脓毒血症是一种严重威胁生命的感染，精准、快速的病原学诊断可帮助临床医师优化抗菌药物的使用。目前，基于病原菌培养的方法仍是脓毒血症病原学诊断的主要手段，但具有耗时长、灵敏度低等不可忽视的缺点。近年来出现了一些不依赖培养的病原学诊断方法，其中基于聚合酶链反应(polymerase chain reaction，PCR)的方法已发展较为成熟。但PCR只能检测已知的特定病原体，临床定量PCR仅用于检测病毒及少数细菌，脓毒血症中的病原体PCR多仅为定性检测。目前，二代测序技术不断成熟并用于临床，成为病原学诊断的有力手段。与血培养等传统病原学检测方法相比，其具有快速、非选择性、可定量或半定量分析的优点。现阶段二代测序仍存在公认判读标准缺乏、测序结果与治疗关系不明确、耐药基因检测困难等不足，亦缺乏较大规模的二代测序与传统诊断方法比较验证的研究结果，尚有待更高级的循证医学证据支持。

Sepsis is a life-threatening infection demanding accurate and fast diagnosis of pathogen for right antibiotics. In current practice, culture-based methods are still the standard approach of etiological diagnosis of sepsis. Practically, these methods have disadvantages including long turnaround time and low sensitivity. Although molecular assays, such as polymerase chain reaction (PCR)-based methods have been established for sepsis, the spectrum of the detection is limited to known pathogens listed on the panel. In the last few years, the next-generation sequencing (NGS) has been constantly improving and stepped into clinical practice, providing a powerful tool with advantages including short turnaround time, full spectrum and semiquantitative analysis. Although NGS has clear advantages compared with culture-based methods, we are facing the following challenges: few acknowledged criteria of sequencing result explanation have been established; the relationship between sequencing results and treatment effects is undetermined; and the drug resistance gene detection is usually with low sensitivity. Besides, large-scale study comparing NGS with traditional etiological diagnosis methods is missing. Thus, NGS still needs high-level evidence to support its clinical application.