Bacterial Endospore
* Endospores are dormant structures formed by vegetative bacterial cells when they find themselves in unfavorable environments.
* Endospore formation is most common in gram-positive Bacillus and Clostridium, but can also be formed in less common genera of bacteria including Desulfotomaculum, Sporocarcina, Sporolactobacillus, Oscillospira and Thermoactinomyces.
* Endospores are resistant to heat, pressure, drying, radiation, starvation, antibiotics and various chemical disinfectants and can survive for thousands of years in this dormant state.
* An endospore is a dormant, though and temporarily mom-reproductive structure produced by certain bacteria. The name "endospore" is suggestive of a spore or seedlike form (endo means within), but it is not a true spore.
* The endospore becomes important when the bacterium is experiencing an environment that us tough to the usual vegetative state of the bacterium, such as in desiccating conditions.
* Endospores enable bacterium to survive periods of environmental stress lasting at least several thousand years and revival of spores many millions of year old has been claimed.
* When the environment becomes more favourable, the endospore can reactivate itself to the vegetative state. Most types of bacteria cannot change to the endospore form, bit examples include Bacillus and Clostridium.
Types of spore arrangement
Variations in endospore morphology : (1,4) central endospore; (2,3,5) terminal endospore; (6) lateral endospore.
• The endospore consists of the bacterium's DNA and part of its cytoplasm, surrounded by a very tough outer coating. Endospore can survive without nutrients.
• They are resistant to ultraviolet radiation, desiccation, high temperature, extreme freezing and chemical disinfectants.
• Common anti-bacterial agents that work by destroying vegetative cell walls don't work on endospore.
• The position of the endospore differs among differs among bacterial species and is useful in identificatiom. The main types within the cell are terminal, subterminal, subterminal and centrally placed endospore.
• 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.
• 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.
Architechture of endospores
• Endospores can be divided into several important parts. The center of the endospores contains the core and it consists of the cytoplasm, DNA, ribosomes, enzymes and everything that is needed to function once returned to the vegetative state.
• The core is dehydrated, which is essential for heat resistance, long-term dormancy and full chemical resistance.
• Calcium dipicolinate is a major component of the core and has been shown to play a role in resistance to wet heat and UV light. Dipicolinic acid (pyridine-2,6-dicarboxylic acid or PDC) is a chemical compound which composes 5% to 15% of the dry weight of bacterial spores. It is responsible for the heat resistance of the endospore.
• Dipicolinic acid (pyridine-2,6-dicarboxylic acid or PDC) is a chemical compound which composes 5% to 15% of the dry weight of bacterial spores. It is implicated as responsible for the heat resistance of the endospore.
• The cortex surrounds the core and is composed of two layers a thin more densely staining layer that is similar in structure to the vegetative cell wall and a thicker less dense layer containing modified peptidoglycan. Two major modifications are present.
• First, there is less cross-linking with only 3% of the muramic acid present in the peptidoglycan of the cortex participating, in comparison to 40% of muramic acid in the vegetative cell wall.
• Second much of the muramic acid is modified to a muramic-β-lactam structure. Both of these modifications of the cortex appear to be important in germination.
• Muramic-β-lactam serves as a specific target for lytic enzymes that are activated during germination and the lower cross-linking enables easier outgrowth.
• Outside of the cortex is the spore coat containing several protein layers that are impermeable to most chemicals.
• The coat is composed of more than two dozen different types of proteins and there is some evidence that these proteins are connected by cross-links. This covalent connection between coat proteins probably contributes to the spores resistance.
Sporulating Process
• Under conditions of starvation, especially the lack of carbon and nitrogen sources, a single endospores form within some of the bacteria. The process is called sporulation.
• Endospores are the most heat-resistant, living organisms known. They form through a complex series of events as shown below.
• First the DNA replicates and a cytoplasmic membranes septum forms at one end of the cell.
• A second layer of cytoplasmic membrane then forms around one of the DNA molecules (the one that will become part of the endospore) to form a forespore.
• Both of these membrane layers then synthesize peptidoglycan in the space between them to form the first protective coat, the cortex.
• Calcium dipocolinate is incorporated into the forming endospore.
• A spore coat composed of keratin-like protein then forms around the cortex. Sometimes an outer membrane composed of lipid and protein and called an exosporium is also seen.
• Finally, the remainder of the bacterium is degraded and the endospore is released. Sporulation generally takes around 15 hours.
• The completed endospore consists of multiple layers of resistant coats (including a cortex, a spore coat, and sometimes an exosporium) surrounding a nucleoid, some ribosomes, RNA molecules and enzymes.
• Bacterial endospores are resistant to antibiotics, most disinfectants and physical agents such as radiation, boiling and drying.
• The impermeability of the spore coat is thought to be responsible for the endospore's resistance to chemicals.
• Once the endospore is formed, the vegetative portion of the bacterium is degraded and the dormant endospore is released.
Although harmless themselves until they germinate, they are involved in the transmission of some disease to humans. Infections transmitted to humans by endospores include :
• Anthrax, caused by Bacillus anthracis;
• Tetanus, caused by Clostridium tetani;
• Botulism, caused by Clostridium botulinum; and
• Gas Gangrene, caused by Clostridium perfringens.
Table showing Comparison of cells and endospores contents
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Destruction of Endospores
• Endospores are resistant to most agents that would normally kill the vegetative cells they formed from.
• While resistant to extreme heat and radiation, endospores can be destroyed by burning or by autoclaving.
• 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 autoclave.
• Prolonged exposure to ionising radiation, such as x-rays and gamma rays, will also kill most endospores.
Germination of Endospores
• Although viability of spores decreases with the passage of time, organisms have been cultured from spores of bacilli over 300 years old.
• The process by which a spore is converted into a germinative cell is called germination.
• Three stages of germination have been distinguished, activation, initiation (germination proper) and outgrowth, or post germinative development.
Activation, Germination & Growth
• Some bacterial spores germinate spontaneously in a favourable medium.
• Others remain dormant even if placed inoptical conditions for germination.
• i) Activation may be brought about by traumatic agents such as heat, low pH and an a SH composed.
• Certain chemicals such as L-alanine, adenosine, glucose and some reducing agents can also bring about activation:
• Heat shock or heat activation appears to be the most general mechanism for activation spores. Heating of spores in an aqueous fluid for 15-60 minutes at 65°C results in the activation of most spores. Higher temperatures of 105 to 120°C are required for thermophilic spores.
• Heat activation is a reversible process. The induced germ inability declines if spores are returned to lower temperatures for some days.
• Reactivation of 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.
• ii) Germination involves the dormant endospore staring 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.
• iii) 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.
• Bacterial spores have no metabolic activity and are adopted for prolonged survival under adverse conditions spores can remain dormant for several years.
Ribosomes
Ribosomes are small granular bodies of 10-20 nm in diameter freely lying in the cytoplasm and composed of ribosomal ribonucleic acid (rRNA) and proteins. Bacterial ribosomes are thought to contain about 80-85% of the bacterial RNA. Sometimes, they are found in small groups called polyribosomes or polysomes, which are formed when several ribosomes begin to translate a single mRNA molecule. Each ribosome has sedimentation coefficient of 70 S and a mass of 2.8 x 106 daltons, and is made up of two subunits of 50 S and 30S, each subunit consisting of roughly equal amounts of rRNA and proteins. Ribosomes are functional only when the two subunits are combined together.
The association and dissociation of two subunits of ribosomes depend on the concentration of Mg++ ions. Each 50 S subunit (mass of 1.8 x 106 daltons) contains one molecule of 23 S rRNA (having approximately 3200 nucleotides), one molecule of 5S rRNA (having only about 120 nucleotides) and 34 different proteins designated as L1 to L34; while the 30 S subunit (mass of 0.9 x 106 daltons) contains one molecule of 16rRNA (having approximately 1540 nucleotides) and 21 different proteins designated as S1 to S21.
As the eukaryotes, ribosomes are the sites of protein synthesis and therefore, antibiotics such as streptomycin and chloramphenicol specifically inhibit protein synthesis by attacking ribosomes. Generally, the ribosomes are a few hundred in number in each bacterial cell, but when the cell undertakes active protein synthesis, they increase in number to as many as 15,000-20,000 per cell, about 15% of the cell mass.
Plasmids
A plasmid is a DNA molecule that is separate from and can replicate independently of, the chromosomal DNA. The term plasmid was first introduced by the American molecular biologist Joshua Lederberg in 1952. They are double stranded and, in many cases, circular. Plasmids usually occur naturally in bacteria, but are sometimes found in eukaryotic organisms (e.g., the 2-micrometre-ring in Saccharomyces cerevisiae).
Plasmid size varies from 1 to over 1,000 kilobase pairs (kbp). The number of identical plasmids within a single cell can range from one to even thousands under some circumstances. Plasmids are often associated with conjugation, a mechanism of horizontal gene transfer. Plasmids are considered transferable genetic elements, or "replicons", capable of autonomous replication within a suitable host. Plasmids can be found in all three major domains, Archea, Bacteria and Eukarya.
The typical ds-DNA circular plasmid is less than 1/20 the size of the bacterial chromosome. Plasmids, the extra-chromosomal genetic materials, existing independently of the bacterial chromosome and are present in many bacteria (they are also present in some yeasts and other fungi). The replicate autonomously as they possess their own replication origins. Because of the small size of plasmid DNA relative to the bacterial chromosome, the whole replication process takes place very quickly, perhaps in 1/10 or less of the total time of cell division cycle. Plasmids have relatively few genes, generally less than 30 and their genetic information is not essential for the bacteria because the latter lacking them function normally. Those plasmids that can reversibly integrate into the bacterial chromosome are called episomes. The plasmids share many characteristics with viruses.
Types of Plasmids
• F-Plasmids tor F-factors
These are the first described plasmids that play major role in conjugation in bacteria.
• R-Plasmids
These are the most widespread and well-studied group of plasmids conferring resistance (hence called resistant plasmids) to antibiotics and various other growth inhibitors. R-plasmids typicallyhave genes that code for enzymes able to destroy and modify antibiotics.
• Virulence-Plasmids
These confer pathogenicity on the host bacterium. They make the bacterium more pathogenic as the bacterium is better able to resist host defence or to produce toxins.
• Col-Plasmids
These plasmids carry genes that confer ability to the host bacterium to kill other bacteria by secreting bacteriocins, a type of proteins. Bacteriocins often kill cells by creating channels in the plasma membrane thus increasing its permeability.
Mesosomes
In most of the bacteria cells (particularly fram-positive ones) the plasma membrane shows characteristic infoldings either superficially or significantly deep, invading the cytoplasm. These infoldings are called Mesosomes, the term coiled by Fitzjames. The bacterial DNA (chromosome) is always attached to or closely associated with Mesosomes. Mesosomes are considered to play in important role in the intiation or replication of bacterial DNA and the septa formation at the time of cell division (Higgins and Shockmann, 1971). They acts as sites of respiratory activity as well.
Mesosomes are folded invaginations in the plasma membrane of bacteria that are produced by the chemical fixation techniques used to prepare samples for electron microscopy. Although several functions were proposed for these structures in the 1960s, they were recognized as artifacts by the late 1970s and are no longer considered to be part of the normal structure of bacterial cells.
Mesosomes Real Structures
Although many functions have been proposed for Mesosomes, it has been found during the recent past that the bacterial cells with no apparent Mesosomes were not defective for such functions. The promoted to reevaluate the evidence for the existence of Mesosomes were always observed attached to or closely associates with bacterial DNA in only electron microscopic observations were frozen in liquid nitrogen for electron microscopy. No Mesosomes were observed elsewhere in cells. This suggests that the observed Mesosomes were artifacts of preparations for electron microscopic observation, formed by DNA pulling on the plasma membrane when the cell were dehydrated. The current view, therefore, is that Mesosomes are artifacts rather than real structures of the bacterial cell with definite functions.