Week 4: Biofilms Flashcards
Define a biofilm?
community encased in a extracellular matrix, adhere to each other or a surface.
highly regulated process
structured community of microbial cells encased in an extracellular matrix and adhere to each other or a surface
a ubiquitous behaviour of bacteria and fungi
Why would microbes form biofilms?
- Adhesion - stay where want to be
- Access nutrients - accumulate on surface
- Collective behaviour- cooperative outcomes, chaired labour, protection from predators
- Transfer between environments - shedding as a group to allow establishment in a new niche. e.g. vibrio cholera
- Facilitate genetic exchange- close contact and eDNA in environment
What is the biofilm cycle?
microbe sense (through mechanosensors) and detect when on surface.
trigger cascades of gene expression, allow matrix molecules to accumulate.
matric molecules allow a habitat that allow the microbes to grow, divide and thrive by attracting nutrients. Protecting them from external threats such as chemicals. Internal threats from our immune system, when they are larger the immune cells cant detect and engulf them.
Once they are in conditions which are favourable to remain.
Either move as clumps or individuals, depend on what is happening.
planktonic cells –> biofilm towers
What is the role of biofilm matrix?
Structure to the community and establishes heterogeneous environments with gradients innutrition, pH, oxygen
Adhesion to neighbours and / or surface
Desiccation tolerance
Allow emergent properties to develop - shielding from immune cells
Space filling in the community
Communal digestion system - trapping nutrients and enzymes
Scaffold for minerals to protect cells from shear force
Absorb water - help with protection
Surface spreading - pressure building
What molecules are found in the matrix?
eDNA and eRNA
Polysaccharides Proteins - adhesions on cell surface, many have fibrous/film formation tendencies
Lipids
Ions and minerals formed by local biomineralisation processes
Host proteins
Water
glycoproteins
What is the purpose of big fibres of proteins in the biofilm matrixes?
structure
allows some cells to become close together.
What are gradients within the biofilm?
aerobic/oxygen gradient.
outside more oxygen and less towards the centre.
How do microbes benefit from one another in a biofilm?
division of labour
What are key features of bacillus subtilis?
- Gram-positive bacterium
- Soil-dwelling
- Shows differentiation of cell fate during growth
- Spore forming
- Genetically tractable
How do Bacillus subtilis grow?
starts from single colony
grow as a collective
division of labour
in order to be able to survive and thrive in an environment.
What populations does division of labour occur?
isogenic or clonal
How did the study label different promoters of bacillus subtilis?
label each one green or red
they have the ability to have the both promoters activated.
heterogeneity
How are 3 ways to study bacillus subtilis biofilms?
colony
pellicle
submerged
Can you track the bacillus subtilis biofilms?
YES
How can you visualise the bacillus subtilis macrostructure?
merge the bright field and GFP microscopy and can see the structure
see how molecules are shares and how cl sae distributed within the colony biofilm.
Is the bacillus subtilis biofilm hydrophobic?
HIGHLY
What are 3 molecules required to make the bacillus subtilis matrix?
ESP
TasA
BslA
What gene encodes TasA?
TapA-sipW-tasA
What happens if you mutate the gene encoding EPS, TasA or BslA?
lack the structure
lack the ability to grow up and come off the surface of the agar plate
What gene encodes ESP
epsA-O 15 member operon
Are the ESP, TasA and BlsA mutants the same phenotype?
no, provide different structures to the community
What are root interactions dependent on?
matrix molecules to allow the bacteria to adherence to the plant root.
this adherence is a form of a biofilm.
What is EPS?
soluble polysaccharide, exerts pressure to expand the community (biofilm matrix).
What happens if you overproduce the biofilm matrix molecule EPS?
hyperwrinkled biofilm phenotype .
cells clump together
What happens of you underproduce EPS phenotype?
remove biofilm formation.
What is TasA?
protein fibres
What is the role of TasA?
gold can bind
make a network
help connect different cells together.
forms fibres in external environment
What microbe has TasA?
bacillus subtilis
Do all microbes share the same structures?
No
exact molecules differ between species
protein fibres differ
matric components differ
What is the struggle with making one molecules to target all microbes?
The components of the matrix differs.
needs to be specific to microbes
What is an experiment which shows the shared resources of the matrix?
- bacillus can divide its labour shown y the red and green staining
division of labour
- combination of eps and tasA mutants/deletion.
mutants are able to restore the complex biofilm
they can share the molecules and work together.
public goods
How else would you describe division of labour?
public goods
How can you keep the bacillus subtilis matrix contained?
add water droplet as it is hydrophobic
impermeable surface for gases
Is bacillus subtilis the only hydrophobic microbe?
no, all microbes are hydrophobic.
What protein is needed to maintain b. subtilis hydrophobic?
bslA
What happens when you mutate bslA?
wetting of biofilm
unstructured and fully wetting biofilm.
What can BslA form?
layer around the b. subtilis community
When does BslA respond?
responds environmentally
structural rearrangement
Why can the BslA rearrange?
BslA forms a well-ordered and robust 2D rectangular lattice on a surface
- BslA exposes its hydrophobic cap at an air-water interface
rearrange a set of hydrophobic amino acids at a particular region on each monomer.
monomers array together to form a film.
Is BslA a public good?
YES
Why would microbes want to regulate transcription?
metabolically expensive
What might microbes use to control transcription?
sigma factors
2-component systems
What are ways to look at transcription? population level…
NGS
microarrays
northern blots
population level methods.
What is the problem with NGS for population level?
you would know if some of the population are wearing glasses.
wouldn’t know if 2 people have a really big pair of glasses or who has no glasses.
no individual level info
How can you look at single cells?
flow cytometry
What is the advantage of single cell transcription analysis?
look at individual geno/phenotypes
then look at population and understand population in more detail.
How to examine transcription in the biofilm?
grow biofilm over time and analyse them.
flow cytometry or microscopy.
How to carry out flow cytometry?
collect material
break it up
flow through cytometer
It detects the florescent signal form each cell.
look at heterogenous gee transcription.
GFP on/off against cell number.
How to carry out microscopy?
stain for gene expression
What info can you get from single cell and population level analysis?
retain the spatial organisation via imaging
The matrix components are produced by a subset of cells.
The macromolecules can be shared in the community.
division of labour in a clonal population.
Describe how biofilm formation is a communal process?
- working together through the Differentiation of isogenic cells.
- Production of “common goods”
–> biofilm matrix
—> Biofilm matrix and biofilm coat.
How can B. subtilis be used in industry?
How to get it to form biofilms and more efficiently.
- Biocontrol
- Exploiting matrix molecules
- Probiotics (b. subtilis)
- Microbes as surface cleaners
- New discoveries…..?
- BslA is a waterproof fabric
