Analyzing Persister Physiology with Fluorescence-Activated Cell Sorting.

TitleAnalyzing Persister Physiology with Fluorescence-Activated Cell Sorting.
Publication TypeJournal Article
Year of Publication2016
AuthorsOrman, MA, Henry, TC, DeCoste, CJ, Brynildsen, MP
JournalMethods Mol Biol
Volume1333
Pagination83-100
Date Published2016
ISSN1940-6029
KeywordsAnti-Bacterial Agents, Drug Resistance, Multiple, Bacterial, Escherichia coli, Flow Cytometry, Humans, Microbial Sensitivity Tests
Abstract

<p>Bacterial persisters are phenotypic variants that exhibit an impressive ability to tolerate antibiotics. Persisters are hypothesized to cause relapse infections, and therefore, understanding their physiology may lead to novel therapeutics to treat recalcitrant infections. However, persisters have yet to be isolated due to their low abundance, transient nature, and similarity to the more highly abundant viable but non-culturable cells (VBNCs), resulting in limited knowledge of their phenotypic state. This technical hurdle has been addressed through the use of fluorescence-activated cell sorting (FACS) and quantification of persister levels in the resulting sorted fractions. These assays provide persister phenotype distributions, which can be compared to the phenotype distributions of the entire population, and can also be used to examine persister heterogeneity. Here, we describe two detailed protocols for analysis of persister physiology with FACS. One protocol assays the metabolic state of persisters using a fluorescent metabolic stain, whereas the other assays the growth state of persisters with use of a fluorescent protein.</p>

DOI10.1007/978-1-4939-2854-5_8
Alternate JournalMethods Mol. Biol.
PubMed ID26468102
PubMed Central IDPMC4908830
Grant ListR21 AI105342 / AI / NIAID NIH HHS / United States
R21AI105342 / AI / NIAID NIH HHS / United States