5 - Bacterial Endospores Flashcards
Endospores
- Resistant, dormant spores formed in some genera of bacteria
- Resistant to chemicals, temperature, and radiation
- Confer resistance, survival and dormancy to host bacteria
Which bacteria form endospores
- Only members of the low GC subdivision of gram +ves
- Make a single endospore per cell that later germinates to form a single cell
Genera of endospore forming bacteria
- Bacillus (rods, aerobic)
- Clostridium (rods, anaerobic)
- Sporosarcina (cocci, aerobic)
Swollen sporangium
Spore is wider than the width of the vegetative cell
Different locations of endospores
- Terminal
- Subterminal
- Central
- Each location can also be swollen
Human significance of endospores
- Set the sterilisation standard
- Medical importance
- Bioterrorism
- Developmental biology
- Pest control in agriculture
How did endospores set the sterilisation standard
- Endospores survive hours of boiling, and most chemical disinfectants
- Sterilisation processes must kill endospores (autoclave)
Medical importance of endospores
- Several spore-forming species are dangerous pathogens (e.g. tetanus)
- Spores provide a source of contamination & infection
Endospore bioterrorism
Use of Bacillus anthracis as a bioterror weapon is based on production and spread of its endospores
Endospore developmental biology
increased understanding of differentiation in bacteria
Endospore pest control
- Bacillus thuringiensis is
an insect biopesticide - Produces a toxic protein
What triggers sporulation
- Bacterial response to unfavourable conditions
- Occurs in stationary phase
unfavourable conditions that can trigger sporulation
- Absence of nutrients (C, N, P)
- Desiccation (removal of moisture)
- Presence of toxins or antibiotics
- Overcrowding
Structure enclosing
cytoplasm in vegetative cells vs endospores
- Vegetative cells: Typical G+ve cell (CW, PM)
- Endospores: Complex of layers
Microscopic appearance in vegetative cells vs endospores
- Vegetative cells: non refractile
- Endospores: refractile
Enzymatic activity in vegetative cells vs endospores
- Vegetative cells: High
- Endospores: Absent
Metabolism in vegetative cells vs endospores
- Vegetative cells: High
- Endospores: Absent
Macromolecular synthesis in vegetative cells vs endospores
- Vegetative cells: Present
- Endospores: Absent
Resistance (heat, radiation, chemical) in vegetative cells vs endospores
- Vegetative cells: Low
- Endospores: High
Sensitivity to lysozyme in vegetative cells vs endospores
- Vegetative cells: Sensitive
- Endospores: Resistant
Sustainability in vegetative cells vs endospores
- Vegetative cells: Easy to stain
- Endospores: Special methods needed
Cytoplasmic pH in vegetative cells vs endospores
- Vegetative cells: ~7
- Endospores: ~5.5-6
Endospore Exosporium
- Outermost layer in SOME species
- Thin glycoprotein layer outside spore coast
- Hair like projections and a basal layer
- Mediates environmental inerations (e.g. adhesion)
Endospore Spore coat
- Keratin-like protein layers rich in S-S bonds
- HIghly impermeable
- Provides resistance to chemicals (e.g. lysozyme)
Four layers of spore coat
Basement, inner, outer, crust
Endospore cortex
- Peptidoglycan with reduced cross linking of NAM residues because cortex PG if modified
Function of endospore cortex
Loosely cross-linked flexible cortex may allow spore envelope to expand and contract in response to humidity change
Endospore core wall
- Typical cell wall (but no teichoic acids)
- Provides osmotic stability
at spore germination - Acts as a primer for regrowth of the vegetative cell wall
Endospore inner membrane
Like PM is a good permeability barrier
Endospore core
- Cell components typical of
bacterial cell (nucleoid DNA,
ribosomes, rRNA, tRNA, etc) - Components unique to spore
Contents of endospore core
- Calcium dipicolinate
- Water content
- Small acid soluble DNA binding proteins (SASPs)
- Phosphoglyceric acid
Calcium dipicolinate
- Replaces water
- Absent in normal vegetative cells
- Inserts between DNA bases, helps stabilise the spore DNA and protect it
Water content
Low water and lower pH contribute to dormancy and spore survival
Small acid soluble DNA binding proteins (SASPs)
- High concentrations
- Alter 3D DNA structure fro, B to A form which is less susceptible to UV damage
- Provide amino acids, carbon and energy for biosynthesis
Phosphoglyceric acid
Source of P for ATP generation on germination
Endospore ATP, mRNA, amino acids and protein synthesis enzymes
- Absent or insignificant in endospores
- Reflects lack of metabolism
Seven stages of endospore formation
- Axial filament formation
- Septum formation
- Engulfment of forespore
- Cortex formation
- Coat synthesis
- Maturation
- M0ther cell lyses and releases endospore
Is endospore cell division symmetrical or asymmetrical
Asymmetrical
Coat synthesis
a second impermeable
protective layer forms outside the cortex, and the exosporium forms
Axial filament formation
DNA has replicated but not separated
How long does endospore formation take
8 hours
Genes that control Endospore formation
- spo genes (control of overall sporulation process)
- ssp genes (act inside the endospore, encode SASPs)
- cot genes (act in mother cell, encode spore coat proteins)
How have mutation studies elucidated the processes
controlling endospore formation
mutations in particular genes arrest process at different structural and functional stages
what systems control sporulation
Controlled by phosphorelay in a two component regulatory system and alternate sigma factors
What senses unfavourable environmental conditions (e.g. nutrient depletion)
Sensor kinase proteins such as KinA
KinA
- KinA autophosphorylates
- Phosphoryl group then transferred between
Spo proteins in a phosphorelay - Eventually Spo0A is phosphorylated
spo0A
- Master regulator
- Response regulator protein
- Phosphorylated Spo0A represses genes required for vegetative growth, and activates genes required for
sporulation
Sigma factor genes
- σF (early sporulation gene transcription) and σG (Late sporulation gene transcription)
- Transcribe genes needed in the forespore or the mother cell
Two component regulatory system of control of endospore formation
System with with a sensor protein (KinA) and a response regulator protein (spo0A)
3 stages involved in breaking endospore dormancy
- Activation
- Germination
- Outgrowth
Activation
- Prepares spores for germination
- Requires some form of shock
- Involves damage to spore coat (germination may not occur unless there is coat damage)
- Reversible if a germination agent is absent
Germination
- The dormant endospore starts metabolic activity
- Triggered by water and a germination agent (e.g. sugar or amino acid) penetrating the exosporium and coat
- Germination agent interacts with inner membrane receptors, signal sent to activate germination
Germination events
- Cations secreted
- Spore swells (H2O replaces Ca-DPA)
- Cortex peptidoglycan is degraded by lytic enzymes (core rehydrates)
- Initiation of metabolic activity
- Degradation of coat by proteases
- Loss of heat resistance
- SASPs degraded (supplying amino acids, DNA repaired)
- ATP generated from 3 phosphoglycerate
Outgrowth
- Cell components are made in the core (SASPs act as carbon and energy source - makes proteins, RNA, DNA)
- Protoplast emerges
- Vegetative growth is resumed
