Development of a hydrolysis probe-based real-time assay for the detection of tropical strains of Fusarium oxysporum f. sp. cubense race 4.
Development of a hydrolysis probe-based real-time assay for the detection of tropical strains of Fusarium oxysporum f. sp. cubense race 4.
复制标题
DOI：
10.1371/journal.pone.0171767
复制
发表时间：
2017
期刊：
影响因子：
3.7
通讯作者：
Ioos R
中科院分区：
文献类型：
--
作者：
Aguayo J;Mostert D;Fourrier-Jeandel C;Cerf-Wendling I;Hostachy B;Viljoen A;Ioos R
香蕉枯萎香蕉黑胫病菌（Foc）是全球香蕉生产的重要威胁之一。缺乏控制病原体的战略，如果有抗性来源，植物抗性是唯一的长期解决方案。因此，防止将Foc引入无病地区仍然是继续可持续香蕉生产的关键战略。近年来，影响卡文迪什香蕉的Foc菌株摧毁了亚洲和中东一些国家以及一个非洲国家的种植园。其中一个营养体相容性组（VCG）01213/16被认为是热带和亚热带气候条件下香蕉的主要威胁。然而，其他遗传相关的VCG，如0121，可能会危及香蕉文化，如果他们被引入到无病地区。为了防止将这些VCG引入无病害的卡文迪什香蕉种植国家，开发了一种实时PCR测试来准确检测这两种VCG。使用先前描述的推定毒力基因开发用于检测热带Foc 4号小种菌株的水解探针/引物的特异性组合。实时PCR参数进行了优化，通过以下的统计方法依赖于正交表和田口方法，试图提高灵敏度，并确保高特异性的测定。本研究还评估了关键性能标准，如重复性、再现性、耐用性和专属性，包括一组136 F。尖孢菌株，包括73个Foc致病菌株代表24个VCG。验证数据表明，新的检测方法可用于香蕉植物材料的监管测试应用，并有助于防止影响卡文迪什香蕉的Foc菌株在热带地区的引入和传播。
Fusarium oxysporum f. sp. cubense (Foc) is one of the most important threats to global banana production. Strategies to control the pathogen are lacking, with plant resistance offering the only long-term solution, if sources of resistance are available. Prevention of introduction of Foc into disease-free areas thus remains a key strategy to continue sustainable banana production. In recent years, strains of Foc affecting Cavendish bananas have destroyed plantations in a number of countries in Asia and in the Middle East, and one African country. One vegetative compatibility group (VCG), 01213/16, is considered the major threat to bananas in tropical and subtropical climatic conditions. However, other genetically related VCGs, such as 0121, may potentially jeopardize banana cultures if they were introduced into disease-free areas. To prevent the introduction of these VCGs into disease-free Cavendish banana-growing countries, a real-time PCR test was developed to accurately detect both VCGs. A previously described putative virulence gene was used to develop a specific combination of hydrolysis probe/primers for the detection of tropical Foc race 4 strains. The real-time PCR parameters were optimized by following a statistical approach relying on orthogonal arrays and the Taguchi method in an attempt to enhance sensitivity and ensure high specificity of the assay. This study also assessed critical performance criteria, such as repeatability, reproducibility, robustness, and specificity, with a large including set of 136 F. oxysporum isolates, including 73 Foc pathogenic strains representing 24 VCGs. The validation data demonstrated that the new assay could be used for regulatory testing applications on banana plant material and can contribute to preventing the introduction and spread of Foc strains affecting Cavendish bananas in the tropics.