Biofilms and microbiol communities

Question 1

Biofilms in terms of environment

Answer 1

Sites for major microbial activities e.g. carbon production, mineralisation, biofilms on living surfaces.

Catalysts for adaptation and evolutionary events e.g. gene transfer and provision of unique selection pressures.

Biodegradation.

Question 2

Biofilms in terms of industrial

Answer 2

Marine fouling

Fouling of hydroelectric, water reticulation, heat exchange and food processing pipelines.

Corrosion of metal surfaces

Wastewater treatment

Trickling filters.

Activated sludge.

Fluidised-bed reactors.

Bioremediation.

Question 3

What is biocorrosion and what does it do?

Answer 3

caused by bacteria results in pitting, crevice corrosion and stress corrosion cracking.

Question 4

Wastewater treatment pipeline

Answer 4

Primary - Removal of large objects e.g. stones, etc.

Secondary - Activated sludge (flocs)
Trickling filters
Biofilms
Bacteria convert dissolved or suspended solids to settleable solids

Tertiary - Biological or chemical removal of nitrate/ammonia and phosphates
Virus removal
Trace chemicals removed

Question 5

What is a trickling filter

Answer 5

Microorganisms are encouraged to grow on stones/plastic over which the sewage is trickled i.e. to form biofilms. The microorganisms remove dissolved organic carbon through metabolism to CO2 or incorporation as biomass.
Aeration is critical.

Question 6

Biofilms in terms of medical/dental

Answer 6

Dental plaque.
Chronic wounds.
Cystic fibrosis.
Medical implants.

Question 7

Bacterial biofilm as a major barrier to wound healing

Answer 7

Bacteria protected from topical agents

Low oxygen in biofilm niches

host defence unable to clear infection

impaired migration and proliferation of keratinocytes

Bacteria protected from systemic antibiotics

Question 8

Define a biofilm

Answer 8

microbial, sessile community characterised by cells that are:
Irreversibly attached to a substratum, interface or to each other.

Are embedded in a self-produced matrix of extracellular polymeric substances (EPS), and

In comparison to planktonic cells, they exhibit an altered phenotype with respect to:
-Growth rate
-Gene transcription

Question 9

Biofilms refer to many different microbial aggregates which all share the following characteristics:

Answer 9

embedded in hydrogel formed by EPS

Long retention time of cells next to each other = microconsortia

Heterogeneity in space and time

High biodiversity: strong gradients, high habitat variability, subpopulations

Retention of exoenzymes and nucleic acids – large gene pool

Increased resistance to biocides, desiccation + other stress

Access to degradation of particulate matter

Sorption of dissolved and particulate nutrients from environment

Physiological differences between planktonic and biofilm cells

Question 10

parts of Biofilm development to target

Answer 10

Host factors: environmental cues, biosurfactants, Quorum-sensin jamming

Initial adhesion: Biosurfactants, surface blanketing

microcolony formation, biofilm maturation: Adhesiom down-regulation, matrix degradation, quorum sesning jamming

Question 11

Oxygen gradient profile in a biofilm

Answer 11

Anaerobic habitats below aerobic colonies due to respiratory oxygen consumption

Question 12

What is EPS and what does it consist of?

Answer 12

The glue: extracellular polymeric 
substances (EPS) – the ‘house’ of biofilm cells.

Biopolymers of microbial origin
Polysaccharides
Proteins
Glycolipids, phospholipids, LPS
Nucleic acids
Biofilm cells are embedded in EPS, which fundamentally influence their microenvironment.

Question 13

Biofilm morphology: due to presence of alginate..

Answer 13

cells stick together from the beginning and, therefore, form clusters and, later, mushrooms

Question 14

Pseudomonas strains

Answer 14

Pseudomonas aeruginosa SG 81:
mucoid strain, grown in a flow-through cell

Pseudomonas aeruginosa SG 81 R:
Non-mucoid mutant

Question 15

Biofilm life-cycle

Answer 15

Attachment -> proliferation -> biofilm maturation -> cell death and dispersal -> Flagella and type IV pili

Question 16

Differentiaton of biofilms

Answer 16

Bacterial biofilm effluents are autotoxic at time of cell death.
Autotoxic activity is detected in biofilm effluent.
For some bacteria, the differentiation is quorum sensing regulated.

Question 17

Two strains that have multicellular traits

Answer 17

Bacillus subtilis

Pseudomonas aeruginosa

Question 18

Conserved autoinducer signals 
and quorum sensing systems (communication)

Answer 18

Acylated homoserine lactone, AHL (also called AI-1) mediated regulation in Gram negative bacteria.

AI-2 signalling system (luxS, luxR) in Gram negative and Gram positive bacteria.

Peptide mediated regulation in Gram positive bacteria (competency, gene transfer, sporulation).

Question 19

AHL mediated biofilm formation (behavioural aspects that are impacted by quorum sesning)

Answer 19

1. Rhizobium sp. - Nodulation
2. Agrobacterium tumefaciens - Colonisation, Ti plasmid transfer and invasion
3. Erwinia carotovora - Biofilm formation, exoenzyme production and invasion in plant tissue
4. Pseudomonas aeruginosa - Swarming, twitching, biofilm formation, virulence factors, dispersal.
5. Serratia marcescens - Attachment, exoenzymes, swarming (surface motility), biofilm formation, dispersal.
6. Aeromonas hydrophila - Exoenzymes and biofilms (in mixed communities).

Question 21

AI-2 mediated biofilm formation

Answer 21

E.coli

Vibro cholerae

V.vulnificus

Bacillus subilis

Question 22

How do quorum sensing blockers work?

Answer 22

block quorum sensing to stiop biofilms acting as a community

Question 23

Dispersal of biofilm cells

Answer 23

active process of release of cells

Question 24

Recolonisation of biofilms

Answer 24

Small colony variants (SCV) show enhanced biofilm formation and microcolony development

Question 25

Appplications of biofilms

Answer 25

Treatment of biofilms in industrial, environmental and medical settings with autotoxic compounds.

Quorum sensing blockers may be added to systems to reduce biofilm load or used in polymers to prevent attachment and biofilm development.

Addition of compounds that induce early dispersal of biofilms.
e.g. nitric oxide

Question 26

Reversible and irreversible attachment

Answer 26

Forces that operate over
Long distances 5 – 20 nm
Short distances 0.2 – 2 nm

The importance of these depends on attractive van der Waals forces, repulsive electrostatic forces and polymer bridging by bacterial surface structures.

Question 27

‘Attachment’ at long distances

Answer 27

Adhesion at a distance of 5 – 20 nm. A result of forces that operate at long distances, i.e. van der Waals forces

Reversible binding

Little energy needed to remove bacteria e.g. the kinetic energy produced by turning the flagella leads to desorption

Question 28

‘Attachment’ at short distances

Answer 28

Bacteria bind at short distances (0.2 – 2 nm)

a) Non-specific binding
b) Specific binding (irriversable)

Binding is mediated by polymer bridging, achieved by reduced radius of body

Irreversible binding.

Question 29

Bacteria possess a range of surface structures that allow for polymer bridging - name them

Answer 29

Exopolymers (exopolysaccharides, fibrillar proteins)
Fimbriae
Flagella
Stalks
Lipoteichoic acids (LTA)
Lipopolysaccharides (LPS)
Surface localised proteins
Surface localised pigments
A-layers
S-layers

Question 30

Two types of binding using surface structures for polymer bridging

Answer 30

Non- specific irreversible adhesion
- Hydrophobic interaction, ion-, hydrogen-, and covalent binding.

Specific irreversible adhesion = ‘key-lock’
- Defined as structure mediated binding that can be blocked by an analogue.

Question 31

Key-lock medated adhesion

Answer 31

Pili or Fimbraie

or

amfimbrial adhesins (adhesins in bacterial cell wall and host cell membrane has adhesin receptors)

Question 32

Fimbriae mediated binding

Answer 32

Large variations in fimbriation between species

2 – 10 nm in width -> 4 µm in length

Consist of identical protein subunits held together by H-bonds and hydrophobic interaction

Flexible fimbriae have many subunits/turn (so called fibrillae). Rigid fimbriae have fewer subunits/turn

Question 33

Ecological advantages of Fimbriae mediated binding

Answer 33

Gene clusters on plasmids

∼10 genes (in addition to regulatory domains). Gene transfer.

Phase variation = individual cells switch between expression and nonexpression.

∼1/1000 cells per generation.
- Regulation includes quantitative control that responds to environmental conditions.

Question 34

Key-lock binding often occurs on living surfaces because:

Answer 34

Epithelial cells expose
- Lipids: generally glycolipids integrated in cell membrane.
- Proteins: glycoproteins and proteins integrated and peripheral.
- Mucus layer, mucus, lipids and proteins.
And these present:
Receptor sites:

Carbohydrates
-Great diversity of monosaccharides
-Mannose, in di- and trisaccharides, and digalactose are common receptors.
-Bacteria use adhesins (lectins) to bind to internal sequence in the macromolecular chain.

Peptide sequences
- Internal and terminal locations.

Question 35

The bacterial surface has several organelles that facilitate interactions with substrates, including..

Answer 35

curli fibers, pili (also called fimbriae), and flagella

Question 36

roperties of the surface such as _______ interact with physico-chemical properties of bacterial cells and influence ______

Answer 36

charge, hydrophobicity, topography, and the identity of the exposed chemical groups

attachment

Question 37

The proximity of bacteria to a surface causes changes in _____ that are sensed by cells

Answer 37

pH, osmolality, and flagella rotation

Question 38

Receptors that interact with E.coli’s pili on host monocyte and host epithelial cells

Answer 38

Host monocyte - CD48

Host epithelial cell - Mannose, TLR4

Question 39

What is pitting?

Answer 39

Biofilm attaches to the surface of the metal, and as you go down through the biofilm, you get an O2 gradient

outer layer is atmospheric O2 levels, as you go down, you get more and more anaerobic

when you get to the base, the action of the anerobic cells will be to eat into the metal itself

Question 40

mucoid vs non-mucoid

Answer 40

non = doesnt produced mucus with algenate ect.