C239S Mutation in the β-Tubulin of Phytophthora sojae Confers Resistance to Zoxamide
C239S Mutation in the β-Tubulin of Phytophthora sojae Confers Resistance to Zoxamide
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DOI：
10.3389/fmicb.2010.00762
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发表时间：
2016-05-20
影响因子：
5.2
通讯作者：
Tyler, Brett M.
中科院分区：
文献类型：
--
作者：
Cai, Meng;Miao, Jianqiang;Tyler, Brett M.
唑沙胺是唯一注册用于控制卵菌病原体的β -微管蛋白抑制剂。本研究考察了zoxamide对大豆疫霉菌的活性，建立了基线敏感性，平均EC50为0.048 μ g/ml。这些数据对于监测实地唑胺敏感性的变化至关重要。在经唑沙胺修饰的培养基上筛选获得了3个稳定的高抗性突变体。虽然抗性发生的频率较低，但在获得性突变体的生长速度、产孢量、萌发和致病性方面没有明显的适合度损失。基于其生物学特性和较低的突变率，大豆对唑沙胺的抗性风险为低至中等。进一步研究发现，所有抗唑胺突变体的β -微管蛋白都有C239S点突变。Zoxamide对大多数种的拮抗也表现出较高的活性，其中只有phhanidermatum对Zoxamide具有天然抗性，并且在β -微管蛋白中含有天然点突变S239。携带突变等位基因（S239）的大豆反转化证实，C239S突变可诱导对唑沙胺的抗性，且抗性水平与突变基因的表达水平呈正相关。相比之下，野生型基因的过表达不能引起唑沙胺抗性。本文首次报道了唑沙胺在卵菌中的耐药分子机制。根据我们的研究，C239可能是zoxamide的关键靶点，是zoxamide区别于苯并咪唑类药物的关键靶点，也是zoxamide耐药风险低的原因。该结果可为今后设计新型β -微管蛋白抑制剂提供参考。
Zoxamide is the sole beta-tubulin inhibitor registered for the control of oomycete pathogens. The current study investigated the activity of zoxamide against Phytophthora sojae and baseline sensitivity was established with a mean EC50 of 0.048 mu g/ml. The data is critical for monitoring changes in zoxamide-sensitivity in the field. Three stable resistant mutants with a high resistance level were obtained by selection on zoxamide amended media. Although the development of resistance occurred at a low frequency, there were no apparent fitness penalty in the acquired mutants in terms of growth rate, sporulation, germination and pathogenicity. Based on the biological profiles and low mutagenesis rate, the resistance risk of P sojae to zoxamide can be estimated as low to medium. Further investigation revealed all the zoxamide-resistant mutants had a point mutation of C239S in their beta-tubulin. Zoxamide also exhibited high activity against most species from the genus Pythium in which only Pythium aphanidermatum was found naturally resistant to zoxamide and harboring the natural point mutation S239 in the beta-tubulin. Back-transformation in P sojae with the mutated allele (S239) confirmed the C239S mutation can induce resistance to zoxamide, and the resistance level was positively related to the expression level of the mutated gene. In contrast, the overexpression of the wild type gene was unable to cause zoxamide resistance. It is the first report on the resistance molecular mechanism of zoxamide in oomycetes. Based on our study, C239 is supposed to be a key target site of zoxamide, which distinguishes zoxamide from benzimidazoles and accounts for its low resistance risk. The result can provide advice on the design of new beta-tubulin inhibitors in future.