Cytamoeba bacterifera from urodeles (Batrachoseps attenuatus, Aneides lugubris, A. flavipunctatus and Dicamptodon ensatus) show a thin peripheral ring of DNA and a less definite, heterogeneous central portion after Feulgen treatment; toluidine blue reveals a similar configuration. Exposure to DNA-ase and RNA-ase, followed by toluidine blue and Feulgen treatment, substantiates the localization of DNA; RNA is present in small quantity and restricted to the periphery of the parasite.

Mitochondria (2–5μ long rod-shaped, central bodies) can be detected with Janus Green B, and osmium tetroxide yields a dark central mass of rods and granules as well as peripheral granules; Sudan Black IV is without action. Zymohexase and acid phosphatase can be detected by cytochemical means, but peroxidase, lipase, urease and alkaline phosphatase were not noted.

From the evidence presented, C. bacterifera is considered a living entity and is tentatively relegated to the Piroplasmida.

The writer would like to express his appreciation to Dr R. S. Bray of the American Foundation for Tropical Medicine, Harbel, Liberia, for his counsel and suggestions.

1. A local population of Gammarus pulex, infected with Heliospora longissima and Rotundula gammari, was found to have an intracellular developmental stage.

2. Experimental infections of G. pulex were made with the two species of gregarines. In the case of R. gammari infections, large numbers of intracellular developmental stages were found.

3. The taxonomic relationship of the genera Cephaloidophora and Rotundula is discussed.

Thanks are due to Dr P. Tate for constant encouragement and guidance during the course of this work and for the many helpful suggestions in the preparation of the manuscript. Thanks are also due to Mr D. W. T. Crompton for kindly giving me the infected material. The work was completed during the tenure of a Royal Society and Nuffield Foundation Commonwealth Bursary.

Stages seen in the life-cycle of Diplocystis tipulae sp.nov. parasitic in Tipula paludosa Mg., included a structure that was probably a trophozoite within the midgut epithelium of one host larva and gamonts that developed on the midgut wall in the haemocoele of the larvae. Syzygy occurred, usually in pairs, when the gamonts were about 200–300 μm in diameter. Initially, syzygous gamonts were bilobed but they quickly became spherical and were enclosed by a fine membrane to form a gametocyst 450–550 μm in diameter. Early stages of gametogony were seen in one pupa; sporogony occurred in pupae and adults. The mean size of the round–oval oocysts was 7·14 × 6·26 μm. Individuals of Tipula oleracea L. are also susceptible to infection with D. tipulae. This gregarine is considered to be a new species, independent of the gregarines of the gut lumen.

R. carnivoralis has been shown to be capable of transmitting T. parva under experimental conditions and stages of the protozoon have been found in sections of salivary glands of the tick.

All instars of the pentastomid Porocephalus crotali, in the tissues of rat intermediate hosts and the lung of rattlesnake definitive hosts, are covered by a vesicular or stacked membranous secretory product which is synthesized in sub-parietal cells (SPC) and channelled to the cuticle via multitudinous chitin-lined ducts. In rats this enveloping foam of vesicles survives for the duration of a cuticle. We have purified and partially characterized the lipid and protein composition of lamellate droplets from SPC of infective nymphs. Lipids in the droplets comprised a mixture of neutral and polar lipid classes with cholesterol being the major neutral lipid and phosphatidylcholine the dominant polar lipid. In addition, Triton X-114 phase separation of the protein component of droplets partitioned these into an aqueous phase (with a major band at 60 kDa), a detergent phase with two hydrophobic polypeptides (24 and 16 kDa) and an insoluble pellet containing several minor proteins and major bands at 31 and 107 kDa. Western blotting, with rabbit anti-lamellate droplet antiserum, strongly label only the 60 kDa and the two hydrophobic proteins. Significantly no proteins at all label with serum from infected rats from 50–150 days post-infection, a finding endorsed by indirect fluorescent antibody tests on sectioned V–VII stage nymphs. Possible immunomodulatory functions of SPC-derived surface membranes are discussed and, in this regard, they are compared with pulmonary surfactant, a product of alveolar Type II cells which lines all vertebrate lungs.

An in vitro culture of Plasmodium falciparum (isolate FCUP-1/RSA) was arrested in the trophozoite stage with α-difluoromethylornithine (DFMO), followed by sorbitol treatment to liberate intracellular parasites from infected erythrocytes. Most of the unlysed erythrocytes and ghost membranes were removed by filtration, after agglutination with anti-erythrocyte monoclonal antibodies or wheat-germ agglutinin. A highly purified parasite preparation was obtained after chromatography on an immuno-affinity column consisting of polystyrene particles to which anti-erythrocyte monoclonal antibodies had been adsorbed. Isolated parasites were free of surrounding erythrocyte membranes and structurally intact as assessed by transmission electron microscopy, SDS–PAGE and ELISA.

Two types of exflagellating bodies of the malaria parasite are described in the infected blood of the Himalayan flying squirrel (Petaurista magnificus). The first type, which is larger, is characterized by a blue-staining cytoplasm. These produce blue-microgametes with red chromatin dots. The second type, which is smaller, stains red, and also produces red microgametes. Certain large schizonts occurring in the peripheral blood are described. It is speculated that these might represent the primary tissue stage of Rayella rayi.

Cysteine (20–100 mM) and reduced glutathione (20–80 mM) potentiated the development of the exsheathed third-stage larva of Haemonchus contortus to the fourth stage. The potentiating effect of cysteine was greatest during the short period before and after the exsheathment of the third-stage larva. Two hundred mM cysteine and 1 mM p–chloro-benzoate inhibited development. This inhibition was reversible. Ten mM iodoacetate and 0·5 and 5 mM n–ethyl-maleimide inhibited development of the larvae. The larvae appeared to be most susceptible to the inhibiting effects of these reagents during the period of development approaching the third moult. Development was either little affected or potentiated, when larvae were incubated with these reagents for short periods immediately after exsheathment.

The effects of the phenothiazine, Stelazine, on Hymenolepis diminuta were investigated. The cestode was incubated for 10 min at 37 °C with 1 mM trifluoperazine, in the presence and absence of Ca2+. Assay of brush border enzymes showed that drug treatment lowered the activities of alkaline phosphatase, Ca2+-ATP'ase, 5′-nucleotidase and type 1 phosphodiesterase. This occurred in parallel with a significant reduction in tegumental protein. Under these conditions gross changes in ultrastructural appearance and cellular organization were observed. There was a lack of ordered microtriches and the distal cytoplasm was absent. Glycogen granules were scattered throughout the cytoplasm within the subtegumental layer. The connective tissue also appeared to be in some disarray. The effects of Stelazine appeared to be dependent on time and were significantly increased when Ca2+ was included in the incubation medium. Incubation with the less hydrophobic phenothiazine trifluoperazine sulphoxide had minimal effect on the integrity of the cestode. The results reported here support the premise that certain phenothiazines may be considered as potential cestocidal agents.

Guanylate kinase, a nucleoside monophosphate kinase of Brugia malayi which is involved in reversible transfer of phosphate groups from ATP to GMP, was cloned, expressed and characterized. The native molecular mass of BmGK was found to be 45 kDa as determined by size exclusion chromatography and glutaraldehyde cross-linking which revealed that the protein is homodimer in nature. This is a unique characteristic among known eukaryotic GKs. GMP and ATP served as the most effective phosphate acceptor and donor, respectively. Recombinant BmGK utilized both GMP and dGMP, as substrates showing Km values of 30 and 38 μm, respectively. Free Mg+2 (un-complexed to ATP) and GTP play a regulatory role in catalysis of BmGK. The enzyme showed higher catalytic efficiency as compared with human GK and showed ternary complex (BmGK-GMP-ATP) formation with sequential substrate binding. The secondary structure of BmGK consisted of 45% α-helices, 18% β-sheets as revealed by CD analysis. Homology modelling and docking with GMP revealed conserved substrate binding residues with slight differences. Differences in kinetic properties and oligomerization of BmGK compared with human GK can provide the way for design of parasite-specific inhibitors.

Trypanosoma equiperdum is a protozoan parasite responsible for causing Dourine, a debilitating neglected veterinary disease, found worldwide affecting equids. It is the only pathogenic trypanosome species that does not require an invertebrate vector for transmission, thus being passed from animal to animal via coitus. At present, there is no officially recognized form of chemotherapeutic treatment and therefore all confirmed (or suspected) cases of infected animals must be slaughtered immediately. For many global communities and farming populations, which rely heavily on their animals for their livelihood, such stringent regulations can seriously enhance the socio-economic problems attributing to poverty. Two reference drugs, together with 37 novel diamidine compounds were tested in vitro using a 72 h drug sensitivity assay to determine their efficacy against two axenically adapted T. equiperdum strains. Further in vivo investigations in mouse models of infection against 4 ‘true’ T. equiperdum strains were performed using the 17 most active diamidines. Single bolus doses of 10 mg kg−1, given i.p. were administered to NMRI mice infected with one of the 4 T. equiperdum strains. The results obtained from this study show that experimentally T. equiperdum can indeed be effectively treated with chemotherapy using in vivo mouse models of infection.

Despite the profound health implications of Necator americanus infection in humans, many aspects of its interaction with the host immune system are poorly understood. Here we investigated the early events at the interface of N. americanus larvae (L3) and human dendritic cells (DCs). Our data show that co-culturing DCs and the larvae trigger ex-sheathing of hookworms rapidly where a majority of DCs are sequestered onto the larval sheath allowing the ex-sheathed larvae to migrate away unchallenged. Intriguingly, DCs show negligible interaction with the ex-sheathed larvae, alluding to differences between the surface chemistry of the larva and its sheath. Furthermore, blocking of two key C-type lectin receptors on DC surface (i.e. DC-SIGN and mannose receptor) resulted in inhibition of ex-sheathing process and DC sequestration, highlighting the importance of C-type lectins on DCs in the induction of the ex-sheathing. Analyses of DC phenotype and cytokine profile after co-culture with the N. americanus larvae showed an immature phenotype as evidenced by the low expression of the maturation markers and cytokines. These data provide new insights into early events at the interface of human DCs and N. americanus larvae and could explain how L3 evade immune recognition upon initial interaction with DCs.

The prevalence and community composition of haemoparasites can substantially differ among avian host populations, which may lead to different selection pressures. Therefore, information about these parameters is crucial for understanding, e.g. the inter-population variation in host life history traits. Here, we molecularly screened a population of a long-distance migrant, the pied flycatcher Ficedula hypoleuca, from central Poland for the presence of three genera of blood parasites: Haemoproteus, Plasmodium and Trypanosoma. The infection rate in this population was the highest for haemosporidians (86·8%) and one of the highest for trypanosomes (39·7%) among the thus far screened breeding populations of this species. The haemosporidian community was composed of six Haemoproteus/Plasmodium lineages, and the trypanosome community – 4 species and a parasite assigned to genus level. Trypanosomes were dominated by T. culicavium, a recently described species, corroborating the prediction that insectivorous songbirds are vertebrate hosts of this parasite. Host sex and age did not explain variation in infection incidence except for the higher trypanosome infection rates in males. A comparison of the study population with three other breeding populations previously screened molecularly for haemosporidians showed some geographic differences. This study confirms the importance of examining local parasite communities across a host distribution range.

Tritrichomonas foetus is a protist that causes bovine trichomoniasis and presents a well-developed Golgi. There are very few studies concerning the Golgi in trichomonads. In this work, monoclonal antibodies were raised against Golgi of T. foetus and used as a tool on morphologic and biochemical studies of this organelle. Among the antibodies produced, one was named mAb anti-Golgi 20.3, which recognized specifically the Golgi complex by fluorescence and electron microscopy. By immunoblotting this antibody recognized two proteins with 60 and 66 kDa that were identified as putative beta-tubulin and adenosine triphosphatase, respectively. The mAb 20.3 also recognized the Golgi complex of the Trichomonas vaginalis, a human parasite. In addition, the nucleotide coding sequences of these proteins were identified and included in the T. foetus database, and the 3D structure of the proteins was predicted. In conclusion, this study indicated: (1) adenosine triphosphatase is present in the Golgi, (2) ATPase is conserved between T. foetus and T. vaginalis, (3) there is new information concerning the nucleic acid sequences and protein structures of adenosine triphosphatase and beta-tubulin from T. foetus and (4) the mAb anti-Golgi 20.3 is a good Golgi marker and can be used in future studies.

Transforming growth factor beta (TGF-β) cytokine is involved in Chagas disease establishment and progression. Since Trypanosoma cruzi can modulate host cell receptors, we analysed the TGF-β receptor type II (TβRII) expression and distribution during T. cruzi – cardiomyocyte interaction. TβRII immunofluorescent staining revealed a striated organization in cardiomyocytes, which was co-localized with vinculin costameres and enhanced (38%) after TGF-β treatment. Cytochalasin D induced a decrease of 45·3% in the ratio of cardiomyocytes presenting TβRII striations, demonstrating an association of TβRII with the cytoskeleton. Western blot analysis showed that cytochalasin D significantly inhibited Smad 2 phosphorylation and fibronectin stimulation after TGF-β treatment in cardiomyocytes. Trypanosoma cruzi infection elicited a decrease of 79·8% in the frequency of cardiomyocytes presenting TβRII striations, but did not interfere significantly in its expression. In addition, T. cruzi-infected cardiomyocytes present a lower response to exogenous TGF-β, showing no enhancement of TβRII striations and a reduction of phosphorylated Smad 2, with no significant difference in TβRII expression when compared to uninfected cells. Together, these results suggest that the co-localization of TβRII with costameres is important in activating the TGF-β signalling cascade, and that T. cruzi-derived cytoskeleton disorganization could result in altered or low TGF-β response in infected cardiomyocytes.