(582ca) Real-Time Monitoring Of Bacterial Growth and Biofilm Formation With Surface Plasmon Resonance Imaging

In this study,
for the first time we used a Surface Plasmon Resonance imaging (SPRi) system to
monitor bacteria in real time on the sensor surface as they move, grow and form
biofilms. SPRi provides label-free and sensitive detection of changes in
refractive index that take place within approximately 200 nm of the gold sensor
surface. Changes in the refractive index are caused by biomolecules displacing
water at the sensor surface. Difference images are generated over time, which
show the changes that occur at the surface during the experiment when compared
to a reference image chosen at the beginning of the experiment. Combining
microfluidics with our SPRi system provides added control while studying
bacterial behavior. By placing microchannels on the surface of the gold surface
we limit the bacteria to two dimensions and keep them close to the surface,
which allows us to study bacterial growth and movement inside the channels.

Two
different strains of Pseudomonas aeruginosa PA14 were used for this
experiment: Wild Type, which produce biofilm, and PelA mutant, which grow
without producing biofilm. The two bacteria were injected into separate microchannels
and placed into the SPRi device. The instrument obtained a difference image of
the sensor surface every three seconds. We continuously monitored the channels for
up to 24 hours as bacteria grew and formed biofilms. We were able to differentiate
between the wild type bacteria and the mutant strain. The wild type bacteria
attached to the surface and quickly formed large bright spots as they produced
extracellular matrix around them, while the mutant bacteria attached to the
surface and grew as small discrete bright spots. For the first time, without
functionalizing the surface we continuously monitored two different strains of
the same bacteria in a single experiment over 1-cm-square area of the sensor
surface. This study provides valuable information about bacterial growth and
biofilm formation. This new technique can be used to study the effect of
different chemicals and surface chemistries on bacterial growth behavior.