Figure 8: Representative microphotographs of 48h-old biofilms
exposed to CIP for 6h: A) untreated biofilm grown in ALI; B) and C)
biofilms grown in ALI and treated with 50µg/mL and 1600µg/mL of CIP,
respectively; D) untreated biofilm was grown in LLI; E) and F) biofilms
grown in LLI and treated with 50µg/mL and 1600µg/mL of CIP,
respectively.
Conventional methods to determine antibiotic efficacy are routinely
performed under the standard liquid culture condition using the broth
microdilution technique, and 96-well microtiter plates [28-31] and
the standard protocol for measuring minimal inhibitory concentration
(MIC), which required to inhibit growth or kill planktonic bacteria
[32] and the minimal biofilm eradication concentration (MBEC)
[33] were proposed. Several novel methods, such as the Calgary
Biofilm Device [34] and microfluidic devices [35], were
developed to improve the measurement of MBEC as well. However, the
biofilms were mostly cultured by submerging in the liquid, and
antibiotics can only be delivered through one approach (biofilm
submerged in antibiotics). In our study, injecting the antibiotic
solution into the basal chamber and interacting with biofilm from the
substrate attaching side illustrates an alternative way to mimic the
antimicrobial mechanism that the drug being administrated through oral
or parenteral routes and taken to the biofilm site through the systemic
circulation. The MBEC of 3-day PAO1 biofilm using CIP reported by Wu et.
al. using the microtiter plate method was 64μg/mL [31], while our
results show that significantly higher CIP concentration is needed to
achieve a 50% decrease in the biofilm. The differences across models
highlight the importance of having a model that can closely mimic the
environmental conditions where these communities persist. Thus,
recognizing the differences in biofilms thrived on various interfaces
could shed light on designing more effective and efficient control
methods.
Conclusions
This paper introduces a novel reusable dual-chamber microreactor to
provide multiple interfaces for biofilm culture and test. The reusable
assembling technique provides greater flexibility to grow biofilms and
to screen the effects of different interfaces on biofilms, both
quantitatively and qualitatively. Protocols for culturing biofilm on ALI
and LLI in static condition and following test methods are feasible and
repeatable. PAO1 biofilms grow on ALI and LLI show differences in
morphology and resistance to physical interruptions. Our dual chamber
device also enables efficacy testing of antibiofilm strategies in an
environment that closely mimics in vivo biofilm growth
conditions, producing more accurate test results. 48h-old PAO1 biofilms
cultured on ALI and LLI show high resistance when antibiotic delivered
from the substrate side.