3.2. Preliminary screening of isolates by 24-well plate test
In this study, gradual vanishing of oil filmwas used as the criterion to
evaluate bacteria biodegradation. Oil film gradually decreased from
center to border with the consumption of lubricating oil. The surface
color of cell culture turned to oysterwhite, and finally very little residual
lubricating oil distributed over hole border with cell culture turning to
turbid in good biodegradation tests. However, a disadvantage of well
plate testwas found in tests: the resultswere different for the same isolate,
so only the isolates with at least two good results in resting cell
tests and one good result in cell culture tests as inoculumwere reserved.
Those isolates with two good results in cell culture tests and only one
good result in resting cell tests were discarded for their weak biodegradation
potentials (Table 2), because resting cells presented poor activity
by comparison with cell culture. Resting cells reflect bacteria essential
attributes more clearly.
Cell density can be regarded as a supplemented criterion to assist
isolate screening. 2.1 ml cell culture was sampled to determine optical
density using 1 cmcuvette. Table 2 shows a regularity that OD600 values
are consistentwith the change of oil film, that is, the higher the cell concentration,
themore obviously the oil film disappears. In good biodegradation
tests, optical density values were 0.20–0.30 in subculture tests
and 0.10–0.22 in resting cell tests with more substrates consumed to
overcome substrate inhibition on cells. The phenomenon reflected on
not only long biodegradation time but also less biomass. In view of the
defect of well platemethod, 15 isolateswere reserved after two repeated
tests, which decreased workload. Such sample capacity would not
omit target strains but decrease workload. The shortage of this method
could be covered by following shaking flask tests.