Could antibiotic cocktails be the answer?
An in vitro study into antibiotic combinations with
enhanced activity against Escherichia coli
Rachel Hickman
Degree project in biology, Master of science (2 years), 2011
Examensarbete i biologi 30 hp till masterexamen, 2011
Biology Education Centre and Department of Cell and Molecular Biology, Uppsala University
Supervisor: Diarmaid Hughes
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µg/mL tetracyline added after). Both the susceptible wild type and transformed wild type E.
coli cultures were grown in Mueller Hinton II (MHBII) broth (22g in a 1 litre of distilled
water, autoclaved, pH 7.3 in accordance to Difco TM). The over-night cultures were used in
the minimum inhibition concentration experiments, fractional inhibition concentration test
experiments and the time-kill curve experiments. To create an overnight culture each were
grown in 15mL falcon tubes with 2mL of MHBII with constant agitation to ensure good
aeration at 37°C for 18 hours. The cultures in the minimum inhibition concentration tests and
the fractional inhibition concentration tests were grown in 96-well microtiter plates without
shaking at 37°C for 18 hours. Solid media was used in the time-kill curve experiments when
samples were taken out and plated; the plates used were Muller Hinton agar plates (38g in a 1
litre of distilled water, autoclaved, pH 7.3 in accordance to Difco TM).
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antibiotic of interest from left column to the right columns. The overnight culture of the test
organism was diluted 1000-fold (~5 x 105CFU/mL) if correct turbidity was observed and 10
µL of this was added to each well on the 96-well microtitre plate.
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IF5.*C!+'()$*!6,&)*!&3'!@&3!(.90!23',!),!&3'!B&3!(.9!"YY µL of MHBII mixed in and mixed
with the pre-existing liquid in the well then "YY µL was removed. Once the desired
concentration of antibiotics for each of the wells was created, each well of the 96-well plate
had 5 µL of diluted 1000-fold overnight culture (~5 x 105CFU/mL) if correct turbidity was
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