Abstract：
Sepsis is a major health concern globally, and identification of the causative organism usually takes several days. Furthermore, molecular amplification using whole blood from patients with sepsis remains challenging because of primer cross-reactivity with human DNA, which can delay appropriate clinical intervention. To address these concerns, we designed primers that could reduce cross-reactivity. By evaluating these primers against human DNA, we confirmed that the cross-reactivity observed with conventional primers was notably absent. In silico PCR further demonstrated the specificity and efficiency of the designed primers across 23 bacterial species that are often associated with sepsis. When tested using blood samples from sepsis patients, the designed primers showed moderate sensitivity and high specificity. Surprisingly, our method identified bacteria even in samples that were detected at other sites but tested negative using conventional blood culture methods. Although we identified some challenges, such as contamination with Acetobacter aceti due to the saponin pretreatment of samples, the developed method demonstrates remarkable potential for rapid identification of the causative organisms of sepsis and provides a new avenue for diagnosis in clinical practice.
摘要：
脓毒症是全球主要的健康问题，和病原体的鉴定通常需要几天。此外，由于引物与人类DNA的交叉反应，使用脓毒症患者的全血进行分子扩增仍然具有挑战性，这可能会延迟适当的临床干预。为了解决这些问题，我们设计了可以降低交叉反应性的引物.通过评估这些针对人类DNA的引物，我们证实与常规引物观察到的交叉反应性明显缺失.计算机PCR进一步证明了设计的引物对23种细菌的特异性和效率，这些细菌通常与败血症有关。当使用败血症患者的血液样本进行测试时，设计的引物灵敏度中等，特异性高。令人惊讶的是,我们的方法甚至在其他部位检测到但使用常规血培养方法检测为阴性的样本中也能识别细菌.尽管我们发现了一些挑战，如由于样品的皂苷预处理而导致的醋酸醋杆菌污染，所开发的方法显示出快速鉴定脓毒症致病生物的巨大潜力，并为临床实践中的诊断提供了新的途径。
