Poultry Institute, Chinese Academy of Agricultural Sciences, Yangzhou, China.
Key Laboratory for Poultry Genetics and Breeding of Jiangsu Province, Jiangsu Institute of Poultry Sciences, Yangzhou, China.
Microbiol Spectr. 2024 Oct 3;12(10):e0127124. doi: 10.1128/spectrum.01271-24. Epub 2024 Aug 27.
serovar Typhimurium (ST) is a predominant serovar causing foodborne illnesses worldwide. Traditional detection methods often face challenges, including the need for specialized equipment, skilled operators, and lengthy procedures. To address these limitations, we developed a rapid, sensitive, and specific ST detection method by integrating loop-mediated isothermal amplification (LAMP) with the clustered regularly interspaced short palindromic repeats and associated protein 12b (CRISPR/Cas12b) system, all within a single tube. Our results indicate that the LAMP-CRISPR/Cas12b reaction can be completed isothermally in under 1 h without requiring specialized instruments. The platform's limit of detection (LoD) is 12.5 copies per reaction. Additionally, the system demonstrated 100% inclusivity and exclusivity when tested against 30 reference strains, highlighting its specificity. In practical applications, the LoDs for ST in pure nucleic acid and contaminated fecal samples were 2.32 and 23.2 CFU/mL, respectively, with higher sensitivity observed in pure nucleic acid samples. Overall, our findings underscore the potential of the one-tube LAMP-CRISPR/Cas12b platform as a rapid, sensitive, and specific tool for ST detection, particularly in resource-limited settings.
Here, we have provided a novel one-step method for Typhimurium detection in one pot by integrating the LAMP assay with the CRISPR/Cas12b system, offering significant advantages in terms of simplicity, speed, and accuracy.
鼠伤寒沙门氏菌（ST）是一种主要的血清型，导致全球范围内的食源性疾病。传统的检测方法常常面临挑战，包括需要专用设备、熟练的操作人员和冗长的程序。为了解决这些限制，我们开发了一种快速、敏感和特异的 ST 检测方法，该方法将环介导等温扩增（LAMP）与成簇规律间隔短回文重复序列和相关蛋白 12b（CRISPR/Cas12b）系统集成在一个单一的管内。我们的结果表明，LAMP-CRISPR/Cas12b 反应可以在不到 1 小时的时间内恒温完成，而不需要专用仪器。该平台的检测限（LoD）为每个反应 12.5 个拷贝。此外，该系统在测试 30 个参考菌株时表现出 100%的包容性和排他性，突出了其特异性。在实际应用中，ST 在纯核酸和污染粪便样本中的 LoD 分别为 2.32 和 23.2 CFU/mL，在纯核酸样本中观察到更高的灵敏度。总的来说，我们的研究结果强调了单管 LAMP-CRISPR/Cas12b 平台作为一种快速、敏感和特异的 ST 检测工具的潜力，特别是在资源有限的环境中。
在这里，我们通过将 LAMP 测定法与 CRISPR/Cas12b 系统整合在一个管中，提供了一种新颖的一步法用于在一个管中检测鼠伤寒沙门氏菌，在简单性、速度和准确性方面具有显著优势。
