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Control of helminth infections is a major task in livestock production to prevent health constraints and economic losses. However, resistance to established anthelmintic substances already impedes effective anthelmintic treatment in many regions worldwide. Thus, there is an obvious need for sensitive and reliable methods to assess the resistance status of at least the most important nematode populations. Several single nucleotide polymorphisms (SNPs) in the beta-tubulin isotype 1 gene of various nematodes correlate with resistance to benzimidazoles (BZ), a major anthelmintic class. Here we describe the full-length beta-tubulin isotype 1 and 2 and alpha-tubulin coding sequences of the cattle nematode Ostertagia ostertagi. Additionally, the Cooperia oncophora alpha-tubulin coding sequence was identified. Phylogenetic maximum-likelihood analysis revealed that both isotype 1 and 2 are orthologs to the Caenorhabditis elegans ben-1 gene which is also associated with BZ resistance upon mutation. In contrast, a Trichuris trichiura cDNA, postulated to be beta-tubulin isotype 1 involved in BZ resistance in this human parasite, turned out to be closely related to C. elegans beta-tubulins tbb-4 and mec-7 and would therefore represent the first non-ben-1-like beta-tubulin to be under selection through treatment with BZs. A pyrosequencing assay was established to detect BZ resistance associated SNPs in beta-tubulin isotype 1 codons 167, 198 and 200 of C. oncophora and O. ostertagi. PCR-fragments representing either of the two alleles were combined in defined ratios to evaluate the pyrosequencing assay. The correlation between the given and the measured allele frequencies of the respective SNPs was very high. Subsequently laboratory isolates and field populations with known resistance status were analyzed. With the exception of codon 167 in Cooperia, increases of resistance associated alleles were detected for all codons in at least one of the phenotypically resistant population. Pyrosequencing provides a fast, inexpensive and sensitive alternative to conventional resistance detection methods.

Managing infections of sheep with gastrointestinal nematode parasites (worms) and problems of resistance to anthelmintic treatments continue to be major challenges for graziers on the Northern Tablelands of New South Wales, Australia. The whole-farmlet study of grazing enterprises undertaken by the Cicerone Project tested the broad hypotheses that compared with typical management (farmlet B), internal parasites can be more effectively managed with improved nutrition (farmlet A) or by intensive rotational grazing (farmlet C). Further aims were to identify the major sources of variation in faecal worm egg count (WEC) over the 6-year period and to examine the efficacy of the various anthelmintic treatments used during the experiment. This paper describes the management of sheep worms at the whole-farmlet level during the experiment, and analyses data from the routine WEC monitoring (5644 records) and larval differentiation tests (322 records) carried out on behalf of the Cicerone Management Board and by a doctoral candidate. It complements more detailed investigations published elsewhere. Over the period from July 2000 to December 2006, worm infections in ewes, lambs, hoggets and wethers were, with some exceptions, successfully controlled on the farmlets through a combination of regular monitoring of WEC, treatment with a wide array of anthelmintics and grazing management. Farmlet C had lower mean WEC (444 epg) and annual anthelmintic treatment frequency (3.1 treatments/year) over the whole experimental period than farmlets B (1122 epg, 4.3 treatments/year) or A (1374 epg, 4.7 treatments/year). The main factors influencing WEC were the time since the last anthelmintic treatment, and the anthelmintic used at that treatment. The magnitude of these effects dwarfed those of climatic and management factors that might be expected to influence the epidemiology of gastrointestinal nematode infections via environmental or host-mediated mechanisms. Nevertheless management factors associated with stocking rate and grazed proportion (proportion of each farmlet grazed at any one time), and climatic indicators of both temperature and moisture availability had significant effects on WEC. The results show that, in a region with Haemonchus contortus as the major sheep nematode, improved host nutrition in a higher input system (farmlet A) did not provide more effective control of gastrointestinal nematodes than typical management (farmlet B); however, it was observed that gastrointestinal nematode control was no worse on farmlet A than on farmlet B in spite of farmlet A supporting a 48% higher stocking rate by later in the trial period (2005). The study provided strong support for the proposition that intensive rotational grazing (farmlet C) provides more effective control of gastrointestinal nematodes than typical management (farmlet B) as evidenced by significantly lower WEC counts and anthelmintic treatment frequency. Tactical worm control based on routine monitoring of WEC provided adequate control of worms on all three farmlets for much of the experimental period but failed to prevent significant spikes in WEC to values associated with significant production loss on multiple occasions, and significant ewe mortality on farmlets A and B on one occasion.