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.

Employing a simple method of growing ookinetes of Plasmodium falciparum in culture, 40% of mature gametocytes convert to macrogametes, 4% reach the retort-form ookinete stage and 0·45% become mature ookinetes. A single-step gradient centrifugation method on 12·5% Nycodenz® enriches both gametocytes and ookinetes.

Yellow-eyed penguins on Stewart Island were identified with a Leucocytozoon spp. of a novel lineage in association with a high regional incidence of chick mortality (n=32, 100% mortality) during the November 2006 to January 2007 breeding season. Fourteen chicks from Stewart Island were examined post-mortem and histologically for Leucocytozoon infection. In addition, a survey of blood to detect Leucocytozoon spp. infections using PCR was performed on 107 yellow-eyed penguins from 4 distinct nesting areas on the South Island (Oamaru, Otago Peninsula, and Catlins) (n=95), and Stewart Island (n=12). The results of the study revealed that 2 of the 14 (14%) chicks necropsied showed severe, disseminated megaloschizont formation in the liver, spleen, lung, kidney and other tissues characteristic of leucocytozoonosis. Eighty-three percent (83%) of blood samples collected from Stewart Island penguins contained Leucocytozoon DNA, whereas samples from the 3 other nesting areas were negative for the blood parasite. Leucocytozoon spp. DNA sequences isolated from blood and tissues of adults (n=10) and chicks (n=7) were similar and grouped with other published Leucocytozoon spp. sequences but in a distinct cluster together with closely related isolates from a Western march harrier (Circus aerginosus) and common loon (Gavia immer). These findings suggest that yellow-eyed penguins on Stewart Island are infected with a regionally isolated, host-specific Leucocytozoon spp. which may contribute to the high chick mortality observed during this period.

Infections by the protozoan parasite Toxoplasma gondii are widely prevalent in humans and animals in Turkey but little is known of the burden of their clinical toxoplasmosis. Many early papers on toxoplasmosis in Turkey were published in Turkish and often not available widely. Here, we review prevalence, clinical spectrum, epidemiology and diagnosis of T. gondii in humans and animals in Turkey. This knowledge should be useful to biologists, public health workers, veterinarians and physicians. Although one-third of the human population in Turkey is seropositive, the rate of congenital toxoplasmosis is unknown and no information is available in children 12 years old or younger. One large outbreak of acute toxoplasmosis has been reported in 14–18-year old school children in Turkey. An alarming rate (36%) of T. gondii tissue cysts were reported in tissues of sheep and water buffalo meats destined for human consumption; these reports require verification. Genetically, T. gondii strains from domestic cats and wild birds in Turkey were generally classical type II and III, like those prevalent in Europe. A separate genotype, Type 1 Africa, was isolated from two congenitally infected children and a domestic cat in Turkey.

This study assessed the anthelmintic activity of plant-derived compounds against gastrointestinal nematodes of goats using the egg hatch and larval motility assays. The compounds tested were saponins (digitonin and aescin) and their respective sapogenins (aglycones), hecogenin acetate and flavonoids (catechin, hesperidin, isocordoin and a mixture of isocordoin and cordoin). Additionally, cytotoxicity of active substances was analysed on Vero cell through 3-4,5-dimethylthiazol-2-yl,2,5diphenyltetrazolium bromide (MTT) and propidium iodide (PI) tests. Significant reduction on the egg hatching (P < 0.05) was seen only in the treatments with aescin (99%/EC50 = 0.67 mg mL−1) and digitonin (45%). The compounds that reduced the larval motility (P < 0.05) were digitonin (EC50 = 0.03 mg mL−1 and EC90 = 0.49 mg mL−1) and the hecogenin acetate (75%). The other sapogenins showed low anthelmintic activity. All the flavonoids showed low ovicidal (4–12%) and larvicidal (10–19%) effects. The aescin and digitonin showed low toxicity in PI test (viable cells >90%). Nevertheless, higher cytotoxicity was observed in the MTT assay, with IC50 of 0.20 mg mL−1 (aescin) and 0.0074 mg mL−1 (digitonin). Aescin and digitonin have a pronounced in vitro anthelmintic effect and the glycone portion of these saponins plays an important role in this activity.

The rate of expulsion of penetration gland material was determined from photomicrographs of Hymenolepis diminuta hexacanths stained with neutral red dye at intervals after hatching in vitro. Identical secretory rates were recorded for hexacanths incubated in either Tyrode's solution or Tyrode's solution plus an extract of the midgut of Tenebrio molitor. In both media the reduction in stained material observed in the glands after 135 min incubations was equivalent to that observed for hexacanths that had had opportunity to penetrate in vivo during the same time period. These results were interpreted as indicating that the in vivo secretory pattern was similar to that observed in vitro, and that chemical stimuli from the tissue extracts were not required to initiate secretion. Microdensitometric readings also demonstrated a quantitative decrease in dye within the glands as incubation time increased. Ultrastructural examination of the glands showed that their secretory inclusions were exported via ducts to the epithelial cytoplasm where they accumulated in discrete blebs enclosed by the surface membrane. These inclusion-filled, membrane-bounded blebs were apparently formed and released continuously until, after approximately 2 h, only a few inclusions remained in the penetration glands.

Variations in the general protein and 40 enzymes of the 8 European pathotypes of the potato cyst nematode Globodera rostochiensis and G. pallida have been examined by isoelectric focusing (IEF) and cluster analysis. Differences in the protein patterns were observed and the presence of a major band at pH 8·0 allowed the two species to be discriminated. Extra isozymed of peptidase (EC 3.4. 11.*) and esterase (EC 3. 1. 1. 1) were also found in G. rostochiensis. Intra-species variation was seen within G. rostochiensis for superoxide dismutase (EC 1. 15. 1. 1) and within G. pallida for phosphoglucomutase (EC 2. 7. 5. 1) and glucose phosphate isomerase (EC 5. 3. 1. 9). The results suggest that the two species are very similar and the prospect for recognizing their pathotypes by IEF is discussed.

In a survey of animal trypanosomiasis in Musoma District, carried out in October and November 1970, 6344 G. swynnertoni were collected from six different localities of the Ikoma-Serengeti area and 623 G. pallidipes from Ikoma. These were dissected and examined for trypanosome infections. The mean infection rates of vivax-type in G. swynnertoni and G. pallidipes were 12.6% and 7.5%, respectively. The congolense group infection rates were 2.0% in G. swynnertoni and 1.8% in G. pallidipes. No salivary gland infection was encountered. The incidence of vivax- and congolense-type infections in general increased with age of both the tsetse species, suggesting that the latter can become infected with these trypanosome types at all ages. The infection rate among female tsetse was higher than among males in the six wing-fray categories. This was due to the slower rate of fraying of the wings with age of the former, so that at each wing-fray category the females were generally older than the males. Vivax-type greatly exceeded congolense-type infection rate in both G. swynnertoni and G. pallidipes. It is suggested that this probably reflects the known greater infectivity of the former group of trypanosomes to Glossina. It is concluded that in Ikoma, where game functions as the reservoir of animal trypanosomiasis and where infected tsetse are abundant, the domestic animals are exposed to a continuous trypanosome challenge.

Clinical immunity to Plasmodium falciparum malaria develops after repeated exposure to the parasite. At least 2 P. falciparum variant antigens encoded by multicopy gene families (var and rif) are targets of this adaptive antibody-mediated immunity. A third multigene family of variant antigens comprises the stevor genes. Here, 4 different stevor sequences were selected for cloning and expression in Escherichia coli and His6–tagged fusion proteins were used for assessing the development of immunity. In a cross-sectional analysis of clinically immune adults living in a malaria endemic area in Ghana, high levels of anti-STEVOR IgG antibody titres were determined in ELISA. A cross-sectional study of 90 nine-month-old Ghanaian infants using 1 recombinant STEVOR showed that the antibody responses correlated positively with the number of parasitaemia episodes. In a longitudinal investigation of 17 immunologically naïve 9-month-old infants, 3 different patterns of anti-STEVOR antibody responses could be distinguished (high, transient and low). Children with high anti-STEVOR-antibody levels exhibited an elevated risk for developing parasitaemia episodes. Overall, a protective effect could not be attributed to antibodies against the STEVOR proteins chosen for the study presented here.

A nutritional characteristic of trypanosomatid protozoa is that in vitro they need a haem-compound as a growth factor, which is supplied as haemoglobin, haematin or haemin. Because haemin and related porphyrins are an important source of oxidative stress in biological systems, the effect of haemin on growth, protein content and the antioxidant defence system in Trypanosoma cruzi was evaluated. We have observed that, in epimastigotes grown under different haemin concentrations in the culture medium (0–30 mg/l), 5 mg/l was the haemin concentration yielding optimum growth. Above 15 mg/l there was a clear decrease in growth rate, producing the epimastigote to amastigote transformation. Such morphological change was observed together with a marked injury of the enzymatic machinery of the parasite, leading to diminished protein synthesis as well as lower activity of the antioxidant enzymes (superoxide dismutase, ascorbate peroxidase and trypanothione reductase), reduced total thiol content and a marked increase in the HaemOx-1 activity and expression. The current work demonstrates that there is a correlation between higher haemin concentrations in the culture medium and oxidative damage in the cells. Under these conditions induction of HaemOx-1 would indicate the important role of this enzyme as an antioxidant defence response in Trypanosoma cruzi.

Phyllobothrium piriei sp.nov. (Cestoda: Tetraphyllidea) from the intestine of Raja naevus in the North Sea resembles P. dagnallium, P. radioductum and P. britannicum more closely than any other known species of the genus but differs from these mainly in its host, in having distinctly bifid bothridia with faint marginal loculi and about 150 testes in each proglottid. Brief comments are made on the preference shown by the adult for the posterior region of the intestine and on a modification of its bothridium for attachment to up to 12 villi.

This work was supported by generous financial aid from the Natural Environment Research Council. I wish also to record my gratitude to Professor V. C. Wynne-Edwards and Dr P. A. Orkin for reading and suggesting improvements to the manuscript and to the Director and staff Marine Laboratory, Aberdeen, in particular Dr B. B. Rae and Mr Stuart Pirie, after whom this species has been named, for providing live rays. Alasdair McVivar of this department, who has now begun an independent study of the intestine of Raja naevus as a habitat for helminth parasites, kindly provided information for Fig. 9.

Oocyst wall formation in Eimeria maxima was studied during the macrogamete stage in intestinal cells of the chick and in unsporulated oocysts isolated from faeces. The outer of the 2 membranes bounding the mature macrogamete separated from the surface but remained as a veil surrounding the developing oocyst throughout the whole intracellular process. Wall-forming bodies Type I were initially applied to the limiting membrane of the zygote cytoplasm; a layer of material similar to their contents was then formed around the zygote. As this occurred a new double membrane was formed surrounding the oocyst cytoplasm. The outer wall layer was initially homogenous in appearance but later developed into 2 zones, an outer amorphous region and an inner osmiophilic region. The inner layer of the oocyst wall was formed from the contents of the wallforming bodies Type II which dispersed between the outer wall and the limiting membranes of the oocyst cytoplasm. There was evidence of an additional membrane formed external to the outer wall. The outer membranes were not present around the wall of oocysts passed in the faeces of chicks, but the same wall zonation was evident, although the inner osmiophilic zone of the outer wall layer was markedly thinner in comparison with the same zone seen in the tissues.

The transmission of Hammondia heydorni among sheep, goats, cattle, dogs and coyotes is described. Oocysts of H. heydorni, obtained from dog faeces by feeding naturally infected goat meat, were fed to an experimental goat. After 312 days tissues from this animal were fed to 2 dogs. One dog, which received goat muscle, shed oocysts, whereas the other dog which received brain, spleen, kidneys, lungs and liver did not. Oocysts from the dog fed goat muscle were fed to an experimental sheep. After 73 days sheep tissues were fed to a dog and a coyote, both of which shed oocysts 5–8 days later. A second isolate of H. heydorni was obtained by feeding naturally infected beef to a coyote. These oocysts were fed to a single goat and sheep and after 71 days, muscle from these 2 animals was fed to 2 dogs and 2 coyotes, all of which shed oocysts 5–8 days later. None of the animals inoculated with H. heydorni became ill and no Hammondia-like organisms were found in tissue sections from the herbivores. Another dog shed H. heydorni-like occysts between 7 and 10 days after ingesting naturally infected muscles from a moose. These results indicate that H. heydorni infects goats, sheep, moose and coyotes in addition to cattle and dogs.

1. Standard conditions are described for preparing suspensions of washed Trypanosoma lewisi and T. equiperdum in modified Ringer–phosphate solution.

2. Oxygen consumption was measured with differential manometers, using microflasks containing 2–5 × 107 trypanosomes in 0·9 ml. of reaction mixture. Measurements of oxygen uptake were carried out at 37° C.

3. T. lewisi respired slowly in the absence of substrate for up to 2 hr. The trypanosomes suffered little damage when stored at 5° C. for 24 hr. without substrate. No oxygen uptake was observed with T. equiperdum in the absence of substrate. The trypanosomes were viable after 24 hr. at 5° C. with glucose or glycerol as substrate, but not in the absence of substrate.

4. With glucose as substrate, the rate of oxygen consumption by T. lewisi increased with the age of infection. This change was more marked with glutamine as substrate.

5. With glucosamine as substrate, the oxygen uptake of T. lewisi was at a slightly lower rate than with glucose. The rate of oxygen uptake was still lower with Na l-glutamic acid, asparagine, aspartic acid, casein hydrolysate, yeast extract and Difco Bacto-peptone. Thirteen other amino-acids had no effect on the motility of the trypanosomes.

6. With glycerol as substrate, the oxygen uptake of T. equiperdum was at a slightly lower rate than with glucose. The rate of oxygen uptake was very low with yeast extract, casein hydrolysate and Difco Bacto-peptone. No oxygen uptake or motility was recorded with glutamine, Na l-glutamic acid, glucosamine, asparagine, aspartic acid, dl-alanine, or Na acetate. Thirteen other amino-acids had no effect on the motility of the trypanosomes.

7. Ammonia was liberated from glutamine by adult and reproductive phase T. lewisi.

Studies on immuno-conglutinin levels in animals infected with T. brucei have shown that immuno-conglutinin appears in rabbits infected with T. brucei ‘M’ variant within 7 days after infection and reaches its peak titre within 30 days. In rabbits infected with the more virulent ‘R’ variant immuno-conglutinin levels rose more rapidly and reached peak titres within 10–14 days after infection.

High levels of immuno-conglutinin are detectable in the sera of rabbits and cats infected with trypanosomes before a significant amount of neutralizing antibody is produced. This difference is most marked in animals infected with the antibody-resistant variant ‘R’ which is more virulent for rabbits.

In animals treated effectively the immuno-conglutinin level dropped to its pre-infection level, while the titre of neutralizing antibody only decreased to a slightly lower level.

No change in the titres of immuno-conglutinin and neutralizing antibody was observed after treatment of animals infected with a drug-resistant strain.

These results are very promising for further field trials and introduce a new approach and method for testing the efficiency of chemotherapy in trypanosomiasis.

The entry of Theileria parva merozoites into bovine erythrocytes in vivo is described and compared to sporozoite invasion of lymphocytes. Merozoites make initial contact with erythrocytes with any part of their surface and invasion of the host cell does not require the re-orientation of the apical end of the merozoite towards the surface of the erythrocyte. After the initial attachment the merozoite and host cell membranes form a continual close junction with the two apposed membranes separated by a 6–8 nm gap containing moderately dense material. The progressive circumferential ‘zippering’ of these closely apposed membranes leads to the movement of the parasite into the erythrocyte. The newly internalized merozoite which is completely surrounded by the erythrocyte plasma membrane escapes from this enclosing membrane by a process involving the discharge of at least the rhoptries; whether the merozoite also contain other types of secretory organelles (e.g. micronemes, microspheres or dense bodies) remains to be determined. Morphologically, the events involved in merozoite invasion of erythrocytes are almost identical to the process of sporozoite invasion of lymphocytes but differ significantly from the entry process of the invasive stages of other Apicomplexan parasites.

The present material of buffalo–dog origin could not be transmitted to the laboratory animals, such as white rats, white mice, rabbits, golden hamsters, guinea-pigs and chicks.

Secondary development took place only in mice.

The strobilar phase was infective to a cat but did not show sexual differentiation even at 30 days, and worms were further observed to be at a degenerative stage.

The buffalo was highly suited to the perpetuation of the parasite, forming larger cysts and massive production of scoleces; nearly 90% of the cysts were fertile.

The strobila had mostly two segments and not more than three.

The number and distribution of testes were more posterior to the genital pore and were also observed posterior to the vitellaria, occasionally.

The position of the genital pore in the gravid as well as mature segments was both anterior and posterior but with an average of 52.8%.

Uterine sacculations were observed to range from 28 to 54.

Echinococcus material of buffalo–dog origin does not appear to fall into a definite ‘subspecies’ pattern. It is suggested that it be merely recognized as a further ‘mutant’, the existence of which can be accounted for on theoretical grounds.

The authors are thankful to the Director, Indian Veterinary Research Institute, and to Dr H. D. Srivastava, Head of Division of Parasitology, I.V.R.I., for providing the necessary laboratory facilities.

Natural infection with the cephalobid nematode, Daubaylia potomaca, is reported in Helisoma antrosum pericarinatum ( = H. anceps pericarinatum) collected in the vicinity of the University of Michigan Biological Station on Douglas Lake, Cheboygan, Michigan.

The life-cycle of D. potomaca, which appears to be unique, has been determined and reproduced experimentally utilizing Australorbis glabratus and other snails as hosts. The cycle involves escape from the host by gravid females, invasion by the females of new hosts and the reproduction therein of generation after generation of parasites, and the eventual emergence of female worms from dying hosts to the external aquatic environment. The parasite does not lead a saprozoic existence, and no dauer larvae were seen. Except for the newly emerged female, no stage of D. potomaca is ‘free-living’, and there is no evidence that transmission can occur except through the agency of these females. However, parasitic females removed at any time from their hosts by dissection are fully capable of infecting new hosts under experimental conditions.

Dissection of A. glabratus infected with known numbers of female D. potomaca disclosed that the gravid nematodes deposit about two eggs per day in the tissues, that larvae may be found after 5 days of infection, and that the first ‘new’ generation of adult males and females appears within the host 11 or more days after infection. Continued reproduction in the host may give rise to very large nematode populations. A. glabratus exposed to five female D. potomaca usually became infected and all infected snails were killed within 44 days. Death of infected snails was preceded, in nearly every case, by the emergence of female D. potomaca from their hosts. It is suggested that this phenomenon reflects an adaptive response of the worms to changes in the internal milieu of the snail.

Attempts to infect certain physid and lymnaeid snails with D. potomaca failed, and varying degrees of success were achieved in attempts to infect several planorbids. Of the latter, the most suitable hosts appeared to be A. glabratus (Puerto Rico) and Helisoma sp. (Michigan); the least suitable host was Helisoma caribaeum (Puerto Rico and St Croix), in which an intense tissue reaction killed all nematode eggs before they could hatch. Other susceptible planorbids (in declining order of apparent suitability as hosts) were Helisoma trivolvis ? macrostomum (Canada), Bulinus truncatus (Egypt) and Planorbarius corneus (Massachusetts).

Some of the problems associated with the possible use of D. potomaca as a means of biological control for A. glabratus are discussed.

It is a pleasure to thank the persons whose names appear in the text for aid or counsel graciously extended. I am also grateful to Dr George L. Graham (University of Pennsylvania) for certain citations in the literature, to Dr M. R. Tripp (now at the University of Delaware) for receiving the shipments of snails in Boston, and to Dr A. H. Stockard, Director of the University of Michigan Biological Station, for facilitating the field work.

Leishmaniasis is a complex of zoonotic diseases caused by parasites of the genus Leishmania, which can develop in domestic as well as wild animals and humans throughout the world. Currently, this disease is spreading in rural and urban areas of non-endemic regions in Brazil. Recently, bats have gained epidemiological significance in leishmaniasis due to its close relationship with human settlements. In this study, we investigated the presence of Leishmania spp. DNA in blood samples from 448 bats belonging to four families representing 20 species that were captured in the Triangulo Mineiro and Alto Paranaiba areas of Minas Gerais State (non-endemic areas for leishmaniasis), Brazil. Leishmania spp. DNA was detected in 8·0% of the blood samples, 41·6% of which were Leishmania infantum, 38·9% Leishmania amazonensis and 19·4% Leishmania braziliensis. No positive correlation was found between Leishmania spp. and bat food source. The species with more infection rates were the insectivorous bats Eumops perotis; 22·2% (4/18) of which tested positive for Leishmania DNA. The presence of Leishmania in the bat blood samples, as observed in this study, represents epidemiological importance due to the absence of Leishmaniasis cases in the region.

Indirect immunofluorescent studies have been used to localize a glycoprotein (gp32) in the amphids of 2nd-stage juveniles (J2) of Meloidogyne incognita parasitic in the roots of tomato plants, male Meloidogyne javanica and some young adult female M. incognita. Immuno-electron microscopic studies showed an apparent absence of gp32 in the amphids of mature adult female M. incognita. Chemoattraction experiments demonstrated that the binding of gp32 antibodies to the amphidial secretions of infective J2 M. incognita partly impaired the ability of the nematodes to locate tomato roots.