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Extracellular Matrix Reorganization during Cryo Preparation for Scanning Electron Microscope Imaging of Staphylococcus aureus Biofilms

Published online by Cambridge University Press:  04 July 2014

Yong Wu ,
Jing Liang ,
Kim Rensing ,
Tseng-Ming Chou  and
Matthew Libera
Yong Wu
Affiliation:
Department of Chemical Engineering and Materials Science, Stevens Institute of Technology, Hoboken, NJ 07030, USA
Jing Liang
Affiliation:
Department of Chemical Engineering and Materials Science, Stevens Institute of Technology, Hoboken, NJ 07030, USA
Kim Rensing
Affiliation:
Leica Microsystems Inc., 1700 Leider Lane, Buffalo Grove, IL 60089, USA
Tseng-Ming Chou
Affiliation:
Department of Chemical Engineering and Materials Science, Stevens Institute of Technology, Hoboken, NJ 07030, USA
Matthew Libera*
Affiliation:
Department of Chemical Engineering and Materials Science, Stevens Institute of Technology, Hoboken, NJ 07030, USA
*
*Corresponding author. [email protected]
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Abstract

Biofilms are three-dimensional communities of bacteria distributed in a highly hydrated extracellular matrix (ECM). They can be visualized by scanning electron microscopy (SEM), but the requisite SEM sample preparation can modify the biofilm morphology. Here, four different approaches to prepare biofilms of hydrated Staphylococcus aureus for SEM imaging are compared. In order of increasing cooling effectiveness these are: (1) drying in air; (2) plunging in liquid nitrogen; (3) plunging in liquid ethane; and (4) high pressure freezing with liquid nitrogen. These different methods give rise to markedly different biofilm morphologies, which are revealed by cryo-SEM imaging. Significantly, high-pressure frozen biofilms exhibit a rich network of nanoscale ECM fibers surrounding individual bacteria throughout the biofilm thickness. This structure is entirely lost when similar biofilms are dried in air, and it is substantially modified when these biofilms are plunged into liquid nitrogen or liquid ethane.

Keywords

biofilmextracellular matrixscanning electron microscopycryoS. aureus
Type
Biological Applications
Information
Microscopy and Microanalysis , Volume 20 , Issue 5 , October 2014 , pp. 1348 - 1355
Copyright
© Microscopy Society of America 2014 

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