Lesson 2 Flemming et al., 2016

Environmental Biotechnology

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🌱 What are Biofilms?

Definition: Aggregates of microorganisms embedded in a self-produced matrix of extracellular polymeric substances (EPS) (polysaccharides, proteins, lipids, DNA).
They stick to surfaces 🪨 or float as flocs 🌊.
They show emergent properties — new traits not seen in free-living bacteria.

🌍 Why are Biofilms Important?

Everywhere: Found in water, soil, sediments, subsurface, and inside hosts.
Roles:
- 🔄 Drive biogeochemical cycles.
- 🧼 Used in wastewater treatment & biocatalysis.
- ⚠️ Cause persistent infections, contamination of implants, corrosion, and biofouling.

🧩 The EPS Matrix: The “House” of the Biofilm

Mostly water (up to 97%).
Components: polysaccharides, proteins, extracellular DNA, amyloids, cellulose, pili, flagella.
Functions:
- Provides architecture and stability 🏰.
- Creates pores/channels → nutrient flow like a “rudimentary circulation system.”
- Protects against desiccation (hydrogel barrier 💧).
- Stores enzymes → acts as an external stomach 🍽️.

🏞️ Biofilms as Habitat Formers

Like corals, beavers, or forests 🌳, biofilms create new habitats at microscale.
They shelter bacteria, trap nutrients, and change local environments.

🧪 Resource Capture

The EPS matrix acts like a sponge 🧽:
- Sorbs nutrients, metals (Ca²⁺, Fe²⁺, Mn²⁺), organic & inorganic particles.
- Even traps pollutants (antibiotics, pharmaceuticals, benzene!).
Recycling ♻️: dead cells release DNA & debris, reused as nutrients.
Supports lithification → rock formation (stromatolites 🪨).

🍽️ External Digestion

Enzymes secreted into EPS stay trapped, not diluted like in free water.
Matrix acts as a communal digestive system.
Enzymes stabilized against heat, drying, and proteolysis → long activity.
Benefits the whole community, not just the secreting cell → a shared kitchen 🍳.

Biofilms are not uniform:
- Gradients of oxygen, pH, nutrients form layers (aerobic at top, anaerobic below).
- Cells adopt different states: active, dormant, persisters, or dead.
Social interactions:
- Cooperation 🤝: sharing metabolites, nitrification chains, synergistic pollutant degradation.
- Communication: quorum sensing (chemical signals) & even electrical signals ⚡ via ion channels/nanowires.
- Competition ⚔️: antibiotics, toxins, vesicles, secretion systems.
Overall: a microscopic society with politics, teamwork, and conflict.

🏰 Biofilm as a Fortress

Tolerance vs Resistance:
- Tolerance: temporary, lost if bacteria disperse (due to slow growth, EPS reactions).
- Resistance: permanent genetic changes (mutations, plasmid exchange).
Mechanisms:
- EPS neutralizes antimicrobials (diffusion-reaction inhibition).
- Dormancy → cells survive attacks (persisters, VBNC state).
- Horizontal gene transfer skyrockets in biofilms (plasmids, eDNA, secretion systems).
Net effect: antibiotics often fail → persistent infections ⚠️.

🔮 Future Directions

Need deeper study of EPS diversity and regulation.
How do biofilms share, store, and partition captured resources?
Can we manipulate EPS to prevent biofouling or infections?
Big unknowns: role of dormancy, inter-kingdom biofilms (with fungi, viruses, protozoa).

📌 Quick Recap (Mnemonic Style)

Think of biofilms as “H.A.R.D. S.H.E.L.L.s” 🐢:

Habitat formation
Architecture (EPS)
Resource capture
Digestion (external enzymes)
Social interactions (cooperation & competition)
Heterogeneity (gradients & niches)
Enhanced tolerance/resistance
Lithification & recycling
Long-term persistence
Shield (fortress against threats)

Quiz

Score: 0/30 (0%)

Q0. What is the main structural component that defines a biofilm?

Flagella

Extracellular polymeric substances (EPS)

Lipid bilayers

Capsule polysaccharides

Q1. Which of the following best describes an emergent property of biofilms?

A property predictable from single-cell studies

A unique feature that arises only when cells form a community

A genetic mutation common to biofilm-forming bacteria

A random environmental artifact

Q2. Which of the following is NOT typically a component of the EPS matrix?

Polysaccharides

Extracellular DNA

Proteins

RNA

Q3. What is the approximate water content of most biofilm matrices?

10%

50%

80%

97%

Q4. Which function does extracellular DNA (eDNA) play in the biofilm matrix?

Acts as a metabolic energy source only

Crosslinks structural polymers and provides stability

Serves as the main carbon source

Acts as a membrane transport molecule

Q5. What role does the EPS matrix play in desiccation tolerance?

Acts as a hydrogel retaining water

Repels water to reduce swelling

Enhances DNA uptake from environment

Increases oxygen diffusion

Q6. What ecological analogy is used to describe biofilms in Box 1?

They are likened to coral reefs and forests as habitat formers

They are compared to deserts

They are compared to single-celled algae

They are likened to sand dunes

Q7. Which of the following processes is enhanced in biofilms due to cell proximity?

Transcription

Translation

Horizontal gene transfer

Chromosome replication

Q8. Which mechanism allows Pseudomonas aeruginosa to maintain structural integrity in its biofilm matrix?

Hydrophobin proteins

Pel polysaccharide crosslinking eDNA

Curli protein fibers

Chitin synthesis

Q9. Why is EPS production considered energetically expensive yet evolutionarily beneficial?

It reduces gene expression cost

It ensures faster replication

It provides essential structural and protective functions

It enhances motility

Q10. What allows biofilms to capture nutrients more efficiently than planktonic cells?

Active transport

Diffusion barriers

Sorption by EPS polymers

Flagellar motion

Q11. What describes the biofilm’s role as an 'external digestive system'?

Biofilms import digestive enzymes from the environment

Retained extracellular enzymes digest external substrates

Bacteria internalize large molecules directly

Only intracellular digestion occurs

Q12. How does quorum sensing benefit from the biofilm matrix?

Signals diffuse away rapidly

Matrix concentrates signaling molecules

Signals are blocked by EPS

It suppresses all communication

Q13. Which process is an example of cooperative metabolism in biofilms?

Oxygen diffusion

Nitrification involving ammonia- and nitrite-oxidizers

Flagellar rotation

Binary fission

Q14. Which mechanism contributes most to antibiotic tolerance in biofilms?

Rapid metabolism

EPS-mediated diffusion–reaction inhibition

Genetic mutations

High oxygen exposure

Q15. Biofilms are defined as aggregates of microorganisms embedded in a self-produced matrix of EPS.

True

False

Q16. The majority of a biofilm’s mass is composed of microbial cells.

True

False

Q17. EPS molecules are static structures that do not change once formed.

True

False

Q18. eDNA contributes to biofilm architecture by crosslinking with polysaccharides.

True

False

Q19. Biofilms can retain extracellular enzymes, effectively forming an external digestion system.

True

False

Q20. Gradients of oxygen and nutrients are uniform throughout the biofilm.

True

False

Q21. Electrical signaling has been observed as a form of communication within biofilms.

True

False

Q22. Slow growth and dormancy in biofilms make some cells tolerant to antibiotics.

True

False

Q23. Horizontal gene transfer is less frequent in biofilms compared to planktonic states.

True

False

Q24. The EPS matrix provides no role in metal ion binding or stabilization.

True

False

Q25. Mixed-species biofilms can show higher stress tolerance than single-species biofilms.

True

False

Q26. Biofilms are major contributors to lithification and sediment stabilization.

True

False

Q27. Antimicrobial tolerance in biofilms is a heritable trait that persists after dispersal.

True

False

Q28. Persister and VBNC cells contribute to the survival of biofilms under stress.

True

False

Q29. The study emphasizes that future research should focus on interkingdom biofilm interactions.

True

False