Single-Cell Microbiology: Tools, Technologies, and Applications

Abstract

The field of microbiology has traditionally been concerned with and focused on studies at the population level. Information on how cells respond to their environment, interact with each other, or undergo complex processes such as cellular differentiation or gene expression has been obtained mostly by inference from population-level data. Individual microorganisms, even those in supposedly “clonal” populations, may differ widely from each other in terms of their genetic composition, physiology, biochemistry, or behavior. This genetic and phenotypic heterogeneity has important practical consequences for a number of human interests, including antibiotic or biocide resistance, the productivity and stability of industrial fermentations, the efficacy of food preservatives, and the potential of pathogens to cause disease. New appreciation of the importance of cellular heterogeneity, coupled with recent advances in technology, has driven the development of new tools and techniques for the study of individual microbial cells. Because observations made at the single-cell level are not subject to the “averaging” effects characteristic of bulk-phase, population-level methods, they offer the unique capacity to observe discrete microbiological phenomena unavailable using traditional approaches. As a result, scientists have been able to characterize microorganisms, their activities, and their interactions at unprecedented levels of detail

Deconvolution of gene expression noise into physical dynamics of cognate promoters

“…In these engineered bacteria, 54 the two components of the regulatory device under study 55 (Ps→xylS/Pm→GFP) are thereby designed adjacent to where Ps-xylS and Pm-GFP are separated. The data 10 shown in Supplemental Figure S3 indicated that both 11 strains are virtually indistinguishable as no significant dif-12 ferences were noticed in either physical quality. Once this 13 was clarified, we repeated with strain P. putida KT-BGS 14 the same induction experiment with 3MBz that was done 15 previously with the reference P. putida mt-2-Pm.…”

“…10 However, the range and intensity of expression noise of 11 given promoters appears in some cases as an adaptive 12 trait that frames the dynamic properties of promoter acti-13 vation [7][8][9]. The onset of single-cell technologies [10][11][12] 14 has shed some light on the various mechanisms behind 15 noise generation.…”

“…10 However, the range and intensity of expression noise of 11 given promoters appears in some cases as an adaptive 12 trait that frames the dynamic properties of promoter acti-13 vation [7][8][9]. The onset of single-cell technologies [10][11][12] 14 has shed some light on the various mechanisms behind 15 noise generation. A major source of noise in virtually 16 every prokaryotic promoter is the so-called bursting ef-17 fect [13,14], a pulse-like activity that largely results from 18 discontinuous topological changes of DNA caused by the 19 progression of RNA polymerase (RNAP) through tran-20 scribed DNA [15,16].…”

Coupling FACS and Genomic Methods for the Characterization of Uncultivated Symbionts

“…Other methods such as serial dilution (Zhang, et al, 2006), micromanipulation (Kvist, et al, 2007;Hongoh, et al, 2 2008), laser capture microdissection (Navin, et al, 2011), Raman tweezers (Brehm-Stecher & Johnson, 2004;Huang, et al, 2009) and microfluidics (Marcy, et al, 2007;Blainey, et al, 2011) have also been used effectively to separate cells prior to genetic or genomic analysis. However, for characterizing uncultivated symbionts these methods fall short of FACS in a number of ways.…”

Methods for Whole Cell Detection of Microorganisms

“…Although the capacity to detect cell-specific nucleic acids without the need for culture may be considered an advantage of such acellular techniques, it is not without cost. The individual cell is the "...fundamental unit of biological organization..." (16,17). Additional layers of information are therefore intrinsically linked to the "granular" or corpuscular nature of microbes.…”