Bacterial Spores: Structure and Spore-Forming Bacteria

Bacterial spores are highly resistant, dormant structures (i.e. no metabolic activity) formed in response to adverse environmental conditions. As bacterial spores are formed within the parent cell, these are called endospores.

When vegetative cells of certain bacteria such as Bacillus spp and Clostridium spp are subjected to environmental stresses such as nutrient deprivation, they produce metabolically inactive or dormant form-endospore. The formation of endospores circumvents the problems associated with environmental stress and ensures the survival of the organisms.

Note: Spores of fungi have a reproductive role. 

Endospores’ size, shape, and location are particularly useful for identifying Clostridium, Bacillus, and related species. Bacterial spores (e.g., spores of Bacillus stearothermophilus) are used as an indicator for proper sterilization of autoclave.


Bacterial cell undergoes spore formation in nutritionally deprived conditions, and this process is called sporulation. Spore develops from a portion of protoplasm (forespore) near one end of the cell. The remaining part of the cell is called sporangium. Spore-forming bacilli form endospores during unfavorable conditions (especially when carbon and nitrogen become depleted or unavailable).

Structure of the Bacterial Spore

An endospore is structurally and chemically more complex than a vegetative cell.  It contains more layers than vegetative cells. Resistance of Bacterial spores may be mediated by dipicolinic acid, a calcium ion chelator found only in spores.  Following are the constituents of bacterial spores;

Structure of Bacterial Spore
Structure of Bacterial Spore


Exosporium, a thin protein covering, is the outermost layer of a bacterial endospore.

Spore coat

It lies below the exosporium. The spore coat is composed of layers of spore-specific proteins.


The cortex lies below the spore coat and consists of loosely cross-linked peptidoglycan.


The core is the innermost region of a bacterial endospore surrounded by a core wall. Cytoplasmic membrane, cytoplasm, nucleoid, ribosomes, and other cellular essentials are found inside the core wall. The core of the mature endospore differs greatly from the vegetative cell from which it was formed.

  1. Dipicolinic acid, absent in vegetative cells, accumulates in the bacterial endospore’s core.
  2. Endospores are also enriched in calcium, most of which are complexed with dipicolinic acid. The calcium-dipicolinic acid complex represents about 10% of the dry weight of the endospore, thus helping to dehydrate it. This complex intercalates (inserts between bases) in DNA and stabilizes the DNA against heat denaturation.
  3. The core of a mature endospore has only 10-25% of the water content of the vegetative cell, and thus the consistency of the core cytoplasm is that of a gel.
  4. In addition to the endospore’s low water content, the core’s pH is about one unit lower than the vegetative cell cytoplasm.

Spore Germination

The process of conversion of a spore into a vegetative cell under suitable conditions is known as germination. When favorable conditions prevail (i.e., availability of water, appropriate nutrients), spores germination occurs, forming vegetative cells of pathogenic bacteria.

This process involves three steps: activation, germination, and outgrowth. Activation occurs when endospores are heated for several minutes at an elevated but sub-lethal temperature. Activated endospores can germinate when placed in a suitable culture medium. Germination is a rapid process that involves loss of microscopic refractility of the endospore, increased ability to be stained by dyes, and loss of resistance to heat and chemicals. The final stage, outgrowth, involves visible swelling due to water uptake and synthesis of RNA, proteins, and DNA.


Bacterial spores are highly resistant to ordinary boiling, disinfectants, and heating, but spores of all medically important bacteria are destroyed by autoclaving. Bacterial spores are highly resistant to;

  • Heat
  • Dehydration
  • Radiation and
  • Chemicals

The following factors/constituents play major roles in the resistance of bacterial spore: 

  • Calcium dipicolinate in core
  • Keratin spore coat
  • New enzymes (i.e., dipicolinic acid synthetase, heat-resistant catalase)
  • Increases or decreases in other enzymes.
Spore of Clostridium botulinum

A mature endospore contains a complete set of genetic material (DNA) from the vegetative cell, ribosomes, and specialized enzymes.

Mature endospores are released from the vegetative cell to become free endospores.  When the free endospores are placed in an environment that supports growth, the endospores will revert to vegetative cells in a process called germination.  It should be noted that unlike the process of binary fission observed with vegetative cells, endospore formation is not a reproductive process but a process of differentiation that provides the bacteria with a mechanism for survival.

Medical Importance of Bacterial Spores

Important features of SporesMedical Implications
Spores are highly resistant to heating; spores are not killed by boiling (100°C) but are killed at 121°C.Medical supplies must be heated to 121°C for at least 15 minutes to be sterilized.
Spores are highly resistant to many chemicals, including most disinfectants.The only solution designated as sporicidal will kill spores.
Spores can survive for many years in soil and other inanimate objects.Wounds contaminated with soils can be infected with spores and cause diseases such as tetanus and gas gangrene.
Spores do not exhibit measurable metabolic activity.Antibiotics are ineffective against spores.
Spores are formed only when nutrients are insufficient.Spores are not often found at the site of infection because nutrients are not limited.

Examples of spore-forming bacteria

Most endospore-forming bacteria are found in soil or aquatic environments.  However, some species of Bacillus and Clostridium have medical significance. Clostridium perfringens, C. botulinum (a potential agent of bioterrorism), and C. tetani are the causative agents of gas gangrene, botulism, and tetanus, respectively.  Bacillus anthracis and Bacillus cereus are the causative agents of anthrax and self-limiting food poisoning, respectively.

Demonstration of Endospore

Endospores cannot be stained by ordinary methods, such as simple and Gram staining, because dyes do not penetrate the endospore wall. But endospores can be seen as an unstained refractile body within the cell, in gram-stained smears, or unstained preparations. Without a special stain, it’s hard to differentiate endospores from inclusions of stored material.

Though the modified Ziehl-Neelsen method can be used for endospore staining, the most commonly used endospore stain is the Schaeffer-Fulton.

Positions of Bacterial Spores

The shape and the position of spores vary in different species and can be useful for classification and identification purposes. The position of the spores can be seen in the smear using the endospore staining method. Endospores may be located in the middle of the bacterium (central), at the end of the bacterium (terminal), near the end of the bacteria (subterminal), and maybe spherical or elliptical.

Spore location
  1. Central endospores are located within the middle of the vegetative cell.  
  2. Terminal endospores are located at the end of the vegetative cell.  
  3. Sub-terminal endospores are located between the middle and the end of the cell.


  1. Central or equatorial, giving the bacillus a spindle shape (eg. Clostridium bifermentans)
  2. Sub-terminal, the bacillus appearing Club shaped (eg. Clostridium perfringens)
  3. Oval and terminal, resembling a tennis racket (eg. Clostridium tertium)
  4. Spherical and terminal, giving a drumstick appearance (Clostridium tetani)

References and further readings

  1. Madigan, M., Martinko, J., Stahl, D., & Clark, D. (2012). Brock Biology of Microorganisms (13th ed). Pearson Education
  2. Bailey & Scott’s Diagnostic Microbiology, Forbes, 11th edition
  3. Review of Medical Microbiology and Immunology, Lange Medical Books, 13th edition

Acharya Tankeshwar

Hello, thank you for visiting my blog. I am Tankeshwar Acharya. Blogging is my passion. As an asst. professor, I am teaching microbiology and immunology to medical and nursing students at PAHS, Nepal. I have been working as a microbiologist at Patan hospital for more than 10 years.

Recent Posts