The Dominance of Coinfecting Parasites' Indirect Genetic Effects on Host Traits.

AbstractIndirect genetic effects (IGEs) exist when there is heritable variation in one organism's ability to alter a second organism's traits. For example, parasites have antigens that can induce a host immune response, as well as disparate strategies to evade or suppress host immunity; among-parasite genetic variation in these antigens generates among-host variation in immune traits. Here, we experimentally show that the cestode parasite Schistocephalus solidus exerts an IGE on an immune trait (peritoneal fibrosis) in its threespine stickleback host: stickleback developed strong fibrosis after exposure to some parasite genotypes but not others. A complication arises during coinfection, when two or more parasite genotypes may impose conflicting IGEs on the same host trait. What parasite-controlled trait will the host express? Will the host trait reflect the more immune-stimulatory parasite genotype or the more immune-evasive genotype? These alternatives can be quantified by estimating the dominance coefficient, as if a coinfected host were a heterozygote. We experimentally estimated the dominance of S. solidus IGEs by coinjecting antigens from different parasite genotypes. Contrary to our a priori hypotheses, coinjected antigens induced an overdominant effect, stronger than either parasite's antigens alone. We present a mathematical model showing that the value of this IGE dominance is biologically important, affecting the evolutionary dynamics of parasites in a density- and frequency-dependent manner. The model indicates that overdominance would be detrimental to immigrants when resident prevalence is high. This combination of experimental data and modeling provides an example of a parasite IGE on host traits and the evolutionary significance of IGE dominance.

Allergenic response induced by Pterobothrium crassicolle (Cestoda: Trypanorhyncha) extracts in murine model.

The aim of this study was to determine the allergenic activity of components present in crude extracts of Pterobothrium crassicolle plerocerci (CPE) and blastocysts (CBE) obtained from Micropogonias furnieri in a murine model. Two groups of seven animals each received 50 µg of CPE or CBE on days 1, 35 and 120. Serum samples were tested by ELISA and Immunoblotting. Specific IgG and IgE levels were detected by ELISA, showing specific humoral responses for the primary immunization for both immunoglobulins and continuously growing titers for IgE. Positive Passive Cutaneous Anaphylaxis tests in rats sensitized with anti-CBE sera and tested by CBE, showed biologically, the allergenic activity of the extracts. The CPE and CBE showed some different recognition regions but both experimental groups recognized all regions of the extracts when tested for cross reactions, showing that CPE and CBE could share antigenic recognition sites.

Population-level immunologic variation in wild threespine stickleback (Gasterosteusaculeatus).

Wild organisms are regularly exposed to a wide range of parasites, requiring the management of an effective immune response while avoiding immunopathology. Currently, our knowledge of immunoparasitology primarily derives from controlled laboratory studies, neglecting the genetic and environmental diversity that contribute to immune phenotypes observed in wild populations. To gain insight into the immunologic variability in natural settings, we examined differences in immune gene expression of two Alaskan stickleback (Gasterosteus aculeatus) populations with varying susceptibility to infection by the cestode Schistocephalus solidus. Between these two populations, we found distinct immune gene expression patterns at the population level in response to infection with fish from the high-infection population displaying signs of parasite-driven immune manipulation. Further, we found significant differences in baseline immune gene profiles between the populations, with uninfected low-infection population fish showing signatures of inflammation compared to uninfected high-infection population fish. These results shed light on divergent responses of wild populations to the same parasite, providing valuable insights into host-parasite interactions in natural ecosystems.

Investigations on the occurrence of tapeworm infections in German horse populations with comparison of different antibody detection methods based on saliva and serum samples.

BACKGROUND: Effective and sustainable worm control in horses would benefit from detailed information about the current regional occurrence of tapeworms. Different diagnostic methods are currently available to detect Anoplocephala spp. infections in horses. However, the format as well as the sensitivity and specificity of the methods vary considerably. METHODS: A coprological, serological and questionnaire study was conducted to investigate the prevalence and risk factors of tapeworm infections on 48 horse farms in the region of Berlin and Brandenburg, Germany. In total, faecal samples of 484 horses were analysed using the double centrifugation/combined sedimentation-flotation and mini-FLOTAC. Serum (n = 481) and saliva (n = 365) samples were analysed by ELISAs to determine antibody levels against Anoplocephala spp. 12/13 kDa excretory/secretory (E/S) antigens. RESULTS: Cestode eggs were detected in 0.6% of faecal samples (farm prevalence 6.3%) without differences between the two methods. In contrast, antibodies against Anoplocephala spp. were detected in 16.2% (farm prevalence 52.1%) and in 29.5% (farm prevalence 75.7%) of the serum and saliva samples, respectively. Both ELISA based methods for detection of tapeworms reported a greater number of infected animals requiring treatment than were positively identified by coproscopy. Logistic regression analysis identified permanent pasture access, large pastures and regular pasture changes and high strongyle egg counts as risk factors for positive serum antibody responses to Anoplocephala spp. while last treatment with praziquantel was protective. Other protective factors were the presence of foals and high numbers of horses on the farm. Daily removal of faeces from the pasture and horse age did not have a significant effect. CONCLUSIONS: The findings of the present serological investigation indicate that tapeworm prevalence in Berlin/Brandenburg horse farms is much higher than would be anticipated by using conventional/coproscopic analyses. Moreover, the majority of tapeworm-positive horses had not received a cestocidal drug at their last treatment. Considering the already known low sensitivity of the coproscopic detection, the equine veterinary diagnostics can be enhanced by the use of antibody detection methods such as the saliva-based ELISA.

The gut microbiota response to helminth infection depends on host sex and genotype.

Vertebrates' gut microbial communities can be altered by the hosts' parasites. Helminths inhabiting the gut lumen can interact directly with their host's microbiota via physical contact, chemical products, or competition for nutrients. Indirect interactions can also occur, for instance when helminths induce or suppress host immunity in ways that have collateral effects on the microbiota. If there is genetic variation in host immune responses to parasites, we would expect such indirect effects to be conditional on host genotype. To test for such genotype by infection interactions, we experimentally exposed Gasterosteus aculeatus to their naturally co-evolved parasite, Schistocephalus solidus. The host microbiota differed in response to parasite exposure, and between infected and uninfected fish. The magnitude and direction of microbial responses to infection differed between host sexes, and also differed between variants at autosomal quantitative trait loci. These results indicate that host genotype and sex regulate the effect of helminth infection on a vertebrate gut microbiota. If this result holds in other taxa, especially humans, then helminth-based therapeutics for dysbiosis might need to be tailored to host genotype and sex.

Analysis of caecal mucosal inflammation and immune modulation during Anoplocephala perfoliata infection of horses.

Anoplocephala perfoliata is the commonest equine tapeworm, the adult parasites are attached in groups close to the ileocaecal valve causing marked inflammatory pathology. This work aimed to characterize the nature of the in vivo mucosal immune response to A perfoliata, and to investigate the role of A perfoliata excretory-secretory components in modulating in vitro immune responses. Real-time PCR detected elevation of IL13 and TGFß transcription in early-stage A perfoliata infection. In late-stage infection, IL-13, IL4 and Ifn transcripts were reduced while the regulatory cytokines, TGFß, IL10 and the transcription factor FOXP3 were increased in tissue close to the site of A perfoliata attachment; indicating downregulation of T-cell responses to A perfoliata. In vitro, A perfoliata excretory-secretory products induced apoptosis of the Jurkat T-cell line and premature cell death of ConA stimulated equine peripheral blood leucocytes. Analysis of cytokine transcription patterns in the leucocyte cultures showed a marked inhibition of IL-1 and IL-2 suggesting that a lack of T-cell growth factor transcription underlies the mechanism of the induced equine T-cell death. These preliminary findings suggest A perfoliata may have the ability to down-regulate host T-cell responses.

Disseminated Metacestode Versteria Species Infection in Woman, Pennsylvania, USA1.

A patient in Pennsylvania, USA, with common variable immunodeficiency sought care for fever, cough, and abdominal pain. Imaging revealed lesions involving multiple organs. Liver resection demonstrated necrotizing granulomas, recognizable tegument, and calcareous corpuscles indicative of an invasive cestode infection. Sequencing revealed 98% identity to a Versteria species of cestode found in mink.

Macrophages treated with antigen from the tapeworm Hymenolepis diminuta condition CD25+ T cells to suppress colitis.

Macrophages play central roles in immunity as early effectors and modulating adaptive immune reponses; we implicated macrophages in the anticolitic effect of infection with the tapeworm Hymenolepis diminuta. Here, gene arrays revealed that H. diminuta antigen (HdAg) evoked a program in murine macrophages distinct from that elicited by IL-4. Further, HdAg suppressed LPS-evoked release of TNF-α and IL-1ß from macrophages via autocrine IL-10 signaling. In assessing the ability of macrophages treated in vitro with an extract of H. diminuta [M(HdAg)] to affect disease, intravenous, but not peritoneal, injection of M(HdAg) protected wild-type but not RAG1-/- mice from dinitrobenzene sulphonic acid (DNBS)-induced colitis. Administration of splenic CD4+ T cells from in vitro cocultures with M(HdAg), but not those cocultured with M(IL-4) cells, inhibited DNBS-induced colitis; fractionation of the T-cell population indicated that the CD4+CD25+ T cells from cocultures with M(HdAg) drove the suppression of DNBS-induced colitis. Use of IL-4-/- or IL-10-/- CD4+ T cells revealed that neither cytokine alone from the donor cells was essential for the anticolitic effect. These data illustrate that HdAg evokes a unique regulatory program in macrophages, identifies HdAg-evoked IL-10 suppression of macrophage activation, and reveals the ability of HdAg-treated macrophages to educate ( i.e., condition) and mobilize CD4+CD25+ T cells, which could be deployed to treat colonic inflammation.-Reyes, J. L., Lopes, F., Leung, G., Jayme, T. S., Matisz, C. E., Shute, A., Burkhard, R., Carneiro, M., Workentine, M. L., Wang, A., Petri, B., Beck, P. L., Geuking, M. B., McKay, D. M., Macrophages treated with antigen from the tapeworm Hymenolepis diminuta condition CD25+ T cells to suppress colitis.

Environmental temperature variation influences fitness trade-offs and tolerance in a fish-tapeworm association.

BACKGROUND: Increasing temperatures are predicted to strongly impact host-parasite interactions, but empirical tests are rare. Host species that are naturally exposed to a broad temperature spectrum offer the possibility to investigate the effects of elevated temperatures on hosts and parasites. Using three-spined sticklebacks, Gasterosteus aculeatus L., and tapeworms, Schistocephalus solidus (Müller, 1776), originating from a cold and a warm water site of a volcanic lake, we subjected sympatric and allopatric host-parasite combinations to cold and warm conditions in a fully crossed design. We predicted that warm temperatures would promote the development of the parasites, while the hosts might benefit from cooler temperatures. We further expected adaptations to the local temperature and mutual adaptations of local host-parasite pairs. RESULTS: Overall, S. solidus parasites grew faster at warm temperatures and stickleback hosts at cold temperatures. On a finer scale, we observed that parasites were able to exploit their hosts more efficiently at the parasite's temperature of origin. In contrast, host tolerance towards parasite infection was higher when sticklebacks were infected with parasites at the parasite's 'foreign' temperature. Cold-origin sticklebacks tended to grow faster and parasite infection induced a stronger immune response. CONCLUSIONS: Our results suggest that increasing environmental temperatures promote the parasite rather than the host and that host tolerance is dependent on the interaction between parasite infection and temperature. Sticklebacks might use tolerance mechanisms towards parasite infection in combination with their high plasticity towards temperature changes to cope with increasing parasite infection pressures and rising temperatures.

Recent evolution of extreme cestode growth suppression by a vertebrate host.

Parasites can be a major cause of natural selection on hosts, which consequently evolve a variety of strategies to avoid, eliminate, or tolerate infection. When ecologically similar host populations present disparate infection loads, this natural variation can reveal immunological strategies underlying adaptation to infection and population divergence. For instance, the tapeworm Schistocephalus solidus persistently infects 0-80% of threespine stickleback (Gasterosteus aculeatus) in lakes on Vancouver Island. To test whether these heterogeneous infection rates result from evolved differences in immunity, we experimentally exposed laboratory-reared fish from ecologically similar high-infection and no-infection populations to controlled doses of Schistocephalus We observed heritable between-population differences in several immune traits: Fish from the naturally uninfected population initiated a stronger granulocyte response to Schistocephalus infection, and their granulocytes constitutively generate threefold more reactive oxygen species in cell culture. Despite these immunological differences, Schistocephalus was equally successful at establishing initial infections in both host populations. However, the no-infection fish dramatically suppressed tapeworm growth relative to high-infection fish, and parasite size was intermediate in F1 hybrid hosts. Our results show that stickleback recently evolved heritable variation in their capacity to suppress helminth growth by two orders of magnitude. Data from many natural populations indicate that growth suppression is widespread but not universal and, when present, is associated with reduced infection prevalence. Host suppression of helminth somatic growth may be an important immune strategy that aids in parasite clearance or in mitigating the fitness costs of persistent infection.

Molecular diagnosis of allergy to Anisakis simplex and Gymnorhynchus gigas fish parasites.

BACKGROUND: There has been an increase in the prevalence of hypersensitivity to Anisakis simplex. There are fish parasites other than Anisakis simplex whose allergenicity has not yet been studied. OBJECTIVE: To assess IgE hypersensitivity caused by fish parasite allergens in patients with gastro-allergic symptoms after consumption of fish, shellfish or cephalopods, compared with healthy subjects, pollen allergic individuals and children with digestive symptoms after eating marine food. METHODS: We carried out in vivo tests (skin prick) and in vitro tests (specific IgE determination, Western blot) and component resolved diagnostics (CRD) using microarray analysis in all patients. RESULTS: CRD better detected sensitisation to allergens from marine parasites than skin prick tests and determination of specific IgE by CAP. Sensitisation to Gymnorhynchus gigas was detected in 26% of patients measured by skin prick tests and 36% measured by IgE. CONCLUSIONS: The prevalence of hypersensitivity to marine parasite allergens other than Anisakis simplex should be studied, and the most appropriate technique for this is CRD.

An experimental approach to the immuno-modulatory basis of host-parasite local adaptation in tapeworm-infected sticklebacks.

The evolutionary arms race of hosts and parasites often results in adaptations, which may differ between populations. Investigation of such local adaptation becomes increasingly important to understand dynamics of host-parasite interactions and co-evolution. To this end we performed an infection experiment involving pairs of three-spined sticklebacks and their tapeworm parasite Schistocephalus solidus from three geographically separated origins (Germany, Spain and Iceland) in a fully-crossed design for sympatric and allopatric host/parasite combinations. We hypothesized that local adaptation of the hosts results in differences in parasite resistance with variation in parasite infection rates and leukocyte activation, whereas parasites from different origins might differ in virulence reflected in host exploitation rates (parasite indices) and S. solidus excretory-secretory products (SsESP) involved in immune manipulation. In our experimental infections, sticklebacks from Iceland were more resistant to S. solidus infection compared to Spanish and German sticklebacks. Higher resistance of Icelandic sticklebacks seemed to depend on adaptive immunity, whereas sticklebacks of German origin, which were more heavily afflicted by S. solidus, showed elevated activity of innate immune traits. German S. solidus were less successful in infecting and exploiting allopatric hosts compared to their Icelandic and Spanish conspecifics. Nevertheless, exclusively SsESP from German S. solidus triggered significant in vitro responses of leukocytes from naïve sticklebacks. Interestingly, parasite indices were almost identical across the sympatric combinations. Differences in host resistance and parasite virulence between the origins were most evident in allopatric combinations and were consistent within origin; i.e. Icelandic sticklebacks were more resistant and their S. solidus were more virulent in all allopatric combinations, whereas German sticklebacks were less resistant and their parasites less virulent. Despite such differences between origins, the degree of host exploitation was almost identical in the sympatric host-parasite combinations, suggesting that the local evolutionary arms race of hosts and parasites resulted in an optimal virulence, maximising parasite fitness while avoiding host overexploitation.

Virulence in the three-spined stickleback specific parasite Schistocephalus solidus is inherited additively.

Parasite virulence is a key trait in host-parasite interactions and plays a crucial role in infection dynamics. Our study system offers the rare opportunity to study the virulence of an individual macroparasite (Schistocephalus solidus) in its vertebrate fish host (Gasterosteus aculeatus). The size of the tapeworm in the fish can be regarded as a good proxy for individual parasite virulence, as parasite size correlates negatively with fitness traits of the stickleback host (i.e. the bigger the parasite, the lower the host's reproductive success) as well as directly with the number of parasite offspring to be expected. To investigate how virulence is inherited, laboratory bred, parasite-naïve stickleback were infected with a cross of two S. solidus populations of either high or low virulence, as well as one hybrid cross between the two. The relative weight of the parasite as expressed in the parasite index served as a measure of virulence. Furthermore, we measured several condition and immune related traits in the fish host to assess parasite impact on the stickleback. We hypothesized that parasite virulence is to a large extent genetically determined and correlated with several fitness traits in the stickleback host. We found that virulence is inherited additively in S. solidus, with hybrids of high and low virulence parasites displaying intermediate levels. However, contrary to expectation, infection rate of S. solidus in three-spined stickleback is not related to virulence. Even though the presence of the parasite caused differences in host condition, these were indistinguishable between the different levels of virulence in this experiment. Fish immune traits also showed a response to infection but had no correlation with level of parasite virulence. With this experiment we have taken the first step towards understanding how virulence is inherited and how it is driven in the Schistocephalus-stickleback system, even though virulence, as measured here, does not directly translate into cost for the host. A better understanding of the costs inflicted on the host by S. solidus infection is needed to understand this interaction in greater detail.

Validation of a novel saliva-based ELISA test for diagnosing tapeworm burden in horses.

BACKGROUND: Tapeworm infections pose a significant threat to equine health as they are associated with clinical cases of colic. Diagnosis of tapeworm burden using fecal egg counts (FECs) is unreliable, and, although a commercial serologic ELISA for anti-tapeworm antibodies is available, it requires a veterinarian to collect the blood sample. A reliable diagnostic test using an owner-accessible sample such as saliva could provide a cost-effective alternative for tapeworm testing in horses, and allow targeted deworming strategies. OBJECTIVES: The purpose of the study was to statistically validate a saliva tapeworm ELISA test and compare to a tapeworm-specific IgG(T) serologic ELISA. METHODS: Serum samples (139) and matched saliva samples (104) were collected from horses at a UK abattoir. The ileocecal junction and cecum were visually examined for tapeworms and any present were counted. Samples were analyzed using a serologic ELISA and the saliva tapeworm test. The test results were compared to tapeworm numbers and the various data sets were statistically analyzed. RESULTS: Saliva scores had strong positive correlations with both infection intensity (0.74) and serologic results (Spearman's rank coefficients; 0.74 and 0.86, respectively). The saliva tapeworm test was capable of identifying the presence of one or more tapeworms with 83% sensitivity and 85% specificity. Importantly, no high-burden (more than 20 tapeworms) horses were misdiagnosed. CONCLUSIONS: The saliva tapeworm test has statistical accuracy for detecting tapeworm burdens in horses with 83% sensitivity and 85% specificity, similar to those of the serologic ELISA (85% and 78%, respectively).

Reciprocal cross infection of sticklebacks with the diphyllobothriidean cestode Schistocephalus solidus reveals consistent population differences in parasite growth and host resistance.

BACKGROUND: In host-parasite evolutionary arms races, parasites are generally expected to adapt more rapidly, due to their large population sizes and short generation times. There exist systems, though, where parasites cannot outpace their hosts because of similar generation times in both antagonists. In those cases concomitant adaptation is expected. METHODS: We tested this hypothesis in the three-spined stickleback-Schistocephalus solidus tapeworm system, where generation times are comparable in both organisms. We chose two populations of sticklebacks which differ prominently in the prevalence of S. solidus and consequently in its level of selective pressure. We performed a full factorial common garden experiment. Particularly, Norwegian (NO) and German (DE) sticklebacks, as well as hybrids between both stickleback populations and in both parental combinations, were exposed each to a single S. solidus originating from the same two host populations. RESULTS: We found the infection phenotype to depend on the host population, the parasite population, but not their interaction. NO-parasites showed higher infectivity than DE-parasites, with NO-sticklebacks also being more resistant to DE-parasites than to the sympatric NO-parasite. Reciprocally, DE-hosts were more susceptible to the allopatric NO-parasite while DE-parasites grew less than NO-parasites in all stickleback groups. Despite this asymmetry, the ratio of worm to host weight, an indicator of parasite virulence, was identical in both sympatric combinations, suggesting an optimal virulence as a common outcome of parallel coevolved systems. In hybrid sticklebacks, intermediate infection rates and growth of S. solidus from either origin suggests a simple genetic basis of resistance. However, comparison of infection phenotypes in NO-maternal and DE-maternal hybrid sticklebacks indicates local adaptation to the sympatric counterpart in both the host and the parasite. CONCLUSIONS: Host-parasite systems with similar generation time show evidence for concomitant reciprocal adaptation resulting in parasite optimal virulence and host parasite specific resistance.

Immunogenic activity of the fish tapeworm Pterobothrium heteracanthum (Trypanorhyncha: Pterobothriidae) in BALB/c mice.

The aim of this study was to verify the immunogenicity of Pterobothrium heteracanthum (Cestoda: Trypanorhyncha) crude protein extract (PH-CPE) in BALB/c mice. The parasites were obtained from Micropogonias furnieri (Osteichthyes: Sciaenidae). Groups of six mice were each immunized with 10, 50 or 100 µg of PH-CPE, on days 0 and 35. Both specific IgG and IgE responses were developed after immunization. The immunoblot assay revealed that specific IgG recognizes PH-CPE proteins with two molecular weight ranges, 60-75 and 30-40 kDa, and that IgE recognizes larger proteins over 120 kDa. This appears to be the first report on the immunogenicity of metacestodes within the Pterobothriidae and that PH-CPE is a potential inducer of a specific IgE response.

In vitro effects of prostaglandin E2 on leucocytes from sticklebacks (Gasterosteus aculeatus) infected and not infected with the cestode Schistocephalus solidus.

Many helminth parasites have evolved strategies to evade the immune response of their hosts, which includes immunomodulation. Prostaglandin E2 (PGE2) is one of the best-described immunomodulators in mammalian helminth parasite infections. We hypothesized that also in teleost fish anti-helminthic immune responses are regulated via PGE2. We used a model system consisting of the tapeworm Schistocephalus solidus and its host, the three-spined stickleback (Gasterosteus aculeatus), to investigate in vitro effects of PGE2 on head kidney leucocytes (HKL) derived from sticklebacks that were experimentally infected with S. solidus. PGE2 was tested alone or in combination with either S. solidus antigens or bacterial lipopolysaccharides (LPS). After in vitro culture, cell viability and changes in leucocyte subpopulations (granulocytes to lymphocytes ratios) were monitored by flow cytometry and HKL were tested for their capacity to produce reactive oxygen species (ROS) with a chemiluminescence assay. In short term (2 h) HKL cultures PGE2 did not change the total numbers of live HKL, but the production of ROS decreased significantly with high (0.1 µmol L(-1)) PGE2 concentrations. In long-term (96 h) cultures high PGE2 concentrations induced a sharp decrease of leucocytes viability, while low (0.1 pmol L(-1)) and intermediate (0.1 nmol L(-1)) concentrations of PGE2 caused elevated leucocyte viability compared to controls. This coincided with reduced ROS production in cultures with high PGE2 and elevated ROS production in cultures with low PGE2. Granulocyte to lymphocyte ratios increased with high PGE2 concentrations alone and in combination with S. solidus antigens and LPS, most prominently with HKL from S. solidus infected sticklebacks. The present study supports the hypothesis that PGE2 might be an immunomodulator in tapeworm-fish parasite-host interactions.

Perch liver reaction to Triaenophorus nodulosus plerocercoids with an emphasis on piscidins 3, 4 and proliferative cell nuclear antigen (PCNA) expression.

Histopathological lesions caused by plerocercoids of Triaenophorus nodulosus within the liver of perch, Perca fluviatilis, from Lake Trasimeno were studied. Livers harbored 1-3 parasite larvae and pathological alterations were more marked in those with 3 plerocercoids. In the liver, larvae were encysted, surrounded by a capsule of host tissue; two of 14 plerocercoids were necrotic. In infected livers, some hepatocytes showed degenerative changes, i.e. swelling and hydropic degeneration, notably those in close proximity to larvae. By comparison, hepatocytes in uninfected livers or in regions away from the point of infection appeared normal. The occurrence of macrophage aggregates (MAs) distributed among the mast cells (MCs) was observed around the encysted larvae. The cellular elements involved in the immune response within liver were assessed by immunohistochemical techniques and by the use of antibodies against the antimicrobial peptides piscidins 3 and 4, which revealed a sub-population of positive MCs. In infected livers, numerous MCs that were immunopositive to P4 and a few that were positive to P3 were found around T. nodulosus larvae. Histological sections of both uninfected and infected liver were immunostained with proliferative cell nuclear antigen (PCNA) antibody. Within the capsule and in close proximity to the parasite larvae, various cell types (i.e., MCs, fibroblasts and epithelioid cells) and a significantly higher number of PCNA-positive hepatocytes that were immunoreactive to PCNA were found compared to uninfected livers (ANOVA, P<0.05). No parasites of any type were found in gill, spleen, kidney or gonad of P. fluviatilis and the intestine of 3 perch were infected with few specimens of Acanthocephalus lucii.

Systemic and mucosal IgE antibody responses of horses to infection with Anoplocephala perfoliata.

Infection of horses with Anoplocephala perfoliata induces a severe inflammatory reaction of the caecal mucosa around the site of parasite attachment adjacent to the ileocecal valve. Lesions show epithelial erosion or ulceration of the mucosa with infiltration by eosinophils, lymphocytes and mast cells leading to oedema, gross thickening and fibrosis of the caecal wall. Despite this evidence of an inflammatory reaction to A. perfoliata within the mucosa of the caecum there is little information about the nature of the local immune response to A. perfoliata. An ELISA which assays serum IgG(T) antibodies to A. perfoliata excretory/secretory antigens has been developed as a diagnostic test. However, the specificity of the ELISA remains sub-optimal and the role of other isotypes in the immune response to A. perfoliata has not been reported. This study measured IgA, IgE and IgG(T) antibody responses to A. perfoliata excretory/secretory antigens in sera of 75 horses presented for slaughter. The prevalence of A. perfoliata infection, as confirmed by the presence of parasites in the terminal ileum, caecum or proximal colon, was 55%. A. perfoliata-specific IgG(T) and IgE antibodies were significantly elevated in infected horses compared to controls; IgA antibodies were also detected but did not differ between infected and control horses. Diagnosis by serum IgG(T) ELISA had a sensitivity of 78% and a specificity of 80%, by comparison the serum IgE ELISA had a sensitivity of just 44% with a specificity of 82% and therefore did not provide an improved diagnostic test. Western blots with sera from infected horses demonstrated IgE-binding to at least 10 separate components of excretory/secretory (E/S) antigens. A similar pattern was also found with IgG(T). Around 30% of horses had high levels of serum IgE which bound fucose-containing carbohydrate antigens on the parasite surface but this was unrelated to the presence of A. perfoliata infection. Immunoperoxidase staining detected numerous IgE-positive cells within lymphoid follicles in the caecal mucosa close to the site of A. perfoliata attachment and quantitative RT-PCR detected high levels of IgE transcription in the caecal mucosa of all horses. Mucosal synthesis of antibodies was confirmed by the demonstration of A. perfoliata-specific IgG(T) and IgE in the supernatant of lamina propria explant cultures that discriminated clearly between infected and uninfected horses. We conclude that there is an active immune response to A. perfoliata within the caecal mucosa involving local production of both IgG(T) and IgE antibody isotypes; but it remains unclear whether this immune response can reduce or eliminate parasite burden.