Bacterial spores are highly resistant dormant structures formed by certain genera of bacteria. Sporulation occurs when nutrients become limited and involves the formation of a protective spore wall and coat. This allows the spore to survive extreme heat, chemicals, and lack of water. When conditions become favorable again, the spore can germinate into a new vegetative bacterial cell through a multi-step process. Certain spore-forming bacteria like Bacillus and Clostridium are used as indicators of proper sterilization.

1. Unit 2
General characteristics of Microbes

2. Spores
◌Highly resistant dormant state of bacteria found in certain genera
◌They are not destroyed by ordinary methods of boiling ofr several
hours
◌They are killed when autoclaved at 15lb pressure (pound per square
inch) at 121*C for 20 minutes.

3. Spores
◌Bacterial spores are small oval or spherical structures that are very
resistant to high temperatures, radiation, desiccation, and
chemical agents.
◌When they are formed intracellularly, they are called endospore. The
bacterial cell producing spore is called vegetative cell.

4. Formation Spores
◌Related to depletion of exogenous nutrients
◌Sporulation involves the production many new structures, enzymes
and metabolites along with the disappearance of many vegetative cell
components
◌It can be induced by PO4 S,C,N and Fe from culture medium

5. SPORULATION
◌The process of production of spores is called sporulation or
sporogenesis.
◌The one vegetative cell forms a single spore, which, after
germination, developed into a new cell.
◌ It takes 8hrs-19h

6. FORMATION OF SPORE
◌Appearance of clear area near one end of cell
◌Gradually becomes more opaque to form
forespore

8. FORMATION OF SPORE
◌Fully develoed spore has:
◌core nuclear body surrounded by spore
wall, a delicate membrane (future cell
wall)
◌Outside this is spore cortex
◌Which is enclosed by multilayered
spore coat

9. FORMATION OF SPORE
◌Spore cortex contains an unusual type of
peptidoglycan sensitive to lysozyme
◌Spore coat is formed by keratin like
protein which is impervious to
antibacterial chemical;
Exosporium

10. FORMATION OF SPORE
◌Additional outer covering called
Exosporium (having ridges and
grooves)
◌Exosporium is a lipoprotein membrane
with some carbohydrate residue
Exosporium

11. TYPES OF SPORE
TWO TYPES OF SPORES
◌ENDOSPORE
◌EXOSPORE
Exosporium

12. ENDOSPORE
◌Formed inside the parent vegetative cell.
◌Endospore are highly durable dehydrate cells, which can
◌survive extreme heat, lack of water, freezing and exposure
◌to many toxic chemicals and radiation.

13. EXOSPORE
◌Formed outside the vegetative cell by
◌budding at one end of the cell.
◌The Exospore do not contain dipicolinic acid.

14. Shape and positions of Bacterial spores

15. SPORE FORMING BACTERIA
◌ GRAM POSITIVE BACILLI:
◌Obligate aerobes- genus Bacillus
◌eg.B.anthracis and B. subtilis
◌Oblicate anaerobes- genus clostridia
◌eg. Cl.tetani, Cl welchii, Cl. Botulinium, Cl perfringes
◌Other bacteria :
◌Gram positive coccus: spore sarcina
◌Gram negative Bacillus: Coxiella burneti

16. GERMINATION OF SPORE
◌When transferred to a favorable conditions of growth spores
germinates.
◌The spore loses its refractility and swells.
◌The spore wall is shed and germ cell appears by rupturing the
spore coat
◌The germ cell elongates to form vegetative bacterium

17. GERMINATION OF SPORE
◌Process occurs in Three stages
1. Activation
2. Initiation
3. Outgrowth

18. GERMINATION OF SPORE
Activation :
• Most endospore cannot germinate immediately after they have formed.
• Germinate after they have rested for several days or are first activated
in a nutritionally rich medium (heat,{60*C for 1hour}, low pH,
sulfhydryl, abrasion, etc) by one or another agent that damages the
spore coat and germinates.

19. GERMINATION OF SPORE
Initiation:- After activation, a spore will initiate germination if the
environmental conditions are favorable
Binding of effector substance to spore coat, activates an autolysin
which destroys peptidoglycan of the cortex.

20. GERMINATION OF SPORE
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.

21. Demonstration
◌Gram staining:- spores appear as an unstained refractile body
within the cell.
◌Modified Ziehl-Neelsan(ZN) staining:-spores appear as acid-fast
(red colour). ZN staining with 0.25-0.5% sulphuric acid as
decolouring agent is used for spore staining.

22. USE OF SPORE
◌Spores of certain species of bacteria are employed as indicator for
proper sterilisation eg. Bacillus stearothermophilus which is
destroyed at a temperature of 121∘C for 10-20 min.
◌ These spores may be kept in autoclave prior to its use.
◌ Absence of the spores after autoclaving indicates proper
sterilization.

23. SPORE
◌Slender