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    Benzimidazole resistance survey for Haemonchus, Teladorsagia and Trichostrongylus in three European countries using pyrosequencing including the development of new assays for Trichostrongylus (2016)

    Art
    Zeitschriftenartikel / wissenschaftlicher Beitrag
    Autoren
    Ramünke, Sabrina (WE 13)
    Melville, Lynsey
    Rinaldi, Laura
    Hertzberg, Hubertus
    de Waal, Theo
    von Samson-Himmelstjerna, Georg (WE 13)
    Cringoli, Giuseppe
    Mavrot, Fabien
    Skuce, Philip
    Krücken, Jürgen (WE 13)
    Demeler, Janina (WE 13)
    Forschungsprojekt
    7. FP der EU: Innovative and sustainable strategies to mitigate the impact of global change on helminth infections in ruminants
    Quelle
    International journal for parasitology. Drugs and drug resistance; 6(3) — S. 230–240
    ISSN: 2211-3207
    Verweise
    DOI: 10.1016/j.ijpddr.2016.10.002
    Pubmed: 27821282
    Kontakt
    Institut für Parasitologie und Tropenveterinärmedizin

    Robert-von-Ostertag-Str. 7-13
    Gebäude 35, 22, 23
    14163 Berlin
    Tel.+49 30 838 62310 Fax.+49 30 838 62323
    email:parasitologie@vetmed.fu-berlin.de

    Abstract / Zusammenfassung

    Resistance to benzimidazoles (BZs) in trichostrongyloid nematodes is a worldwide problem for livestock production, particularly regarding small ruminants. Sensitive and reliable methods are required to assess anthelmintic resistance status. Currently available methods for BZ resistance detection can be divided into three main groups, in vivo (e.g. faecal egg count reduction test), in vitro (e.g. egg hatch assay) and molecular tests. Three single nucleotide polymorphisms (SNPs) in the isotype-1 β-tubulin gene of various nematode species correlate with BZ resistance. While PCR-based methods have been reported for the three most economically important nematodes of sheep, namely, Trichostrongylus, Haemonchus and Teladorsagia, pyrosequencing assays are so far only available for the latter two. Here, the design and evaluation of pyrosequencing assays for isotype-1 and isotype-2 β-tubulin genes of Trichostrongylus colubriformis are described. PCR fragments carrying the susceptible and corresponding resistant genotype were combined in defined ratios to evaluate assay sensitivity and linearity. The correlation between the given and the measured allele frequencies of the respective SNPs (codons F167Y, E198A and F200Y) was very high. Pyrosequencing assays for Haemonchus, Teladorsagia and Trichostrongylus were subsequently used for a BZ resistance survey, carried out in the three European countries, namely Ireland, Italy and Switzerland. Larval cultures obtained from field survey samples in 2012 and 2013 were used for pyrosequencing. The test was applied when the target species represented at least 10% of the sample. Trichostrongylus and Teladorsagia were detected in all countries' samples whereas Haemonchus was not detected in samples from Ireland. SNPs in isotype-1 associated with resistance were detected for all three species, with frequencies at codon F200Y far exceeding those at codons F167Y and E198A. Elevated SNP frequencies in isotype-2 of Trichostrongylus were only rarely detected. Farms with BZ resistance-associated SNP frequencies above 10% were most often found in Switzerland followed by Ireland and Italy.