Development of Persister-FACSeq: a method to massively parallelize quantification of persister physiology and its heterogeneity
Development of Persister-FACSeq: a method to massively parallelize quantification of persister physiology and its heterogeneity
复制标题
DOI：
10.1038/srep25100
复制
发表时间：
2016-05-04
影响因子：
4.6
通讯作者：
Brynildsen, Mark P.
中科院分区：
文献类型：
--
作者：
Henry, Theresa C.;Brynildsen, Mark P.
细菌持续存在被认为是慢性感染复发的基础。持久性生理学的知识将阐明治疗干预的途径;然而，这种知识仍然是难以捉摸的，因为持久性还没有从其他细胞类型分离到足够的纯度。这个技术障碍阻碍了理解持久性的进展。在这里，我们开发了Persister-FACSeq，这是一种使用荧光激活细胞分选，抗生素耐受性测定和下一代测序来询问持续生理学及其异质性的方法。作为概念验证，我们使用Persister-FACSeq在报告基因库中研究非生长大肠杆菌中的基因表达分布，发现氧氟沙星的持久性与非生长细胞合成蛋白质的能力呈负相关。由于Persister-FACSeq可用于研究任何环境中任何抗生素对任何细菌的持久性，因此我们预计它将产生对这种有害表型的前所未有的知识。
Bacterial persisters are thought to underlie the relapse of chronic infections. Knowledge of persister physiology would illuminate avenues for therapeutic intervention; however, such knowledge has remained elusive because persisters have yet to be segregated from other cell types to sufficient purity. This technical hurdle has stymied progress toward understanding persistence. Here we developed Persister-FACSeq, which is a method that uses fluorescence-activated cell sorting, antibiotic tolerance assays, and next generation sequencing to interrogate persister physiology and its heterogeneity. As a proof-of-concept, we used Persister-FACSeq on a library of reporters to study gene expression distributions in non-growing Escherichia coli, and found that persistence to ofloxacin is inversely correlated with the capacity of non-growing cells to synthesize protein. Since Persister-FACSeq can be applied to study persistence to any antibiotic in any environment for any bacteria that can harbor a fluorescent reporter, we anticipate that it will yield unprecedented knowledge of this detrimental phenotype.