细菌异质性耐药是指引起感染的细菌群体中存在对特定抗生素不同程度耐药性的现象，其在临床感染病例中难以检测，常导致抗菌治疗方案的失败，贻误病情。现有的检测方法均基于体外纯培养菌株的扩增分析，不能真实反映患者体内感染菌株群体的异质性耐药特征。本项目拟在前期对单细胞拉曼技术分析细菌菌株的研究基础上，开发不依赖于培养扩增的异质性耐药原位分析方法，并结合单细胞分选和高通量测序技术，研究临床样品细菌群体的异质性耐药机制。项目采用逐层深入的研究方案，分别对不同MIC菌株混合样品、已知异质性耐药菌株、尿路感染标本和反复感染抗菌效果不良患者尿液标本进行单细胞拉曼检测，建立单细胞拉曼原位检测临床标本异质性耐药的技术，并通过对异质性耐药细胞亚群的分选和扩增，结合高通量测序技术，揭示患者体内异质性耐药的机制。本项目将建立细菌异质性耐药的单细胞拉曼原位检测和机制分析方法，为临床开展快速准确的抗菌治疗提供检验依据。

Bacterial heteroresistance describes a phenomenon where bacterial subpopulations exhibit a range of susceptibility to a particular antibiotic. Detection of bacterial heteroresistance in clinical infection is difficult and this may lead to antimicrobial treatment failure. Current detection methods depend on the in vitro amplification of pure culture strains and cant not represent the in vivo status of bacterial population infected patients. This study aims to develop an in situ culture-independent method on the basis of our previous results on bacterial analysis by single cell raman spectroscopy. Single cell sorting and high throughput sequencing are also applied to investigate the mechanism of resistance in the bacterial subpopulations. A step-by-step experimental procedure will be used to analyze the mix bacterial culture with different MICs, known strains with heteroresistance, urinary tract infection samples, and urine samples from repeated infection without good antimicrobial effects, by single cell raman spectroscopy. In situ detection of bacterial heteroresistance in clinical samples will be established, and the in vivo resistance mechanism will be revealed by further cell sorting and sequencing. This study will develop the methods for in situ detection of bacterial heteroresistance, which provides experimental evidence for clinical antimicrobial treatment.