Rice false smut spores (RFSS), which are airborne spores caused by Ustilaginoidea virens (U. virens), not only cause severe yield loss and grain quality reduction, but also produce toxins that are harmful to humans and animals. Nucleic acid detection has become the main method for RFSS monitoring due to its high specificity and sensitivity. However, nucleic acid detection requires multiple steps of spore collection, DNA extraction, nucleic acid amplification and detection, which has a high demand for personnel and is hard to link with other intelligent equipment to achieve automation. Microfluidic chip has become an important approach for integrated detection of pathogens owning to miniaturization and integration in recent years. Yet there is a lack of portable methods that integrate the collection of airborne fungal spores and nucleic acid detection. Because RFSS have thick cell walls and require liquid nitrogen grinding to extract DNA, breaking the walls on-chip is difficult. Therefore, the realization of RFSS wall breaking on-chip is a major difficulty and also a very meaningful study. This study uses RFSS as the research object and provides a novel method of culturing RFSS on-chip to solve the problem of hard wall breaking, realizing the integrated detection of RFSS. The mycelium grown by RFSS germination could be easily broken to release DNA for on-chip detection, which eliminates the need for manual DNA extraction and resolves the issue of difficult wall breaking. This chip can collect RFSS based on the aerodynamic theory and achieve gas–liquid coupling through a simple microvalve structure. A micromixer is constructed to mix the liquid, and then accomplish detection quickly by recombinase polymerase amplification and lateral flow dipsticks (RPA-LFD). The detection sensitivity of this method is 1 × 10²–1 × 10⁵ CFU ml⁻¹. It can realize the “sample in and answer out” detection of RFSS due to its simple operation, independence from precision instruments, high sensitivity and specificity. The result shows that it can be used for the early detection of RFSS, has great application prospects and is expected to promote the development of on-site instant detection equipment.

中文翻译：

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基于微流控芯片的水稻假黑曲病孢子采集培养一体化核酸检测方法

水稻假黑穗病孢子 (RFSS)，是由黑曲病菌 (U. virens) 引起的空气传播的孢子)，不仅造成严重的减产和粮食品质下降，而且还会产生对人畜有害的毒素。核酸检测以其高特异性和敏感性成为RFSS监测的主要方法。但核酸检测需要孢子采集、DNA提取、核酸扩增检测等多个步骤，对人员要求高，且难以与其他智能设备联动实现自动化。近年来，微流控芯片以其小型化、集成化成为病原体综合检测的重要手段。然而，缺乏将空气中真菌孢子的收集和核酸检测结合起来的便携式方法。由于 RFSS 的细胞壁较厚，需要液氮研磨才能提取 DNA，打破芯片上的墙壁是困难的。因此，片上实现RFSS破壁是一大难点，也是一项非常有意义的研究。本研究以RFSS为研究对象，提供了一种片上培养RFSS的新方法，以解决硬壁破裂问题，实现RFSS的集成检测。RFSS发芽生长的菌丝体很容易破碎释放出DNA用于片上检测，无需人工提取DNA，解决了破壁难的问题。该芯片基于空气动力学原理采集RFSS，通过简单的微阀结构实现气液耦合。构建微混合器混合液体，然后通过重组酶聚合酶扩增和侧流试纸（RPA-LFD）快速完成检测。² –1 × 10 ^{5 个}菌落形成单位毫升^-1。它具有操作简单、不依赖精密仪器、灵敏度高、特异性强等特点，可实现RFSS的“进样出”检测。结果表明，可用于RFSS的早期检测，具有广阔的应用前景，有望推动现场即时检测设备的发展。