基于新型荧光纳米机器的双耐药菌同时诊断及其耐药性检测研究

细菌感染是导致人类疾病和死亡的主要原因之一且耐药问题日益严峻。目前已报道的细菌传感方法大多只能检测单一靶标，临床上对细菌耐药性检测耗时费力，亟需发展细菌感染及耐药性高效诊断技术。DNA纳米材料因设计灵活、精准可控、可功能化、生物相容性高而具有重要应用潜力。本项目在申请人前期研究基础上，拟设计荧光标记DNA组合且串联形成链置换通路，构建新型DNA纳米机器，实现目标诱导荧光级联放大和顺序释放功能DNA；并结合双致病菌特异性适配体，赋予适配体靶向和开关双功能，构建免DNA提取、无酶介导的荧光纳米传感器，实现在单一均相中同时诊断两种临床细菌感染的目标；利用适配体只能靶向结合活菌的优势，通过检测抗生素处理前后细菌诱导的荧光变化，实现在单一均相中同时检测两种耐药菌药敏性的目标。本项目为发展功能化DNA纳米机器开辟了新思路，为临床快速诊断致病菌及耐药性提供了新方法，对预防和控制细菌感染具有重要临床意义。

Bacterial infection is one of the main causes of human disease and death, and the problem of drug resistance is becoming increasingly severe. Most of the bacterial sensing technologies reported so far can only detect a single target. Clinically, the detection of bacterial drug resistance is time-consuming and laborious, and it is urgent to develop high-efficiency diagnostic technologies for bacterial infection and drug resistance. DNA nanomaterials have important application potential due to their flexible design, precise controllability, functionalization, and high biocompatibility. Based on the applicant's preliminary research, this project plans to design a combination of fluorescently labeled DNA and connect them in series to form a strand replacement pathway, construct a new type of DNA nanomachine, to achieve target-induced fluorescence cascade amplification and release of functional DNA in order. Combining with dual pathogen-specific aptamers, enabling the aptamers with dual functions of targeting and switching, constructing a DNA extraction-free and enzyme-free fluorescent nanosensor, realizing the goal of simultaneous diagnosis of two clinical bacterial infections in a single homogeneous phase. Aptamers can only precipitate and bind live bacteria, by taking advantage of it, and detecting the fluorescence changes induced by bacteria before and after antibiotic treatment, the goal of simultaneously detecting the susceptibility of two drug-resistant bacteria in a single homogeneous phase is achieved. This project opens up new ideas for developing functional DNA nanomachines, provides a new method for clinical rapid diagnosis of pathogenic bacteria and drug resistance, and has important clinical significance for the prevention and control of bacterial infections.