Science of the Total Environment 502 (2015) 426–433

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Science of the Total Environment

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

Anaerobic degradation of Polychlorinated Biphenyls (PCBs) and

Polychlorinated Biphenyls Ethers (PBDEs), and microbial community
dynamics of electronic waste-contaminated soil
Mengke Song a,e, Chunling Luo a,⁎, Fangbai Li b, Longfei Jiang a,c, Yan Wang a, Dayi Zhang d, Gan Zhang a
a
Guangzhou Institute of Geochemistry, Chinese Academy of Sciences, Guangzhou 510640, China
b
Guangdong Institute of Eco-environmental and Soil Sciences, Guangzhou 510650, China
c
College of Life Sciences, Nanjing Agricultural University, Nanjing 210095, China
d
Lancaster Environment Centre, Lancaster University, Lancaster LA1 4YQ, UK
e
Graduate University of Chinese Academy of Sciences, Beijing 100039, China

H I G H L I G H T S

• The biodegradation PCBs and PBDEs in e-waste contaminated soils was studied.
• DIRB and arylhalorespiring bacteria were responsive to dehalogenation respiration.
• Soil bacteria and Fe ion cycling play synergistic roles in dehalogenation.

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

Article history: Environmental contamination caused by electronic waste (e-waste) recycling is attracting increasing attention
Received 9 June 2014 worldwide because of the threats posed to ecosystems and human safety. In the present study, we investigated
Received in revised form 12 September 2014 the feasibility of in situ bioremediation of e-waste-contaminated soils. We found that, in the presence of lactate
Accepted 15 September 2014
as an electron donor, higher halogenated congeners were converted to lower congeners via anaerobic
Available online 29 September 2014
halorespiration using ferrous ions in contaminated soil. The 16S rRNA gene sequences of terminal restriction frag-
Editor: Eddy Y. Zeng ments indicated that the three dominant strains were closely related to known dissimilatory iron-reducing bac-
teria (DIRB) and those able to perform dehalogenation upon respiration. The functional species performed the
Keywords: activities of ferrous oxidation to ferric ions and further ferrous reduction for dehalogenation. The present study
E-waste links iron cycling to degradation of halogenated materials in natural e-waste-contaminated soil, and highlights
PCBs the synergistic roles of soil bacteria and ferrous/ferric ion cycling in the dehalogenation of polychlorinated biphe-
PBDEs nyls (PCBs) and polybrominated biphenyl ethers (PBDEs).
Biodegradation © 2014 Elsevier B.V. All rights reserved.
Microbial community

1. Introduction
In the past few decades, uncontrolled electronic waste (e-waste)
recycling has become common in China. The primitive technologies
used, including acid leaching and burning of wire in the open,
have released many organic pollutants into the environment, as
polychlorinated biphenyls (PCBs) and polybrominated biphenyl ethers
(PBDEs). Extremely high levels of PCBs and PBDEs are found in soil,
water, air, sediment, and vegetation on/around e-waste recycling sites
(Wong et al., 2007). PCBs and PBDEs are persistent, bioaccumulative
and toxic, disrupting endocrine systems and posing serious threats to
local ecosystems and human health (Egloff et al., 2011; Frye et al.,
⁎ Corresponding author. Tel.: +86 20 85290290; fax: +86 20 85290706.
E-mail address: clluo@gig.ac.cn (C. Luo).
2012; Hamlin and Guillette, 2011; Purser, 2001; She et al., 2013;
Wang et al., 2010; Zhou et al., 2001).
Bioremediation is an efficient method to remove organic pollutants
from soils. Aerobic and anaerobic biodegradation of such compounds
have been investigated extensively (Bedard, 2003; Chang et al., 2013;
Chen et al., 2010; Cutter et al., 2001; Dabrowska and Rosinska, 2012;
Deng et al., 2011; Fennell et al., 2004; Li et al., 2005; Payne et al., 2011,
2013; Rayne et al., 2003; Shih et al., 2012). Aerobic microbial degrada-
tion is often possible only when halogenated compounds have a maxi-
mum of four-to-five halogen atoms, although an aerobic bacterium
responsible for deca-BDE debromination was isolated from sediment
(Deng et al., 2011). Generally, the biodegradability of PCBs and PBDEs
congeners decreased as the number of halogen atoms rose (Furukawa,
2000). More evidence was identified in PCB-contaminated soils where
lower chlorinated PCB congeners were the prior substrates for

http://dx.doi.org/10.1016/j.scitotenv.2014.09.045
0048-9697/© 2014 Elsevier B.V. All rights reserved.
 M. Song et al. / Science of the Total Environment 502 (2015) 426–433
indigenous microorganisms (Bedard et al., 1986), and the amount of
PCBs degrading microbes decreased progressively with the increasing
number of chlorine atoms (Abramowicz, 1990; Bedard et al., 1986).
Anoxic dehalogenation by microorganisms plays an important role
in elimination of environmental PCBs and PBDEs. Dehalogenation
of these compounds has been observed in soil, sewage sludge, and
estuarine and marine sediment, under different redox conditions.
Higher halogenated organic compounds are usually first reductively
dehalogenated to less halogenated molecules under anaerobic condi-
tions in the presence of organic compounds. During dehalogenation
process, the common electron donors include lactate, acetate and glu-
cose (Rhee et al., 2003; Boyle et al., 1999). The metabolites are further
degraded to non-toxic compounds by aerobic microorganisms
(Bedard, 2003; Bedard et al., 2005; Boyle et al., 1999; Bzdusek et al.,
2006a,b; Li et al., 2005, 2012; Magar et al., 2005a,b; Pakdeesusuk et al.,
2005; Payne et al., 2013; Sanford et al., 1996; Yan et al., 2006).
Dehalorespiration is critical for effective dehalogenation, and the effect
of dissimilatory iron reduction on dehalogenation has been discussed
(Li et al., 2008; Wu et al., 2010). During dehalorespiration, halogenated
compounds act as electron acceptors, resulting in accumulation of
lower halogenated congeners and halogen-free compounds (Fetzner,
1998; Holliger et al., 1998; Wohlfarth and Diekert, 1997). A sulfidogenic
2-bromophenol-degrading consortium enriched from estuarine sedi-
ment could use 2-bromophenol as the sole electron acceptor when lac-
tate and acetate were present as energy sources (Rhee et al., 2003).
Another anaerobic bacterium isolated from estuarine sediment used
lactate as electron donor and 2,4,6-tribromophenol as electron acceptor,
indicating that 2,4,6-tribromophenol was converted to phenol via
dehalorespiration (Boyle et al., 1999).
Under anaerobic conditions, the role of dissimilatory iron reducing
bacteria (DIRB) on dehalogenation has been well recognized and under-
stood (Li et al., 2008; Wu et al., 2010). In terms of dissimilatory iron re-
duction, halogenated compounds are bioreduced by DIRB via an
electron-shuttling mechanism in the presence of Fe(III) oxides (Feng
et al., 2013; McCormick et al., 2002; Tobler et al., 2007; Wu et al.,
2010). Here, Fe(III) species serve as electron shuttles, accepting elec-
trons created during biotic oxidation of organic matter by DIRB and do-
nating these to target contaminants (Feng et al., 2013). Carbon
tetrachloride reduction is associated with the biogenic iron species pro-
duced by Geobacter sulfurreducens (DIRB) and the electron shuttle is ac-
tive under iron-reducing conditions (Maithreepala and Doong, 2009). In
addition, some DIRB, including Geobacter and Shewanella, utilize haloge-
nated compounds as direct terminal elector acceptors (Feng et al.,
2013). Recently, Comamonas koreensis CY01 is reported as DIRB to re-
duce hydrated Fe(III) oxides and 2,4-dichlorophenoxyacetic acid, ex-
tending the diversity of iron-reducing bacteria associated with
dechlorination (Wu et al., 2010).
To date, though many studies have invested the dehalogenation pro-
cess in the field, little is known about their anaerobic bioremediation
process at e-waste sites, leaving the research gaps of the natural
dehalogenation in soils. The objectives of the present study were to in-
vestigate the degradation of native PCBs and PBDEs in e-waste contam-
inated soils, and to explore the dynamics of the microbial community
responsible for degradation process under anaerobic conditions with
lactate as the exotic elector donor and energy source. The work yielded
useful information on mechanisms of PCBs and PBDEs dehalogenation
in anaerobic bioreactors or biopilings and further bioremediation en-
hancement possibility.
2. Materials and methods
2.1. Soil collection and characterization
Soils were collected from the e-waste recycling town of Qingyuan,
Guangdong province, South China (23.57° N, 113.0° E). Topsoil samples
(from 0 to 15 cm depth) from an e-waste burning site were collected,
427
placed in polythene zip-bags, and transported immediately to the labo-
ratory. Soils were homogenized, sieved through a screen of 2 mm pore
size, and stored at 4 °C prior to study. The soil properties are given in
Tables S1–S3.
2.2. Experimental set-up
Both sterile controls and non-sterile samples were tested in this
study. All samples were prepared in triplicates, and three vials were an-
alyzed at each time point. In detail: (1) 15 g amounts of soil (dry
weight) were placed in sterile 100 mL serum bottles; (2) 50 mL of
30 mM piperazine-N-N′-bis-2-ethanesulfonic acid (PIPES) buffer
(pH 7.0) with lactate (10 mM) were added to each bottle; and
(3) each bottle was sealed with a rubber stopper and an aluminum
seal, and incubated at 25 °C in the dark. All experiments were conducted
in an anaerobic glove box under 99.999% N2 at a flow rate of 80 L min−1.
All stock solutions were filtered through 0.2-μm pore-size filters and
stored in dark-brown containers. Stock solutions were deoxygenated
under 99.999% nitrogen for 2 h prior to use in the anaerobic box. To
prepare sterile controls, soils were γ-irradiated (50 kGy) for 2 h before
use. Of the three carbon sources (acetate, lactate and glucose), lactate
had the best performance for PCBs/PBDEs removal and selected as the
electron donor in all the other treatments (see Fig. S1).
2.3. Sampling
Samples were taken on 0, 24, 40, 60, and 90 d, subjected to chemical
analysis (chloride ions, bromide ions, HCl-extractable Fe(II), HCl-
extractable Fe(III), PCBs, and PBDEs) and DNA extraction. At each time
point, the soil samples were taken from the incubator and kept in the
anaerobic box. Three biological replicates were conducted for both ster-
ile control and PCBs/PBDEs treatments. After vigorous mixing, 5 mL of
soil suspension was directly taken for HCl-extractable Fe(II) and Fe(III)
analysis. The rest of the samples passed through 0.2 μm pore-size mem-
branes prior to the analysis of chloride and bromide ions. The soil resi-
dues were at −20 °C for further chemical and biological analyses.
2.4. Chemical analysis
2.4.1. Chloride and bromide ions
Chloride and bromide ions were measured via ion chromatography
(ICS-90, DIONEX) coupled with an RFICTM Ion Pac®AG14A-7 μm
Guard Column (50 mm length and 4 mm i.d), an AMMS III
micromembrane suppressor, an IonPac®AS14A-7 μm Analytical Col-
umn (250 mm; 4 mm i.d), and a DS5 Detection Stabilizer conductivity
detector. The eluent (8.0 mM Na2CO3 and 1.0 mM NaHCO3) was
pumped at 1.0 mL min−1. Sulfuric acid (0.05 M) was used to regenerate
suppression of eluent conductivity. Chromeleon software was used for
data analysis.
2.4.2. HCl-extractable Fe(II) and Fe(III) species
All HCl-extractable Fe(II) species were extracted into 0.4 M HCl at
25 ± 1 °C in the dark for 1.5 h, filtered through a 0.2 μm pore-size mem-
brane, and measured using the 1,10-phenanthroline method. To mea-
sure HCl-extractable iron levels, 1 mL of solution was extracted into
0.4 M HCl at 25 ± 1 °C in the dark for 1.5 h, filtered through a 0.2 μm
pore-size membrane, and placed in a centrifuge tube containing 1 mL
hydroxylamine hydrochloride (10%, w/v), 4 mL sodium acetate
(10%, w/v), and 4 mL 1,10-phenanthroline (0.1%, w/v); iron levels
were next determined using the 1,10-phenanthroline method (Heron
and Christensen, 1995). The HCl-extractable Fe(III) content was taken
to be the difference between the HCl-extractable total iron and Fe(II)
levels.
428 M. Song et al. / Science of the Total Environment 502 (2015) 426–433

2.4.3. PCBs and PBDEs (5′-GGTTACCTTGTTACGACTT) (Operon Biotechnologies) using the fol-
Freeze-dried soil samples were homogenized, pulverized, spiked lowing polymerase chain reaction (PCR) program: 94 °C for 10 min
with surrogate standards (TCmX, PCB30), and extracted into dichloro- for initial melting; 30 amplification cycles of 94 °C for 30 s, 54 °C for
methane (DCM) in a Soxhlet apparatus, for 48 h, with addition of acti- 30 s; and 72° for 1.5 min; and a final extension at 72 °C for 10 min.
vated copper to remove sulfur. The extract was concentrated to 0.5 mL After amplification, PCR products (150 ng) were purified using an
after solvent exchange to hexane. The soil extracts were purified on a Omega ENZA Cycle-Pure kit following the manufacturer's instructions,
multilayer silica gel/alumina column filled with anhydrous Na2SO4; and digested with HaeIII for 4–5 h at 37 °C. One nanogram of each la-
50% (w/w) sulfuric acid silica-gel; neutral silica gel (3% w/w; beled PCR product was analyzed on an ABI 3730 Genetic Analyzer run-
deactivated); and neutral alumina (3% w/w; deactivated) (from top to ning the Peak Scanner software version 1.0; the ROX500 set of internal
bottom), via elution with 16 mL hexane/DCM (1:1, v/v). After concen- standards was used. The percentage abundance of each fragment was
tration to 50 μL under a gentle stream of N2, a known amount of 13C la- determined.
beled PCB-141 was added prior to analysis.
A total of 17 PCB congeners (PCB-8, - 28, - 49, - 60, - 66, - 77, - 82, 2.4.4.2. Sequencing. For 16S rRNA gene sequencing, DNA was amplified
-101, -156, -166, -170, -179, -180, -183, -189, -198, and -209) was de- as above except that the forward primer was unlabeled (27 F 5′-AGAG
tected via GC–EI-MS using a 50-m capillary column (Varian, CP-Sil 8 CB, TTTGATCMTGGCTCAG). The PCR products were purified using an
0.25 mm i.d., 0.25 μm film thickness). The initial oven temperature was Omega ENZA Cycle-Pure kit. The recovered fragments were cloned
set to 150 °C for 3 min, raised to 290 °C at 4 °C min−1, and held at that into Escherichia coli JM 109 using a TA cloning kit. E. coli clones were
temperature for 10 min. The temperatures of the MSD source and the grown on Luria-Bertani medium solidified with 15 g agar L− 1 and
quadrupole were 230 °C and 150 °C, respectively. 50 μg mL−1 ampicillin for 16 h at 37 °C. The plasmids of positive clones
Eight PBDE congeners (BDE-28, -47, -99, -100, -153, -154, -183, and were extracted with EZNA plasmid mini-kit and the insertion was con-
- 209) were detected via GC–NCI-MS (an Agilent GC7890 column firmed by 0.8% agarose gel electrophoresis. The right clone was directly
coupled with a 5975C MSD). A DB5-MS (30 m, 0.25 mm i.d., 0.25 μm sequenced on an ABI 3730 genetic analyzer using M13 universal
film thickness) capillary column was used to analyze the first seven con- primers (Luo et al., 2009). Sequence similarity searches and alignments
geners; BDE 209 was analyzed separately on a CP-Sil 13 CB column were performed with the aid of BLAST and Molecular Evolutionary Ge-
(15 m, 0.25 mm i.d., 0.2 μm film thickness). The oven was set to netics Analysis, version 5. The sequences of all TRFs were deposited in
130 °C for 1 min, ramped at 12 °C min−1 to 155 °C, at 4 °C min−1 to GenBank under accession numbers KF535193–KF535198.
215 °C, and at 3 °C min−1 to 300 °C; this temperature was held for
10 min. 2.5. Statistical analysis
Mixed standards (17 PCB and 8 PBDE congeners) were used to quan-
tify PCBs and PBDEs. Instrumental performance was subjected to quality Statistical analysis was performed using the SPSS package (version
control calibration by the standards, after each set of eight samples had 11.0). Values are the means of data from three independent replicates.
been analyzed. Six PCB and PBDE concentrations were used to derive Analysis of variance (ANOVA) followed by Duncan's test was per-
calibration curves. Concentrations in samples were corrected by refer- formed. A p value b0.05 was considered to indicate statistical signifi-
ence to surrogate recovery levels. cance. The equality and normality of data were tested by Brown–
Forsythe and Shapiro–Wilk test respectively, and the null hypothesis
2.4.4. Biotic analysis was rejected for p value less than 0.05.
The content variation of individual PCBs and PBDEs on 90 d followed
2.4.4.1. T-RFLP of 16S rRNA genes. After freeze-drying the slurry soil sam- Eq. (1):
ples, total genomic DNA of the microcosm was extracted using a MoBio
Powersoil DNA isolation kit according to the manufacturer's instruction. C 90 −C 90s
Content variation ð%Þ ¼  100% ð1Þ
DNA concentrations were determined using an ND-1000 UV–vis spec- C0
trophotometer. Total DNA was subjected to terminal restriction frag-
ment (TRF) length polymorphism analysis using standard procedures where C90 represents the concentration of individual molecule in sam-
(Liu et al., 1997). DNAs were amplified with 27 F-FAM (5′-AGAGTTTG ples after 90 d treatment, C90s refers to that in sterile treatment, and
ATCMTGGCTCAG; 5′ end-labeled with carboxyfluorescein) and 1492R C0 is the concentration of individual PCBs/PBDEs molecule on day 0.

200 sterile90d 0d 24d 40d 60d 90d

350
180
content variation (%)

300
250
160 200
150
100
Concentration (ng g-1)

140 50
0
120 -50
-100
8

00

54

53

83

09
E2

E4

E9
E1

E1

E1

E1

E2

100
BD

BD

BD
BD

BD

BD

BD

BD

80

60

40

20

0
BDE28 BDE47 BDE100 BDE99 BDE154 BDE153 BDE183 BDE209

Fig. 1. The change of PBDEs against time. The subfigure shows the content variation of individual compounds on 90 d in samples because of the microbial activity. The stars show significant
changes with time with p b 0.05. Error bars denote the standard deviation of the mean of triplicate samples.
 M. Song et al. / Science of the Total Environment 502 (2015) 426–433 429

400
sterile90d 0d 24d 40d 60d 90d
200
350

content variation (%)

150

100
300
50

Concentration (ng g-1)

0
250
-50

-100
200

150

100

50

Fig. 2. The change of PCBs against time. The subfigure shows the content variation of individual compounds on 90 d in samples because of the microbial activity. The stars show significant
changes with time with p b 0.05. Error bars denote the standard deviation of the mean of triplicate samples.

3. Results and discussion
3.1. Anaerobic dehalogenation of native PCBs and PBDEs in
e-waste-contaminated soil
The changes of PBDEs concentration against time were illustrated in
Fig. 1. No obvious change was observed in sterile controls (p N 0.05,
ANOVA test), and then only the 90 d PBDEs concentrations in sterile
control are presented (see Fig. S3). The concentrations of BDE-209 and
BDE-47 decreased gradually, and 39.7% and 29.5% was removed respec-
tively by 90 d. The concentrations of BDE-183, BDE-158, BDE-154 and
BDE-99 had limited increase over time. The levels of less-brominated
congeners, including BDE-28 and BDE-100, were steady throughout
the experiment. From previous evidence on the BDE-209 degradation
attribution to the production of less-brominated congeners (He et al.,
2006; Kim et al., 2012; Lee and He, 2010; Tokarz et al., 2008), the loss
of deca-BDE in this study might be associated with the rise in hepta-,
hexa-, and penta-chlorobiphenyls under anaerobic conditions. For ex-
ample, hepta-BDEs and octa-BDEs were produced by debromination
of deca-BDE, and BDE-154, BDE-99, and BDE-49 were among the
debromination products (He et al., 2006). Increases in the levels of
nona-, octa-, hepta- and hexa-PBDEs were found upon debromination
of BDE-209 in sediment (Tokarz et al., 2008). In this study, BDE-47
was degraded, whereas no significant degradation was observed for
the other measured congeners including BDE-158, BDE-154, BDE-99,
BDE-28 and BDE-100. The results were similar to what was found in
Nan-Kan River sediment, but different from the Er-Jen River work
(Yen et al., 2009). The main products of BDE-47 degradation were iden-
tified as BDE-17 or BDE-28, dependent on the functional bacteria
No obvious change was observed in the levels of lower chlorinated con-
geners including PCB-66 (p = 0.991), PCB-60 (p = 0.986), PCB-28
(p = 0.972), and PCB-8 (p = 0.893). The control PCB profiles did not
change during the entire experiment, and thus only the PCB concentra-
tions at 90 d were shown. Supported by the previous study that tetra-,
penta-, hexa- and hepta-chlorobiphenyls were produced during the
dechlorinated process of deca- and hepta-chlorobiphenyls (Alder
et al., 1993; Dabrowska and Rosinska, 2012; Payne et al., 2011), the
rise of tetra-, penta-, hexa-, and hepta-chlorobiphenyls in this study
may attribute to the loss of deca- and hepta-chlorobiphenyls. It was
also found that approximately 65% of the meta and para chlorines
were removed in 2 months in the sediment cultures (Alder et al.,
1993). Dabrowska and Rosinska (2012) found that the levels of higher
chlorinated congeners fell, and those of lower congeners rose, under
anaerobic conditions. In a study on reductive dechlorination of commer-
cial PCBs upon bioaugmentation with a dehalorespiring bacterium,
higher chlorinated PCB levels in mesocosms decreased by about 56%
(by mass) after 120 d of incubation (Payne et al., 2011).
Both PCBs and PBDEs exhibited the same trend that higher haloge-
nated compounds were reduced in amount; the levels of lower haloge-
nated compounds were stable; and compounds with four to seven
chlorides or bromides accumulated over 90 d, with the exception
of BDE-47. The results might be explained by the fact that the
dehalogenation often occurs among the compounds with high number
700 ferrous ion
ferric ion
600
sterile ferrous ion
Concentration (ng g-1)

(Robrock et al., 2008). The debromination capacity and efficiency 500 sterile ferric ion
depended on both the number of bromide atoms and the congener
structure/substitution. Bromides at the meta and para positions were 400
prior to be removed than those at the ortho position via bacterial func-
300
tions (Alder et al., 1993; He et al., 2006). It was often observed that
ortho-bromine substituted congeners such as BDE-183, BDE-154, BDE- 200
153 and BDE-99 were generated from the octa-BDE mixture (Robrock
et al., 2008) and BDE-209 (He et al., 2006; Tokarz et al., 2008), which 100
was consistent with the increase of BDE-183, BDE-154, BDE-153 and
0
BDE-99 in this study. 0 20 40 60 80 100
The biodegradation process of PCBs against time was shown in Fig. 2. Time (day)
On day 90, the levels of highly chlorinated congeners including PCB-
209, PCB-189, PCB-183, and PCB-179 were reduced, whereas the Fig. 3. The concentrations of Fe2+ and Fe3+ at different time point. Error bars denote the
concentrations of PCBs with four to seven chloride atoms had risen. standard deviation of the mean of triplicate samples.
430 M. Song et al. / Science of the Total Environment 502 (2015) 426–433

0d

Abandance (%)
6
24d

40d
4
90d
2

0
75bp 182bp 191bp 198bp 201bp 220bp
Granulicella Anaeromyxobacter Geothrix fermentans Acidiferrobacter Denitratisoma Xylophilus ampelinus
sapmiensis strain dehalogenans strain strain H5(100%); thiooxydans strain m- oestradiolicum strain strain BPIC 48(100%);
S6CTX5A(85%); 2CP-1(86%); Desulfotalea arctica 1(93%); AcBE2-1(91%); Aquincola
Geothrix fermentans Anaeromyxobacter strain LSv514 (80%); Methylococcus Thiobacter subterraneus tertiaricarbonis strain
strain H5(84%); dehalogenans 2CP-C Geoalkalibacter capsulatus strain strain C55(91%); L10(96%);
Geobacter (86%); ferrihydriticus strain Z- Texas(89%); Nitrosospira multiformis Rubrivivax gelatinosus
grbiciae(83%) Anaeromyxobacter sp. 0531(80%) Methylocaldum gracile strain Nl13(90%) strain ATH 2.2.1(96%)
Fw109-5 (86%) strain VKM 14L(87%)

Fig. 4. The abundance of main T-RFLP fragments in different time. The names below the x-axis were strains with the strongest BLAST hits with the responsive bacteria measured in this
work, the number in the bracket were the similarity.

of substituted halogens (Chang et al., 2013; Payne et al., 2013; Shih et al., significantly removed from parent compounds. Further chloride ion
2012). Besides, the dehalogenation pathways are dependent on the analysis demonstrated that chloride concentration increased from
functions of various reductive dehalogenase-homologous in different 93.84 ± 1.30 mg L− 1 on 0 d to 104.40 ± 1.11 mg L−1 on 24 d and
bacterial species (Futagami et al., 2013). It has been suggested that 112.53 ± 1.57 mg L− 1 on 60 d, confirming the dehalogenation
dehalogenation may be microbial population-specific in terms of the of PCBs. Comparing to the bromine background in the soil, the
substitution positions attacked and the numbers of halogen atoms re- debromination contributed only trace bromine ions, consequently caus-
moved, and that different microbial populations may generate different ing the stable bromine level.
dehalogenation patterns (Alder et al., 1993; Yen et al., 2009).

3.2. Generation of chloride and bromide ions 3.3. Changes in ferrous and ferric ion levels

Chloride ions increased significantly during the dechlorination pro-
cess (p b 0.05, ANOVA test), whereas bromide ions kept stable through-
out the experiment (Fig. S4). This behavior was similar to the reductive
transformation of pentachlorophenol (Li et al., 2008; Wu et al., 2010). In
sterile controls, both chloride and bromine ions did not change
(p N 0.05, ANOVA test). The results suggested that chloride ions were
As shown in Fig. 3, ferrous iron showed a cycling behavior, reaching
the peak concentration at 40 d and 90 d, whereas the trough was at 60 d.
Notably, the changes in ferric iron levels were the opposite as declining
at 40 d, rising at 60 d, and declining again at 90 d. In the sterile controls,
changes in both ferric and ferrous iron levels were similar to those in
test samples at 20 d and 40 d except that the ferrous iron level decreased

100 220bp (KF535193)

100 Aquincola tertiaricarbonis L10(NR043913.1)
82
Xylophilus ampelinus BPIC 48(NR036931.1)
100
Denitratisoma oestradiolicum AcBE2-1(NR043249.1)
100 201bp (KF535194)
198bp (KF535196)

94 Methylocaldum gracile VKM-14L(NR026063.1)

66
100 Methylococcus capsulatus Texas (NR042183.1)
79 Geobacter grbiciae TACP-5(NR041826.1)
Desulfotalea arctica LSv514 (NR024949.1)
97 182bp (KF535198)

60 Anaeromyxobacter dehalogenans 2CP-1(NR027547.1)

100 Anaeromyxobacter sp. Fw109-5(NR074968.1)
191bp (KF535195)
75bp (KF535197)
99 Geothrix fermentans H5 (NR036779.1)

0.02

Fig. 5. Phylogenetic relationships among the unclassified functional bacteria and closely related type strains. GenBank accession numbers are in parentheses. The tree was constructed
using the neighbor-joining algorithm. Bar, 2% estimated sequence divergence.
 M. Song et al. / Science of the Total Environment 502 (2015) 426–433
from 21 d to 40 d; neither level changed at the last sample times after
40 d.
Soil originally exposed to air was transferred to reductive anaerobic
conditions, facilitating redox reactions. Hence, the ferric iron level ini-
tially decreased and that of ferrous iron rose in both sterile and non-
sterile samples. In sterile control without biological activity, the slight
decrease of ferrous iron from 0 d to 40 d might attribute to the changing
redox equilibrium from aerobic (0 d before experiment) to anaerobic
condition (40 d) (Yang, 2001; Gotoh and Patrick, 1974; Brennan and
Lindsay, 1998). In test samples, changes in iron levels were also affected
by biological factors. Ferrous iron acts as an electron donor in
dehalogenation and some microorganisms can oxidize ferrous iron,
generating ferric iron (McCormick et al., 2002). Also, dehalogenation
can also be driven by dissimilatory iron reduction (Feng et al., 2013;
McCormick et al., 2002; Wu et al., 2010). On the other hand, DIRB can
transfer electrons to ferric iron when oxidizing organic material. There-
fore, both ferrous and ferric iron are generated and consumed continu-
ously in test samples if energy is available.
3.4. Dynamics of microbial composition and function
Terminal restriction fragment length polymorphism (T-RFLP) analy-
sis (Fig. 4) showed that the relative abundances of the principal TRFs
changed over time. Six prominent fragments were observed. The 16S
rRNA sequencing suggested that the two organisms yielding TRFs of
75- and 191-bp belonged to the family Geobacteraceae, and the organ-
ism yielding the 191-bp TRF was closely related to Geothrix fermentans
H5 (similarity, 100%) (Fig. 5). However, they had different relative
abundance behavior that the 191-bp TRF was dominant at 0 d and rap-
idly declined at 24 d, whereas the relative abundance of the 75-bp TRF
increased at 24 d and 40 d, subsequently declining at 90 d. The
Geobacteraceae are strictly anaerobic dissimilatory Fe (III)-reducing
bacteria which, in the presence of organic matter, form ferrous iron spe-
cies abiotically reducing certain chlorinated hydrocarbons (Feng et al.,
2013; McCormick et al., 2002; Wu et al., 2010). DIRB bacteria such as
Geobacter and Shewanella directly use chlorinated hydrocarbons as ter-
minal electron acceptors, and both Fe(III) oxides and bacterial action
promote contaminant bioreduction via electron shuttling (Lonergan
et al., 1996; Tobler et al., 2007; Wu et al., 2010).
The 16S rRNA gene sequence of the organisms yielding the 182-bp
TRF was closely related to that of the myxobacteria, which are
unique arylhalorespiring facultative anaerobic organisms. Its relative
abundance of increased throughout the whole experiment. Such
chlororespiring myxobacteria can use 2,6-dichlorophenol, 2,5-dichloro-
phenol and 2-bromophenol as terminal electron acceptors, and lactate
as an electron donor (Sanford et al., 2002).
The relative abundance of the 201-bp TRF has the cycling behavior,
and its peak and trough was at 24 d (90 d) and 40 d, respectively. The
16S rRNA sequence indicated its high similarity to Denitratisoma
oestradiolicum AcBE2-1 (denitrifying bacteria) which can grow on
fatty acids (C2 to C6) with nitrate as the electron acceptor (Fahrbach
et al., 2006).
The relative abundance of 198- and 220-bp TRF was similar, domi-
nant at 0 d and dramatically declining at 24 d. The 198-bp TRF was
closely related to methanotrophic bacteria, some of which grow under
fully aerobic conditions whereas others are facultative aerobes. The or-
ganism yielding the 220-bp TRF matched with the aerobic Xylophilus
ampelinus BPIC 48 (similarity, 100%) and Aquincola tertiaricarbonis L10,
neither of which perform dehalogenation.
The microcosm environment greatly influences microbial communi-
ty composition. When soil lacks oxygen, aerobic (and some facultative)
bacteria grow poorly, explaining the sharp falls in the relative
abundances of the 220- and 198-bp TRFs in the early experiment. An-
aerobic bacterial numbers rose over time, eventually becoming pre-
dominant. Thus, the relative abundance of organisms yielding 75- and
182-bp TRFs increased to 90 d during all the sample time points. The
431
organisms represented by the 191-bp TRF were acetate-oxidizing
Fe(III)-reducing bacteria, and variation in the level of this TRF was prob-
ably attributable to changes in the growth environment and acetate
level (Sanford et al., 2002).
Previous studies showed that aerobic bacteria play an important role
in the biodegradation of halogenated compounds with less than five
halogen atoms (Chang et al., 2013; Chen et al., 2010). Aerobic bacterium
with the ability of debrominating deca-BDE was also isolated from sed-
iment by Deng (Deng et al., 2011); while for the microbial degradation
of higher halogenated compounds, it was proved by different researches
that the compounds were dehalogenated under anaerobic conditions by
various anaerobic bacteria (Bedard, 2003; Bedard et al., 2005; He et al.,
2006; Tokarz et al., 2008; Yen et al., 2009). The dehalogenation of PCBs
and PBDEs observed in this work was attributed to the biochemical
process, which probably involved the actions of dissimilatory Fe(III)-
reducing and arylhalorespiring bacteria. The ferrous iron concentration
increased as the amount of dissimilatory Fe(III)-reducing bacteria rose
in test samples. A clear peak in ferrous ion concentration was evident
at 40 d. Furthermore, arylhalorespiring bacteria represented by the
182-bp TRF grew rapidly. These growth patterns may have caused de-
creases in the levels of highly chlorinated and brominated congeners in-
cluding PCB-209, PCB-198, and BDE-209, from 40 d to 90 d. Work with
pure cultures showed that myxobacteria can exert a dehalogenating ac-
tivity under anaerobic conditions (Sanford et al., 2002). The suggestion
that dehalogenation involved the action of ferrous ions is supported by
the decrease in ferrous ion concentration, with a concomitant increase
in ferric ion concentration, from 40 d to 60 d. Ferric ions may serve as
electron shuttles accepting electrons from biotic oxidations of organic
matters by DIRB, donating such electrons to halogenated compounds.
This is consistent with earlier data (Wu et al., 2010), which reported
that the maximum decline in 2,4-dichlorophenoxyacetic acid concen-
tration coincided with the fall in Fe(II) concentration. Next, Fe(III)-re-
ducing bacteria (with 75- and 191-bp TRFs) reduce ferric ions,
decreasing ferric ion and increasing ferrous ion concentrations between
60 d and 90 d.
3.5. Possible solutions for bioremediation implication
Our results have revealed that PCBs and PBDEs could be biodegraded
under anaerobic conditions with lactate as electron donor. Dissimilatory
Fe(III)-reducing and arylhalorespiring bacteria are important in this
context. Hence, measures could be taken to increase the activities of
such species in the bioreactor or biopiling. For example, the restricted
carbon source (such as lactate) could be dosed in the contaminated
soils to stimulate reductive dechlorination by DIRB, and so could iron
oxide. Also, humic substances serve as electron shuttles in DIRB and
arylhalorespiring bacteria. These microorganisms transfer electrons to
dissolved humic substances, and the metabolites can rapidly reduce
Fe(III) oxides or halogenated compounds (Cervantes et al., 2002;
Lovley et al., 1996; Nevin and Lovley, 2000).
Thus, our present study extends the state of knowledge on
dehalogenation processes under anoxic conditions in bioreactor or
biopiling system, and offers possible solutions for bioremediation impli-
cations of e-waste-contaminated soils. Further efforts, to adjust the
growth environments of functional bacteria and gain insight into
the complex chemical–biological mechanisms of dehalogenation in
e-waste-contaminated soils, will aid in extrapolation of our laboratory
results to field conditions.
Acknowledgments
This study was supported by the Joint Funds of the National Natural
Science Foundation of China and the Natural Science Foundation of
Guangdong Province, China (no. U1133004), and the National Natural
Science Foundation of China (nos. 41173082 & 41322008).
432 M. Song et al. / Science of the Total Environment 502 (2015) 426–433
Appendix A. Supplementary data
Detailed information can be found in the supporting document on
soil properties, initial PCBs/PBDEs contamination, effects of different
electron donor, and PCBs/PBDEs concentration change in sterile control.
Supplementary data are available online at http://dx.doi.org/10.1016/j.
scitotenv.2014.09.045.
References
Abramowicz DA. Aerobic and anaerobic biodegradation of PCBs — a review. Crit Rev
Biotechnol 1990;10:241–9.
Alder AC, Haggblom MM, Oppenhelmer SR, Young LY. Rductive dechlorination of
polychlorinated-biphenyls in anaerobic sediments. Environ Sci Technol 1993;27:
530–8.
Bedard DL. Polychlorinated biphenyls in aquatic sediments: environmental fate and out-
look for biological treatment. Dehalogenation Microb Process Environ Appl 2003:
443–65.
Bedard DL, Unterman R, Bopp LH, Brennan MJ, Haberl ML, Johnson C. Rapid assay
for screening and characterizing microorganisms for the ability to degrade
polychlorinated-biphenyls. Appl Environ Microbiol 1986;51:761–8.
Bedard DL, Pohl EA, Bailey JJ, Murphy A. Characterization of the PCB substrate range of mi-
crobial dechlorination process LP. Environ Sci Technol 2005;39:6831–8.
Boyle AW, Phelps CD, Young LY. Isolation from estuarine sediments of a Desulfovibrio
strain which can grow on lactate coupled to the reductive dehalogenation of 2,4,6-
tribromophenol. Appl Environ Microbiol 1999;65:1133–40.
Brennan EW, Lindsay WL. Reduction and oxidation effect on the solubility and transfor-
mation of iron oxides. Soil Sci Soc Am J 1998;62:930–7.
Bzdusek PA, Christensen ER, Lee CM, Pakdeesusuk U, Freedman DL. PCB congeners and
dechlorination in sediments of Lake Hartwell, South Carolina, determined from
cores collected in 1987 and 1998. Environ Sci Technol 2006a;40:109–19.
Bzdusek PA, Lu JH, Christensen ER. PCB congeners and dechlorination in sediments of
Sheboygan River, Wisconsin, determined by matrix factorization. Environ Sci Technol
2006b;40:120–9.
Cervantes FJ, de Bok FAM, Tuan DD, Stams AJM, Lettinga G, Field JA. Reduction of humic
substances by halorespiring, sulphate-reducing and methanogenic microorganisms.
Environ Microbiol 2002;4:51–7.
Chang Y-C, Takada K, Choi D, Toyama T, Sawada K, Kikuchi S. Isolation of biphenyl and
polychlorinated biphenyl-degrading bacteria and their degradation pathway. Appl
Biochem Biotechnol 2013;170:381–98.
Chen C-Y, Wang C-K, Shih Y-H. Microbial degradation of 4-monobrominated diphenyl
ether in an aerobic sludge and the DGGE analysis of diversity. J Environ Sci Health
B 2010;45:379–85.
Cutter LA, Watts JEM, Sowers KR, May HD. Identification of a microorganism that links its
growth to the reductive dechlorination of 2,3,5,6-chlorobiphenyl. Environ Microbiol
2001;3:699–709.
Dabrowska L, Rosinska A. Change of PCBs and forms of heavy metals in sewage sludge
during thermophilic anaerobic digestion. Chemosphere 2012;88:168–73.
Deng D, Guo J, Sun G, Chen X, Qiu M, Xu M. Aerobic debromination of deca-BDE: Isolation
and characterization of an indigenous isolate from a PBDE contaminated sediment.
Int Biodeterior Biodegrad 2011;65:465–9.
Egloff C, Crump D, Chiu S, Manning G, McLaren KK, Cassone CG, et al. In vitro and in ovo
effects of four brominated flame retardants on toxicity and hepatic mRNA expression
in chicken embryos. Toxicol Lett 2011;207:25–33.
Fahrbach M, Kuever J, Meinke R, Kampfer P, Hollender J. Denitratisoma oestradiolicum gen.
nov., sp. nov., a 17 beta-oestradiol-degrading, denitrifying betaproteobacterium. Int J
Syst Evol Microbiol 2006;56:1547–52.
Feng C, Yue X, Li F, Wei C. Bio-current as an indicator for biogenic Fe(II) generation driven
by dissimilatory iron reducing bacteria. Biosens Bioelectron 2013;39:51–6.
Fennell DE, Rhee SK, Ahn YB, Haggblom MM, Kerkhof LJ. Detection and characterization of
a dehalogenating microorganism by terminal restriction fragment length polymor-
phism fingerprinting of 16S rRNA in a sulfidogenic, 2-bromophenol-utilizing enrich-
ment. Appl Environ Microbiol 2004;70:1169–75.
Fetzner S. Bacterial dehalogenation. Appl Microbiol Biotechnol 1998;50:633–57.
Frye CA, Bo E, Calamandrei G, Calza L, Dessi-Fulgheri F, Fernandez M, et al. Endocrine dis-
rupters: a review of some sources, effects, and mechanisms of actions on behaviour
and neuroendocrine systems. J Neuroendocrinol 2012;24:144–59.
Furukawa K. Biochemical and genetic bases of microbial degradation of polychlorinated
biphenyls (PCBs). J Gen Appl Microbiol 2000;46:283–96.
Futagami T, Morono Y, Terada T, Kaksonen AH, Inagaki F. Distribution of dehalogenation
activity in subseafloor sediments of the Nankai Trough subduction zone. Philos Trans
R Soc Lond B Biol Sci 2013;368:20120249.
Gotoh S, Patrick WH. Transformation of iron in a waterlogged soil as influenced by redox
potential and pH. Soil Sci Soc Am J 1974;38:66–71.
Hamlin HJ, Guillette Jr LJ. Embryos as targets of endocrine disrupting contaminants in
wildlife. Birth Defects Res C Embryo Today 2011;93:19–33.
He J, Robrock KR, Alvarez-Cohen L. Microbial reductive debromination of polybrominated
diphenyl ethers (PBDEs). Environ Sci Technol 2006;40:4429–34.
Heron G, Christensen TH. Impact of sediment-bound iron on redox buffering in a landfill
leachate polluted aquifer (Vejen, Denmark). Environ Sci Technol 1995;29:187–92.
Holliger C, Hahn D, Harmsen H, Ludwig W, Schumacher W, Tindall B, et al. Dehalobacter
restrictus gen. nov. and sp. nov., a strictly anaerobic bacterium that reductively
dechlorinates tetra- and trichloroethene in an anaerobic respiration. Arch Microbiol
1998;169:313–21.
Kim Y-M, Murugesan K, Chang Y-Y, Kim E-J, Chang Y-S. Degradation of polybrominated
diphenyl ethers by a sequential treatment with nanoscale zero valent iron and aero-
bic biodegradation. J Chem Technol Biotechnol 2012;87:216–24.
Lee LK, He J. Reductive debromination of polybrominated diphenyl ethers by
anaerobic bacteria from soils and sediments. Appl Environ Microbiol 2010;76:
794–802.
Li J, Mgonella MK, Bzdusek PA, Christensen ER. PCB congeners and dechlorination
in sediments of Upper Sheboygan River, Wisconsin. J Great Lakes Res 2005;31:
174–86.
Li F, Wang X, Li Y, Liu C, Zeng F, Zhang L, et al. Enhancement of the reductive transforma-
tion of pentachlorophenol by polycarboxylic acids at the iron oxide-water interface.
J Colloid Interface Sci 2008;321:332–41.
Li Z, Inoue Y, Suzuki D, Ye L, Katayama A. Long-term anaerobic mineralization of penta-
chlorophenol in a continuous-flow system using only lactate as an external nutrient.
Environ Sci Technol 2012;47:1534–41.
Liu WT, Marsh TL, Cheng H, Forney LJ. Characterization of microbial diversity by deter-
mining terminal restriction fragment length polymorphisms of genes encoding 16S
rRNA. Appl Environ Microbiol 1997;63:4516–22.
Lonergan DJ, Jenter HL, Coates JD, Phillips EJP, Schmidt TM, Lovley DR. Phylogenetic anal-
ysis of dissimilatory Fe(III)-reducing bacteria. J Bacteriol 1996;178:2402–8.
Lovley DR, Coates JD, BluntHarris EL, Phillips EJP, Woodward JC. Humic substances as elec-
tron acceptors for microbial respiration. Nature 1996;382:445–8.
Luo C, Xie S, Sun W, Li X, Cupples AM. Identification of a novel toluene-degrading bacte-
rium from the candidate phylum TM7, as determined by DNA stable isotope probing.
Appl Environ Microbiol 2009;75:4644–7.
Magar VS, Brenner RC, Johnson GW, Quensen JF. Long-term recovery of PCB-
contaminated sediments at the Lake Hartwell superfund site: PCB dechlorination. 2.
Rates and extent. Environ Sci Technol 2005a;39:3548–54.
Magar VS, Johnson GW, Brenner RC, Quensen JF, Foote EA, Durell G, et al. Long-term recov-
ery of PCB-contaminated sediments at the Lake Hartwell superfund site: PCB dechlo-
rination. 1. End-member characterization. Environ Sci Technol 2005b;39:3538–47.
Maithreepala RA, Doong RA. Transformation of carbon tetrachloride by biogenic iron spe-
cies in the presence of Geobacter sulfurreducens and electron shuttles. J Hazard Mater
2009;164:337–44.
McCormick ML, Bouwer EJ, Adriaens P. Carbon tetrachloride transformation in a model
iron-reducing culture: relative kinetics of biotic and abiotic reactions. Environ Sci
Technol 2002;36:403–10.
Nevin KP, Lovley DR. Potential for nonenzymatic reduction of Fe(III) via electron shuttling
in subsurface sediments. Environ Sci Technol 2000;34:2472–8.
Pakdeesusuk U, Lee CM, Coates JT, Freedman DL. Assessment of natural attenuation
via in situ reductive dechlorination of polychlorinated bipheny is in sediments
of the twelve mile creek arm of Lake Hartwell, SC. Environ Sci Technol 2005;
39:945–52.
Payne RB, May HD, Sowers KR. Enhanced reductive dechlorination of polychlorinated bi-
phenyl impacted sediment by bioaugmentation with a dehalorespiring bacterium.
Environ Sci Technol 2011;45:8772–9.
Payne RB, Fagervold SK, May HD, Sowers KR. Remediation of polychlorinated biphenyl
impacted sediment by concurrent bioaugmentation with anaerobic halorespiring
and aerobic degrading bacteria. Environ Sci Technol 2013;47:3807–15.
Purser D. Toxicity of fire retardants in relation to life safety and environmental hazards.
Fire Retard Mater 2001:69–127.
Rayne S, Ikonomou MG, Whale MD. Anaerobic microbial and photochemical degradation
of 4,4′-dibromodiphenyl ether. Water Res 2003;37:551–60.
Rhee SK, Fennell DE, Haggblom MM, Kerkhof LJ. Detection by PCR of reductive
dehalogenase motifs in a sulfidogenic 2-bromophenol-degrading consortium
enriched from estuarine sediment. FEMS Microbiol Ecol 2003;43:317–24.
Robrock KR, Korytar P, Alvarez-Cohen L. Pathways for the anaerobic microbial debromination
of polybrominated diphenyl ethers. Environ Sci Technol 2008;42:2845–52.
Sanford RA, Cole JR, Loffler FE, Tiedje JN. Characterization of Desulfitobacterium
chlororespirans sp. nov., which grows by coupling the oxidation of lactate to the re-
ductive dechlorination of 3-chloro-4-hydroxybenzoate. Appl Environ Microbiol
1996;62:3800–8.
Sanford RA, Cole JR, Tiedje JM. Characterization and description of Anaeromyxobacter
dehalogenans gen. nov., sp. nov., an aryl-halorespiring facultative anaerobic
myxobacterium. Appl Environ Microbiol 2002;68:893–900.
She Y-Z, Wu J-P, Zhang Y, Peng Y, Mo L, Luo X-J, et al. Bioaccumulation of polybrominated
diphenyl ethers and several alternative halogenated flame retardants in a small her-
bivorous food chain. Environ Pollut 2013;174:164–70.
Shih Y-h, Chou H-L, Peng Y-H. Microbial degradation of 4-monobrominated diphenyl
ether with anaerobic sludge. J Hazard Mater 2012;213:341–6.
Tobler NB, Hofstetter TB, Straub KL, Fontana D, Schwarzenbach RP. Iron-mediated micro-
bial oxidation and abiotic reduction of organic contaminants under anoxic conditions.
Environ Sci Technol 2007;41:7765–72.
Tokarz III JA, Ahn M-Y, Leng J, Filley TR, Nies L. Reductive debromination of
polybrominated diphenyl ethers in anaerobic sediment and a biomimetic system.
Environ Sci Technol 2008;42:1157–64.
Wang FX, Wang J, Dai JY, Hu GC, Wang JS, Luo XJ, et al. Comparative tissue distribution,
biotransformation and associated biological effects by decabromodiphenyl ethane
and decabrominated diphenyl ether in male rats after a 90-day oral exposure
study. Environ Sci Technol 2010;44:5655–60.
Wohlfarth G, Diekert G. Anaerobic dehalogenases. Curr Opin Biotechnol 1997;8:290–5.
Wong MH, Wu SC, Deng WJ, Yu XZ, Luo Q, Leung AOW, et al. Export of toxic chemicals — a
review of the case of uncontrolled electronic-waste recycling. Environ Pollut 2007;149:
131–40.
 M. Song et al. / Science of the Total Environment 502 (2015) 426–433 433

Wu C-Y, Zhuang L, Zhou S-G, Li F-B, Li X-M. Fe(III)-enhanced anaerobic transformation of
2,4-dichlorophenoxyacetic acid by an iron-reducing bacterium Comamonas koreensis
CY01. FEMS Microbiol Ecol 2010;71:106–13.
Yan T, LaPara TM, Novak PJ. The effect of varying levels of sodium bicarbonate on
polychlorinated biphenyl dechlorination in Hudson River sediment cultures. Environ
Microbiol 2006;8:1288–98.
Yang C. Microbial reduction characteristic of ferric iron in some paddy soils. Xian Yang: N. W.
Agric. For. Univ.; 2001.
Yen JH, Liao WC, Chen WC, Wang YS. Interaction of polybrominated diphenyl ethers
(PBDEs) with anaerobic mixed bacterial cultures isolated from river sediment. J Hazard
Mater 2009;165:518–24.
Zhou T, Ross DG, DeVito MJ, Crofton KM. Effects of short-term in vivo exposure to
polybrominated diphenyl ethers on thyroid hormones and hepatic enzyme activities
in weanling rats. Toxicol Sci 2001;61:76–82.