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New codon 198 β-tubulin polymorphisms in highly benzimidazole resistant Haemonchus contortus from goats in three different states in Sudan

BACKGROUND: Benzimidazole (BZ) resistance in gastrointestinal nematodes is a worldwide problem for livestock production, particularly in small ruminants. Assignment of the emergence of resistance using sensitive and reliable methods is required to adopt the correct strategies for control. In Sudan,...

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Autores principales: Mohammedsalih, Khalid M., Krücken, Jürgen, Khalafalla, Amna, Bashar, Ahmed, Juma, Fathel-Rahman, Abakar, Adam, Abdalmalaik, Abdalhakaim A. H., Coles, Gerald, von Samson-Himmelstjerna, Georg
Formato: Online Artículo Texto
Lenguaje:English
Publicado: BioMed Central 2020
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7053126/
https://www.ncbi.nlm.nih.gov/pubmed/32122383
http://dx.doi.org/10.1186/s13071-020-3978-6
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author Mohammedsalih, Khalid M.
Krücken, Jürgen
Khalafalla, Amna
Bashar, Ahmed
Juma, Fathel-Rahman
Abakar, Adam
Abdalmalaik, Abdalhakaim A. H.
Coles, Gerald
von Samson-Himmelstjerna, Georg
author_facet Mohammedsalih, Khalid M.
Krücken, Jürgen
Khalafalla, Amna
Bashar, Ahmed
Juma, Fathel-Rahman
Abakar, Adam
Abdalmalaik, Abdalhakaim A. H.
Coles, Gerald
von Samson-Himmelstjerna, Georg
author_sort Mohammedsalih, Khalid M.
collection PubMed
description BACKGROUND: Benzimidazole (BZ) resistance in gastrointestinal nematodes is a worldwide problem for livestock production, particularly in small ruminants. Assignment of the emergence of resistance using sensitive and reliable methods is required to adopt the correct strategies for control. In Sudan, BZ resistant Haemonchus contortus populations were recently reported in goats in South Darfur. This study aimed to provide additional data regarding albendazole efficacy and to describe the prevailing molecular BZ resistance mechanisms. METHODS: Faecal egg count reduction and egg hatch tests (EHT) were used to evaluate albendazole efficacy in three different areas of South Darfur using naturally (Rehed Al-Birdi and Tulus) and experimentally infected (Tulus and Um Dafuq) goats. Using samples from Central, East and South Darfur, pyro- and Sanger sequencing were used to detect the polymorphisms F167Y, E198A and F200Y in H. contortus isotype 1 β-tubulin in DNA extracted from pooled third-stage larval (L3) samples (n = 36) on days 0 and 10 during trials, and from pooled adult male H. contortus (treated goats, n = 14; abattoirs, n = 83) including samples from populations previously found to be resistant in South Darfur. RESULTS: Albendazole efficacies at 5, 7.5 and 10 mg/kg doses were 73.5–90.2% on day 14 in natural and experimental infections while 12.5 mg/kg showed > 96.6% efficacy. EC(50) in the EHT were 0.8 and 0.11 µg/ml thiabendazole in natural and experimental infection trials, respectively. PCRs detected Haemonchus, Trichostrongylus and Cooperia in L3 samples from albendazole-treated goats. Haemonchus contortus allele frequencies in codons 167 and 200 using pyrosequencing assays were ≤ 7.4% while codon 198 assays failed. Sanger sequencing revealed five novel polymorphisms at codon 198. Noteworthy, an E198L substitution was present in 82% of the samples (L3 and adults) including all post-treatment samples. Moreover, E198V, E198K and potentially E198I, and E198Stop were identified in a few samples. CONCLUSIONS: To our knowledge, this is the first report of E198L in BZ resistant H. contortus and the second where this is the predominant genotype associated with resistance in any strongyle species. Since this variant cannot be quantified using pyrosequencing, the results highlight important limitations in the general applicability of pyrosequencing to quantify BZ resistance genotypes. [Image: see text]
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spelling pubmed-70531262020-03-10 New codon 198 β-tubulin polymorphisms in highly benzimidazole resistant Haemonchus contortus from goats in three different states in Sudan Mohammedsalih, Khalid M. Krücken, Jürgen Khalafalla, Amna Bashar, Ahmed Juma, Fathel-Rahman Abakar, Adam Abdalmalaik, Abdalhakaim A. H. Coles, Gerald von Samson-Himmelstjerna, Georg Parasit Vectors Research BACKGROUND: Benzimidazole (BZ) resistance in gastrointestinal nematodes is a worldwide problem for livestock production, particularly in small ruminants. Assignment of the emergence of resistance using sensitive and reliable methods is required to adopt the correct strategies for control. In Sudan, BZ resistant Haemonchus contortus populations were recently reported in goats in South Darfur. This study aimed to provide additional data regarding albendazole efficacy and to describe the prevailing molecular BZ resistance mechanisms. METHODS: Faecal egg count reduction and egg hatch tests (EHT) were used to evaluate albendazole efficacy in three different areas of South Darfur using naturally (Rehed Al-Birdi and Tulus) and experimentally infected (Tulus and Um Dafuq) goats. Using samples from Central, East and South Darfur, pyro- and Sanger sequencing were used to detect the polymorphisms F167Y, E198A and F200Y in H. contortus isotype 1 β-tubulin in DNA extracted from pooled third-stage larval (L3) samples (n = 36) on days 0 and 10 during trials, and from pooled adult male H. contortus (treated goats, n = 14; abattoirs, n = 83) including samples from populations previously found to be resistant in South Darfur. RESULTS: Albendazole efficacies at 5, 7.5 and 10 mg/kg doses were 73.5–90.2% on day 14 in natural and experimental infections while 12.5 mg/kg showed > 96.6% efficacy. EC(50) in the EHT were 0.8 and 0.11 µg/ml thiabendazole in natural and experimental infection trials, respectively. PCRs detected Haemonchus, Trichostrongylus and Cooperia in L3 samples from albendazole-treated goats. Haemonchus contortus allele frequencies in codons 167 and 200 using pyrosequencing assays were ≤ 7.4% while codon 198 assays failed. Sanger sequencing revealed five novel polymorphisms at codon 198. Noteworthy, an E198L substitution was present in 82% of the samples (L3 and adults) including all post-treatment samples. Moreover, E198V, E198K and potentially E198I, and E198Stop were identified in a few samples. CONCLUSIONS: To our knowledge, this is the first report of E198L in BZ resistant H. contortus and the second where this is the predominant genotype associated with resistance in any strongyle species. Since this variant cannot be quantified using pyrosequencing, the results highlight important limitations in the general applicability of pyrosequencing to quantify BZ resistance genotypes. [Image: see text] BioMed Central 2020-03-02 /pmc/articles/PMC7053126/ /pubmed/32122383 http://dx.doi.org/10.1186/s13071-020-3978-6 Text en © The Author(s) 2020 Open AccessThis article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated in a credit line to the data.
spellingShingle Research
Mohammedsalih, Khalid M.
Krücken, Jürgen
Khalafalla, Amna
Bashar, Ahmed
Juma, Fathel-Rahman
Abakar, Adam
Abdalmalaik, Abdalhakaim A. H.
Coles, Gerald
von Samson-Himmelstjerna, Georg
New codon 198 β-tubulin polymorphisms in highly benzimidazole resistant Haemonchus contortus from goats in three different states in Sudan
title New codon 198 β-tubulin polymorphisms in highly benzimidazole resistant Haemonchus contortus from goats in three different states in Sudan
title_full New codon 198 β-tubulin polymorphisms in highly benzimidazole resistant Haemonchus contortus from goats in three different states in Sudan
title_fullStr New codon 198 β-tubulin polymorphisms in highly benzimidazole resistant Haemonchus contortus from goats in three different states in Sudan
title_full_unstemmed New codon 198 β-tubulin polymorphisms in highly benzimidazole resistant Haemonchus contortus from goats in three different states in Sudan
title_short New codon 198 β-tubulin polymorphisms in highly benzimidazole resistant Haemonchus contortus from goats in three different states in Sudan
title_sort new codon 198 β-tubulin polymorphisms in highly benzimidazole resistant haemonchus contortus from goats in three different states in sudan
topic Research
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7053126/
https://www.ncbi.nlm.nih.gov/pubmed/32122383
http://dx.doi.org/10.1186/s13071-020-3978-6
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