Monoclonal antibodies reacting with pellicular antigens of Toxoplasma gondii tachyzoites have been selected among hybridomas produced against this organism by immunofluorescence assay. These antigens have been further characterized by immunofluorescence on living zoites, Western immunoblotting and immunoprecipitation of lactoperoxidase surface radio-iodinated tachyzoite lysates. The simultaneous characterization of 5 different surface antigens (P43, P35, P30, P23, P22) some of which have already been studied individually allowed a better definition of these antigens and the characterization of a yet undescribed surface molecule (P23).

Monte Carlo simulations were developed to assess the potential impact of parasite pathology on vector salivary function as well as of host haemostasis on transmission. Assuming that a proportion of desisting vectors switch host following failure to locate blood, we demonstrate three possible consequences: (1) infected vectors contact more hosts than non-infected hosts, (2) non-infected vectors are biased to infected hosts, independently of attraction, and (3) an exponential relationship exists between parasite load and transmission. We discuss possible epidemiological implications.

Doses from 1 to 10 sporulated oocysts of a cloned strain of Eimeria hungaryensis were used to infect laboratory-reared, coccidia-free wood mice (Apodemus sylvaticus). Total oocyst production, reproductive indices (output/dose) and daily patterns of oocyst production were compared. At doses of 10 oocysts or less, observed yields exceeded expected yields calculated using details of endogenous stages. At doses above 10 oocysts, the converse occurred. From infections initiated with 10 oocysts, total outputs were not significantly different in male and female wood mice, host age had no apparent effect and repeated infections, at 10-day intervals, of both sexes, resulted in progressive decreases in output at each successive infection, suggesting development of resistance.

An acoustic actograph was used to monitor for 17 days after infection the spontaneous flight activity of the mosquito Anopheles stephensi parasitized with the rodent malaria Plasmodium yoelii. Activity fell to approximately two-thirds of control levels at about day 10 post-infection – when oocysts were reaching maximum size and starting to rupture (mean number of oocysts = 92) – and thereafter remained at this reduced level. The circadian activity pattern was not affected by the parasitism.

The DNA coding for parts of the repetitive amino acid sequence of the Plasmodium falciparum circumsporozoite protein has been spliced to a sequence encoding part of the precursor to the major merozoite surface antigens, to produce a hybrid gene. Expression in Escherichia coli produces a protein with antigenic determinants from both malaria proteins. Antibodies raised against the expressed material react with both a peptide derived from the circumsporozoite repeat sequence, and the merozoite surface molecule. Hybrid molecules of this type may be the basis of a malaria vaccine.

The fate of 75Se-labelled parasites and their released pre-synthesized macromolecules has been followed in three murine infection models. Parasite numbers in specific tissues were determined by autoradiography, and released material was estimated by gamma-counting of tissues, with adjustment for the presence of parasite-associated radiolabel. Marked differences were found between the three models. The pattern of migration of normal schistosomula was similar to that previously reported. In addition we have described the transit of parasites through the lymph nodes draining the infection site. Significant quantities of released material were detected in the skin, draining lymph nodes, bloodstream and liver. The circulating material was of parasite origin, macromolecular, and hence potentially antigenic. In comparison to the normal infection, radiation-attenuated parasites (inducing a high level of resistance to challenge) persisted in the skin, draining lymph nodes and lungs, releasing a proportionally greater amount of material in the nodes. In mice exposed to attenuated parasites and treated with the compound ROl1–3128 at 24 h (inducing a low level of resistance) there was an early death and rapid clearance of the parasites whilst still in the skin. This situation resulted in the highest levels of released material in the skin, bloodstream and liver, but negligible levels in the draining lymph nodes. We suggest that the persistence of radiation-attenuated parasites in the skin and draining lymph nodes, together with the prolonged release of antigen in the latter site, compared to the normal situation, are major factors in the induction of resistance.

Attempts were made to immunize goats by infection with large numbers of metacyclic trypanosomes of a clone of Trypanosoma vivax, followed by chemotherapy. Five groups of 6 goats each were infected intradermally with 5 different doses of cultured metacyclics of T. vivax, ranging from 102 to 106 trypanosomes/goat. Four weeks after infection, the goats were treated with 10 mg/kg diminazene aceturate (Berenil, Hoechst A.G.). Three weeks after treatment, 3 goats in each group were challenged intradermally with 104 homologous metacyclics derived from culture. The remaining 3 goats in each group were challenged by 20 tsetse infected with the homologous clone. Five out of 30 goats were resistant to homologous challenge; 4 of the goats that had been challenged with cultured parasites, and 1 that had been challenged by tsetse. In each group 1 goat was protected. Protection was therefore not apparently influenced by the number of trypanosomes used to establish the primary infection. In another experiment, 6 goats were each infected by feeding 100 tsetse on the goats for 15 consecutive days. Three weeks after infection the goats were treated with Berenil and 3 weeks later challenged by 20 tsetse infected with the homologous clone. Three out of the 6 goats resisted challenge. The susceptible goats in both experiments, however, showed a reduction in the peak of parasitaemia following challenge compared with both challenge controls and the initial infections. Lytic antibodies to cultured metacyclics of T. vivax were detected in goats that resisted challenge after a primary infection with cultured metacyclics, and in resistant and susceptible goats after a primary infection by tsetse. All infected goats produced lytic antibodies to live bloodstream forms, as well as antibodies to bloodstream form lysates (demonstrated by ELISA). It is suggested that the immunity that had been induced in some of the experimental animals is due to antibody responses to both metacyclic and bloodstream variable antigen types (VATs) expressed during infection.

Cystosporogenes operophterae, a microsporidian parasite of the winter moth, Operophera brumata, was cultured in a Spodoptera frugiperda cell line. The infected cultures were achieved by addition of haemolymph from larvae infected with C. operophterae. The infection rates for the cell line ranged from 80 to 95% after incubation for 10 days at 25 or 20 °C. There was no development in cultures incubated at 30 °C. Light and electron microscopy were used to observe the development of C. operophterae in vitro.

The development of granulomatous reactions against moulting nymphal pentastomids (Porocephalus crotali) in the tissues of rat and mouse intermediate hosts is described. Adipose tissue and lungs are favoured sites for encystment accounting for 70% of larvae. Six moults separate the primary larva from the final infective stage which first appears about 80 days post-infection (p.i.) and is fully infective by day 120. Larvae, and particularly their cast cuticles, are the foci of granulomatous reactions characterized by an intense eosinophilia. During ecdysis, large numbers of eosinophils permeate the entire lesion but, significantly, degranulation is limited to the underside of cast cuticles where the resultant debris is endocytosed by macrophage/epithelioid cells. A pronounced asymmetry in the granulomatous lesion, evident even in the earliest cysts, results from the accumulation of individual epithelioid granulomas associated with cuticle fragments close to the ventral side of the developing parasite; each is circumscribed by fibrosis. External to this region are extensive tracts of tissue composed of mature plasma cells. Particularly in rats, large numbers of partially degranulated mast cells ( = globule leucocytes) also surround cuticle granulomas, and mast cell granules can accumulate within macrophages and fibroblasts. Inflammation slowly subsides once the infective stage is attained. This 1 cm-long larva resides in a thin, fibrotic, C-shaped cyst and can remain viable for years: uniquely this instar retains its last moulted cuticle as a protective sheath. Nymphal instars II-VI feed predominantly upon eosinophils but we do not yet know whether this requirement is obligate.

A steer was infected with Theileria parva parva Kilae stabilate; nymphal Rhipicephalus appendiculatus ticks were applied to its ears so that they completed repletion when the steer had a high piroplasm parasitaemia. The engorged nymphs were subsequently incubated at 28°C for 26–29 days to complete moulting, when the adult ticks were divided into two groups; one was incubated at 18°C for 20 days and the other at 18°C for 14 days and then at 37°C for 6 days. Groups of ticks incubated at 37 and 18°C were triturated and each resultant supernatant fluid inoculated into a steer. Both steers became infected, but the 37°C supernatant group showed a much shorter pre-patent period to schizonts. Groups of ticks incubated at 37 or 18°C were applied to pairs of cattle for 24, 48 and 72 h and then removed. There was a more rapid transmission of theileriosis to cattle by ticks kept at high ambient temperatures compared to those kept at low ambient temperatures. All cattle on which ticks treated at 37°C were applied developed acute and fatal T. parva infection irrespective of the duration of tick application, while only 1 animal receiving ticks treated at 18°C and fed for 72 h developed infection. The pre-patent period for macroschizonts was very short in all the groups receiving ticks incubated at 37°C. Mature sporozoites were detected in the salivary glands of unfed ticks incubated at 37°C and after they were applied to cattle for 24, 48 and 72 h but in those incubated at 18°C sporozoites were detected, only at 48 and 72 h after application.

Two species of Schizotrypanum, T. (S.) dionisii and T. (S.) vespertilionis, were identified from British bats. Laboratory studies on stocks of isolated trypanosomes from 5 species of bat (Pipistrellus pipistrellus, Nyctalus leisleri, N. noctula, Eptesicus serotinus and Myotis brandti) indicated that the predominant species was T. d. dionisii. Collections and dissection of the bat bug Cimex pipistrelli from bat roosts revealed flagellate infection in a total of 12 out of 20 bugs; 7 of these bugs had metacyclic trypanosomes present. C. pipistrelli and the human bed bug, C. lectularius were reared in the laboratory and allowed to feed on wild-caught bats known to be infected with T. d. dionisii. Development occurred in both species of Cimex. Cimex spp. could be used to detect subpatent Schizotrypanum infections by xenodiagnosis. This technique was used to test the parasitological status of bats collected in the wild or reared in captivity. On a single occasion an apparent transmission of T. d. dionisii to an uninfected (by xenodiagnosis) laboratory reared bat was achieved. A stock of Schizotrypanum isolated from a wild-caught C. pipistrelli collected in a N. leisteri roost was identified by DNA buoyant density centrifugation as T. (S.) vespertilionis. A P. pipistrellus known to be infected with T. d. dionisii was found to have cyst-like structures in thoracic skeletal muscle containing amastigotes. The study provided the strongest evidence yet that C. pipistrelli is the vector of Schizotrypanum in British bats.

CBA/Ca mice infected with the tetrathyridia of Mesocestoides corti were exposed to Schistosoma mansoni cercariae either simultaneously, or at varying intervals after the initial M. corti infection. Cohorts were infected with either parasite alone. The dual infected mice and the mice harbouring M. corti alone were significantly heavier than either naive controls or mice carrying the S. mansoni infection only. The livers and spleens from dual infection mice were also found to be significantly heavier than those from other groups. Free M. corti tetrathyridia were reduced in number in the peritoneal cavity of dual infected mice, as compared with mice harbouring a single infection. Furthermore, the intensity of the initial M. corti infection, as measured by the number of capsules surrounding parasites in the liver, was reduced when the mice experienced an S. mansoni infection 21 days later. The mice which were exposed to M. corti only exhibited more mast cells and eosinophils around encapsulated tetrathyridia in the liver than did dual infection mice, while cells surrounding S. mansoni egg granulomas in the liver were significantly increased in dual infection mice. An increase in serum alkaline phosphatase levels was detected in both the mice receiving the dual infection and the mice with the S. mansoni only infection, but not in mice harbouring M. corti alone.

Shared antigens between Brugia malayi and Aedes aegypti were studied. The experiments carried out with sera from infected Mastomys natalensis indicated that an immunological response against A. aegypti antigens (Mr 185, 35, 32 kDa) appeared often when animals became microfilaraemic and increased progressively in intensity during the time-course of infection. Sera of animals immunized with B. malayi reacted with the crude extract of mosquitoes and conversely, antibodies from animals immunized with A. aegypti reacted with the surface of B. malayi microfilariae. The implications of these findings for the natural history of B. malayi infection are discussed.

The host—parasite relationships of two geographical isolates of Schistosoma haematobium in CBA mice are described and compared to previous reports on this parasite in other experimental hosts and in man. The mean percentage establishment of worms in mice was 17% and was not affected by the age or sex of the host. Adult worm burdens remained constant over 20 weeks, but were reduced after 18 months of infection. Male and female worms reached mean maximum lengths of 4·78 and 5·9 mm respectively. Egg laying commenced 9·5 weeks after infection and eggs accumulated in the tissues throughout the period of infection. A large increase in the rate of egg accumulation occurred coincidental with the appearance of eggs in the bladder of some mice. Faecal eggs were first observed in some mice at 12·5 weeks and most mice excreted a few eggs by 17 weeks p.i. (post-infection). Eggs were not found in the urine of infected mice. Excreted eggs and eggs isolated from the livers of infected mice hatched, but the resulting miracidia were unable to infect appropriate snail hosts. The development of hepatic granulomas and egg-induced pathology in the bladder of mice is described.

Relapse of parasitaemia after drug treatment of trypanosome infections is normally attributed to drug-resistance on the part of the parasite, under-dosage of the drug or reinfection of the host. In addition, inaccessibility of parasites to drug through sequestration in privileged extravascular sites has been shown in the past to occur with Trypanosoma brucei, and we have obtained evidence that extravascular foci of T. vivax can also serve as a source of relapsing infections. Infection of goats with a West African stock of T. vivax resulted in severe illness, which was fatal if untreated. During the terminal stage of an acute infection, clinical signs of central nervous system involvement were apparent. Histologically, the choroid plexus was swollen and oedematous, and in some cases meningitis or meningoencephalitis was seen. Trypanosomes could be detected in the cerebrospinal fluid, and also extravascularly in the choroid plexus and meninges. In three cases they were present in the aqueous humor, associated with corneal cloudiness or opacity. Treatment of 2 goats with the trypanocidal drug diminazene aceturate eliminated parasitaemia, but infections in both relapsed about 6 weeks later, despite trypanosomes being undetectable in the bloodstream during the intervening period. We conclude that the relapse infections were caused by re-emergence of trypanosomes from the CNS and/or the eye, where sequestered parasites may have been inaccessible to the trypanocide.

We have shown here that collagen type I bound efficiently to the trypomastigote surface. In addition, monoclonal and polyclonal antibodies against collagen types I and III inhibited the infection of fibroblasts by the parasite. These results suggested the presence of collagen-binding protein(s) on the parasite surface. This protein was identified from trypomastigote surface antigens using affinity chromatography on a Gelatin Ultrogel column (denatured form of collagen). These collagen-binding proteins were revealed as a low-affinity gelatin binding protein (LAG Bp) of 98 kDa, and a high-affinity binding protein (HAG Bp) of 58 and 68 kDa under non-reducing and reducing conditions respectively. In addition, HAG Bp and LAG Bp bound to collagen type I. The 58/68 kDa protein was purified to homogeneity on a wheat germ agglutinin Sepharose column. A polyclonal antibody to this glycoprotein, as well as a monoclonal antibody (McAb) 155D3 produced against the HAG Bp, immunoprecipitated two parasite surface antigens of 160 and 58 kDa under non-reducing conditions which migrated at a position of 80–85 and 68 kDa when reduced. However, only the 80–85 kDa component could be precipitated from [S] methionine-labelled trypomastigote antigens under reducing conditions. The antibodies to the 58/68 kDa glycoprotein as well as McAb 155D3 diminished the invasion of fibroblasts by parasites. Taken together these results suggest that the same receptor binds fibronectin and/or collagen and that both the 80–85 and 58/68 kDa glycoproteins form part of the same receptor. These trypomastigote surface molecules may interact with the host cell fibronectin and/or collagen during the initial phase of parasite-cell recognition.

We have obtained 5 specific DNA probes for African trypanosomes of the subgenera Trypanozoon and Nannomonas. Each probe consists of one repeat unit of the major repetitive DNA (satellite DNA) of each species or intra-specific group. One probe hybridized with all members of subgenus Trypanozoon (except T. equiperdum which was not tested). In subgenus Nannomonas, one probe recognized T. simiae, but 3 probes were needed to identify all stocks of T. congolense available. Each of the 3 latter probes recognized trypanosomes from one of the 3 major groups of T. congolense previously defined by isoenzyme characterization, i.e. savannah, forest and Kenya coast types. As few as 100 trypanosomes could be unequivocally identified by dot blot hybridization and individual trypanosomes could be identified by in situ hybridization. We show how this simple methodology can be used in the field for the identification of immature and mature trypanosome infections in tsetse.

A preparation highly enriched in tegument membranes from adult worms has been used to probe rodent responses to schistosome surface antigens. All exposure regimes in mice stimulated levels of anti-tegument IgG, measurable by ELISA. Plateau levels of circulating antibody attained were similar after mixed-sex and male-only infections, and vaccination with tegument membranes. Female-only infections and exposure to irradiated cercariae stimulated lower levels of anti-tegument antibody. Responses in rats were similar, but occurred more rapidly, probably due to the large initial parasite exposure. Although both mice and rats could be stimulated to produce high levels of anti-tegument antibody by multiple vaccination, consistent protection against a challenge was not obtained. Paradoxically, rats and mice vaccinated with irradiated cercariae developed low levels of antibody to tegument membrane antigens but were highly resistant to challenge. It was concluded that, in the range of sera examined, antibody responses to schistosome tegument membranes did not correlate with the immune status of the host.

Transmission of the monogenean Pseudodiplorchis americanus is restricted to the brief period when its host, the desert toad Scaphiopus couchii enters temporary water to breed. This study followed parasite recruitment into one host population in Arizona, USA, during the spawning season (July). Torrential rainfall triggered 3 successive assemblies. Amongst males, which may enter each assembly, the 3 pulses of invasion led to an approximate doubling of mean worm burdens at each exposure, culminating in 100% prevalence and a mean intensity of over 100 larvae/host. Females, which generally enter only one assembly, acquired a mean of about 40 larvae/host. Each exposure is limited to a maximum of 7 h by strictly nocturnal host activity, and the force of infection increased exponentially: around 10% of recruitment occurred in the first 3 h, and more than 30% in the last 1 h. Correlation of recruitment into males with parasite reproductive potential suggests that individual oncomiracidia had a 30% chance of invasion. Although invasion of the desert host relies on an aquatic infective stage, host behaviour ensures that targets are concentrated in space and time, and the parasite's annual reproductive output is synchronized with a period of host vulnerability totalling less than 24 h/year.

We have determined the molar content of guanine + cytosine (GC content) of DNA of the filarial nematode (Brugia malayi, Brugia pahangi and Dirofilaria imitis) and of the free-living soil nematodes Caenorhabditis elegans and have analysed the DNA for the presence of methylcytosine. Two independent methods, thermal denaturation and direct analysis of base content by HPLC following enzymatic hydrolysis, reveal that the GC content of filarial nematodes is 26–28%. We have been unable to find methylcytosine in the DNA of B. malayi.