Plasmodium falciparum was grown in vitro in blood taken from naturally infected Gambian patients, and the development of the cultured sexual parasites was studied by light and electron microscopy. The young (Stage II and III) female gametocytes undergo a single cryptomitotic nuclear division. This division immediately follows the S phase which Sinden & Smalley (1979) have demonstrated in the Stage I and II gametocytes of both sexes. The male gametocytes, by contrast, do not undergo mitosis during their maturation period in the erythrocyte and thus remain polyploid. Hence the cell cycles of the male and female gametocytes differ significantly. The ultrastructural basis of the characteristic changes in shape of the developing gametocyte are shown to be due to the assembly and subsequent loss of components of the sub-pellicular membranous and microtubular cytoskeleton. Stage I–III gametocytes synthesize numerous ribosomes and endoplasmic reticulum. This correlates with the active synthesis of RNA and protein in these young parasites (Sinden & Smalley, 1979). The marked reduction in macromolecular synthesis in the mature parasites is paralleled by a reduction in cytoplasmic ribosome density in the male gametocyte only. In the female, however, this reduction in activity is correlated with the appearance of a nucleolus. These changes suggest that different mechanisms are being used to control RNA synthesis in the two sexes of gametocyte.

An electron microscopical study was made on the development of Eimeria dispersa in the small intestine of the domestic turkey. Turkey poults, 10–14 days of age, were inoculated with oocysts and pieces of intestinal tissue were fixed at intervals between 3·5 and 114 h after inoculation. Sporozoites were occasionally seen in enterocytes but more often in ‘pale’ cells closely resembling lymphocytes. These cells were insinuated between enterocytes and in this study are referred to as intestinal intra-epithelial leucocytes (IEL). Maturation of the first-generation of schizogony occurred in this type of cell. Other generations of schizonts and gametogony occurred in epithelial cells and were characterized by the presence of ‘spines’ arranged around the parasitophorous vacuole extending into the cytoplasm of the host cell. A limited study of the parasite in the Bobwhite quail, Colinis virginianus, showed that development occurred in similar cells to those of the turkey. The ‘spines’, which were characteristic of later stages of E. dispersa in the turkey, were also present in this host and seemed to be a characteristic of E. dispersa, not of the host species.

The Houghton (H) strain of Eimeria acervulina was attenuated by serial passage through chickens of the first oocysts produced during infection. This selection pressure resulted in a reduction in the pre-patent period of the parasite, shown to be due to the selection of a line predominantly with only 3 instead of 4 generations of schizonts. The precocious line had a reproductive potential much lower than that of the parent strain and it was significantly less pathogenic. Chickens given oocysts of the precocious line were almost completely immune to challenge with the Houghton strain.

Eimeria funduli, a coccidium infecting the liver and pancreas of killifishes and requiring an intermediate host, was allowed to develop at 22±2 °C for 5, 10, 15 or 20 days (schizont through zygote stages) in Fundulus grandis and was then exposed to 10·0±0·5 °C for 20 days or to 7·0±0·5 °C for 5 or 20 days. Both low temperature treatments inhibited all developmental stages, except for the case of fish infected for 5 days before exposure to 7 °C in which infections were eliminated. Returning fish back to a temperature of 22 °C for 20 days resulted in resumed development of all stages at the same or somewhat reduced rate. In all cases, intensity of infections was greatly reduced during exposure to low temperatures. Many organisms exhibited abnormalities. Gamonts and zygotes were atrophied or disintegrated within their parasitophorous vacuoles in all exposures and, in fish exposed to 7 °C for 20 but not 5 days, about 30% of the oocysts were abnormal, with from 1–8 sporoblasts. Areas of degeneration or necrosis incorporated aggregates and single parasites in both the pancreas and the hepatic parenchyma. The granulocytic inflammatory response, which at 22 °C was normally restricted to the period spanning development of gamonts and formation of the oocyst wall, was inhibited or delayed from infiltrating infected, diseased areas at low temperatures. The response intensified after cold-exposed fish were returned to 22 °C. A fibroid, non-granulocytic response occurred in fish introduced 5 days post-infection to 10°C before being returned to warm water. Two other types of lesion occurred in the pancreas. The first could develop into necrosis and was associated with low temperature; it did not always involve infected tissue. The other, a degenerating focal alteration, was not necessarily associated with infections or low temperatures.

Regular samples made on the 1978 cohort of brook sticklebacks (Culaea inconstans) from a swamp in Ile Perrot, Quebec, Canada were examined for Apatemon gracilis metacercariae. The prevalence of the parasite rapidly reached 100% in the fish population. The mean parasite burden increased from zero to a plateau of about 44 parasites/fish. Over-dispersion of the frequency distribution of parasites in the fish host, as measured by variance to mean ratios, increased to a peak and then decreased significantly while the mean parasite burden remained constant. The effects of parasite burden on the survival of the stickleback host are discussed, as well as the validity of the use of changes in over-dispersion for demonstrating parasite-induced mortality.

The processes which govern the age structure of Fasciola hepatica populations in sheep were investigated by means of a simple mathematical model. The mortality of the metacercariae on the pasture was shown to be a factor of major importance. The mortality rate depends on the micro-climate that prevails at the pasture surface, and thus the age distribution curve of a population of flukes is also determined in part by the weather. This has important implications for the chemotherapy of fascioliasis: flukicides are not equally effective over the whole range of age-classes of parasite and so the factors which govern the age distribution curve of the parasite population within individual hosts ultimately determine the efficiency of chemotherapy.

The cirrus of a monogenean Diplectanum aequans was isolated by treatment with sodium carbonate solution and studied with a scanning electron microscope. The method may be used in studies of the functional morphology and taxonomy of other organisms.

Monoclonal antibodies have been produced after fusion of NS-1 murine myeloma cells with spleen cells from mice immunized either by chronic primary infection or with irradiated cercariae: in both cases, animals were challenged with live cercariae 7 days before fusion. The initial cultures were screened for anti-schistosomular antibodies both by a radioimmunoassay with whole schistosomulum extracts and by immunofluorescence. There was no correlation between the two techniques and subsequent screening was carried out by immunofluorescence. Cloning was carried out in soft agar and 7 cloned cell lines, from 5 initial cultures, were selected for detailed study. Products of 6 of these 7 lines were monoclonal, as judged by isoelectricfocusing of [35S]methionine-labelled supernatant fluids, and their binding to live schistosomula was specific. None of the antibodies showed detectable activity in mediating eosinophil- or complement-dependent damage to schistosomula in vitro. However, 2 antibodies were successfully used to isolate surface proteins with an apparent molecular weight of 24000 on SDS-polyacrylamide gel electrophoresis.

Two hybridomas secreting monoclonal IgM antibody to Schistosoma mansoni have been isolated following fusion of spleen cells from Balb/c mice immunized with living S. mansoni and NS1 myeloma cells. One monoclonal IgM antibody (WP66.4) mediated about the same level of passive protection against a challenge infection as immune serum from mice with a chronic S. mansoni infection. The other monoclonal antibody (WP66.2) did not give a significant level of passive protection. This result indicates that the effective monoclonal antibody recognizes an antigen which may be a valuable candidate for experimental vaccination. In vitro one monoclonal antibody (WP66.4) caused a much higher level of complement-dependent cytotoxicity than the other (WP66.2), suggesting a possible mechanism for the effect observed in vivo. With indirect immunofluorescence both monoclonal antibodies reacted with surface determinants on living S. mansoni schistosomula, adult worms and miracidia but these determinants were not detected on cercariae or lung schistosomula. Neither monoclonal antibody cross-reacted with S. haematobium schistosomula or Fasciola hepatica metacercariae, indicating a possible use for these reagents in differential diagnosis of S. mansoni infections.

Mice infected with 25 to 35 unsexed Schistosoma mansoni cercariae became resistant to homologous challenge by 8 weeks, and the degree of resistance that was acquired correlated with the size of the worm burden and the tissue egg burden. Mice given 100 male or 100 female cercariae alone failed to become resistant even after 30–40 weeks of infection. When a super-infection of 100 male or 100–300 female cercariae was given to mice with 25 unsexed cercariae, the degree of resistance that was acquired was generally lower than in mice given 25 unsexed cercariae alone. In the super-infected mice there was no correlation between the size of the worm burden (mainly unpaired male or female worms) and the degree of acquired resistance. In mice super-infected with male worms there was an inverse correlation between the number of male worms and the number of eggs deposited in the liver and intestine by each mature worm pair. A positive correlation was, however, still found between the degree of acquired resistance and total tissue egg counts in mice super-infected with single sex cercariae. Mice injected intraperitoneally or subcutaneously with intact S. mansoni eggs failed to become resistant to S. mansoni challenge. Transfer of lymphoid cells or serum, separately or together, from heavily infected and demonstrably resistant donor mice to naive normal or T-cell deprived recipients failed to render the recipients resistant to S. mansoni challenge. The results indicate that resistance to re-infection in mice with recently patent S. mansoni infections is dependent on the number of S. mansoni eggs entrapped in the tissues, and that excessive numbers of unpaired S. mansoni male or female worms can inhibit the fecundity of established worm pairs. The apparent acquired resistance to S. mansoni that has been observed in this model system may be largely a consequence of non-specific factors interfering with normal patterns of challenge worm migration and maturation.

These studies were aimed at giving a clearer idea of the source of the antigens responsible for the induction of resistance by larval schistosomes. Firstly, the resistance induced by radiation-attenuated (20 krad.) infections was compared with that induced by the larval phase of unirradiated infections. The superiority of the irradiated larvae was demonstrated by infecting mice with 500 female cercariae which were either unirradiated or irradiated, and challenging them 3 weeks later with male cercariae. At perfusion, when primary and challenge worms could be distinguished by sex, significantly greater resistance was demonstrated in the groups vaccinated with irradiated larvae (56 cf. 31% and 48 cf. 26% in two experiments). Secondly, in order to establish how long immunizing larvae need to survive to stimulate optimum resistance, mice were either treated with schistosomulicidal drugs or had the site of infection excised at various intervals post-vaccination. Treatment or excision at up to 24 h did not result in significant resistance to challenge (6–16%). Similar treatments at 1 week did not prevent significant resistance developing (26–37%) but the levels of resistance were significantly lower than those manifest by vaccinated, untreated mice (51–71%). It is concluded that the immunizing larvae need to persist for between 1 and 2 weeks in order to stimulate optimum protection and that antigens released during protracted survival and/or death of the larvae in either the skin or the lungs are probably key factors in the induction of resistance.

Freeze fracture study of Schistosoma mansoni membrane differentiation enabled us to describe the characteristic features of the tegumental membrane complex at various stages of development of this parasite. The observation of a membrane exuviation during the 10–20 day period of infection of the definitive host led us to search for possible hormonal control. The development of an appropriate radioimmunoassay has allowed the detection of significant production of ecdysteroids in a human platyhelminth. This hormonal material undergoes level variation during the development of post-cercarial stages of S. mansoni and its precise chemical characterization is presently being investigated.

The influence of temperature and salinity on miracidial longevity and miracidial infectivity of the digenean, Himasthla militaris, has been examined. At 14, 25 and 30 °C the half-life of the miracidia was 1200, 630 and 420 min respectively, and infection of the first intermediate host, Hydrobia ventrosa, only occurred at 25 and 30 °C, for both temperatures 52% became infected. In the range 2·1 to 34‰ (2·1, 4·2, 8·5, 17 and 34‰) the miracidia had a minimal and maximal half-life of 60 and 630 min in water with a salinity of 2·1 and 17‰ respectively, while the infection of the snail host was possible only in water with a salinity of 8·5 and 17‰.

Acid proteases hydrolysing haemoglobin, first described by Timms & Bueding (1959), were studied to investigate the interspecific and intraspecific distribution in adult parasitic helminths. Acid protease activity was detected in a variety of adult helminths, irrespective of their habitats, namely, parasitic nematodes, Angiostrongylus cantonensis, Dirofilaria immitis, Trichuris muris and Ascaris suum, a trematode, Paramphistomum sp. and cestodes, Diphyllobothrium erinacei and Hymenolepis nana. Much higher specific activity was observed in the blood nematodes, A. cantonensis and D. immitis than in the gastro-intestinal helminths examined. The intraspecific distribution of the acid protease activity within organs was studied in those nematodes from which the organs could be anatomically separated. The highest specific activity was observed in the intestine of A. cantonensis, D. immitis and A. suum. The activity was also found in other organs of these nematodes, namely, the reproductive organs and the body wall, but to a much lesser extent. The pH optima for the hydrolysis of haemoglobin were found to be in the pH range of 3·1 to 4·6. Each of the enzyme solutions from the 7 species of helminths hydrolysed haemoglobin faster than the other proteins examined but some activity on myoglobin, casein, albumin and other proteins was demonstrated. Pepstatin, a potent inhibitor of carboxyl proteases, inhibited the hydrolysis of haemoglobin by the acid proteases of the 7 species of the parasites. Acid protease activity from A. cantonensis and A. suum was not inhibited in the presence of thiol-, serine- and metallo-protease inhibitors at a concentration of 10−6 M. This fact, together with the inhibitory effect of pepstatin, suggests that the enzyme activity is likely to be due to carboxyl protease(s). Acid proteases such as cathepsin D and pepsin were simultaneously used for comparison of some enzymological characteristics. The substrate specificity of the enzyme solutions from the helminths seemed to resemble that of cathepsin D rather than that of pepsin. The effects of pH and pepstatin on the acid protease activity of the helminths were similar to those on pepsin and cathepsin D.

The amastigote form of the human pathogen Leishmania mexicana contains high proteinase activity, some 20 times greater than that in the promastigote form and macrophages and appreciably higher than the activity in other flagellate protozoa. The main amastigote enzymes are soluble, whereas those of the promastigote are particulate, and have inhibitor sensitivities characteristic of cysteine proteinases. The very high soluble proteinase activity of L. mexicana amastigotes may be a primary factor in the survival and growth of this mammalian stage in its potentially degradative intracellular habitat.

Survival of Hymenolepis diminuta eggs (as measured by their infectivity to the intermediate host) was found to be age-dependent with an expected value of 11 days when the eggs were retained within the faecal pellet at 10 °C. The expected life-span of eggs under experimental conditions (i.e. extracted from faecal material and placed on filter paper at 30 °C) was estimated as 33 min. The mean parasite burden of populations of Tribolium exposed to known densities of H. diminuta eggs was found to rise to a plateau with increasing exposure time. The experimental results provided a method of estimating the instantaneous rate of parasite transmission: a value of 0·0004/egg/min/host/13 cm2 was obtained. The mean parasite burden of populations of T. confusum exposed to known densities of H. diminuta eggs was found to decrease exponentially with increasing host density. A second estimate of the instantaneous rate of parasite transmission of 0·004/egg/min/host/13 cm2 was obtained from the experimental results. No differences in susceptibility to infection between 2-week-old male and female beetles were found. A marked decrease, however, was apparent with increasing beetle age up to 14 weeks post-eclosion.

Fifteen children aged from 6 to 12 years who were infected with Ascaris lumbricoides were selected for the study. The number of eggs/g (e.p.g.) of stool was determined using Beaver's direct smear technique, Stoll's dilution egg count technique and Katz's modified thick smear technique. Beaver's direct smear method consistently gave a higher e.p.g. than the other two methods. Results of this study show that Katz's modified thick smear technique was the best of the three methods used. The e.p.g. depends on several factors: the amount of stool passed daily, the concentration of eggs within a stool sample, the daily egg output, the worm load, the age of the worm and the technique used for determining the egg load. The daily average egg output/female A. lumbricoides was estimated to be 238722 (range 134462–358 750) with an average e.p.g. of 3540. The estimation of worm load based on egg count is useful in determining the intensity of infection in an individual. The male/female worm ratio was estimated to be 1: 1·4.

‘Each organism must be examined as a biochemical entity before any reasonable understanding of helminth metabolism can be attained’ (Saz, 1969).