Molecular evidence of widespread benzimidazole drug resistance in Ancylostoma caninum from domestic dogs throughout the USA and discovery of a novel β-tubulin benzimidazole resistance mutation.
Molecular evidence of widespread benzimidazole drug resistance in Ancylostoma caninum from domestic dogs throughout the USA and discovery of a novel β-tubulin benzimidazole resistance mutation.
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DOI：
10.1371/journal.ppat.1011146
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发表时间：
2023-03
期刊：
影响因子：
6.7
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中科院分区：
文献类型：
--
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犬钩虫是一种重要的人畜共患犬胃肠道线虫，与人类钩虫关系密切。我们最近报道了美国赛狗感染了A。通常对多种驱虫药耐药的犬。苯并咪唑抗性。犬中的F167Y（TTC> TAC）同种型-1 β-微管蛋白突变频率较高。在这项工作中，我们表明，苯并咪唑电阻是非常普遍的A。从美国各地的家犬身上提取的。首先，我们鉴定并显示了一种新的苯并咪唑同种型-1 β-微管蛋白抗性突变Q134H（CAA> CAT）的功能意义。几种苯并咪唑抗性A.来自灰狗的犬分离物具有低频率的F167Y（TTC> TAC）突变，具有高频率的Q134H（CAA> CAT）突变，该突变以前未从该领域的任何真核病原体中报道。结构建模预测，Q134残基直接参与苯并咪唑药物结合，134 H取代将显著降低结合亲和力。将Q134H取代引入C.通过CRISPR-Cas9编辑，线虫β-微管蛋白基因ben-1赋予了与ben-1无效等位基因相似的抗性水平。对A.来自685个钩虫阳性宠物狗粪便样本的犬卵显示，这两种突变在美国广泛存在，F167 Y（TTC> TAC）和Q134 H（CAA> CAT）的患病率分别为49.7%（总平均频率54.0%）和31.1%（总平均频率16.4%）。典型密码子198和200苯并咪唑耐药突变缺失。F167 Y（TTC> TAC）突变在美国西部的患病率和频率显著高于其他地区，我们假设这是由于避难所的差异。这项工作对伴侣动物寄生虫控制和人类钩虫耐药性的潜在出现具有重要意义。尽管在家畜中越来越常见，但迄今为止，在任何伴侣动物或人类胃肠道线虫寄生虫中均未证实广泛的驱虫剂耐药性报告。犬钩虫是一种常见的犬肠道人畜共患寄生虫，对幼犬有严重的临床影响，控制依赖于定期使用驱虫剂。我们最近报道了A.来自美国多个地点的灰狗的犬分离株可能是由犬舍长期强化治疗方案引起的。在这项研究中，我们研究了苯并咪唑抗性在A。在美国各地的宠物狗中发现了这种病毒。我们还发现了一个新的苯并咪唑同种型-1 β-微管蛋白抗性突变，并显示了其功能意义。犬从灰狗，没有以前在该领域的任何生物体的报告。然后，我们确定这种新的突变，以及先前表征的耐药突变，在许多A.全美国的犬科动物这项研究报告了伴侣动物的任何寄生线虫广泛耐药性的第一个证据，并说明了分子方法在一个地区快速评估驱虫剂耐药性的能力。
Ancylostoma caninum is an important zoonotic gastrointestinal nematode of dogs worldwide and a close relative of human hookworms. We recently reported that racing greyhound dogs in the USA are infected with A. caninum that are commonly resistant to multiple anthelmintics. Benzimidazole resistance in A. caninum in greyhounds was associated with a high frequency of the canonical F167Y(TTC>TAC) isotype-1 β-tubulin mutation. In this work, we show that benzimidazole resistance is remarkably widespread in A. caninum from domestic dogs across the USA. First, we identified and showed the functional significance of a novel benzimidazole isotype-1 β-tubulin resistance mutation, Q134H(CAA>CAT). Several benzimidazole resistant A. caninum isolates from greyhounds with a low frequency of the F167Y(TTC>TAC) mutation had a high frequency of a Q134H(CAA>CAT) mutation not previously reported from any eukaryotic pathogen in the field. Structural modeling predicted that the Q134 residue is directly involved in benzimidazole drug binding and that the 134H substitution would significantly reduce binding affinity. Introduction of the Q134H substitution into the C. elegans β-tubulin gene ben-1, by CRISPR-Cas9 editing, conferred similar levels of resistance as a ben-1 null allele. Deep amplicon sequencing on A. caninum eggs from 685 hookworm positive pet dog fecal samples revealed that both mutations were widespread across the USA, with prevalences of 49.7% (overall mean frequency 54.0%) and 31.1% (overall mean frequency 16.4%) for F167Y(TTC>TAC) and Q134H(CAA>CAT), respectively. Canonical codon 198 and 200 benzimidazole resistance mutations were absent. The F167Y(TTC>TAC) mutation had a significantly higher prevalence and frequency in Western USA than in other regions, which we hypothesize is due to differences in refugia. This work has important implications for companion animal parasite control and the potential emergence of drug resistance in human hookworms. Although increasingly common in livestock, no reports of widespread anthelmintic resistance are confirmed in any companion animal or human gastrointestinal nematode parasite to date. The canine hookworm is a common intestinal zoonotic parasite of dogs with severe clinical impacts in young dogs, and for which control is dependent on regular anthelmintic use. We recently reported multiple anthelmintic drug resistance in A. caninum isolates from greyhounds derived from multiple locations in the USA likely caused by long standing intensive treatment regimens in kennels. In this study, we investigated benzimidazole resistance in A. caninum in pet dogs across the USA. We also identified and showed the functional significance of a novel benzimidazole isotype-1 β-tubulin resistance mutation in A. caninum from greyhounds that has not been previously reported in the field for any organism. We then determined that this novel mutation, as well as a previously characterized resistance mutation, were present, often at high frequency, in many A. caninum populations across the USA. This study reports the first evidence of widespread drug resistance for any parasitic nematode of companion animals and illustrates the power of molecular approaches to rapidly assess anthelmintic resistance in a region.
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影响因子：
64.8
作者：
Jumper J;Evans R;Pritzel A;Green T;Figurnov M;Ronneberger O;Tunyasuvunakool K;Bates R;Žídek A;Potapenko A;Bridgland A;Meyer C;Kohl SAA;Ballard AJ;Cowie A;Romera-Paredes B;Nikolov S;Jain R;Adler J;Back T;Petersen S;Reiman D;Clancy E;Zielinski M;Steinegger M;Pacholska M;Berghammer T;Bodenstein S;Silver D;Vinyals O;Senior AW;Kavukcuoglu K;Kohli P;Hassabis D
通讯作者：
Hassabis D
影响因子：
4
作者：
Avramenko, Russell W.;Redman, Elizabeth M.;Gilleard, John S.
通讯作者：
Gilleard, John S.
影响因子：
48
作者：
Callahan BJ;McMurdie PJ;Rosen MJ;Han AW;Johnson AJ;Holmes SP
通讯作者：
Holmes SP
DOI：
10.1016/j.ijpddr.2021.08.005
发表时间：
2021-12
期刊：
International journal for parasitology. Drugs and drug resistance
影响因子：
--
作者：
Jimenez Castro PD;Venkatesan A;Redman E;Chen R;Malatesta A;Huff H;Zuluaga Salazar DA;Avramenko R;Gilleard JS;Kaplan RM
通讯作者：
Kaplan RM
影响因子：
8.3
作者：
BECKER, SV;SELBY, LA;HACKER, DV
通讯作者：
HACKER, DV