Comparison of three artificial digestion methods for detection of non-encapsulated Trichinella pseudospiralis larvae in pork.

In a ring trial involving five laboratories (A, B, C, D, and E), three different methods of artificial digestion were compared for the detection of non-encapsulated Trichinella pseudospiralis larvae in minced meat. Each sample panel consisted of ten 1g minced pork samples. All samples in each panel were derived from a bulk meat preparation with a nominal value of either 7 or 17 larvae per g (lpg). Samples were tested for the number of muscle larvae using the magnetic stirrer method (labs A, B, and E), stomacher method (lab B), and Trichomatic 35 (labs C and D). T. pseudospiralis larvae were found in all 120 samples tested. For samples with 7 lpg, larval recoveries were significantly higher using the stomacher method versus the magnetic stirrer method, but there were no significant differences for samples with 17 lpg. In comparing laboratory results irrespective of the method used, lab B detected a significantly higher number of larvae than lab E for samples with 7 lpg, and lab E detected significantly less larvae than labs A, B, and D in samples with 17 lpg. The lowest overall variation for quantitative results (i.e. larval recoveries which were outside the tolerance range) was achieved by using the magnetic stirrer method (22%), followed by the stomacher method (25%), and Trichomatic 35 (30%). Results revealed that T. pseudospiralis larvae in samples with a nominal value of 7 and 17 lpg can be detected by all three methods of artificial digestion.

Population dynamics and intra-litter transmission patterns of Isospora suis in suckling piglets under on-farm conditions.

The aim of this study was to investigate the intra-litter infection dynamics of Isospora suis under natural conditions, and to study any association between parasite transmission and the contamination level of the farrowing pen by applying different interventions in order to reduce the transmission of I. suis infection within the litter. The study was divided in 2 trials including in total 22 litters (254 piglets). The first trial included 4 litters (where standard procedures practiced routinely on the farm piglets were applied) and the piglets were followed coprologically from farrowing until 2 weeks after weaning. The sows of those litters were also examined at various intervals before and after farrowing. The second trial included the application of 3 different management procedures: (A) standard farm hygiene and management procedures, (B) standard farm hygiene and management procedures+the first piglets found to excrete I. suis oocysts in each pen were removed from the pen, and (C) reduced cleaning. Each procedure was studied in 2 litters. This was replicated 3 times to yield a total of 18 litters. The results suggested that (i) the sow does not play an important role in transmission of I. suis in the farrowing pen; (ii) in natural infections, both the age of the piglet age at onset of oocyst excretion and the oocyst excretion patterns may vary considerably; (iii) the course of oocyst excretion or development of diarrhoea is related to the time of initial infection and (iii) piglets, which are heavy at birth, are more prone to acquire I. suis infection. Moreover, it was demonstrated that cleaning could be an effective means of restricting the spread of the parasite within the litter and thus the development of diarrhoea.

Interpretation of serum antibody response to Anoplocephala perfoliata in relation to parasite burden and faecal egg count.

REASON FOR PERFORMING STUDY: Increased knowledge is needed to assist in the interpretation of presently available diagnostic techniques for infection by the tapeworm Anoplocephala perfoliata in horses. HYPOTHESIS: The suggested cut-off level of an A. perfoliata specific ELISA may not adequately reflect the actual infection level. Hence, faecal egg counts may be a more useful diagnostic test for individual horses than previously reported. METHODS: Eighty-four horses admitted for slaughter at a Danish abattoir were examined for the presence of A. perfoliata. The number of tapeworms, their stage of development and gross pathological mucosal lesions were recorded and compared with serum antibody responses and faecal egg counts. Faecal egg counts were determined in samples from A. perfoliata infected horses using a semi quantitative centrifugation/flotation technique. Blood samples collected at slaughter were analysed by ELISA to determine serum antibody levels against A. perfoliata 12/13 kDa excretory/secretory antigens. RESULTS: Macroscopically visible tapeworms were detected in 24 (29%) of the horses. The overall sensitivity of the faecal egg count was found to be 0.46; however, if the detection limit was increased to above 20 tapeworms, sensitivity increased to 0.89. There was a correlation of 0.71 between worm burden and egg count. The antibody levels correlated significantly with infection intensity despite a wide variation among horses with similar levels of infection. The optimal cut-off value was determined using receiver operating characteristic analysis. If cut-off was chosen at optical density (OD) = 0.7, sensitivity was 0.68 and specificity 0.71. CONCLUSIONS: Both diagnostic methods were capable of revealing potentially pathogenic infections, with the faecal egg count being more applicable on the individual horse level. POTENTIAL RELEVANCE: In the population of Danish horses investigated the serum ELISA test should be interpreted such that horses in need of anti-Anoplocephala treatment have an OD = 0.7 or above.

Molecular and phylogenetic characterization of Cryptosporidium and Giardia from pigs and cattle in Denmark.

The genetic diversity of Cryptosporidium spp. and Giardia duodenalis from dairy cattle and pigs in Denmark was determined in the present study. Faecal samples from 1237 pigs and 1150 cattle originating from 50 sow herds and 50 dairy herds, respectively, were analysed for the presence of the two parasites by immunofluorescence microscopy. A large proportion of the (oo)cyst containing samples were selected for molecular characterization. Sequencing and phylogenetic analysis of the 18S rDNA locus and/or the HSP70 gene of 183 pig and 154 cattle isolates of Cryptosporidium revealed the presence of C. suis, pig genotype II, C. parvum (cattle genotype), C. bovis, Cryptosporidium deer-like genotype and a novel C. suis-like genotype. For both cattle and pigs, a host age-related change in distribution of species/genotypes was observed. The zoonotic C. parvum (cattle genotype) was most prevalent in young calves. For Giardia, 82 and 145 isolates from pigs and cattle, respectively, were analysed at the 18S rDNA locus and/or the gdh gene. Giardia isolates belonging to the zoonotic Assemblage A was found in both young and older calves, as well as in weaners and piglets, whereas cows seemed to be infected purely by isolates of the livestock group, Assemblage E.

World association for the advancement of veterinary parasitology (WAAVP): second edition of guidelines for evaluating the efficacy of anthelmintics in swine.

Guidelines are provided for evaluating the efficacy of anthelmintics in swine which, in conjunction with other sets of guidance such as those of the International Cooperation on Harmonization of Technical Requirements for Registration of Veterinary Medicinal Products (VICH GL7 and VICH GL16), should encourage the adoption of uniform registration requirements globally. Testing of efficacy should be carried out according to the principles of "Good Clinical Practice" (VICH GL9, 2000). Data obtained according to these guidelines should be internationally acceptable for the registration of anthelmintics for swine. Further, the use of the guidelines should expedite development, government review, and approval of anthelmintics for swine, as well as contribute towards reducing costs and the number of experimental animals used for drug testing.

Helminths of red foxes (Vulpes vulpes) in Denmark.

An epidemiological study of helminths in 1040 red foxes collected from various localities in Denmark during 1997-2002, revealed 21 helminth species at autopsy, including nine nematode species: Capillaria plica (prevalence 80.5%), Capillaria aerophila (74.1%), Crenosoma vulpis (17.4%), Angiostrongylus vasorum (48.6% from Northern Zealand (endemic area)), Toxocara canis (59.4%), Toxascaris leonina (0.6%), Uncinaria stenocephala (68.6%), Ancylostoma caninum (0.6%), and Trichuris vulpis (0.5%); seven cestodes: Mesocestoides sp. (35.6%), a number of Taeniid species (Taenia pisiformis, T. hydatigena, T. taeniaeformis, T. crassiceps, and unidentified Taenia spp.) (22.8%), and Echinococcus multilocularis (0.3%); four trematodes: Alaria alata (15.4%), Cryptocotyle lingua (23.8%), Pseudamphystomum truncatum (3.6% from Northern Zealand), and Echinochasmus perfoliatus (2.4% from Northern Zealand); one acanthocephalan: Polymorphus sp. (1.2%). Significant difference in prevalence was found for T. canis and A. vasorum according to host sex, and for T. canis, U. stenocephala, Mesocestoides sp., Taenia spp., A. alata, A. vasorum, and Capillaria spp. according to age groups (adult, young or cub). Prevalence and average worm intensity for each helminth species varied considerably according to geographical locality, season, and year. Aggregated distribution was found for several helminth species. The two species E. multilocularis and E. perfoliatus are first records for Denmark.

Meat inspection for Trichinella in pork, horsemeat and game within the EU: available technology and its present implementation.

A new EU directive relating to meat inspection for Trichinella, expected to come into force in 2006, imposes important modifications to current legislation. Nevertheless, several issues need more attention. Optimisation of methods, especially concerning sensitivity and digestibility of the meat to be inspected, along with further simplification of the legislation with regard to the number of techniques accepted, is recommended to guarantee that all member states of the EU will be given tools to perform inspection of consumer meat at the same high level. Additionally, there is a need for guidelines and protocols regarding optimal proficiency testing procedures. This paper presents an overview of the current methods for Trichinella meat inspection and their implementation in the EU, listing advantages and disadvantages for each method, including some suggestions for specific points of improvement.

Meat inspection for Trichinella in pork, horsemeat and game within the EU: available technology and its present implementation.

A new EU directive relating to meat inspection for Trichinella, expected to come into force in 2006, imposes important modifications to current legislation. Nevertheless, several issues need more attention. Optimisation of methods, especially concerning sensitivity and digestibility of the meat to be inspected, along with further simplification of the legislation with regard to the number of techniques accepted, is recommended to guarantee that all member states of the EU will be given tools to perform inspection of consumer meat at the same high level. Additionally, there is a need for guidelines and protocols regarding optimal proficiency testing procedures.

Cryptosporidium and cryptosporidiosis in Denmark--current status.

The genus Cryptosporidium comprises a group of protozoan parasites that infect a broad variety of vertebrates causing severe diarrhoeal illness in immunocompromised as well as immunocompetent hosts. Although molecular heterogeneity of the genus is being increasingly recognised, traditional diagnostic methods do not discriminate all species/subtypes, and population genetic studies of these parasites, using discriminatory molecular markers, have only been published recently. In Denmark, Cryptosporidium research has focussed mainly on detection methods, pathogenicity and veterinary aspects. The present paper gives an overview of recent and ongoing Cryptosporidium research in Denmark with an emphasis on molecular approaches to study epidemiology and transmission.