Electron microscopy of malachite green--glutaraldehyde fixed bacteria

Area of Science:

• Microscopy
• Cell Biology
• Bacteriology

Background:

• Lipid complexes in eukaryotic cells are preserved using malachite green and glutaraldehyde fixatives.
• Electron microscopy requires specific fixation techniques for optimal visualization of cellular structures.

Purpose of the Study:

• To investigate the efficacy of malachite green-glutaraldehyde as a fixative for bacterial cell structures.
• To identify and characterize granular inclusions revealed by this new fixation method.

Main Methods:

• Bacteria were prefixed with a mixture of malachite green and glutaraldehyde.
• Comparison of fixation with glutaraldehyde alone versus the combined fixative.
• Electron microscopy was used to examine thin sections of preserved bacterial cells.

Main Results:

• The malachite green-glutaraldehyde mixture revealed granular electron-dense inclusions in all tested bacterial cultures.
• These inclusions were absent or electron-transparent when glutaraldehyde alone was used.
• Polyhydroxybutyrate granules in Aquaspirillum serpens were well preserved.
• Inclusions were located near the nucleoid and cell membrane (Gram-negative) or associated with nucleoid and mesosomes (Gram-positive).
• Malachite green complexes were observed extracellularly; capsules were not preserved or stained.

Conclusions:

• Malachite green-glutaraldehyde is a superior fixative for revealing and preserving specific granular inclusions in bacteria.
• This method enhances the visualization of polyhydroxybutyrate granules, improving their preservation in electron microscopy studies.
• The fixative offers new insights into bacterial ultrastructure and the nature of these inclusions.