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Focusing on the research and development and application of isothermal nucleic acid amplification technology

重组酶介导的核酸等温扩增荧光法快速检测日本血吸虫感染性钉螺-江苏血防所

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[Abstract] Objective: To establish a recombinase-aided amplification (RAA) assay combined with fluorescence detection for the rapid identification of Schistosoma japonicum-infected Oncomelania snails, and to optimize the sample preparation method for these snails. Methods: Snail samples were divided into three groups, each containing seven subgroups. Six subgroups consisted of mixtures comprising 1, 2, 3, 4, 5, or 10 infected snails embedded within 50 uninfected snails, while one subgroup included a mixture of 1 infected snail mixed among 100 negative snails. Each group was processed using three different methods: the crushed-shell nucleic acid extraction kit method, the crushed-shell crude nucleic acid extraction method, and the direct-crushed-shell crude nucleic acid extraction method. Both fluorescence-based RAA and PCR assays were then performed on these samples, and the results were compared. Results: A fluorescence-based RAA assay was successfully established, operating at a temperature of 39°C and capable of detecting S. japonicum-infected snails within 30 minutes. When the crushed-shell nucleic acid extraction kit method was used, the fluorescence RAA assay could detect as few as 1 infected snail mixed among 100 negative snails, whereas the PCR assay required at least 1 infected snail mixed among 50 negative snails. In contrast, the crushed-shell crude nucleic acid extraction method allowed detection of 1 infected snail mixed among 100 negative snails via fluorescence RAA and 3 infected snails mixed among 50 negative snails via PCR. Finally, when the direct-crushed-shell crude nucleic acid extraction method was applied, the fluorescence RAA assay could detect up to 10 infected snails mixed among 50 negative snails, while the PCR assay identified 10 infected snails even in the presence of 50 negative snails. Conclusion: A rapid fluorescence RAA assay has been successfully developed for detecting S. japonicum-infected Oncomelania snails in population samples, offering advantages such as speed, high sensitivity, and ease of operation. Among the three sample preparation methods tested, the crushed-shell crude nucleic acid extraction method emerged as the optimal approach for processing snail samples.

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重组酶介导的核酸等温扩增荧...血吸虫感染性钉螺的效能评价_叶钰滢

重组酶介导的华支睾吸虫特异性核酸等温扩增方法的建立及初步评价-江苏血防所

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重组酶介导的核酸等温扩增荧...血吸虫感染性钉螺的效能评价_叶钰滢

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重组酶介导的华支睾吸虫特异性核酸等温扩增方法的建立及初步评价-江苏血防所