Despite evidence for the existence of interspecific interactions between helminth species, there has been no theoretical exploration of their effect on the distribution of the parasite species in a host population. We use a deterministic model for the accumulation and loss of adult worms of 2 interacting helminth species to motivate an individual-based stochastic model. The mean worm burden and variance[ratio ]mean ratio (VMR) of each species, and the correlation between the two species are used to describe the distribution within different host age classes. We find that interspecific interactions can produce convex age-intensity profiles and will impact the level of aggregation (as measured by the VMR). In the absence of correlated exposure, the correlation in older age classes may be close to zero when either intra- or interspecific synergistic effects are strong. We therefore suggest examining the correlation between species in young hosts as a possible means of identifying interspecific interaction. The presence of correlation between the rates of exposure makes the interpretation of correlations between species more difficult. Finally we show that in the absence of interaction, strong positive correlations are generated by averaging across most age classes.

Reliable detection, discrimination and quantification of parasites are important for host-parasite studies and diagnostics. Microsporidial infections are problematic in this respect. Their discrimination and quantification using light microscopy is difficult because spores are the only light microscopically visible form of the parasite and they offer few distinct characters. We developed a quantitative PCR (qPCR) assay based on SYBR Green chemistry to quantify the microsporidia Glugoides intestinalis, Octosporea bayeri and Ordospora colligata in their host, the freshwater crustacean Daphnia magna. The assay allows the quantification of infection intensities in whole animals and is more than an order of magnitude more sensitive than light microscopy. Sampling and DNA extraction account for more than 90% of the residual variance in infection intensity data and this variance considerably impairs the resolution of qPCR. Where higher resolution is required, we propose using the ratio of parasite to host DNA as the measure of infection intensity. We show that this measure is robust and greatly improves resolution of qPCR. Additionally, this method can be applied to compare samples of unequal volume.

Leishmania (Viannia) braziliensis is the main cause of highly disfiguring mucocutaneous leishmaniasis (MCL) in South America. The related species L. (V.) peruviana has only been identified in simple cutaneous lesions (CL). Hybrids between L. braziliensis and L. peruviana have been reported although genetic exchange in Leishmania is considered to be rare. Here we compared growth in vitro, adaptive capacity under thermal and oxidative stress and behaviour in a hamster model, of L. braziliensis, L. peruviana, and their putative hybrids. At 24°C, the optimal temperature for in vitro growth, L. braziliensis had the highest growth rate. In in vitro studies hybrid clones presented heterogeneous phenotypes, from slower growth rates, similar to L. peruviana, to higher growth rates, as observed in L. braziliensis. Hamsters infected with hybrid strains, presented the highest parasite densities and aggressive relapses at a later stage of infection. Hybrids generally presented higher plasticity and phenotypic diversity than the putative parental species, with potential eco-epidemiological implications, including an impact on the success of disease control.

Glycosomes are specialized peroxisomes found in all kinetoplastid organisms. The organelles are unique in harbouring most enzymes of the glycolytic pathway. Matrix proteins, synthesized in the cytosol, cofactors and metabolites have to be transported across the membrane. Recent research on Trypanosoma brucei has provided insight into how these translocations across the membrane occur, although many details remain to be elucidated. Proteins are imported by a cascade of reactions performed by specialized proteins, called peroxins, in which a cytosolic receptor with bound matrix protein inserts itself in the membrane to deliver its cargo into the organelle and is subsequently retrieved from the glycosome to perform further rounds of import. Bulky solutes, such as cofactors and acyl-CoAs, seem to be translocated by specific transporter molecules, whereas smaller solutes such as glycolytic intermediates probably cross the membrane through pore-forming channels. The presence of such channels is in apparent contradiction with previous results that suggested a low permeability of the glycosomal membrane. We propose 3 possible, not mutually exclusive, solutions for this paradox. Glycosomal glycolytic enzymes have been validated as drug targets against trypanosomatid-borne diseases. We discuss the possible implications of the new data for the design of drugs to be delivered into glycosomes.

Hematodinium is a genus of parasitic dinoflagellates whose species have caused significant mortalities in marine crustacean fisheries worldwide. A species of Hematodinium infects the blue crab, Callinectes sapidus on the eastern seaboard of the USA. The mode of transmission of the parasite in blue crabs is unknown. We established several continuous in vitro cultures of Hematodinium sp. isolated from the haemolymph of infected blue crabs. One isolate has been continuously maintained in our laboratory through serial subcultivation for over 12 months, and is capable of infecting new hosts when inoculated into healthy crabs. Cells of the parasite undergo characteristic developmental changes in vitro consistent with the identifiable stages of Hematodinium sp.: filamentous trophonts, amoeboid trophonts, arachnoid trophonts and sporonts, sporoblasts, prespores and dinospores (macrospores and microspores). Additionally, we describe an unusual shunt in the life cycle wherein presumptive schizonts derived from arachnoid sporonts developed into filamentous and arachnoid trophonts that can then initiate arachnoid sporonts in new cultures. This may explain the rapid proliferation of the parasite in blue crab hosts. We also found that temperature and light intensity affected the growth and development of the parasite in vitro.

Major sperm protein (msp) genes were isolated from complementary (cDNA) and genomic DNA libraries prepared from the parasitic nematode, Oesophagostomum dentatum, characterized at the nucleotide and amino acid (aa) levels, and their expression was investigated. Three different msp cDNA and 2 genomic sequences were determinedThe nucleotide sequences reported in this article have been deposited in the EMBL, GenBank and DDJB databases under the Accession numbers AJ627869–AJ627873., each with an open reading frame (ORF) of 381 nucleotides. Nucleotide variation was detected at 30 positions in the ORF among all 5 sequences. Conceptual translation of the full-length msp sequences inferred 4 different MSPs each of 126 aa. These predicted MSPs differed at aa positions 15 (serine<->threonine), 101 (alanine<->glycine), 103 (glutamine<->leucine) and 126 (proline<->leucine). Southern blot analysis of O. dentatum genomic DNA, digested separately with various restriction endonucleases, displayed multiple (n=7–13) bands for each enzyme, providing support for a multigene family. Also, at the genomic level, sequence tracts consistent with a ‘substitute’ TATA box sequence motif were identified within a region (−1 to −123 nt) preceding the 2 msp genes. In contrast to other species of nematode investigated to date, no GATA transcription factor binding motif was detected immediately upstream of the msp coding region. Real-time PCR analysis demonstrated that msp mRNA was expressed exclusively in the males of both fourth-stage larvae (L4s) and adults of O. dentatum (raised in pigs after intragastric inoculation). The magnitude of expression in male O. dentatum raised in pigs in the presence of female worms was the same as in males in the absence of females. Comparative analyses showed aa sequence conservation among MSPs from various nematodes, suggesting similar functional roles for these proteins.

The establishment of N. dubius juveniles was more successful in mice maintained on a diet inadequate to support the full growth potential of the animals than in mice maintained on an adequate diet. The distribution of encysted juveniles along the duodenum was considerably more extensive in the former group of mice.

More juveniles were established in male than in female mice within the respective dietary groups, although the pattern of distribution was similar.

The distribution of adult nematodes at 10 days after infection was confined to a smaller area of the duodenum than that of the encysted juveniles at 5 days.

The differences in distribution of nematodes after 5 and 10 days between the HP and LP mice was not evident after 21 days. The survival of adult nematodes in the HP mice was more successful than in the LP animals.

The female nematodes in the LP mice achieved a greater length but were less prolific egg producers than those in the HP mice.

A diet inadequate to support the full growth potential of mice was therefore associated with the increased ability of Nematospiroides dubius juveniles to establish themselves in mice compared with the situation in adequately fed animals. The low plane diet, on the other hand, was associated with a decreased ability of the adult nematodes to survive.

This work was supported by grants from the Commonwealth Scholarship and Fellowship Plan and the Australian Wool Board. I should like to express my appreciation for the supervision of this work by Professor J. F. A. Sprent and to thank Miss Sharon McFeeter and Mrs Anne McKeown for their most competent technical assistance.

Twenty-one day old Fasciola hepatica were recovered from the livers of infected mice and cultured for 5–7 days in a serum-free medium containing either [C]leucine, [C]isoleucine or [S]methionine, or in a medium containing [C]leucine and serum from a sheep vaccinated with excretory– secretory (ES) antigens of juvenile F. hepatica. All three labelled amino acids were incorporated into fluke proteins. Labelled proteins also appeared in the culture medium. Three major polypeptides detected in the culture media had apparent molecular weights of 26000, 24000 and 23000. All were immunoprecipitated from [C]leucine-labelled culture medium using antisera against fluke somatic antigens raised in rabbits or from sheep vaccinated with ES antigens of juvenile F. hepatica. A polypeptide of molecular weight 27 000 was also prominent in the culture medium when [C]isoleucine was used. This polypeptide was present as a minor component when [C]leucine and [S]methionine were included in the culture media; it did not appear to be immunoprecipitated by the above antisera from [C]leucine-labelled culture medium. In the presence of serum from vaccinated sheep, the ES antigens formed immune complexes which contained the polypeptides mentioned above, together with several higher molecular weight polypeptides. Additionally, a number of minor bands of varying molecular weight were present. After micro-Ouchterlony gel immunodiffusion, 2 precipitin lines formed between the labelled ES antigens and antisera. Electrophoresis of these indicated that the 23000, 24000 and 26000 Dalton labelled polypeptides were present in each. The higher molecular weight and the 27000 Dalton labelled polypeptides were also present in one of the lines.

A comparison was made of the major excretory products when adult Haemonchus contortus worms were incubated with D-[U-14C]glucose under aerobic and anaerobic conditions. Catabolites measured were propan-1-ol, acetate, n-propionate and CO2 and the only major difference was that nearly twice as much CO2 both in terms of quantity and radioactivity was excreted under aerobic than anaerobic conditions. The worms were also much more physically active under aerobic conditions. When worms were incubated under aerobic conditions with increasing amounts of fluoroacetate their CO2 production was progressively reduced to the anaerobic level. Their movement and their ability to clump together was also progressively reduced. After aerobic incubation with fluoroacetate and D-[U-14C]g1ucose the quantity and radioactivity of citrate within worms increased greatly. When worms were similarly incubated anaerobically no increase in citrate occurred, no radioactivity was associated with the citrate and the worms appeared physically unaffected. When worms were incubated aerobically with fluoro[1-14C]acetate they produced radioactive fluorocitrate.

5-HT is a candidate for the excitatory transmitter at the neuromuscular junction in trematodes including Fasciola hepatica. This study has determined the response of a muscle strip preparation from Fasciola hepatica to 5-HT and a range of agonists that distinguish between the vertebrate receptor 5-HT subtypes. 5-HT increased the resting tone and the rhythmic activity of the muscle strip. Of the 19 compounds tested, only 10 had an effect similar to 5-HT and all but 2 of these were tryptamine compounds. 5-HT was more potent than tryptamine whilst 4-OH-tryptamine had no effect, suggesting that the response is mediated by a 5-HT rather than a tryptamine receptor. 5-Fluorotryptamine and 5-carboxyamidotryptamine were the most potent agonists. 8-OH-DPAT also mimicked the effect of 5-HT, though less potently. Assuming that these agents elicit their response through a common receptor, this suggests the presence of a 5-HT receptor with similar properties in terms of agonist recognition as the vertebrate 5-HT1 class of receptor involved in controlling Fasciola muscle motility.

A study on the immune and mating interactions between Schistosoma intercalatum and S. mansoni has been carried out. The immunologically mediated interactions and mating patterns observed between the two species demonstrate that S. mansoni is competitively dominant over S. intercalatum. When S. mansoni females are absent, S. mansoni males are more competitive than S. intercalatum males at pairing with S. intercalatum females. The use of a mathematical model shows how the abundance of S. mansoni worms per host could dramatically affect the mating probability of S. intercalatum females with their homologous males. It appears that the competitive exclusion between these two species of schistosomes may probably be an important factor restricting the distribution of S. intercalatum in Africa.

Haemogregarines (Apicomplexa: Adeleiorina) are a diverse group of haemoparasites reported from almost all vertebrate classes. The most commonly recorded haemogregarines to parasitize anurans are species of Hepatozoon Miller, 1908. To date 16 Hepatozoon species have been described from anurans in Africa, with only a single species, Hepatozoon hyperolli (Hoare, 1932), infecting a member of the Hyperoliidae. Furthermore, only two Hepatozoon species are known from South African anurans, namely Hepatozoon theileri (Laveran, 1905) and Hepatozoon ixoxo Netherlands, Cook and Smit, 2014, from Amietia delalandii (syn. Amietia quecketti) and three Sclerophrys species, respectively. Blood samples were collected from a total of 225 individuals representing nine hyperoliid species from several localities throughout northern KwaZulu-Natal, South Africa. Twenty frogs from three species were found positive for haemogregarines, namely Afrixalus fornasinii (6/14), Hyperolius argus (2/39), and Hyperolius marmoratus (12/74). Based on morphological characteristics, morphometrics and molecular findings three new haemogregarine species, Hepatozoon involucrum Netherlands, Cook and Smit n. sp., Hepatozoon tenuis Netherlands, Cook and Smit n. sp. and Hepatozoon thori Netherlands, Cook and Smit n. sp., are described from hyperoliid hosts. Furthermore, molecular analyses show anuran Hepatozoon species to be a separate monophyletic group, with species isolated from African hosts forming a monophyletic clade within this cluster.

Multiplication of Plasmodium parasites within human erythrocytes is essential to malarial disease. The cell-division cycle of this organism, however, is still poorly understood. In other eukaryotes, various techniques for (apparent) cell-cycle synchronization have been used to shed light on the mechanisms involved in cell division and its control. Thus far there is no technique for cell-cycle synchronization (as opposed to selection of parasites of a limited age-range) in Plasmodium. We therefore investigated the possibility that inhibitors of DNA synthesis, the mitotic spindle, or cell-cycle control elements (such as cyclin-dependent kinases) could be used to synchronize P. falciparum cultures to a particular cell-cycle phase. Surprisingly, most of these compounds did not cause a block at a specific phase. Three compounds, Hoechst 33342, roscovitine and L-mimosine, did block development at the trophozoite–schizont transition (S or G2 phase). The block caused by the latter 2 inhibitors was reversible, suggesting that they might be used as synchronizing agents. However, a consideration of the perturbing effects of inhibitors and problems with ‘batch’ synchronization techniques in general lead us to believe that any results obtained using roscovitine- or L-mimosine-treated parasites may not be reflective of the normal cell cycle.

Strong resistance to reinfection developed in pigs as a result of trickle inoculations with Trichuris suis (250 infective eggs twice weekly for 4 weeks). At challenge (4000 eggs) 8 weeks after the last trickle inoculation, we observed a 96% reduction in the mean number of established challenge worms in immunized (TRI/CHA) compared with non-immunized controls (CON/CHA) (74 v. 1753; P = 0·004). Moreover, the total faecal egg excretion was significantly lower in immunized pigs compared with controls (P = 0·0003). The trickle inoculations depressed pig weight gains over the 8-week course of immunization (P = 0·042). It was observed that a small number of pigs developed a weaker level of resistance, reflecting the importance of genetic determinants. The relevance of these data to the population biology of porcine and human whipworm infections is discussed.

Habitat modification may facilitate the emergence of novel pathogens, and the expansion of agricultural frontiers make domestic animals important sources of pathogen spillover to wild animals. We demonstrate for the first time that Plasmodium juxtanucleare, a widespread parasite from domestic chickens, naturally infects free-living passerines. We sampled 68 wild birds within and at the border of conservation units in central Brazil composed by Cerrado, a highly threatened biome. Seven out of 10 passerines captured in the limits of a protected area with a small farm were infected by P. juxtanucleare as was confirmed by sequencing a fragment of the parasite's cytochrome b. Blood smears from these positive passerines presented trophozoites, meronts and gametocytes compatible with P. juxtanucleare, meaning these birds are competent hosts for this parasite. After these intriguing results, we sampled 30 backyard chickens managed at the area where P. juxtanucleare-infected passerines were captured, revealing one chicken infected by the same parasite lineage. We sequenced the almost complete mitochondrial genome from all positive passerines, revealing that Brazilian and Asian parasites are closely related. P. juxtanucleare can be lethal to non-domestic hosts under captive and rehabilitation conditions, suggesting that this novel spillover may pose a real threat to wild birds.

The localization of the 28 kDa Schistosoma mansoni glutathione S-transferase (Sm28 GST) has been investigated using immunohistochemistry and electron microscopy and the results compared with previously published data. This study confirms the wide distribution of this antigen in the parasite. In male and female worms, Sm28 GST is localized in the tegument, the parenchyma, the oesophageal epithelium and in genital organs. Sm28 GST was clearly detected in germinal and sustentacular cells. The decrease of staining intensity during the differentiation of germinal cells suggests a down-regulated expression of the molecule. At the ultrastructural level, this antigen was abundant in nuclei and less present in the cytoplasm. The marked heterogeneity observed in the staining of individual worms indicates that Sm28 GST seems to be closely associated with the parasite’s metabolism. The results are discussed in relation to the biological and protective functions of the protein.

Goldfish were experimentally infected with different population densities of the acanthocephalan Pomphorhynchus laevis, and maintained at different temperatures. The density of infection had no effect upon the establishment of the parasite, but a 12°C rise in water temperature reduced the recovery after one week by 30%. Thereafter there was a continuous loss of parasites during the course of infection. The rate of this loss was independent of worm burden and temperature, but increased under conditions of host starvation. Both male and female parasites attached themselves in the same region of the alimentary tract, with a mean position about 19% along its length. They remained in that region throughout the course of infection, and the population did not migrate down the intestine with increasing age. In starved fish, the region of attachment was nearer the oesophagus, at low temperatures nearer the rectum, and when crowded the occupied region extended anteriorly. Male worms were unable to establish as easily as females, but once established survived better, and the sex ratio after two weeks was in favour of males. The results are compared with the results of studies on other acanthocephalans and on fish-cestode systems. It is recognized that although the establishment of fish acanthocephala is affected by temperature to a lesser extent than that of some fish cestodes, temperature in conjunction with changes in fish diet is a major control upon the flow of parasites through acanthocephalan-fish parasite systems. The significance of the results to the interpretation of data based on field observations is discussed.

Through the kindness of Dr S. W. Kemp, F.R.S., and of the Discovery Committee, the writer has recently had the opportunity of examining examples of a species of Uncinaria from the intestine of a sea-lion. These were collected by Mr J. E. Hamilton, of the Discovery staff, at Cape Dolphin in the Falkland Islands, in April 1931. The host was a sea-lion pup, about three months old, which was found dead on the beach, and it was suspected that the worms were concerned in the causation of its death.

Observations on the larvae of sheep trichostrongyles on hill pastures showed:

1. A reduction in larval populations occurred when the ‘moving-flock’ type management was used. The large proportion of eggs passed in faeces is deposited on a part of the pasture where little or no grazing occurs.

2. Local concentrations of larvae occurred on the grazed portions of a pasture when grazing was uneven. Thus, the apparent low rate of stocking on hill pastures does not give a true picture of the potential dangers of parasitic disease. Estimates of pasture contamination by Taylor's method of sampling were deceptively low.

3. The number of larvae on grass blades was lowest when the grass was wet with dew. Sampling of pastures between 12 noon and 5 p.m g.m.t. gave maximal and most constant estimates.

4. There was an increase in the number of larvae on a pasture during the warmer months of the year. While low temperatures prevent or delay hatching, the increase in population was not entirely due to the increased rate of development when temperature was high. The increase in numbers was partly due to an increase in the number of adult worms in the sheep hosts.

5. Removal of sheep for 3 weeks reduced the number of larvae on a pasture. This reduction was not apparent, however, until after the first 12 days. After 3 weeks the pasture population was reduced by 55 %, while after 4 weeks (i.e. 7 days after the return of the sheep) the reduction was 90%. Two weeks later there was a small increase in the larval population, but this was again reduced with the onset of colder conditions.

1. A qualitative paper chromatographic analysis of the free amino acids in Rhodnius urine and excreta was made: eleven amino acids or related compounds were detected, cystine and cysteic acid in urine, and histidine, histamine, leucine, taurine, glycine, valine, phenylalanine and alanine in the excretory fractions.

2. In addition to these, three yellow pigment spots were separated from the excreta; one of these is of unknown composition and two are peptides.

3. A possible relationship between certain of the excretory amino acids and haemolymph ones is discussed.

I am greatly indebted to the late Professor P. A. Buxton, C.M.G., F.R.S., for his stimulating criticism during this study, and to the members of the School for very useful discussion, and to Mr Paul Baily for performing the column chromatographic analyses on my behalf. The study forms part of a Thesis for the degree of Ph.D. approved by the University of London, and was financed by the South African Council for Scientific and Industrial Research. I wish to acknowledge their generous financial support.