Measuring bacterial adaptation dynamics at the single-cell level using a microfluidic chemostat and time-lapse fluorescence microscopy

The Analyst ◽  
2014 ◽  
Vol 139 (20) ◽  
pp. 5254-5262 ◽  
Author(s):  
Zhicheng Long ◽  
Anne Olliver ◽  
Elisa Brambilla ◽  
Bianca Sclavi ◽  
Marco Cosentino Lagomarsino ◽  
...  

We grewE. coliin a microfluidic chemostat and monitored the dynamics of cell dimensions and reporter GFP expression in individual cells during nutritional upshift or downshift.

2013 ◽  
Vol 79 (18) ◽  
pp. 5643-5651 ◽  
Author(s):  
Robyn T. Eijlander ◽  
Oscar P. Kuipers

ABSTRACTSingle-cell methods are a powerful application in microbial research to study the molecular mechanism underlying phenotypic heterogeneity and cell-to-cell variability. Here, we describe the optimization and application of single-cell time-lapse fluorescence microscopy for the food spoilage bacteriumBacillus cereusspecifically. This technique is useful to study cellular development and adaptation, gene expression, protein localization, protein mobility, and cell-to-cell communication over time at the single-cell level. By adjusting existing protocols, we have enabled the visualization of growth and development of singleB. cereuscells within a microcolony over time. Additionally, several different fluorescent reporter proteins were tested in order to select the most suitable green fluorescent protein (GFP) and red fluorescent protein (RFP) candidates for visualization of growth stage- and cell compartment-specific gene expression inB. cereus. With a case study concerningcotDexpression during sporulation, we demonstrate the applicability of time-lapse fluorescence microscopy. It enables the assessment of gene expression levels, dynamics, and heterogeneity at the single-cell level. We show thatcotDis not heterogeneously expressed among cells of a subpopulation. Furthermore, we discourage using plasmid-based reporter fusions for such studies, due to an introduced heterogeneity through copy number differences. This stresses the importance of using single-copy integrated reporter fusions for single-cell studies.


Lab on a Chip ◽  
2018 ◽  
Vol 18 (23) ◽  
pp. 3668-3677 ◽  
Author(s):  
Witold Postek ◽  
Pawel Gargulinski ◽  
Ott Scheler ◽  
Tomasz S. Kaminski ◽  
Piotr Garstecki

We separate emulsions with an immiscible oil phase to identify reaction conditions by the location of emulsion in emulsion series.


2014 ◽  
pp. 2791 ◽  
Author(s):  
Giovanni Maria Severini ◽  
Lorella Pascolo ◽  
Barbara Bortot ◽  
Nuria Benseny-Cases ◽  
Alessandra Gianoncelli ◽  
...  

2013 ◽  
Vol 64 (1) ◽  
Author(s):  
Rachel J. Errington ◽  
Sally C. Chappell ◽  
Imtiaz A. Khan ◽  
Nuria Marquez ◽  
Marie Wiltshire ◽  
...  

2019 ◽  
Vol 86 (3) ◽  
Author(s):  
Wenchao Zhang ◽  
Yan Wang ◽  
Huining Lu ◽  
Qin Liu ◽  
Chuandong Wang ◽  
...  

ABSTRACT The predatory behavior of Myxococcus xanthus has attracted extensive attention due to its unique social traits and inherent biological activities. In addition to group hunting, individual M. xanthus cells are able to kill and lyse prey cells; however, there is little understanding of the dynamics of solitary predation. In this study, by employing a bacterial tracking technique, we investigated M. xanthus predatory dynamics on Escherichia coli at the single-cell level. The killing and lysis of E. coli by a single M. xanthus cell was monitored in real time by microscopic observation, and the plasmolysis of prey cells was identified at a relatively early stage of solitary predation. After quantitative characterization of their solitary predatory behavior, M. xanthus cells were found to respond more dramatically to direct contact with live E. coli cells than heat-killed or UV-killed cells, showing slower predator motion and faster lysing of prey. Among the three contact-dependent killing modes classified according to the major subareas of M. xanthus cells in contact with prey, leading pole contact was observed most. After killing the prey, approximately 72% of M. xanthus cells were found to leave without thorough degradation of the lysed prey, and this postresidence behavior is described as a lysis-leave pattern, indicating that solitary predation has low efficiency in terms of prey-cell consumption. Our results provide a detailed description of the single-cell level dynamics of M. xanthus solitary predation from both prey and predator perspectives. IMPORTANCE Bacterial predation plays multiple essential roles in bacterial selection and mortality within microbial ecosystems. In addition to its ecological and evolutionary importance, many potential applications of bacterial predation have been proposed. The myxobacterium Myxococcus xanthus is a well-known predatory member of the soil microbial community. Its predation is commonly considered a collective behavior comparable to a wolf pack attack; however, individual M. xanthus cells are also able to competently lead to the lysis of a prey cell. Using a bacterial tracking technique, we are able to observe and analyze solitary predation by M. xanthus on Escherichia coli at the single-cell level and reveal the dynamics of both predator and prey during the process. The present study will not only provide a comprehensive understanding of M. xanthus solitary predation but also help to explain why M. xanthus often displays multicellular characteristic predatory behaviors in nature, while a single cell is capable of predation.


2010 ◽  
Vol 77 (1) ◽  
pp. 67-72 ◽  
Author(s):  
M. C. Konopka ◽  
T. J. Strovas ◽  
David S. Ojala ◽  
L. Chistoserdova ◽  
M. E. Lidstrom ◽  
...  

ABSTRACTThe ability to detect specific functions of uncultured microbial cells in complex natural communities remains one of the most difficult tasks of environmental microbiology. Here we present respiration response imaging (RRI) as a novel fluorescence microscopy-based approach for the identification of microbial function, such as the ability to use C1substrates, at a single-cell level. We demonstrate that RRI could be used for the investigation of heterogeneity of a single microbial population or for functional profiling of microbial cells from complex environmental communities, such as freshwater lake sediment.


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