host of C. australis is an oystercatcher, we first discuss our results in the context of transmission strategy. Comparisons with other studies on more or less related trematode species parasitic in bivalves and evolving under similar constraints for their transmission, shed light on the origin of this adaptation in C. australis.

Light and electron microscopy were used to study the ultrastructure of the cement apparatus, namely cement glands and cement ducts of mature specimens of the acanthocephalan parasite Pomphorhynchus laevis Müller, 1776, recovered from the digestive tracts of fish Leuciscus cephalus Risso, 1826. In addition, the cement glands of immature P. laevis found within the body cavity of the fish Alburnus alburnus alborella De Filippi, 1844 were examined. In a mature male of P. laevis the 6 cement glands are rounded to oval in shape and each of them has an outer cytoplasmic layer containing nuclei and surrounding a space for storage of the cement material within the gland. The nuclei have an irregular outline and the cytoplasm of the cells contains round, membrane-bound secretory granules approximately 1 μm in diameter. Nuclei surrounded by secretory granules were present inside the gland lumen. Within the gland ducts of mature males, granules were present within the wall thickness and, inside the luminal area, mitochondria were encountered. In contrast, within the cement glands of immature P. laevis there were no secretory granules and the chromatin of the nuclei appeared condensed. The nature of the secretory product of the cement glands was investigated with histological and electrophoretic methods. A protein with molecular weight of 23 kDa was recorded as a major component of cement.

In the savanna areas of tropical Africa, cattle are frequently infected with the filaria Onchocerca ochengi. This parasite is closely related to Onchocerca volvulus, the causative agent of human onchocerciasis (river blindness), and is capable of developing in the same vector, Simulium damnosum s.l. In North Cameroon, where both O. ochengi and O. volvulus are endemic, we carried out a field study (reported in this and 2 following papers) to examine to which extent the transmission of the 2 parasite species overlap and what influence this has on the epidemiology of human onchocerciasis. In this paper we report our experiments to determine which of the S. damnosum species in North Cameroon act as vectors of O. ochengi, how efficiently they do so and whether other Simulium species play a vector role. To this end, infected cattle were exposed near 5 rivers in different geographical areas. Among 14 Simulium species identified as aquatic and/or adult stages at these rivers, only 6 (S. squamosum, S. damnosum s.s., S. sirbanum, S. bovis, S. wellmanni and S. hargreavesi) were found to bite cattle in important numbers in at least 1 of the sites. The 3 species of the S. damnosum complex were all capable of ingesting microfilariae (mf) of O. ochengi and developing a proportion of them to infective larvae (L3). Whereas S. squamosum and S. damnosum s.s., the prevailing vectors in the Guinea and Sudan savanna respectively, showed a high vector competence (17% of ingested mf developed to L3), S. sirbanum, which was much rarer in both areas, appeared to have a much lower susceptibility (2%). Other boophilic Simulium species were only seen in certain sites and seasons, being either incapable of ingesting important numbers of O. ochengi mf from body regions where these mf were abundant (S. bovis, S. hargreavesi); not able to support the development of ingested mf to L3 (S. wellmanni), or bit cattle preferentially in the ears, where O. ochengi mf do not occur (S. hargreavesi). We conclude that in North Cameroon members of the S. damnosum complex are the only important vectors of O. ochengi, with S. squamosum and S. damnosum s.s. being the main vectors.

Transmission electron microscopy was used to study Toxoplasma gondii infections in the small intestines of Swiss-Webster mice at 2–48 h post-feeding of oocysts (p.f.). Sporozoites passed through intestinal epithelial cells (enterocytes and goblet cells) and infected all cells except red blood cells in the lamina propria. Parasites in intestinal epithelial cells or in cells in the lamina propria were located within a single type of parasitophorous vacuole, which contained exocytosed electron-dense material and well-developed tubulovesicular membranous networks. Sporozoites did not infect intraepithelial lymphocytes (IELs), but at 48 h p.f. IELs had become infected with tachyzoites arising from those that had developed in the lamina propria. At 48 h p.f., the lamina propria contained numerous tachyzoites, much cellular debris, and few intact cells. The intestinal epithelium exhibited limited cytopathological changes except for villar fusion, slight vacuolation, and cell separation at the bases of enterocytes.

A fluorescent dye monochlorobimane (MCB) that binds glutathione (GSH) was used as a tool for measuring the concentration of GSH in skin and mechanically-transformed schistosomula. The specificity of MCB binding to GSH was confirmed by thin layer chromatography (TLC) and high performance liquid chromatography (HPLC). The MCB binding to GSH is an energy-dependent process since no labelling could be seen at low temperature. When 24-h-old schistosomula were depleted of GSH by buthionine sulfoximine (a specific inhibitor of GSH synthesis) for 18 h, a significant decrease (P<0·001) in fluorescence was observed. PZQ treatment of the schistosomula after first labelling the parasites with MCB did not greatly affect MCB binding to GSH. However, when the 24-h-old schistosomula were first PZQ treated and afterwards labelled with MCB, the pattern of labelling was identical to that of those of the non-labelled parasites. When 24-h-old schistosomula were first PZQ treated, washed and labelled in the presence of 1 mM GSH, the level of fluorescence was recovered. These results suggest that PZQ depletes GSH from schistosomula, and may render them susceptible to the host's immune system.

The effect of trypanosome infection on vector survival was observed in a line of Glossina morsitans morsitans selected for susceptibility to trypanosome infection. The differential effects of midgut and salivary gland infections on survival were examined by exposing flies to infection with either Trypanosoma congolense which colonizes midgut and mouthparts or Trypanosoma brucei rhodesiense which colonizes midgut and salivary glands. A comparison of the survival distributions of uninfected flies with those exposed to infection showed that salivary gland infection significantly reduces tsetse survival; midgut infection had little or no effect on the survival of tsetse. The significance of these findings is discussed in relation to the vectorial capacity of wild flies.

Schistosome cercariae of the genus Trichobilharzia are the causative agent of swimmers' itch. In order to characterize the changes in parasites during and after the penetration of the host skin, in vitro and in vivo (in ducks and mice) transformations of T. szidati cercariae to schistosomula were performed. Ultrastructural observation revealed that cercariae possess a simple outer tegumental membrane with a thick glycocalyx. As with human schistosomes, the latter structure disappears during transformation and a new double membrane with putative protective function is formed. Our biochemical and immunological observations showed that the carbohydrate-rich glycocalyx of cercariae is readily bound by lectins and antibodies. The in vitro transformation to schistosomula can be detected by enhanced reactivity of 2 lectin probes (PNA and ConA) with the surface. The in vivo-transformed (skin and lung) schistosomula appear to have few surface ligands for the 12 lectin probes being tested. Similarly, the cercarial surface and its remnants on the in vitro-produced schistosomula is recognized by sera from immunized mice and humans with cercarial dermatitis; the tissue schistosomula fail to react with these antibodies. The loss of surface targets as a part of parasite immune evasion within the host is discussed.

Saliva of blood-feeding arthropods promotes infection by the vector-borne pathogens they transmit. To investigate this phenomenon in vitro, cultures of mouse L cells were treated with a salivary gland extract (SGE) prepared from feeding ticks and then infected with vesicular stomatitis virus (VSV). At low input doses of VSV, viral yield was increased 100-fold to 10000-fold by 16–23 h post-infection compared with untreated cultures, and depending on the SGE concentration. SGE-mediated acceleration of viral yield corresponded with the earlier appearance of VSV nucleocapsid protein as detected by 2-dimensional electrophoresis of infected cells. The observation that physiological doses of virus (i.e. doses likely to be inoculated by an infected arthropod vector into its vertebrate host during blood-feeding) respond to SGE treatment in vitro provides a new opportunity for identifying the factors in tick saliva that promote virus transmission in vivo.

Host-parasitoid interactions are abundant in nature and offer great scope for the study of coevolution. A particularly fertile area is the interaction between internal feeding parasitoids and their hosts. Hosts have evolved a variety of means of combating parasitoids, in particular cellular encapsulation, while parasitoids have evolved a wide range of countermeasures. Studies of the evolution of host resistance and parasitoid virulence are reviewed, with an emphasis on work involving Drosophila and its parasitoids. Genetic variation in both traits has been demonstrated using isofemale line and artificial selection techniques. Recent studies have investigated the fitness costs of maintaining the ability to resist parasitoids, the comparative fitness of flies that have successfully defended themselves against parasitoids, and the degree to which resistance and virulence act against one or more species of host or parasitoid. A number of studies have examined geographical patterns, and sought to look for local adaptation; or have compared the traits across a range of species. Finally, the physiological and genetic basis of change in resistance and virulence is being investigated. While concentrating on Drosophila, the limited amount of work on different systems is reviewed, and other possible areas of coevolution in host-parasitoid interactions are briefly discussed.

Epidemiological models are used to analyse 8 published data sets reporting age–prevalence curves for trypanosome infections of the tsetse fly Glossina pallidipes. A model assuming a fixed maturation period and a rate of infection which is independent of fly age is adequate for Trypanosoma vivax-type infections, explaining 98% of observed variance in prevalence by site and age, allowing that the rate of infection may be site dependent. This model is not adequate for T. congolense-type infections and the fit can be improved by allowing (i) the rates of infection to decline with age (although non-teneral flies remain susceptible), (ii) a fraction of resistant flies, which may vary between sites, (iii) increased mortality of infected flies and (iv) variation in the maturation period. Models with these features can explain up to 97% of observed variance. Parameter estimates from published experimental data suggest that all may contribute in practice but that (i) and/or (ii) are likely to be the most important.

The competence of 7 different stocks of Rhipicephalus appendiculatus and R. zambeziensis to transmit 2 different stocks of Theileria parva was compared by feeding nymphae of each tick stock simultaneously on infected cattle and assessing the infections in the salivary glands of the resultant adult ticks. There were significant differences in the patterns of infection of the 2 stocks (T. parva Muguga and T. parva Boleni) in the different stocks of ticks, and these differences were shown to be reproducible. The Muguga tick stock from Kenya and the Zambia tick stock from Eastern Province had the highest infections of T. parva Muguga and T. parva Boleni respectively. The Zambia Southern Province tick stock and the Zimbabwe Mashonaland West tick stock had the lowest infections of T. parva Muguga and T. parva Boleni respectively. The difference in mean abundance of infection between the most and least efficient vector for T. parva Muguga was 63·3 while that for T. parva Boleni was 54·4 infected acini. The implications of these results for laboratory transmission of T. parva and for the epidemiology of theileriosis are discussed.

In order to assess the prevalence of the cattle filaria Onchocerca ochengi in onchocerciasis vectors (Simulium damnosum s.l.) in North Cameroon, we searched for a means to morphologically identify its developing larvae, which closely resemble those of O. volvulus. To this end microfilariae of the 2 Onchocerca species were isolated from slaughter cattle in Ngaoundéré and injected into neonate Simulium species. Whereas the early developmental stages (sausage stage, L2 and pre-infective larva) were indistinguishable, the infective larvae (L3) of O. ochengi were longer (median: 740 μm), more slender (diameter = 19·3 μm = 2·6% of body length) and had a relatively shorter tail (4·9% of body length) than those of O. volvulus (680 μm, 20·5 μm, 3·0% and 5·8% respectively). The tail of O. ochengi L3 was thick and rounded, whereas it was slightly tapering in O. volvulus L3. O. ochengi L3 produced by feeding flies on infected cattle in a different area in North Cameroon (Sora Mboum) showed the same features as intrathoracically produced O. ochengi L3 from Ngaoundéré, but were even longer (785 μm). On the basis of the differences in length, relative diameter, length of the tail and shape of the tail, a simple key for the separation of O. volvulus and O. ochengi L3 was elaborated, and 248 L3 found in wild S. damnosum s.l. were separated into ‘O. ochengi’ (160 L3) and ‘O. volvulus’ (88 L3) following this key. Sequential dot blot hybridization of each of the 248 larvae with a DNA probe which reacts with O. ochengi and O. volvulus but not with other Onchocerca species (pOo5/1) and with an O. volvulus-specific DNA probe (pOv12) revealed that the morphological identification had been correct in 86–91% of the cases. Only a small proportion (6–9%) of the dot blots did not react with either probe. Since this proportion was equal in experiments using experimentally produced L3 and in experiments using wild L3, the non-hybridization was certainly due to a loss of L3 during washing of the filters and not due to the presence of other unknown L3 species resembling O. volvulus and O. ochengi. Our study shows that in Cameroon it is possible to identify O. volvulus and O. ochengi infective larvae during routine fly dissections by morphology alone.

The presence of naturally portacaval shunts has been investigated in the vasculature of normal and Schistosoma mansoni-infected Rattus rattus. Using the technique of injecting Polystyrene microspheres in the superior mesenteric vein, we demonstrated that the presence of adult schistosomes in the lungs of R. rattus was not due to an innate anomaly of the rat vasculature but resulted from the formation of portacaval shunts during infection. In rats harbouring a bisexual infection, microspheres were only detected in the lungs from week 7. The development and increasing size of the shunts were maximal between weeks 7 and 10 and coincident with the translocation of adult worms from the portal tract to the lungs. At weeks 20–25, only 1–2% of the microspheres were recovered from the lungs, suggesting that the portacaval anastomoses have regressed due to reduction in portal hypertension after worm translocation. R. rattus with a male-only schistosome infection harboured adult worms in the lungs, indicating that the development of shunts does not solely depend upon egg deposition in the liver to generate hypertension. The relationships between the presence of the schistosomes in the lungs, the portacaval shunting and the resistance to reinfection is discussed.