Key Laboratory of the Ministry of Agriculture for Integrated Pest Management on Tropical Crops, Institute of Environmental and Plant Protection, Chinese Academy of Tropical Agricultural Sciences, Haikou, Hainan, China.
FEMS Microbiol Lett. 2013 Dec;349(2):127-34. doi: 10.1111/1574-6968.12305. Epub 2013 Nov 6.
Fusarium wilt caused by Fusarium oxysporum f. sp. niveum (Fon) is one of the major limiting factors for watermelon production worldwide. Rapid and accurate detection of the causal pathogen is the cornerstone of integrated disease management. In this paper, a real-time fluorescence loop-mediated isothermal amplification (RealAmp) assay was developed for the rapid and quantitative detection of Fon in soil. Positive products were amplified only from Fon isolates and not from any other species or formae speciales of F. oxysporum tested, showing a high specificity of the primer sets. The detection limit of the RealAmp assay was 1.2 pg μL(-1) genomic DNA or 10(3) spores g(-1) of artificially inoculated soil, whereas real-time PCR could detect as low as 12 fg μL(-1) or 10(2) spores g(-1). The RealAmp assay was further applied to detect eight artificially inoculated and 85 field soil samples. No significant differences were found between the results tested by the RealAmp and real-time PCR assays. The RealAmp assay is a simple, rapid and effective technique for the quantitative detection and monitoring of Fon in soil under natural conditions.
由尖孢镰刀菌绵腐专化型（Fon）引起的枯萎病是全球西瓜生产的主要限制因素之一。快速准确地检测致病病原体是综合疾病管理的基石。本文开发了一种实时荧光环介导等温扩增（RealAmp）检测方法，用于快速定量检测土壤中的 Fon。阳性产物仅从 Fon 分离物中扩增，而不从测试的任何其他 F. oxysporum 种或专化型中扩增，显示出引物对的高特异性。RealAmp 检测方法的检测限为 1.2 pg μL(-1) 基因组 DNA 或 10(3) 个接种土壤中的孢子 g(-1)，而实时 PCR 可以检测低至 12 fg μL(-1) 或 10(2) 个孢子 g(-1)。该 RealAmp 检测方法进一步应用于检测 8 个人工接种和 85 个田间土壤样品。通过 RealAmp 和实时 PCR 检测方法检测到的结果之间没有发现显著差异。RealAmp 检测方法是一种简单、快速、有效的技术，可用于定量检测和监测自然条件下土壤中的 Fon。
