Pseudomonas
aeruginosa
Human opportunistic pathogen
– exploits some break in hosts’
defenses in which to initiate
infection
Pseudomonas Antibiotic Resistance
Immunocompromised
individuals
2nd most common cause of
ventilator-associated pneumonia
(HAI)
Cystic fibrosis infection
•less than 1yr – 21%,
•greater than 19yrs – 80%
•Responsible for death of majority
4th most common cause
of catheter-associated
urinary tract infection
(HAI)
Nosocomial infections originate or occur in a
hospital or hospital-like
setting
[also called hospitalacquired infections (HAI)]
Grows sometimes
on skin, can infect
through injury
Quorum Sensing

LasI

LasR


LasI
Signal diffuses
Cells and signal
accumulate
Signal binds and
activate LasR, a
transcriptional
regulator
Virulence gene
expression
LasR
LasI
Signal synthase
LasR
Signal receptor
Discovery of anti-activator

LasI

QteE
LasR
LasR


LasI

QteE
LasR
Signal produced
QteE inhibits the
binding of the signal
and LasR
Signal accumulates
Signal binds with
and activates LasR,
the transcriptional
regulator
Virulence gene
expression
LasR
LasI
Signal synthase
LasR
Signal receptor
To differentiate
strains, integrate
mCherry
fluorescent
protein into
chromosome
Insert the GFP
plasmid into
strains for QS
determination
Co-culture
strains together
and use
microscopy to
identify
fluorescence
Integrating mCherry
 Confirm integration:
att site
 Tecan – measure
fluorescent excitation with
a machine
 Microscopy – check cellular
fluorescence with a
fluorescent microscope
Microscopy Image
The green fluorescent
protein is controlled
by a QS promoter
The cells will
fluoresce green when
LasR binds the signal
and induces
transcription of QS
genes
GFP
plasmid
Strain 2
Name
Strain 2 Function
Predicted
Results
Initial
Predicted
Results
Final
WT
ΔlasI
Doesn’t produce
LasI signal
synthase, QS can
only respond to
WT-produced
signal
Neither
strain
green
Both turn
green at
the same
time
Double mutant,
requires signal
from qteE-,
depends on
diffusion for
signal and QS
qteE- is
green,
lasIqteE- is
not
Both green
Control
Strain 1 Strain 1
Name
Function
(red)
Test
qteEQteE
Produces all
proteins, QS
proceeds as
normal
LasI
lasIqteEDoesn’t
produce QteE,
can “shortLasI
circuit”
QteE