Sterilization and disinfection
Dipicolinic acid is present in (DNB June 2010)
|A||Gram- positive cell wall|
|B||Gram- negative cell wall|
a. Dipicolinic acid is exclusively found in the bacterial spores.
b. It is present in the spore core.
c. It is responsible for heat-resistance of spores.
- Sporing bacteria: Bacillus, Clostridium, Sporosarcina
- Each bacterium forms one endospore which germinates into single vegetative cell
- Method of preservation not replication
- Highly resistant resting stage formed during nutrient depletion
- Layers : (from inside out) spore cytoplasm, spore wall (inner membrane), spore cortex, spore coat (outer membrane), exosporium
- Round, oval, elongated ; terminal , subterminal, central ; bulging, non bulging
- Dormant for many years
- Extremely resistant to physical agents may be due to dipicolinic acid, a calcium ion chelator found only in bacterial spores
- Killed by moist heat 121c, 15min, 15 psi: autoclaving
- Demonstrated by: modified ZN staining (0.5% sulphuric acid as decoloriser) and by Ashby’s stain,
- Unstained in gram stain, In unstained preparation : refractile
Fig: The stages of endospore formation. (Reproduced with permission from Merrick MJ: Streptomyces.
The germination process occurs in three stages: activation, initiation, and outgrowth.
1. Activation—Most endospores cannot germinate immediately after they have formed. But they can germinate after they have rested for several days or are first activated, in a nutritionally rich medium, by one or another agent that damages the spore coat. Among the agents that can overcome spore dormancy are heat, abrasion, acidity, and compounds containing free sulfhydryl groups.
2. Initiation—Once activated, a spore will initiate germination if the environmental conditions are favorable. Different species have evolved receptors that recognize different effectors as signaling a rich medium: Thus, initiation is triggered by L-alanine in one species and by adenosine in another. Binding of the effector activates an autolysin that rapidly degrades the cortex peptidoglycan. Water is taken up, calcium dipicolinate is released, and a variety of spore constituents are degraded by hydrolytic enzymes.
3. Outgrowth—Degradation of the cortex and outer layers results in the emergence of a new vegetative cell consisting of the spore protoplast with its surrounding wall. A period of active biosynthesis follows; this period, which terminates in cell division, is called outgrowth. Outgrowth requires a supply of all nutrients essential for cell growth.