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An endospore is a dormant, tough, and non-reproductive structure produced by certain bacteria from the Firmicute phylum. The name "endospore" is suggestive of a spore or seed-like form (endo means within), but it is not a true spore (i.e., not an offspring). It is a stripped-down, dormant form to which the bacterium can reduce itself. Endospore formation is usually triggered by a lack of nutrients, and usually occurs in Gram-positive bacteria. In endospore formation, the bacterium divides within its cell wall. One side then engulfs the other. Endospores enable bacteria to lie dormant for extended periods, even centuries. Revival of spores millions of years old has been claimed. When the environment becomes more favorable, the endospore can reactivate itself to the vegetative state. Most types of bacteria cannot change to the endospore form. Examples of bacteria that can form endospores include Bacillus and Clostridium.
The endospore consists of the bacterium's DNA and part of its cytoplasm, surrounded by a very tough outer coating.
Endospores can survive without nutrients. They are resistant to ultraviolet radiation, desiccation, high temperature, extreme freezing and chemical disinfectants. According to scientist Dr. Steinn Sigurdsson, "There are viable bacterial spores that have been found that are 40 million years old on Earth - and we know they're very hardened to radiation." Common anti-bacterial agents that work by destroying vegetative cell walls do not affect endospores. Endospores are commonly found in soil and water, where they may survive for long periods of time.
Some classes of bacteria can turn into exospores, also known as microbial cysts, instead of endospores. Exospores and endospores are two kinds of "hibernating" or dormant stages seen in some classes of microorganisms.
Bacteria produce a single endospore internally. The spore is sometimes surrounded by a thin covering known as the exosporium, which overlies the spore coat. The spore coat, which acts like a sieve that excludes large toxic molecules like lysozyme, is resistant to many toxic molecules and may also contain enzymes that are involved in germination. The cortex lies beneath the spore coat and consists of peptidoglycan. The core wall lies beneath the cortex and surrounds the protoplast or core of the endospore. The core contains the spore chromosomal DNA which is encased in chromatin-like proteins known as SASPs (small acid-soluble spore proteins), that protect the spore DNA from UV radiation and heat. The core also contains normal cell structures, such as ribosomes and other enzymes, but is not metabolically active.
Up to 20% of the dry weight of the endospore consists of calcium dipicolinate within the core, which is thought to stabilize the DNA. Dipicolinic acid could be responsible for the heat resistance of the spore, and calcium may aid in resistance to heat and oxidizing agents. However, mutants resistant to heat but lacking dipicolinic acid have been isolated, suggesting other mechanisms contributing to heat resistance are also at work.
Visualising endospores under the light microscope can be difficult due to the impermeability of the endospore wall to dyes and stains. While the rest of a bacterial cell may stain, the endospore is left colourless. To combat this, a special stain technique called a Moeller stain is used. That allows the endospore to show up as red, while the rest of the cell stains blue. Another staining technique for endospores is the Schaeffer-Fulton stain, which stains endospores green and bacterial bodies red. The arrangement of spore layers is as follows:
- Spore coat
- Spore cortex
- Core wall
The position of the endospore differs among bacterial species and is useful in identification. The main types within the cell are terminal, subterminal, and centrally placed endospores. Terminal endospores are seen at the poles of cells, whereas central endospores are more or less in the middle. Subterminal endospores are those between these two extremes, usually seen far enough towards the poles but close enough to the center so as not to be considered either terminal or central. Lateral endospores are seen occasionally.
Examples of bacteria having terminal endospores include Clostridium tetani, the pathogen that causes the disease tetanus. Bacteria having a centrally placed endospore include Bacillus cereus, and those having a subterminal endospore include Bacillus subtilis. Sometimes the endospore can be so large the cell can be distended around the endospore, this is typical of Clostridium tetani.
Formation and destruction
When a bacterium detects environmental conditions are becoming unfavourable it may start the process of endosporulation, which takes about eight hours. The DNA is replicated and a membrane wall known as a spore septum begins to form between it and the rest of the cell. The plasma membrane of the cell surrounds this wall and pinches off to leave a double membrane around the DNA, and the developing structure is now known as a forespore. Calcium dipicolinate is incorporated into the forespore during this time. Next the peptidoglycan cortex forms between the two layers and the bacterium adds a spore coat to the outside of the forespore. Sporulation is now complete, and the mature endospore will be released when the surrounding vegetative cell is degraded.
Endospores are resistant to most agents that would normally kill the vegetative cells they formed from. Nearly all household cleaning products, alcohols, quaternary ammonium compounds and detergents have little effect. However, alkylating agents (e.g. ethylene oxide), and 10% bleach are effective against endospores. To kill anthrax spores, standard household bleach (with 5% sodium hypochlorite) diluted freshly 1:10 in water (100ml of bleach in 900ml of water) must be in contact with the spore for at least 5 minutes. This contact time is important as a very small fraction of spores can survive longer than 10 minutes while soaked in this bleach solution. Higher concentrations of bleach is not more effective, and can cause some types of bacteria to aggregate and thus survive.
While resistant to extreme heat and radiation, endospores can be destroyed by burning or by autoclaving. Endospores are able to survive boiling at 100°C for hours, although the longer the number of hours the fewer that will survive. An indirect way to destroy them is to place them in an environment that reactivates them to their vegetative state. They will germinate within a day or two with the right environmental conditions, and then the vegetative cells can be straightforwardly destroyed. This indirect method is called Tyndallization. It was the usual method for a while in the late 19th century before the advent of inexpensive autoclaves. Prolonged exposure to ionising radiation, such as x-rays and gamma rays, will also kill most endospores.
In hospitals, endospores on delicate invasive instruments (eg, video endoscopes) are killed by low-temperature and non-corrosive, non-toxic, plasma-activated concentrated hydrogen peroxide vapor in sterilizers. In contrast, "High level disinfection" does not kill endospores but is used for instruments that don't enter sterile bodily cavities (eg, a colonoscope). This latter method involves only warm water, enzymes, and detergents.
Reactivation of the endospore occurs when conditions are more favourable and involves activation, germination, and outgrowth. Even if an endospore is located in plentiful nutrients, it may fail to germinate unless activation has taken place. This may be triggered by heating the endospore. Germination involves the dormant endospore starting metabolic activity and thus breaking hibernation. It is commonly characterised by rupture or absorption of the spore coat, swelling of the endospore, an increase in metabolic activity, and loss of resistance to environmental stress.
Outgrowth follows germination and involves the core of the endospore manufacturing new chemical components and exiting the old spore coat to develop into a fully functional vegetative bacterial cell, which can divide to produce more cells.
Endospores can stay dormant for a very long time. For instance, Endospores were found in the tombs of the Egyptian Pharaohs. When placed in appropriate medium, under appropriate conditions, they were able to be reactivated. Other viable endospores were recovered from bees trapped in amber that was about 25 million years old.
As a simplified model for cellular differentiation, the molecular details of endospore formation have been extensively studied, specifically in the model organism Bacillus subtilis. These studies have contributed much to our understanding of the regulation of gene expression, transcription factors, and the sigma factor subunits of RNA polymerase.
Endospores of the bacterium Bacillus anthracis were used in the 2001 anthrax attacks. The powder found in contaminated postal letters was composed of extracellular anthrax endospores. Inhalation, ingestion or skin contamination of these endospores led to a number of deaths.
According to WHO veterinary documents, B. anthracis sporulates when it sees oxygen instead of the carbon dioxide present in mammal blood; this signals to the bacteria that it has reached the end of the animal, and an inactive dispersable morphology is useful.
Sporulation requires the presence of free oxygen. in the natural situation, this means the vegetative cycles occur within the low oxygen environment of the infected host and, within the host, the organism is exclusively in the vegetative form. once outside the host, sporulation commences upon exposure to the air and the spore forms are essentially the exclusive phase in the environment. 
Geobacillus stearothermophilus endospores are used as biological indicators when an autoclave is used in sterilization procedures.
Bacillus subtilis spores are useful for the expression of recombinant proteins and in particular for the surface display of peptides and proteins as a tool for fundamental and applied research in the fields of microbiology, biotechnology and vaccination.
Examples of endospore-forming bacteria include the genera:
- C.Michael Hogan. 2010. Bacteria. Encyclopedia of Earth. eds. Sidney Draggan and C.J. Cleveland, National Council for Science and the Environment, Washington DC
- Cano RJ, Borucki MK (1995) Revival and identification of bacterial spores in 25- to 40-million-year-old Dominican amber. Science 268: 1060-1064. doi:10.1126/science.7538699
- "endospore" at Dorland's Medical Dictionary
- BBC Staff (23 August 2011). "Impacts 'more likely' to have spread life from Earth". BBC. Retrieved 2011-08-24.
- Prescott, L. (1993). Microbiology, Wm. C. Brown Publishers, ISBN 0-697-01372-3.
- Heninger, Sara; Christine A. Anderson, Gerald Beltz, and Andrew B. Onderdonk (January 1, 2009). "Decontamination of Bacillus anthracis Spores: Evaluation of Various Disinfectants". Appl Biosaf. 14 (1): 7–10. PMC 2957119. PMID 20967138.
- Abel-Santos, E (editor) (2012). Bacterial Spores: Current Research and Applications. Caister Academic Press. ISBN 978-1-908230-00-3.
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