In vitro hatching of Trichuris suis eggs.

Eggs of the pig whipworm, Trichuris suis ova (TSO), are currently tested in human clinical trials for their potential immunomodulatory capacity. The biological potency of TSO (egg viability and infectivity) is traditionally assessed in Göttingen minipigs as the establishment of intestinal larvae after inoculation with a known number of eggs. To minimize testing in animal models, development of an in vitro egg hatching assay is proposed as a reliable, cost-effective, and a faster alternative to test the egg viability. The present study aimed to investigate the influence of different chemical, physical, and biological factors on egg hatching. Thus, in a series of experiments and in different combinations, the eggs were stimulated with glass beads, artificial gastric juice, bile salt and trypsin solution, fermentation gut medium, or stimulated with mucosal scrapings from the ileum and the large intestine of the infected and uninfected Göttingen minipig. Mechanical stimulation with glass beads presented a simple and reproducible method for egg hatching. However, incubation of eggs with mucosal scrapings from the ileum, caecum, and colon for 24 h at 38 °C significantly increased hatching.

The jejunal cellular responses in chickens infected with a single dose of Ascaridia galli eggs.

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.

Viability of Ascaris suum eggs in stored raw and separated liquid slurry.

Separation of pig slurry into solid and liquid fractions is gaining importance as a way to manage increasing volumes of slurry. In contrast to solid manure and slurry, little is known about pathogen survival in separated liquid slurry. The viability of Ascaris suum eggs, a conservative indicator of fecal pollution, and its association with ammonia was investigated in separated liquid slurry in comparison with raw slurry. For this purpose nylon bags with 6000 eggs each were placed in 1 litre bottles containing one of the two fractions for 308 days at 5 °C or 25 °C. Initial analysis of helminth eggs in the separated liquid slurry revealed 47 Ascaris eggs per gramme. At 25 °C, egg viability declined to zero with a similar trend in both raw slurry and the separated liquid slurry by day 308, a time when at 5 °C 88% and 42% of the eggs were still viable in separated liquid slurry and raw slurry, respectively. The poorer survival at 25 °C was correlated with high ammonia contents in the range of 7.9-22.4 mM in raw slurry and 7.3-23.2 mM in liquid slurry compared to 3.2-9.5 mM in raw slurry and 2.6-9.5 mM in liquid slurry stored at 5 °C. The study demonstrates that at 5 °C, A. suum eggs have a higher viability in separated liquid slurry as compared to raw slurry. The hygiene aspect of this needs to be further investigated when separated liquid slurry is used to fertilize pastures or crops.

Population dynamics of Ascaridia galli following single infection in young chickens.

The population dynamics of Ascaridia galli was studied in 70 ISA Brown layer pullets, 42 of them were each experimentally infected with 500 embryonated A. galli eggs and 28 chickens were kept as uninfected controls. Six chickens from the infected group and 4 from the control group were necropsied at 3, 7, 10, 14, 21, 28 and 42 days post-infection (d.p.i.). The mean worm recovery varied from 11-20% of the infection dose with the highest recovery at 3 d.p.i. and the lowest at 21 and 42 d.p.i. (P < 0·05). More larvae were recovered from the intestinal wall than from the content (P < 0·0001) and intestinal content larvae were longer than those from the wall (mean length 1·6 and 1 mm, respectively, P < 0·0001). Although larvae were growing over time, a population of small-sized larvae (length < 1 mm) was recovered at all d.p.i. During the first week of infection most of the larvae were located in the anterior half of the jejunoileum but they moved posteriorly with the age of infection. Thus, a subpopulation of larvae mainly in the lumen grew with time while another subpopulation remained small and associated with the mucosa. During the infection both subpopulations moved to a more posterior localization in the gastrointestinal (GI) tract.

Ascaridia galli in chickens: intestinal localization and comparison of methods to isolate the larvae within the first week of infection.

This study was conducted to observe the localization and to compare methods for isolation of minute Ascaridia galli larvae in chicken intestine. Firstly, six 7-week-old layer pullets were orally infected with 2,000 embryonated A. galli eggs and necropsied either at 3, 5 or 7 days post infection (dpi). More than 95 % of the recovered larvae were obtained from the anterior half of the jejunoileum, suggesting this part as the initial predilection site for A. galli larvae. Secondly, the intestinal wall of one layer pullet infected with 20,000 A. galli eggs 3 days earlier was digested in pepsin-HCl for 90 min. The initial 10 min of digestion released 51 % of the totally recovered larvae and the last 30 min of continuous digestion yielded only 5 %. This indicates that the majority of larvae were located superficially in the intestinal mucosa. Thirdly, 48 7-week-old layer pullets were infected with 500 A. galli eggs and necropsied at 3 dpi to compare three different larval isolation methods from the intestinal wall, viz., EDTA incubation, agar-gel incubation and pepsin-HCl digestion, resulting in mean percentages of the recovered larvae: 14.4, 18.2 and 20.0 %, respectively (P = 0.15). As conclusion, we recommended Pepsin-HCl digestion as the method of choice for larval recovery from the intestinal wall in future population dynamics study due to high efficiency and quick and simple detection. The agar-gel method was considered to be a prerequisite for molecular and immunological investigations as the larvae were more active and fully intact.

Trichuris suis ova therapy for allergic rhinitis: a randomized, double-blind, placebo-controlled clinical trial.

BACKGROUND: Parasitic helminth infections can protect against allergic airway inflammation in experimental models and have been associated with a reduced risk of atopy and a reduced course of asthma in some observational studies. Although no clinical evidence exists to support the use of helminth therapy for allergic disease, the helminth Trichuris suis has demonstrated efficacy in treatment of inflammatory bowel disease. OBJECTIVE: To determine efficacy of helminth therapy for allergic rhinitis. METHODS: We conducted a double-blind, placebo-controlled, parallel group trial in which 100 subjects age 18 to 65 years with grass pollen-induced allergic rhinitis were randomly assigned to ingest a total of 8 doses with 2500 live T suis ova or placebo with an interval of 21 days. The primary outcome was a change in mean daily total symptom score for runny, itchy, sneezing nose (maximum change, 9.0) or in percentage of well days during the grass pollen season. RESULTS: Treatment with T suis ova (N = 49) compared with placebo (N = 47) caused transient diarrhea peaking at day 41 in 33% of participants (placebo, 2%), and increased eosinophil counts (P < .001) and T suis-specific IgE (P < .05), IgG (P < .001), IgG(4) (P < .003), and IgA (P < .001), whereas there was no significant change in symptom scores (0.0; 95% CI, -0.5 to 0.4; P = .87), well days (3%; 95% CI, -9% to 14%; P = .63), total histamine (P = .44), grass-specific IgE (P = .76), or diameter of wheal reaction on skin prick testing with grass (P = .85) or 9 other allergens. CONCLUSION: Repeated treatment with the helminth T suis induced a substantial clinical and immunologic response as evidence of infection, but had no therapeutic effect on allergic rhinitis.

Prevalence of gastrointestinal nematodes in growing pigs in Kabale District in Uganda.

During the last 30 years, pig production in Uganda and neighbouring counties has increased markedly. Pigs are mainly kept as a source of income for small-scale farmers; however, the pig production is subject to several constraints, one of them being worm infections. A study was carried out in rural communities in Kabale District in the South Western part of Uganda in September and October 2007 in order to estimate the prevalence of gastrointestinal nematode parasites in pigs based on coprological examination. Fifty-six households were randomly selected and visited. Housing system and deworming history were recorded. Faeces was sampled from rectum of one to five pigs (age, 3-12 months) per household. A total of 106 pigs were examined coprologically of which 91% excreted nematode eggs. The following prevalences of nematode eggs were recorded: strongyles (89%), Ascaris suum (40%), Trichuris suis (17%) and spiruroid eggs (48%). On household level, rearing pigs on slatted floors in pens significantly reduced the faecal egg excretion of strongyle eggs with almost 80% (p=0.010) and a significant interaction between floor type and anthelmintic treatment was found for spiruroids (p=0.037). Fifteen T. suis egg positive pigs were selected for post-mortem examination of the gastrointestinal tract. The post-mortem examinations revealed that 93% pigs were infected with Oesophagostomum spp. (worm burden, min-max 10-2,180), 73% with A. suum (1-36), 67% with T. suis (6-58), and 20% with Hyostrongylus rubidus (worms not quantified). In general, nematode infections were widespread and polyparasitism common in pigs in Uganda. However, worm burdens were moderate which may be related to recent deworming or to the practice of rearing pigs on slatted floors in wooden elevated pens.

Survival of Isospora suis oocysts under controlled environmental conditions.

Isospora suis is a coccidian parasite infecting piglets soon after birth. While the gross epidemiology of I. suis is well known, little knowledge exists on the ecology of the oocysts. To study the development and survival of oocysts of I. suis under controlled laboratory conditions, known numbers of oocysts ( approximately 200 in each of 4 replicates) were exposed to all combinations of 4 relative humidities (53-100% RH) and 3 temperatures (20 degrees , 25 degrees , 30 degrees C). Determination of viability was based on morphological and fluorescent properties of the oocyst as well as on the permeability of the oocyst wall characterized by inclusion/exclusion of the fluorescent dye propidium iodide. The viability of the oocysts was studied over time by fluorescence and light microscopy until <5% of the oocysts were considered to be viable. The sporulation rate increased with temperature, however, the infective sporocyst stage was reached within 24h at all temperatures, while RH did not seem to affect sporulation. Results show a rapid reduction in viable oocysts exposed to high temperatures (25 degrees C and 30 degrees C) in combination with low relative humidities (53% RH and 62% RH), at which conditions oocysts died within 24h. Viability was higher when oocysts were exposed to higher relative humidities (75% RH and 100% RH) as well as a lower temperature (20 degrees C). However, even at 75% RH the oocysts died within 24-60 h at 30 degrees C to 20 degrees C, respectively, while the most favourable condition appeared to be 100% RH and 25 degrees C at which condition the percentage of viable oocysts decreased from 100% to 17% in 96 h. The results indicate that it may be possible to reduce the infection pressure of I. suis in modern sow herds by changing the environmental conditions and/or the management within the farrowing pens, and thereby increase animal welfare without relying on the use of routine medication.

A novel technique for identification of Ascaris suum cohorts in pigs.

The objective of the present study was to develop a fast, cheap and reliable technique for identifying different cohorts of the swine parasite, Ascaris suum. A polymerase chain reaction linked restriction fragment length polymorphism (PCR-RFLP) technique on mt-DNA was used to identify unique haplotypes of four gravid A. suum females on agarose gels after eggs were recovered from each of the worms. Each of four pigs was inoculated with 2000 embryonated eggs originating from one of the four identified Ascaris haplotypes, respectively. Ascaris larvae were isolated from the small intestine at day 14 post-infection using an agar technique. Single larvae from each pig were transferred to 96-well PCR plates and a simple DNA extraction using a worm lysis buffer was carried out and followed by the PCR-RFLP analysis. More than 100 larvae from each of the four pigs were analysed and all were found to have the same haplotype as the parental female. We conclude that unique haplotypes of female A. suum and offspring can be identified by means of PCR-RFLP on mt-DNA and suggest that this method can be used in future research on Ascaris population biology using cohorts with distinct mt-DNA profile.

The effect of fermentable carbohydrates on experimental swine dysentery and whip worm infections in pigs.

An experiment was conducted to study the effect of diets with contrasting fermentability in the large intestine on experimental infections with Brachyspira hyodysenteriae, the causative agent of swine dysentery, and the whip worm, Trichuris suis, in pigs. Two diets with organically grown ingredients were composed. Both diets were based on triticale and barley and supplemented with either rape seed cake (Diet 1) or dried chicory root and sweet lupins (Diet 2). The study had a three-factorial design, with eight groups of pigs receiving Diet 1 or Diet 2, +/-B. hyodysenteriae, and +/-T. suis. Pigs fed Diet 2 and challenged with B. hyodysenteriae did not develop swine dysentery and B. hyodysenteriae was not demonstrated in any of the pigs during the study. In contrast, 94% of the B. hyodysenteriae challenged pigs fed Diet 1 showed clinical symptoms of swine dysentery and all the pigs were shedding B. hyodysenteriae in faeces at some points in time during the experiment. The number of T. suis was lower in pigs fed Diet 2 compared to pigs fed Diet 1, but the differences were not significant. Pigs on Diet 1 and challenged with both pathogens showed clinical symptoms of SD for a longer period than pigs inoculated with B. hyodysenteriae only. The study showed that diets supplemented with highly fermentable carbohydrates from dried chicory roots and sweet lupins can protect pigs against developing swine dysentery, but do not have any significant influence on T. suis.

A time course study of immunological responses in Trichuris suis infected pigs demonstrates induction of a local type 2 response associated with worm burden.

In order to investigate immunological changes over time in pigs infected with Trichuris suis, we inoculated 40 pigs with 5000 infective T. suis eggs and left 40 pigs as uninfected controls. Equal numbers of pigs from both groups were sacrificed every other week from 1 to 11 weeks p.i. At necropsy tissue samples were collected from all pigs and their worm burdens were determined. In the proximal colon of T. suis-infected pigs infiltration of eosinophils peaked 5 weeks p.i. and mast cell infiltration developed from 5 to 11 weeks p.i. Histological evaluation of the proximal colon revealed that the presence of T. suis was closely associated with intestinal histopathological changes such as crypt hyperplasia, goblet cell hyperplasia and a general hypertrophy of mucosa. The crypt lengths were positively associated with worm burdens. Real-time PCR analysis of genes related to immune function indicate a local increased transcription of genes coding for CCR3, ARG1, MUC5AC, IL-4, IL-5, IL-13, FcepsilonR1alpha, and IL-13Ralpha2 and decreased expression of genes coding for iNOS, TNF-alpha, IL-10, CD3epsilon, CD80, CD86, IL-4Ralpha, IL-13Ralpha1 and CD40 in the proximal colon of pigs infected with T. suis. This local T-helper cell Type 2-like gene-expression pattern indicates that the Type 2 immune response characteristic of helminth infections in both mouse and humans also develops in pigs infected with T. suis. The results from this study expand our knowledge of the immunomodulatory effect of T. suis, a parasite that has proven effective in treating inflammatory bowel disease, when its eggs are administered regularly to patients.

Trichuris suis population dynamics following a primary experimental infection.

Trichuris suis population kinetics was studied by experimentally infecting 40 pigs with 5000 T. suis infective eggs. Six pigs were sacrificed every 2 weeks from 1 to 9 weeks post-inoculation (p.i.) and the remaining 10 pigs were sacrificed 11 weeks p.i. to estimate worm burdens. An equal number of uninfected control pigs were sacrificed at the same time points for comparison. Egg excretions from each pig were evaluated every week from 5 to 11 weeks p.i. Peripheral blood eosinophilia and basophilia were also evaluated every 2 weeks throughout the experimental period. After an initial almost 100% establishment of T. suis an expulsion phase followed approximately 9 weeks p.i., resulting in an aggregated distribution of the worms in the pig population, as it is characteristic for most helminth infections. By 11 weeks p.i. almost all worms had been expelled. Egg excretion peaked 7 weeks p.i. and a significant non-linear relationship between worm burdens and egg excretion was found. The predilection site for T. suis was the caecum and proximal colon and the relative distribution of worms in the large intestine did not change over time until expulsion. Peaking peripheral blood eosinophil and basophil levels were observed in the T. suis infected pigs 5 and 7 weeks p.i., respectively, reflecting the host activated immune response against the parasite. We here describe the course of a primary T. suis infection in pigs by detailed worm counts, demonstrating an effective expulsion that results in an almost 100% clearance of infection as previously indicated by monitoring faecal egg excretion.

Temperature dependent embryonic development of Trichuris suis eggs in a medicinal raw material.

The therapeutic potential of infective pig whipworm eggs, Trichuris suis ova (TSO), is currently tested in several clinical trials on immune-mediated diseases. This paper studied the embryonic development of TSO in a medicinal raw product, where the parasite eggs were suspended in sulphuric acid (pH1). Unembryonated T. suis egg batches were stored at 5, 10, 15, 20, 25, 30, and 40°C (±1°C) and examined at 2, 4, 8, and 14 weeks. Subsequently, sub-batches from each temperature were allowed to embryonate for additional 14 weeks at 25°C, and selected samples were tested for infectivity in Göttingen minipigs. Both male and female pigs were used to evaluate eventual gender specific infectivity. Storage at 30°C up to 14 weeks and subsequent embryonation for 14 weeks at 25°C did not significantly reduce the overall larval establishment in minipigs, as compared to storage at 5°C and subsequent embryonation at 25°C. As marked impairment of egg development was observed during storage at 40°C, a second set of unembryonated egg batches were incubated at 30, 32, 34, 36, 38, and 40°C (±1°C) for 1-8 weeks. The development of the eggs was repeatedly examined by manual light microscopy, multispectral analysis (OvaSpec), and an egg hatching assay prior to the final testing in minipigs (Trial 1). These methods showed that the development started earlier at higher temperatures, but the long-term storage at higher temperature affected the egg development. The present study further documents tolerance of the TSO to storage at temperature 5-15°C, at which temperature development of larvae is not initiated.

Population structure in Ascaris suum (Nematoda) among domestic swine in Denmark as measured by whole genome DNA fingerprinting.

We here analyze the population structure in the pig roundworm, Ascaris suum, among domestic pigs in Denmark using a whole-genome DNA fingerprinting technique, "amplified fragment length polymorphism" (AFLP) analysis. With these data, we can extract absolute gene frequency variance components and G-statistics for 135 independent nucleotide polymorphisms. The average proportion of total variance partitioned between Jutland and Zealand is less than 3% of the total variance, implying no restriction in gene flow between worms from different regions in Denmark. The average gene frequency difference between two farms widely separated in Jutland represents 5% of the total genetic variance of these two farms combined. Conversely, worms from different hosts within these two farms are more subdivided, with an average of 12% of the total variance in gene frequencies within farms being distributed between hosts. This result implies substantial single generation inbreeding due to founder effects in the establishment of adult worms in single hosts. Absolute variance components extracted from the gene diversities also showed significant differences, with the among-host variance being greater that the between-farm and between-region values. This little geographical variation is discussed in relation to the hierarchic structure of the Danish swine production system. Comparison of our results with other studies on parasitic roundworms, suggests that patterns of host dispersal effectively control patterns of worm gene flow. Furthermore, the potential spread of anthelminth resistance among A. suum may thus be rapid, due to the flow of infected hosts within the domestic swine stocks in Denmark.

Dose-dependent establishment of Trichuris suis larvae in Göttingen minipigs.

Embryonated eggs of the pig whipworm Trichuris suis (TSOee) constitute the active pharmaceutical ingredient (API) in a medicinal product explored in human clinical trials against several immune-mediated diseases. The measurement of TSO biological potency (hatchability and infectivity) is a requirement for the assessment of TSO's pharmacological potency in human clinical trials. The present study aims to validate the dose-dependent establishment of T. suis larvae in Göttingen minipigs and eventual clinical implication of a dose range (1000-10,000 TSO). Four groups of 5 minipigs were inoculated with doses of 1000, 2500, 7500, and 10,000 TSOee, respectively, to evaluate a range of concentrations of TSOee in a minipig infectivity model. Unembryonated eggs (TSOue) were added to keep the total egg number in the inoculum constant at 10,000 eggs. Two groups received 2500 and 7500 TSOee per pig without the addition of TSOue as controls. The intestinal larval establishment at 21 days post inoculation (dpi) demonstrated a clear positive linear dose-response relationship between numbers of inoculated TSOee and recovered larvae. There was a low level of variation in larval counts in all study groups. Thus, the infectivity model in minipigs within the tested dose range offers a reliable, sensitive and accurate assay for testing biological potency of TSO.

Helminth infections in Danish organic swine herds.

In nine organic swine herds, faecal excretion and pasture contamination by parasite eggs/larvae were studied in a period from March to October 1999. It was shown that the organic pigs were infected with Ascaris suum (28% of weaners, 33% of fatteners, 4% of sows), Trichuris suis (4% of weaners, 13% of fatteners, <1% of sows) and Oesophagostomum spp. (5% of weaners, 14% of fatteners, 20% of sows) whereas no infections with Hyostrongylus rubidus, Metastrongylus spp. or Strongyloides ransomi were detected. Moreover, no pigs showed clinical signs of infestations with scabies or lice. In the soil samples, very few Trichuris eggs were found throughout the season, whereas Ascaris eggs were found in 14% of the soil samples from sow pastures and in 35% from slaughter pig pastures, with the first infective eggs being recorded in July and the maximum number in August. Infective Oesophagostomum larvae were found in the grass samples in increasing numbers from May to October. Single herd cases of exceptionally high parasite infection levels are described in relation to herd management procedures.

Phasmid ultrastructure of an ascaridoid nematode Hysterothylacium auctum.

Here, for the first time in an ascaridoid (Hysterothylacium auctum), we present structural features of the phasmids, paired sense organs, positioned in a bilateral manner close to the point of the tail; the features were obtained using scanning and transmission electron microscopy. We found that each phasmid consists of a single ciliated dendritic process situated in a phasmidial canal surrounded by 2 supporting cells, a socket and a sheath cell. The socket cell contains clusters of electron-dense fibrous material in its apical region and covers the phasmidial canal along its whole length. The sheath cell is characterized by a well-developed endoplasmic reticulum. The phasmidial canal is lined with a thin layer of cuticle that becomes incomplete at the base of the ciliated dendritic process. In this region, the dendritic process consists primarily of a high number of microtubule singlets and some peripheral microtubule doublets. The base of the dendritic process, containing numerous striated rootlets, gives off a large number of fingerlike offshoots, villi, invading the surrounding sheath cell. The systematic significance and functional implication of the phasmid in nematodes are also discussed.

Excretion in feces and mucosal persistence of Salmonella ser. Typhimurium in pigs subclinically infected with Oesophagostomum spp.

OBJECTIVE: To determine interactions between Oesophagostomum spp and Salmonella ser. Typhimurium in pigs. ANIMALS: 30 healthy 5- to 6-week-old pigs. PROCEDURE: Pigs were allotted to 3 groups (n = 10 pigs/group) and treated as follows: group A was given Oesophagostomum dentatum and O quadrispinulatum; group B was given O dentatum, O quadrispinulatum, and S Typhimurium; and group C was given S Typhimurium only. Pigs in groups A and B were trickle infected with Oesophagostomum spp 3 times weekly throughout the study. After 19 days, groups B and C were inoculated once with S Typhimurium. One pig from each group was euthanatized on the day of Salmonella exposure and 2 and 4 days after Salmonella exposure. The remaining pigs were euthanatized on days 16 and 17 after Salmonella exposure. RESULTS: Pigs with dual infections of nematodes and bacteria (group B) excreted significantly higher amounts of S Typhimurium in feces, compared with nematode-free pigs (group C). In addition, group-B pigs excreted S Typhimurium on more days than pigs in group C. Salmonella Typhimurium was detected in the cecum and colon in the majority of pigs in group B, whereas S Typhimurium was only detected in the colon in pigs in group C. Immunohistochemical examination detected S Typhimurium in 7 of 9 pigs in group B but only 2 of 9 pigs in group C. CONCLUSIONS AND CLINICAL RELEVANCE: Interactions between intestinal nematodes and bacteria may play an important role in the dynamics of S Typhimurium infections.

Parasite population dynamics in pigs infected with Trichuris suis and Oesophagostomum dentatum.

The aim of the present study was to investigate the population dynamics and potential interactions between Trichuris suis and Oesophagostomum dentatum in experimentally co-infected pigs, by quantification of parasite parameters such as egg excretion, worm recovery and worm location. Forty-eight helminth naïve pigs were allocated into four groups. Group O was inoculated with 20 O. dentatum L3/kg/day and Group T with 10 T. suis eggs/kg/day. Group OT was inoculated with both 20 O. dentatum L3/kg/day and 10 T. suis eggs/kg/day, while Group C was kept as an uninfected control group. All inoculations were trickle infections administered twice weekly and were continued until slaughter. Faecal samples were collected from the rectum of all pigs at day 0, and twice weekly from 2 to 9 weeks post first infection (wpi). Six pigs from each group were necropsied 5 wpi and the remaining 6 pigs from each group were necropsied 10 wpi. The faecal egg counts (FEC) and total worm burdens of O. dentatum were dramatically influenced by the presence of T. suis, with significantly lower mean FECs and worm burdens at 5 and 10 wpi compared to single infected pigs. Furthermore, in the presence of T. suis we found that O. dentatum was located more posteriorly in the gut. The changes in the Trichuris population were less prominent, but faecal egg counts, worm counts 5 wpi (57% recovered vs. 39%) and the proportion of infected animals at 10 wpi were higher in Group OT compared to Group T. The location of T. suis was unaffected by the presence of O. dentatum. These results indicate an antagonistic interaction between T. suis and O. dentatum which is dominated by T. suis.

OvaSpec - A vision-based instrument for assessing concentration and developmental stage of Trichuris suis parasite egg suspensions.

BACKGROUND: OvaSpec is a new, fully automated, vision-based instrument for assessing the quantity (concentration) and quality (embryonation percentage) of Trichuris suis parasite eggs in liquid suspension. The eggs constitute the active pharmaceutical ingredient in a medicinal drug for the treatment of immune-mediated diseases such as Crohn׳s disease, ulcerative colitis, and multiple sclerosis. METHODS: This paper describes the development of an automated microscopy technology, including methodological challenges and design decisions of relevance for the future development of comparable vision-based instruments. Morphological properties are used to distinguish eggs from impurities and two features of the egg contents under brightfield and darkfield illumination are used in a statistical classification to distinguish eggs with undifferentiated contents (non-embryonated eggs) from eggs with fully developed larvae inside (embryonated eggs). RESULTS: For assessment of the instrument׳s performance, six egg suspensions of varying quality were used to generate a dataset of unseen images. Subsequently, annotation of the detected eggs and impurities revealed a high agreement with the manual, image-based assessments for both concentration and embryonation percentage (both error rates <1.0%). Similarly, a strong correlation was demonstrated in a final, blinded comparison with traditional microscopic assessments performed by an experienced laboratory technician. CONCLUSIONS: The present study demonstrates the applicability of computer vision in the production, analysis, and quality control of T. suis eggs used as an active pharmaceutical ingredient for the treatment of autoimmune diseases.