Ecology and viability of eggs of the pig round worm (Ascaris suum) – on-farm and laboratory studies

Ascaris suum is a highly prevalent parasitic helminth in pig production systems worldwide. In most industrialized countries production systems are highly intensive and fully in-doors (low prevalence of parasites) but there is a growing market for pigs produced out-door or under organic pig farming conditions (high prevalence of parasites). In organic pig production there is a number of factors related to housing and management practices that predispose for A. suum infections. These practices include restricted use of anthelmintics, late weaning of piglets (after min seven weeks of age) and access to outdoor runs. Provision of bedding material, which might be conducive for development and survival of free-living stages of helminths, is mandatory in organic production systems but the amount and type of bedding material vary between farm and housing systems. The differences in A. suum egg development in different housing systems have been poorly described. Biodegradation of organic matter in deep litter housing systems produces physico-chemical conditions which may affect the development and survival of A. suum eggs, but little is known about such effects. Understanding the ecology and transmission dynamics of A. suum may help prevent infections in organic farming. Pig slurry is used to fertilize crops in organic as well as conventional farming and may be treated to inactivate a variety of pathogens present in it before application. To achieve adequate pathogen inactivation, slurry may need to be stored for around one year which may not be ideal logistically and economically. Chemical treatment of slurry may reduce storage time needed to adequately inactivate pathogens. The eggs of A. suum are commonly used as a conservative indicator in estimating pathogen inactivation during slurry treatment as they are more resistant to environmental stress than bacterial and viral pathogens. The eggs of the poultry ascarid, Ascaridia galli are ubiquitous, resistant and available even in places where the eggs of A. suum cannot be found, e.g., areas without pig keeping, but little is known about the suitability of A. galli eggs as a conservative hygiene indicator. Currently, viability of A. suum eggs is estimated by their ability to embryonate which takes up to six weeks depending on temperature. There is a need to develop more rapid viability tests and in addition, embryonation cannot be used as a viability criterion for already embryonated eggs. A quantitative egg hatch test may thus be helpful for already embryonated eggs. The present Ph.D. project therefore addressed the above issues by investigating in six separate studies, the ecology and transmission of A. suum eggs in organic pig farms, including determination of conditions under which A. suum eggs are inactivated

ØKOLOGISKE GRISE HAR MANGE SPOLORM

Svinets spolorm er uhyre veltilpasset til grisenes levevis i de økologiske besætninger, da ormene smitter via meget hårdføre æg. Et igangværende projekt (PAROL) søger derfor at mindske smittepresset og mindske brugen af ormemidler, så bæredygtigheden styrkes

Antagonistic effect of chitin-degrading microfungi on thick-shelled pig helminth eggs

The rising demand for pigs produced in organic farming systems, which require access to outdoor areas and associated increased risk of gastrointestinal nematode infections, is creating growing interest in alternative control strategies. Also development of anthelminthic resistance due to extensive use of anthelmintics is a concern in intensive pig production. Use of chitin-degrading microfungi is one of the options. Extensive research has been conducted on biological control of plant pathogenic nematodes using such fungi, and some fungal species are commercially available for plant protection. In contrast, there are only limited laboratory studies conducted on eggs from nematodes which are animal parasites. The aim of present study is to investigate the antagonistic effect of chitin-degrading microfungi on thick-shelled pig helminth eggs (i.e. Ascaris suum and Trichuris suis) both in the laboratory as well as in the field. We have performed a pilot study of the ovicidal effect of Paecilomyces lilacinus alone and in combination with chitin supplement on A. suum eggs over 4 weeks in 2% water agar. Within 1 week more than 90% of the eggs were hatched in both cases within the Petri dish as compared to controls, indicating that the fungus is capable of degrading the chitin wall so the larvae are becoming free. Soon we will proceed for full-fledged study with 2 fungal species (P. lilacinus and Pochonia chlamydosporia) which were proven to have ovicidal activity against nematode eggs. This will be done in the laboratory using soil samples amended with A. suum eggs, under semi-natural (pasture plot) conditions and finally on heavily contaminated pastures

KONTROL AF SPOLORM I ØKOLOGISKE SVIN. Resultater fra PAROL projektet: Parasitter i økologiske produktionsdyr: innovative løsninger til nye udfordringer

Spolorm er meget almindelige i danske økologiske svin og kan reducere tilvækst, foderudnyttelse og effekt af vacciner hos smittede grise samt være årsag til kassering/værditab af levere. Spolorm smitter via hårdføre æg og projektet har kortlagt smittemønstre i 5 besætninger og undersøgt hvor smittepresset er størst og hvordan vi bedst kan spredningen i danske økologiske (og frilands-) besætninger

Evaluation of soil microfungi as biological control agents against ascarid eggs

Thick-shelled ascarid eggs have been reported to remain infective in the environment for several years, thus posing a prolonged risk of infection to animals and/or humans. The following in vitro study was therefore conducted to evaluate the negative impact of two species of naturally occuring soil microfungi (Pochonia chlamydosporia and Paecilomyces lilacinus), on the viability of Ascaridia galli, Toxocara canis and Ascaris suum eggs. Approximately 150 fresh eggs of individual ascarid species were embryonated on a 2% water agar in Petri dishes with or without a fungus (P. chlamydospria or P. lilacinus). On days 7, 14, 21, 28, 35 and 42 post experimental set up (p.s.), the viability of the eggs in each experimental group was evaluated (destructive sampling). By day 14 p.s., P. chlamydosporia had reduced the viability of A. galli and T. canis eggs by 70-86% and 52-67%, respectively, compared to the controls. In contrast, P. lilacinus had reduced the viability of A. galli and T. canis eggs by only 17-30% and 6-28%, respectively. Neither fungal species was found to be effective against A. suum eggs (<4% reduction in both cases). These results indicate interspecies differences in the susceptibility of ascarid eggs to microfungi. Ascaridia galli and T. canis eggs seemed to have been degenerated mainly due to hydrolysis of shells by fungal enzymes. The present study demonstrates that P. chlamydosporia may potentially be utilized as a biological control agent against A. galli and T. canis eggs in the environment

Development and survival of Ascaris suum eggs in deep litter of pigs

Indoor transmission of Ascaris suum partly depends on the physico-chemical conditions in bedding material. Temperature,pH, aqueous ammonia, moisture, occurrence and development of A. suum eggs were therefore compared in different areas(resting, intermediate and latrine) of two deep litter pens on an organic farm in four seasons. There was some variation, but mean ammonia levels were generally very low (1·0–2·6mM) and pH levels were moderate (8·04–8·88) in all three areas. Relatively, resting areas were characterized by overall moderate moisture (36%) and moderately high temperature (35·7 °C)levels. The area contained few eggs (50 eggs g−1 DM) of which 17% were viable, and though only 4% were larvated and 0·7% appeared infective, it was more than in the other areas. Intermediate areas had moderate moisture (43%) and high temperature (43·6 °C) levels. There were many eggs (523 eggs g−1 DM), but overall viability was very low (5%) and few eggs were larvated (0·004%) or even infective (0·002%). Latrines typically had high moisture (79%) and moderate temperature 30 °C) levels. The concentration of eggswas very high (1444 egg g−1DM)and though 32%were viable, none had developed larval stages. The large majority of A. suum eggs appear to die and only few become infective while in the deep litter. However, a large fraction of eggs may remain viable for some time and could thus contaminate agricultural land and develop to infectivity, if the manure is not composted appropriately

Evaluation of soil microfungi as biological control agents against eggs of animal parasitic nematodes

Thick-shelled eggs of ascarid nematodes have been reported to remain infective in the environment for several years, thus posing a prolonged risk of infection to animal livestock and/or humans. An in vitro study was therefore conducted to evaluate the negative impact of two species of soil microfungi, Pochonia chlamydosporia and Purpureocillium lilacinum (syn. Paecilomyces lilacinus), on the viability of Ascaridia galli, Toxocara canis and Ascaris suum eggs. Approximately 150 fresh eggs of individual ascarid species were embryonated on a 2% water agar in Petri dishes with or without a fungus (P. chlamydospria or P. lilacinum). On days 7, 14, 21, 28, 35 and 42 post experimental set up (p.s.), the viability of the eggs from each experimental group was evaluated (destructive sampling). By day 14 p.s., P. chlamydosporia had reduced the viability of A. galli and T. canis eggs by 70-86% and 52-67%, respectively, compared to the controls. In contrast, P. lilacinum had reduced the viability of A. galli and T. canis eggs by only 17-30% and 6-28%, respectively. Neither fungal species was found to be effective against A. suum eggs (<4% reduction in both cases). These results indicate interspecies differences in the susceptibility of ascarid eggs to soil microfungi. Ascaridia galli and T. canis eggs seemed to have been degenerated mainly due to hydrolysis of shells by fungal enzymes. The present study demonstrates that P. chlamydosporia may potentially be utilized as a biological control agent against A. galli and T. canis egg contaminations in the soil environment

Survival of Ascaris suum and Ascaridia galli eggs in liquidmanure at different ammonia concentrations andtemperatures

tEggs of Ascaris suum from pigs are highly resistant and commonly used as a conservativeindicator of pathogen inactivation during slurry storage. Eggs of Ascaridia galli, the poultryascarid, are also known to be highly resistant but the suitability as an indicator of pathogeninactivation has never been tested. Pig slurry has to be stored for several months to inac-tivate pathogens but chemical treatment of slurry may reduce this time. The suitability ofA. galli as an indicator of slurry sanitation was tested by comparing the survival of eggsof A. suum and A. galli in pig slurry. In addition, the effect of urea treatment on inactiva-tion of ascarid eggs in relation to storage time was also tested. Nylon bags with 10,000eggs 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, 36 h at 40◦C or2 h at 50◦C. At all the temperatures in both slurry types, A. galli eggs were inactivated ata significantly faster rate (P < 0.05) compared to A. suum eggs. For each 10◦C raise in tem-perature from 20◦C, T50(time needed to inactivate 50% of eggs) for both types of eggswas reduced markedly. At all temperatures, viability of eggs of both species was signifi-cantly higher (P < 0.05) in untreated slurry compared to urea-treated slurry except A. gallieggs at 20◦C where no significant difference was detected. In untreated slurry, the levelsof pH (6.33–9.08) and ammonia (0.01–1.74 mM) were lower (P < 0.0001) compared to thatof urea-treated slurry (pH: 8.33–9.28 and ammonia 1–13 mM). The study demonstratedthat 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 ofpathogens that are more sensitive than A. galli eggs. Addition of urea may markedly reducethe storage time of slurry needed to inactivate A. suum and A. galli eggs

Overlevelse af sundhedsskadelige mikroorganismer (bakterier/vira) eller parasitter ved slangeudlægning og nedfældning af gylle?

De fleste æg af parasitter vil overleve lang tid ved lave temperaturer i gylle, men dør hurtigt ved høje temperaturer (>35oC). Spredning af gylle vil medføre risiko for smitte, hvis afgrøder benyttes/afgræsses kort tid efter. Nedfældning øger risiko for nedsivning og vil med stor sandsynlighed øge overlevelsen af parasitter

Prevalence of the protozoan parasite Cryptosporidium on three organic pig farms in Denmark

Pigs are potential sources of contamination with Cryptosporidium spp., which can lead to infection in humans. Cryptosporidiosis in humans is primarily caused by two species, C. hominis and the zoonotic C. parvum, however, other species including C. suis and C. scrofarum are able to cause zoonotic infection. The oocysts can survive for long periods in the environment and are able to resist most disinfectants. In order to estimate the prevalence of Cryptosporidium spp. in organic pigs and to improve knowledge of the epidemiology, the oocyst excretion was monitored at quarterly intervals during the period September 2011 to June 2012 in each of three organic, Danish pig farms. Faecal samples for examination of Cryptosporidium spp. were collected from 994 pigs allocated into four age groups, piglets (n = 161), weaners (n = 315), fatteners (n = 232) and sows (n = 286), distributed on the three farms and four sampling times. Oocysts were quantified by immunofluorescence microscopy, and the overall prevalence of Cryptosporidium spp. was found to be 38%. The prevalence varied significantly between age groups with 45% piglets, 65% weaners, 40% fatteners and 3% sows found positive. No significant difference in the overall prevalence was detected between farms or seasons. Intensity of infection was age dependent with piglets and weaners having the highest oocyst excretion. No clear differences in the intensity of infections were seen between the four sampling times, but an apparent correlation was observed between number of positive sows and number of piglets with massive oocyst excretion (>105 oocysts per gram faeces ( OPG). Nine of the ten animals with highest OPG originated from the same farm which also had the highest prevalence in sows. Molecular characterisation of the collected Cryptosporidium isolates is ongoing and will reveal the zoonotic potential as well as the genetic variation between farms, seasons and age groups