The cellular distribution of a vertically transmitted, feminizing microsporidian was followed in its host Gammarus duebeni. In adult females the parasite was restricted to gonadal tissue, in particular primary and secondary follicle cells. Spores were diplokaryotic with a thin spore wall and a short polar filament, characteristics typical of ‘early’ spores involved in autoinfection. The diplokaryotic life-cycle, absence of spore groupings and of a pansporoblast membrane typify the genus Nosema. However, the unusual globular polaroplast of the spore and restriction of this stage to host ovarian tissue have not previously been described in Nosema. Sporogony occurred only in follicle cells adjacent to developing oocytes and was in synchrony with the process of vitellogenesis. Oocytes were infected after formation of intracellular connections with follicle cells but harboured only vegetative stages of the parasite. Parasites were associated with the perinuclear cytoplasm and, in developing embryos, segregated to daughter cells along the axis of the spindle. In juvenile animals there was no evidence of pathology linked with feminization and the parasite was found at low density in cells under the cuticle. The parasite is highly adapted to transovarial transmission with an efficient mechanism of oocyte infection and no evidence of pathology.

Aiming to better understand the ecological aspects of Trypanosoma cruzi transmission cycles, wild carnivores, small mammals and dogs were examined for T. cruzi infection in the Serra da Canastra National Park region, Brazil. Isolates were genotyped using mini-exon gene and PCR-RFLP (1f8 and H3) genomic targets. Trypanosoma cruzi transmission was well established in the area and occurred in both wild and peridomestic environments. Dog seroprevalence was 29·4% (63/214) and TcI and TcII genotypes, besides mixed infections were observed. Only TcI was detected in wild mammals. Marsupials displayed lower relative abundance, but a high prevalence of positive haemocultures (4/22), whereas rodents displayed positive haemocultures (9/113) mainly in the abundant Akodon montensis and Cerradomys subflavus species. The felid Leopardus pardalis was the only carnivore to display positive haemoculture and was captured in the same region where the small mammal prevalence of T. cruzi infection was high. Two canid species, Chrysocyon brachyurus and Cerdocyon thous, were serologically positive for T. cruzi infection (4/8 and 8/39, respectively), probably related to their capacity to exploit different ecological niches. Herein, dog infection not only signals T. cruzi transmission but also the genotypes present. Distinct transmission strategies of the T. cruzi genotypes are discussed.

We investigated whether host specificity is linked to variability within species of Lamellodiscus monogeneans, which are gill ectoparasites of the Sparidae. We sampled fish parasites in the northeastern part of the Mediterranean Sea: 4 specialist species, using 1 single host species, and 3 generalist species, using 2 distinct host species. Intraspecific variability was assessed from 2 different datasets. Morphometric variability of the attachment organ, called the haptor, was estimated first from measurements of several sclerified haptoral parts on 102 individuals. Genetic variability was calculated based on comparisons of sequences derived from the first internal transcribed spacer (ITS-1) of nuclear ribosomal DNA of 62 individuals. Morphometric variances in the specialist versus generalist species were compared via principal component analysis and F-tests, and uncorrected genetic distances (p-distances) were estimated within each species. We showed that the inter-individual variance of morphometric characters, as well as p-distances, are clearly greater within generalist species than specialist ones. These findings suggest that a relative increase in morphological and molecular variability enhances the possibility to colonize new host species in monogeneans, and supports the hypothesis that intraspecific variability could be a potential determinant of host specificity.

Leishmania donovani causes visceral disease (kala-azar), a major health problem throughout the tropics with 500 000 new cases every year. Leishmania differentiates from the promastigote to the amastigote form to establish infection in a mammalian host. To understand the process of differentiation, we assessed the global variation in gene expression in promastigotes, an intermediate stage of differentiation (PA24) and axenic amastigotes in culture using an L. donovani genomic microarray with 4224 clones printed in triplicate. During an intermediate stage of differentiation 24 h after shifting the promastigotes into amastigotes (PA24), there were 41 (∼1%) clones with expression ⩾2·0-fold higher than promastigotes, whereas in terminally differentiated amastigotes there were 130 (∼3%) such clones. Of particular interest were certain genes that exhibited a transient increase or decrease in expression at the PA24 stage. Kinases showed a transient increase, and surface molecules, PSA and amino acid permease, were prominent clones among those showing a brief decrease at the PA24 stage. The microarray results have been validated using Northern blots or RT-PCR. In summary, our results provide important clues about the genes involved in the differentiation process of L. donovani that may contribute to virulence.

Neuronal abnormalities have been described in the intestine of helminth-infected rats. However, the physiological ramifications of these changes have not been determined. Here, we examined epithelial ion secretion, indicated by increases in short-circuit current (Isc), evoked by electrical transmural stimulation (TS) of enteric nerves in Ussing-chambered jejunal tissues from Nippostrongylus brasiliensis-infected rats. Rats were examined at 10 and 35 days post-infection (p.i.); non-infected rats served as controls. TS resulted in significantly reduced ion secretion in jejunum from 10 day p.i. rats compared to controls or jejunum from 35 day p.i. rats. The TS response in tissue from infected rats had, unlike controls, no cholinergic component. Tissues from both non-infected and infected rats were equally responsive to the muscarinic agonist bethanechol, suggesting that the cholinergic defect was neuronal and not an inability of the epithelium to respond to cholinergic stimulation. However, increases in Isc evoked by exogenous substance P (SP) in tissue from rats 10 day p.i. were reduced in magnitude to approximately 25% of control values. Concomitant with these physiological changes, tissue from infected rats contained increased amounts of substance P immunoreactivity and intestinal sections displayed increased numbers of substance P-immunoreactive nerve fibre profiles at both 10 and 35 days p.i. Thus, following N. brasiliensis infection there is a shift in the enteric nervous system away from cholinergic to non-cholinergic regulation, associated with increased amounts of the pro-inflammatory neuropeptide, substance P. We speculate that changes in neuronal structure and function are intimately involved in the co-ordinated multicellular response to intestinal parasitic infection and subsequent gut recovery.

The penetration barrier presented to the miracidium by the snail epithelium can be divided into three layers. The chemical composition and physical configuration of the outermost of these plays an important part in the initial attachment response of the miracidium. Attachment can be stimulated in the absence of the snail by pure chemicals in solution. However, the surface to which the miracidium attaches must have the correct physical configuration otherwise the miracidium is unable to form a stable attachment.

In vivo, the miracidial body begins to contract and relax following attachment to the snail. This coincides with the start of secretion by the apical gland and accessory gland cells. The snail's columnar epithelium is rapidly cytolysed so that 10 min after attachment the anterior of the miracidium has reached the underlying connective tissues.

As the miracidium penetrates the snail, its ciliated epithelial cells are shed in sequence from anterior to posterior. This shedding removes a protective barrier against osmosis which is probably the acid mucopolysaccharide present in the epithelial cells. The mechanism of shedding is not understood but involves the reversal of binding by the desmosomal mucosubstance which attaches the epithelial cells to surrounding intercellular ridges.

The miracidium metamorphoses into the sporocyst as it penetrates the snail, by forming a new body surface. The material for this is extruded from the vesiculated cells which lie beneath the musculature of the body wall. The process of surface formation coincides with cell shedding and moves backwards as cells are shed. At not more than 2·5 h after attachment the extruded cytoplasm forms a thin continuous layer over the surface of the organism. Contacts with underlying cells appear to have been broken and the cytoplasm is underlain by a thin fibrous basal lamella. In the first 24 h after penetration the surface of this syncytium becomes thrown into folds and metamorphosis into the sporocyst can be considered complete.

1. All stages of Argas persicus are negatively phototropic. In addition, they are also susceptible to pronounced differences in the intensity of the illumination and select the darker places.

2. In the gorged state, the ticks are slightly positively geotropic. In the unfed state, this property is not developed.

3. At all stages the tick endeavours to bring as much of its surface as possible in contact with its surroundings.

4. A. persicus is attracted by heat.

5. In all stages an olfactory sense is well developed.

6. “Haller's organ” is olfactory in function and constitutes a means by which a tick is able to recognise its host. By depriving ticks of this organ it is possible by suitable means to cause them to feed on media other than blood, thus showing that a sense of taste is absent. Argas persicus, Ornithodorus moubata and Hyalomma aegyptium have all given similar results with regard to the latter point and we believe that this constitutes a method by which perhaps other blood-sucking arthropods, after being deprived of the organ or organs necessary for the recognition of their hosts, may be made to feed on any desired medium.

A comparative study has been made of the progress in the control of ovine echinococcosis caused by Echinococcus granulosus and the ovine cysticercoses caused by Taenia hydatigena and T. ovis in New Zealand. The methods of control included an educational and a dog-dosing programme. In the Styx field trial, the Otago/Southland Surveillance Programme and the national control programme, E. granulosus declined towards extinction. In contrast, this same control effort transformed the cysticercoses from hyper-endemic to endemic status with focal epidemics. In this unstable state, the density-dependent constraint was negligible and superinfection occurred.

Sporozoites of Eimeria tenella were disrupted by sonication and subcellular fractions were separated by sucrose gradient ultracentrifugation. Fractions from gradients were characterized by electron microscopical appearance and their polypeptide and antigenic profiles determined by PAGE and Western blotting with antisera to sporozoites and 1st- and 2nd-generation merozoites. Fractions containing micronemes, rhoptries or membranes showed markedly different polypeptide content and antigenic reactivity. Microneme epitopes were strongly conserved between sporozoites and 2nd-generation merozoites whereas the majority of rhoptry epitopes and many membrane epitopes were sporozoite specific. The only polypeptide of sporozoites which was strongly recognized by antisera raised to 1st generation merozoites was a microneme antigen of molecular weight approximately 100 kDa.

1. It was found that larvae of the genera Trichostrongylus, Haemonchus, Ostertagia and Chabertia migrated upwards on the blades of sedge (Kylinga sp.) and of rye grass (Lolium perenne) in greatest numbers when subjected to a light of an intensity about 62 f.c.

2. Moisture on the grass was found to favour laval migration. More than 0·12 ml. of water per sq. cm. of soil surface, however, retarded movement. More than 0·2 ml. per sq. cm. tended to prevent larval migration up the grass.

3. Of the genera examined Ostertagia spp. were most favoured by the presence of moisture on the grass; Haemonchus contortus was most hindered by its presence.

4. Increasing quantities of moisture in the soil up to 85% saturation assisted larvae to move on to the grass. Above this value larval migration was retarded.

5. Ostertagia spp. were best able to migrate from wet soil while Haemonchus contortus was least able to migrate from wet soil.

6. It was found that the presence of water on the soil surface tended to lessen the ability of larvae to move on to the grass.

7. Haemonchus contortus was found to migrate on relatively dry grass at a faster rate than Trichostrongylus spp. and Ostertagia spp.

8. In the temperature range examined larvae were found to move up the grass in largest numbers at 5 and 45° C.

9. Haemonchus contortus was the most active species at higher temperatures.

10. The cycle of larval movement during the day, under conditions approximating to those of the field, was investigated. The greatest number of larvae were recovered from the grass in early morning. Reactions in the evening were much less marked.

Murine monoclonal antibodies (Mabs) were produced by vaccination of Balb/c mice with live Plasmodium falciparum infected red cells (iRBC). The iRBC Mabs recognized altered forms of the human erythrocyte membrane protein band 3; however, these Mabs did not recognize the band 3 molecule in uninfected or ring-infected red cells. The location of epitopes was determined by studying the binding of the iRBC Mabs after selective proteolysis of band 3 as well as by the reactivity of these Mabs to synthetic peptides that corresponded to putative exofacial regions of band 3. Treatment of uninfected red cell membranes with trypsin under low ionic strength conditions resulted in exposure of cryptic epitopes of band 3 which were recognized by the iRBC Mabs. Several of the anti-iRBC Mabs (two of which were described previously) inhibited the in vitro adherence of infected erythrocytes to C32 amelanotic melanoma cells. A mouse polyclonal serum against a synthetic peptide based on an amino acid sequence motif of band 3 reacted (by immunostaining) only with the surface of iRBC and blocked adhesion. Thus, it appears that cryptic residues of the band 3 protein become exposed upon parasitization, and their presence contributes to the increased adhesiveness of the P. falciparum-infected red cell.

The numerical relationships between a population of Lepeophtheirus pectoralis and its host population (plaice) have been described. There was no significant difference between infection levels in male and female plaice. In most of the monthly samples (75%) there was no significant relationship between host length and either the incidence or intensity of infection. The samples in which significant positive correlations occurred were usually taken within the period November to April. It is probable that the immigration of smaller fish, with lower levels of infection, from shallower water produces this effect.

The parasite population was found to have an overdispersed distribution (variance > mean) within its habitat, the host population. The frequency distributions of numbers of parasites per fish were calculated and the negative binomial distribution was fitted to these. The fit was good in 22 of the 24 samples and the value of k, the exponent, was found to be directly related to the mean density of the parasite population. Iwao's (1970) method of dispersion analysis indicated that the observed negative binomial was generated by a model of randomly distributed clumps or colonies of individuals. A possible mechanism of producing this pattern was proposed based on evidence from studies of mortality in the parasite population (which indicated that all-or-none type mortality operates on the clumps at certain periods during the year), the infection process (which demonstrated that a single wave of infection produces an overdispersed spatial distribution) and the examination of parasites from individual fish (which displayed a clumping of parasites at certain stages rather than an even spread over the range of possible stages). The mechanism proposed is that the adaptations for host location of the infective copepodid stage tend to produce aggregations of these larvae in favourable areas of a heterogeneous habitat. Fish moving randomly into these areas will tend to be infected by more than one larva (a clump of larvae) at any one time when available for infection. The fish can be regarded as equivalent to random samples of the free larval population and the aggregated distribution of the parasite population on its hosts is therefore directly related to the aggregated distribution of the free-swimming infective larvae.

The protective efficacy of a Schistosoma japonicum, Chinese strain, triose-phosphate isomerase (TPI) plasmid DNA vaccine was examined in naïve pigs. Pigs were vaccinated with the TPI DNA-plasmid alone, or in conjunction with IL-12 as pcDNA3.1-P35, pcDNA3.1-P40 plasmids via intramuscular injection. Control pigs were immunized with equivalent amounts of pcDNA3.1. Pigs were immunized 3 times at 21-day intervals and challenged 30 days after the final boost. Forty-five days post-challenge, pigs were sacrificed and perfused to compare adult worm burdens, female worm burdens, liver egg burdens and granuloma size. We found that pigs vaccinated with SjCTPI DNA alone had adult worm burdens reduced by 48·3% and that a further decrease in adult worm burdens was not seen in the group vaccinated with SjCTPI DNA in conjunction with IL-12 (46·2% reduction). The SjCTPI DNA vaccines had a more pronounced effect on reducing female worm burdens i.e. 53·6% SjCTPI alone and 59·6% for SjCTPI+IL-12. Vaccination with SjCTPI-DNA reduced liver eggs by 49·4% and this response was significantly enhanced by the addition of IL-12 (65·8% reduction in liver eggs). In addition to the dramatic protective effects seen in vaccinated pigs, we also noted that granuloma size was reduced by 42% in both groups. Thus, vaccination of pigs and other large animals in China with SjCTPI DNA vaccine will likely reduce transmission by reducing adult worm burdens and worm egg output and simultaneously reduce hepatic egg-associated pathology.

1. The seasonal incidence of Nematodirus infection in sheep was followed by faecal egg counts, post-mortem examination and pasture larval studies.

2. It was shown that only one generation of adult worms was produced each year and that the life history required a prolonged resting stage on the pasture, which took 12 months to complete.

3. The influence of this type of life cycle on the parasitic disease caused by Nematodirus, and on its control, are discussed.

Lungworms of the genus Dictyocaulus from cattle, roe deer, and moose in Sweden were subjected to morphological and molecular analyses. The objectives of the study were to investigate whether mixed or monospecific Dictyocaulus infections occur in Swedish cattle and whether wild cervids may act as reservoirs. The morphological characters examined were thickness and shape of the buccal capsule wall (BCW) and total spicular length (TSL). Morphometry was also done on the total body length, and BCW thickness and length. In the molecular identification, we used a PCR-linked hybridization assay to probe worm DNA with species-specific oligonucleotide probes to the second internal transcribed spacer (ITS2). The results showed that the BCW shape was the most reliable morphological character for identification. Significant differences were observed in this character, but an overlap occurred between lungworms from each of the host species. With the hybridization assay, all lungworms from cattle were identified as D. viviparus, whereas those from roe deer represented a novel Dictyocaulus species demonstrating that each host had a monospecific lungworm infection. In moose, 61 (78·2%) worms belonged to the new species and 17 (21·8%) were D. eckerti. This study shows the usefulness of hybridization assay as an epidemiological tool for the specific identification of lungworms of cattle and wild cervids.

Parasitism by the flea Synosternus cleopatrae does not affect the body mass of its principal rodent host, Gerbillus andersoni under natural infestation levels. We hypothesized that the lack of negative effects of flea parasitism on rodent body mass could be related either to the low level of natural infestation or to the differential susceptibility of rodent age cohorts to flea parasitism. We tested these hypotheses by measuring body mass change under flea parasitism in (a) adult rodents infested with fleas above the natural infestation level (the first hypothesis) and (b) juvenile rodents infested with fleas at natural infestation levels (the second hypothesis). Adult individuals parasitized by a number of fleas higher than in nature lost body mass at higher rates than non-parasitized control individuals. Parasitism significantly affected daily body mass change of juvenile gerbils. Juvenile rodents parasitized by fleas at the natural level of infestation lost body mass faster and gained body mass slower than control animals. We suggest that some regulating mechanisms may limit natural flea densities at a point at which the negative effect on hosts is below the accuracy of our measurements. However, natural flea densities are sufficiently high to harm the more susceptible, juvenile cohort.

Guidelines recommend praziquantel (PZQ) for the treatment and control of schistosomiasis, with no real alternative. Metrifonate was still widely used against Schistosoma haematobium in the 1990s, and then withdrawn. Experimental studies and clinical trials suggest that artemisinin compounds are active against S. haematobium. In a Cochrane systematic review assessing the efficacy and safety of drugs for treating urinary schistosomiasis, 24 randomized controlled trials (n=6315 individuals) met our inclusion criteria. These trials compared a variety of single agent and combination regimens with PZQ, metrifonate or artemisinin derivatives. The review confirmed that both the standard recommended doses of PZQ (single 40 mg/kg oral dose) and metrifonate (3×7·5–10 mg/kg oral doses administered fortnightly) are efficacious and safe in treating urinary schistosomiasis, but there is no study comparing these two regimens head-to-head. There is currently not enough evidence to evaluate artemisinin compounds. Most of the studies included in the Cochrane systematic review were insufficiently powered, lacked standardization in assessing and reporting outcomes, and had a number of methodological limitations. In this paper we discuss the implications of these findings with respect to public health and research methodology and propose priority research needs.

Mathematical models of disease dynamics tend to assume that individuals within a population mix at random and so transmission is random, and yet, in reality social structure creates heterogeneous contact patterns. We investigated the effect of heterogeneity in host contact patterns on potential macroparasite transmission by first quantifying the level of assortativity in a socially structured wild rodent population (Apodemus flavicollis) with respect to the directly-transmitted macroparasitic helminth, Heligmosomoides polygyrus. We found the population to be disassortatively mixed (i.e. male mice mixing with female mice more often than same sex mixing) at a constant level over time. The macroparasite H. polygyrus has previously been shown to exhibit male-biased transmission so we used a Susceptible-Infected (SI) mathematical model to simulate the effect of increasing strengths of male-biased transmission on the prevalence of the macroparasite using empirically-derived transmission networks. When transmission was equal between the sexes the model predicted macroparasite prevalence to be 73% and infection was male biased (82% of infection in the male mice). With a male-bias in transmission ten times that of the females, the expected macroparasite prevalence was 50% and was equally prevalent in both sexes, results that both most closely resembled empirical dynamics. As such, disassortative mixing alone did not produce macroparasite dynamics analogous to those from empirical observations; a strong male-bias in transmission was also required. We discuss the relevance of our results in the context of network models for transmission dynamics and control.

Migratory birds contribute to the movement of avian parasites between distant locations, thereby influencing parasite distribution and ecology. Here we analyse the prevalence, diversity and interaction patterns of Haemosporida parasites infecting Blackcap (Sylvia atricapilla) populations in a recently established migratory divide of southwestern Germany across 4 years. We hypothesize that the temporal and spatial isolation provided by 2 sympatric Blackcap breeding populations (migratory divide) might modify ecological interactions and thus create differences in the structure of the parasite community according to migratory route. We used a fragment of the mitochondrial DNA cytochrome b gene to determine haemosporidian haplotypes. We detected an overall infection prevalence of 70·3% (348 out of 495 blackcaps sampled from 2006 to 2009), and prevalence rates were significantly different among years and seasons. We observed a total of 27 parasite haplotypes infecting blackcaps, from them 6 new rare Haemoproteus haplotypes were found in 2 mixed infections. H. parabelopolskyi haplotypes SYAT01 (35·7%) and SYAT02 (20·8%) comprised most of the infections. An association analysis suggests that SYAT01 and SYAT02 are interacting negatively, implying that they are either competing directly for host resources, or indirectly by eliciting a cross-immune response. Molecular data show no clear difference between the parasite communities infecting blackcaps with different migratory routes, despite some temporal and spatial isolation between the two sympatric blackcap populations.

An in vitro system is described in which over 85% of a population of Leishmania mexicana mexicana amastigotes transforms to promastigotes within 48 h. The differentiation process involves 3 morphologically and biochemically distinct intermediates, including a division stage. Cell division is shown to be necessary for complete development to promastigotes. Foetal calf serum (FCS) has been found to be an essential component of the medium for high percentage transformation to be achieved. One of the important components of the FCS has been identified as non-esterified fatty acids, and these support a relatively high percentage of amastigotes through transformation in the absence of FCS, possibly due to their use as energy substrates. Only small numbers of amastigotes transform to promastigotes if glucose or amino acids are the only energy substrates available. Transformation is inhibited by a number of metabolic inhibitors including anti-leishmanial and other anti-protozoal drugs. The stage at which inhibition is apparent varies with the inhibitor. It is suggested that the system described for the transformation in vitro of L. m. mexicana amastigotes to promastigotes may be the best method available at present for studying the metabolism and drug sensitivity of amastigotes free from possible interference by host macrophage components.