Lecture 3: Quorum Sensing Flashcards
1
Q
Quorum sensing
A
- is a form of cell-to-cell communication: enables single-celled organisms to participate in cooperative group behaviour (allows them to monitor their own population density)
- characterized by the secretion and detection of small molecules (termed autoinducers) within a bacterial population (either by diffusion or active secretion)
- detection results in coordinated behaviours once a quorum is reached
- methods of secretion and detection vary depending on the precise nature of the autoinducer molecule
2
Q
Why do bacteria (and fungi) perform quorum sensing?
A
- bacteria produce many secreted factors that are important for growth and/or survival, or play a role in infection - PROTEASES and HAEMOLYSINS
- proteases break down host cell proteins, providing carbon source for growth (can also break down parts of the immune response)
- haemolysins lyse red blood cells, releasing a source of iron for bacteria
- individual bacterial cells cannot produce sufficient quantities to gain much benefit
- however, populations of bacteria can produce sufficient quantities of these factors into the extracellular environment, if they co-ordinate their behaviour through quorum sensing
- therefore, proteases and haemolysins are under the control of QS
3
Q
AHLs/HSLs: Acyl homoserine lactones
A
- Two main family of proteins make AHL: LuxI and LuxR
- AHLs are produced by LuxI family proteins and passively diffuse out of the bacterial cell
- once external AHL concentration reaches threshold (the point at which the concentration gradient is reversed), AHL diffuses back into cell, and binds with transcriptional regulator LuxR
- AHL induces a conformational change in LuxR and then it becomes transcriptionally active (QS gene switch on!)
- particularly found in gram negative bacteria
4
Q
QS in bioluminescence
A
- LuxI/LuxR nomenclature reflects the fact that QS was first recognized as regulating bioluminescence of Aliivibrio fischeri: LUCIFERASE GENE enzyme produces the bioluminescence
1. Autoinducer synthesis leads to high-level expression of lux operon
2. Low cell densities result in basal level transcription of lux operon
3. High cell densities: high AI levels and elevated transcription of lux operon - Autoregulation of LuxI/R genes
5
Q
Quorum sensing is widespread
A
- since being identified within Vibrio species, the phenomenon of QS has been identified in many other microbial species including Gram-positive and Gram-negative bacteria and also fungi
- QS regulates diverse processes and employs several different types of autoinducer molecule
6
Q
Key differences in QS of Gram-positives: AIPs
A
- ‘Auto-inducing peptides’ (AIP) instead of AHLs
- produced as propeptide before processing and active secretion
- AIP does not re-enter cell, but triggers signalling via two-component signalling pathway
-e.g. see AGR system in staph aureus
AgrB secretes the AIP
AgrC (histidine kinase: one half of a two component system) binds the AIP - triggers autophosphorylation
AgrA is phosphorylated, gene regulator turns on QS gene expression
7
Q
Pseudomonas aeruginosa
A
- Gram-negative, aerobic bacilli
- Commonly found in soil & water, but of increasing importance as an opportunistic pathogen, particularly in hospital setting
- One of the leading causes of healthcare-associated infection - can infect any site in immunocompromised
- Urinary tract infections, respiratory infections, dermatitis, soft tissue infections, bacteraemia, bone & joint infections, GI infections and various systemic infections (burns/AIDS/cancer)
- The dominant pathogen within cystic fibrosis (CF) patients
8
Q
QS systems of P. aeruginosa: one of the best studied QS and biofilm bacterium
A
- Within Pseudomonas aeruginosa, there are at least three distinct quorum sensing systems (not unusual to have multiple QS systems):
- two of them are based on the AHL system: LasIR and RlhIR
- the third is the distinct PQS system (Pseudomonas quinolone signal).
- LasIR and RlhIR (LuxI family proteins! conserved) produce AHL (slightly different, lactone ring conserved but the carbon tail is different length with different oxygenation), can diffuse freely in aq environment
- The PQS system uses a structurally distinct signal molecule that is extremely hydrophobic in nature. This means that for it cannot passively diffuse within/through a bacterial population, it is instead packaged into membrane vesicles that bud off from the bacterial cell, and it is these PQS-containing vesicles that enable PQS-mediated cell-cell communication (actively trafficked)
- 5% of the genome of P. aeruginosa can be regulated by QS. Amongst the genes activated are a multitude of potential virulence factors that contribute to infectious process.
9
Q
Quorum sensing regulon of P. aeruginosa
A
- Elastase degrades elastin and other matrix proteins of the human lung, leading to tissue damage and destruction of lung structure. It is also a potent inflammatory factor.
- Alkaline Protease can proteolytically inactivate cytokines and other host defence proteins, thus interfering with the host immune response
- Superoxide dismutase is an important anti-oxidant protein (detoxifying), protecting the cell from oxidative stress. Oxidative stress is a key component of the innate immune response, particularly within phagocytic cells, so the ability of bacteria to resist this stress can promote its survival.
- LasA is a metalloendopeptidase which has strong anti-staphylococcal activity, and can also degrade elastin (bugs can compete against each other)
- HCN (hydrogen cyanide) is toxic to cells, inhibiting cellular respiration. Within the lungs of cystic fibrosis patients (who are commonly infected with Pseudomonas aeruginosa), the detection of HCN in CF sputum has been directly associated with poorer lung function.
- Rhamnolipids are haemolytic glycolipids with detergent-like activity. They demonstrate cytolytic activity against monocyte-derived macrophages and neutrophils.
- Pyocyanin has very diverse functions. It is ‘redox active’, resulting in the generation of reactive oxygen species that are damaging to the host cells (this is achieved by disrupting the cellular respiration of the host cells). It also has diverse immunological effects by disrupting the cytokine and chemokine response to infection, thus disrupting the ability of the host to respond appropriately.
- Biofilm architecture evades immune response
10
Q
Importance of QS for virulence of P. aeruginosa
A
- P. aeruginosa QS mutants have been studied in a variety of infection models including models of burn wounds, acute pneumonia and chronic lung infection
- All highlighted the profound involvement of QS in virulence, as mutants are associated with:
- Less tissue destruction
- Reduced cases of pneumonia
- Reduction in disseminated infection
- Reduced mortality
- many QS knockout bacteria are avirulent
- Despite this, it is important to stress that QS mutants are not completely avirulent
11
Q
Biofilms
A
- Biofilms are an aggregate of bacterial cells attached to a surface and encased in an adhesive matrix that is secreted by the cells
- Responsible for up to 80% of infections within the human body, and are a major problem on indwelling devices (Prosthetic heart valves, catheters, IV lines, tracheal tubes)
- Biofilm-associated infections are difficult to treat due to the protection conferred against antibiotics and the immune system:
Impaired diffusion of certain antibiotics
Reduced metabolic activity within biofilms (cells embedded deep in the matrix do not have ready access to nutrients and oxygen so tolerate AB, also PERSISTERS) - Biofilms are often negatively charged - if an AB has a positive charge it will not penetrate the matrix
- Control strategies include the use of indwelling devices that have been impregnated with antimicrobials
- QS has a role in biofilm formation
12
Q
Azithromycin
A
- an established antibiotic that inhibits QS
- Although azithromycin has no bactericidal or bacteriostatic activity against P. aeruginosa, its use has seen clinical benefits
- Azithromycin inhibits QS activity in patients
13
Q
Inter-species communication via autoinducers
A
- AHLs Acyl homoserine lactones produced by different bacteria share the same basic structure but differ in:
1. Length of the R-group side-chain (vary from 4 to 18 carbon atoms)
2. Substitution of a carbonyl at the third carbon - Similarity between AHL molecules of different species can enable inter-species communication
- Inter-species communication is particularly favoured within mixed biofilms (i.e. different microbial species co-existing within the biofilm)
- P. aeruginosa & Burkholderia cepacia form mixed biofilms in the CF lung
- B. cepacia can respond to the quorum sensing molecules produced by Pseudomonas
14
Q
The significance of inter-species communication
A
- Non-pathogenic bacteria harboured by the host can modulate the behaviour of pathogenic bacteria, thus increasing the severity of disease
- e.g. pseudomonas and oropharyngeal bacteria - pPA responds to the autoinducers produced by OF and becomes much more pathogenic
15
Q
Conclusions
A
- Quorum sensing enables populations to co-ordinate their activities through use of autoinducer molecules
- Acylhomoserine lactones are widespread in Gram-negative bacteria, and freely diffuse in and out of the bacterial cell
- Autoinducing peptides are used by Gram-positive bacteria; they are actively secreted and bind to cell-surface receptor
- QS plays a central role in virulence regulation (including biofilm formation), and as such is an attractive antimicrobial target
- QS can also occur between different species, particularly within mixed biofilms
