Biology Reference
In-Depth Information
Bradford et al. (2007) for carboxyl latex microspheres with a diameter of 1.1 mm suspended in
solutions with ionic strengths up to 31mM (the ionic strength of the solutions we used was 4.7
mmol/L only). Total porosity was determined gravimetrically to be 0.40. Prior to the
experiments, to remove impurities, the sand was rinsed sequentially with acetone, hexane and
concentrated HCl, followed by repeated rinsing with de-mineralized water until the electrical
conductivity was very low (<3 S/cm).
The column consisted of a 5 m transparent perspex tube with an inner diameter of 10 cm, and
with sampling ports placed at 10-50 cm intervals along the tube. A stainless steel grid for
supporting the sand was placed at the bottom of the tube. The column was gently filled with the
clean quartz sand under saturated conditions, while the sides of the column were continuously
tapped during filling, to avoid layering or trapping of air. The column was connected both at the
funnel shaped effluent end and influent end with two Masterflex pumps
(Console Drive Barnant
Company Barrington Illinois, USA) via teflon tubes, and the pumps were adjusted to a fluid
approach velocity of 1.16×10
-4
m/s, coinciding with flushing the column with 1 pore volume per
working day. Prior to a column experiment, the column was flushed for two days with either DI
or AGW to arrive at stable fluid conditions inside the column. Bacteria influent suspensions were
prepared by washing and centrifuging (3000 rpm) three times in either DI or AGW, and then
diluting 1000 times to arrive at bacteria cell concentrations of approximately 10
5
cells/mL.
Experiments were conducted by applying a pulse of 0.15 PV (approximately 2.2L) of bacteria
influent suspension to the column, followed by bacteria free DI or AGW. Samples were taken at
7 distances from the column inlet, and immediately plated (0.1 mL) in duplicate on Chromocult
agar (Merck). Decay was assessed in all experiments by plating samples of the influent at an
hourly frequency during the entire experiment. All plates were incubated at 37 °C for at least 18
hours.
After each experiment, to clean the sand in the column, and to prepare for the next experiment, a
pulse of 5 L 1.9 M HCl followed by a pulse of 5 L 1.5 M NaOH was flushed through the column,
followed by flushing with DI water until the electrical conductivity of the effluent was well
below 3 S/cm.
To determine auto-aggregation, 15 ml of freshly grown bacteria were centrifuged (14000 xg) and
washed three times in AGW, and then allowed to stand for 180 minutes at a temperature of 4 °C.
A sample of 1 mL 1cm below the surface of the suspension was obtained, immediately and 180
minutes after washing. The optical density of the samples was measured at 254 nm, and the auto-
aggregation was determined as the ratio of the final over the initial optical density (in %).
4.2.2 Determining the sticking efficiency in each column slice
Crucial in assessing the characteristics of the sticking efficiency distribution of each
E. coli
strain, including determining the value of the minimum sticking efficiency, is the way in which
the sticking efficiencies are calculated. Instead of considering the entire column length, we
determined the sticking efficiency for each slice of column (Martin et al., 1996), in between two
sampling ports:
2
d
M
(4.1)
Α
= −
c
ln
i
(
)
0
i
3 1
−
Θ Η
L
M
i
i
−
1
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