Ultra-Rapid Drug Susceptibility Testing for Klebsiella pneumoniae Clinical Isolates in 60 Min by SYBR Green I/Propidium Iodide Viability Assay.
Ultra-Rapid Drug Susceptibility Testing for Klebsiella pneumoniae Clinical Isolates in 60 Min by SYBR Green I/Propidium Iodide Viability Assay.
复制标题
通过 SYBR Green I/碘化丙啶活力测定在 60 分钟内对临床分离株进行超快速药物敏感性测试
DOI：
10.3389/fmicb.2021.694522
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发表时间：
2021
影响因子：
5.2
通讯作者：
Zhang Y
中科院分区：
文献类型：
--
作者：
Chen J;Wang X;Wang S;Chen C;Zhang W;Zhang Y
我们的目的是使用 89 个肺炎克雷伯菌临床分离株，通过 SYBR Green I/PI (SG-PI) 方法优化和验证药物敏感性试验 (DST)，与传统 DST 方法相比，用于治疗这种细菌感染的三种最重要的抗生素（包括亚胺培南、头孢美唑和庆大霉素）。通过使用 SYBR Green I 和 PI 染料染色，分别与活细胞和死细胞或膜受损细胞的百分比线性相关的绿色荧光和红色荧光用于产生两条标准曲线，以计算每个对数期和稳定期培养物的相对细胞膜不透率。稳定期肺炎克雷伯菌细胞用于亚胺培南和头孢美唑SG-PI DST测定，而对数期细胞用于庆大霉素测定。采用常规肉汤微量稀释法作为药敏试验的金标准进行比较。数据显示，不同浓度的亚胺培南、头孢美唑、庆大霉素抗生素治疗30~60 min后，耐药肺炎克雷伯菌的存活细胞数显着高于敏感菌株，其中耐药临床菌株的平均相对膜不透率分别为88.5%、92.5%和103.8%，临床菌株的平均相对膜不透率为9.1%、49.3%和103.8%。敏感菌株分别为71.5%。总体而言，超快速SG-PI方法与传统最低抑菌浓度测定法诊断亚胺培南、头孢美唑和庆大霉素耐药性的总一致性较高，分别为96.6%（86/89）、95.4%（83/87）和95.5%（85/89）。我们证明，我们的新型 SG-PI 测定可以在 60-90 分钟内以超快速的方式准确、稳定地检测肺炎克雷伯菌临床分离株对不同抗生素的耐药性。
We aimed to optimize and validate the drug susceptibility test (DST) assay by SYBR Green I/PI (SG-PI) method using a panel of 89 Klebsiella pneumoniae clinical isolates in comparison with the conventional DST method to three most important antibiotics used for treatment of this bacterial infection, including imipenem, cefmetazole, and gentamicin. By staining with SYBR Green I and PI dyes, green fluorescence and red fluorescence, which linearly correlated with the percentages of live and dead or membrane damaged cells, respectively, were used to produce two standard curves to calculate the relative cell membrane impermeable rates for each log and stationary phase cultures. Stationary phase K. pneumoniae cells were used in imipenem and cefmetazole SG-PI DST assay whereas log phase cells were used in the gentamicin assay. The conventional broth microdilution method was used as a gold standard for DST for comparison. Data showed that after antibiotic treatment for 30–60 min, the antibiotic-resistant K. pneumoniae strains had significantly higher numbers of surviving cells than the susceptible strains at different concentrations of imipenem, cefmetazole, and gentamicin, where the average relative membrane impermeable rates were 88.5, 92.5, and 103.8% for resistant clinical strains, respectively, and 9.1, 49.3, and 71.5% for susceptible strains, respectively. Overall, the total concordances between the ultra-rapid SG-PI method and conventional minimal inhibitory concentration assay in diagnosing imipenem, cefmetazole and gentamicin resistance were high and were 96.6% (86/89), 95.4% (83/87), and 95.5% (85/89), respectively. We demonstrate that our novel SG-PI assay can accurately and stably detect resistance to different antibiotics in clinical isolates of K. pneumoniae in an ultra-fast manner in 60–90 min.
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作者：
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通讯作者：
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