Intracellular forms of Trypanosoma lewisi have been found in the mid-gut cells of the flea, Nosopsyllus fasciatus. These stages have been found only infrequently, and in only two of the four strains of T. lewisi studied.

Attempts to find similar stages in Trypanosoma nabiasi, evotomys and microti were not successful.

The possible significance of these forms is discussed.

The financial assistance of the Agricultural Research Council is gratefully acknowledged. The author is also indebted to Dr P. Tate for his enthusiasm, help and guidance throughout the course of the study.

1. The oxygen uptake of Trypanosoma lewisi and T. equiperdum was measured with differential manometers using microflasks of 5 ml. capacity. Each flask contained 2–5 × 107 trypanosomes.

2. The oxygen uptake of trypanosomes in heparinized infected blood was compared with that of washed trypanosomes suspended in Ringer–phosphate–glucose solution with or without added serum or blood. With T. lewisi, the oxygen uptake began at a higher rate in the presence of blood or serum than with Ringer–phosphate–glucose solution. With T. equiperdum, the oxygen uptake in the presence of blood or serum began at almost the same rate as with Ringer–phosphate–glucose solution, but the rate of oxygen uptake decreased more rapidly in the absence of blood or serum than when it was present.

3. High concentrations of iodoacetic acid (e.g. 3·3 × 10−2m) caused an increase in oxygen uptake when tipped into heparinized normal rat blood. Similar concentrations caused an inhibition of oxygen uptake when 2·0 × 10−4m methylene blue was also present or when the blood had been treated with sodium nitrite to convert haemoglobin into methaemoglobin.

4. Iodoacetic acid caused little inhibition of oxygen uptake when T. lewisi was suspended in Ringer–phosphate solution containing glutamine or glutamine plus glucose, or when heparinized infected blood was tested. Inhibition was greatest when the substrate was glucose, while glucosamine, glycerol, or glucose plus serum gave intermediate results.

5. Iodoacetic acid caused least inhibition of oxygen uptake when T. equiperdum was suspended in Ringer–phosphate solution containing glycerol, or when heparinized infected blood was tested. Inhibition was greatest when the substrate was glucose, and was slightly less marked when the substrate was glucose plus serum or glucose plus glycerol.

6. Neoarsphenamine inhibited the oxygen uptake of T. lewisi to a similar extent whether glucose or glutamine was present.

7. Neoarsphenamine and reduced stovarsol thioglycollate inhibited the oxygen uptake of T. equiperdum slightly less when the substrate was glycerol than when it was glucose. Sodium arsenite was as active as reduced stovarsol thioglycollate when the substrate was glucose, but 10−4m sodium arsenite had little effect when the substrate was glycerol.

Using the ability to sporulate as a measure of viability, the effects of exposure of unsporulated oocysts of 10 species of coccidia of chickens, rabbits and cattle to saturated NaCl solution has been studied. Although appreciable deformation and collapse of the oocyst occurred after 1–2 days contact, the effect was reversible after washing free from salt and incubating. Some reduction in ability to sporulate following several days contact with saturated salt was noted in most species, although no effect was seen with Eimeria stiedai following 7 days exposure, Eimeria tenella was one of the more sensitive species studied. Culture titration experiments in chickens with E. tenella indicated that oocysts which had sporulated following prolonged exposure to salt were in no way inferior in virulence or ability to retain virulence on prolonged storage to oocysts prepared with minimal contact with salt. No evidence was obtained to contra-indicate the use of salt-flotation methods for the separation of oocysts from faeces.

Many studies have been carried out on the structure and chemical composition of trematode and cestode egg envelopes. Among the nematodes, much work has been done on the eggs of ascarids but there have been few studies on the egg envelopes of species in the Superfamily Trichostrongyloidea. Zawadowsky et al. (1929) reported on the biology of five species of this family, namely Trichostrongylus instabilis (T. colubriformis), T. extenuatus (T. axei), T. probolurus, Cooperia pectinata and Ostertagia mentulata. They described the egg envelope as consisting of three or possibly four layers. The physical and chemical properties of these layers were investigated and these workers concluded that the structure of the egg envelope was similar to that of Ascaris. However, the solubility of the inner layer differed from that of Ascaris in dissolving only partially in absolute alcohol and very little in ether.

The genus Astiotrema Looss is parasitic in reptiles (tortoises and snakes), except Ast. impletum Looss, which is known from a fish Tetradon fahaka. Hitherto three species of this genus have been recorded, two from Egypt and one from Naples (Italy). The present paper contains an account of two more species obtained from tortoises at Allahabad. The allied genus Glypthelmins Stafford is parasitic in Amphibia in North America.

The host of this gregarine is the larva of a Ceratopogonid: Dasyhelea obscura Winnertz, which, as I have previously mentioned, lives in the decomposed sap filling the infected wounds of the Elm and Horse-chestnut trees in Cambridge (at Newnham and along the Queen's Road). The parasitised larvae were found late in the season, about the end of September, they were all young and never heavily infected, containing from three to eight and exceptionally twelve parasites in different developmental stages. The gregarine seems to be rare as among several hundred of these larvae examined only twelve yielded this interesting parasite.

We compared 26 hand-reared grey partridges given an experimental infection of the caecal nematode Heterakis gallinarum with 26 uninfected ones. Under laboratory conditions after 91 days, there were no measurable clinical effects of the infection. We found no effect of treatment on the amount of food eaten or on caecal dropping production. However, treated birds, in particular females, developed slightly lower body mass (around 2%) compared to the controls. At post-mortem examination, we found a positive relationship between breast muscle mass and the number of worms collected from the caeca of treated birds. Treated birds with no worms when examined had smaller breast muscle mass (4·6%) compared to the uninfected control birds. These results are largely different to those found in a similar study that documented significant negative impacts on most of these factors in 8 infected birds compared to 6 controls. Its findings were used in a published model to support a hypothesis that H. gallinarum maintained in the environment by common pheasants, the primary host for this worm, could negatively affect wild grey partridge productivity and survival. In the same model our data would not support this hypothesis. Possible explanations for the different results from the 2 experiments are discussed. Together they suggest that only in certain, as yet unidentified circumstances, could experimental H. gallinarum infections have deleterious effects on hand-reared grey partridges.

Bovine besnoitiosis is a chronic and debilitating disease, caused by the apicomplexan parasite Besnoitia besnoiti. Infection of cattle by B. besnoiti is governed by the tachyzoite stage, which is related to acute infection, and the bradyzoite stage gathered into macroscopic cysts located in subcutaneous tissue in the skin, mucosal membranes and sclera conjunctiva and related to persistence and chronic infection. However, the entire life cycle of this parasite and the molecular mechanisms underlying tachyzoite-to-bradyzoite conversion remain unknown. In this context, a different antigenic pattern has been observed between tachyzoite and bradyzoite extracts. Thus, to identify stage-specific proteins, a difference gel electrophoresis (DIGE) approach was used on tachyzoite and bradyzoite extracts followed by mass spectrometry (MS) analysis. A total of 130 and 132 spots were differentially expressed in bradyzoites and tachyzoites, respectively (average ratio±1·5, P<0·05 in t-test). Furthermore, 25 differentially expressed spots were selected and analysed by MALDI-TOF/MS. As a result, 5 up-regulated bradyzoite proteins (GAPDH, ENO1, LDH, SOD and RNA polymerase) and 5 up-regulated tachyzoite proteins (ENO2; LDH; ATP synthase; HSP70 and PDI) were identified. The present results set the basis for the identification of new proteins as drug targets. Moreover, the role of these proteins in tachyzoite-to-bradyzoite conversion and the role of the host cell environment should be a subject of further research.

In Medium 199 supplemented with calf serum (+ 10%), adult Schistosoma mansoni produced a mean total of 400 eggs/worm pair during maintenance for 10 days in vitro. During the period of egg deposition in vitro, the dry weight of worm pairs decreased by 35%. Biomass decreases were proportionally greater in female worms than in males. Declines in the dry weight of female worms were mainly due to decreases in protein content, while biomass decreases in males resulted from losses of both protein and glycogen. Glucose was depleted from the maintenance medium at a rate of approximately 200 μg/worm pair/day during each of the first 3 days in vitro, while concentrations of glucose in the medium of above 1 g/litre did not affect the degree of glycogen depletion observed in adult S. mansoni during culture for 24 h. The contribution of the loss of schistosome gut contents in vitro to the observed changes is discussed. The work described represents part of an attempt at logical design of in vitro culture.

By means of ultra-thin sections for electron microscopy it has been possible to observe intra-nuclear changes in the polyhedrosis of larvae of Tipula paludosa. The virus rods arise in the centres of the greatly enlarged nuclei of blood cells. Thin-walled vesicles, apparently filled with fluid, form round individual virus rods and collect in masses on the inner wall of the nuclear membrane. Crystallization then begins, starting at the edge against the nuclear membrane. The vesicles are gradually compressed round the rods, eventually forming a capsule. Sections through free virus particles show them to be enclosed in a membrane, a second inner membrane round the substance of the rod may also be present.

There appears to be no capsule round the free virus particles such as encloses the virus particles inside the polyhedral crystal.

The prevalence of Trypanosoma cobitis was recorded in species of small fish from the River Lee at Enfield Lock during 33 months of sampling. Trypanosome infections were more numerous during the summer months with 50–70% of bottom-living ‘inactive’ fish, bullheads (Cottus gobio), stone loach (Nemacheilus barbatulus) and gudgeon (Gobio gobio) infected, compared to only 6 and 18% of the more active pelagic fishes, stickle-backs (Gasterosteus aculeatus) and minnows (Phoxinus phoxinus).

Highly purified antigen and appropriate controls are essential for antigen-specific immunoassays. In the case of Isospora suis, the causative agent of neonatal porcine coccidiosis, the only current source of antigen is oocysts isolated from faeces. The aim of this study was to develop a procedure for high-grade purification of I. suis oocysts from piglet faeces to obtain both antigen and representative controls suitable for in vitro re-stimulation of lymphocytes. This was achieved by use of filtration, density-gradient centrifugation and fluorescence-activated cell sorting (FACS). The feasibility for immunological studies was demonstrated with IFN-γ ELISPOT assays after in vitro re-stimulation of lymphocytes from previously infected swine using the obtained antigen. The developed method allowed the production of highly purified antigen and representative controls from faeces with an oocyst recovery rate of 14%. Regarding the application of the obtained material it could be shown that lymphocytes from I. suis-infected pigs react in an antigen-specific manner in terms of an in vitro recall response by the production of IFN-γ. This demonstrates the suitability of the developed method for the production of antigen and controls for sensitive immunological readout systems. Moreover, the detected specific IFN-γ response encourages further functional studies on the cellular immune response to I. suis.

This review of protozoan and helminth infections in pregnancy focuses on the impact on the immune response in the newborn infant to maternal infection. Studies of protozoan and helminth infections in pregnant women and in their offspring have shown that children exposed to antigens or microorganisms during pregnancy often have a reduced immune response to these infections. The most common finding is a reduced IFNγ response to specific antigens regardless of specific infection studied. In some studies the impaired immune response disappeared before the age of one year, while in other studies the impaired immune response was present as much as two decades after birth. Data from chronic viral infections like Rubella, cytomegalovirus and hepatitis B also show that congenital or perinatal infections may result in a life-long inability to control the infections. Studies of both helminth and protozoan infections show that children exposed to antigens during gestation have a microorganism-specific impaired immune response which is characterized by reduced IFN-γ and stimulation of responses to specific antigens.

Babesia caballi and Theileria equi are biological agents responsible for equine piroplasmosis (EP). We conducted a robust and extensive epidemiological study in Nigeria on the prevalence and risk factors of EP. Blood (468, both horses and donkeys) and ticks (201 pools) were screened using polymerase chain reaction (PCR). DNA of equine piroplasms was observed in tick pools with B. caballi amplified in Rhipicephalus evertsi evertsi only [minimum infection rate (MIR) of 7.6%] while T. equi was observed in R. e. evertsi (MIR, 61.6%), Hyalomma dromedarii (MIR, 23.7%) and H. truncatum (MIR, 50.0%). Overall results showed that 196/468 (41.9%) animals were positive for equine piroplasms (both B. caballi and T. equi). The prevalence for T. equi was 189/468 (40.4%) compared to 7/468 (1.5%) for B. caballi. In the horses and donkeys, respectively, the prevalence for T. equi was (39.9%; 112/281) and (41.2%; 77/187) compared with (1.4%; 4/281) and (1.6%; 3/187) due to B. caballi. Our analysis showed that location (Jigawa state), Talon breed, horses used for work and reproduction, unsatisfactory husbandry practices, contact with other mammals are risk factors that associated positivity to T. equi infection in horses, whilst horses kept on intensive management appeared to be less prone to infection. On the other hand, Jangora breed of donkeys and location (Jigawa state) are risk factors to infection with T. equi in donkeys. Findings suggest the persistence of EP in equids and ticks in Nigeria.

Three superoxide dismutases (SOD) (EC 1.15.1.1) were detected in homogenates of Ascaris suum. Each of the three forms of SOD was purified by a sequence of multiple differential centrifugations, ammonium sulphate precipitation, ion-exchange chromatography and G-75 Sephadex column chromatography. The three forms of SOD were present in different cellular locations; one in the cytoplasmic fraction, sensitive to cyanide and hydrogen peroxide, and two in the mitochondrial fraction, one of which was cyanide sensitive. The SOD forms presented clear differences in their electrophoretic patterns. The sexual organs of females showed the highest SOD activities of all the tissues examined. The finding of such high levels of superoxide dismutase in A. suum reflects the importance of this enzyme in the metabolism of this helminth parasite.

A combined interdisciplinary research strategy is even more crucial in immunology than in many other biological sciences in order to comprehend the closely linked interactions between cell proliferation, molecular signalling and gene rearrangements. Because of the multi-dimensional nature of the immune system, an abundance of different experimental approaches has developed, with a main focus on cellular and molecular mechanisms. The role of metabolism in immunity has been underexplored so far, and yet researchers have made important contributions in describing associations of immune processes and metabolic pathways, such as the central role of the l-arginine pathway in macrophage activation or the immune regulatory functions of the nucleotides. Furthermore, metabolite supplement studies, including nutritional administration and labelled substrates, have opened up new means of manipulating immune mechanisms. Metabolic profiling has introduced a reproducible platform for systemic assessment of changes at the small-molecule level within a host organism, and specific metabolic fingerprints of several parasitic infections have been characterized by 1H NMR spectroscopy. The application of multivariate statistical methods to spectral data has facilitated recovery of biomarkers, such as increased acute phase protein signals, and enabled direct correlation to the relative cytokine levels, which encourages further application of metabolic profiling to explore immune regulatory systems.

Trypanosoma cruzi has three biochemically and morphologically distinct developmental stages that are programmed to rapidly respond to environmental changes the parasite faces during its life cycle. Unlike other eukaryotes, Trypanosomatid genomes contain protein coding genes that are transcribed into polycistronic pre-mRNAs and have their expression controlled by post-transcriptional mechanisms. Transcriptome analyses comparing three stages of the T. cruzi life cycle revealed changes in gene expression that reflect the parasite adaptation to distinct environments. Several genes encoding RNA binding proteins (RBPs), known to act as key post-transcriptional regulatory factors, were also differentially expressed. We characterized one T. cruzi RBP, named TcZH3H12, which contains a zinc finger domain and is up-regulated in epimastigotes compared to trypomastigotes and amastigotes. TcZC3H12 knockout (KO) epimastigotes showed decreased growth rates and increased capacity to differentiate into metacyclic trypomastigotes. Transcriptome analyses comparing wild type and TcZC3H12 KOs revealed a TcZC3H12-dependent expression of epimastigote-specific genes such as genes encoding amino acid transporters and proteins associated with differentiation (PADs). RNA immunoprecipitation assays showed that transcripts from the PAD family interact with TcZC3H12. Taken together, these findings suggest that TcZC3H12 positively regulates the expression of genes involved in epimastigote proliferation and also acts as a negative regulator of metacyclogenesis.

Six myxosporidian species were found in chub (Leuciscus cephalus) originating from Lower Austrian rivers. The frequency of the parasites and their localization was recorded. In all chub, independent of size and origin, Myxobolus cyprini occurred predominantly in the macrophage centres (MCs) of the haematopoietic organs, spleen and kidney. Exclusively in the head kidney of young fish not yet described vermicular plasmodia containing spores of M. cyprini were found. In muscle tissue the prevalence of M. cyprini was comparatively low. Other species of Myxobolus characterized by plasmodial cysts frequently occurred in gills and swimbladder but were rarely detected, and only in small numbers, in the haematopoietic organs. The number of M. cyprini spores and the relative volume of MCs in the haematopoietic organs were estimated in order to examine possible correlations. Significant interrelated changes were found only in juvenile fish up to a size of 15cm. In bigger fish, the number and size of macrophage aggregates were highly variable and independent of infection intensity and fish size, but the number of spores never exceeded that of the aggregated macrophages. The data suggest that due to an early date of infection M. cyprini is the only species which is closely associated with macrophage aggregation.

Recent have shown the relationship between Ecto-Nucleoside-Triphosphate-Diphosphohydrolases (Ecto-NTPDases or ecto-nucleotidases) and virulence and infectivity in trypanosomatids. In this work, the inhibition of the ecto-ATPase activities and promastigote growth of Leishmania amazonensis by CrATP was characterized. Furthermore, this compound was used to investigate the role of ecto-nucleotidase in the interaction of L. amazonensis with resident peritoneal macrophages obtained from BALB/c mice. CrATP partially inhibits the ecto-ATPase activity, presenting Ki values of 575·7±199·1 and 383·5±79·0 μm, in the presence or absence of 5 mm MgCl2, respectively. The apparent Kms for ATP (2·9±0·5 mm to Mg2+-dependent ecto-ATPase and 0·4±0·2 mm to Mg2+-independent ecto-ATPase activities) are not significantly altered by CrATP, suggesting a reversible non-competitive inhibition of both enzymes. When CrATP was added to the cultivation medium at 500 μm, it drastically inhibited the cellular growth. The interaction of promastigote forms of L. amazonensis with BALB/c peritoneal macrophages is strongly affected by CrATP. When the parasites were treated with 500 μm CrATP before interacting with macrophages, the adhesion and endocytic indices were strongly reduced to 53·0±14·8% and 39·8±1·1%, respectively. These results indicate that ecto-nucleotidase plays an important role in the infection process caused by Leishmania amazonensis.