Department of Emergency Medicine, Stanford University, Stanford, CA, USA.
Department of Mathematical and Computational Science, Stanford University, Stanford, CA, USA.
J Antimicrob Chemother. 2020 Jul 1;75(7):1747-1755. doi: 10.1093/jac/dkaa078.
Traditional antimicrobial susceptibility testing (AST) is growth dependent and time-consuming. With rising rates of drug-resistant infections, a novel diagnostic method is critically needed that can rapidly reveal a pathogen's antimicrobial susceptibility to guide appropriate treatment. Recently, RNA sequencing has been identified as a powerful diagnostic tool to explore transcriptional gene expression and improve AST.
RNA sequencing was used to investigate the potential of RNA markers for rapid molecular AST using Klebsiella pneumoniae and ciprofloxacin as a model. Downstream bioinformatic analysis was applied for optimal marker selection. Further validation on 11 more isolates of K. pneumoniae was performed using quantitative real-time PCR.
From RNA sequencing, we identified RNA signatures that were induced or suppressed following exposure to ciprofloxacin. Significant shifts at the transcript level were observed as early as 10 min after antibiotic exposure. Lastly, we confirmed marker expression profiles with concordant MIC results from traditional culture-based AST and validated across 11 K. pneumoniae isolates. recA, coaA and metN transcripts harbour the most sensitive susceptibility information and were selected as our top markers.
Our results suggest that RNA signature is a promising approach to AST development, resulting in faster clinical diagnosis and treatment of infectious disease. This approach is potentially applicable in other models including other pathogens exposed to different classes of antibiotics.
传统的抗菌药物敏感性测试（AST）依赖于生长且耗时。随着耐药感染率的上升，迫切需要一种新的诊断方法，能够快速揭示病原体对抗菌药物的敏感性，从而指导进行适当的治疗。最近，RNA 测序已被确定为一种强大的诊断工具，可用于探索转录基因表达并改进 AST。
使用肺炎克雷伯菌和环丙沙星作为模型，通过 RNA 测序来研究 RNA 标志物在快速分子 AST 中的应用潜力。应用下游生物信息学分析进行最佳标志物选择。使用定量实时 PCR 对另外 11 株肺炎克雷伯菌进行了进一步验证。
从 RNA 测序中，我们确定了在接触环丙沙星后被诱导或抑制的 RNA 特征。在抗生素暴露后 10 分钟左右即可观察到转录水平的显著变化。最后，我们通过传统基于培养的 AST 确认了标记物表达谱与 MIC 结果一致，并在 11 株肺炎克雷伯菌中进行了验证。recA、coaA 和 metN 转录本具有最敏感的药敏信息，被选为我们的最佳标志物。
我们的研究结果表明，RNA 特征是一种很有前途的 AST 开发方法，可加快临床诊断和传染病治疗。这种方法可能适用于包括其他病原体暴露于不同类别的抗生素的其他模型。
