Environmental Engineering Reference
In-Depth Information
detect moving bacterial bands. For random motility quantification, similar
experiments can be performed with no chemoattractant.
Temporal bacterial concentration profiles can be obtained from digitized
images and transformed into dimensionless bacterial concentration versus
position plots (Fig. 7.1B). The area under the crest or above the trough repre-
sents bacterial concentration and is proportional to the number of migrated
bacteria. Ford et al. [18] show that the area above the trough is linearly
proportional to the square root of time. The following expression can be used
to determine the random motility coefficient [15]:
r
4
0
t
p
b
0
2
¼
A
exp
(7
:
1)
where b
0
is initial bacterial concentration in the injected solution, m
0
is the random
motility coefficient, t is elapsed time, and A
exp
is the experimentally determined
area under the bacterial concentration versus position curve. The random moti-
lity coefficient can be calculated using slope of the A
exp
/b
0
versus t curve and Eq.
(7.1). A detailed procedure for determining chemotaxis parameters is provided by
Lewus and Ford [15]. One of the main advantages of using this method is that it
provides well-defined boundary conditions for attractant and bacterial concen-
trations in both spatial and temporal coordinates, and ease in mathematical
analysis of experimentally obtained data. In addition, the initial chemical con-
centration is easily adjusted, enabling controlled studies of the chemotactic
response at varying contaminant concentrations. The SFDC has been used to
confirm that P. putida F1 exhibits both positive and negative chemotaxis toward
benzene at low and high chemical concentrations, respectively.
7.3.3 Agarose Plug Assay
The agarose-in-plug bridge method was first developed by Yu and Alam [19]
for studying chemotaxis. In this method, an attractant or repellent is mixed
with low-melting temperature agarose and a drop of mixture is placed on
the top of a microscope slide. A cover slip supported by plastic strips at
both ends is placed on top to form a chamber around the agarose plug
(Fig. 7.1C). A bacterial suspension is flooded into the chamber around the
plug. A characteristic chemotactic band of bacterial accumulation is visua-
lized surrounding the plug—a small distance from its edge—using light scat-
tering microscopy. The limitation of this method is its poorly defined
boundary conditions due to variability in the shape of the plug, making it
difficult to model mathematically. This method has been used for toluene [20]
and TCE [20, 21], and is particularly useful for volatile compounds, as the
system is partially closed and therefore minimizes volatilization losses of the
chemical [7].
