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Helminths are known to be strong modulators of host immune responses and inflammation to ensure their own survival within their hosts. Paradoxically, deliberate infection with helminths has been shown to alleviate symptoms of autoimmune disorders in animal models, and also human patients with some proof-of-concept with live-worm therapy. Identification of the mechanisms responsible for this immunomodulation may be a major step towards design of effective vaccines for helminth infection, and for identification of helminth-derived molecules which may be developed as novel anti-inflammatory agents. Ascaris suum is a swine helminth and one of the most common helminths present in pig production, and closely related to the human A. lumbricoides. Ascaris infections are known to result in pronounced modulation of host immune responses, and molecules derived from A. suum have shown some promise in treating inflammatory disorders in mouse models. In order to investigate possible mechanisms driving the immunomodulatory capabilities of A. suum, the aim of this study was to investigate if A. suum adult body fluid (ABF) has immunomodulatory effects on human monocyte-derived macrophages (Mɸ) in vitro. Mɸs play a crucial role in innate and adaptive immunity and are major mediators of the inflammatory response. The experiments were divided in three groups: 1) Mɸ were differentiated into classically activated (M1), alternatively activated (M2) and uncommitted (M0) phenotypes with granulocyte macrophage colony-stimulating factor (GM-CSF), macrophage colony-stimulating factor (M-CSF) and Human Serum, respectively, and stimulated with A. suum ABF, lipopolysaccharide (LPS) or both; 2) A. suum ABF was added to M0 Mɸs which were co-incubated with interferon-gamma (IFN-γ) plus LPS for the M1 phenotype or interleukin (IL)-4 for M2; 3) Monocytes were differentiated into Mɸs in the presence of A. suum ABF and GM-CSF, and after Mɸ differentiation LPS was added. The pro-inflammatory cytokines tu

Background: A common characteristic of Trichuris spp. infections in humans and animals is a low to varied efficacy of single-dose benzimidazoles, used in mass drug administration programmes against human trichuriasis. The bacillary band, a specialised morphological structure of Trichuris spp., as well as the unique partly intra- and extracellular habitat of adult Trichuris spp., may be involved in restricted drug absorption, resulting in a low drug accumulation. However, the function of the bacillary band is still unknown. Methodology: We studied whether adult worms of T. muris were dependent on glucose and/or amino acids for survival in vitro and if the bacillary band has an absorptive function. The viability of the worms was evaluated microscopically using a motility scale from 0 - 3, and by measuring emission in response to metabolic activity utilizing the colorimetric assay Alamar Blue. The absorptive function of the bacillary band was explored using a fluorescent glucose analogue (6-NBDG) detected in the living worms by confocal microscopy. To exclude oral ingestion of 6-NBDG, the oral cavity of T. muris worms was sealed. Principal Findings: Adult worms of T. muris incubated in media devoid of glucose were immotile after 17 hours incubation and generated an emission signal similar to controls (RPMI media without worms). The fluorescent glucose analogue was detected in the pores of the bacillary band and accumulated inside the worms, primarily within the stichocytes in living worms with and without a sealed oral cavity. Conclusions/Significance: The whipworm T. muris is dependent on glucose for survival in vitro, and the bacillary band has an absorptive function in relation to the glucose analogue 6-NBDG which accumulates within the stichocytes of the worms. This new insight into the absorptive function of the bacillary band may encourage for further explorations of the bacillary band in relation to anthelmintics.

Background The pig parasite Ascaris suum plays and important role in veterinary medicine and represents a suitable model for A. lumbricoides, which infects over 800 million people. In pigs, continued exposure to Ascaris induces immunity at the level of the gut, protecting the host against migrating larvae. The objective of this study was to identify and characterize parasite antigens targeted by this local immune response that may be crucial for parasite invasion and establishment and to evaluate their protective and diagnostic potential. Methodology/Principal Findings Pigs were immunized by trickle infection for 30 weeks, challenged with 2,000 eggs at week 32 and euthanized two weeks after challenge. At necropsy, there was a 100% reduction in worms recovered from the intestine and a 97.2% reduction in liver white spots in comparison with challenged non-immune control animals. Antibodies purified from the intestinal mucus or from the supernatant of cultured antibody secreting cells from mesenteric lymph nodes of immune pigs were used to probe L3 extracts to identify antibody targets. This resulted in the recognition of a 12kDa antigen (As12) that is actively shed from infective Ascaris L3. As12 was characterized as a phosphorylcholine-containing glycolipid-like antigen that is highly resistant to different enzymatic and chemical treatments. Vaccinating pigs with an As12 fraction did not induce protective immunity to challenge infection. However, serological analysis using sera or plasma from experimentally infected pigs or naturally infected humans demonstrated that the As12 ELISA was able to detect long-term exposure to Ascaris with a high diagnostic sensitivity (98.4% and 92%, respectively) and specificity (95.5% and 90.0%) in pigs and humans, respectively. Conclusions/Significance These findings show the presence of a highly stage specific, glycolipid-like component (As12) that is actively secreted by infectious Ascaris larvae and wh

BACKGROUND: Trichuris suis and T. trichiura are two different whipworm species that infect pigs and humans, respectively. T. suis is found in pigs worldwide while T. trichiura is responsible for nearly 460 million infections in people, mainly in areas of poor sanitation in tropical and subtropical areas. The evolutionary relationship and the historical factors responsible for this worldwide distribution are poorly understood. In this study, we aimed to reconstruct the demographic history of Trichuris in humans and pigs, the evolutionary origin of Trichuris in these hosts and factors responsible for parasite dispersal globally.METHODS: Parts of the mitochondrial nad1 and rrnL genes were sequenced followed by population genetic and phylogenetic analyses. Populations of Trichuris examined were recovered from humans (n = 31), pigs (n = 58) and non-human primates (n = 49) in different countries on different continents, namely Denmark, USA, Uganda, Ecuador, China and St. Kitts (Caribbean). Additional sequences available from GenBank were incorporated into the analyses.RESULTS: We found no differentiation between human-derived Trichuris in Uganda and the majority of the Trichuris samples from non-human primates suggesting a common African origin of the parasite, which then was transmitted to Asia and further to South America. On the other hand, there was no differentiation between pig-derived Trichuris from Europe and the New World suggesting dispersal relates to human activities by transporting pigs and their parasites through colonisation and trade. Evidence for recent pig transport from China to Ecuador and from Europe to Uganda was also observed from their parasites. In contrast, there was high genetic differentiation between the pig Trichuris in Denmark and China in concordance with the host genetics.CONCLUSIONS: We found evidence for an African origin of T. trichiura which were then transmitted with human ancestors to Asia and further to South

Background The Internal Transcribed Spacer 2 (ITS2) is a candidate diagnostic marker of the pathogenic cattle nematode Ostertagia ostertagi. The aims of this study were: (i) to document and quantify how the development of O. ostertagi eggs affects ITS2 copies under different storage conditions, and (ii) to suggest optimal storage conditions for faecal samples in a diagnostic pipeline that involves detection and semi-quantification by real-time semi-quantitative polymerase chain reaction (qPCR). Findings Eggs of Ostertagia ostertagi were obtained from fresh faeces and stored at 4 °C or 25 °C under aerobic or anaerobic (vacuum packing) conditions. Development was monitored by microscopy for up to 336 h, and the ITS2 copies were determined by qPCR from a fixed number of parasites. Under aerobic conditions at 25 °C, embryonation and a significant increase of ITS2 copies (P < 0.0001) were observed after 12 h. At 4 °C, embryonation occurred after 168 h with a trend towards increased ITS2 copies. Anaerobic conditions inhibited egg development at both temperatures and no significant increase in ITS2 copies was noticed (P = 0.90). ITS2 copies were analysed for each parasite stage: first-stage larvae (L1) exhibited significantly higher copy numbers (20,353 ± 1,950) than unembryonated eggs (568 ± 168; P < 0.0001) with lower coefficient of variation (33 vs 266 %). Conclusions Aerobic storage of O. ostertagi eggs at 25 °C led to a significant increase in ITS2 copies after 12 h due to embryonation and subsequent hatching. In contrast, anaerobic storage (vacuum packing) at 25 °C completely inhibited egg development and any undesirable semi-quantification bias for up to 336 h. Hence, vacuum packing is an optimal storage strategy prior to molecular diagnostic analyses. Alternatively, aerobic storage at 4 °C for up to 72 h can be used. Due to high copy numbers and lower genetic variation, the L1 stage may be considered for diagnostics and further molecular research., Background: The Internal Transcribed Spacer 2 (ITS2) is a candidate diagnostic marker of the pathogenic cattle nematode Ostertagia ostertagi. The aims of this study were: (i) to document and quantify how the development of O. ostertagi eggs affects ITS2 copies under different storage conditions, and (ii) to suggest optimal storage conditions for faecal samples in a diagnostic pipeline that involves detection and semi-quantification by real-time semi-quantitative polymerase chain reaction (qPCR). Findings: Eggs of Ostertagia ostertagi were obtained from fresh faeces and stored at 4 °C or 25 °C under aerobic or anaerobic (vacuum packing) conditions. Development was monitored by microscopy for up to 336 h, and the ITS2 copies were determined by qPCR from a fixed number of parasites. Under aerobic conditions at 25 °C, embryonation and a significant increase of ITS2 copies (P < 0.0001) were observed after 12 h. At 4 °C, embryonation occurred after 168 h with a trend towards increased ITS2 copies. Anaerobic conditions inhibited egg development at both temperatures and no significant increase in ITS2 copies was noticed (P = 0.90). ITS2 copies were analysed for each parasite stage: first-stage larvae (L1) exhibited significantly higher copy numbers (20,353 ± 1,950) than unembryonated eggs (568 ± 168; P < 0.0001) with lower coefficient of variation (33 vs 266 %). Conclusions: Aerobic storage of O. ostertagi eggs at 25 °C led to a significant increase in ITS2 copies after 12 h due to embryonation and subsequent hatching. In contrast, anaerobic storage (vacuum packing) at 25 °C completely inhibited egg development and any undesirable semi-quantification bias for up to 336 h. Hence, vacuum packing is an optimal storage strategy prior to molecular diagnostic analyses. Alternatively, aerobic storage at 4 °C for up to 72 h can be used. Due to high copy numbers and lower genetic variation, the L1 stage may be considered for diagnostics and further molecular research.

microRNAs (miRNAs) are recently discovered as key regulators of gene translation and are becoming increasingly recognized for their involvement in various diseases. This study investigates the miRNA profile in pig serum during the course of an infection with the gastrointestinal parasite, Trichuris suis. Of this panel, the expression of selected miRNAs in serum from T. suis infected and uninfected pigs were determined by quantitative real time PCR using Exiqon Human Panel assays at 0, 2, 4, 6, 8 and 10 weeks post first infection (wpi). One miRNA, ssc-let-7d-3p, was significantly up-regulated in infected pigs 8 wpi. Interestingly, ssc-let-7d-3p shows high complementary to tsu-let-7a, which is the most highly transcribed miRNA in T. suis. The let-7 family miRNAs have been shown to post-transcriptionally regulate the translation of the helminth-controlling cytokine, IL-13, in a murine model for asthma and we hypothesize possible interactions between these host- and parasite-derived miRNAs and their immunomodulating roles., microRNAs (miRNAs) are recently discovered as key regulators of gene translation and are becoming increasingly recognized for their involvement in various diseases. This study investigates the miRNA profile in pig serum during the course of an infection with the gastrointestinal parasite, Trichuris suis. Of this panel, the expression of selected miRNAs in serum from T. suis infected and uninfected pigs were determined by quantitative real time PCR using Exiqon Human Panel assays at 0, 2, 4, 6, 8 and 10 weeks post first infection (wpi). One miRNA, ssc-let-7d-3p, was significantly up-regulated in infected pigs 8 wpi. Interestingly, ssc-let-7d-3p shows high complementary to tsu-let-7a, which is the most highly transcribed miRNA in T. suis. The let-7 family miRNAs have been shown to post-transcriptionally regulate the translation of the helminth-controlling cytokine, IL-13, in a murine model for asthma and we hypothesize possible interactions between these host- and parasite-derived miRNAs and their immunomodulating roles.

Summary Humans can be infected by a range of different parasites including both protozoa and helminths. The interactions between parasites and the host immune system are of great interest, as parasites are known to modulate the host immune response to ensure their own survival, utilising a range of different mechanisms to do so. As co-infections commonly occur between diverse parasites, interactions and immuno-modulatory mechanisms attract much interest. One of the main cell types at the forefront of the host immune response are dendritic cells (DCs). Interactions between DCs and parasite antigens could be important in understanding parasite-host cell interactions. The aim of this study was to investigate qualitative differences of how DCs respond to antigens from diverse parasites and whether protozoan and helminth antigen interacted to modulate DC function. Immature human DCs were matured with lipopolysaccharide in the presence or absence of protozoan antigens from Giardia duodenalis or Plasmodium falciparum and from helminth antigens Ascaris suum or Trichuris suis. Cytokine secretion and cell surface marker expression were quantified to measure inflammatory responses and maturation status respectively. To determine interactive effects, analysis of additivity was carried out. Overall, the results showed a remarkably conserved anti-inflammatory response of DCs to diverse parasite material originating from diverse protozoa and helminths. This supported previous findings of all four antigens suppressing pro-inflammatory cytokine production. Secretion of the anti-inflammatory cytokine interleukin (IL)-10 was enhanced by all except A. suum. Exposure of DCs to G. duodenalis resulted in significant apoptosis which was not observed for the other parasites. Interestingly, DCs did not respond in an additive fashion to co-stimulation with the different antigens. The exact causes of this are unknown but could possibly be due to different mechanisms of

Four species of ancylostomatids are considered as canine hookworms, Ancylostoma caninum, A. braziliense, A. ceylanicum and Uncinaria stenocephala. Despite their veterinary relevance, they are zoonotic and able to cause different clinical manifestations and patent infections in humans. Little is known about the hookworm species present in Africa and their distribution. Consequently the main objective of this thesis was to identify, by the use of molecular techniques, the hookworm species present Abstract in canine faecal samples from Morogoro, Tanzania. A second (and minor) objective was to identify gastrointestinal helminth infections in the collected samples by copromicroscopy. The fieldwork was based in Sokoine University of Agriculture (SUA), Morogoro, Tanzania where faecal samples from 160 dogs were collected in four different areas. Hookworm positive samples were processed in order to recover larvae for further molecular characterisation. The molecular work took place at University of Copenhagen, Denmark where 68 samples brought from Morogoro were analyzed. Two different molecular approaches were used PCR-RFLP and newly designed species-specific primers coupled with sequencing. Three different species of gastrointestinal helminths were found. Hookworms were present in 58% of the samples followed by Toxocara spp (6%) and Dipylidium caninum (1%). The species and proportions found by copromicroscopy were in line with previous prevalence reports. By the use of PCR-RFLP we were only able to detect the presence of the ubiquitous A. caninum. However, by sequencing some of the species-specific PCR positive we successfully identify A. braziliense, U. stenocephala and A. ceylanicum. This is the first study, to my knowledge, documenting the presence of the zoonotic A. ceylanicum and U. stenocephala by molecular characterisation in Africa.

Human trichuriasis is a neglected tropical disease which affects hundreds of millions of people worldwide and is particularly prevalent among children living in areas where sanitation is poor. This review examines the current knowledge on the taxonomy, genetics and phylogeography of human Trichuris and its relationship to whipworm parasites in other host species. The evidence for zoonotic transmission of Trichuris and the emergence of anthelmintic resistance is assessed. In addition, the implications of the recent publication of the genomes and transcriptomes of multiple Trichuris species are discussed. Finally, priorities for future research in Trichuris genetics are proposed.

This histopathological study was carried out in order to investigate the cellular response in the jejunum to Ascaridia galli during the first 7 weeks of infection. Fourty-two ISA Brown chickens (7 weeks old) were infected orally with 500 embryonated A. galli eggs each while 28 chickens were left as uninfected controls. Six infected and four control chickens were necropsied at each time point 3, 7, 10, 14, 21, 28 and 42 days post-infection (dpi). Samples for histopathology were taken from three sites of the jejunoileum. Significantly higher eosinophil counts were seen in infected chickens compared to uninfected at 3, 7, 10, 14 and 28 dpi (P < 0.01). In both groups, the initial number of mast cells was high, but this high level of mast cells remained for a longer period in the infected group compared to the control group. Significantly higher counts were thus found in the infected group at 21 (P < 0.001), 28 (P < 0.01) and 42 dpi (P < 0.05). A. galli infection induced changes in the mucosal thickness as reduced villi length at 7, 10, 14, 21 and 28 dpi and in the degree of general cellular infiltration in the lamina propria of the mucosal layer. No adult worms were seen during the experiment; therefore, A. galli larvae have elicited a moderate cellular response in the lamina propria, mainly consisting of eosinophils in the early phase and later of mast cells.

BACKGROUND: The whipworms Trichuris trichiura and Trichuris suis are two parasitic nematodes of humans and pigs, respectively. Although whipworms in human and non-human primates historically have been referred to as T. trichiura, recent reports suggest that several Trichuris spp. are found in primates.METHODS AND FINDINGS: We sequenced and annotated complete mitochondrial genomes of Trichuris recovered from a human in Uganda, an olive baboon in the US, a hamadryas baboon in Denmark, and two pigs from Denmark and Uganda. Comparative analyses using other published mitochondrial genomes of Trichuris recovered from a human and a porcine host in China and from a françois' leaf-monkey (China) were performed, including phylogenetic analyses and pairwise genetic and amino acid distances. Genetic and protein distances between human Trichuris in Uganda and China were high (~19% and 15%, respectively) suggesting that they represented different species. Trichuris from the olive baboon in US was genetically related to human Trichuris in China, while the other from the hamadryas baboon in Denmark was nearly identical to human Trichuris from Uganda. Baboon-derived Trichuris was genetically distinct from Trichuris from françois' leaf monkey, suggesting multiple whipworm species circulating among non-human primates. The genetic and protein distances between pig Trichuris from Denmark and other regions were roughly 9% and 6%, respectively, while Chinese and Ugandan whipworms were more closely related.CONCLUSION AND SIGNIFICANCE: Our results indicate that Trichuris species infecting humans and pigs are phylogenetically distinct across geographical regions, which might have important implications for the implementation of suitable and effective control strategies in different regions. Moreover, we provide support for the hypothesis that Trichuris infecting primates represents a complex of cryptic species with some species being able to infect both humans and non-human

A humoral immune response following helminth infection in pigs is well documented. However, it has been difficult to confirm the existence of antibody mediated resistance against the large roundworm, Ascaris suum, and whipworm, Trichuris suis, in experimental settings by correlating worm burdens or egg excretion with specific antibody levels. We set out to investigate the association between worm load and T. suis and A. suum specific serum antibody levels (IgG1, IgG2 and IgA) against excretory-secretory products of adults and third stage larvae, respectively, measured at 0, 7 and 14 weeks p.i. in a trickle-infected F1-resource-population of crossbred pigs (n=195). Furthermore, we wanted to determine the heritability of these antibody isotypes during the course of infection. Most pigs remained infected with A. suum throughout the experiment while they expelled T. suis between 7 and 14 weeks post infection (p.i.). Parasite specific IgG1 and IgA were significantly (P<0.001) elevated after 7 and 14 weeks of infection, whereas parasite specific IgG2 levels only changed slightly at 14 weeks p.i.. However, the observed association between specific antibody isotype levels and faecal egg counts and macroscopic worm load was weak. The relative heritabilities of the different parasite specific isotypes were assessed and resulted in significant heritability estimates for parasite specific IgG1 and IgA. The highest heritabilities were found for A. suum specific IgG1 (h(2)=0.41 and 0.46 at 7 and 14 weeks p.i., respectively). Thus, the present study demonstrates that host genetic factors influence the IgG1 and IgA antibody isotype responses specific to two of the most common gastrointestinal nematodes of swine whereas specific antibody levels were poorly associated with egg excretion and the presence of macroscopic worms.

Few studies have been carried out in Africa toestimate the prevalence of Taeniahydatigena. With theaim to determine theprevalence of T. hydatigena inslaughtered pigs and small ruminants (goats and sheep) in Mbeya, Tanzania, two cross-sectional surveys were carried out investigating pigs in Aprilto May 2014 and small ruminants in September 2012. In total, 243 pigs were examined post-mortem for T. hydatigena cysts which were found in 16 (6.6%) pigs. The majority(80%) of cysts were found on the omentum and the rest on the liver (20%), allon the visceral surface. Two pigs were also found infected with T. solium, but showed no signs of otherinfections. A total of 392 goats and 27 sheep were examined post-mortem and theprevalence of T. hydatigena wassimilar in goats and sheep with 45.7% and 51.9%, respectively. DNA sequencing of the mitochondrial cytochrome c oxidase subunit 1 gene (cox1) from a subsample of metacestodesfrom goats and sheep confirmed the T. hydatigena infection. Theprevalence found in small ruminants was comparable to other studies conductedin Africa, but for pigs it is one of the highest recorded to date. The presentstudy also confirms the occurrence of T.hydatigena and T. solium in pigsfrom Mbeya. Further studies are needed to determine the impact of T. hydatigena on production undersub-Saharan conditions and the financial consequences for smallholder farmers.

Trichuris suis is a common parasitic helminth of pigs. As with many other parasites, T. suis ensures its own survival by evading host immune responses, but little is known about how this is achieved. MicroRNAs (miRNA) have been shown to be involved in various immunological processes by post-transcriptional regulation of specific genes, and the potential of using these molecules as biomarkers of disease is currently being examined. It has recently been shown that parasites may secrete extracellular structures such as exosomes and microvesicles, containing proteins and miRNA. The fusion of these structures with host cells has been demonstrated, and a role of exosome-derived miRNA in host gene regulation has been suggested. In the present study, we show that exosome- and microvesicular-like structures are secreted by T. suis L1 larvae and also demonstrate the presence of miRNA-sized RNA inside these structures. A potential role of these molecules in host-parasite interactions is suggested. In addition, an electron-dense layer covering the surface of the larvae was observed, which may play a function in host immune evasion.

The aims of this study were to assess how the development of Ostertagia ostertagi eggs into first-stage larva (L1) affects the copy numbers of the Internal Transcribed Spacer region 2 (ITS2) of the ribosomal DNA; and based on these results, to suggest optimal storage conditions for faecal samples prior to detection and quantification by real-time quantitative polymerase chain reaction (qPCR). Fresh O. ostertagi eggs were isolated from cattle faeces and stored at 4°C or 25°C under aerobic or anaerobic conditions. Embryonation was monitored by microscopy and the ITS2 copies were determined by qPCR at predetermined intervals for up to 336 h. Under aerobic conditions, L1 was observed after 24 h at 25°C, while development to L1 took 336 h at 4°C. A corresponding significant increase of the ITS2 copies was also observed (p < 0.0001). However, anaerobic conditions inhibited embryonation at both temperatures and no significant effect of storage on ITS2 copies was noticed (p =0.8984). ITS2 copies were significantly higher in L1 compared with copies in unembryonated eggs (p <0.0001) and with lower coefficients of variation for L1 (33%) compared with unembryonated eggs (266%). In conclusion, storage conditions affect the outcome of qPCR analysis for the quantitative determination of O. ostertagi eggs in cattle faeces. Cold storage at 4°C for up to 3 days or anaerobicvacuum packing at 25°C for up to 336 h will entail no undesirable effects on ITS2 copies.

BACKGROUND: Since the nematodes Trichuris trichiura and T. suis are morphologically indistinguishable, genetic analysis is required to assess epidemiological cross-over between people and pigs. This study aimed to clarify the transmission biology of trichuriasis in Ecuador.FINDINGS: Adult Trichuris worms were collected during a parasitological survey of 132 people and 46 pigs in Esmeraldas Province, Ecuador. Morphometric analysis of 49 pig worms and 64 human worms revealed significant variation. In discriminant analysis morphometric characteristics correctly classified male worms according to host species. In PCR-RFLP analysis of the ribosomal Internal Transcribed Spacer (ITS-2) and 18S DNA (59 pig worms and 82 human worms), nearly all Trichuris exhibited expected restriction patterns. However, two pig-derived worms showed a "heterozygous-type" ITS-2 pattern, with one also having a "heterozygous-type" 18S pattern. Phylogenetic analysis of the mitochondrial large ribosomal subunit partitioned worms by host species. Notably, some Ecuadorian T. suis clustered with porcine Trichuris from USA and Denmark and some with Chinese T. suis.CONCLUSION: This is the first study in Latin America to genetically analyse Trichuris parasites. Although T. trichiura does not appear to be zoonotic in Ecuador, there is evidence of genetic exchange between T. trichiura and T. suis warranting more detailed genetic sampling.

One of the most common zoonotic helminth infections is caused by species in the genus Toxocara, particularly Toxocara canis and T. cati (Syn. T. mystax). However, their relative contribution to toxocarosis in humans remains largely unknown because causative larvae are seldom recovered and uncertainties regarding the validity of existing serological assays. In this study, we used sera from a pig model experimentally infected with T. canis and T. cati to evaluate whether a Western blot could discriminate between the two species. No proteins were observed that could be used as a diagnostic tool. In addition, a heterogenic protein pattern between individual hosts was found, which was most pronounced in the T. cati-infected pigs. There is therefore an urgent need to optimize and validate current methods or develop new species-specific serological methods in order to implement appropriate control measures.

This study investigated the changes in establishment rates during the time course of a 6week trickle infection of chickens with Ascaridia galli at two different dose levels, using a molecular marker. To differentiate early and late infection, two different egg cohorts (haplotype a and haplotype b, genetically identified using PCR-linked restriction fragment length polymorphism on the cox1 gene of the mitochondrial DNA) were used. Cohort-specific egg batches were produced by harvesting eggs from the uteri of female worms of the specific cohort. Fifty-six 8week old Lohmann Brown Lite chickens were divided into seven groups and the infectivity of the egg batches was compared between two groups of chickens (P=0.6). The remaining chickens were allocated to four infection regimes and one control group. Group ab100 was trickle infected for 3weeks with 100 eggs of haplotype a (twice weekly) followed by the same dose of eggs of haplotype b for another 3weeks. Group ba100 was treated similarly but in the opposite order (haplotype b preceding a). A similar infection regime was applied for groups ab25 and ba25 but with a lower inoculation dose (25 eggs). All of the birds in these five groups (four infected and one control) were euthanased 2weeks after the last inoculation. It was found that in the low-dose groups both the early and late infections established equally well, whereas in the high-dose groups the early infection was recovered in a significantly (P<0.001) higher proportion of chickens than the late infection, irrespective of genetic cohorts. Moreover, relatively higher proportions of the larvae from both the early and late infections were found in the posterior section of the small intestine. This result indicates the presence of dose-dependent resistance against reinfection and this resistance seems to act by reducing the establishment of late infection and by relocating the larvae from early infection.

Toxocara canis is a zoonotic parasite of major socioeconomic importance worldwide. In humans, this nematode causes disease (toxocariasis) mainly in the under-privileged communities in developed and developing countries. Although relatively well studied from clinical and epidemiological perspectives, to date, there has been no global investigation of the molecular biology of this parasite. Here we use next-generation sequencing to produce a draft genome and transcriptome of T. canis to support future biological and biotechnological investigations. This genome is 317 Mb in size, has a repeat content of 13.5% and encodes at least 18,596 protein-coding genes. We study transcription in a larval, as well as adult female and male stages, characterize the parasite's gene-silencing machinery, explore molecules involved in development or host-parasite interactions and predict intervention targets. The draft genome of T. canis should provide a useful resource for future molecular studies of this and other, related parasites.

Whipworms (Trichuris spp.) infect a variety of hosts, including domestic animals and humans. Of considerable interest is the porcine whipworm, T. suis, which is particularly prevalent in outdoor production systems. High infection levels may cause growth retardation, anaemia and haemorrhagic diarrhoea. A significant proportion of the variation in Trichuris faecal egg count (FEC) has been attributed to the host's genetic make-up. The aim of the present study was to identify genetic loci associated with resistance to T. suis in pigs. We used single nucleotide polymorphism (SNP) markers to perform a whole-genome scan of an F1 resource population (n=195) trickle-infected with T. suis. A measured genotype analysis revealed a putative quantitative trait locus (QTL) for T. suis FEC on chromosome 13 covering ∼4.5Mbp, although none of the SNPs reached genome-wide significance. We tested the hypothesis that this region of SSC13 harboured genes with effects on T. suis burden by genotyping three SNPs within the putative QTL in unrelated pigs exposed to either experimental or natural T. suis infections and from which we had FEC (n=113) or worm counts (n=178). In these studies, two of the SNPs (rs55618716, ST) were associated with FEC (P<0.01), thus confirming our initial findings. However, we did not find any of the SNPs to be associated with T. suis worm burden. In conclusion, our study demonstrates that genetic markers for resistance to T. suis as indicated by low FEC can be identified in pigs.

Ancient parasite eggs were recovered from environmental samples collected at a Viking-age settlement in Viborg, Denmark, dated 1018-1030 A.D. Morphological examination identified Ascaris sp., Trichuris sp., and Fasciola sp. eggs, but size and shape did not allow species identification. By carefully selecting genetic markers, PCR amplification and sequencing of ancient DNA (aDNA) isolates resulted in identification of: the human whipworm, Trichuris trichiura, using SSUrRNA sequence homology; Ascaris sp. with 100% homology to cox1 haplotype 07; and Fasciola hepatica using ITS1 sequence homology. The identification of T. trichiura eggs indicates that human fecal material is present and, hence, that the Ascaris sp. haplotype 07 was most likely a human variant in Viking-age Denmark. The location of the F. hepatica finding suggests that sheep or cattle are the most likely hosts. Further, we sequenced the Ascaris sp. 18S rRNA gene in recent isolates from humans and pigs of global distribution and show that this is not a suited marker for species-specific identification. Finally, we discuss ancient parasitism in Denmark and the implementation of aDNA analysis methods in paleoparasitological studies. We argue that when employing species-specific identification, soil samples offer excellent opportunities for studies of human parasite infections and of human and animal interactions of the past.