biofilm

Question 1

What is a biofilm

Answer 1

A biofilm is a layer of micro organism such as bacteria or alien that are surrounded by self produced mucus attached to a surface. The different micro organism are held together by an extracellular matrix.

It is marked by the excretion of extracellular polymeric substances (EPS° and they can exist as multiple or single microbial species.

mixed biofilms are predominantly found in environmental settings and single biofilms are most of the time found in infection related biofilm

Question 2

where are biofilms found

Answer 2

they can be formed on solid substrates in contact with moisture, on tissues of living organism and at liquid air interfaced.

These can then be found anywhere from the aquatic environment such as on algae, rocks ships, in industry setting such as pipelines or plastics and in medicines on catheters, wounds, teeth etc.

Question 3

beneficial biofilms

Answer 3

for waste water treatment, for bioremediation of contaminated soils (bacteria degrade it), for microbial leaching (use of bacteria to remove compounds such as copper).

In public health it is beneficial in human ear for protection against infection, in vagina as protective role, in the digestive tract for protection against opportunistic pathogens.

Question 4

step by step of biofilm formation

Answer 4

pseudomonas adhere to the surface (reversible) (MICROBIAL conditioning)
Microbial attachement
after a while, the adhesion becomes irreversible and

Biofilm development
from micro colonies you can see EPS formation. Once the colonies are covered with EPS, there is an irreversible binding and part of the

detachment
biofilm can be sheared and adhere somewhere else.

Question 5

which factors are involved In biofilm formation

Answer 5

Conditioning of substrate
microbial attachement
microbial accumulation
biofilm growth and maturation
dispersal and reattachment

Question 6

conditionnant of substrate

Answer 6

Environmental conditions are important for initial adhesion.

It is dependent on hydrodynamic flow, shearing force and the surface characteristic such as roughness, topography etc.

Question 7

Microbial attachement

Answer 7

the surface chemistry is also important such as the charges, the electrostatic interactions, acid base interactions

on conditionnied materials or tissues, ligand receptor binding I relevant aswell, ph too

For exemples, the coating the coating of titanium with human serum albumin decrease adhesion of a staphylococcus

some bacteria can be ore hydrophobic than others. Some MO can attach to hydrophobic surfaces through a process called van Der Waals forces which are attractive forces between molecules.

Question 8

microbial accumulation

Answer 8

depends on cell division
the production of EPS aching the bacteria to each other and the substrate
the formation of the complex 3D communities.

The EPS is secreted by the bacteria and is made of plenty of different glycans, poluglutamic acid, eDNA (bacteria that are dead and their DNA which is free)

Question 9

biofilm maturation

Answer 9

is dictated by environmental conditions (different forms of biofilms depending on the environment and varying morphology are seen)

depends on nutrients available (if less nutrients, is takes more times to form biofilms and the final structure will e very different. If sufficient supply, the biofilm is very compact)

the presence of other organism
physics chemical properties of the surface
the process related by the organism
hydrodynamics of surrounding bulk phases

the mature biofilms has a structural heterogeneity with interstitial spaces, with the ability of internal mass transports. It makes it protective against many other microbial agents and hots defense mechanism.

Question 10

Dispersal and reattachement

Answer 10

the bacteria can feel good if it is a good or bad biofilm maturation. It can go out of the biofilms and disperse again if not good.

It can disperse in different way: it can be fluid associated though swarming (use of flagella) and clumping (clumps together to get away)

it can be surface associated by twitching (go to the surface and use type 4 vili to move over the substrat), it can rot (micol acid) and it can slide

the detachment can occurs passively or indices by the bacteria which produce product to stimulate detachment

Question 11

bacterial communication

Answer 11

they use quorum sensing

the cells can sense their own population density. It can be intra or interspecies and they perceived other cell in the community through signal molecules secreted by the bacteria
it regulates gene expression that affect various physiological process such as biofilm formation, virulence factors expression, antibiotics productions,exo enzyme etc

when there is an increase in cells density - there is more signal molecule = autoinducers

the level of signal molecules trigger the gene expression required to a minimum bacterial populations (= quorum)

Question 12

quorum sensing

Answer 12

Quorum sensing is the regulation of gene expression in response to fluctuations in cell-population density. Quorum sensing bacteria produce and release chemical signal molecules called autoinducers that increase in concentration as a function of cell density.

a signal molecule can be homoserine lactone (HSL)
it is a type of small molecule that is produced by many different bacteria and is used to coordinate a variety of behavior such as biofilm, virulence and antibiotic production

when it reaches a certain threshold, it activated specific genes In bacteria that responds to the signals. This can results in changes in the gene expression and the production of various proteins and enzymes that allow them to coordinate together (eg luciferace in vibrio fisher)

Question 13

QS in virbrio fisheri

Answer 13

i is a gram negative bacterium that leaves in symbiont with fish, jellyfish or squid.

They are present in the light organs with is a nutrient rich environment for the bacteria. At a high bacterial concentration, they start to produce Luciferase and therefore luminescence.

Once the light organ emptied, the bacteria start to grow again until they reach the quorum

the signal molecules bind to the regulators which is an activator of the luciferase

Question 14

signal molecules in gram negative

Answer 14

HSL is the signal peptide - acyl homoserine lactones more specifically.

it diffuses through the ells membraned through envelope or assisted by transporter.

Question 15

signal molecules in gram positive

Answer 15

they use oligopeptide which creates cysteine bridges. they are auto inducing peptides.

Gram positive bacteria have an additional layer, they therefore need to use active transport through the ABC transporter to get the oligo peptide out.

they can then bind to sensors kinase which phosphorylate a response regulator that leads to the transcription of target genes. (this is a two component signal transduction system (TCSTS)

Question 16

interspecies communication

Answer 16

a test was done with multiples bacteria and a vibrio Harvey mutant (this one cannot be self activates - so needs other bacterial signal molecules) they discovered that it was glowing in the presence of two pathogenic bacteria!

there has been seen interspecies communication via the AL-2 molecules (alloinducers)

this AL-2 molecules is induced by LuxS homologues, which are found in many gram positive and gram negative species

QS of LuxS can therefore give an indication of expression virulence factors/biofilm formation.

Question 17

biofilm and antibiotic resistance

Answer 17

Why is there more and more resistance to antibiotic therapy

1) delayed penetration

the EPS is basically a diffusional barrier, it influence the rate of transport of molecule inside the biofilms and EPS can actually react to an antimicrobial by neutralizing it

2) there is an altered environment. In the inner layer of the biofilm, there is a depletion of n nutrients. The biofilm embedded cells grow significantly slower which means they ahem to reduce their metabolic activity. This decrease their susceptibility to antimicrobials and their effectivity. (dont forget that biofilm is characterized by a chemical heterogeneity - the outside of the biofilm is not the same as the inner layers)

3) stress response - biofilm embedded cells can induce a stress response on time by antiipitation of antibiotic action. The platonic cells will get overwhelmed by the antimicrobial challenge but for biofilms, the diversity in gene expression allows to give time to anticipates and up regulate the stress response with an effective deployment.

4) persister cells (super fits) - these are specialized survivors that are in a formant states. they are phenotypic variants of the wild type (not mutant) that make out for 0.1 to 10% of the cells in planktonic cultures and in biofilm. Their formation is largely dependent on toxins and antitoxins modules that reversibly block important process. This makes them highly resistant to antibiotics

the role of these cells in plantokinc cultures to be free floating cells that can rapidly revert to a susceptible state

in a biofilm, their role is to easily accumulate and be retained in the EPS. After antibiotic levels drops the persisters cells become active and can repopulate the biofilms :)

5) horizontal gene transfer - there is a possibly to exchange genes. The bacteria can take up DNA and transform, conjugate between bacteria or use the transduction process. In biofilms QS is used to activate genetic competence that facilitate gene exchange between species ( up to 600 fold more competent in a biofilm)

Question 18

biofilm resistance to host response

Answer 18

For planktonic cells, you can use antibodies or antibiotics and the immune cells will recognize them. However, with biofilms these agents cannot reach the bacteria dand the Immune system either. The IS gets overwhelm, keep producing cytokines and chemokine which leads to break down of tissues

the advantages of biofilm is that they are fixed to a surface, they have the EPS proteins, they can work with a social behavior, adapt to different environmental conditions

they basically greatly resistance to antibiotics and the immune system, which results in chronic and persistent infection.

Question 19

BIOfilms infections on host tissues

Answer 19

topically are metal plaques shish can lead to caries, gingivitis, periodontitis

CARIES - in the dental plaques biofilms, there is more than 1000 different species. they varies in composition in function of their locations and need salary components to form biofilm. the growth of the metal plaques is made up of different bacteria.

in caries, you see the formation of cariogenic biofilms?. They cause localized destructions of dental hard tissues by acidic by product (bacterial fermentation of dietary sugars). At pH lower than 5.5 you get demineralization of enamel.

GINGIVITIS - PERIONDITITS - usually caused by gram neg anaerobes that induce inflammation of periodontal tissues. because more gingival fluid is produces, this is nutrient for bacteria and start growing. They case bone loss around teeth and finally can loose it (as well as gum)/ RF include smoking, pool controlled diabetes

OTITIS MEDIA - it is a middle ear infections. It is usually infections that originally cause sore throats, cold etc that then spread. Children are more affected due to their sell IS developing, their Eustachian tube being shorter and straighter and larger adenoids that can interferes.

In can of chronic OM, you can implant a tympanovstomy tubes that helps with the pressures builds up. However, the tubes are also vulnerable to biofilm formation.

URINARY TRACT INFECTIONS - A MAJORS CauSE IS ECHERICHI COLI STRains
. THEY ARE GENERALLY CONSDERED AS ACUTE SELF LIMITING INFECTIONS. HOWEVER, THERE IS A PREVALENCE TO RECURRENCY. there are different stages - invasion of superficial , large umbrella cells, replication in cells and formation of intracellular bacterial community with biofilm like traits, flagellation of bacteria and then spreadsThe granulocyte are recruited but they fail to gain access to the IBC

CYSTIC FIBROSIS - LUNG INFECTION - they have a mutation in CFTR gene which leads to a defective cAMP dependent chloride channel, which decrease the airway surface liquid volume and increase in mucus secretion. The thick mucus layer can be colonized by a number of pathogens. different colonizers will develop in the thin k mucus . CF patient are predisposed for pseudonomonas aeruginoza colonization and biofilm formation.

P Aeruginosa ssems to blunts the immune repose by down regulate the production of highly immunogenic fledgeling and by converting mucoid phenotype (by producing large amount of alginate EPS that attenuate the immune repose.Over time, accumulation promote persistence in CF lungs

WOND INFECTIONS - contaminating - non replicating bacteria

the you see colonization - replicating MO without injury to the host

infection - replication of MO with a wound causing host injury

Question 20

biofilm infection on medical devices

Answer 20

everything that enters the body are basically exposed to bacterial biofilm formation

CONTACT LENSES - they really adhere easily to the lens and also contact storage. they can ends up to cause keratitis - S aureus

CATHETERS - staphylococcus areus. they can come from the baxter, if not cleaned correctly, the bacteria form the blood vessels can stay attached to the catheters and form a biofilm that attract other bacteria

Question 21

prevention biofilm infection

Answer 21

1) prevent initial attachment
by using various materials, modifying their surface properties, hydrophobic, roughness but not the best. Therefore use of anti adhesive surfaces, which are coated with slowly releasing antibiotics and antimicrobial compounds, as well as heparin

use of anti adhesions antibodies (adhesion analogues that blocks the binding sites, receptors analogues, however, bacterial adhesion to cell surface can be very complex and variables

2) target EPS - can use alginate that is known to disrupt the matrix produced by certain bacteria however EPS is different in function of species so hard to find a general solutions.

“3) prevent QS - basically do quorum quenching

either inhibit signal molecules synthase (LUXL homologues)
degrades the signal molecules
block the signal receptors sung molecules analogues

No occurrence of bacterial resistance due to absence of selective pressure????