Cytoplasmic Granules in Bacteria

Cytoplasmic granules or inclusion bodies are concentrated deposits of certain substances. Their location is in the cytoplasm of certain bacteria. They serve as storage areas for nutrients, e.g. volutin granules are high energy reserves stored in the form of polymerized metaphosphate.

Types of Cytoplasmic Granules

The number and nature of the inclusions vary depending on the bacterial species and the nutritional state of the organism’s environment. Common types of granules that are microscopically visible in certain bacteria stained with specific dyes are:

• Polysaccharide granules (glycogen): a storage form of glucose and
• polyphosphate granules: a storage form for inorganic phosphates (volutin granules, also known as metachromatic granules, are composed of polyphosphate)
• poly-β-hydroxybutyrate (PHB) granules: a reserve carbon and energy source.
• Elemental sulfur: Intracellular globules of elemental sulfur may accumulate in certain bacteria growing in environments rich in hydrogen sulfide.
• Magnetosomes: Some bacteria have specialized membrane-bound organelles called magnetosomes that contain magnetic crystals, usually magnetite (iron oxide) or greigite. These magnetosomes allow bacteria to orient themselves along magnetic field lines, aiding their navigation within aquatic environments.

Cytoplasmic granules/inclusion bodies stain characteristically with certain dyes;

1. Volutin granules appear as “metachromatic” granules, stains intense reddish-purple color with methylene blue dye (instead of blue, as one would expect), and can be observed by light microscopy.
2. Poly-β-hydroxybutyrate (PHB) granules can be stained with lipid-soluble dyes such as Nile blue.
3. Glycogen can be stained brown with iodine.

Metachromatic granules are characteristics of Corynebacterium diphtheriae

Staining Methods Used for Observing Cytoplasmic Granules in Bacteria

Methylene Blue Staining

Purpose: Methylene blue is a basic dye that stains negatively charged components in bacterial cells.

Results: Cytoplasmic granules like polyphosphate may take up the stain and appear blue.

Application: Provides a quick and general overview of bacterial morphology, including some granules.

Nile Red Staining

Purpose: Nile Red is a fluorochrome that selectively stains lipids.

Results: Lipid granules within bacterial cells will fluoresce when stained with Nile Red.

Application: Useful for detecting lipid-rich structures.

Sudan Black B Staining

Purpose: Sudan Black B is a lipophilic dye that stains lipids.

Results: Lipid granules in bacterial cells will appear black or dark blue.

Application: Similar to Nile Red, Sudan Black B detects lipid accumulation.

Gram Staining

Purpose: Gram staining is a differential method for categorizing bacteria based on their cell wall characteristics.

Results: Bacteria with thick peptidoglycan cell walls may have granules that retain the crystal violet stain, appearing purple.

Application: While primarily used for cell wall characteristics, Gram staining may indirectly reveal specific granules in bacterial cells.

Acid-Fast Staining

Purpose: Used to identify bacteria with mycolic acid in their cell walls, such as Mycobacterium species.

Results: Acid-fast bacteria retain the primary stain and appear red or pink.

Application: Useful for identifying bacteria with specific lipid-rich structures.

References

1. Obruca, S., Sedlacek, P., Mravec, F., Krzyzanek, V., Nebesarova, J., Samek, O., Kucera, D., Benesova, P., Hrubanova, K., Milerova, M., & Marova, I. (2017). The presence of PHB granules in cytoplasm protects non-halophilic bacterial cells against the harmful impact of hypertonic environments. New biotechnology, 39(Pt A), 68–80. https://doi.org/10.1016/j.nbt.2017.07.008