MIIM30011 2024 L8 - Bacterial Invasion & Dissemination Strategies PDF

 MIIM30011 – Medical Microbiology: Bacteriology Lecture 8: Bacterial Invasion and Dissemination Strategies Dr Sacha Pidot Learning objectives At the completion of this lecture students should be able to: Use examples to compare and contrast the two different mechanisms...

 MIIM30011 – Medical Microbiology: Bacteriology Lecture 8: Bacterial Invasion and Dissemination Strategies Dr Sacha Pidot Learning objectives At the completion of this lecture students should be able to: Use examples to compare and contrast the two different mechanisms of bacterial invasion Describe actin-mediated motility and why this is an advantageous trait for an intracellular bacterial pathogen Discuss the main ways in which intracellular pathogens can replicate, with examples of each Colonisation can involve invasion Why invade? – Disseminate into deeper tissues – Hide from immune response – Steal nutrients from host (obligate intracellular bacteria)  Getting inside host cells Two main mechanisms: Zipper Exploits host cell pathways normally used for adhesion of epithelial cells to extracellular matrix Induces receptor immobilization and clustering High affinity binding of bacterial ligand to host receptor Receptor mediated endocytosis Trigger Resembles cell ruffling induced by growth factors and hormones Brief contact by bacteria induces large scale actin polymerization and formation of ruffles on cell surface Ruffles fold over bacteria and engulf them Macropinocytosis Croinin and Backert, Frontiers in Cellular and Infection Microbiology, 2012  Zipper mechanism – Listeria monocytogenes L. monocytogenes = Gram positive, food borne pathogen CNS infections, maternofetal infections May induce abortion in pregnant women Internalisation requires the actin cytoskeleton and bacterial protein internalin A (InlA) Beads coated with InlA will enter cells Cossart P, et al. Trends Cell Biol. 2003  Zipper mechanism – Listeria monocytogenes InlA interacts with E-cadherin – E-cadherin involved in tight junction formation InlA + E-cadherin interaction → receptor clustering – Causes cytoskeletal changes – Leads to membrane curvature Bonazzi et al, 2009, Cold Spring Harb Perspect Biol Hamon et al. Nature Reviews Microbiology 2006 Loh et al, Cells, 2019  Zipper mechanism – Listeria monocytogenes Actually, it’s not that straight forward… L. monocytogenes has 2 invasins – InlA and InlB – Differences in human/animal receptors leads to differential infectivity/ability of Listeria to bind and invade Hamon et al. Nature Reviews Microbiology 2006  Trigger mechanism – Shigella flexneri Gram negative Water/person to person spread Bacillary dysentery (bloody diarrhoea) Invasive pathogen Invasion requires T3SS Cossart and Sansonetti, Science, 2004  Trigger mechanism – Shigella flexneri During Shigella infection: – Passes through M-cells – Induces cell death in macrophages – From basal membrane, can then invade epithelial cells and undergo intracellular life cycle BUT, this is also an alarm to the host!! Schroeder G., Clin. Microbiol. Rev. 2008  Trigger mechanism – Shigella flexneri T3SS in Shigella – genes on a plasmid T3SS secreted proteins directly assist invasion T3SS effectors target cytoskeleton (IpaA) and host cell GTPases that regulate actin (IpaC, IpgB1 and IpgB2) T3SS effectors target phosphoinositide signaling (IpgD) Schroeder G., Clin. Microbiol. Rev. 2008  Trigger mechanism – Salmonella Also enteric pathogen – causes diarrhoea SPI-1 invasion – encoded on Salmonella pathogenicity island 1 – encodes a type 3 secretion system T3SS effectors that also induce membrane ruffling  Trigger mechanism – Shigella vs Salmonella Compare the two processes in these bacteria Both use the trigger mechanism Both use T3SS to inject effector proteins that induce ruffling Ruffling facilitates internalisation Gruenheid and Finlay, Nature, 2003  Intracellular pathogens Why do some pathogens reside inside host cells? Advantages: – privileged environment (no competition) – inaccessible to attack by complement and antibodies – no longer need to adhere to host cells to maintain infection – ready access to nutrients – require host cell environment for survival (intracellular parasitism)  Intracellular pathogens Why do some pathogens reside inside host cells? Disadvantages: – Very hostile environment if pathogen is not adapted – Immune signalling from infected cells → apoptosis – What must a bacterial cell do to survive intracellularly?  Intracellular pathogens Two approaches to being an intracellular pathogen: 1. Enter and multiply in non-phagocytic cells induced endocytosis or macropinocytosis 2. Entry and multiplication in phagocytic cells (phagocytosis) neutrophils macrophages  Intracellular pathogens Three different pathways to be intracellular: – Intralysosomal – live in the lysosome – organelle designed to kill bacteria! Eg Coxiella burnetii – Intravacuolar – live in your own vacuole – Cytosolic – live in the cytosol  Cytosolic pathogens – Listeria and Shigella Invade cells Escape vacuole and proliferate Recruit host cell cytoskeletal proteins, induce actin polymerisation Formation of actin tail (comet like tail) Propel bacteria into adjacent cell Welch and Way, 2013, Cell Host Microbe  Actin tails – promoting cell-cell spread Actin tails promote bacterial spread within organs/tissues Drive bacteria into protrusions Protrusions are engulfed by neighbouring cell Why??? – To avoid host immunity! Welch and Way, 2013, Cell Host Microbe  Actin tails – promoting cell-cell spread (Listeria) Listeria escapes vacuole with listeriolysin O (LLO) – pore-forming toxin ActA mimics WASP → Recruits Asp2/3 → drives actin polymerisation Spreads to neighbouring cell Cycle repeats… Lamason and Welch, 2017, Curr Opin Microbiol Cossart PNAS 2011  Actin tails – promoting cell-cell spread (Shigella) Shigella uses IcsA IcsA recruits N-WASP → Recruits Asp2/3 → drives actin polymerisation Spreads to neighbouring cell Burkholderia, Rickettsia also spread via actin tails Cossart, Cell Microbiol. 2000 Lamason and Welch, 2017, Curr Opin Microbiol  Actin tails – promoting cell-cell spread (Listeria) Shigella actin tails Listeria actin tails https://www.youtube.com/watch?v=6EAAvwTaHKE https://www.youtube.com/watch?v=HAqAWJOP8Ko  Intravacuolar pathogens Salmonella, Mycobacteria, Chlamydia, Legionella Establish intracellular niches by constructing membrane- encompassed compartments Protect cells from cytoplasmic immune sensing molecules Must maintain integrity of the membrane compartment to survive – Have weapons to maintain vacuolar-lifestyle  Legionella pneumophila – life cycle Legionella pneumophila – Gram-negative, aerobic rod – Aquatic organism – Lives in amoebae in environment; human macrophages during infection Two lifecycle phases: – Replicative phase – non-motile – Infectious/transmissive phase– motile  Legionella pneumophila – life cycle Infectious phase – Flagellated form enters host cells – When in vacuole, secretes effectors type IV secretion involved >330 identified effectors (!!) Allow LCV to mature – Prevents lysosome fusion Stops digestion of bacteria – Recruits vesicles from ER and mitochondria Make itself look like another ER  Legionella pneumophila – vesicular manipulation Vesicles are how proteins/lipids move around within a eukaryotic cell Effectors important in modulating Effectors manipulate LCV surface host vesicle trafficking molecules – Prevents LCV-lysosome fusion – Makes itself look more like Golgi apparatus Qiu and Luo, 2017, Nat Rev Microbiol  Intralysosomal pathogens Phagocytes – macrophages and neutrophils Phagocytosis facilitated by opsonisation Killing follows phagosome-lysosome fusion – Oxygen-dependent - reactive oxygen species – Oxygen–independent – enzymes (lysozyme, phospholipases, etc) Intralysosomal pathogens include Coxiella burnetii https://www.youtube.com/watch?v=w0-0Bqoge2E  Intralysosomal pathogens – Coxiella burnetii Coxiella burnetii – obligate intracellular parasite – Causes Q fever – flu-like symptoms Acute and chronic forms – chronic = endocarditis Vaccine available – only approved for use in Australia! – Outbreak in goat farms in the Netherlands in 2007-2011 >4000 human infections, thousands of animals slaughtered as containment – Associated with animal products and contaminated milk – Single organism can cause disease! – Class B biological weapon  Coxiella burnetii life cycle Coxiella taken up by phagocytosis – Enters a Coxiella containing vacuole (CCV) – At this stage cell is small cell variant (SCV) Resistant to heat, dessication; metabolically inactive Enters vacuole, which fuses with endosomes/lysosomes – Becomes increasingly acidic Coxiella grows optimally at pH