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Basic studies and applications on bioremediation of DDT: A review

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DOI: 10.1016/j.ibiod.2011.07.011

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International Biodeterioration & Biodegradation 65 (2011) 921e930

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International Biodeterioration & Biodegradation

journal homepage: www.elsevier.com/locate/ibiod

Review

Basic studies and applications on bioremediation of DDT: A review

Adi Setyo Purnomoa, Toshio Morib, Ichiro Kameic, Ryuichiro Kondob, *
a
Department of Chemistry, Faculty of Mathematics and Natural Sciences, Institut Teknologi Sepuluh Nopember (ITS), Kampus ITS Sukolilo, Surabaya 60111, Indonesia
b
Department of Agro-Environmental Sciences, Faculty of Agriculture, Kyushu University, 6-10-1 Hakozaki, Higashi-ku, Fukuoka 812-8581, Japan
c
Faculty of Agriculture, University of Miyazaki, 1-1 Gakuen Kibanadai-nishi, Miyazaki 889-2192, Japan

a r t i c l e i n f o a b s t r a c t

Article history: The persistent insecticide DDT (1,1,1-trichloro-2,2-bis (4-chlorophenyl) ethane) has been widely used for
Received 7 June 2011 pest control in the management of mosquito-borne malaria and is still used for that purpose in some
Received in revised form tropical countries. Considering the potential for negative effects due to DDT contamination, it is neces-
12 July 2011
sary to determine effective methods of remediation. Several methods have been used to degrade or
Accepted 12 July 2011
Available online 16 September 2011
transform DDT into less toxic compounds. Bacteria and white-rot fungi (WRF) have been shown to
enhance the degradation process in soil using both pure and mixed cultures. Recently, a biological
approach has been used as an environmentally-friendly treatment, using new biological sources to
Keywords:
Bioremediation
degrade DDT, e.g. brown-rot fungi (BRF), cattle manure compost (CMC) and spent mushroom waste
Degradation (SMW). In this review, the abilities of BRF, CMC and SMW to degrade DDT are discussed, including the
DDT mechanisms and degradation pathways. Furthermore, application of these sources to contaminated soil
Brown-rot fungi is also described. The review discusses which is the best source for bioremediation of DDT.
Compost Ó 2011 Elsevier Ltd. All rights reserved.
Spent mushroom waste

Contents

1. Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 922
2. Degradation of DDT by brown-rot fungi . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 922
2.1. Identification of metabolic products . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 922
2.2. Involvement of the Fenton reaction in DDT degradation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 922
2.3. Degradation of DDT and its metabolites by the chemical Fenton reaction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 923
2.4. Mechanisms of involvement of the Fenton reaction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 923
2.5. DDT degradation pathway . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 924
3. DDT degradation potential of cattle manure compost . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 924
3.1. DDT degradation by cattle manure compost . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 924
3.2. Isolation and identification of fungi from cattle manure compost . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 925
3.3. DDT degrading assay on isolated fungi . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 925
4. Degradation of DDT by mushroom waste medium . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 925
4.1. Degradation and mineralization of DDT . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 926
4.2. Ligninoliytic enzymes activities . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 926
5. Application on contaminated soil . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 926
5.1. Application of brown-rot fungi in artificially DDT-contaminated soil . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 926
5.2. Degradation of artificially DDT-contaminated soil by cattle manure compost and its isolated fungi . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 927
5.3. Degradation of artificially DDT-contaminated soil by spent mushroom waste of Pleurotus ostreatus . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 927
5.4. Degradation of DDT in historically contaminated soil by spent mushroom waste of Pleurotus ostreatus . . . . . . . . . . . . . . . . . .. . . . . . . . . . . . . . . . 928
6. Conclusions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 928
Acknowledgments . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 928
References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 928

* Corresponding author. Tel./fax: þ81 92 642 2811.

E-mail addresses: adi.spurnomo@yahoo.com, adi_setyo@chem.its.ac.id (A.S. Purnomo), ryukondo@agr.kyushu-u.ac.jp (R. Kondo).

0964-8305/$ e see front matter Ó 2011 Elsevier Ltd. All rights reserved.
doi:10.1016/j.ibiod.2011.07.011
 Author's personal copy
922 A.S. Purnomo et al. / International Biodeterioration & Biodegradation 65 (2011) 921e930
1. Introduction
DDT (1,1,1-trichloro-2,2-bis (4-chlorophenyl) ethane) was intro-
duced as an insecticide during World War II to combat mosquitoes
which spread malaria and typhus (Busvine 1989; Boul 1995; Foght
et al. 2001). DDT was the first synthetic insecticide and gained
popularity all over the world. However, its use has recently been
prohibited in most countries because of its negative impact on wildlife
and its ill effects on human health via the food web (Kale et al. 1999;
Foght et al. 2001). Low direct exposure to DDT leads to symptoms such
as headache, nausea, vomiting, confusion, and tremors. Furthermore,
accumulation of DDT in the body can affect the nervous system,
increase tumor production, and is associated with pancreatic cancer
(Turusov et al. 2002; Gautam and Suresh 2007). The presence of
chlorine atoms in DDT and its metabolites, in conjunction with their
low solubility and tendency to partition preferentially into the lipo-
philic phase, makes them highly ecotoxic, especially to higher
organisms. However, DDT is still being used in some developing
countries for essential public health purposes (Foght et al. 2001). The
United States Environmental Protection Agency (EPA) has classified
DDT and its metabolite products, DDD (1,1-dichloro-2,2-bis (4-
chlorophenyl) ethane) and DDE (1,1-dichloro-2,2-bis (4-
chlorophenyl) ethylene), as priority pollutants (Sayles et al. 1997;
Foght et al. 2001). Considering the potential for negative effects, it is
necessary to address the environmental persistence of this insecticide
and to find effective methods of remediation.
The removal of DDT from contaminated soils has become an
environmental priority, and both physicochemical and biological
remediation processes have been studied. Although chemical and
physical treatments are more rapid than biological treatments, they
are generally more destructive and intrusive to affected soils, more
energy intensive, and often more expensive than bioremediation
(Foght et al. 2001). Considering the bioremediation options, several
bacteria and white-rot fungi (WRF) have been shown to enhance
degradation processes in soil, using both pure and mixed cultures
(Guenzi and Beard 1967; Wedemeyer 1967; Subba-Rao and Alex-
ander 1985; Bumpus and Aust 1987; Fernando et al. 1989; Bumpus
et al. 1993; Xu et al. 1994; Boul 1996; Hay and Focht 2000; Kama-
navalli and Ninnekar 2004; Thomas and Gohil 2011; Xie et al. 2011).
Recently, bioremediation of DDT was performed using an
environmentally-friendly biological approach by means of some
new biological sources to degrade DDT, including brown-rot fungi
(BRF, Purnomo et al. 2008, 2010a, 2011), cattle manure compost
(CMC, Purnomo et al. 2010b) and spent mushroom waste (SMW,
Purnomo et al. 2010c). BRF including Gloeophyllum trabeum, Fomi-
topsis pinicola and Daedalea dickinsii have shown a good ability to
degrade DDT, with the Fenton reaction being an important part of
the system (Purnomo et al. 2008, 2010a, 2011, Villa et al. 2008).
Mucor circinelloides and Galactomyces geotrichum were isolated from
CMC, which is good at degrading DDT in artificially contaminated
soil (Purnomo et al. 2010b). In addition, mineralization of DDT by
SMW of Pleurotus ostreatus in artificially contaminated soil has been
reported, and proposed as a promising source of DDT bioremedia-
tion (Purnomo et al. 2010c). Previously, there have been two reviews
on microbial degradation of DDT: one by Aislabie et al. (1997) which
covers literature up to 1997 about microbial degradation of DDT and
its residues, and the other by Foght et al. (2001) which covers
literature up to 2001 about bioremediation of DDT-contaminated
soil. In this article, we have updated this knowledge by reviewing
recent works on the bioremediation of DDT.
2. Degradation of DDT by brown-rot fungi
Unlike the process using bacteria and WRF, DDT biodegradation
by BRF has received relatively little attention. Because BRF do not
have ligninolytic enzymes, it has been proposed that they use
hydroxyl radicals produced via the Fenton reaction for degradation
of various compounds (Purnomo et al. 2008, 2010a). In BRF,
extracellular Fenton-type mechanisms have been reported to be
involved in the degradation of several xenobiotic compounds
(Wetzstein et al. 1997; Kerem et al. 1999; Wetzstein et al. 1999;
Schlosser et al. 2000; Jensen et al. 2001; Newcombe et al. 2002).
Twelve species of BRF were screened for their ability to degrade
DDT in which G. trabeum, F. pinicola and D. dickinsii showed the
greatest abilities to degrade DDT (Purnomo et al. 2008). In this
section, characterization of the major metabolic products, clarifi-
cation of the degradation pathways, and an examination of the role
of the Fenton reaction is discussed in detail.
2.1. Identification of metabolic products
The metabolic products were identified by HPLC and GC/MS on
the basis of the retention time and absorption maximum at specific
wavelengths in comparison with standards (Table 1). DDE, DDD,
DBP (4,4-dichlorobenzophenone) and DBH (4,40 -dichlorobenzhy-
drol) were detected as metabolic products of DDT degradation by
G. trabeum. However, when D. dickinsii was used the metabolic
products were DDE, DDD and DDMU (1-chloro-2,2-bis (4-
chlorophenyl) ethylene), while only DDD was detected when
F. pinicola was used. These results show that the different fungi have
different transformation pathways of DDT (Purnomo et al. 2008,
2010a).
The mineralization of [14C]DDT by G. trabeum, F. pinicola and
D. dickinsii was evaluated. Less than 0.25% of the initial [14C]DDT
(378 kdpm) was mineralized to [14C]CO2. Because these minerali-
zation rates are very low, it is unlikely that the fungi can mineralize
DDT (Purnomo et al. 2008, 2010a). Bumpus and Aust (1987)
reported that the mineralization of [14C]DDT by G. trabeum
was < 0.1% in both nitrogen-deficient and nitrogen-sufficient
cultures. Thus, it appears that BRF cannot mineralize DDT. In the
case of the DDT degradation pathway in the WRF P. chrysosporium,
DBP appears to be an end-intermediate compound, which is
subsequently converted by “unknown” ring cleavage processes to
produce CO2 (Bumpus and Aust 1987).
2.2. Involvement of the Fenton reaction in DDT degradation
DDT degradation by G. trabeum was enhanced by an iron-
dependent reaction, resulting in the production of different meta-
bolic products. DBP was not detected as a metabolic product of DDT
in the medium without FeSO4, indicating that DBP was produced by
the iron-dependent reaction (Purnomo et al. 2008). For F. pinicola,
where DDD was detected as a metabolic product, degradation of
DDT was not enhanced by an iron-dependent reaction. Degradation
Table 1
DDT degradation ability of brown-rot fungi in pure culture (Purnomo et al. 2008,
2010c).
Brown-rot DDT decrease Metabolites (%)
fungi (%)
DDE DDD DBP DBHa DDMUa
G. trabeum 61.8  1.4 30.1  1.5 22.6  3.8 13.9  0.8 þ 
F. pinicola 63.1  4.2  61.8  3.1   
D. dickinsii 46.8  4.2 24.1  3.9 19.9  3.1   þ
Initial concentration of DDT is 0.25 mmol. Data are presented as mean  standard devi-
ations (n ¼ 3). DDT (1,1,1-trichloro-2,2-bis (4-chlorophenyl) ethane); DDE (1,1-dichloro-
2,2-bis (4-chlorophenyl) ethylene); DDD (1,1-dichloro-2,2-bis (4-chlorophenyl) ethane);
DBP (4,4-dichlorobenzophenone); DBH (4,4-dichlorobenzhydrol); DDMU (1-chloro-2,2-
bis (4-chlorophenyl) ethylene).
a
The data were determined by GC/MS: () undetectable; (þ) detectable.
 Author's personal copy
A.S. Purnomo et al. / International Biodeterioration & Biodegradation 65 (2011) 921e930
of DDT by D. dickinsii also did not require an iron-dependent
reaction (Purnomo et al. 2010a).
For further evidence of the involvement of the Fenton reaction,
the concentration of hydroxyl radicals was determined. F. pinicola
and D. dickinsii produced lower concentrations of hydroxyl radicals
compared with G. trabeum (Purnomo et al. 2008, 2010a). Further-
more, the presence of mannitol as a hydroxyl radical scavenger
inhibited DDT degradation by G. trabeum but not by D. dickinsii and
F. pinicola. Since D. dickinsii and F. pinicola produced low amounts of
hydroxyl radicals, this suggests that the production of hydroxyl
radicals alone is not enough to degrade DDT (Purnomo et al. 2010a).
For F. pinicola, addition of Fe2þ to the culture caused the production
of hydroxyl radicals to increase. However, DDT degradation and
DDD production decreased. Since an increase in hydroxyl radicals
caused a decrease in DDT degradation and DDD production, this
suggests that hydroxyl radicals might inhibit the ability of fungus to
degrade DDT. F. pinicola might be sensitive to hydroxyl radicals
since the radicals are known to attack many biomolecules due to
their high reactivity. Besides, increasing of Fe2þ concentration
enhanced DDT degradation by D. dickinsii due to an increase in the
number of hydroxyl radicals. The results indicate that if the
required amount of hydroxyl radical is reached, D. dickinsii can
degrade DDT via the Fenton reaction (Purnomo et al. 2010a). A
similar tendency was seen in G. trabeum. The added Fe2þ might be
oxidized rapidly to yield Fe3þ as the predominant form of iron in
the cultures. G. trabeum can reduce Fe3þ and thus drive the Fenton
reaction extracellularly (Jensen et al. 2001). Furthermore, an
extracellular metabolite; 2,5-dimethoxy-1,4-benzoquinone
(DMBQ) was detected from G. trabeum as a key component for
the Fenton reaction cycle. This finding was supported by Kerem
et al. (1999), who identified DMBQ as an important component of
extracellular oxidative systems in G. trabeum to operate a redox
cycle. The fungal mycelium reduced DMBQ to 2,5-dimethoxy-1,4-
hydroquinone (DMHQ). DMHQ reduces Fe3þ to Fe2þ and produces
extracellular H2O2 via a nonenzymatic reaction. This pathway
provides evidence that G. trabeum uses a benzoquinone-driven
Fenton reaction to produce the hydroxyl radical (Kerem et al.
1999). However, DMBQ was not detected in the F. pinicola and
D. dickinsii cultures. This demonstrates that these fungi produce
Fig. 1. Transformation of DDT (R ¼ Cl) or DDD (R ¼ H) or DDE (R ¼ OH) to DBP by the Fenton reaction (Purnomo et al. 2008).
923
a low level of hydroxyl radicals due to their lack of a Fenton reaction
cycle system. Supplemented Fe2þ could be used directly to stimu-
late the Fenton reaction causing oxidation to Fe3þ. However, since
the fungi lack a redox system, the reaction will not occur for long
(Purnomo et al. 2010a).
Different levels of hydroxyl radical production in cultures may
also be caused by different amounts of H2O2 production by fungi.
G. trabeum produced about 50e70 mM of H2O2 (Purnomo et al.
2008). However, lower amounts of H2O2 were detected from
F. pinicola (16 mM) and D. dickinsii (21 mM) (Purnomo et al. 2010a).
Since G. trabeum has a Fenton reaction cycle system, the higher
amount of hydroxyl radical production in G. trabeum is caused by
the higher amount of H2O2. These findings support the supposition
that a benzoquinone-driven Fenton reaction cycle is important for
G. trabeum to degrade DDT.
2.3. Degradation of DDT and its metabolites by the chemical Fenton
reaction
To prove that DBP was produced by the Fenton reaction, DDT
and its metabolites (DDE, DDD and DBP) were treated via the
chemical Fenton reaction (CFR) in the same manner as in the fungal
culture (70 mM H2O2 and 100 mM FeSO4). DDT, DDE and DDD
(0.25 mmol) were degraded approximately 32%, 34%, and 44%,
respectively. DBP was detected as the main metabolic product
approximately 20%, 31%, and 37%, respectively (Purnomo et al.
2008). These results indicate that a Fenton reaction is possibly
involved in the DDT degradation pathway, particularly in the
transformations of DDT, DDE and DDD to DBP. In application,
approximately 75% of initial DDT in slurry soil (1.6 mg g1) was
degraded by CFR (Villa et al. 2008).
2.4. Mechanisms of involvement of the Fenton reaction
A mechanism of the direct transformations of DDT, DDE and
DDD to DBP was proposed (Fig. 1). For DDT, the benzylic hydrogen
located between the two rings is expected to be easily attacked by
the hydroxyl radical, yielding a resonance-stabilized, carbon-
centered benzylic radical. This radical will react with oxygen,
 Author's personal copy
924 A.S. Purnomo et al. / International Biodeterioration & Biodegradation 65 (2011) 921e930
yielding a peroxyl radical, which can then attack hydrogen from the
surroundings to yield the peroxide. In the presence of Fe2þ, the
peroxide will be reduced to an alkoxyl radical, which will undergo
beta scission to yield the benzophenone (DBP) and trichloromethyl
radical (CCl3). The same pathway could likely operate in the case of
DDD, yielding DBP and dichloromethyl radical (CHCl2) (Purnomo
et al. 2008). For DDE, the transformation mechanism involving
the hydroxyl radical is based on the anti-Markovnikov rule. The
hydroxyl radical is assumed to be electron-deficient and electro-
philic. The alkene double bond is attacked by the hydroxyl radical,
yielding a resonance-stabilized, carbon-centered benzylic radical.
This radical will react with oxygen, yielding a peroxyl radical, which
can then attack hydrogen from the surroundings to yield the
peroxide. In the presence of Fe2þ, the peroxide will be reduced to an
alkoxyl radical, which will undergo beta scission to yield benzo-
phenone (DBP) and a dichlorohydroxymethyl radical (COHCl2,
Purnomo et al. 2008).
2.5. DDT degradation pathway
On the basis of the identification of the metabolic products, the
DDT degradation pathway in BRF is proposed as shown in Fig. 2.
G. trabeum initially dehydrochlorinates DDT to form DDE, followed
by hydrogenation to DDD. DDD then undergoes oxidative
dechlorination to DBP, followed by reduction to DBH (Purnomo
et al. 2008, 2010a). This pathway differs from the proposed
pathways in bacteria and other fungi described above, particularly
in the transformation of DDE to DDD (Purnomo et al. 2008). As
with G. trabeum, D. dickinsii also initially dehydrochlorinates DDT
to form DDE, followed by hydrogenation to DDD, and then
undergoes dechlorination to DDMU. This pathway differs from the
proposed pathway in G. trabeum described above, particularly in
the transformation of DDD to DDMU. F. pinicola dechlorinates DDT
to DDD where DDD is the end product. These results indicate that
DDT degradation pathways differ between BRF (Purnomo et al.
2010a).
DDT, DDD and DDE may also be directly transformed to DBP via
the involvement of a Fenton reaction (Purnomo et al. 2008, 2010a).
A benzoquinone-driven Fenton reaction cycle is a key to BRF
degradation of DDT. Because BRF do not have lignin-degrading
enzymes, it is also possible that BRF use an enzymatic system
that differs from that of WRF.
Fig. 2. Proposed DDT degradation pathway of brown-rot fungi. Thin white, open, and dotted arrows indicate transformation by G. trabeum, D. dickinsii and F. pinicola, respectively.
Thick black arrows show transformation by Fenton reaction. DDT (1,1,1-trichloro-2,2-bis (4-chlorophenyl) ethane); DDE (1,1-dichloro-2,2-bis (4-chlorophenyl) ethylene); DDD (1,1-
dichloro-2,2-bis (4-chlorophenyl) ethane); DBP (4,4-dichlorobenzophenone); DBH (4,4-dichlorobenzhydrol); DDMU (1-chloro-2,2-bis (4-chlorophenyl) ethylene) (Purnomo et al.
2008, 2010a).
3. DDT degradation potential of cattle manure compost
Since composts are rich sources of xenobiotic-degrading
microorganisms, they can be used to degrade pollutants or to
transform pollutants into less toxic substances. Composts are
reportedly capable of degrading a variety of organic contaminants
(Laine and Jorgensen 1996; Semple and Fermor 1997; Semple et al.
1998; Eggen 1999; Reid et al. 2002; Lau et al. 2003; Puglisi et al.
2007). However, DDT biodegradation by composts has received
relatively little attention (Kantachote et al. 2003; Lourencetti et al.
2007). In this section, the ability of cattle manure compost (CMC) to
degrade DDT is discussed. A detailed investigation into degradation
processes and the biodegradation ability of each composting stage
(mesophilic, thermophilic, and maturation) was performed. DDT-
degrading fungi were isolated, identified, and tested for their
ability to degrade DDT.
3.1. DDT degradation by cattle manure compost
Under optimal conditions, composting proceeds through
three stages: (1) the mesophilic stage, which occurs for a few
days; (2) the thermophilic stage, which can last from a few days
to several months; and (3) the maturation stage, which lasts for
several months (Tuomela et al. 2000). Cattle manure compost
(CMC) was made from cattle manure, rice bran, and pearlite.
After mixing, some of the composting material was taken and
used as the mesophilic stage material. The remaining compost
material was placed into a reactor for composting, with air
supplied at a rate of 0.5 L min1. Once the temperature increased,
some of the compost material was taken from the reactor and
used as the thermophilic stage material. After the temperature
dropped and remained at a relatively constant level, the compost
material was taken and used as maturation stage material
(Purnomo et al. 2010b).
During composting, the CMC passed from the mesophilic to the
thermophilic stage after 36 h. The maximum temperature occurred
at 9 h and then steadily dropped. After 120 h, the temperature
stabilized, indicating the maturation stage. During the initial 45 h,
the mesophilic and thermophilic stages were critical points in the
DDT degradation process when thermophilic microbes degrade
DDT more effectively than those present during the mesophilic and
maturation stages (Purnomo et al. 2010b). At the beginning of
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A.S. Purnomo et al. / International Biodeterioration & Biodegradation 65 (2011) 921e930 925

composting, mesophilic bacteria predominate, but after the Table 2

temperature increases to over 40  C, thermophilic bacteria and DDT degradation ability of isolated fungi from cattle manure compost at different
temperature (Purnomo et al. 2010b).
fungi also appear in the compost. When the temperature exceeds
60  C, microbial activity decreases dramatically, but after the Strain no. Name of species Degradation rate (%)
compost has cooled, mesophilic bacteria and actinomycetes again 30  C 60  C
dominate (McKinley and Vestal 1985; Strom 1985). Mesophilic
To investigate the process of DDT degradation by CMC in 1CMC1 Galactomyces geotrichum 83.6  3.4aA 90.1  2.9aA
detail, the ability of the materials at each stage of the CMC 1CMC2 Mucor circinelloides 81.9  7.2aA 72.9  1.2bB
1CMC3 Mucor circinelloides 85.9  2.2aA 81.9  5.2aA
process was investigated. The experiments were conducted at 30
1CMC4 Mucor circinelloides 82.1  3.7aA 79.6  3.2aA
or 60  C, temperatures that are representative of mesophilic/ 1CMC5 Mucor circinelloides 89.4  2.7aA 55.4  5.9bB
maturation or thermophilic conditions, respectively. The ther- 1CMC6 Mucor circinelloides 95.3  1.4aA 88.7  4.1bB
mophilic stage material demonstrated the highest degradation 1CMC7 Mucor circinelloides 89.9  1.5aA 80.4  3.4aB
ability, indicating that thermophilic stage materials degraded DDT 1CMC8 Mucor circinelloides 88.5  8.2aA 82.9  4.2aA
more effectively than materials at the other stages. The ability of Thermophilic
thermophilic microbes to degrade DDT may be more effective 2CMC1 Galactomyces geotrichum 85.1  2.1aA 91.9  1.1aB
than those present during the mesophilic/maturation stages 2CMC2 Galactomyces geotrichum 80.4  3.9aA 93.5  2.6aB

(Purnomo et al. 2010b). In the composting environment, ther- Maturation

mophilic microbes are an important biodegradation agent, even
in small populations (McKinley and Vestal 1985; Strom 1985;
Tuomela et al. 2000). The results provide direct evidence that
during the composting process, DDT was degraded to a greater
extent during the thermophilic stage, with DDD was detected as
a major metabolic product (Baczynski et al. 2010). However, since
DDD production was not equivalent to DDT degradation, it is
likely that some other metabolic by-products were produced. It is
possible that DDD could be transformed into other metabolic
products such as DBP (Baczynski et al. 2010; Purnomo et al.
2010b).
The mineralization of [14C]DDT (378 kdpm) by materials at each
stage of the CMC at 30 and 60  C was also studied. The thermophilic
stage showed the highest mineralization ability at 60  C. The results
also provide direct evidence that during the composting process,
DDT was mineralized to a greater extent during the thermophilic
stage (Kantachote et al. 2003; Purnomo et al. 2010b). However,
because the mineralization rate is very low (less than 1.00%), it is
assumed that CMC could not mineralize DDT (Purnomo et al.
2010b).
3.2. Isolation and identification of fungi from cattle manure
compost
Fungi play a very important role in the biodegradation and
conversion processes during composting. In addition, fungi use
many carbon sources and can survive in extreme conditions, and
are mainly responsible for compost maturation (Sparling et al.
1982; McKinley and Vestal 1985; Strom 1985; Wiegant 1992;
Tuomela et al. 2000). However, several bacteria has been isolated
and identified from contaminated soils such as Staphylococcus
(Songkong et al. 2008), Basillus, Clostridium, Escherichia (Purnomo
et al. 2010b), and Alcaligenes (Xie et al. 2011). Bacteria were more
sensitive to DDT than actinomycetes and fungi. DDT-resistant
bacterial strains showed more promise in degrading DDT than
isolated fungal strains (Kantachote et al. 2003).
Several fungi were isolated from CMC, which most of the fungi
isolated from the mesophilic and maturation stages were 99e100%
identical at the nucleotide level to Mucor circinelloides (Table 2). M.
circinelloides is one of the most commonly occurring fungi of the
Mucor genus (Purnomo et al. 2010b). Most of the fungi belonging to
the Mucor genus have been reported to possess the ability to
degrade xenobiotic compounds (Anderson et al. 1970; Anderson
and Lichtenstein 1971, 1972; Shetty et al. 2000; Su et al. 2006;
Szewczyk and D1ugon  ski 2009). In addition, all isolated fungi
from the thermophilic stage were 100% identical with Galactomyces
geotrichum (Table 2) (Purnomo et al. 2010b). This fungus has been
reported to degrade dyes (Jadhav et al. 2008a,b).
3CMC1 Mucor circinelloides 84.5  1.2aA 69.8  1.4bB
3CMC2 Mucor circinelloides 81.9  5.2aA 75.1  4.8aA
Initial concentration of DDT is 0.25 mmol. Data are presented as mean  standard
deviations (n ¼ 3). Data followed by the same minor letter on each column or by the
same capital letter on each row are not statistically different from each other
(P < 0.05).
3.3. DDT degrading assay on isolated fungi
The ability of isolated fungi to degrade DDT in PDB medium at
30 or 60  C was determined. The fungi isolated from materials at
the thermophilic stage (G. geotrichum) exhibited a significantly
higher degradation ability at 60  C (Table 2) (Purnomo et al. 2010b).
This indicated that thermophilic fungi have greater activity at
a higher temperature although they have adequate activity at
a lower temperature. Most of the fungi isolated from materials at
the mesophilic and maturation stages (M. circinelloides) demon-
strated a greater ability to degrade DDT at 30  C (Table 2). These
results indicate that even though they have activity during the
thermophilic stages, a lower temperature is more suitable for their
activity. This finding demonstrate that most of the fungi isolated
from CMC materials possess the ability to degrade DDT at 30 and
60  C. Approximately 16% and 6% of DDD and DBP were detected as
metabolic products, respectively (Purnomo et al. 2010b). Based on
identification of metabolic products, DDT was initially reductively
dechlorinated to form DDD, followed by oxidative dechlorination to
DBP. The processes involve single electron transfer, removal of
a chlorine ion, and formation of an alkyl radical (Archer 1973; Foght
et al. 2001). In addition, soil fungus Fusarium solani had ability to
degrade DDT and its metabolites such as DDD, DDE, DDOH and DBP
(Mitra et al. 2001).
4. Degradation of DDT by mushroom waste medium
There have been several studies of DDT degradation by WRF in
liquid cultures, in which the process has been assumed to be
mediated by the fungal lignin-degrading system (Wedemeyer
1967; Bumpus and Aust 1987; Fernando et al. 1989; Bumpus et al.
1993; Boul 1996; Hay and Focht 2000; Kamanavalli and Ninnekar
2004; Chung et al. 2009; Purnomo et al. 2010c, Xiao et al. 2011).
WRF species produce different enzymes depending on their genetic
and growth conditions. Key degradation enzymes include lignin
peroxidase (LiP), manganese peroxidase (MnP), versatile peroxi-
dase (VP) and laccase. The lignin and xenobiotic degradation
potential of these enzymes are well documented (Lamar 1992; Vyas
et al. 1994; Zhao and Yi 2010). However, bioremediation by fungi in
liquid cultures requires costly preparations prior to application,
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926 A.S. Purnomo et al. / International Biodeterioration & Biodegradation 65 (2011) 921e930
especially for the medium. To evaluate the effectiveness of biore-
mediation of DDT-contaminated soil, several parameters must be
investigated first, and the degradation ability might be reduced in
contaminated soil compared with liquid cultures (Purnomo et al.
2010c). In mushroom production, 5 kg of spent mushroom waste
(SMW) is generated from the production of every kilogram of
mushrooms (Semple et al. 2001). High levels of residual nutrients
and enzymes are still present in SMW. When using WRF for
bioremediation, the availability of fungal inocula is a practical
consideration and the use of SMW could be advantageous due to
the low cost and environmentally-friendly treatment (Purnomo
et al. 2010c). SMWs have been well documented in their degrada-
tion of some xenobiotics (Semple et al. 1998; Eggen 1999; Law et al.
2003; Puglisi et al. 2007; Chung et al. 2009).
4.1. Degradation and mineralization of DDT
Since high levels of residual nutrients and enzymes remain in
SMW, it might be used advantageously as a low-cost bioremedia-
tion tool to degrade DDT. Various cultivated fungal inocula were
screened for their ability to degrade DDT. Fungal substrate from
Pleurotus ostreatus showed the highest ability to degrade DDT
(Chung et al. 2009; Purnomo et al. 2010c). Since >25% of worldwide
mushroom production is Pleurotus mushrooms, it would be
advantageous to use Pleurotus SMW. Thus, the fungal substrate
generated before mushroom production and SMW generated after
mushroom production from P. ostreatus were used in the detailed
investigation. SMW was significantly better at degrading DDT than
the fungal substrate. Eggen (1999) has reported that degradation of
PAHs by SMW from P. ostreatus was higher than that by the fungal
substrate before mushroom production.
Mineralization of [U-14C]DDT was also investigated. [U-14C]DDT
(378 kdpm) was mineralized by 4.4% and 5.1% during the 56
d incubation by the P. ostreatus fungal substrate and SMW,
respectively (Purnomo et al. 2010c). Bumpus and Aust (1987)
reported that [U-14C]DDT was 6% mineralized during a 30 d incu-
bation at room temperature in a low nitrogen liquid culture of
P. ostreatus. This suggests that the mineralization of [U-14C]DDT in
solid and liquid matrices is not very different. Based on the
degradation and mineralization results, the degradation of DDT by
P. ostreatus SMW is more effective than by the fungal substrate
before production (Purnomo et al. 2010c).
4.2. Ligninoliytic enzymes activities
Bioremediation activity is dependent on the ability of the fungal
species colonized in the substrate to produce oxidative ligninolytic
enzymes such as laccase, VP, MnP, and LiP. These enzymes are
responsible for the degradation of lignin as well as of pollutants
(Buswell et al. 1993; Rodríguez et al. 2004). Therefore, the lig-
ninolytic enzyme activities of P. ostreatus fungal substrate and SMW
were determined. Among the ligninolytic enzymes, the MnP
activity showed high activities for both the fungal substrate and
SMW (Purnomo et al. 2010c). The degradation of some pollutants
by WRF has been well correlated with the ligninolytic activity of the
fungi (Bumpus and Aust 1987). LiP and MnP had an influence on the
efficiency of DDT degradation (Chung et al. 2009). It is also possible
that VP may be involved in DDT degradation. It has been reported
that VP can oxidize phenolic and non-phenolic aromatic
compounds as well as Mn2þ to Mn3þ and that it act as a diffusible
oxidizing agent (Rodríguez et al. 2004). However, since the lig-
ninolytic enzyme activities in the SMW were significantly different
than in the fungal substrate, but the mineralization rate was not
substantially different, the results indicate that the higher miner-
alization rate of DDT by P. ostreatus SMW was not caused by the
higher ligninolytic enzyme activities. However, the high MnP
activity in both fungal substrate and SMW suggests that MnP might
be involved in DDT degradation even though it is not the main
mechanism (Purnomo et al. 2010c). Further investigation into DDT
degradation by directly ligninolytic enzymes is needed.
It is also possible that intracellular enzymes may be involved in
DDT degradation. It has been reported that cytochrome P450
monooxygenase (P450) is involved in the degradation of some
pollutants. DDT is metabolized under aerobic conditions by the
P450 enzyme to DDD, dicofol, FW-152, 2,2-bis(4-chlorophenyl)
acetic acid (DDA) and DBH (Purnomo et al. 2010c; Suhara et al. in
press). It can be assumed that the intracellular P450 also has an
important role in the metabolism of persistent aromatic
compounds, alongside ligninolytic enzymes.
5. Application on contaminated soil
Soils are a complex environment containing mixed populations
of microorganisms with synergistic and antagonistic activities. Soil
is not an inert matrix, and its properties and the prevailing envi-
ronmental conditions will influence the behavior of both microor-
ganisms and DDT (Deepthi et al. 2007; Mwangi et al. 2010).
Therefore, it is important to examine DDT degradation in soil, both
in the laboratory under controlled conditions and in the field
(Guenzi and Beard 1967; Xu et al. 1994; Boul 1996). Firstly, the DDT
degradation abilities of BRF, CMC and SMW of P. ostreatus were
investigated in artificially contaminated soils in the laboratory
under controlled conditions. The best source was selected and
applied on historically contaminated soil.
5.1. Application of brown-rot fungi in artificially
DDT-contaminated soil
G. trabeum, F. pinicola and D. dickinsii all displayed a good ability
to degrade DDT in pure culture (Purnomo et al. 2008, 2010a). In this
section, the ability of these fungi to degrade DDT in soil was
investigated to find out whether these fungi are suitable for
bioremediation purposes. DDT was degraded by G. trabeum,
F. pinicola and D. dickinsii in both sterilized (SL) and un-sterilized
(USL) soils (Table 3). Considering the DDT degradation rates in
both SL and USL soils, G. trabeum has the best ability to degrade DDT
in artificially contaminated soil (Purnomo et al. 2011). By
comparing the DDT degradation rates in SL and USL soils, the
degradation of DDT by G. trabeum in USL soil was higher than in SL
soil (Table 3), indicating that soil microorganisms enhanced the
ability of G. trabeum to degrade DDT. However, the degradation of
DDT by F. pinicola in USL soil was lower than in SL soil (Table 3),
indicating that soil microorganisms may inhibit the ability of
F. pinicola to degrade DDT. Finally, the degradation of DDT by
D. dickinsii in USL and SL soils did not show any significant differ-
ence, which indicates that soil microorganisms did not inhibit or
enhance the ability of this fungus to degrade DDT (Table 3,
Purnomo et al. 2011). In the USL soil, DDD was detected as
a metabolic product. Because DDD was not detected in the SL soil,
the production of DDD might be related to the involvement of soil
microorganisms. Some soil bacteria have been reported to trans-
form DDT to DDD via reductive dechlorination (Katayama et al.
1993; Megharaj et al. 2000).
The use of Fenton reaction system for the treatment of
contaminated soils was initially investigated by Watts et al. (1990),
who was the first to observe pentachlorophenol mineralization.
Subsequently, several other works appeared which emphasized the
efficiency of Fenton processes for the remediation of soils
contaminated with other organic pollutants (Watts et al. 1994; Villa
and Nogueira 2006; Villa et al. 2008). As the Fenton reaction
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A.S. Purnomo et al. / International Biodeterioration & Biodegradation 65 (2011) 921e930
Table 3
DDT degradation rates in artificially contaminated sterilized (SL) and un-sterilized (USL) soils by brown-rot fungi during a 14 d incubation period (Purnomo et al. 2011).
Brown-rot fungi DDT decreased (%)a
Sterilized soil
2þ
Without addition Fe
G. trabeum 36.2  1.8aA
F. pinicola 8.8  1.0aB
D. dickinsii 11.5  1.0aC
Initial concentration of DDT is 0.25 mmol. The data were determined by HPLC. Data are presented as mean  standard deviations (n ¼ 3). Data followed by the same lower case
letter on each row or by the same capital letter in each column are not significantly different (P < 0.05). DDT (1,1,1-trichloro-2,2-bis (4-chlorophenyl) ethane).
a
vs. Control.
enhanced DDT degradation by BRF in pure cultures (Purnomo et al.
2008, 2010a), the effect of the addition of Fe2þ as a source for the
Fenton reaction in DDT contaminated soil was also investigated.
Although the soil system may already contain iron, the concen-
tration of iron may not be high enough to assist in degradation of
DDT because of its use in other reactions, especially for F. pinicola
and D. dickinsii. The Fe concentrations in soil found differed in
a range from few mM Fe to about 200 mM Fe (Ammari and Mengel
2006). The supplement of Fe2þ (10 mM) was expected to enrich
the soil and also enhance DDT degradation. In comparison to the
DDT degradation without the addition of Fe2þ, G. trabeum degraded
DDT better with the addition of Fe2þ than without it both SL and
USL soils. Similarly, D. dickinsii and F. pinicola also degraded DDT
better with the addition of Fe2þ than without it (Table 3, Purnomo
et al. 2011). In the USL soil, DDD was produced by G. trabeum,
F. pinicola and D. dickinsii. The addition of Fe2þ to F. pinicola and
D. dickinsii enhanced DDT degradation significantly, causing an
increase in DDD production (Purnomo et al. 2010a, 2011).
5.2. Degradation of artificially DDT-contaminated soil by cattle
manure compost and its isolated fungi
The greatest amount of degradation occurred by mesophilic
stage material in both SL and USL soils (Purnomo et al. 2010b).
Higher degradation in the USL soil was caused by the activities of
soil microorganisms. Since active soil microbes alone cannot
degrade DDT, the synergistic actions between compost and soil
microorganisms might enhance DDT degradation (Purnomo et al.
2010b). Three isolated fungi from each stage of the composting
process exhibited an ability to degrade DDT at 30  C:
M. circinelloides 1CMC6, G. geotrichum 2CMC1, and M. circinelloides
3CMC1. These fungi were selected to examine degradation of DDT
in artificially contaminated soil. The fungi degraded DDT
(0.25 mmol) by approximately 50% and 85% in SL and USL soils,
respectively (Purnomo et al. 2010b). These results indicate that soil
microorganisms enhanced the abilities of these fungi to degrade
DDT.
In the USL soil, DDD and DBP were detected as metabolic
products. It seemed that DDD, which is produced by soil microor-
ganisms, was transformed into DBP by the fungi (Purnomo et al.
2010b). This would suggest that the fungi prefer to degrade DDD
over DDT. Approximately 27% and 5% of DDD and DBP were also
detected as metabolic products in the SL soil, respectively. The
isolated fungi and soil microorganisms possess a comparable ability
to transform DDT into DDD. However, since the amount of DDD in
the USL soil was higher than in the SL soil, it appears that soil
microorganisms produce DDD to a greater extent than the fungi,
with the transformation of DDT to DDD by soil microorganisms
dominating (Kantachote et al. 2003, 2004; Lourencetti et al. 2007;
Purnomo et al. 2010b). Based on these results, isolated fungi
demonstrated a high ability to degrade DDT in both SL and USL
927
Un-sterilized soil
2þ
With addition Fe Without addition Fe2þ With addition Fe2þ
40.8  0.9bA 41.6  1.0aA 42.8  0.8aA
8.7  0.2aB 4.3  1.1aB 28.7  3.7bB
15.1  0.4bC 11.0  0.4aC 32.3  0.7bC
soils, showing that isolated fungi can be used for the bioremedia-
tion of DDT contamination in soil. This supports the evidence that
CMC contains fungi which have the ability to degrade DDT in soil.
5.3. Degradation of artificially DDT-contaminated soil by spent
mushroom waste of Pleurotus ostreatus
P. ostreatus fungal humus showed the ability to degrade DDT
(0.25 mmol) in the soil with an efficiency of 50e70% after 30
d (Purnomo et al. 2010c). Furthermore, DDT was degraded by about
40% and 80% by P. ostreatus SMW in SL and USL soils, respectively.
DDD was detected as the major metabolic product in the USL soil.
Since DDD was not detected in the SL soil, the production of DDD
might be caused by the involvement of soil microorganisms
(Purnomo et al. 2010c). The mineralization of [U-14C]DDT in arti-
ficially contaminated soil was also investigated. During the 56
d incubation, [U-14C] DDT (378 kdpm) was mineralized by 5.1% and
8.0% by P. ostreatus SMW in SL and USL soils, respectively. Several
studies have found that mineralization of DDT in soils is typically
very low (<3% after a 42 d incubation (Nair et al. 1992; Boul 1996)
or longer (Guenzi and Beard 1968; Zayed et al. 1994)). Mineraliza-
tion activity in the control was 1.1%, which indicated mineralization
by soil microorganisms (Purnomo et al. 2010c). This result provides
further evidence for the potential role of soil microbes to enhance
DDT degradation (Deepthi et al. 2007).
The distribution of [U-14C] intermediate metabolic products
was investigated. From both SL and USL soils; DDD, DDMU and
DDMS (1-chloro-2,2-bis (4-chlorophenyl) ethane) were detected
in the acetone and n-hexane extracts. Additionally, DDA (bis (4-
chlorophenyl) acetic acid) was detected as a metabolic product
in the water extract. From the CO2 formation results, the lower
level of intermediate compounds in the water-fraction of the USL
soil might be due to the increased formation of CO2. This suggests
Mineralization rate (%)
100 10
Degradation rate (%)
80 8
60 6
40 4
20 2
0 0
G. trabeum CMC P. ostreatus
SMW
Fig. 3. Degradation and mineralization of DDT in contaminated soil by G. trabeum,
CMC and P. ostreatus SMW during 28 d incubation period. Bars represent degradation
rate and triangles represent mineralization rate. Data points represent means and
standard deviations (n ¼ 3).
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928 A.S. Purnomo et al. / International Biodeterioration & Biodegradation 65 (2011) 921e930
Fig. 4. Degradation of DDT in historically contaminated soil by spent mushroom waste
of P. ostreatus during 28 d incubation period. Data points represent means and stan-
dard deviations (n ¼ 3). The same minor letter on each line indicates no statistical
difference (P < 0.05).
that soil microorganisms enhanced the transformation of water-
soluble intermediate compounds to CO2 (Purnomo et al. 2010c).
However, the possibility of synergistic interactions between
P. ostreatus SMW and soil microorganisms needs further detailed
investigation.
5.4. Degradation of DDT in historically contaminated soil by spent
mushroom waste of Pleurotus ostreatus
Even though BRF and CMC have a high ability to degrade DDT in
artificially contaminated soil, these sources lack mineralization
ability (Fig. 3). Since SMW of P. ostreatus has a good ability to
degrade and mineralize DDT in artificially contaminated soil
(Fig. 3), this source was applied to historically contaminated soil
which has higher concentrations of DDT. In historically contami-
nated soil, the DDT concentration (0.8 mmol) is more than three
times higher than the DDT concentration in the artificially
contaminated soil (0.25 mmol). Compared with the control, about
70% of DDT was degraded by SMW of P. ostreatus in historically
contaminated soil (Fig. 4). This means that about 30% of the DDT
remained in the soil, at a concentration of 42.5 ppm. This study
gives evidence that SMW of P. ostreatus is a potential natural source
which can be used for the bioremediation of DDT in “real world”
application.
Further detail investigation on degradation pathway and
mechanism is needed to achieve complete mineralization of DDT.
Since mineralization rate was still low, developing method is
needed to obtain complete mineralization of DDT. Therefore,
some methods are needed to stimulate Hiratake for perfect
mineralization of DDT. The use of mixed cultures of some WRF
could be advantageous to gain perfect mineralization rate. Since
WRF have ligninolytic enzymes system to degrade DDT, mixed
cultures can be used to enhance mineralization ability. Besides,
the use mixed cultures of WRF along with bacteria could be
advantageous to enhance DDT mineralization. Involvement of
biosurfactant, and effect of co-metabolic compounds will be also
of interest. Since ring-cleavage bacteria have good ability to
cleave DDT, further degradation to CO2 should proceed easily by
associated with WRF.
6. Conclusions
Recent work used a biological approach for an environmentally-
friendly treatment using biological sources to degrade DDT and
also applied it for bioremediation in contaminated soil. Some BRF
were screened for their ability to degrade DDT with G. trabeum,
F. pinicola and D. dickinsii proposed as new candidates for DDT
bioremediation. In application on artificially contaminated soil,
G. trabeum showed the highest ability to degrade DDT in both SL
and USL soils. The ability of CMC to degrade DDT was also inves-
tigated, with DDT degraded during composting and DDD detected
as a metabolic product. Some fungi were isolated and identified
from CMC, and most of them were closely related to
M. circinelloides and G. geotrichum. These fungi demonstrated
a high ability to degrade DDT at both 30 and 60  C in pure culture
and artificially contaminated soil. Furthermore, SMW of P. ostreatus
has the ability to degrade and mineralize DDT in pure culture and
artificially contaminated soil. Since SMW of P. ostreatus has a high
ability to degrade and mineralize DDT in contaminated soil
compared with other sources, SMW from P. ostreatus is proposed
as the best bioremediation source in DDT-contaminated environ-
ments. Mineralization is important in bioremediation, indicating
that the process of degradation of the pollutant into a non toxic
compound (CO2) is complete. After application to historically
contaminated soil, about 70% of the initial DDT concentration in
the soil was degraded by SMW of P. ostreatus, indicating that this
source is very suitable for bioremediation of DDT in soil. Further
research in situ and ex situ are needed to promote DDT bioreme-
diation as a viable treatment option.
Acknowledgments
This work was supported by a grant from the Research Project
for Ensuring Food Safety from Farm to Table, Ministry of Agricul-
ture, Forestry and Fisheries, Japan (PO-3216).
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