Tjandra Kristel C, Ram-Mohan Nikhil, Abe Ryuichiro, Wang Tza-Huei, Yang Samuel
Department of Emergency Medicine, Stanford University School of Medicine, Palo Alto, CA, USA.
Departments of Mechanical Engineering and Biomedical Engineering, The Johns Hopkins University, Baltimore, Maryland, USA.
bioRxiv. 2023 Mar 1:2023.03.01.530513. doi: 10.1101/2023.03.01.530513.
(NG) is an urgent threat to antimicrobial resistance (AMR) worldwide. NG has acquired rapid resistance to all previously recommended treatments leaving ceftriaxone monotherapy as the first and last line of therapy for uncomplicated NG. The ability to rapidly determine susceptibility, which is currently nonexistent for NG, has been proposed as a strategy to preserve ceftriaxone by using alternative treatments. Herein, we used a DNA-intercalating dye in combination with NG-specific primers/probes to generate qPCR cycle threshold (Ct) values at different concentrations of 2 NG-relevant antimicrobials. Our proof of concept dual-antimicrobial logistic regression model based on the differential Ct measurements achieved an AUC of 0.93 with a categorical agreement for susceptibility of 84.6%. When surveying the performance against each antimicrobial separately, the model predicted 90% and 75% susceptible and resistant strains respectively to ceftriaxone and 66.7% and 83.3% susceptible and resistant strains respectively to ciprofloxacin. We further validated the model against the individual replicates and determined the accuracy of the model in classifying susceptibility agnostic of the inoculum size. We demonstrated a novel PCR-based approach to determine phenotypic ciprofloxacin and ceftriaxone susceptibility information for NG with reasonable accuracy in under 30 min, a significant improvement compared to the conventional method which takes 3 days.
淋球菌（NG）对全球的抗菌药物耐药性（AMR）构成了紧迫威胁。NG已对所有先前推荐的治疗方法迅速产生耐药性，使得头孢曲松单药治疗成为无并发症NG感染的一线和最后一线治疗方法。目前不存在快速确定NG药敏性的方法，有人提出通过使用替代治疗方法来保留头孢曲松，以此作为一种策略。在此，我们使用一种DNA嵌入染料与NG特异性引物/探针相结合，在两种与NG相关的抗菌药物的不同浓度下生成定量聚合酶链反应（qPCR）循环阈值（Ct）值。我们基于差异Ct测量的概念验证双抗菌药物逻辑回归模型的曲线下面积（AUC）为0.93，药敏性分类一致性为84.6%。当分别检测针对每种抗菌药物的性能时，该模型对头孢曲松的敏感菌株和耐药菌株的预测分别为90%和75%，对环丙沙星的敏感菌株和耐药菌株的预测分别为66.7%和83.3%。我们进一步针对单个重复样本验证了该模型，并确定了该模型在不考虑接种量的情况下对药敏性进行分类的准确性。我们展示了一种基于PCR的新方法，可在30分钟内以合理的准确性确定NG对环丙沙星和头孢曲松的表型药敏性信息，与需要3天的传统方法相比有显著改进。
