Midgut proteases contribute to the success or failure of Plasmodium infection of the mosquito. This paper examines the reciprocal effect of Plasmodium yoelii nigeriensis on midgut trypsin, chymotrypsin, aminopeptidase and carboxypeptidase in the mosquito Anopheles stephensi. The total protein ingested and the rate of protein digestion were unaffected by the parasite, but more protein was ingested at the first than the second bloodmeal. All peptidases were unaffected by the presence of the parasite during the first gonotrophic cycle, when ookinetes were penetrating the midgut. In the second gonotrophic cycle, trypsin and chymotrypsin were unaffected by growing oocysts, but aminopeptidase activity was reduced in the midguts of infected mosquitoes. Chymotrypsin activity was depressed and aminopeptidase activity elevated during the second gonotrophic cycle. Plasmodium infection has a negligible effect on bloodmeal digestion and does not limit the availability of the protein for egg production. The significance of changes in aminopeptidase activity when oocysts are present is discussed.

The diversity and biology of Cryptosporidium that is specific for rats (Rattus spp.) are not well studied. We examined the occurrence and genetic diversity of Cryptosporidium spp. in wild brown rats (Rattus norvegicus) by microscopy and polymerase chain reaction (PCR)/sequencing targeting the small subunit rDNA (SSU), actin and HSP70 genes. Out of 343 faecal samples tested, none were positive by microscopy and 55 were positive by PCR. Sequence analysis of SSU gene revealed the presence of Cryptosporidium muris (n = 4), C. andersoni (n = 3), C. ryanae (n = 1), C. occultus (n = 3), Cryptosporidium rat genotype I (n = 23), Cryptosporidium rat genotype IV (n = 16) and novel Cryptosporidium rat genotype V (n = 5). Spherical oocysts of Cryptosporidium rat genotype I obtained from naturally-infected rats, measuring 4.4–5.4 μm × 4.3–5.1 μm, were infectious to the laboratory rats, but not to the BALB/c mice (Mus musculus) nor Mongolian gerbils (Meriones unguiculatus). The prepatent period was 3 days post infection and the patent period was longer than 30 days. Naturally- and experimentally-infected rats showed no clinical signs of disease. Percentage of nucleotide similarities at the SSU, actin, HSP70 loci between C. ratti n. sp. and the rat derived C. occultus and Cryptosporidium rat genotype II, III, IV, and V ranged from 91.0 to 98.1%. These genetic variations were similar or greater than that observed between closely related species, i.e. C. parvum and C. erinacei (93.2–99.5%). Our morphological, genetic and biological data support the establishment of Cryptosporidium rat genotype I as a new species, Cryptosporidium ratti n. sp.

The development of the free-living infective larvae of a homogonic strain Strongyloides ratti is described.

The larvae develop only between 15 and 34 °C. Transfer experiments show the temperature block to be in the preparation for the second moult.

Within the temperature range 15–34 °C, increasing the temperature speeds up the rate of development of all the larval stages equally, the Q10 for development being 2·5.

The maximum percentage development occurs at 20 °C. The percentage development is highest in faeces–peat culture (95% development at 20 °C), whilst the percentage development in charcoal and vermiculite cultures is about the same (75% development at 20 °C.).

Larvae grown on charcoal cultures are larger than those grown on vermiculite, which are larger than those grown on peat. No significant difference was found in the length:oesophagus and length:width ratios or in the variability of larvae grown at different temperatures or on different culture media.

Different worm densities in the cultures of from 2000 to 10000 larvae per g of culture did not affect either the size of the infective larve or the percentage development.

The optimum temperature for survival is 15 °C. Worms grown at 20 °C lived longer than worms grown at any other temperature. There was no evidence of temperature adaptation by the larvae.

The infective larvae are positively thermotactic, and show maximum activity at 37 °C.

I should like to thank my supervisor, Dr Tate, for his advice and encouragement. The work was carried out during the tenure of a Medical Research Council Scholarship.

Evidence for the existence of 2 first intermediate host-specific sibling species of the 45 collar-spined echinostome Echinoparyphium recurvatum is presented. Experimental studies on their life-cycles were carried out under controlled laboratory conditions. The two entities were found to be morphologically indistinguishable in all major respects, yet they exhibited distinct biological characteristics. One of the sibling species utilizes the freshwater lymnaeid pulmonate snail Lymnaea peregra as first intermediate host, and the adults occur in the anterior small intestine of the wildfowl experimental definitive host Anas platyrhynchos. The other utilizes the freshwater mesogastropod prosobranch snail Valvata piscinalis as first intermediate host, and the adults occur in the posterior small intestine and rectum of A. platyrhynchos. The existence of the two sibling species in sympatry may be explained by the fact that the life-cycles of the two forms represent two distinct cycles of transmission serving to reduce excessive competition between them.

Dirofilaria immitis is an important filarial parasite of dogs and cats, and a useful model for human filariasis. Current diagnostic tests for heartworm infection in animals rely on the presence of fecund female worms (usually found 6·5 months post-infection or later) and therefore fail to detect pre-patent infections. Putative pepsin inhibitors from 2 filarial parasites of humans namely Onchocerca volvulus (Ov33, Oc3.6, OvDSB) and Brugia malayi (Bm33), have been shown to be useful in diagnosis of onchocerciasis and lymphatic filariasis, respectively. Previous studies have suggested that a homologue exists in D. immitis (DiT33), which may have potential in diagnosis of heartworm infection. In this study, the isolation and characterization of a cDNA clone encoding DiT33 is described.‡ This cDNA contains 12 bases of the nematode-specific 22 nucleotide spliced leader sequence and encodes a 26·4 kDa-protein with a high level of similarity (87–89%) to other filarial members of the family. DJT33 was over-expressed in E. coli as a fusion with the maltose-binding protein and serological analysis was performed using a panel of clinically defined dog sera. The findings of this study indicate that DiT33 is a promising antigen for the early detection of D. immitis and may be a valuable tool in the control and management of heartworm infection.

Drugs that kill or inhibit the sexual stages of Plasmodium in order to prevent transmission are important components of malaria control programmes. Reducing gametocyte carriage is central to the control of Plasmodium falciparum transmission as infection can result in extended periods of gametocytaemia. Unfortunately the number of drugs with activity against gametocytes is limited. Primaquine is currently the only licensed drug with activity against the sexual stages of malaria parasites and its use is hampered by safety concerns. This shortcoming is likely the result of the technical challenges associated with gametocyte studies together with the focus of previous drug discovery campaigns on asexual parasite stages. However recent emphasis on malaria eradication has resulted in an upsurge of interest in identifying compounds with activity against gametocytes. This review examines the gametocytocidal properties of currently available drugs as well as those in the development pipeline and examines the prospects for discovery of new anti-gametocyte compounds.

Trypanosomatids of the genera Angomonas and Strigomonas (subfamily Strigomonadinae) have long been known to contain intracellular beta-proteobacteria, which provide them with many important nutrients such as haem, essential amino acids and vitamins. Recently, Kentomonas sorsogonicus, a divergent member of Strigomonadinae, has been described. Herein, we characterize the genome of its endosymbiont, Candidatus Kinetoplastibacterium sorsogonicusi. This genome is completely syntenic with those of other known Ca. Kinetoplastibacterium spp., but more reduced in size (~742 kb, compared with 810–833 kb, respectively). Gene losses are not concentrated in any hot-spots but are instead distributed throughout the genome. The most conspicuous loss is that of the haem-synthesis pathway. For long, removing haemin from the culture medium has been a standard procedure in cultivating trypanosomatids isolated from insects; continued growth was considered as an evidence of endosymbiont presence. However, we demonstrate that, despite bearing the endosymbiont, K. sorsogonicus cannot grow in culture without haem. Thus, the traditional test cannot be taken as a reliable criterion for the absence or presence of endosymbionts in trypanosomatid flagellates. It remains unclear why the ability to synthesize such an essential compound was lost in Ca. K. sorsogonicusi, whereas all other known bacterial endosymbionts of trypanosomatids retain them.

Although co-infection is the norm in most human and animal populations, clinicians currently have no practical tool to assist them in choosing the best treatment strategy for such patients. Given the vast range of potential pathogens which may co-infect the host, obtaining such a practical tool may seem an intractable problem. In ecology the joint concepts of functional groups and guilds have been used to conceptually simplify complex ecosystems, in order to understand how their component parts interact and may be manipulated. Here we propose a mechanism by which to apply these concepts to pathogen co-infection systems. Further, we describe how these groups could be incorporated into a mathematical modelling framework which, after validation, could be used as a clinical tool to predict the outcome of any particular combination of pathogens co-infecting a host.

The main aim of this study is to overview the past and present situations of human taeniases and cysticercosis in Indonesia and including future perspectives. Through joint projects from 1996, we have confirmed the occurrence of Taenia saginata (beef tapeworm) in Bali, of Taenia solium (pork tapeworm) mainly in Papua and sporadically in Bali, and of Taenia asiatica in North Sumatra. These taeniases were caused through eating uncooked pork and pig viscera for T. solium and T. asiatica, respectively, and beef for T. saginata. The distribution of these tapeworms in Indonesia is basically highly restricted by the traditional cultural and religious backgrounds in each island. T. saginata is relatively common in Bali although people consume pork ‘lawar’ more than beef ‘lawar’. Taeniases due to T. saginata or T. asiatica and T. solium and cysticercosis due to T. solium have also been sporadically reported in some other islands. Among these species, T. solium is exceptional since humans can be infected not only by larval stages (cysticerci) in pork but also by eggs released from human tapeworm carriers. Cysticercosis has been confirmed in Indonesia in humans, pigs and even dogs.

The total lipids and the amount of neutral lipids of male and female N. brasiliensis, from 7-day and 15-day infections, have been studied quantitatively. There is an increase in total lipids from 10·9 ± 2 to 15·5 ± 2 % in females and an increase from 8·4 ± 1 to 14·6 ± 2 % in males. In both sexes there is a rise in cholesterol and trigylceride content and a fall in free fatty acid content. These changes are discussed with reference to the development of immunity of the host to this nematode.

I wish to thank Mrs B. Fisher, Mr A. W. Pearson and Mr N. Sykes for assistance during this work. These results form part of a programme of work supported by the A.R.C. while I was at the Molteno Institute, University of Cambridge, and continued at the Houghton Poultry Research Station. I wish to thank Dr P. Tate for accommodation at the Molteno Institute during the early part of this work.

By a variety of microscopical techniques, an initial survey has been made of the functional organization of ovarian balls from mature, inseminated female Moniliformis dubius and Polymorphus minutus. On the basis of evidence from observations made with the transmission electron microscope, we have concluded that the ovarian ball consists of three components; these are two separate, multinucleate syncytia and a cellular zone. The inner region is considered to be an oogonial syncytium from which the germ-line cells arise to form the cellular zone. The oogonial syncytium and the cellular elements are embedded in a supporting syncytium which also forms the boundary of the ovarian ball. Details of the ultrastructure of these components, observations on fertilization and a hypothesis to account for the main events occurring in an acanthocephalan ovarian ball are also presented.

Acanthocephalans are dioecious parasites which become sexually mature in the alimentary tract of vertebrates. After insemination, individual female worms begin to release embryonated eggs at rates which have been estimated to vary, on average, from 2000 per day in the case of Polymorphus minutus (Crompton & Whitfield, 1968) to 260000 per day in the case of Macracanthorhynchus hirudinaceus (Kates, 1944). Irrespective, however, of the species of acanthocephalan and the number of eggs produced, a most interesting mode of ovarian development and multiplication is believed to exist (see Bullock, 1969). Within the female worm, the primordial ovarian tissue gives rise to separate ovaries which are often termed ovarian balls. These in turn give rise to more ovarian balls and the process continues until large numbers have been formed. The ovarian balls of a mature female have no permanent attachment to the tissues of the worm. They are contained in the fluid of the body cavity either freely, or loosely constrained in large membranous chambers known as ligament sacs.

Few investigations have been made into the structure and cytology of the ovarian balls of the Acanthocephala. Early observations on the histology of the germ-line constituents of ovarian balls were made by Hamann (1891), Kaiser (1893) and Meyer (1928), each of whom suggested that some of the developing oogonial stages were arranged in a syncytial manner. In a study of the embryology of P. minutus. Nicholas & Hynes (1963) interpreted preparations of ovarian balls, which had been subjected to the Feulgen reaction, as indicative of syncytial tissue in the central region of the ball. They also recorded their uncertainty about whether an ovarian ball has a bounding membrane or not. More recently, Robinson (1964, 1965) described mitotic and meiotic divisions in oogonia and oocytes from ovarian balls of Mac. hirudinaceus and Moniliformis dubius and Stranack (1972) included an electron micrograph of part of an ovarian ball of Pomphorhynchus laevis in a description of the nature of the egg envelopes of that species. In this paper, we present a general survey, based largely on evidence obtained with the electron microscope, of the functional organization of the ovarian balls of mature M. dubius and P. minutus.

Plasmodium falciparum parasites use multiple ligand-receptor interactions to invade human erythrocytes. Variant expression levels of members of the PfRh and PfEBA ligand families are associated with the use of different erythrocyte receptors, defining invasion pathways. Here we analyse a major polymorphism, a large sequence deletion in the PfRh2b ligand, and erythrocyte invasion profiles in uncultured Senegalese isolates. Parasites vary considerably in their use of sialic acid-containing and protease-sensitive erythrocyte receptors for invasion. The erythrocyte selectivity index was not related to invasion pathway usage, while parasite multiplication rate was associated with enhanced use of a trypsin-resistant invasion pathway. PfRh2b protein was expressed in all parasite isolates, although the PfRh2b deletion was present in a subset (~68%). Parasites with the PfRh2b deletion were found to preferentially utilize protease-resistant pathways for erythrocyte invasion. Sialic acid-independent invasion is reduced in parasites with the PfRh2b deletion, but only in isolates derived from blood group O patients. Our results suggest a significant role for PfRh2b sequence polymorphism in discriminating between alternative erythrocyte receptors for invasion and as a possible determinant of virulence.

The CAF01 adjuvant has previously been shown to be safe for human use and to be a potent adjuvant for several vaccine antigens. In the present work, we sought to optimize the Leishmania amazonensis antigens (LaAg) intranasal vaccine in an attempt to enhance the protective immune responses against Leishmania (infantum) chagasi by using the CAF01 association. LaAg/CAF01 vaccinated mice that were challenged 15 days after booster dose with L. (infantum) chagasi showed a significant reduction in their parasite burden in both the spleen and liver, which is associated with an increase in specific production of IFN-γ and nitrite, and a decrease in IL-4 production. In addition, LaAg/CAF01 intranasal delivery was able to increase lymphoproliferative immune responses after parasite antigen recall. These results suggest the feasibility of using the intranasal route for the delivery of crude antigens and of a human-compatible adjuvant against visceral leishmaniasis.

Histological studies using horse-radish peroxidase (HRP) as a marker, injected iontophoretically through a recording electrode, have indicated the origins of potentials encountered upon advancement of the electrode into and beneath the dorsal surface of adult male Schistosoma mansoni. The first potential encountered, having a value of – 51 ± 0·6 mV originates across the outer tegumental membrane. The next potential has a value of — 28 ± 0·6 mV and originates in the muscle masses underlying the tegument. Finally, a potential having the value — 10 ± 0·5 mV originates within the basal lamina and the interstitial fibres and extracellular space surrounding the muscle. Altering ion concentrations in the bathing medium (i.e. high K+, low Na+, zero Ca2+, low Cl−, high Li+) depolarizes all three potentials. External applications of ouabain and the anti-schistosomal, praziquantel, also cause depolarization of the potentials. It appears that the muscle potential and the tegumental potential are primarily K+-dependent. The depolarizing effects of ouabain, LiCl and low Na+ suggest that active transport is important in the maintenance of the muscle potential, just as is the case for the tegumental potential. There appears to be a close correlation between changes in the tegumental, muscle and extracellular space potentials. The correlation between tegument and muscle potential changes might be explained by junctional complexes between tegumental cell bodies and muscle cell bodies.

We are studying early events in the establishment of Trypanosoma brucei in the tsetse midgut using fluorescent trypanosomes to increase visibility. Feeding flies with the lectin-inhibitory sugars D-glucosamine (GlcN) or N-acetyl-glucosamine (GlcNAc) has previously been shown to enhance fly susceptibility to infection with trypanosomes and, as expected, we found that both sugars increased midgut infection rates of Glossina morsitans morsitans with T. brucei. However, GlcNAc did not show the inhibitory effect on salivary gland infection rate reported previously for GlcN. Both sugars significantly slowed the movement of the bloodmeal along the midgut. GlcN also significantly increased the size of the bloodmeal taken and fly mortality. The most surprising finding was that GlcNAc stimulated trypanosome growth not only in the midgut, but also in vitro in the absence of any factor derived from the fly. Thus our direct comparison of the effects of GlcN and GlcNAc on the trypanosome-tsetse interaction has shown that these sugars impact on trypanosome growth and tsetse physiology in different ways and are not interchangeable as suggested in the literature. The sugars cause multiple effects, not restricted solely to the inhibition of midgut lectins. These findings have implications for current models of tsetse susceptibility to trypanosome infection.

In the ideal situation, the development of new amoebicides, or more accurately anti-amoebic compounds which are compounds with activity against Entamoeba histolytica, should initially proceed with the study of parasite-specific metabolic pathways and their inhibition, followed by whole parasite in vitro studies, experimental in vivo models and finally clinical trial. However, there are considerable gaps in our knowledge which will be discussed below, and consequently many investigators consider that empirically selected compounds should be tested experimentally in addition to specifically designed compounds. Before clinical trials can begin, extensive examination of the candidate amoebicide in experimental animals is required in order to investigate possible toxicological hazards.

In addition to inhibiting the amoebic parasite, the drug has to reach the parasite in several different sites in the body, thus there is also a problem of pharmacokinetics and distribution. Prior to the discovery of the nitroimidazole class of amoebicides, the multi-site attack was solved by the use of several drugs, sometimes in sequence during the treatment of an individual case (Powell, 1972). The discovery of the nitroimidazole class of compound changed the situation dramatically and these have shown a clinical and parasitological effect against extra-intestinal and intestinal wall infections. The effect on intralumenal infection (that is mildly symptomatic or asymptomatic infections) of the large intestine is, however, less certain (Finegold, 1977; Spillman, Ayala & Sanchez, 1976).

Although the treatment of amoebiasis appears to be satisfactory at the present time, it is difficult to predict problems which might arise in the future, and therefore it is valuable to continue the pre-clinical development, especially the investigation of parasite metabolism, in order to define parasite-specific points of chemotherapeutic attack.

The chemotherapy of amoebiasis was reviewed comprehensively by Woolf (1963, 1965). The present account of the development of amoebicides therefore starts from the Woolfe reviews.

The metacercaria of Holostephanus lühei Szidat (1936) is described together with comments on both its linear and tissue distribution within the host (Gasterosteus aculeatus L.), host-parasite reaction and numbers of cysts in relation to the size of the fish.

The adult of Holostephanus lühei Szidat (1936) (syn. H. bursiformis Szidat, 1936) obtained experimentally from the duck and pigeon, is re-described in detail. It is noted that this species is amphitypic and it is suggested that H. dubius (Szidat, 1936) (syn. Cyanocotyloides Dubius, Szidat, 1936) should be reduced to synonymy with H. lühei.

Grateful acknowledgement is expressed to Dr G. Dubois for his interest, advice and valuable opinions given in connexion with this study.

The immature stages of Caraphractus cinctus Walker are described.

Four larval stages have been observed which may correspond to three or four instars. In no stage have mandibles been observed.

The first-instar larva is elongated and active. In the second-instar oral lobes are present. In the next stage the larva becomes shapeless and rather transparent. In the last stage the wall of the mid-gut shows conspicuous opaque spots which consist of single cells containing spherical concretions, probably the products of excretion.

These concretions later become free in the gut of the pupa and are discharged in the meconium when the imago emerges from the host egg.

The effects of parasitism on the host eggs of different ages is discussed. In newly laid host eggs all the contents are consumed by the parasitic larvae. In advanced eggs of Agabus the egg of Caraphractus is laid in the mid-gut of the host embryo where the parasitic larva develops, the hard parts of the host larva remaining intact. Parasitism in advanced eggs of Dytiscus is rarely successful.

The larvae of other Mymaridae are discussed and their main features compared. The first-instar larva of Caraphractus does not correspond to any of the previously described types of mymarid larvae.

The seasonal history is described, with the sequence of hosts parasitized throughout the year. From four to six generations of Caraphractus have been bred indoors during the year. The winter is passed as a diapausing prepupa. In some cases diapause may be prolonged. Fertility is high: 121 imagines have been bred from one female of Caraphractus.

I have much pleasure in acknowledging my gratitude to the late Dr W. D. Hincks for his kindness in reading the typescript of this paper: the comments by such a well-known authority on Mymaridae have been most valuable. I am very grateful to Mr O. Bakkendorf for allowing me to reproduce several of his figures of Mymarid larvae from the valuable work he devoted to the biology of Danish hymenopterous egg-parasites. I am also grateful to Miss B. A. Trott of the Commonwealth Institute of Entomology for the help she has so kindly given me with references.

We compared the distribution of the digenean Pholeter gastrophilus in the stomach of 27 harbour porpoises, Phocoena phocoena, 27 striped dolphins, Stenella coeruleoalba, 18 bottlenose dolphins, Tursiops truncatus, and 100 long-finned pilot whales, Globicephala melas. The stomach of these species is composed of 4 chambers of different size, structure and function. In all species, P. gastrophilus was largely restricted to the glandular region of the stomach, but the parasite tended to favour the fundic chamber in bottlenose dolphins and harbour porpoises, the pyloric chamber in pilot whales, and none in striped dolphins. However, predictability at infrapopulation level was generally low, suggesting a weak preference of P. gastrophilus for any of the chambers. Three hypotheses were tested to investigate a common cause for the distribution of P. gastrophilus in all host species, namely, colonization of chambers was (1) sequential, (2) dependent on chamber size, or (3) dependent on the passage time of food through the whole stomach. The latter hypothesis was indirectly tested by assuming, based on previous evidence from other vertebrates, that the greater the size of the stomach and/or the energy content of prey, the greater the delay of food passage. We found no compelling evidence that chamber colonization was sequential, or related to chamber size in any species. However, the distribution of P. gastrophilus was significantly more anteriad when the host species had larger stomachs and, particularly, when hosts fed on prey with higher caloric content. Accordingly, the stomach distribution of P. gastrophilus at this scale seems to be passively driven by features of the diet and digestive physiology of each host species. This study provides a general framework to formulate null hypotheses in future studies on microhabitat choice by parasites.

E. praeox has been isolated in Britain for the first time and is similar in regard to its oocyst size, prepatent time, life-cycle and the development of host immunity to the reports of Tyzzer et al. (1932). Oocysts had mean dimensions of 20·4 × 17·45 μm and the first oocysts were discharged 83½ h after infection.

Oocyst production and life-cycle studies suggest that at least three generations of schizogony precede gametogony and that at least one further generation of schizogony is needed to explain the oocyst reproduction resulting from a small dose of oocysts. Young chickens 1½–3 weeks of age were not such suitable hosts as older chickens (6 weeks) and the reproduction of the parasite was seriously reduced when large numbers (e.g. 105, 106) were used to infect chickens. A high degree of resistance to reinfection occurred after a single exposure to infection.

Sporozoite infections of the caeca or cloaca resulted in the infection of the usual small intestine site; there was no development at the site of inoculation and attempts to infect the embryo allantois did not succeed.

I wish to thank Dr M. Elaine Rose for the surgery required for the caecal infections, for her interest throughout and for help with the manuscript; Dr R. F. Gordon for help with the manuscript and permission to publish and Mr B. J. Millard for skilled technical assistance.