The reproductive kinetics of Protopolystoma xenopodis primary and secondary infections in Xenopus laevis were monitored in a 3-year study. Thirty-five naïve, lab-raised, full-sib X. laevis from 1 spawning were each exposed to 30 P. xenopodis eggs. The course of infections at 20 °C was monitored by screening isolated hosts for parasite egg production. Ninety-four percent of toads supported the development of gravid parasites. Infections became patent 9–19 weeks p.i., lasted 3–30 months and produced estimated totals of 1–7152 eggs/host. Variation in primary infection characters was discontinuous: a subgrouping of hosts (16%) was characterized by extended infection duration and low reproductive rate. In order to test the effect of long-term infection history on a subsequent challenge, each host was re-exposed to P. xenopodis infective stages (30 eggs/host) 6 months after the loss of its original infection. Establishment of patent infection was significantly lower (15%), and pre-patent period (12–28 weeks) longer, than in primary infections of the same hosts, and than in concurrently exposed naïve controls (contemporary full-sibs of the primary/secondary infection group, maintained in parallel; n = 28). There was no relationship between primary infection characteristics and secondary infection outcome. Overall reproductive output per initial infective stage for the primary exposure exceeded that for the secondary exposure by a ratio of 15[ratio ]1. Results suggest that primary infection with P. xenopodis can elicit strong, long-term protective immunity against re-infection in X. laevis.

A procedure is described for the isolation of sub-cellular fractions from bloodstream forms of Trypanosoma brucei. The method leaves intact most of the nuclei, mitochondria and microbodies. All the fractions have been chemically characterized and tested for 10 enzymatic markers. About 5% of total cell protein was isolated as a microsomal fraction containing mostly plasma membranes and endoplasmic reticulum vesicles. Plasma membranes were purified by high-speed centrifugation on magnesium-containing Dextran, and on linear sucrose-density gradients. The yield of membranes was approximately 0·3% of the total cell protein. The purified material had a sucrose density of 1·14 g/cm3 and consisted of smooth vesicles. Specific activity of the membrane markers Na+, K+, ouabain-sensitive ATPase and adenylate cyclase were 26-and 20-fold higher, respectively, than in total cells. Neither DNA nor RNA was detected. The sum of the cholesterol and phospholipid content was 0·99 mg/mg protein. The cholesterol/phospholipid molar ratio was 1 : 2.

Malaria is one of the most serious infectious diseases with ~250 million clinical cases annually. Most cases of severe disease are caused by Plasmodium falciparum. The blood stage of Plasmodium parasite is entirely responsible for malaria-associated pathology. Disease syndromes range from fever to more severe complications, including respiratory distress, metabolic acidosis, renal failure, pulmonary oedema and cerebral malaria. The most susceptible population to severe malaria is children under the age of 5, with low levels of immunity. It is only after many years of repeated exposure, that individuals living in endemic areas develop clinical immunity. This form of protection does not result in sterilizing immunity but prevents clinical episodes by substantially reducing parasite burden. Naturally acquired immunity predominantly targets blood-stage parasites and it is known to require antibody responses. A large body of epidemiological evidence suggests that antibodies to Plasmodium antigens are inefficiently generated and rapidly lost in the absence of ongoing exposure, which suggests a defect in the development of B cell immunological memory. This review summarizes the main findings to date contributing to our understanding on cellular processes underlying the slow acquisition of humoral immunity to malaria. Some of the key outstanding questions in the field are discussed.

One of the ways of human parasitic infection is the accidental ingestion of vegetables contaminated with parasites, which represents a major human health hazard. This non-exhaustive review aims to evaluate studies carried out on five types of vegetables (lettuce, parsley, coriander, carrot and radish) since 2000, particularly the methods used for recovery, concentration, detection and identification of protozoan parasites such as Toxoplasma gondii, Giardia duodenalis and Cryptosporidium spp., and the results of each work. Various studies have determined the presence of pathogenic parasites in fresh vegetables with different rates; this variation in rate depends particularly on the detection method used which is related to each parasite and each vegetable type. The variation in parasitic prevalence in food could be due to different factors such as the geographical location, the size of analysed samples and the methods used for parasite detection.

Worm free lambs 2–4 months old were given a single infection of 50 000 Haemonchus contortus larvae. The changes in packed red cell volume were compared with those of uninfected sheep and the infected sheep were divided into two groups according to the rate of change of packed cell volume. In group 1, packed cell volume fell sharply from the 10th day of infection and the sheep eventually died. In group 2, the fall was less marked and the packed cell volume returned to normal after about 40 days. The rates of increase of parasite body length and dry weight were compared for the two groups. Parasites in group 1 sheep grew more slowly than their group 2 contemporaries. Accurate planning of metabolic studies on H. contortus was made possible by using this information to predict the course of an infection and the size of the worms. Parasite development was also measured in sheep for which the packed cell volume was not recorded. Computer analysis showed that adult worm length increase did not follow a simple growth pattern starting from the last ecdysis, but consisted of a rapid elongation followed by a simple negative exponential increase. Dry weight increase also followed a negative exponential during the second phase. Measurements of dry weight as a percentage of wet weight indicated that the rapid elongation was possibly linked with cell enlargement by the uptake of water.

A reproducible and standardized assay for measuring the cytoadherence of knobby Plasmodium falciparum-infected red cells to amelanotic melanoma cells was developed. Adhesion was dependent on temperature, haematocrit, and parasitaemia. Addition of EDTA to the binding medium reduced adhesion. Removal of protease-sensitive molecules on the surface of the infected cell abolished cytoadherence, whereas removal of carbohydrate residues by treatment of cells with neuraminidase or galactosidase promoted adhesion. Calcium, magnesium, fibrinogen or fibronectin in the medium had no effect on adhesion nor was there any enhancement of adhesion by pre-loading infected cells with calcium. Serum was essential for good adhesion. Adhesion was species specific for target cells; human endothelial or amelanotic melanoma cells were suitable target cells whereas bovine cells were not. The amelanotic melanoma cell could be formalin-fixed and still retain its adhesion properties. The binding properties of formalin-fixed amelanotic melanoma cells were not identical to those of endothelial or unfixed target cells.

When Prof. E. Hindle took over the direction of the Biological Laboratory of the Egyptian Government School of Medicine, Cairo, he found there a large collection of parasitic worms, most of which had been accumulated by Prof. Looss while in charge of the Laboratory for a number of years prior to the Great War. As a large proportion of this material was unnamed, and as the named portion of it contained many specimens possibly of considerable value as original types or co-types, which had been in a neglected condition for some time, it was thought advisable to send duplicates of the collection to England, in order that part of it might be kept there in safer custody. It was also felt to be desirable that arrangements should be made for the examination and determination of the contents of the unnamed part of the collection.

Eggs of three species of Diclidophora were incubated in alternating 12 h periods of light and darkness at 13°C. Eggs of D. merlangi collected at Arbroath hatched during the illumination period with most larvae being recovered in the first 4–6 h; some evidence of a seasonal difference in hatching of these eggs was found. Eggs of D. merlangi collected at Plymouth hatched with a peak of larval recovery in the 2 h period before the light came on. Eggs of D. luscae hatched over ‘dusk’ while those of D. denticulata hatched after the light was switched off. Neither mechanical disturbance nor the proximity of host tissue caused hatching in D. merlangi or D. luscae. Observations on the behaviour of the host fishes suggest that the hatching rhythms are adapted to specific host behaviour patterns.

Plasmodium vinckei chabaudi (which has a 24 h cycle with high synchronicity) has a peak of schizogony in the early morning at about 02.00–04.00. This cycle is orientated to the cycle of the host (mouse or thamnomys). The male gametocytes become ripe for exflagellation in a well-marked wave from 22.00 to 08.00 with peak about 00.00–04.00. This timing presumably corresponds to the hours when the vector mosquitoes are most likely to suck blood, and it is adapted to facilitate transmission. The male gametocytes remain ripe for exflagellation for a mean period of about 6 h and then apparently degenerate.

The virulence of the malaria parasite Plasmodium falciparum is due, in part, to its ability to cytoadhere in deep vascular beds. Our inability to quantify the load of sequestered parasites hampers our understanding of the pathophysiological mechanisms involved in disease progression and complicates diagnosis. In this study we evaluate potential biochemical markers of sequestered load by comparing them with estimates of the sequestered load from a statistical model fitted to longitudinal patterns of peripheral parasite densities in a series of 22 patients with severe Plasmodium falciparum malaria. The markers comprised the host factors: haematocrit, circulating host DNA, sTNF-R75 and parasite derived products HRP2, pLDH, pigments and circulating parasite DNA. We investigated the suitability of these markers in determining sequestered loads in patients on quinine treatment. Observed peripheral parasitaemia, plasma levels of sTNF-R75 and circulating parasite DNA were most strongly correlated with estimates of sequestered loads on admission. However the dynamics of both sTNF-R75 and circulating parasite DNA during follow-up were very different from those of the estimated sequestered mass. These analyses suggest that none of the markers gave reliable estimates of the current sequestered load, though they may reflect the history of infection. Longitudinal analyses are needed that allow for the clearance rates of the marker molecules and for variations between hosts in the history of parasitaemia.

An artificial skin of hardened gelatine was used to examine the factors affecting penetration of skin by cercariae of the bird schistosome, Austrobilharzia terrigalensis.

The percentage of cercariae able to penetrate through a gelatine membrane was increased by a factor of 3–4 by coating the membrane with a thin layer of lipid collected from the surface of chicken skin.

The free sterol fraction, isolated from chicken skin surface lipid by thin-layer chromatography, stimulated penetration to the same extent as whole skin lipid.

Cholesterol was detected in the sterol fraction by mass spectrometry and pure cholesterol had the full stimulating effect on cercarial penetration.

Skin lipid, from which free sterols had been removed, lost the stimulatory effect on cercariae, but full activity was recovered by adding cholesterol to the sterol-free lipid. Fractions of skin lipid containing free fatty acids or triglycerides, wax esters and sterol esters similarly failed to stimulate penetration.

These results establish that penetration of A. terrigalensis cercariae is greatly stimulated by the free sterols present in the surface lipid of chicken skin but cholesterol may not be the only active sterol. Cholestanol and the plant sterols campesterol and β-sitosterol were also detected in chicken skin surface lipid. These sterols were not tested for activity on cercariae because samples free from cholesterol could not be obtained.

Some cercariae were able to penetrate plain gelatine membranes not coated with cholesterol but small amounts of free sterol were detected in the gelatine itself. This sterol could not be completely removed by prolonged solvent extraction and consequently it is not known whether any cercariae are able to penetrate in the complete absence of sterols.

Temperature had a marked effect on penetration of cercariae; lowering the temperature from 40 to 25 °C reduced the number of successful penetrants by a factor of 4.

I am grateful to Miss V. Bowen for excellent technical assistance and to Dr J. McLeod of the Research School of Chemistry, Australian National University, who performed and interpreted the mass spectrometry. This investigation was partly supported by a research grant from the World Health Organization.

Myxosporean parasites are significant parasites of fishes not only for their apparent high diversity but also for their potential impact on fish health and/or marketability. Regardless, our knowledge of most myxosporeans, especially those found in elasmobranch hosts, is superficial. A study of multivalvulidan diversity in a range of elasmobranchs from Queensland, Western Australia and the Northern Territory (Australia) was conducted to address this knowledge gap. Specimens were collected from a total of 3 orders, 9 families and 31 species of elasmobranchs. Myxosporean infections referable to the genus Kudoa were discovered in host muscle and characterized morphologically and genetically. Both small subunit (SSU) and large subunit (LSU) rDNA sequences were used in molecular phylogenetic analyses. Kudoa spp. infected 27 of the 31 species of elasmobranchs examined, representing new records of this parasite genus in 26, of the 27, host species. Kudoids were observed in all 3 orders, and 7 out of the 9 families of elasmobranchs investigated. This paper reports the first 2 multivalvulidan species to be formally described from elasmobranchs, Kudoa hemiscylli n.sp. characterized from Hemiscyllium ocellatum (and 8 other host species) and Kudoa carcharhini n. sp. characterized from Carcharhinus cautus (and 2 other host species). Phylogenetic analyses revealed that kudoids from elasmobranchs form a separate lineage to those of teleosts, but are anchored within the overall kudoid clade.

One of a large number of Lymnaea peregra (Mull.) from a pond near Glasgow was found to be infested with Cercaria diplostomi gasterostei.

C. d. gastrostei has been found to penetrate Gasterosteus aculeatus and develop into a diplostomulum in the retina of the eyes.

Large numbers of G. aculeatus from the pond have been found with Diplostomulum gasterostei occurring in the retina.

Infested heads of fish were fed to domestic ducklings and pigeons, and adult stages of a worm considered to be a new species and named D. gasterostei were obtained.

Morphological details of the cercaria, diplostomulum and the adult worm are described.

The author wishes to thank Dr C. A. Hopkins for reading and criticising the manuscript, Dr D. A. Erasmus and Dr G. Rees for valuable suggestions. My thanks are due also to the technical staff of the Department who assisted me in the collection of snails and fishes, and to Mr Francis Deen who did the sectioning of the specimens. This work was in part supported by a grant from Shell Oil Company (London).

TaqMan minor groove binder probes were evaluated as to their suitability for the real-time allelic discrimination of the β-tubulin codon 200 TTC/TAC single nucleotide polymorphism in cyathostomin species. Amplification of titrated cloned full-length β-tubulin cDNA revealed that the TaqMan minor groove binder PCR is capable of specifically detecting as few as 10 copies. Testing of DNA from single adult and larval stages of several different species of cyathostomin allowed reproducible genotyping of individual worms. Using the real-time PCR approach, the throughput of samples was considerably increased compared with conventional post-PCR readout procedure. Only 7·8% homozygous TAC L3 were found among 102 L3 which were genotyped from phenotypically BZ-resistant small strongyle populations. The percentages of the homozygous TTC and heterozygous TTC/TAC were 41·3% and 50·9%, respectively. This resulted in a total TAC-allele percentage of only 33·3%. These findings correspond to data obtained by genotyping of an experimentally selected BZ-resistant cyathostomin population. It is concluded that the β-tubulin codon 200 polymorphism is not the sole mechanism involved in the process of BZ resistance in cyathostomins.

Entobdella diadema produces spermatophores from a pair of follicular glands lying one on each side of the body alongside the main nerve cord, and the secretion of these glands is stored in a two-chambered reservoir within the cirrus-sac.

The vagina is made up of two regions, a wide distal part to receive, accommodate and digest the spermatophores, and a proximal narrow part connected through the vitelline reservoir with the oviduct.

E. diadema Monticelli, 1901, is shown to be distinct from E. bumpusii Linton, 1900, and has been recorded from two new hosts, Dasyatis pastinaca at Plymouth, and Myliobatis aquila at Naples.

We wish to thank the directors and staffs at the Plymouth Marine Laboratory and at the Stazione Zoologica, Naples, for providing excellent facilities for the above studies to be carried out. The work was assisted by a grant to one of us (J. L.) from the Trustees of the Browne Fund, to whom grateful acknowledgement is made.

A cross-sectional sero-epidemiological study was performed in Magoda, Tanzania, an area where malaria is holoendemic. Blood samples were collected from children (1–4 years) and tested for IgG antibody reactivity against 2 recombinant protein fragments of Plasmodium falciparum Rhoptry-Associated Protein-1 (rRAP-1). The data were related to the prevalence of malarial disease and single P. falciparum or mixed Plasmodium infections. Fever ([ges ]37·5 °C) in combination with parasite densities >5000/μl were used to distinguish between children with asymptomatic malaria infections and those with acute clinical disease. Furthermore, C-reactive protein (CRP) was applied as a surrogate marker of malaria morbidity. The prevalence of Plasmodium infections was 96·0%. Eleven children were defined as clinical malaria cases, all with single P. falciparum infections. The density of P. falciparum was significantly lower in children with mixed Plasmodium infections compared to those with single P. falciparum infections. Children with asymptomatic P. falciparum infections had higher IgG reactivities to rRAP-1, compared to IgG reactivities of children with malarial disease. Children with mixed Plasmodium infections generally showed elevated IgG reactivity to rRAP-1, when compared to children with single P. falciparum infections. The possible relationship between mixed species infections, clinical outcome of the disease and antibody responses to RAP-1 is discussed.

Chagas disease (CD) is a neglected parasitic condition endemic in the Americas caused by Trypanosoma cruzi. Patients present an acute phase that may or not be symptomatic, followed by lifelong chronic stage, mostly indeterminate, or with cardiac and/or digestive progressive lesions. Benznidazole (BZ) and nifurtimox are the only drugs approved for treatment but not effective in the late chronic phase and many strains of the parasite are naturally resistant. New alternative therapy is required to address this serious public health issue. Repositioning and combination represent faster, and cheaper trial strategies encouraged for neglected diseases. The effect of imatinib (IMB), a tyrosine kinase inhibitor designed for use in neoplasias, was assessed in vitro on T. cruzi and mammalian host cells. In comparison with BZ, IMB was moderately active against different strains and forms of the parasite. The combination IMB + BZ in fixed-ratio proportions was additive. Novel 14 derivatives of IMB were screened and a 3,2-difluoro-2-phenylacetamide (3e) was as potent as BZ on T. cruzi but had low selectivity index. The results demonstrate the importance of phenotypic assays, encourage the improvement of IMB derivatives to reach selectivity and testify to the use of repurposing and combination in drug screening for CD.

In Part I the musculature of the internal organs of the male blow-fly, C. erythrocephala, testis, vas efferens, accessory gland, vas deferens, and ejaculatory sac and duct, is described, together with the movements observed in them and the passage of secretions down the system. It is suggested that the testicular and accessory gland secretions are ejaculated separately. The musculature, mechanics and co-ordinated movements of the external organs and associated structures are also described.

In Part II the generative organs of the female, except the ovaries, with special reference to their musculature and movements, and the musculature of the ovipositor are described and compared with those of M. domestica. It is suggested that the male accessory gland secretion is kept separate from the testicular secretion, the former passing into the lateral sacs of the female and the latter into the spermathecae.

In Part III the relation of the parts of the two sexes during coitus is discussed.

The uptake of thiamine by isolated second-generation schizonts of Eimeria tenella and by host intestinal cells was found to consist of two components. One was passive and the other apparently an active process. The kinetic constants of the latter were compared in host (Km = 0·36 μm) and parasite (Km = 0·07 μm) and found to be significantly different. Both systems were competitively inhibited by amprolium but showed different affinities for the drug. (Host Ki = 326 μm; parasite Ki = 7·6 μm). Further differences were observed in schizonts of a drug-resistant line of E. tenella. These findings are discussed in terms of the inhibition of thiamine uptake being the basis of the anticoccidial activity of amprolium.

We investigated the diversity of Bartonella in Apodemus agrarius, an important rodent of peri-domestic habitats, which has spread into Europe in the past 1000 years. Spleen samples of 344 A. agrarius from Eastern Slovakia were screened for the presence of Bartonella spp. using 16S–23S rRNA internal transcribed spacer region and bacteria were detected in 9% of rodents. Based on sequencing of three housekeeping genes (gltA, rpoB and groEL) Bartonella genotypes were ascribed to the species typical for mice and voles: B. grahamii, B. taylorii and B. birtlesii. However, the study also confirmed presence of genotypes belonging to the B. clarridgeiae/B. rochalimae clade, and the B. elizabethae/B. tribocorum clade, which are not commonly found in woodland rodents. In addition, a potential recombination event between these two genotypes was noted, which highlights an important role of A. agrarius in shaping Bartonella diversity and evolution.