Lecture 7 - Virulence factors II and Evading phagocytosis Flashcards
What are superantigens?
Superantigens
Stimulate large numbers of immune cells: damage by extensive inflammatory reaction.
* Proteins that act indirectly on host cells stimulating over production of T-cells
* They bind to the Vβ domain of the T-cell receptor which is outside the normal antigen binding region.
* They activate 5-25% of all T-cells (normal antigens only activate 0.05% of all T-cells
* T-cells produce cytokines-overproduce IF-γ- activates macrophages
* The activated macrophages then over-produce more cytokines, IL-1, IL-6 and TNF-α
* This leads to severe inflammation, shock and multi-organ failure
Examples (streptococci and staphylococci)
Staphylococcus aureus causes toxic shock syndrome toxin 1 (TSST 1) which causes the secretion of fluids and electrolytes from capillaries leading to reduced blood volume and pressure
* Very resistant to proteases
* Stable at 60C
Can survive pH 2.5-11
What are the two methods of exposure to toxin?
- Intoxication
Up-take of a pre-formed toxin - usually in food- Colonisation and in situ toxin production
High density of pathogens required - form biofilms
Toxin production then initiated by quorum sensing
- Colonisation and in situ toxin production
What are Sierophores?
Both animals and bacteria require iron for metabolism and growth. Animals withhold iron from tissue fluids by binding it to haemoglobin, transferrin or lactoferrin. Pathogens can normally access iron in haemoglobin.
The low concentration of iron triggers the synthesis of siderophores by the pathogen (to capture iron)
Siderophore binding constants are so high that iron bound to transferrin and lactoferrin is confiscated and taken up by the bacterial cells via surface receptor proteins that recognise Fe-siderophores.
Corynebacterium diphtheriae
This bacteria is not QS controlled and is instead controlled by Fe availability. It is a Gram +ve actinobacterium that causes diphtheria.
The exotoxin is encoded in the DNA of a temperate phage.
β-phage encodes for the tox gene. The bacterial repressor protein (DTxR) controls expression of the tox gene and genes for siderophore production. When iron is no longer limited the tox gene expression increases (not QS controlled)
How do Pathogens interact with phagocytes?
Phagocytosis - is the actin-dependent engulfment of a particle, larger than 0.5um, which is recognised by receptors on the phagocytic cell (ingestion)
Phagosome maturation - is the process by which internalised particles (such as bacteria and apoptotic cells) are trafficked into a series of increasingly acidified membrane bound structures, ultimately leading to particle degradation (digestion).
Phagocytic cells that feed on the human biofilm
* Immune system: Macrophages, monocytes, neutrophiles and dendritic cells
* Feed on sloughed biofilm or individual pathogens
* One amoeba - Entamoeba coli
Environmental biofilms
* Amoebae
* Feed directly on the biofilm
* Similar phagocytic mechanisms to immune cells
* Bacteria practice on amoebae to develop pathogenicity
Describe phagocytosis in immune cells.
- Phagocytosis of antigen
- Lysosome fuses with phagosome (simplified)
- Antigen and enzyme mix in phagolysosome
- Antigen is degraded
- Antigen residue is voided by exocytosis
- Some processed antigen fragments (epitopes) are displayed on the macrophage surface
Amoeba don’t present these fragments
Describe injestion in phagocytosis
Ingestion
* Opsonic - prey particle is coated with immunoglobulins to enhance recognition by phagocytes (immune phagocytosis only) - disrupted with IgA proteases
* Non-opsonic - Particle is not coated with immunoglobulins and the phagocyte receptors recognise specific ligands on the prey cell (both immune phagocytes and amoebae)
Ingestion is receptor-mediated
The ligands: Pathogens associated molecular patterns (PAMPs) include:
* Adhesins (Pilli, fimbrae, flagella, lipoteichoic acid, attachment proteins)
* O-antigens of the LPs
* Peptidoglycan
They bind to sugar components of specific receptors.
The receptors: Toll-like receptors (TLRs) (not in amoebae)
* Don’t directly induce phagocytosis
* Indirectly activate phagocytic cells in immune system
Pattern recognition receptors (in amoebae)
* Two main groups of glycoproteins on surface of phagocyte- directly induces phagocytosis
○ Mannose receptor - CD206
○ Class A and B scavenger receptors
How do bacteria avoid injestion.
- Alteration of PAMPs
○ Antigenic variation
○ Acetylation/deacetylation/glycosylation of MurNAc/GlcNAc
○ Alanylation/glycosylation of Lipoteichoic acid- Defensive virulence factors
○ Being in a biofilm
○ Being encapsulated - Offensive virulence factors
○ Production of leukocidins - kill predator cells such (e.g. Cytotoxins that kill white blood cells)
- Defensive virulence factors
How do bacteria avoid digestion?
Phagolysosome also contains acid phosphatases, hydrogen peroxide, superoxide anions, lipases, nucleases etc.
Avoiding Digestion
* Tolerance - Some prey survive the digestive process and emerge unharmed
* Avoidance - Some prey prevent the digestive biochemistry occurring. They do not divide by are egested unharmed. They can divide in the food vacuole and then burst the host cell
* Escape - Leave the food vacuole and divide in the host cytoplasm
How do bacteria use tolerance to overcome phagocytosis.
Entry in to the phagosome is a shock therefore bacteria go into a shock response meaning stress proteins (SPs) appear within the cell. Most are molecular chaperones (helper proteins) whose role is to correct folding of polypeptide and re-fold incorrectly folded proteins.
Example - Escherichia coli
The RNA polymerase recognises the alternative sigma factor σ32 instead of the norma sigma factor σ70. Normally σ32 constitutively produced in cells - but broken down by DnaK within a minute so there is no build up and few chaperonins are produced. When stressed DnaK is side-tracked (needed to help folding) and therefore σ32 levels increase which allows transcription of other chaperones and there is a upregulation of chaperonins.
Overall tolerance allows cells to tolerate a short term stress, the changes are totally reversible over time. Egested bacteria may still be in the stress response leading to increased resistance to antimicrobials
Cells are in close association in the food vacuole
* E.coli conjugation is 3 times higher in food vacuoles
There is an increase in QS controlled genes being expressed and up-regulation of virulence.
How do bacteria use avoidance to overcome phagocytosis.
Interference with the biochemistry of digestion. Inhibits lysosomal fusion with the phagosome.
* Prevents digestive enzyme activity
Produces a protease that disrupts the host superoxide anion generation - superoxide dismutase- prevents H2O2 production
* Produces a protease that disrupts the host superoxide anion generation
* Some species do not divide but sit out the more hospitable food vacuole
Legionella pneumophila
* Main virulence system is the Do/Icm - type IV secretion system (attachment protein)
* Secretes effector proteins into host cells
* Divides within the phagosome
* When critical density of cells reached -lyses the host cell
* Natural reservoir = amoebae (e.g. Acanthamoeba castellanii)
Trojan Horse - pre-lysis
How do bacteria escape phagocytosis
Escape
Salmonella typhimurium & Listeria spp. Lyse the enclosing food vacuole membrane
Escape into the cytoplasm and proliferate, some strains recorded as dividing in the contractile vacuole. They eventually burst the host cell
Some increase in virulence and resistance features as with other mechanisms
