Lecture 5 - Bacterial endospores Flashcards
What are endospores and their function?
- Dormant and most resistant biological structure
- Function = resistance, survival, dormancy allowing later germination
Describe the taxonomy of endospores and list important genera
- Low GC G+ve bacteria = Firmicutes from common ancestor
- 12 families, 40 genera incl Bacillus, Clostridium, Sporosarcina
List the locations of endospores in C. tetani, B. anthracis, C. botulinum
- C. tetani = terminal
- B. anthracis = central
- C. botulinum = subterminal swollen
How do endospores set the sterilisation standard?
- Sterilisation must kill endospores by B. subtilis spores confirmation
- Autoclaving = 121 C, 15 psi, 15+ mins to kill all cells and endospores
List the pathogenic endospore-forming bacteria of medical importance and their diseases
- C. botulinum = botulism
- C. tetani = tetanus
- C. perfringens = gas gangrene
- C. difficile = antibiotic associated colitis
- B. anthracis = anthrax (lungs then death)
What endospore-forming bacteria was used for bioterrorism?
B. anthracis
How are endospore forming bacteria used in developmental biology?
To understand bacterial differentiation
What bacteria is used in agricultural pest control and how?
B. thuringiensis as insect biopesticide producing toxin protein crystals
List the endospore structures from outermost to innermost
Exosporium, spore coat, cortex, core wall, inner membrane, core
Describe the S+F of the exosporium
- Thin glycoporein layer
- Hairs or nap and basal layers
- Adhesion to soil or intestinal cells, barrier to penetration by large molecules
Describe the S+F of the spore coat
- 4 keratin like protein layers with S-S bonds and 70+ proteins
- Basement, inner, outer, crust layers
- Impermeable and resists chemicals
Describe the S+F of the cortex
- PG with reduced NAM residue cross-linking (7%)
- Function = core dehydration for dormancy and high temp resistance, loose cross-linking for expanding and contracting in humidity
Why is there reduced NAM cross-linking in the endospore corex compared to vegetative CW?
- 50% NAMs -> muramic lactam (MAL) = no peptide addition
- 15-25% NAMs with shortened peptides (L-ala) = no cross-linking
Describe the S+F of the core wall and inner membrane
- Peptidoglycan CW without teichoic acids
- CW = osmotic stability, primer for regrowth
- Inner membrane = permeability barrier
List the 6 components of the endospore core and what their function is
- Organelles
- High Ca-DPA = bind water for dehydration + heat resistance, inserts beween DNA bases = stabiliy + protection
- Low water and pH = dormancy + survival
- High small acid-soluble DNA binding proteins (SASPs) = saturate DNA + alter 3D A to B = UV protection
- High phosphoglyceric acid = source of P for ATP later
- Absent ATP, mRNA, aa’s, enzymes etc = no metabolism
How long does endospore formation take, what are the stimuli and when in the cell cycle does it occur?
- 8hrs
- Environmental, metabolic, cell cycle stimuli
- Stationary phase
Describe the 7 steps in endospore formation
- Nucleoid divides + stretches = axial filament
- Septum forms asymmerically between forespore and mother
- Engulfment = mother’s PM around forespore = double membrane
- Cortex wall PG made
- Coat layer = forms outisde cortex then exosporium
- Maturation = coat thickens
- Mother lyses = endospore released
What is the trigger for sporulation and how is it imitated in the lab?
- Unfavourable conditions = absen nutrients, desiccation, toxins etc
- High density culture centrifuged, filtrate with quorum sensing oligopeptides added to low density culture
List 3 important genes controlling endospore formation and what they do
- Spo = controls overall
- Ssp = SASPs inside endospores
- Cot = coat proteins in mother
What is the process used to control sporulation, what are the components involved and what does it ensure about sporulation?
- 2-component phosphorelay regulatory system + alternate sigma factors
- KinA sensors and Spo0A responders
- Sporulation only occurs when survivability threatened
Describe the 3 step process of the 2-component phosphorelay regulatory system with alternate sigma factors
- KinA sense unfavourable conditions, autophosphorylates, phosphoryl -> Spo proteins -> Spo0A
- Spo0A = master regulator TF represses vegetative and activates spore genes
- Genes incl sigmaF and G = transcibe genes for forespore and mother
What are the 3 steps for germination to occur?
- Activation/prep = shock and damaged spore coat
- Germination = start metabolism after water and germination agent penetrae exosporium and coat
- Outgrowth = cell components made, protoplast emerges = vegetative growth
List the events in the germination cascade
- Cations secreted for ATP
- ATP from 3-phosphoglycerate
- Water replaces Ca-DPA = swells
- Cortex PG degraded = rehydration
- Initiate metabolism and acivate enzymes
- Proteases degrade coat
- Loss of resistance
- SASPs degraded = aa’s and DNA repaired
What are the current findings/experiments of endospore survival?
- 25-40 million year survival of B. sphaericus in amber preserved bee
- 250 million year old bacterium in salt crystal
- 500 year experiment with B. subtilis spores to be checked every 25 years
