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  Section: General Biotechnology / Biotechnology & Environment
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Environmental Biotechnology

Bioremediation of Xenobiotics
Use of pesticides has benefited the modem society by improving the quantity and quality of the worlds' food production. Gradually, pesticide usage has become an integral part of modern agriculture system. Many of the artificially made complex compounds i.e. xenobiotics persist in environment and do not undergo biological transformation. Microorganisms play an important role in degradation of xenobiotics, and maintaining of steady state concentrations of chemicals in the environment. The complete degradation of a pesticide molecule to its inorganic components that can be eventually used in an oxidative cycle removes its potential toxicity from the environment. However, there are two objectives in relation to biodegradation of xenobiotics: (i) how biodegradation activity arises, evolves and transferred among the members of soil microflora, and (ii) to device bioremediation methods for removing or detoxifying high concentration of dangerous pesticide residues (Gupta and Mukerji, 1998).

The characters of pesticide degradation of microorganisms are located on plasmids and transposons, and are grouped in clusters on chromosome. Understanding of the characters provides clues to the evolution of degradative pathways and makes the task of gene manipulation easier to construct the genetically engineered microbes capable of degrading the pollutants.

Microbial Degradation of Xenobiotics
Biodegradation of pesticides occurs by aerobic soil microbes. Pesticides are of wide varieties of chemicals e.g. chlorophenoxyalkyl caboxylic acid, substituted ureas, nitrophenols, tri-azines, phenyl carbamates, orga-nochlorines, organophosphates, etc. Duration of persistence of her­bicides and insecticides in soil is given in Table 21.2. Otganophos-phates (e.g. diazion, methyl par-athion and parathion) are perhaps the most extensively used insecti­cides under many agricultural sys­tems. Biodegradation through hy­drolysis of p-o-aryl bonds by Pseudomonas diminuta and Flavobacterium are considered as the most significant steps in the detoxification of organophospho-rus compounds. Organomercurials (e.g. Semesan, Panodrench, Panogen) have been practiced in agriculture since the birth of fun­gicides. Several species of Aspergillus, Penicillium and Trichoderma have been isolated from Semesan-treated soil. More­over, they have shown ability to grow over 100 ppm of fungicide in vitro. The major fungicides used in agriculture are water soluble derivatives such as Ziram, Ferbam, Thiram, etc. All these are de­graded by microorganisms.

Outline of aerobic and anaerobic degradation of pentachlorophenol.

Fig. 21.7. Outline of aerobic and anaerobic degradation of pentachlorophenol.

Pentachlorophenol (PCP) is a broad spectrum biocide which has been used as fungicide, insecticide, herbicide, algicide, disinfectant and antifouling agent. Bioreactors containing alginate immobilized + Polyurethane foam immobilized PCP degrading Flavobacterium (ATCC39723) cells have been used to remove PCP from contaminated water. Absorption of PCP by Polyurethane immobilized matrix plays a role in reducing the toxicity of PCP. Flavobacterium re­moved and detoxified PCP (Zhong -Cheng, 1994). In other experiment P. chrysosporium enzyme (ligninase) has been found to dehalogenate PCP. Steps of PCP degradation has been shown in Fig. 21.7.





In situ bioremediation



Intrinsic bioremediation



Engineered in situ bioremediation


Ex situ bioremediation



Solid phase system (composting, composting process)



Slurry phase system (aerated laggons, low shear airlift reactor)



Factors affecting slurry phase bioremediation


Bioremediation of hydrocarbon



Use of mixture of bacteria



Use of genetically engineered bacterial strains 


Bioremediation of Industrial wastes



Bioremediation of dyes



Bioremediation of heavy metals



Bioremediation of coal waste through VAM fungi


Bioremediation of xenobiotics



Microbial degradation of xenobiotics



Gene manipulation of pesticide-degrading microorganisms

Utilization of sewage, and agro-wastes


Production of single cell protein


Biogas from sewage


Mushroom production on agro-wastes



Microbial leaching (bioleaching)   


Microorganisms used in leaching


Chemistry of leaching



Direct leaching



Indirect leaching


Leaching process (slope leaching heap leaching in situ leaching)


Examples of bioleaching



Copper leaching



Uranium leaching



Gold and silver leaching



Silica leaching

Hazards of environmental engineering


Survival of released GMMs in the environment



Adaptive mutagenesis in GMMs



Gene transfer from GMMs into other microorganisms



Gene transfer via conjugative transposons



Effect of environmental factors on gene transfer


Ecological impact of GMMs released into the environment



Growth inhibition of natural strains



Growth stimulation of indigenous strains



Replacement of natural strains


Monitoring of GEMs in the environment



Risk assessment of the GEMs released into the environment

21.2. Duration of persistence of insecticides and herbicides in soil.
Biocides Time taken for
75-100% disappearance

A. Chlorinated insecticides


DDT (l,l,l-trichloro-2,2-bis-(p-chlorophenyl) ethane)

4 years


3 years


5 years


2 years

Lindane (hexachloro-cyclohexane)

3 years

B. Organophosphate insecticides



12 years


1 week


1 week

C. Herbicides


2,4-D (2,4-dichlorophenoxyacetic acid)

4 weeks


30 weeks


40 weeks


48 weeks


1.5 years

Source : Madigan et al. (1997).

DDT (l,l,l-trichlofo-2,2-bis (p-chlorophenyl) ethane) is an insecticide that persists in soil for four years. Degradation pathway of DDT involves an initial dechlorination of the trichloromethyl group to form 1,1-dichloro 2-ethane which then undergoes further dechlorination, oxidation and decarboxylation to form bis methane. Subsequent cleavage of one of the normal aromatic rings yields p-chlorophenyl acetic acid, which may also undergo ring cleavage. Microorganisms associated with DDT degradation are Aspergillus flavus, Fusarium oxysporum, Mucor aternans, P. chrysosporium, Trichoderma viride, etc. Environmental factors including pH, temperature, bioavailability, nutrient supply and oxygen availability affect biodegradation of pesticides.

Gene Manipulation of Pesticide-degrading Microorganisms
Day-by-day the number of xenobiotic-degrading microorganisms is increasing. However, pesticide-degrading genes of only a few microorganisms have been characterized. Most of genes responsible for catabolic degradation are located on the chromosomes, but in a few cases these genes are found on plasmids or transposons. Chakrabarty and Gunsalus (1971) found that camphor degrading genes of Pseudomonas putida are located on plasmid. Soon after, naphthalene (NAH) degrading plasmid was isolated. Discovery of these genes made it possible to construct a new genetically engineered strain of P. putida that alone was potent to degrade camphor, naphthalene, xylene, toluene, octanes and hexanes. For detailed discussion, see Abatement of pollution.

For the first time pesticide degradation through plasmid mediated genetically engineered microorganism was reported by Chakrabarty et al (1981). Nagata et al (1993) have also cloned and sequenced two genes involved in early steps of Y-HCH degradation in UT26. The linAgene encodes Y-HCH dehydrochlorinase which converts Y-HCH to 1,2,4TCB via Y-PCCH and 1,4-TCDN. The linBgene encodes 1,4-TCDN chlorohydrolase which converts 1,4-TCDN to 2,4-DDOL via 2,4,5-DNOL. This gene is a member of the haloalkanedehalogenase family with a broad range specificity for substrate. The genetically engineered P. putida comprises of both linA and linB genes (Nagata et al, 1993).

The pod gene, initially isolated from Flavobacterium sp. ATCC27551 and Pseudomonas diminuta, has been well characterized. It is associated for degradation of pesticides such as parathion, methylparathion, etc. Sims et al (1990) transferred a recombinant DNA plasmid containing pod gene into a fungus, Gliocladium virens which expressed at low level. G. virens is a useful soil saprophyte which has shown a strong mycoparasitic activity against many fungal pathogens. Strains of G. virens are potential for use in the bioremediation of contaminated soil. Optimization of pod expression in bioremedially useful organism such as P. chrysosporium, G. virens, etc. holds a great promise in lessening the pesticide pollution.


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