The tremor disease (TD) caused by Spiroplasma eriocheiris is the most destructive disease of the Chinese mitten crab, Eriocheir sinensis. This study attempts to construct Multienzyme Isothermal Rapid Amplification (MIRA), a quick and simple nucleic acid amplification method that operates at room temperature. Based on the gene sequences of S. eriocheiris, appropriate amplification primers were constructed and screened in this investigation. Both the relevant specific probe and the chosen specific amplification primers were designed and labeled. The MIRA and MIRA-LFD reaction conditions were then optimized. The result showed MIRA and MIRA-FFD could identify S. eriocheiris at 37 °C in 30 min and 15 min, respectively. To investigate the specificity of MIRA and MIRA-LFD, three Gram-negative bacteria (Bacillus subtilis, Bacillus thuringiensis, and Staphylococcus aureus), three Gram-positive bacteria (Escherichia coli, Aeromonas hydrophila, and Salmonella typhimurium) and S. eriocheiris were selected. The result showed MIRA and MIRA-LFD were highly specific to S. eriocheiris and did not react with other six pathogens. The sensitivities of PCR, MIRA, and MIRA-LFD were then evaluated. The result showed the detection limit of PCR is 1 ng/L whereas the detection limit of MIRA and MIRA-LFD is 10 pg/L. Finally, the established MIRA and MIRA-LFD detection methods had the advantages of being quick, sensitive, and specific for S. eriocheiris detection, as well as not requiring any specialized equipment.

由绒毛螺原体引起的震颤病（TD）是中华绒螯蟹最具破坏性的疾病。本研究试图构建多酶等温快速扩增（MIRA），一种在室温下操作的快速、简单的核酸扩增方法。本研究根据马蹄草基因序列，构建并筛选了合适的扩增引物。设计并标记相关的特异性探针和所选的特异性扩增引物。然后优化 MIRA 和 MIRA-LFD 反应条件。结果表明，MIRA 和 MIRA-FFD 在 37 °C 下分别可在 30 分钟和 15 分钟内识别马蹄草。为了研究 MIRA 和 MIRA-LFD 的特异性，选择了三种革兰氏阴性菌（枯草芽孢杆菌、苏云金芽孢杆菌和金黄色葡萄球菌） 、三种革兰氏阳性菌（大肠杆菌、嗜水气单胞菌和鼠伤寒沙门氏菌）和绒毛沙门氏菌。 。结果表明，MIRA和MIRA-LFD对绒毛链球菌具有高度特异性，并且与其他六种病原体不发生反应。然后评估 PCR、MIRA 和 MIRA-LFD 的敏感性。结果显示PCR的检出限为1ng/L，MIRA和MIRA-LFD的检出限为10pg/L。最后，所建立的MIRA和MIRA-LFD检测方法具有快速、灵敏、特异性强、不需要任何专门设备的优点。