Environmental Engineering Reference
In-Depth Information
The initial ratio of participating dioxygenases (BDO and ISP-OHB) therefore
appeared crucial for more difficult substrates when inhibitory effects of
reaction products were suspected (such as the case of 2,2′-CBP). Conse-
quently, LB400(pRO41) inocula for bioaugmentation of PCB-contaminated
soils should be grown on CBA prior to delivery in soil.
6.4.1.8 Growth on defined PCB mixtures
Control strain LB400(pRT1) was grown on BP and inoculated in K1 medium
with 1 m M mix C. Mix C represents the best scenario that would be seen in
anaerobically dechlorinated sediments initially contaminated with Aroclor
1242, 1248, and perhaps 1256. There were no previous reports on growth of
LB400 on any artificial mixes or naturally occurring dechlorination products.
As shown in Figure 6.17, the control strain grew on mix C, and the sum of
released Cl - and yielded 2-CBA, 4-CBA, and 2,4-CBA totaled approximately
1.4 m M Cl - , compared to the theoretically expected 1.5 m M . Of the total 1.4
m M chlorine released, 0.65 m M was released as Cl - and 0.7 m M accumulated
as 2-CBA in accordance with growth of the LB400(pRT1) on individual 2-
and 2,2′-CBP, and with only partial consumption of 2,2′-CBP. In contrast,
recombinant LB400(pRO41) released 1.5 m M Cl - and only negligible
amounts of 4- and 2,4-CBA and chlorocatechol, indicating nearly total min-
eralization. Further flask experiments were done to characterize growth and
degradation of products expected from the anaerobic phase of Aroclor reme-
diation. In these experiments, the inoculum of GEM LB400(pRO41:: ohb ) was
prepared on 2,5-CBA, whereas control strain LB400 was inoculated from
BP-grown culture. Individual flasks in triplicates represented time points.
Production of biomass (optical density, OD, at 600 nm), degradation of PCBs
(GC), accumulation of CBAs (HPLC), and release of Cl - (Bergmann and Sanik
method, 1957) were measured. The recombinant LB400(pRO41:: ohb ) grew
efficiently on both lighter mix C (Figure 6.17) and heavier mix M (Rodrigues
et al., 2005, submitted). Importantly, we observed significantly higher bio-
mass production and Cl - release, greatly diminished accumulation of CBAs,
and potentially toxic intermediates such as (Cl)HOPDAs by the recombinant
LB400(pRO41:: ohb ). It also well sustained the exceptionally high rates of PCB
oxidation characteristic for the parent LB400. These results suggested that
the GEM is superior to its wild-type parent for bioremediation purposes.
6.4.1.9 Validation of PCB remediation strategy in soil
(microcosm studies)
In preparation for a field test, we conducted a series of laboratory experi-
ments both in flasks and in soil microcosms to validate applicability of the
proposed two-phase anaerobic-aerobic PCB remediation employing GEMs.
Based on growth in flasks on defined PCB mixtures M and C, on Aroclor
toxicity testing, and on survival of different GEMs in soil microcosm exper-
iments, we have concluded that using a combination of two GEMs, the
longer-surviving gram-positive Rhodococcus RHA1(pRHD34:: fcb ) and the
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