title: Differential expression of β-tubulin isotypes in different life stages of Parascaris spp after exposure to thiabendazole.
abstract: Anthelmintic resistance (AR) to macrocyclic lactones (ML) has been described in Parascaris of horses world-wide. In contrast, benzimidazoles (BZ) are still effective, although reduced efficacy to this drug class was recently reported. The mode of action of BZ is binding to β-tubulin, which prevents polymerisation of microtubules. In this study, β-tubulin gene expression of isotypes 1 and 2 was investigated at seven time points (0, 6, 24, 72, 96 and 120 h) during embryogenesis and in adult worms. In addition, an in ovo larval developmental test was developed to study β-tubulin gene expression of both isotypes in parasacaris eggs after exposure to different concentrations of thiabendazole (TBZ) for five days at 25 °C. A strong pattern of differential expression of β-tubulin and isotype 1 was observed in all stages, while isotype 2 expression was mainly found at an early phase of the embryogenesis. For isotype 1, a 5-fold increase was observed during the first 48 h, but gene expression gradually decreased after 72, 96 and 120 h. Isotype 2 was only expressed during the first 24h, followed by a 130-fold decrease at (time points) 72, 96 and 120 h. The in ovo larval developmental test, in which we exposed initially unembryonated eggs to increased concentrations of TBZ, did affect isotype 1 gene expression but not isotype 2. This assumes that each isotype has specific functions in different life stages. This is in agreement with the 'multi-tubulin' hypothesis, which states that different tubulin isotypes are required for specialised microtubule functions. Isotype 1 is the most likely drug target for BZs, as isotype 2 was only expressed at very low levels later in development. Increasing concentrations of TBZ altered β-tubulin isotype 1 gene expression after exposure of the eggs for five days, but this was not seen for isotype 2.
authors.affiliation: Department of Biomedical Sciences and Veterinary Public Health, Division of Parasitology, Swedish University of Agricultural Sciences, S-750 07 Uppsala, Sweden. Electronic address: Eva.tyden@slu.se.
authors.forename: Eva
authors.initials: E
authors.lastname: Tydén
authors.affiliation: Department of Biomedical Sciences and Veterinary Public Health, Division of Parasitology, Swedish University of Agricultural Sciences, S-750 07 Uppsala, Sweden.
authors.forename: Moa
authors.initials: M
authors.lastname: Skarin
authors.affiliation: Department of Economics, Swedish University of Agricultural Sciences, S-750 07 Uppsala, Sweden.
authors.forename: Mikael
authors.initials: M
authors.lastname: Andersson-Franko
authors.affiliation: Department of Biomedical Sciences and Veterinary Public Health, Division of Parasitology, Swedish University of Agricultural Sciences, S-750 07 Uppsala, Sweden.
authors.forename: Matilda
authors.initials: M
authors.lastname: Sjöblom
authors.affiliation: Department of Biomedical Sciences and Veterinary Public Health, Division of Parasitology, Swedish University of Agricultural Sciences, S-750 07 Uppsala, Sweden.
authors.forename: Johan
authors.initials: J
authors.lastname: Höglund
mesh_terms: D000818:Animals; D000871:Anthelmintics / Q000494:pharmacology*; D017191:Ascaridida Infections / Q000188:drug therapy / Q000469:parasitology; D001199:Ascaridoidea / Q000145:classification / Q000187:drug effects* / Q000235:genetics / Q000254:growth & development*; D004305:Dose-Response Relationship, Drug; D004351:Drug Resistance; D018507:Gene Expression Regulation, Developmental; D015801:Helminth Proteins / Q000235:genetics* / Q000378:metabolism; D006734:Horse Diseases / Q000188:drug therapy / Q000469:parasitology; D006736:Horses; D007814:Larva / Q000187:drug effects / Q000235:genetics; D020033:Protein Isoforms; D013827:Thiabendazole / Q000494:pharmacology*; D014404:Tubulin / Q000235:genetics* / Q000378:metabolism
chemical_list: D000871:Anthelmintics; D015801:Helminth Proteins; D020033:Protein Isoforms; D014404:Tubulin; D013827:Thiabendazole
keywords: Embryogenesis; Gene expression; Life stages; Parascaris spp; Thiabendazole; β-tubulin