Phenotypic and genotypic characterisation of benzimidazole susceptible and resistant isolates of Haemonchus contortus.

The study was designed to compare the in vitro egg hatch test (EHT) and molecular allele-specific polymerase chain reaction PCR (AS-PCR) methods as tools for detection of benzimidazole resistance in Haemonchus contortus, a nematode parasite of small ruminants. Comparisons were made during the course of an experimental infection and changes in EHT and AS-PCR were monitored to measure the correlation between in vitro and molecular tests. Both methods were carried out according to World Association for the Advancement of Veterinary Parasitology (WAAVP) recommendations. The molecular test was used to discriminate TAC/TTC polymorphism in the beta-tubulin 200 codon of eight (four resistant and four susceptible) isolates of H. contortus. Using DNA from 100 third-stage larvae, the polymerase chain reaction (PCR)-based analysis revealed a decrease of the homozygous TTC/TTC genotype and an increase in heterozygous TTC/TAC and homozygous TAC/TAC individuals in all resistant isolates. Both methods showed comparable and reliable results with regard to detection of benzimidazole resistance. The molecular test has an advantage over the EHT because of its higher sensitivity. On the other hand, EHT is less time-consuming, allows reliable detection of <10% resistance allele frequency and is fairly reliable for the detection of benzimidazole resistance under field conditions.

Application of a Reverse Line Blot hybridisation assay for the species-specific identification of cyathostomins (Nematoda, Strongylida) from benzimidazole-treated horses in the Slovak Republic.

Five horse farms located in eastern Slovakia were investigated for the presence of benzimidazole-resistant strongyles by faecal egg count reduction test and egg hatch assay. Coprocultures were prepared for each farm from faecal samples taken pre- and post-treatment and harvested larvae were molecularly examined with a Reverse Line Blot assay. Faecal egg count reduction values ranged from 0 to 52.5% and all farms were positive for benzimidazole-resistant cyathostomins. Seven benzimidazole-resistant cyathostomin species were molecularly identified on farms before and also after treatment. These data demonstrate that resistance to benzimidazoles is well established in cyathostomin populations from horse farms in the Slovak Republic and that the molecular assay was able to determine the species-specific distribution of resistant cyathostomins under field conditions.

Worm control practices on sheep farms in the Slovak Republic.

A questionnaire to obtain information on worm control practices and sheep management was performed on 49 sheep farms in 2003 and 2004. The majority of Slovak farms kept native breeds Tsigai (22 farms) and Improved Valachian (14 farms). Farms were divided according to their altitude to lowland, upland and lower highland farms. Sizes of pastures and stocking rates for lowland, upland and lower highland farms were 81.5, 269.2, and 316.7 ha and 6.3, 2.6, and 2.9 sheep/ha, respectively. One third of farmers (33.3%) used permanent pastures and two thirds of breeders (66.7%) rotated sheep between pastures. Mean drenching rate for lambs and yearlings/adults was 1.76 and 1.70, respectively. The most frequently used drugs during period from 1999 to 2004 were albendazole and ivermectin. On 13 farms benzimidazole drugs were applied in spring before turn out and macrocyclic lactones in autumn before turn in. Benzimidazoles and macrocyclic lactones were used almost exclusively on 7 and 9 farms, respectively. Visual appraisal was the most common practice to determine weight of animals (87.8% of farmers). Weights of the heaviest animals were used for determination of anthelmintic doses only on 16.7% of farms. Coprological examinations were performed on 47.9% of farms, usually in frequency once per year (75%).