VL14: Biofilms

Question 1

Define biofilm

Answer 1

Surface attached microbial communities composed of microorganisms and a matrix of extra-polymeric substances (exopolysaccharides, proteins, and nucleic acids)

Question 2

Why do bacteria form biofilms?

Answer 2

• self defence (resist physical forces that sweep away unattached cells, resist phagosytosis by immune system cells and resist penetration of toxins (antibiotics))
• allows cells to remin in a favorable niche (help trap nutrients and prevent detachment in flowing systems)
• co-operative interactions

Question 3

Where can biofilms form?

Answer 3

• solid substrates in contact with moisture
• soft tissue surfaces in living organisms
• liquid air interfaces

Question 4

What are the steps in biofilm formation?

Answer 4

1. Planctonic stage
2. Attachment (reversible, then irreversible)
3. Microcolony formation
4. Macrocolony/ cells form extracellular matrix
   (5. Dispersal)

Question 5

Explain how to figure out if your bacteria have formed a biofilm + simple was of characterization

Answer 5

crystal violet assay for biofilm
(grow cells, wash to remove planktonic B, stain remaining cells to visualize attachment pattern
can be used for semi-quantitative assessment)

describe morphology, microscopy

Question 6

How is biofilm formation regulated? Describe 1 vibrio cholerae mutant, and name one
Name one (or three) of the structures (genes) important for biofilm formation (in vibrio cholerae)

Answer 6

many genes/ reg pathway/ environmental sensing neccesary for biofilm formation, integrated in other aspects of the lifestyle

type 4 pili, if you knock out mshA growth is significantly reduced)

(spo0A is master regulator of sporulation) deletion of spo0A inhibits biofilm formation
sigF = sigma factor, forms different kind of biofilm

Question 7

How do biofilms impact human health?

Answer 7

biofilms are THE major bacterial lifestyles

medical/ dental conditions
chronic infections
antibiotic resistances 
treatments are expensive and inefrfective
infections in indwelling medical devices

Question 8

What about showercurtains?

Answer 8

good place for biofilms, potential pathogen source

Question 9

What about dental plaques?

Answer 9

1 acidic film of glycoproteins on teeth
2 colonization by streptococcus
3 bacteria of different species stick to early colonizers
4 multispecies biofilm

Question 10

and kidney stones?

Answer 10

15-20% involve UTI
bacterial infection, biofilm, mineralization
produce urease -> crystal formation

Question 11

and cystic fibrosis? how become biofilm bacteria tolerant?

Answer 11

in CF pseudomonas infections often result in ab resistant biofilms, p.aeruginosa makes extrac/ polysaccarides alignate Pel and Psl

B in biofilms may be tolerant to antimicrobial agents (don’t grow but survive)

-restricted antimicrobial diffusion, differential physiological activity and induction of specific tolerance mechanisms contribute to tolerance

Question 12

What’s the problem with pseudomonas biofilms and antibiotic therapy?

Answer 12

some ab kill only metabolically active cells
colistin interferes with membrane structre/function

only combined treatment gets rid of most bacteria

Question 13

What about biofilms and hearts?

Answer 13

biofilms can develop on heart valves and surrounding tissue = prosthetic valve endocarditis

Question 14

Whats the simple most important medical problem with biofilms?

Answer 14

urinary catheters

can result in bacteremia

Question 15

Which different areas of life are affected by biofilms?

Answer 15

cooling water in power plants
food processing
medical implants
paper manufacturing
drinkng water
oil recovery
teeth

pipe plugging, corrosion, water contamination

biofilms help degraide soil bound contaminants, help cycle nutrients (increase agricultural productivity)

Question 16

What are the key components of the biofilm extracellular matrix?

Answer 16

polysaccharides
proteins
dead cells
dna

Question 17

What does rugose mean?

Answer 17

runzelig (wild type surface of v.cholerae biofilm)

Question 18

what are important matrix proteins (v.cholerae)? What happens if you knock them out?

Answer 18

RbmA RbmC Bap1
if you knock one of them out, biofilm can still form but differently,
if you knock them all out no biofilm forms

Question 19

Which molecule is important for biofilm regulation? How is it made? How degraded? With what receptors does it interact and what are the results?

Answer 19

c-di-GMP (cyclic-di-GMP) remember name!
made by Diguanylate cyclase (contain a GGDEF domain)from 2GTPs
degraded by phosphodiesterases, sensed by receptor proteins

level of c-di-GMP determines if biofilm is formed
it interacts with transcription factors, riboswithches, PilZ domains, Degenerate GGDEF and EAL domains

and what are the outputs?
biofilm +, motility -, virulence -, cell cycle +

major regulator of biofilm formation AND a major global regulator

Question 20

What determines how much c-di-GMP is present?

Answer 20

affected by many different environmental factors
light, oxygen, surface, arginine, nitiric oxide ect

most importantly QS

cell density via QS -> cyclic di-GMP -> Phenotypic changes

Question 21

describe 3 requirements of biofilm growth

Answer 21

attachment (help of adhesin or VPS (vibrio polysaccharide)

requires cell-cell communication (QS)

Requires coordinated gene regulation (cyclic di GMP, regulates polysaccharide formation and switch between motile and biofilm growth)

Question 22

1 example of preventing biofilm infections

Answer 22

seaweed produces furanones and remains slime free

Question 23

Summary of Biofilms

Answer 23

Biofilms are a community of microorganisms embedded in an organic polymer matrix consisting of polysaccharide, proteins and DNA

Biofilms are prevalent in medical, industrial, environmental, agricultural settings

-C-di-GMO, a nucleotide-based intracellular signaling molecule is critical for planktonic to biofilm life-style switch

Question 24

3 important characteristics of biofilms

Answer 24

• usually highly resistant to antibiotics ( high or long term doses are needed to fight biofilms)
• help horizontal gene transfer (closely interacting communities of bacteria including extracellular DNA and dead cells)
• Protect bacteria from phagocytosis (by extracellular matrix, causes phagocytes to release proinflammatory enzymes and cytokines -> inflammation and tissue destruction)