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<p>Anticoccidial drug resistance in <i>Eimeria</i> spp. presents a growing threat to global poultry health and food security, necessitating robust molecular tools for resistance surveillance. In this study, we report the identification and functional validation of a single nucleotide mutation (c.1255 C &gt; T) in the <i>ETH2_0402100</i> gene of <i>Eimeria tenella</i> that confers resistance to maduramicin, a widely used ionophore anticoccidial drug. Using genetic crossing, backcrossing, and QTL-seq, we narrowed the resistance locus to a 76 kb region on chromosome HG994964.1. CRISPR/Cas9-mediated site-directed mutagenesis and gene overexpression confirmed that the leucine-to-phenylalanine substitution in <i>ETH2_0402100</i> confers maduramicin resistance. To translate this discovery into a practical tool, we developed a PCR-based amplification sequencing assay that enables rapid, quantitative detection of maduramicin-resistant alleles from field samples. Together, these findings establish the molecular basis and monitoring strategy for maduramicin resistance in <i>Eimeria</i> spp., and provide a field-deployable molecular diagnostic tool for detecting ionophore resistance.</p>

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Target gene mutation discovery and amplicon sequencing for evaluating maduramicin resistance in Eimeria tenella

  • 期刊论文

  • 出版模式：

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  • 发表日期：

    2025年12月20日

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  • Yang Gao,
  • Xiaojin Li,
  • Meng-di Yuan,
  • Qingbin Guo,
  • Sixin Zhang,
  • Zixuan Wang,
  • Yuehua Xie,
  • Hongyu Han,
  • Xianyong Liu,
  • Yuanyuan Zhang,
  • Xun Suo

摘要

Anticoccidial drug resistance in Eimeria spp. presents a growing threat to global poultry health and food security, necessitating robust molecular tools for resistance surveillance. In this study, we report the identification and functional validation of a single nucleotide mutation (c.1255 C > T) in the ETH2_0402100 gene of Eimeria tenella that confers resistance to maduramicin, a widely used ionophore anticoccidial drug. Using genetic crossing, backcrossing, and QTL-seq, we narrowed the resistance locus to a 76 kb region on chromosome HG994964.1. CRISPR/Cas9-mediated site-directed mutagenesis and gene overexpression confirmed that the leucine-to-phenylalanine substitution in ETH2_0402100 confers maduramicin resistance. To translate this discovery into a practical tool, we developed a PCR-based amplification sequencing assay that enables rapid, quantitative detection of maduramicin-resistant alleles from field samples. Together, these findings establish the molecular basis and monitoring strategy for maduramicin resistance in Eimeria spp., and provide a field-deployable molecular diagnostic tool for detecting ionophore resistance.