Visualization of Phytoplasmas Using Electron Microscopy
The use of electron microscopy, both transmission and scanning, provides reliable and accurate methods for detecting phytoplasmas in plants. Our understanding of these pathogens, their morphology, development, and intracellular location in plants and insect vectors has been greatly increased through the use of these instruments. Development of techniques such as immunolabeling, cryofixation with freeze substitution or plunge freezing with direct transfer to the microscope stage, together with advances in instrumentation is enabling us to study these pathogens under conditions close to their native state. The visualization of fine detail and ultrastructure, using modern and established techniques, can only be appreciated by the magnification and spatial resolution offered in the electron microscopes. Now that the full sequencing of four phytoplasma genomes (to date) has been achieved, electron microscopy can play an important role in identifying and understanding specific gene functions.
Key wordsCryofixation Electron microscopy Scanning electron microscopes Transmission electron microscopes Ultramicrotomy
I would like to thank Prof. Phil Jones, former head of Bioimaging at Rothamsted, for his valued advice and from whom a version of the TEM protocol originated and the use of the TEM images. Also to Prof. John Lucas and Dr Allison van de Meene for helpful comments on the manuscript. Rothamsted Research receives grant-aided support from the Biotechnology and Biological Sciences Research Council (BBSRC) of the UK.
- 4.Horne RW (1970) The ultrastructure of mycoplasma and mycoplasma-like organisms. Micron 2:19–38Google Scholar
- 5.Waters H, Hunt P (1980) The in vivo three dimensional form of a plant mycoplasma-like organism by analysis of serial ultrathin sections. J Gen Microbiol 116:111–131Google Scholar
- 6.Musetti R (2010) Biochemical changes in plants infected by phytoplasmas. In: Weintraub PG, Jones P (eds) Phytoplasmas: genomes, plant hosts and vectors. CAB International, Wallingford, pp 132–146Google Scholar
- 10.Kiernan JA (2000) Formaldehyde, formalin, paraformaldehyde and glutaraldehyde: what they are and what they do. Microsc Today 00–1:8–12Google Scholar
- 12.Dykstra MJ, Reuss LE (2003) Biological electron microscopy: theory, techniques and troubleshooting, vol 2. Springer Verlag GmbH, New York, pp 27–28Google Scholar
- 15.Haggis GH, Sinha RC (1977) Scanning electron microscopy of mycoplasma like organisms after freeze fracture of plant tissues affected with Clover Phyllody and Aster Yellows. J Phytopathol 68:677–680Google Scholar