Rice root nematode, Hirschmanniella oryzae is an important plant-parasitic nematode inflicting substantial destruction on economically important agricultural crops worldwide and invading an extensive range of plant hosts, including aquatic plants. However, morphological identification of the nematode has encountered limitations. This study developed a loop-mediated isothermal amplification (LAMP) assay targeting the genetic sequence of H. oryzae 28S ribosomal RNA gene as an alternative method. The LAMP assay demonstrated remarkable speed that the outcome could be achieved within 40 min at an isothermal temperature, 65 °C. The assay could interpret the fluorescence (SYBR safe) and color (SYBR green I) in a pre-reaction mixture, ensuring reliability and minimizing contamination risk during post-amplification. Our assay has high specificity and enables accurate differentiation from other nematode species in diverse aquatic plant samples. The sensitivity was higher than PCR and real-time PCR as low as 4.61 fg/μL. The development of fluorescent and colorimetric closed tube LAMP assay highlights it as a valuable tool that is a simplified, sensitive and rapid diagnostic method for detecting aquatic pest nematode, H. oryzae, at point-of-service level. The implementation of this method holds promise for future applications in plant quarantine departments and offers a significant advancement in the diagnosis of infected plants.

中文翻译：

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开发闭管环介导的等温扩增 （LAMP） 测定法以检测出口水生植物中的米巨人菌（Tylenchida：Pratylenchidae）


水稻根线虫，米巨曼氏菌是一种重要的植物寄生线虫，对全球具有重要经济价值的农作物造成重大破坏，并侵入包括水生植物在内的广泛植物宿主。然而，线虫的形态学鉴定遇到了局限性。本研究开发了一种靶向米嗜血杆菌 28S 核糖体 RNA 基因基因序列的环介导等温扩增 （LAMP） 测定法作为替代方法。LAMP 分析显示出显著的速度，在 65 °C 等温温度下可在 40 分钟内获得结果。 该分析可以解释反应前混合物中的荧光 （SYBR 安全） 和颜色 （SYBR green I），确保可靠性并最大限度地降低后扩增过程中的污染风险。我们的检测具有高度特异性，能够准确区分不同水生植物样品中的其他线虫物种。灵敏度高于 PCR，实时 PCR 低至 4.61 fg/μL。荧光和比色闭管 LAMP 测定法的发展突出表明它是一种有价值的工具，是一种简化、灵敏和快速的诊断方法，用于在服务点检测水生害虫线虫 H. oryzae。该方法的实施为植物检疫部门的未来应用带来了希望，并为感染植物的诊断提供了重大进展。