Transcript Lecture #14 Bio3124 - University of Ottawa

Medical Microbiology
Microbial Pathogenicity
Lecture #14
Bio3124
Pathogens as Parasites
• pathogens are parasites
– organisms that live on or within a host
organism, metabolically dependent on the host
– Parasitism:
• Ectoparasite: parasite lives on the host
• Endoparasite: parasite lives in the host
Parasitism and disease
• Infection
– growth and multiplication of parasite on or within host
• Infectious disease
– disease resulting from infection
• Pathogen: any parasitic organism that causes infectious
disease
– primary (frank) pathogen – causes disease by direct
interaction with healthy host
– opportunistic pathogen – part of normal flora, causes
disease when gains access to other tissue sites or when
host is immunocompromised
• Pathogenicity
– ability of a parasite to cause disease
Host-parasite relationship and disease outcome
Disease state depends on:
– number of organisms present
– degree of virulence of pathogen
– virulence factors
• e.g., capsules, pili, toxins
– host’s defenses or degree of resistance
Virulence: degree/intensity of pathogenicity
• determined by,
– Invasiveness: ability to spread to adjacent tissues
– Infectivity: ability to establish focal point of infection
– pathogenic potential: degree to which pathogen can cause damage
to host
• Toxigenicity: ability to produce toxins
• Immunopathogenicity: ability to trigger exaggerated immune
responses
Measuring virulence
• lethal dose 50 (LD50)
– number of pathogens
that will kill 50% of
an experimental
group of hosts in a
specified time
• Infectious dose 50
(ID50)
– number of pathogens
that will infect 50% of
an experimental
group of hosts in a
specified time
Infection Cycle
• Mode of entry depends on pathogen
• Mucosal surfaces, wounds, insect bites
• Infection cycle
Route a pathogen takes
to spread
• Spread via direct contact
• Indirect contact
– Contact with fomites
– Horizontal transmission via vectors
• Mosquitoes—Yellow fever, malaria
• Reservoir for disease organism
– May not show disease symptoms
Virulence Factors
• Virulence genes
– Help pathogen to invade host
• Toxins, attachment proteins, capsules
• Pathogenicity islands
– Section of genome
• Contain multiple virulence genes
– Often encode related functions
» protein secretion system, toxin production
– Horizontally transmitted
• Often flanked by tRNA genes; phage or plasmid
genes
• Often have GC content different from rest of genome
Virulence Factors
• Several factors contribute
– Protein secretory systems
• Examples:Type II, type III and type IV
– Adhesins: host attachement & colonization
– Toxins
• Exotoxins
–
–
–
–
–
Membrane active toxins
Protein synthesis inhibitors
Cell signaling inhibitors
Superantigens
proteases
• Endotoxins
– Immune avoidance factors
Role of protein secretory pathways in virulence
• PS Type II (retractable)
– Subunits in inner, outer and
periplasmic space
– G subunit
polymerize/depolymerize
– Extends/retracts past outer
membrane through complex D
– like a piston pushes out the
secreted proteins to periplasmic
space
– Ex. Cholera toxin
• PS Type II mechanism
resemble pili type IV used for
twitching motility
Type III protein secretory system
• many G- bacteria, live in close
association with their hosts
• secrete regulatory proteins via
injectisome directly into host
cells
– to modulate host cell activities
– evolutionary resemblance to
flagellum
• increase virulence potential
– Avoids receptor use
– Avoids dilution of secreted proteins
outside pathogen
Ken Miller talks about PSIII and flagellum
Salmonella SPI-1 and SPI-2 are type III secretory systems
• 12 pathogenicity islands in S. typhi
• SPI-1, a type III secretory system
• Injects 13 different toxins (effector
proteins)
• Subvert signaling, remodel
cytoskeleton
• Induce membrane ruffles, take
S.typhi
• SPI-2: alter vesicle trafficking
– Prevent phogosome-lysosome
fusion
– Pathogen avoids innate immunity
Injectisome: a type III secretory virulence factor
Toxin secretion by type IV secretory system
• Resemble conjugation
apparatus of gram
negative bacteria
• Bordetella pertussis
toxin secreted through
general SecA pathway
to periplasm
• Type IV collects toxin in
periplamic space
• Exports across outer
membrane
General SecA dependent secretory system
Adhesins: Microbial Attachment
• Human body expels invaders
–
–
–
–
Mucosa, dead skin constantly expelled
Liquid expelled from bladder
Coughing, cilia in lungs
Expulsion of intestinal contents
• Adhesins: surface proteins, glycolipids,
glycoproteins
– assist in attachment and colonization of host
tissues
• Pili (fimbriae)
• Hollow fibrils with tips to bind host cells
Adhesins: Pili type I
• e.g. Pyelonephritis pili of
uropathogenic E.coli
• attachment to P-blood group antigen
• upper uninary tract infection
• Pili assemble on outer membrane
• First, general SecA dependent
secretion to periplasm
• PapG,E,F & major subunit Pilin A
• PapD chaperon sorting/delivery to
PapC
• Secretion and pilus formation
• PapG recognizes the digalactoside on
P-blood group antigen of host kidney
cells
Adhesins: Pili type IV
• Found on P. aeruginosa, V.
cholera, pathogenic E. coli & N.
meningitidis
• Mediates attachment and twitching
motility
• Resemble type II secretory system
• Pil A is major structural pilin
• PilC,Y1 tip attachment proteins
• Assembly: PilA preprotein signal
sequence removed by PilD
• PilQ mediates export across outer
membrane
• PilF/T mediates energy dependent
assembly/disassembly of pilus
Type IV pili: bacterial attachment and motility
Exotoxins
• soluble, heat-labile, proteins
• usually released into the surroundings as bacterial
pathogen grows
• most exotoxin producers are gram-positive
• often travel from site of infection to other tissues or
cells where they exert their effects
More About Exotoxins
• Some toxin genes born on plasmids or prophages
• the most lethal substances known
• highly immunogenic
• can stimulate production of neutralizing antibodies
(antitoxins)
• can be chemically inactivated to form immunogenic
toxoids
– e.g., tetanus toxoid
Membrane-disrupting exotoxins
Alpha toxin of S. aureus
•
Forms 7-membered oligomeric beta-barrel
•
Cause cytoplasmic leakage
Phospholipase of Clostridium perfringens
• removes charged head group of phospholipids
in host-cell plasma membranes
– membrane destabilized, cell lyses and dies
– Also called α-toxin or lecithinase
AB type Exotoxins
Composed of two subunits
• “A” subunit – responsible for toxic effect
– ADP-ribosyltion of target proteins eg.
diphtheria toxin
– Cleave 28S rRNA, eg. Shiga toxin
• “B” subunit – binds to target cell,
delivers A subunit
Diphtheria exotoxin
• B subunit mediates receptor binding
• Endocytosis and fusion membrane
vesicles eg. ER or endosomes
• B recycles back to membrane
• “A” escapes and enters cytoplasm
• In the cytoplasm A catalyses ADPribosylation of EF2, halts translation
• Cell death ensues
Diphtheria toxin targets EF2
disrupts translation
Anthrax toxin: a deadly protease
• Anthrax toxin composed of,
– Protective antigen (B
subunit): delivers EF and LF (A
subunits)
– Edema factor raises cAMP
levels
• Causes fluid secretion, tissue
swelling
– Lethal factor cleaves protein
kinases
• Blocks immune system from
attacking
Bacillus anthracis
Animation: anthrax toxin mode of action
Superantigens
• Are bacterial and viral proteins that can activate T-cells
• in the absence of a real bacterial antigen mediate the
binding of MHC-II and T-cell receptors (almost 30% of Tcell population)
• eg. Staphylococcal enterotoxin B (SEB)
• Massive activation results in producing lots of cytokines
• Results in tissue damage and shock and multi-organ failure
Animation: Superantigens
Endotoxins
• lipopolysaccharide in gram-negative cell wall can be
toxic to specific hosts
– called endotoxin because it is bound to bacterium and
released when organism lyses and some is also
released during multiplication
– toxic component is the lipid portion, lipid A
• heat stable
• toxic (nanogram amounts)
• weakly immunogenic
• generally similar, despite source
Immune avoidance mechanisms
• Once inside host cell, how to avoid
death?
– Cell ingests pathogens in phagosome
• Some pathogens use hemolysin to break out
– Shigella dysenteriae, Listeria monocytogenes
– Phagosome fuses with acidic lysosome
• Some pathogens secrete proteins to prevent
fusion
– Salmonella, Chlamydia, Mycobacterium, Legionella
• Some pathogens mature in acidic environment
– Coxiella burnetii—Q fever
Surviving within the Host
Survival inside phagocytic cells
• escape from phagosome before fusion
with lysosome
– microbes use actin-based motility to
move within and spread between
mammalian host cells
Burkholderia pseudomallei forming
actin tails and protrude through
membrane and extend infection to
nearby cells
Surviving within the Host
• Outside host cell, how to avoid death?
– Complement, antibodies bind pathogen
• Some pathogens secrete thick capsule
– Streptococcus pneumoniae, Neisseria meningitidis
• Some pathogens make proteins to bind antibodies
– Staphylococcus aureus cell wall Protein A
» Binds Fc fragment
» Antibodies attach “upside down”
» Prevents opsonization
• Some pathogens cause apoptosis of phagocytes