Comparative Effect of Ascaridia Galli Infection on the Two Laying Hen Lines on the Liver Function and Immune Response.

Chicken production is quickly rising due to the low associated costs and the capability of poultry to convert nutrients into biological protein along with chicken meat accounting for 30% of all animal protein eaten by humans. Despite advances in poultry production, parasitic illnesses in laying hens remain a problem. Farm birds reared in semi-intensive and free-range systems are more prone to parasite infections due to the absorption of polluted water and food from scavenging behaviors and waste droppings. In this study, the effects of Ascaridia galli infection on the immune response and liver function of two laying hen lines are compared, and their infection resistance is determined. In total, 50 laying hens at eight weeks of age were used (25 Lohmann brown-classic and 25 Lohmann lsl-lite), and each line was divided into two groups: an infected group (n=15), which was orally infected with a single dose of 500 A. galli embryonated eggs, and a control group (n=10), which was given normal saline. After four and eight weeks, blood was collected from the wing vein to assess the serum's AST, ALT, total protein, and IgY levels. The results demonstrated that the infected Lohmann brown-classic and Lohmann lsl-lite chickens presented significantly increased (P≤0.05) AST, ALT, and IgY, compared to the respective control group. Moreover, Lohmann brown-classic hens presented a significantly increased (P≤0.05) IgY concentration four weeks after infection, compared to Lohmann lsl-lite hens. From our results, it can be concluded that genetic variation plays a crucial role in the immune response against A. galli, where the Lohmann brown-classic line was found to be more resistant, compared to the Lohmann lsl-lite line.

Ascaridia galli - An old problem that requires new solutions.

Reports of Ascaridia galli in laying hens in Europe have increased since the ban on conventional battery cages in 2012. As this parasite is transmitted directly via the faecal-oral route by parasite eggs containing a larva, it is reasonable to assume that the escalating problem is related to the increased exposure now occurring in modern welfare-friendly cage-free housing systems. On many farms, A. galli reappears in subsequent flocks, even though the birds have no access to the outdoors, biosecurity is high and empty houses are cleaned and disinfected during downtime. Since the egg production cycle lasts only ≈80 weeks and recombinant antigen production for helminth vaccines has not yet been solved, the development of a vaccine seems to be an unrealistic option. Therefore, disrupting the life cycle of the parasite by other means, including the strategic use of dewormers, appears to be the key to controlling infection. Of concern is that only one class of anthelmintics is licenced for poultry in Europe and that are usually administered indiscriminately through the birds' drinking water and often too late when the parasite is already established. If current calendar-based parasite control strategies are not changed, there is a risk that resistance to anthelmintics may develop, as has already been demonstrated with nematodes in livestock. We insist that treatments can be more effective and the risk of developing drug resistance can be mitigated if we invest in a better understanding of A. galli responses to more prudent and judicious use of anthelmintics. This review identifies knowledge gaps and highlights aspects of sustainable parasite control that require further research to support commercial egg producers.

Viability and development of Ascaridia galli eggs recovered in artificial media followed by storage under different conditions.

Eggs oviposited by Ascaridia galli females in artificial media are commonly used as a source of infective material. We investigated the rate of egg production by cultured mature females (n = 223), and changes in egg viability under different storage and incubation conditions. Eggs recovered after 1, 2 or 3 days of culture were subjected to either (1) storage in water at 4°C (1, 4 or 8 weeks) followed by incubation in 0.1 N H2SO4 at 26°C (2, 4 or 6 weeks); or (2) prolonged storage at 4°C (up to 14 weeks). Egg development and viability was assessed by morphology coupled with a viability dye exclusion test of hatched larvae. Of the 6,044 eggs recovered per mature female 49.2, 38.5 and 12.3% were recovered on days 1, 2 and 3 of worm incubation respectively with similar initial viability (≥99%) between days. Eggs recovered on different days had only minor differences in viability after storage. The prolonged storage period at 4°C significantly affected both viability and embryonation ability resulting in decline in viability of 5.7-6.2% per week. A smaller but significant decline in egg (2.0%) and hatched larval (1.4%) viability per week of incubation at 26°C was also observed. We conclude that storage and incubation conditions, not the day of egg recovery, are the main factors affecting A. galli egg viability. Our findings indicate that under aerobic conditions storage at 26°C may be preferable to 4°C whereas other studies indicate that under anaerobic conditions storage at 4°C is preferable.

Effect of a targeted treatment strategy against Ascaridia galli on egg production, egg quality and bird health in a laying hen farm.

Worm control is an important aspect of the successful management of the egg production industry. Of particular concern is Ascaridia galli, which at high parasite loads affect health and production in layers. Application of a targeted treatment strategy (TT) to control A. galli has shown promise. The purpose of this study was to explore the effect of such a strategy on welfare indicators and production performance of layers. Six flocks (F1-6) on a commercial farm were allocated to three treatment groups. Flocks F1 and F4 were treated (TT) with fenbendazole at 22, 27 and 36 weeks post-placement (WPP). Flocks F2 and F5 were treated at 27 WPP (conventional treatment, CT) and hens in flocks F3 and F6 served as untreated (UT) control groups. At 19, 35 and 45 WPP twenty-five hens plus thirty eggs per flock were randomly selected. Hens were weighed and their plumage conditions (PC) were assessed. The eggs were subjected to various external and interior quality analyses. Production data such as number of eggs/hen/week, egg mass and feed conversion ratio (FCR) were calculated from raw data obtained from all flocks on a weekly basis. The number of eggs/hen/week, egg mass and FCR were higher (P < 0.05) in the TT flocks and hens had better PC both at 35 and 45 WPP compared with other flocks. No differences in body weight and physical egg quality were observed between groups except for egg shell strength which was higher (P < 0.05) in the CT flocks. These data suggest that better production performance and plumage, which suggests improved health, can be achieved through the application of a TT strategy. The insights gained from this research should help to justify the extra cost and labor associated with the TT strategy.

The impacts of Ascaridia galli on performance, health, and immune responses of laying hens: new insights into an old problem.

Gastrointestinal nematodes are re-emerging in countries where the popularity of free-range poultry production systems is increasing. Amongst all gastrointestinal nematodes, Ascaridia galli is of significant concern due to the parasite's direct life cycle and ability to survive extreme environmental conditions. In laying hens, A. galli parasites have been associated with reduced health, welfare, immunity, and egg production. Direct losses are caused by obstruction and damage of the intestinal tract in hens when high worm burdens are present. These result in reduction in egg production and body weight of infected laying hens, consequently leading to significant economic losses for farmers. Furthermore, heavy infections with A. galli may lead to increased mortality within the flock. Indirect losses are due to suppression of immune system function which can increase susceptibility to secondary infections. Infection with A. galli can also alter nutrient utilization and absorption. Levels of anti- A. galli serum and egg yolk antibodies increase following A. galli infection. Elevated antibodies can be used as an indicator of current or previous infections and therefore can be used as a diagnostic tool. The impact of A. galli on hen health and welfare manifests through the depletion of liver lipid reserves and increased use of energy reserves to mount immune responses against the parasite. This review highlights the variable effects of A. galli infection on the performance, health, egg quality, and emphasizes especially on immune responses of free-range laying hens as well as it evaluates various potential detection methods and preventive and control measures of this parasitic disease.

Detection of Ascaridia galli infection in free-range laying hens.

Reliable methods for detection of A. galli infection using excreta egg count (EEC) and ELISA assays to determine A. galli specific IgY levels in serum and yolk samples were compared from hens infected naturally and artificially. Artificially infected hens were used to generate samples for analysis of preferred detection methods and to generate contaminated ranges for use in the naturally acquired infection study in which Lohmann Brown hens (n = 200) at 16 weeks of age were randomly assigned to four treatments with five replicate pens. Hens of negative control (NC) ranged on a decontaminated area, hens of low infection, medium infection and positive control (PC) ranged on the areas previously contaminated by hens artificially infected with 250, 1000 and 2500 A. galli eggs/hen, respectively. Additionally, hens of PC were orally infected with 1000 A. galli eggs/hen. Anti A. galli antibody levels in hen serum (SIgY) and yolk (YIgY) were measured before range access, and 2, 7 and 12 weeks after access to the contaminated ranges. In a natural infection study, eggs were detected in the excreta of all hens 4 weeks after range access, with the exception of NC in which no eggs were detected. EEC increased to reach maximum value (2204 ±â€¯307 eggs/g) after 11 weeks of range access and then declined at 12 weeks (905 ±â€¯307eggs/g) (p < 0.01). While SIgY OD values were not different in hens between any groups before range access, after 2 weeks, both SIgY and YIgY gradually increased in hens of PC (1.17 ±â€¯0.03 and 0.88 ±â€¯0.04) and medium infection (1.07 ±â€¯0.03 and 0.96 ±â€¯0.04) compared to low infection (0.38 ±â€¯0.03 and 0.29 ±â€¯0.04) (p < 0.01) and NC. After 12 weeks, SIgY were similar in hens of PC, medium and low groups whereas YIgY was higher in hens of low infection group (p < 0.01). Sensitivity of the serum and egg yolk antibody levels assay to detect A. galli infection was 100% and 96%, respectively, whereas the pooled EEC method yielded a sensitivity of 93%. The results of this study suggest that hens naturally infected with A. galli produce both SIgY and YIgY at different levels depending on the infection intensity and duration of exposure which allows the diagnosis of prior infection or early diagnosis of current infection. Use of the practical and non-invasive method of yolk sample analysis for detecting IgY can be just as informative as using serum samples to detect A. galli infection.

Time- and dose-dependent development of humoral immune responses to Ascaridia galli in experimentally and naturally infected chickens.

Factors affecting the development of Ascaridia galli-specific humoral responses and their protective roles are largely unknown. We investigated the effects of time and infection dose on A. galli-specific IgY antibody levels following experimental infection. The acquisition and development of new infections and reinfections were also monitored by using tracer birds. Relationships between the retrospective IgY and the final worm burden of the birds were investigated to determine whether humoral immune responses generated during infection provide protection to the host animal. Young chickens were infected (+) with either 100 or 1000 embryonated eggs of A. galli (100+: n = 45; 1000+: n = 45) or kept as uninfected controls (CON: n = 10). Uninfected birds were also added to each infection group as tracer (T) birds (T100+; n = 5 and T1000+; n = 5). Faecal egg counts and IgY antibody concentrations in plasma and egg yolk were determined at selected intervals. Final worm burdens were quantified at 28 weeks post infection (wpi). The plasma antibody (PAB) and egg yolk antibody (EAB) levels of experimentally infected birds were compared to those of control and tracer birds throughout the study period, and PAB levels were found to depend initially on the infection dose but thereafter mainly on reinfections. Starting at wpi 2, 1000+ had consistently higher PAB levels than CON did (P < 0.05). With exceptions at wpi 0, 2 and 12, PAB levels were also higher (P < 0.05) or tended to be higher (P < 0.10) in 100+ than in CON. An earlier and higher increase was observed in the PAB levels of T1000+ than in those of T100+, implying that (re-)infection occurrence depended on the infection dose. Although 1000+ showed higher (P < 0.05) EAB levels than CON did at both wpi 14 and 18, EAB levels were higher in 100+ than in CON only at wpi 28 (P < 0.05). The total worm burdens in the initial experimentally infected birds were similar (P = 0.257); they were also highly comparable between experimentally and naturally infected birds, indicating that final worm burden was mainly determined by the naturally occurring infections resulting from continuous exposure. When all available information on the retrospective plasma and egg yolk IgY levels was collectively evaluated to estimate the larval or total worm burdens of the experimentally infected birds, both PAB and EAB levels at particular wpi were significantly associated with worm burden, especially with larval count. In conclusion, our data support the hypothesis that the number of larvae, rather than the number of mature worms, affects the antibody levels in both plasma and egg yolk. Despite the increased levels of A. galli-specific antibodies in plasma and egg yolk throughout the study period, only a weak indication was found that antibodies might be directly associated with protection.

Effect of the nematophagous fungus Pochonia chlamydosporia on soil content of ascarid eggs and infection levels in exposed hens.

BACKGROUND: The nematophagous fungus Pochonia chlamydosporia can degrade ascarid (e.g. Ascaridia galli) eggs in agar and soil in vitro. However, it has not been investigated how this translates to reduced infection levels in naturally exposed chickens. We thus tested the infectivity of soil artificially contaminated with A. galli (and a few Heterakis gallinarum) eggs and treated with P. chlamydosporia. Sterilised and non-sterilised soils were used to examine any influence of natural soil biota. METHODS: Unembryonated eggs were mixed with sterilised (S)/non-sterilised (N) soil, either treated with the fungus (F) or left as untreated controls (C) and incubated (22 °C, 35 days) to allow eggs to embryonate and fungus to grow. Egg number in soil was estimated on days 0 and 35 post-incubation. Hens were exposed to the soil (SC/SF/NC/NF) four times over 12 days by mixing soil into the feed. On day 42 post-first-exposure (p.f.e.), the hens were euthanized and parasites were recovered. Serum A. galli IgY level and ascarid eggs per gram of faeces (EPG) were examined on days -1 and 36 (IgY) or 40 p.f.e. (EPG). RESULTS: Egg recovery in SF soil was substantially lower than in SC soil, but recovery was not significantly different between NF and NC soils. SF hens had a mean worm count of 76 whereas the other groups had means of 355-453. Early mature/mature A. galli were recovered from SF hens whereas hens in the other groups harboured mainly immature A. galli. Heterakis gallinarum counts were low overall, especially in SF. The SF post-exposure IgY response was significantly lower while EPG was significantly higher compared to the other groups. CONCLUSIONS: Pochonia chlamydosporia was very effective in reducing ascarid egg numbers in sterilised soil and thus worm burdens in the exposed hens. However, reduced exposure of hens shifted A. galli populations toward a higher proportion of mature worms and resulted in a higher faecal egg excretion within the study period. This highlights a fundamental problem in ascarid control: if not all eggs in the farm environment are inactivated, the resulting low level infections may result in higher contamination levels with associated negative long-term consequences.

Performance, egg quality, and liver lipid reserves of free-range laying hens naturally infected with Ascaridia galli.

A study was conducted to determine the performance, egg quality, and liver lipid reserves of laying hens exposed to ranges contaminated with Ascaridia galli. Sixteen-week-old Lohmann Brown laying hens (n = 200) were divided into 4 treatments with 5 replicates containing 10 hens per pen. Hens of treatment 1 [negative control (NC)] ranged on a decontaminated area, and hens of treatments 2 (low infection) and 3 (medium infection) ranged on areas previously contaminated by hens artificially infected with 250 and 1,000 embryonated A. galli eggs, respectively. The hens of treatment 4 [positive control (PC)] ranged on areas previously contaminated by hens artificially infected with 2,500 embryonated A. galli eggs, and in addition these hens were orally inoculated with 1,000 embryonated eggs. Results indicated that hens of the medium infection group had a higher number of intestinal A. galli worms and A. galli eggs in the coprodeum excreta (43.9 ± 4.0 and 3,437 ± 459 eggs/g) compared to hens of the low infection group (23.8 ± 4.0 and 1,820 ± 450 eggs/g) (P < 0.01) and similar worm counts to PC hens (34.4 ± 4.0 and 2,918 ± 474) (P > 0.05). Egg production, egg mass, feed intake, and feed conversion ratio (FCR) were not affected by A. galli infection (P > 0.05). Egg quality parameters (egg weight, shell reflectivity, shell weight, shell thickness, shell percentage, shell breaking strength, deformation, albumen height, Haugh unit, and yolk score) were not affected by A. galli infection (P > 0.05). Highly infected hens had lower liver lipid content (2.72 ± 0.51 g) compared to uninfected hens (4.46 ± 0.58 g, P < 0.01). The results indicate that exposure to ranges contaminated with A. galli resulted in infection of the ranging hens, but this did not affect egg production or egg quality. Infection with A. galli lowered the liver lipid reserves of the host significantly, suggesting infected hens use more energy reserves for maintenance and production.

Effect of an artificial Ascaridia galli infection on egg production, immune response, and liver lipid reserve of free-range laying hens.

This study was conducted to determine the effect of Ascaridia galli infection on free-range laying hens. Lohmann Brown laying hens (n = 200) at 17 wk of age were allocated to 4 treatment groups (n = 50 per group), each with 5 replicate pens of 10 hens. Hens in 3 treatment groups were orally inoculated with different doses of embryonated A. galli eggs: low (250 eggs), medium (1,000 eggs), and high (2,500 eggs) levels, whereas hens of the control group were not infected. Infection levels were monitored using excreta egg counts and mature A. galli worm counts in the intestine. Anti A. galli antibody titers (IgY) in the serum were measured prior to infection, and at 6, 11, 15, and 20 wk post infection (PI) and in egg yolk at 11 and 20 wk PI. Parameters evaluated included feed intake, egg production, egg weight, egg mass, FCR, liver weight, liver fat, and intra epithelial immune cell infiltration. The results showed no difference in feed intake, body weight, or FCR among any treatment groups (P > 0.05). Egg production was lower in the low infection group compared to other groups at 20 wk of age (P < 0.01). Serum IgY was higher in the infected groups' hens at 20 wk PI compared to control group hens (P < 0.01). Yolk IgY increased significantly over time and was higher in infected hens compared to hens of the control group at 11 and 20 wk PI (P < 0.001). No differences were observed in liver lipid content or intraepithelial lymphocytes infiltration among treatment groups. Ascaridia galli eggs in the coprodeum content and adult A. galli worm count were higher in infected hens compared to hens of the control group (P < 0.01). In conclusion, the effects of artificial infection with A. galli on the parameters investigated were minor, and egg yolk antibody may be a more reliable indicator of A. galli infection than serum antibody or excreta egg count.

Survival and development of chicken ascarid eggs in temperate pastures.

Eggs of chicken ascarids (Ascaridia galli and Heterakis spp.) are believed to be hardy and survive for long periods. However, this has not been evaluated quantitatively and our study therefore aimed to determine development and recovery of chicken ascarid eggs after burying in pasture soil. Unembryonated eggs were mixed with soil, placed in sealed nylon bags and buried at 7 cm depth in pasture plots April (spring, n = 72) and December 2014 (winter, n = 72). Eight randomly selected bags per season were used to estimate pre-burial egg recovery [0 week post-burial (wpb)]. Eight random bags were removed at 5, 12, 23, 38, 52, 71 wpb per season and additionally at 104 wpb for spring burial. The content of each bag was analysed for numbers and development stages of eggs. Eggs buried in spring were fully embryonated within 12 wpb. In contrast, eggs buried in winter were developing between 23 and 38 wpb, so that all viable eggs seemed to be fully developed by 38 wpb. About 90% eggs disappeared within 23 wpb (spring) and 38 wpb (winter). Small proportions (2-3%) of seemingly viable and infective eggs were still recovered up to 2 years after deposition. In conclusion, most eggs buried in temperate pasture soil seem to experience a heavy mortality within a few months after the deposition, especially during warm periods. However, a small proportion of eggs may survive and remain infective for at least 2 years.

Comparison between anthelmintic treatment strategies against Ascaridia galli in commercial laying hens.

The efficacy of a sustainable deworming strategy based on targeted treatments (TT) against Ascaridia galli was investigated for the first time in laying hen flocks on a Swedish commercial farm. Three experimental protocols with different levels of treatment, e.g. targeted treatment (TT), conventional treatment (CT) and untreated (UT), were tested in randomly allocated flocks of two different bird hybrids. Every second week faecal egg counts (FECs) were determined from pooled faecal materials collected on trays (20×27cm) placed for a maximum of 12h on the litter belts. In the TT, anthelmintic administration (fenbendazole, 1mg/kg body weight for 5days) started at 22 weeks post placement (wpp) and was repeated twice when the pooled FECs surpassed the assigned threshold of 200 egg per gram faeces (EPG). The CT flocks were treated once at 27wpp using the same anthelmintic. Hens in the UT were not dewormed and served as controls. Additionally, FECs on cloacal contents, worm fecundity and worm burdens were determined at 19, 35 and 45wpp. None of the flocks became infected until after 16wpp. The cumulative pooled FECs at the end of the study were significantly (p<0.01) lower in the TT compared to both CT and UT. Although repeated treatment in the TT protocol did not affect the fecundity, a worm density-dependent increase in fecundity was observed. Cloacal FECs and the number of adult A. galli in TT at 35 and 45wpp were significantly lower compared to other flocks. This study provides evidence that the TT strategy is better in terms of lower worm burden and decreased cumulative environmental parasite egg numbers compared to CT strategy. The TT strategy should be considered as an alternative to the CT strategy with regard to A. galli control in commercial laying hens.

The efficacy of flubendazole against different developmental stages of the poultry roundworm Ascaridia galli in laying hens.

Infection with the poultry roundworm Ascaridia galli has increased in European countries due to the ban on battery cages. This study was conducted in two commercial laying hen flocks (F1 & F2) on different farms in central Sweden. The aims were to (1) investigate the efficacy of flubendazole (FLBZ, 1.43 mg/kg administered in drinking water for 7 days) against adult and larval stages including histotrophic larvae of A. galli, and (2) determine how long it took before the flocks were reinfected after deworming. Accordingly, 180 randomly selected hens were sacrificed before drug administration (bd), on day 3 and 7 during drug administration (dd), and on a weekly basis for up to five weeks post drug administration (pd). Intestinal contents and cloacal materials of each hen plus pooled faecal samples from manure belts were investigated to assess the worm burden and the parasite egg per gram faeces (epg). Additionally, drinking water, and serum and gastrointestinal digesta content samples obtained from ten treated animals were analyzed by HPLC to measure FLBZ and its reduced (R-FLBZ) and hydrolyzed (H-FLBZ) metabolites. No parasite eggs were observed in cloacal samples on day 21 and 28 pd on F1 and on day 21 pd on F2. The epg in manure decreased by 65% and 88% on day 3 dd and by 99% and 97% on day 35 pd on F1 and F2 respectively. Mean FLBZ concentrations quantified in duodenal contents ranged between 0.50 and 0.79 µg/g. Although, no histotrophic larvae were found dd, they reappeared one week pd (7 ± 7 F1, 0.5 ± 0.5 F2). Adult worms were found in both flocks before drug administration (44 ± 20 F1, 35 ± 25 F2), on day 3 dd (4 ± 3 F1, 2 ± 2 F2), and then not until day 35 (0.2 ± 0.6) on F1 and day 28 (0.4 ± 0.9) pd on F2. Thus, the only period in which no A. galli were found was on day 7 dd. Although FLBZ was highly efficient our results indicate that the birds were reinfected already within one week pd.

Effects of alcoholic extract of Curcuma longa on Ascaridia infestation affecting chicken.

Ascaridia galli, the common intestinal nematode, remains a major cause of economic loss in the poultry industry in developing countries. Treatments using chemicals are not only expensive but also affect host health. Plant extracts as better alternative is gaining significance. Here, we have studied the effects of alcoholic extract of turmeric, Curcuma longa L. (Zingiberaceae) roots, against A. galli infection in chicken. Different concentrations of C. longa root extract were tested in vitro on 5 groups of adults A. galli worms and in vivo on 6 groups of chicks. The results showed that the turmeric root extract @ 60 mg mL(-1) in vitro significantly (P < 0.001) proved paralytic and fatal against worms (16.80 ± 1.28 h). In vivo, chicken groups (G2-G6) were infected with an average of 300 ± 12 embryonated eggs of A. galli. The G2 was not given any treatment while G3 was treated with piperazine (@ 200 mg kg(-1) body wt.); and Groups 4, 5 and 6 were given turmeric @ 200, 400 and 600 mg kg(-1) body wt., respectively. The mean number of worms extracted at the end of the trial in G2 (untreated) was 18.10 ± 2.42, while the G3 treated with piperazine had no worms. Groups 4 and 5 did not show any significant difference compared to G2. However, G6 that had 3.20 ± 1.33 worms was statistically significant. Higher concentrations of turmeric given to infected chickens significantly reduced the length and weight of worms. The study showed that the worm infestation damaged the intestinal villi, and.treatment with high concentration of C. longa had healing effects and restored the integrity of intestinal mucosa. The results have demonstrated the ameliorating effect of C. longa turmeric on A. galli infested chickens.

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.

Environmental tolerance of free-living stages of the poultry roundworm Ascaridia galli.

The poultry roundworm Ascaridia galli is re-emerging in laying hens in many European countries due to the increase in non-caged housing. A series of in vitro experiments was carried out to study the in ovo larval development (embryonation) under different environmental conditions. Between 83% and 96% of the eggs developed to L3 within 7-21 days of incubation in water between 20 and 30°C. Twenty-six percent completed development at 33°C and 4% at 35°C after 31 days. At 15°C parasite egg development was low with 8% L3 after 56 days. In another trial larval development occurred, when parasite eggs were exposed to freeze-thaw cycle (30' to 12h) followed by incubation for 2 weeks at 25°C. Alkaline and acidic conditions in the range of pH 2.5-12.5 had no adverse effect on development. Oxygen and relative humidity above 70% were necessary for development to occur. Thus, some A. galli eggs may complete development at conditions prevailing in poultry barns in temperate climate zones throughout the year. Although exposure to a 1% or 2% dilution of the broad-spectrum disinfectant chlorocresol for 4h or longer was ovicidal, further work is required to improve the method of application in the field.

Survival of Ascaris suum and Ascaridia galli eggs in liquid manure at different ammonia concentrations and temperatures.

Eggs of Ascaris suum from pigs are highly resistant and commonly used as a conservative indicator of pathogen inactivation during slurry storage. Eggs of Ascaridia galli, the poultry ascarid, are also known to be highly resistant but the suitability as an indicator of pathogen inactivation has never been tested. Pig slurry has to be stored for several months to inactivate pathogens but chemical treatment of slurry may reduce this time. The suitability of A. galli as an indicator of slurry sanitation was tested by comparing the survival of eggs of A. suum and A. galli in pig slurry. In addition, the effect of urea treatment on inactivation of ascarid eggs in relation to storage time was also tested. Nylon bags with 10,000 eggs of either species were placed in 200 ml plastic bottles containing either urea-treated (2%) or untreated pig slurry for up to 120 days at 20°C, 6 days at 30°C, 36h at 40°C or 2h at 50°C. At all the temperatures in both slurry types, A. galli eggs were inactivated at a significantly faster rate (P<0.05) compared to A. suum eggs. For each 10°C raise in temperature from 20°C, T50 (time needed to inactivate 50% of eggs) for both types of eggs was reduced markedly. At all temperatures, viability of eggs of both species was significantly higher (P<0.05) in untreated slurry compared to urea-treated slurry except A. galli eggs at 20°C where no significant difference was detected. In untreated slurry, the levels of pH (6.33-9.08) and ammonia (0.01-1.74 mM) were lower (P<0.0001) compared to that of urea-treated slurry (pH: 8.33-9.28 and ammonia 1-13 mM). The study demonstrated that A. galli eggs are more sensitive to unfavourable conditions compared to A. suum eggs. The use of A. galli eggs as hygiene indicator may thus be suitable to assess inactivation of pathogens that are more sensitive than A. galli eggs. Addition of urea may markedly reduce the storage time of slurry needed to inactivate A. suum and A. galli eggs.

Ascaridia galli infection affects pullets differently when feed is contaminated with the Fusarium toxin deoxynivalenol (DON).

The Fusarium mycotoxin deoxynivalenol (DON) is a common contaminant of cereal grains used as animal feed. DON is known for its cytotoxic and anti-proliferative properties and might adversely affect the health of poultry. The prevalence of the intestinal parasitizing roundworm Ascaridia galli is higher in outdoor housing systems and has been associated with maldigestion and malabsorption. It was hypothesized that ingested DON might not only affect the pullet itself but could also act on the nematode parasitizing in the ingesta. To examine these interactions between A. galli infection and DON contamination of feed 4 groups of 9 pullets in each were tested; non-infected groups were fed either an uncontaminated control (CON-) or a Fusarium toxin contaminated and mainly DON-containing diet (FUS-), and the corresponding A. galli inoculated groups were fed accordingly (CON+, FUS+). A. galli infection significantly reduced the jejunal villi height and increased the thickness of the tunica muscularis with the effect being more pronounced when the DON-containing diet was fed (Group FUS+). Only in this group significantly increased weights of jejunal and ileal tissues and of livers were noticed. Moreover, DON was detected in plasma of the pullets at higher frequencies when they were infected suggesting a facilitated absorption of DON. Group FUS+ was characterized by a significantly higher excretion of A. galli eggs and a concomitant lower proportion of pullets with detectable antibodies against a somatic antigen of A. galli while worm burden and worm characteristics were not affected by diet. Other effects of feeding the FUS diet to the infected pullets included an increased mass per length of male worms. In conclusion, infection of pullets with A. galli might increase the susceptibility towards DON as indicated by an increased DON absorption rate and a compromised antibody formation. The effects of DON on fecundity and worm morphology require further examination.

The complete mitochondrial genomes of three parasitic nematodes of birds: a unique gene order and insights into nematode phylogeny.

BACKGROUND: Analyses of mitochondrial (mt) genome sequences in recent years challenge the current working hypothesis of Nematoda phylogeny proposed from morphology, ecology and nuclear small subunit rRNA gene sequences, and raise the need to sequence additional mt genomes for a broad range of nematode lineages. RESULTS: We sequenced the complete mt genomes of three Ascaridia species (family Ascaridiidae) that infest chickens, pigeons and parrots, respectively. These three Ascaridia species have an identical arrangement of mt genes to each other but differ substantially from other nematodes. Phylogenetic analyses of the mt genome sequences of the Ascaridia species, together with 62 other nematode species, support the monophylies of seven high-level taxa of the phylum Nematoda: 1) the subclass Dorylaimia; 2) the orders Rhabditida, Trichinellida and Mermithida; 3) the suborder Rhabditina; and 4) the infraorders Spiruromorpha and Oxyuridomorpha. Analyses of mt genome sequences, however, reject the monophylies of the suborders Spirurina and Tylenchina, and the infraorders Rhabditomorpha, Panagrolaimomorpha and Tylenchomorpha. Monophyly of the infraorder Ascaridomorpha varies depending on the methods of phylogenetic analysis. The Ascaridomorpha was more closely related to the infraorders Rhabditomorpha and Diplogasteromorpha (suborder Rhabditina) than they were to the other two infraorders of the Spirurina: Oxyuridorpha and Spiruromorpha. The closer relationship among Ascaridomorpha, Rhabditomorpha and Diplogasteromorpha was also supported by a shared common pattern of mitochondrial gene arrangement. CONCLUSIONS: Analyses of mitochondrial genome sequences and gene arrangement has provided novel insights into the phylogenetic relationships among several major lineages of nematodes. Many lineages of nematodes, however, are underrepresented or not represented in these analyses. Expanding taxon sampling is necessary for future phylogenetic studies of nematodes with mt genome sequences.

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.