Various factors that may influence routine and high levels of mosquito infection with cultured Plasmodium falciparum gametocytes are considered in this paper. One of the most important is the choice of an appropriate isolate, with facilities for cryopreservation and a good technique for initiation of cultures. The use of automated culture systems with strict adherence to detail and routine has eliminated much of the variability. The quality of the serum used for the culture of gametocytes and inclusion in the feed material for mosquitoes is of the highest importance. Blood collection for culture purposes must preferably involve alcohol as an antiseptic for cleaning donor skin or suitable receptacles. Mosquito blood meals should not include plasma with citrate phosphate dextrose or sera collected in microtainer tubes or from volunteers on proguanil-chloroquine prophylaxis. Sera of individuals on chloroquine alone do not influence transmission. Haemato-crits of from 5 to 10% permit the culture of equally infective gametocytes. It was impossible to predict the outcome of an infection in mosquitoes based on the number of female gametocytes or gametes. Within any experiment, the oocyst load initially increased, followed by a decline with progressively lower numbers of gametocytes accompanied by a progressive increase in the efficiency of transmission. Some of the variability of mosquito infection within an experiment was due to individual differences in the speed of blood digestion of the mosquitoes. A new membrane feeder is described with three different sizes to accommodate a variety of goals. The new design permitted more mosquitoes to feed within a limited period compared to the older more widely used feeder of equivalent capacity, and mosquitoes feeding on the new-type feeder were frequently better infected than those fed on the older type.

Over the past few years several experienced groups studying malaria have encountered significant problems with their particular rodent malaria–host system. This has involved, in some cases, periods during which the recovery of cryo-preserved parasite stocks and growth of bloodstream parasites was markedly inhibited and, in other cases, periods of drastically increased mortality rates. The common factor linking these incidents was that they coincided with alterations in the experimental animal diet used. in the experimental animal diet used. The inhibition of growth of cryopreserved stabilates or bloodstream parasites was abolished by supplementation with p-aminobenzoic acid (PABA) or by changing the diet used. Although the suppressive effects of diets lacking PABA on parasite growth have been known for over 30 years, the variation of PABA levels in modern laboratory animal feed concentrates is not well recognized. We have not established the exact cause of increased mortality, but it has been overcome by changing the diet used. We are documenting our experiences with this potential variable to forewarn workers in other laboratories of possible problems inherent in the use of different diets.

The characteristics of intra-lymphocytic Theileria isolated from African buffalo and from cattle that were infected with buffalo-derived parasites were evaluated using anti-schizont monoclonal antibodies (mAbs) and DNA probes. Antigenic differences were revealed by the reactivities of 27 mAbs with the buffalo-derived parasites isolated from different animals. Antigenic diversity was also seen with Theileria-infected lymphoblastoid cell isolates taken from the lymph nodes and blood of the same animals. Two DNA probes, selected from a genomic library of T. parva piroplasm DNA cloned in λgt11, showed specific hybridization to parasite DNA in Southern blots of restriction enzyme-digested, lymphoblastoid cells infected with buffalo-derived theilerial parasites. Genotypic differences between the buffalo-derived parasites were revealed by the restriction fragment length polymorphisms seen with hybridization of those probes to DNA from cloned and uncloned Theileria-infected cell lines. The evaluation of theilerial parasites derived from buffalo and from cattle which underwent typical T. p. lawrencei reactions, after being infected with buffalo-derived theilerial parasites, did not show any specific phenotypic or genotypic characteristics of these parasites that would distinguish them from T. p. parva and T. p. bovis parasites. The validity of these subspecies distinctions is discussed.

Parasites with low host specificity are predicted to search for hosts in ways less specific than highly host-specific species. They may also be better dispersers. Unionicolan mites that parasitize mussels are positively phototactic, with some species becoming negatively phototactic when host substances are present. This response has been interpreted as an aid to host location and occurs in species where adult mites have a relatively permanent association with hosts. Three species of Unionicola (Unionicolidae: Acari) that differ in host specificity co-occur in freshwater mussels (Unionidae) in St Mark's River in north Florida.

During 1987 a total of 1393 cattle was examined for Schistosoma nasale infection at the Kandy slaughterhouse, Sri Lanka. The overall prevalence was 12·6% (monthly range 3–17%). Based on the appearance of macroscopic lesions, 6 types (0–5) were recognized; type 5 being the most severe, with cauliflower-like growths obstructing the nasal cavity. Older bovines with 8 permanent incisors were more heavily infected (29·%) than younger ones with no permanent incisors (6·0%). The severity of the lesions increased also with the age of the animals. Observations on the localization of the lesions showed that the first sessile nodular areas appear on the medial septum, on the dorsal edge of the ventral nasal concha and on the lateral wall of the middle meatus of the nasal cavity. Later, they gradually spread over the whole mucosal surface of the anterior part of the cavity but were rarely found further than 10 cm posterior to the nasal opening. The histopathology showed that granuloma formation due to the presence of eggs was the most common feature of the respiratory mucosa.

Mice infected heavily with Schistosoma mansoni cercariae were treated 6 days later with Praziquantel and the parasites studied 1 h post-treatment. Immunofluorescence experiments showed that parasite surface antigens became available for labelling in larvae harvested from Praziquantel-treated mice, but not from untreated mice. Red blood cell antigens acquired from the host were localized on all lung worms studied. The distribution of each set of antigens, as revealed by fluorescence-labelling was, however, quite different. The lungs of heavily infected, drug-treated mice, exhibited severe haemorrhages which occurred in a dose-dependent manner. Fewer haemorrhages were found when heavily infected mice were treated simultaneously with Praziquantel and Aprotinin, a proteinase inhibitor. These results indicate that Praziquantel treatment in vivo induces the exposure of parasite antigens on lung-stage worms and may cause the release of parasite-derived enzymes which induce haemorrhages in the lung tissue of the host.

Sixteen monoclonal antibodies (MABs) were selected for their reactivity with adult schistosome tegument. The distribution of target epitopes in different tissues of the adult and in various developmental stages was investigated by indirect immunofluorescence. The distinct patterns of reactivity of the MABs permitted their classification into 9 groups. The distribution of epitopes in larvae, particularly 3 h schistosomula, generally mirrored that in adults. A change in distribution of epitopes coincided with transformation from cercaria to schistosomulum providing a marker for this process. Two MABs reacted with surface membranes of intact cercariae, and 5 with the surfaces of intact 3 h schistosomula. The target epitopes of these 5 MABs were present in the tegument of adults, but not accessible externally. These observations suggest masking of antigens, not shedding, in the course of development. Indeed, no MAB reacted with intact lung or liver worms unless they were damaged. Three MABs reacted with membranes of the tegument and most other tissues, implying properties in common. Evidence for shared functions between epithelia was provided by the reactivity of 2 MABs with the tegument and protonephridia. Eight MABs bound both to the tegument and its cell bodies. Since the latter contains the machinery for protein synthesis, it is difficult to explain why 5 MABs bound to the tegument alone. Little cross-reactivity was found with the tissues of the tapeworm Hymenolepis, rather more with the closely related Schistosoma japonicum. It was concluded that tegument antigens are to a degree tissue specific but definitely not stage specific.

The localization and distribution of neuropeptides in the nervous system of the liver fluke, Fasciola hepatica at different stages in the development of the adult fluke have been determined by an indirect immunofluorescence technique, using antisera to 19 vertebrate peptides and the invertebrate neuropeptide, FMRFamide. Positive immunoreactivity was obtained with antisera to pancreatic polypeptide (PP), peptide tyrosine tyrosine (PYY), substance P (SP) and FMRFamide. Cell bodies and nerve fibres immunoreactive to the 4 peptides are present in the anterior ganglia and the 3 pairs of longitudinal nerve cords and their commissures in the central nervous system. In the peripheral nervous system, immunoreactivity occurs in the nerve plexuses supplying the subtegumental musculature, the oral and ventral suckers, and the muscular lining of the male and female reproductive ducts, including the ootype, uterus, cirrus pouch and gonopore. Cells displaying immunoreactivity to PYY and FMRFamide lie amongst the Mehlis' gland cells that surround the ootype. Processes from these cells extend into the wall of the ootype. One group of PP-immunoreactive cells occurs at the junction of the vitelline and ovovitelline ducts, whilst another group is situated at the entrance to the uterus from the ootype. The results are discussed in relation to the possible roles of the peptides in the neurophysiology and egg production of the fluke.

The effects of pharmacological concentrations of several gastrointestinal hormones on the rate of sucker activity and the frequency and the amplitude of spontaneous longitudinal muscle contractions have been examined in adult Fasciola hepatica. Caerulein and serum decrease the rate of oral sucker activity; motilin decreases and CCK–PZ increases ventral sucker activity when compared to controls. Caerulein, serum and motilin significantly inhibit the frequency of contractions while bile, caerulein and motilin decrease the amplitude of contractions. These results suggest that F. hepatica can recognize and respond to certain gastrointestinal hormones and there may be adaptive value in these behavioural responses.

Mating was observed in populations of Gyrodactylus turnbulli from guppies. Copulation took place between older (post-1st birth) flukes and resulted in the insemination of spermatozoa into the seminal receptacle. New-born flukes, and flukes which had only recently given birth for the first time (in which the penis was absent or developing), were never inseminated. Flukes were promiscuous, copulating with several partners in succession. The occurrence of mating was related to the size, age and crowding of the G. turnbulli population, taking place most frequently when the parasites were most densely crowded upon the peduncle of the fish on days 5 and 6 of the infection. Maximum population size was positively correlated with the number of flukes initiating an infection. As a result, mating began earlier, and continued for longer in populations founded by several parasites rather than those begun by single flukes. These observations are discussed with respect to the population biology of G. turnbulli: it is likely that sexual and asexual reproduction alternate, with sex being important only during phases of epidemic population growth. The significance of sex in gyrodactylids is discussed, and the potential importance of continuous sexual reproduction in G. turnbulli infections on cultivated guppies is noted.

Trypanosoma musculi are readily killed when phagocytosed by mononuclear phagocytes but the nature of the mediators of this cytotoxicity is unclear. Among the most potent mediators are oxygen-derived species. The generation of chemilumine-scence (CL) by peritoneal macrophages from 12 day T. musculi-infected mice, which phagocytose and kill parasites when opsonizing antibodies are present, was recorded in the presence of antibody-coated trypanosomes. Taurine, a specific quencher of hypochlorous acid (HOCl) inhibited CL production by peritoneal macrophages, showing that HOCl is produced during phagocytosis of T. musculi. In vitro, HOCl alone exerted a powerful trypanocidal activity which was inhibited in the presence of specific quenchers. The role of HOCl generated by phagocytes in trypanosome killing was studied using granulocytes which produce more oxygen-derived species than macrophages when stimulated. Phorbol myristate acetate-triggered granulocytes can destroy T. musculi and trypanosome killing is inhibited in the presence of taurine. These data demonstrate that HOC1 produced by phagocytes can effectively destroy T. musculi.

An in vitro method for studying radiorespiration has been adapted to single macrofilariae. Using this method viable (but not heat-killed) Dipetalonema viteae and Onchocera gibsoni macrofilariae evolved measurable amounts of 14CO2 from L-[U-14C]glutamine. Nonlinear and less uniform rates of 14CO2 evolution were demonstrated with D-[U-14C]glucose, [1-14C]acetate and [1-14C]octanoate. These findings led us to develop an in vitro screen in which inhibition of 14CO2 evolution from L-[U-14C]glutamine was used as a parameter for gauging macrofilaricidal activity. Using this screen we have examined the activity of 17 ‘so-called’ antifilarial standards and found a greater degree of sensitivity than shown by other biochemical criteria. Other data obtained suggest a role for radiorespirometry in determining the viability of fragments of Onchocerca tissue.

All polyopisthocotylean monogeneans previously studied, including representatives of the Polystomatidae infecting anuran amphibians, feed on host blood. However, the present analysis of species of Polystomoides, Polystomoidella and Neopolystoma, polystomatids which infect chelonian reptiles, has shown that this group has diverged nutritionally from related parasites. Histochemical tests failed to demonstrate haemoglobin in the gut caeca, and X-ray microanalysis confirmed the absence of haematin (or high concentrations of bound iron) in the gastrodermis. The chelonian poly-stomatids (and also the single monogenean which infects a mammal, Oculotrema hippopotami) feed on epithelial cells and mucus, the diet typical of monopisthocotyleans. Transmission electron microscopy revealed the same gastrodermal architecture in representatives of Polystomoides from Africa, N. America and S.E. Asia. The organization of the caecal epithelium conforms with that of blood-feeding polyopisthocotyleans, with two components: lamellated cells responsible for intracellular digestion interspersed with elements of a non-lamellated connecting syncytium. In other polyopistho-cotyleans, the syncytium probably has a skeletal, supportive role, related to the problems of intracellular accumulation of haematin, but in polystomatids infecting chelonians the syncytium is extremely reduced and its presence probably reflects an ancestry amongst blood-feeding relatives. The utilization of the presumably more primitive monogenean diet of epithelial cells and mucus by chelonian polystomatids may be related to the scarcity of superficial blood vessels in their oral and urinary bladder habitats.

Predisposition to trichuriasis in mice is reflected in the inability of certain strains, or certain individuals within strains, to express protective immunity. Poor responders fail to expel worms and harbour chronic patent infections. The mechanisms underlying this phenomenon were studied in poor responder mice challenged after abbreviated or prolonged primary infections. Mice exposed to a complete primary infection were fully susceptible when challenged after the removal of the primary infection by anthelmintic. Failure to expel either infection suggests (a) that non-responsiveness to a primary infection does not reflect an inability to expel worms of a certain size, i.e. is not a consequence of the speed of the immune response in relation to parasite growth and (b) that non-responsiveness is long-lasting. Challenge after abbreviation of primary infections at different stages of worm development showed that persistence of larvae beyond day 21 was critical in determining poor response to reinfection. By inference the same conclusion can be drawn about the inability of such mice to expel primary infections. Serological analysis suggested a relationship between low antibody titres, restricted antigen recognition profiles and resistance to infection. It is suggested that the later stages of parasite development are immunosuppressive; the implications for human trichuriasis are discussed.

The course of primary infection with a hamster-adapted strain of Ancylostoma ceylanicum was studied in inbred DSN and randomly bred WO/GD and WO/CR hamsters. Infective larvae were administered orally and began to develop in the small intestine without embarking on a tissue migration. Only the occasional larva was detected in other organ sites. It was concluded that the developing larvae moulted on days 3–4 and again to the pre-adult stage about 9–11 days post-infection. Worm burdens in infected hamsters were stable for at least 11 weeks after infection. There was no sudden expulsive phase and some adult worms survived for over 200 days. Overall the sex ratio of worms in groups of hamsters killed concurrently was about 50% although occasionally the ratio was biased in favour of one sex in individual animals. The blood packed cell volume (PCV) was significantly depressed 2 weeks following infection and continued to decline until a point of stability was achieved 4–5 weeks post-infection. The PCV subsequently remained depressed throughout the period of observation. Infected hamsters lost weight if kept in groups, but not when housed in separate cages. Groups of animals which lost weight did not recover to normal values within 11 weeks of infection. It is suggested that this model of hookworm infection has scope for exploring aspects of the host-parasite relationship which the canine models cannot fulfill adequately.

The in vivo activity of two cyclosporins, cyclosporin A (CsA) and a non-immunosuppressive derivative of dihydrocylosporin A (DHCsA-d) against three tapeworms, Hymenolepis microstoma, H. diminuta and Mesocestoides corti, has been assessed. CsA reversibly reduced the dry weight of H. microstoma in the mouse, briefly delayed oviposition and had a statistically significant effect on worm numbers recovered. Oral and subcutaneous treatments of both CsA and DHCsA-d were effective in reducing worm weight; juvenile worms were most susceptible but worms of all ages responded to drug by a dramatic reduction in weight from which they recovered. Multiple courses of CsA were no more active than single courses of treatment but dose response suggested that a threshold level of drug was necessary to evoke activity. By contrast, H. diminuta in the rat was completely unaffected by CsA but no explanation for the differences in drug response by these two closely related helminths is forthcoming. Mesocestoides corti responded reversibly to CsA in the mouse by a reduction in asexual proliferation of both liver and peritoneal cavity tetrathyridia. The data presented argue in favour of a range of anti-parasitic activities by cyclosporins but the details of the various putative modes of action remain to be defined.

The activity of the gluconeogenic enzyme, alanine-amino-transferase (ALT), in a preparation from the liver of rats was studied by means of an in vitro assay throughout the course of a primary infection of Nippostrongylus brasiliensis, established by a subcutaneous injection of approximately 4000 3rd-stage larvae. The activity was measured on days 1–14 p.i. in both uninfected and infected rats and a marked pattern in the enzyme's activity was observed. In infected rats, the activity increased from 1·46±0·19 U/g liver on day 1 p.i. to a peak on day 4 p.i. of 10·75±1·62 U/g liver, then decreased to a trough of 0·44±0·18 U/g liver on day 10 p.i. before returning to original levels by day 14 p.i., by which time the infection had been largely eliminated. In uninfected rats the activity of the liver enzyme remained constant throughout this period with a value of 2·54±0·12 U/g liver. The activity of the enzyme in vitro was found to be related to the size of the inoculum on days 4 and 10 p.i. It was proposed that these observations could be due to either (1) a direct effect of the parasite, or (2) a consequence of the host immune response to the infection. In order to investigate the second proposition more fully, liver ALT activity was investigated by in vitro assay on selected days p.i. in rats experiencing a secondary N. brasiliensis infection. In these rats the liver ALT activity was observed to reach a peak on day 2 p.i., with an activity of 3·87 ± 0-28 U/g liver, and a trough on day 4 p.i. with an activity of 0·11 ±0·03 U/g liver, returning to similar levels to those measured in uninfected rats by day 7 p.i. When serum prepared from rats having secondary N. brasiliensis infections collected on day 4 p.i. was added to the assay, a reduction in the activity of liver ALT activity from both the infected and uninfected rats was measured by in vitro assay. The results are discussed in relation to protein metabolism and gluconeogenesis in rats infected with N. brasiliensis, and also in relation to the host’s immune response to the parasitic infection.

The results of 4 different assays for haemocytic immune responsiveness in the cockroach, Periplaneta americana, have shown that the presence of larvae of Moniliformis moniliformis within the haemocoele depresses the insect's immune reactivity. Thus, the proportion of haemocytes that phagocytose fluorescent latex beads in vivo, the proportion of haemocytes that can be stimulated in vitro by the β1,3-glucan, laminarin, to produce phenoloxidase, and the number of haemocytic aggregates produced in vivo in response to zymosan stimulation, are significantly depressed compared with control, unparasitized cockroaches. Also, when cockroaches are injected intra-haemocoelically with hatched oncospheres of the tapeworm, Hymenolepis diminuta, a higher prevalence and intensity of tapeworm larvae are found in insects already parasitized by M. moniliformis. All of these assays show that depression rather than total suppression of the immune response occurs; in the wild, this may be sufficient to ensure that the cockroach does not succumb to unrelated infections during the long developmental period of the parasite.

Resistance developed by the immune system in response to blood-stage malaria is complex in nature, involving humoral and non-antibody effector mechnisms. Different species of malarial parasites may vary in their ability to elicit, or their susceptibility to, those immune effectors. This complexity is enhanced by the different results obtained in vaccinated as opposed to drug-controlled infections. It is therefore important that some attempt be made to unravel these interactions. This is particularly so when we have to decide on methods for assessing the potentiality of antigens to induce protective immunity. In this review the limitations of some in vitro assays of immunity, as well as those of various host–parasite models, are discussed. The relative importance of cell-mediated and humoral immunity in laboratory models and natural infections is also considered in the context of vaccine development.