Article

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Biofouling Control with Bead-Entrapped Quorum Quenching

Bacteria in Membrane Bioreactors: Physical and Biological Effects
Sang-Ryoung Kim,† Hyun-Suk Oh,† Sung-Jun Jo,† Kyung-Min Yeon,† Chung-Hak Lee,*,†
Dong-Joon Lim,‡ Chi-Ho Lee,§ and Jung-Kee Lee§
†
School of Chemical and Biological Engineering, Seoul National University, Seoul 151-744, Republic of Korea
‡
School of Chemical Engineering, Yeungnam University, Gyeongsan 712-749, Republic of Korea
§
Department of Life Science and Genetic Engineering, Paichai University, Daejeon 302-735, Republic of Korea
*
S Supporting Information

ABSTRACT: Recently, interspecies quorum quenching by bacterial cells

encapsulated in a vessel was described and shown to be efficient and
economically feasible for biofouling control in membrane bioreactors
(MBRs). In this study, free-moving beads entrapped with quorum
quenching bacteria were applied to the inhibition of biofouling in a MBR.
Cell entrapping beads (CEBs) with a porous microstructure were
prepared by entrapping quorum quenching bacteria (Rhodococcus sp.
BH4) into alginate beads. In MBRs provided with CEBs, the time to
reach a transmembrane pressure (TMP) of 70 kPa was 10 times longer
than without CEBs. The mitigation of biofouling was attributed to both
physical (friction) and biological (quorum quenching) effects of CEBs,
the latter being much more important. Because of the quorum quenching
effect of CEBs, microbial cells in the biofilm generated fewer extracellular
polymeric substances and thus formed a loosely bound biofilm, which enabled it to slough off from the membrane surface more
easily. Furthermore, collisions between the moving CEBs and membranes gave rise to frictional forces that facilitated detachment
of the biofilm from the membrane surface. CEBs bring bacterial quorum quenching closer to being a practical solution to the
problem of biofouling in MBRs.

■ INTRODUCTION
Although membrane bioreactors (MBRs) have been in
commercial use for more than two decades, membrane
biofouling caused by the formation of biocakes (deposited
microbial flocs plus biofilm) on the membrane surface still
remains a bottleneck that limits their widespread use.1,2 Many
researchers have attempted to mitigate biofouling in various
ways, such as by using additives or specific media,3,4 by changes in
the design of the MBR system,5 or modification of the membrane
surface.6 Recently, novel biological approaches have been
attempted to control biofouling using quorum quenching,7−10
that is, via disruption of quorum sensing. Quorum sensing is the
cell−cell communication between microorganisms, which
determines phenotypes such as biofilm formation, secretion of
extracellular polymeric substances (EPS) and virulence.11,12
Yeon et al.7,8 prepared a magnetic enzyme carrier by
immobilizing a quorum quenching enzyme, acylase, onto
magnetic particles and demonstrated its potential as a novel
approach to control biofouling in MBR. Kim et al. 9 immobilized
acylase onto the membrane surface and showed mitigation of
membrane biofouling. These applications, however, have
drawbacks, such as the high cost of enzyme extraction and
purification as well as enzyme instability. As an alternative to
enzymatic quenching, Oh et al. 10 isolated bacteria that produce
quorum quenching enzymes and also developed a microbial
vessel in which quorum quenching bacteria (Rhodococcus sp.
BH4) were encapsulated. They observed that a submerged MBR
equipped with the microvessel has a much lower biofouling
tendency compared with a conventional MBR. In their study,
however, quorum quenching bacteria were confined within a
small vessel that was submerged in a fixed place in the MBR so
that they could degrade only soluble signal molecules that were
able to diffuse into the vessel. As such, the mass transfer of signal
molecules from the mixed liquor to the inside of the microbial
vessel was limited.
The aim of this study was to find an alternative method of
bacterial quorum quenching that is both more efficient than
using a microbial vessel and more feasible than enzymatic
quorum quenching from the viewpoint of practical application.
We prepared free-moving beads using alginate and entrapped
Rhodococcus sp. BH4 into highly interconnected microstructural
pores of the beads, which will be called cell entrapping beads
(CEBs) throughout this article. We placed CEBs directly into a
Received: October 2, 2012
Revised: December 17, 2012
Accepted: December 20, 2012
Published: December 20, 2012

© 2012 American Chemical Society 836 dx.doi.org/10.1021/es303995s | Environ. Sci. Technol. 2013, 47, 836−842
Environmental Science & Technology
submerged MBR and allowed them to move freely together with
other microorganisms in the mixed liquor as well as to contact
the biofilm on the filtration membrane to catch up signal
molecules in the biofilm more easily. It is thought that CEBs
inhibit biofilm formation through interspecies quorum quench-
ing as well as by physical washing through their collisions with the
membrane surface.
■ EXPERIMENTAL SECTION
Quorum Quenching Bacteria. Rhodococcus sp. BH4 was
isolated from a working MBR plant for wastewater treatment
(capacity 18 000 m3/d, Okcheon, Korea), using an enrichment
culture method described by Oh et al.10
Preparation of CEBs. Sodium alginate (Sigma-Aldrich), a
nontoxic substance to bacteria,13 was used as a cell immobiliza-
tion material. The isolated Rhodococcus sp. BH4 were inoculated
in Luria−Bertani (Miller, US) broth at 30 °C for 24 h. The BH4
culture was centrifuged (12 000g, 15 min), washed with water,
and resuspended in 3 mL of water. The BH4 suspension (200 mg
BH4/mL of water) was gently mixed with 97 mL of the sterile
sodium alginate suspension to make a 4% (w/v) BH4−alginate
suspension. The BH4−alginate suspension was dripped into 3%
(w/v) CaCl2 solution through a nozzle at a rate of 1.6 mL/min.
As depicted in Supporting Information Figure S1, the dripping
device consisted of a nozzle, fluid line, and pump with a velocity
controller. The CEBs were formed and left in CaCl2 solution for
3 h before being washed three times with distilled water and dried
at room temperature. The average size and density of CEBs were
approximately 3.5 mm and 1.5 g/mL, respectively. The BH4
content of the CEBs was 6.0 mg BH4/g sodium alginate. Because
the size and mechanical properties of the CEBs can be easily
controlled by changing the diameter of the nozzle or the
concentration of CaCl2, this method offers advantages for the
preparation of diverse CEBs suitable for various types of MBRs.
Measurement of Activity and Stability of CEBs. The
quorum quenching activity and stability of CEBs were evaluated
by the degradation rate of standard C8-HSL (N-octanoyl-DL-
homoserine lactone) (Sigma-Aldrich, USA), which is one of the
dominant signal molecules (autoinducers) in the MBR for
wastewater treatment.7 The degradation rate of C8-HSL was
measured according to the method described by previous
studies.7−10 C8-HSL was added to 50 mM Tris−HCl buffer (pH
7.0, 30 mL) to a final concentration of 200 nM. Twenty
individual CEBs were then added to the Tris−HCl buffer
containing C8-HSL. The activity of the CEBs was measured via
the decrease in the C8-HSL concentration with time. The
stability of the CEBs was measured from the decrease in the C8-
HSL concentration for 30 min and was monitored 13 times
during continuous MBR operation over 30 days. The CEBs were
periodically removed from the mixed liquor, but were returned to
the MBR following activity measurement.
MBR Operation. Two laboratory-scale MBRs, each with a 1.6
L working volume, were operated in parallel. Three sets of
operating schemes were designed for two MBRs, depending on
the presence of either vacant beads or CEBs in each MBR (Figure
1): set 1, control and CEBs (with BH4 cells); set 2, control and
vacant beads (without BH4 cells); set 3, vacant beads and CEBs.
The number of vacant beads or CEBs inserted into each MBR
was 40, and each MBR was always operated under a constant flux
of 28.7 L/(m2 h). Hydraulic retention times and sludge retention
times were set to 5.3 h and 25 d, respectively. The submerged
hollow fiber membrane was made of polyvinylidene fluoride
(ZeeWeed 500, GE-Zenon, USA) with an effective area of 13.4
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Figure 1. Schematic diagram for three sets of operations of MBRs.
cm2. Activated sludge was taken from a wastewater treatment
plant (Tancheon, Korea). Mixed liquor suspended solid in the
MBR was maintained at 12.5−13.0 g/L. The detailed
composition of the synthetic wastewater is given in Supporting
Information Table S1.
Measurement of Loosely and Tightly Bound Biofilms.
For the qualitative and quantitative analysis of biofilm formed on
the surface of membranes, biofilms were detached from the used
membranes and were classified into two types: loosely bound
biofilm (LB biofilm) or tightly bound biofilm (TB biofilm). The
former was defined as biofilm that can be detached only by air-
scouring at a fixed aeration time and rate, whereas the latter, by
sonication and subsequent air-scouring (Supporting Information
Figure S2). The used membranes covered with biofilm were
submerged in an aeration tank filled with 1 L of water. The LB
biofilm was obtained after 80 min of aeration at a rate of 3 L/min.
The remaining biofilm on the same used membrane was further
sonicated for 10 min, followed by an additional 20 min of
aeration to obtain the TB biofilm. The dry weight of each biofilm
in suspension was measured. The total weight of LB and TB
biofilms was regarded as the total attached biomass (TAB).
Extraction of EPS from the biofilm in suspension was carried
out using an ion-exchange resin method.14 A cation exchange
resin in sodium form (CER, Dowex Marathon, Sigma-Aldrich)
was washed for 1 h in phosphate buffer and was added to the
suspension of each biofilm (10 g CER/g detached biofilm). The
mixed suspension was stirred at 300 rpm for 2 h and then
centrifuged at 4000g for 20 min. A pellet composed of cells and
CER formed in the bottom of the tube, and the supernatant
contained EPS. The weight of cells was determined by
subtracting the weight of CER from the weight of the pellet.
The amount of EPS was determined by subtracting the cell
weight from the TAB.
Extraction and Analysis of Acyl Homoserine Lacotones
(AHLs). Standard AHLs and AHL extracts were analyzed by
HPLC (Waters, USA). AHL was extracted from the biofilm on
the used membrane as follows: The used membrane was placed
in 400 mL of deionized water, and the biofilm was detached by
backwashing and sonication. After the membrane was removed,
dx.doi.org/10.1021/es303995s | Environ. Sci. Technol. 2013, 47, 836−842
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Figure 2. SEM microphotographs of the beads: cross section of a vacant bead (a) ×25, (b) ×1000, and (c) ×6000. Cross section of a CEB (d) ×25, (e)
×1000, and (f) ×6000.
the biofilm in suspension was shaken with 100 mL of ethyl
acetate for 2 h. After the organic layer was separated from the
water layer using a separating funnel, it was dried with a vacuum
evaporator. The residue was redissolved with 200 μL of methanol
for HPLC analysis. Gradient elution was performed with a
mixture of water/methanol as a mobile phase, and the UV
detector was set at 210 nm. Standard AHLs were dissolved in
methanol to obtain 1 mg/mL solutions. Aliquots (20 μL) of each
of these solutions were added to 980 μL of methanol−water
(35:65, v/v) with 0.1% formic acid. The column used in the
HPLC was a Gemini C18, 50 mm × 2 mm, 5 μm particle size.
AHL standard mixtures (25 μL), blanks, or AHL extracts were
injected at a flow rate of 0.25 mL/min. The HPLC was connected
to a fraction collector. Fractions were collected every 9 min into a
test tube, reduced in volume, then loaded for the bioassay of AHL
molecules using an indicating agar plate. The detailed analysis
procedure for HPLC is described in the Supporting
Information.15
Analytical Methods. The internal and external morpholo-
gies of the vacant beads and CEBs were identified by SEM. The
live (green) or dead (red) cells inside the CEBs were stained by a
Viability kit according to the manufacturer’s instructions and
were observed using CLSM. The detailed analytical procedures
for SEM and CLSM are described in the Supporting Information.
■
RESULTS AND DISCUSSION
Characterization of CEBs. The vacant beads were almost
spherical, with a smooth surface and uniform size. Entrapment of
quorum quenching bacteria (BH4) into the beads did not result
in any significant change in either the shape or the size of beads
(Supporting Information Figure S3a). The size of CEBs was ∼3.5
mm, and their density was roughly 1.5g/mL. The CEBs were able
to circulate in the mixed liquor under aeration (Supporting
Information Figure S3b).
The cross-sectional SEM image showed the morphologies of
the vacant beads and CEBs as well as the differences between the
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two (Figure 2). The vacant beads and CEBs possess a porous
microstructure, with a high degree of interconnectivity (Figure
2a and d). Since there are many pores and the pore diameter in
CEBs is around 300 μm, CEBs appear to provide enough space
for BH4 colonization as well as low mass transfer resistance.
Inside the pore, no BH4 were observed in vacant beads (Figure
2b), whereas BH4 were spread on the alginate matrix surface
(Figure 2e) in CEBs. The BH4 appear as short rods with an
approximate size of 1.2−2.0 μm in length and 0.5 μm in width
(Figure 2e).
To investigate the viability of BH4 during entrapment, CLSM
images of CEBs were taken after viability staining. Before
entrapment, the proportion of free live BH4 was ∼80% (±3%),
on the basis of the ISA image. After entrapment, the BH4
appeared densely packed and evenly dispersed in the micro-
structure of the CEBs (Supporting Information Figure S4). From
the images, the mean percentage of living cells entrapped in
CEBs was calculated to be 65% (±5%). The damage to live cells
during entrapment indicates that cell immobilization had a
negative effect on cell viability. It is possible that BH4 near the
surface of CEBs were killed by contact with CaCl2 solution, but
the decrease in the amount of live BH4 was not substantial
(Supporting Information Figure S4b). The SEM and CLSM
analysis confirmed that CEBs were successfully constructed by
the combination of alginate, Ca+, and BH4.
Quorum Quenching Activity of Free BH4 and CEBs. The
quorum quenching activity of isolated free BH4 was tested using
a bioassay with C8-HSL, which was most abundant in the
biofilm-formed membranes in MBRs.7 Live BH4 readily
degraded C8-HSL, whereas dead BH4 hardly reduced C8-HSL
levels, despite its potential physicochemical adsorption (Sup-
porting Information Figure S5). The quorum quenching activity
of CEBs was tested using the same method as for free BH4. As
shown in Figure 3, CEBs degraded 91% of C8-HSL, whereas the
vacant beads removed less than 10% of the C8-HSL in 60 min.
The removal by vacant beads was attributed to its physicochem-
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Figure 3. Quantitative quorum quenching activity of control, vacant
beads, and CEBs (n = 4).
ical adsorption because vacant beads have neither quorum
quenching bacteria nor quorum quenching enzyme. A control
was also conducted to check the potential removal of C8-HSL by
its adsorption onto the surface of a glass beaker, but the
adsorption was negligible.
Application of CEBs to the MBR. CEBs were applied to
submerged MBRs to test their potential to inhibit biofouling in
MBRs (set 1 in Figure 1). Two lab-scale MBRs in continuous
mode were operated in parallel under identical operating
conditions except for the addition of CEBs to one MBR at the
start of the operation. The rise of the profiles transmembrane
Figure 4. Comparison of TMP between (a) control and CEBs MBRs, (b) control and vacant beads MBRs, and (c) vacant beads and CEBs MBRs under
the same operating conditions.
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pressure (TMP) of the control and CEB MBRs were compared
to evaluate the inhibition of biofouling by CEBs. As shown in
Figure 4a, it took 1.8 d for the TMP to reach 70 kPa in the first
cycle of the control MBR, whereas it took 18.8 d for the first cycle
of the CEB-treated MBR. Thus, CEBs mitigated the formation of
biofilm and extended the time required to reach the TMP of 70
kPa by ∼10-fold, compared with the control MBR. From a
practical point of view, this is important because the delay in the
rise of TMP is closely associated with a saving of energy in the
operation of the MBR.
The remarkable effect of CEBs to reduce biofouling is better
than that reported by Oh et al.10 They encapsulated the same
quorum quenching bacteria, BH4, into a microbial vessel, applied
it to a submerged MBR, and observed that the microbial vessel
resulted in much lower biofouling compared with a conventional
MBR. Although a direct comparison between results obtained
from CEBs and the microbial vessel is difficult, CEBs appear to
be superior to the microbial vessel in terms of reducing
membrane fouling. The excellent performance of CEBs could
be attributed either to the inhibition of biofilm formation by
quorum quenching or to the sloughing of biofilm from the
membrane surface by collision between moving CEBs and the
submerged membrane in the MBR. To verify this, two
consecutive MBR operations were carried, out as depicted in
sets 2 and 3 in Figure 1.
Physical Washing Effect by Free-Moving CEBs. To
confirm a physical washing effect, the control MBR and the MBR
with the vacant beads were run in parallel under the same
operating conditions (set 2 in Figure 1). As shown in Figure 4b, it
dx.doi.org/10.1021/es303995s | Environ. Sci. Technol. 2013, 47, 836−842
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took 1.8 d to reach the TMP of 70 kPa in the control MBR,
whereas it took 3.1 d to reach the same TMP in the MBR with
vacant beads. Although the vacant beads with porous micro-
structures contain no BH4, they continuously circulate in the
mixed liquor and collide with the surface of submerged
membranes in the MBR. This collision could facilitate the
detachment of biocake already deposited or formed on the
membrane surface. A physical washing effect by moving media in
MBRs has been previously reported.4,16
To quantitatively evaluate the physical washing effect, a
separate experiment was designed. Two control MBRs with
neither vacant beads nor CEBs were run until the TMP reached
70 kPa. The used hollow fiber membrane module was then
removed from each control MBR, and each used membrane
module was immersed in two separated beakers containing 1 L of
water. Forty vacant beads were added to only one beaker, and
each beaker was then aerated for 80 min (3 L/min) to assess how
much biomass would be detached from each used module and to
compare each experiment. Further sonication following aeration
also made it possible to determine the TAB of each used
membrane module. For the five repeating measurements, 0.64 g
(87% of TAB) of biomass was detached in the beaker with the
vacant beads, whereas 0.52 g (72% of TAB) of biomass was
detached in the beaker without the vacant beads (Figure S6). On
the other hand, the average TAB of both membrane modules was
similar: 0.74 (±0.05) g in the beaker with the beads and 0.73
(±0.04) g in the beaker without beads, with a 5% relative
standard deviation of each. Consequently, it can be concluded
that the vacant beads facilitated the detachment and, thus,
increased the amount of detached biomass by ∼15% through
their collision with the membrane.
Quorum Quenching Effect by Moving CEB. To confirm
the quorum quenching effect, one MBR with vacant beads and
the other MBR with CEBs were run under the same operating
conditions (set 3 in Figure 1). During the operation of each
MBR, a used membrane module was replaced by a new one when
the TMP exceeded 70 kPa, and TMP monitoring was
reperformed with the new membrane. As shown in Figure 4c,
it took 17 d to reach the TMP of 70 kPa for the MBR with CEBs,
whereas it took only 2 or 3 d to reach the same TMP for the MBR
with the vacant beads. Eight cycles were thus repeated in the
MBR with the vacant beads during only cycle with the one with
CEBs. Expressed differently, CEBs extended the time required
for the MBR to reach the TMP of 70 kPa by about 7-fold. It is
worth noting that assuming that the physical washing effects of
the vacant beads and CEBs were identical because the same
number of vacant beads or CEBs were added to each MBR, the
large difference in the rate of the TMP rise (set 3 in Figure 1) is
attributable only to the BH4 in the porous microstructural CEBs.
Mechanisms of Quorum Quenching. To investigate the
mechanisms of quorum quenching, biofilms were detached from
the used membrane modules after 75 h of operation in both the
MBR with vacant beads and the MBR with CEBs. Biofilms from
both used modules were analyzed in terms of EPS, TAB, and
adhesiveness and then compared with each other. After 75 h of
operation, the TMP reached 70 kPa in the MBR with vacant
beads, whereas only 7.9 kPa was reached in the MBR with CEBs.
This coincided well with the greater TAB in the former (0.77 g)
compared with the latter (0.24 g), as shown in Table 1: less than
one-third of the biomass developed on the membrane surface in
the MBR with quorum quenching bacteria during the same
operation period.
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Table 1. The Amount of TAB, EPS and Loosely and Tightly
Bound Biofilms in the Used Membrane Modules for the MBR
with Vacant Beads and MBR with CEBs
MBR with vacant
beads MBR with CEBs
a
TMP at the operating time of 70.3 kPa 7.9 kPa
75 h
TABb 0.77 g 0.24 g
TAB EPS 0.36 g (47% of TAB)c 0.07 g (29% of TAB)c
cell 0.41 g 0.17 g
TAB loosely bound 0.53 g (68% of 0.22 g (89% of
biofilm TAB)d TAB)d
tightly bound biofilm 0.24 g (32% of
TAB)d
0.02 g (11% of
TAB)d
a
TMP: transmembrane pressure. bTAB: total attached biomass. cEPS
percentage = (EPS/TAB) × 100. dBound biofilm percentage =
(bound biofilm/TAB) × 100.
To characterize the detached biomass, the TAB was further
divided into two components: EPS and cells. Not only the total
amount, but also the proportion of EPS was much lower in the
MBR with CEBs (0.07 g, 29%) than in the MBR with vacant
beads (0.36 g, 47%). It is known that quorum sensing regulates
the production of EPS through the transcription of target genes
and determines the physiology of the microbial community.11
Previous studies confirmed that enzymatic quorum quenching
decreases EPS production in the biofilm.7−9,17 Rhodococcus sp.
have been reported to generate an enzyme (lactonase) that can
degrade AHLs.18 Consequently, the application of CEBs (i.e.,
BH4) to MBRs inhibits quorum sensing between cells by
reducing the concentration of AHLs and thus decreasing EPS
production in the biofilm. The attached biomass was also
classified into two types: LB biofilm or TB biofilm. As illustrated
in Supporting Information Figure S2, one portion of the TAB
biofilm (TB biofilm) requires more vigorous conditions for
detachment than the other (LB biofilm), indicating that TB
biofilm has stronger cohesive or adhesive forces (or both) than
LB biofilm. In Table 1, both the total amount and the portion of
the TB biofilm was substantially lower in the MBR with CEBs
(0.02 g, 11%) than in the MBR with vacant beads (0.24 g, 32%).
This could be attributed to the lower production of EPS, the key
element for the construction of biofilm due to quorum
quenching by CEBs.19
Identification of Signal Molecules for Quorum Sensing
in MBRs. An important point concerns the demonstration of the
destruction of signal molecules by CEBs. For this purpose, both
MBRs in this study were run for 48 h, and the extracts from
biofilm formed on the membrane surfaces from MBRs with or
without CEBs were then analyzed by HPLC and a bioassay. The
extract from the MBR with vacant beads showed four peaks (blue
line in Figure 5a). Two of the four peaks appeared with a
retention time of 3.2 and 11.8 min and were identified as C8-HSL
and 3oxoC8-HSL, respectively, by comparison with the peaks of
two standard signal molecules (green and yellow lines in Figure
5a). In the extract from the MBR with CEBs (red line in Figure
5a), however, no AHL was detected. To ensure that the
destruction of AHLs occurred by CEBs, we collected two HPLC
fractions, each after 9 min: fraction 1 was eluent collected for the
first 9 min, and fraction 2 was eluent collected for the second 9
min. Both fractions were analyzed by bioassay with A136 as a
reporter strain.7 The two fractions from the MBR with vacant
beads showed blue colors, indicating the presence of AHLs
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Figure 5. Identification of AHLs extracted from the biofilm formed on the used membrane by HPLC. (a) Chromatogram of standard and extracted
AHLs. (b) Bioassay of fractions (1 and 2) collected every 9 min for the MBR with vacant beads. (C) Bioassay of fractions (1 and 2) collected every 9 min
for the MBR with CEBs.

Figure 6. Reconstructed CLSM images of biofilm formed on the membrane surface in (a) control MBR, (b) MBR with vacant beads, and (c) MBR with
CEBs after 48 h operation, stained with SYTO9 (cell; green). Magnification: ×100. Image size: 1212 μm × 1212 μm.

(Figure 5b), whereas those from the MBR with CEBs showed no
blue color, indicating the absence of AHLs (Figure 5c).
Visual Confirmation of the Quorum Quenching Effect
by Mobile CEBs. The quorum quenching effect of CEBs was
confirmed visually using CLSM. Figure 6 represents the
reconstructed CLSM images of biofilm formed on the membrane
surfaces which were removed from the MBRs operated for 48 h.
The amount of biofilm formed in the MBR with CEBs was the
least, whereas that in the control MBR was the greatest, and that
in the MBR with vacant beads was intermediate. In summary,
CEBs induced a physiological change in microorganisms,
including a decrease in EPS production through the disruption
841
of AHLs. Consequently, the cohesion between cells or the
adhesion between cells and membrane was weakened, and thus,
less biomass was attached to the membrane with the CEBs. In
short, CEBs can inhibit biofilm formation by quorum quenching.
Effect of CEBs on MBR Performance. In addition to
variation in TMP, the quality of the permeated water is another
important factor in MBR performance. We monitored the
removal efficiencies of COD in three MBRs on the basis of their
feed and permeate concentrations. Although the influent COD
to three reactors was around 500 mg/L, three reactors exhibited
similar COD concentrations in the permeate with more than
96% of COD removal: 6.2−19.1 mg/L for the control, 6.3−16.4
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Environmental Science & Technology
mg/L with vacant beads, and 6.9−14.2 mg/L for CEBs. There
was no significant difference in the operational parameters
among three MBRs throughout the entire experimental period.
Moreover, taking into account the volume of CEBs added to the
MBR was less than 0.63% of the working reactor volume, the
adsorption of COD or TN on CEBs would be negligible.
Therefore, it has been concluded that quorum quenching with
CEBs mitigates membrane biofouling but does not decrease
microbial activity, at least for the degradation of organic matter in
the MBRs.
Stability of CEBs. The structural integrity and quorum
quenching activity of CEBs were monitored during the operation
of continuous MBR for 30 d. Surprisingly, the quorum quenching
activity of CEBs increased by ∼3% after 30 d of operation, which
is highly favorable for practical applications (Supporting
Information Figure S7a). We also monitored the wet weight of
CEBs and vacant beads during the MBR operation (Supporting
Information Figure S7b). The wet weight of the CEBs increased
by ∼4% after 25 d of operation, whereas that of the vacant beads
did not change substantially. Thus, it can be concluded that the
increase in the quorum quenching activity of CEBs is caused by
the growth of BH4 within the beads. Furthermore, the sizes of
the CEBs and vacant beads were nearly constant and maintained
their structural integrity at least for the test period of 25 d
(Supporting Information Figure S8).
In summary, CEBs prepared in this study demonstrated their
potential for efficient biofouling control in MBRs via both
physical washing and quorum quenching effects. The application
of quorum quenching bacteria entrapped in beads could be
expanded to the plant-scale of MBRs with economic feasibility.
■
*
ASSOCIATED CONTENT
S Supporting Information
Additional information as noted in text. This material is available
free of charge via the Internet at http://pubs.acs.org.
■
AUTHOR INFORMATION
Corresponding Author
*Phone: +82-2-880-7075; fax: +82-2-874-0896; e-mail: leech@
snu.ac.kr.
Notes
The authors declare no competing financial interest.
■
ACKNOWLEDGMENTS
This research is supported by the Korea Ministry of Environment
as ″Converging Technology Project″ (2012001440001).
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