The rapid detection and species-level differentiation of bacterial pathogens facilitates antibiotic stewardship and improves disease management. Here, we develop a rapid bacteriophage-based diagnostic assay to detect the most prevalent pathogens causing urinary tract infections: Escherichia coli, Enterococcus spp., and Klebsiella spp. For each uropathogen, two virulent phages were genetically engineered to express a nanoluciferase reporter gene upon host infection. Using 206 patient urine samples, reporter phage-induced bioluminescence was quantified to identify bacteriuria and the assay was benchmarked against conventional urinalysis. Overall, E. coli, Enterococcus spp., and Klebsiella spp. were each detected with high sensitivity (68%, 78%, 87%), specificity (99%, 99%, 99%), and accuracy (90%, 94%, 98%) at a resolution of ≥10³ CFU/ml within 5 h. We further demonstrate how bioluminescence in urine can be used to predict phage antibacterial activity, demonstrating the future potential of reporter phages as companion diagnostics that guide patient-phage matching prior to therapeutic phage application.

细菌病原体的快速检测和物种水平区分有助于抗生素管理并改善疾病管理。在这里，我们开发了一种基于噬菌体的快速诊断方法，以检测引起尿路感染的最常见病原体：大肠杆菌、肠球菌属和克雷伯氏菌属。对于每种尿路病原体，两个剧毒噬菌体经过基因改造，在宿主感染时表达纳米荧光素酶报告基因。使用 206 份患者尿液样本，对报告噬菌体诱导的生物发光进行定量，以识别菌尿，并将该测定方法与传统尿液分析进行比较。总体而言，大肠杆菌、肠球菌属和克雷伯氏菌属。均以≥10 3 CFU/的分辨率检测到高灵敏度（68%、78%、87%）、特异性（99%、99%、99%）和准确度（90%、94%、98 ^%  ） 5 小时内。我们进一步证明了尿液中的生物发光如何用于预测噬菌体抗菌活性，证明了报告噬菌体作为伴随诊断的未来潜力，可在治疗性噬菌体应用之前指导患者与噬菌体匹配。