Bacterial

Virulence and
Pathogenesis
Host-Parasite Relationships: Pathogenesis of Infections
In any host-pathogen encounter, there are two determinants of the
outcome:
1. Virulence of the parasite
2. Resistance of the host
In some cases, the host-pathogen relationship is very complex:

-Commensal but opportunistic

will take advantage of weakened host and invade tissues setting up a potentially life-
threatening infection

Examples include motile Aeromonads- natural inhabitants of intestine but cause

septicemia when fish is immune suppressed
o Bacteria cause disease by 2 basic mechanisms:
1 Direct damage of host cells
2 Indirectly by stimulating exaggerated host inflammatory/immune response
 Virulence factors are molecular components expressed by a
pathogen that increases its ability to cause disease

Virulence factors can be divided into two categories:

• 1. Those that cause damage to the host (toxins)
• 2. Those that do not directly damage the host but promote colonization
and survival of infecting bacteria
A. Bacterial toxins
1. Exotoxin: protein molecule liberated from intact living bacterium.
a. They are antigenic and can elicit protective antitoxic antibodies. Many of these
toxins can be converted to nontoxic immunizing agents termed toxoids.
b. Three roles of exotoxins in disease:
i. Ingestion of preformed toxin (botulism)
ii. Colonization of wound or surface followed by toxin production (cholera and
diphtheria toxins)
iii. Exotoxin produced by bacteria in tissues to aid growth and spread
(Clostridium perfringens alpha-toxin)
 d.Types of exotoxins:
i. A-B toxins (intracellular acting)
1) Composed of two parts: A and B portions
2) The B portion mediates binding to a specific host cell receptor.
3) After binding to the host cell, the A portion is translocated into host cells and has
biological activity against an intracellular target

or
4) Examples:
a) Diphtheria toxin: ADP-ribosylation of host EF-2; host cells are killed by blocking translation.
b) Cholera toxin: ADP-ribosylation of a cAMP regulatory protein, which causes loss of
ion regulation, water loss, diarrhea.
c) Shiga toxin cleaves host rRNA, which blocks translation and kills the host cell.
d) Clostridium botulinum- large subunit targets neurons, small subunit cleave snare
proteins inhibiting neurotransmitter release from neurons- causes paralysis BoNT- E in
fish (most toxic substance known)
 ii. Membrane disrupting (surface damaging)
1) Cause damage or disruption of plasma membranes, which
leads to osmotic lysis and cell death. Many were originally
termed “hemolysins” because they lyse RBCs.
2) Three types of membrane disrupting toxins:
a) Enzymes that hydrolyze phospholipids:
phospholipase,
sphingomyelinase
b) Toxins with detergent-like surfactant activity that disrupt
by membrane solubilization
c) Pore forming toxins (the most common): proteins that
insert in the host membrane and form a hydrophilic pore

Aeromonas produces up to 4 hemolysins- aerolysin A (AeroA) and

Heat labile hemolysin AHH1- work synergistically, also some
aeromonads produce the pore forming toxin RtxA

Staphylococcus aureus
alpha hemolysin,
looking down the
central pore
 Superantigens
iii.
1) Toxins that bind directly to MHC II on
macrophages (without being processed)
and form a crosslink with T cell
receptors.
2) Crosslinking causes stimulation of up to
1 in 5 T cells in the body (normal
antigens cause stimulation of 1 in
10,000).
3) Excessive IL-2 production results from
the massive stimulation of T helper cells,
4) Stimulation of other cytokines by IL-2
lead to shock.
Example: staphylococcal toxic-shock
syndrome
 iv. Extracellular enzymes: break down host macromolecules.
play an important role in disease development by providing a nutrients or aiding in
dissemination. Can cause extensive tissue damage
Examples:
a) Coagulase – clots fibrin, thus protecting the bacteria.
b) Hyaluronidases and proteases – aid in the spread of bacteria by degrading
extracellular matrix.
c) Collagenase – aids in dissemination
d) DNase – reduces viscosity of debris from dead cells (may help escape DNA net by
neutrophil).

A. hydrophila - Express diverse extracellular enzymes can contribute to virulence including

collagenase, elastase, enolase, lipases (heat stable lipase, pla and Plc), metallo protease,
and serine protease, Rnase R.
 2. Endotoxin- released when cells die: lipopolysaccharide (LPS)
produced by gram- negative bacteria. In gram-positive bacteria
peptidoglycan and teichoic acids.

a. LPS is bound by LPS binding proteins in plasma, which then binds CD14. This complex binds
Toll-like receptor 4 (TLR4) on macrophages and monocytes. TLR2 binds teichoic acids.
TLR1 binds peptidoglycan.
b. Macrophages and monocytes release cytokines (IL-1, IL-6, IL-8, TNF alpha,
Platelet Activating Factor), which subsequently trigger prostaglandin and
leukotriene release
c. The complement and coagulation cascades are activated.
e. endotoxic shock occurs when bacterial products reach high enough levels in the blood to
trigger complement activation, cytokine release, and coagulation cascade activation in
many parts of the body. Circulatory system collapse followed by multiple organ system
failure occurs.
B. Bacterial invasion of host tissues
1. Host damage is caused during invasion by either:
a. direct disruption of function
b. an exaggerated immune response that compromises tissue function.
2. The invasive bacteria are classified as:
a. Facultative Intracellular Parasites
i. FIPs are not confined to cells
ii. Some can multiply in professional phagocytic cells.
iii. When a balance is established between the bacterium and phagocyte, the
bacteria may survive in this intracellular state for months or years (example:
Mycobacterium).
b. Obligate Intracellular Parasites; can only propagate inside host cells.
Examples include chlamydia and rickettsia
c. Extracellular parasites, which cause tissue damage while they are
outside phagocytes and other cells and do not have the ability to survive
long periods in cells.
 3. Steps in bacterial invasion:
a. Motility
i. Flagella are the best characterized; adapted for low
viscosity fluids.
ii. Other types of motility: corkscrew type (Spirochetes--best in
viscous solutions), gliding motility (Flavobacterium columnare
and cytophagas, myxobacteria--movement over solid surfaces).
iii. Chemotaxis is directional swimming using a gradient (especially
nutrients).

A. hydrophila produce lateral flagella for surface movement and polar

flagella for movement in suspension. Glycosylation of polar flagella involved
in biofilm formation, binding to cells and mucosal adherence
 b. Adherence
i. Two common strategies: fimbriae and monomeric protein adhesins.
ii. Fimbriae (pili): receptors are usually carbohydrate residues of glycoproteins or
glycolipids. Attachment is more fragile. Highly specific binding, often mediated by
adhesins, can be blocked by antibodies, often specific for host tissue
type/location.
iii. Monomeric protein adhesins: mediated by cell surface proteins, tighter binding to
host cell, may recognize proteins on host cell surface, may follow looser fimbrial
attachment.

Aeromonas-bundle-forming pilus (encoded by bfp) is a critical internal colonizing

factor
 c. Invasion of host cells (intracellular pathogens)
i. Some invasive bacteria have mechanisms for entering host cells that are
not naturally phagocytic.
ii. Two types of bacterial-mediated invasion:
a. Zippering: bacteria present ligands on their surface allowing them to bind to
host cells and initiate the entry process. It is similar to FcR- and CR3-
mediated phagocytosis, which is characterized by the formation of inclusion
shaped by the bacteria they ingest (Yersinia pestis Ail).
b. Triggering: bacteria inject effectors into host cells via T3SS to regulate
phagocytosis (Salmonella).
iii. Following attachment to host cells, pathogens cause changes in host
cell cytoskeleton (actin) that cause the pathogen to be internalized.
iv. Some pathogens can utilize actin fibers intracellularly to move through host
cells (transcytosis).
v. Invasins may also mediate uptake of bacteria into professional phagocytic cells in
a way that bypasses normal phagosome formation.
d. Manipulation of host cell functions
i. Bacterial pathogens are often very manipulative of host cell functions; both
extracellular and intracellular pathogens will cause host cells to perform functions
favorable to the pathogen.
a. For example, leukotoxin produced by Mannheimia haemolytica (extracellular pathogen)
induces cytokine secretion.
b. Listeria monocytogenes (intracellular pathogen) produces a protein that mobilizes actin to
propel bacteria through the cell and into neighboring cells.
 ii. Some bacterial pathogens have a specialized type III secretion system (TTSS) that forms a
needle-like structure that injects effector proteins directly into the host cell cytoplasm.
a. In some cases, these effector proteins serve as receptors in the host membrane for
bacterial attachment.
b. In some cases, these effector proteins can mobilize cytoskeleton to cause
phagocytosis.
c. In some cases, effector proteins can induce or prevent apoptosis.

Aeromonas express type II, III and VI secretion

systems III and VI can inject effector proteins into
host cells (II is for extracellular release of proteins).
4. Obtaining nutrients
a. Pathogenic bacteria have intricate methods to obtain all essential nutrients.
b. Obligate intracellular bacteria have complex nutrient requirements and
parasitize the living cell for an extended period.
c. Host cytoplasm is a very nutrient rich environment.
i. Extracellular pathogens often lyse cells to obtain nutrients.
ii. Intracellular pathogens will either escape from phagosomes to enter the
nutrient rich cytoplasm or modify the vacuole so they can get nutrients
from the cytoplasm (example: E. ictaluri).
d. Iron
i. Host tissues are very low in iron because it is bound to
transferrin, lactoferrin, ferritin, and heme.
ii. Bacterial strategies for obtaining iron (often induced by low iron
conditions):
1) Siderophores--low MW compounds that chelate iron with very
high affinity; secreted and taken up by bacterial surface receptors
2) Direct binding of host
transferrin, lactoferrin, ferritin,
or heme by bacterial surface
receptors.
3) Exotoxins that lyse host cells (can
be used to obtain other nutrients
as well).
 5. Evasion of host immune response
a. Serum resistance
i. Serum resistance is defined as the ability to prevent bacterial lysis by the C5b-C9
membrane attack complex (MAC).
ii. Capsule mediates resistance to complement by:
1) preventing C3b binding
2) promoting C3bH complex formation instead of C3bBb (mediated by sialic acid in
capsule-this inhibits complement cascade).
iii. Lipopolysaccharide--binds C3b and C5b. However, O polysaccharide can mediate
resistance to complement by:
1) having sialic acid attached to promote C3bH formation
2) having long O polysaccharide side chains that prevent MAC killing after C5b binds
(possibly too far from bacterial outer membrane).
iv. S-layer or outer membrane proteins
Aeromonas encodes an S-layer also and capsule, TagA cleaves C1-esterase inhibitor imparting
serum resistance
 b. Resistance to opsonization/phagocytosis

i. Capsule:
1) prevents C3b-mediated opsonization (by the same
mechanism used to avoid complement-mediated
killing)
2) prevents antibody-mediated opsonization by
masking (hyaluronic acid, sialic acid).

Aeromonas- capsule have anti-phagocytic activity,

provide increased resistance to the complement
system, and increased adherence
b. Resistance to opsonization/phagocytosis
ii. LPS O polysaccharide can prevent opsonization if it has sialic acid
iii. S-layer
iv. Extracellular products: enzymes that inactivate C5a chemoattractant (S.
pyogenes), toxins that kill phagocytes (leukotoxins) (Mannheimia
haemolytica), inhibit migration, or reduce oxidative burst.
c. Strategies for surviving
phagocytosis:
i. Escape from phagosome before fusion with lysosome (example: Listeria
monocytogenes, mediated by listeriolysin)
ii. Prevent phagosome-lysosome fusion-use type 3 secretion system to
influence trafficking
iii. Express factors that allow survival in harsh phagolysosome conditions
(catalase, superoxide dismutase, surface polysaccharides, cell wall)
d. Evading antibody
i. Ig proteases
ii. Antigenic switching or phase variation
iii. Masking (sialic acid, hyaluronic acid, coating with
host proteins such as fibronectin).
6. Virulence factors expression and release are coordinated by
intricate gene regulation and regulated protein function
a. Regulon-coordinated control of group of virulence factors that
are activated or deactivated in response to environmental
signal.
b. Allows bacterial pathogens to adapt to varying host conditions.
c. Virulence gene expression can be triggered when a
pathogen senses environmental cues from the host
environment (examples: pH, iron concentration).
 d. Virulence gene expression is sometimes triggered when
a pathogen detects sufficient bacterial numbers are
present:“quorum sensing”
i. Bacteria with quorum sensing capability secrete a small molecule (for example,
homoserine lactone)
ii. When the quorum sensing molecule reaches a critical concentration, gene
expression is stimulated.
iii. Sometimes quorum sensing regulates virulence genes.

Aeromonads have elaborate quorum

sensing system that regulated biofilm
formation and virulence genes

advancedhealing.com
Biofilm
Definition: a structured community of bacteria enclosed
in a self-produced polymeric matrix and adherent to an
inert or living surface. Can provide resistance to damage
outside of host, can protect against immune processes
inside the host and can provide transient antibiotic
resistance
Resistance is due to:
a. Slower growth rates of bacteria within biofilms
b. Decreased diffusion of antibiotics through
the biofilm (protective matrix)
c. Accumulation of enzymes that contribute to
resistance
Scanning electron
micrograph of E. coli
O157:H7
biofilm bacteria
 Persistence in the presence of antibiotics- regulated phenotypes

Persisters are non- or slow-growing reversible phenotypic variants of the wild

type, tolerant to bactericidal antibiotics.
• tolerance
i.
is due to inhibition of essential cell functions during
antibiotic stress,
• resulting in inactivity of the antibiotic target.
iii.
• ii. Persistence occurs in E. coli, Pseudomonas aeruginosa,
Mycobacterium tuberculosis, and Staphylococcus aureus.
• Persistence requires coordinated metabolic changes; entry and exit
from the persister state is regulated by signal molecules (such as
guanosine pentaphosphate or ppGpp).
Summary

o Bacterial pathogens are highly adapted and specialized.

o Infection constitutes an organized, orchestrated attack.
o Toxins manipulate the host to the bacteria’s advantage.
o Specialized mechanisms are required to invade the host, obtain
nutrients, and avoid the immune response.
Thank you