130
Bioremediation for Sustainable Environmental Cleanup
all three chemicals in a combination of the tod and tol metabolic pathways (Worsey and Williams
1975, Zylstra et al. 1988).
As a result of the inability of catechol 2,3-dioxygenase in the tod pathway to attack
3,6-dimethylpyrocatechol produced from p-xylene and xylene oxygenase in the athway to employ
benzene as a substrate, it is possible that complete biodegradation of the BTX combination through
these two pathways will not be possible (Worsey and Williams 1973, Zylstra et al. 1988, Gibson
et al. 1970, Gibson et al. 1974). Consequently, an existing route must be restructured. In order to
manipulate the genes for two pathways in P. putida, a hybrid metabolic pathway was constructed
around the crucial metabolic step. In P. putida, an attempt was made to mineralize BTX by a
hybrid route. To construct the hybrid pathway, a bridging step between the tod and tol pathways
was identified. The tol pathway destroys cis-glycol chemicals, which are metabolic intermediates
immediately before the catechol molecules in the tod pathway. As the only source of carbon, BCG
was 90% degraded (Lee et al. 1994). P. putida TB101 was generated by introducing the TOL plasmid
pWW0 into P. putida F39/D, a P. putida FI variant incapable of converting cis-glycol chemicals to
catechols. Toluate-cis-glycol dehydrogenase in P. putida TB101 diverted the metabolic flux of BTX
into the tod pathway at the level of cis-glycol molecules, resulting in the simultaneous mineralization
of BTX mixture without accumulation of any metabolic intermediates (Lee et al. 1994). Toluate-cis
glycol dehydrogenase, which is encoded on the TOL plasmid of P. putida mt-2, was considered to
be responsible for the degradation of these cis-glycol compounds due to the comparable chemical
structure of its original substrates, benzoate-cis-glycol (BACG) and TACG. BCG was utilized as
a substrate during the toluate-cis-glycol dehydrogenase experiment. The observation shows that
the reaction product produced by BCG’s toluate-cis-glycol dehydrogenase may be destroyed by
catechol 2,3-dioxygenase, the enzyme that follows toluate-cis-glycol dehydrogenase in the tol
pathway. In P. putida TBlOl, a hybrid pathway was developed for the complete mineralization
of benzene, toluene and p-xylene. Using P. putida TBlOl, simultaneous biodegradation of BTX
combinations was accomplished with maximum specific degradation rates of 0.27, 0.86 and
2.89 mgmg–1 biomass/h for benzene, toluene, and p-xylene, respectively (Lee et al. 1994).
8.3.3 Phenylurea
Phenylureas like diuron linuron isoproturon and chlorotoluron were once used to control weeds, but
they have now contaminated drinking water. Diuron is a phenylurea herbicide that is used to control
weeds in a number of crops (Moretto et al. 2019). It has been found in soil and water (Giacomazzi
and Cochet 2004). Diuron has been linked to kidney disease, haematopoiesis and hemolytic anaemia
(Ihlaseh-Catalano et al. 2014). In high salinity conditions, the CASB3 strain of Stenotrophomonas
rhizophila was used to degrade diuron. Fifty mg L–1 diuron was completely degraded in 48 to
120 hr (Silambarasan et al. 2020). Arthrobacter globiformis D47 can ‘partially’ degrade herbicide
diuron via urea carbonyl group hydrolysis (Cullington and Walker 1999, Turnbull et al. 2001). The
concentration of 3,4-dichloroaniline (DCA) rises as the diuron degrades. A very small amount of
DCA can be converted to 3,3’ 4,4’ tetrachloroazobenzene and the remaining bulk of the DCA continue
bound to the organic matter with slow half-life for several years. Pseudomonas putida strain was
enriched for the demineralization of DCA. The pathway investigation study suggested biodegradion
through 3,4 dichloromuconate, 3 chlorobutenolide, 3 chlorolevulinic acid, 3 chloromelyelacetate
3chloro 4 ketadipate to succinate (You and Bartha 1982).
8.3.4 Methyl Parathion
Microbes are also decomposers that improve soil fertility by degrading pesticides and immobilizing
heavy metals. Toxic pollutants include organophosphate pesticides such as Methyl Parathion (MP)
and heavy metals like cadmium. MP-degrading enzymes are found in microbes such as Pseudomonas
sp. A3 (Ramanathan and Lalithakumari 1999), Plesiomonas sp. M6 (Zhongli et al. 2001), and
Pseudomonas sp. SMSP-1 (Shen et al. 2010). They convert Methylparathion to paranitrophenol