Observation of Bacteriophage Ultrastructure by Cryo-Electron Microscopy

Abstract

Transmission electron microscopy (TEM) is an ideal method to observe and determine the structure of bacteriophages. From early studies by negative staining to the present atomic structure models derived from cryo-TEM, bacteriophage detection, classification, and structure determination have been mostly done by electron microscopy. Although embedding in metal salts has been a routine method for virus observation for many years, the preservation of bacteriophages in a thin layer of fast frozen buffer has proven to be the most convenient preparation method for obtaining images using cryo-electron microscopy (cryo-EM). In this technique, frozen samples are observed at liquid nitrogen temperature, and the images are acquired using different recording media. The incorporation of direct electron detectors has been a fundamental step in achieving atomic resolution images of a number of viruses. These projection images can be numerically combined using different approaches to render a three-dimensional model of the virus. For those viral components exhibiting any symmetry, averaging can nowadays achieve atomic structures in most cases. Image processing methods have also evolved to improve the resolution in asymmetric viral components or regions showing different types of symmetries (symmetry mismatch).

Keywords: Bacteriophage structure; Cryo-electron microscopy; Data acquisition; Fast freezing; Image processing; Three-dimensional reconstruction.