Philippine Journal of Science Research Note

139 (1): 71-78, June 2010

ISSN 0031 - 7683

Characterization and Identification of Congo Red

Decolorizing Bacteria from Monocultures and Consortia

Aileen C. Jalandoni – Buan1, Anna Lynn A. Decena-Soliven, Ernelea P. Cao2,

Virginia L. Barraquio3, and Wilfredo L. Barraquio4

1
Outsourcing and Manufacturing Solutions, Inc. (OMSI), AFP-RSBS Industrial Park,
Western Bicutan, Taguig, Metro Manila
2
Natural Sciences Research Institute, College of Science,
University of the Philippines Diliman, Quezon City 1101
3
Animal and Dairy Sciences Cluster (Dairy Training and Research Institute),
College of Agriculture, University of the Philippines Los Baños, College, Laguna 4031
4
Institute of Biology, College of Science, University of the Philippines Diliman

Congo red is a carcinogenic direct diazo dye used for the coloration of paper products. It is
recalcitrant and is thus found in effluents of paper factories. Bacteria in consortia capable of
decolorizing Congo red were isolated from polluted and non-polluted sites. Members of the
consortia were isolated and those capable of decolorizing Congo red were selected. SB13B was
the fermentative and nitrate reducing monoculture, SB12D and IRRI-1C were the fermentative
monocultures and S22B was the denitrifying monoculture. The toxicity of decolorized medium
and undegraded Congo red was tested on bacteria, yeast, rice, and mungbean plants. The
monocultures were characterized and identified. The decolorizing activities were 97% for SB13B,
96% for SB12D, 92% for IRRI-1C and 96% for S22B. S22B and SB13B were able to degrade
Congo red while reducing nitrate to nitrogen gas and nitrite. The growth of bacteria and yeast
were not inhibited by both the decolorized medium and undegraded Congo red. Decolorization
of Congo red by bacteria diminished the severe toxic effect of the dye to rice and mungbean
plants. SB13B was identified as Escherichia coli by API 20 E, fatty acid analysis and 16S rRNA
sequencing. SB12D was identified as Enterobacter dissolvens by 16S rRNA sequencing. BIOLOG
and 16S rRNA sequencing identified S22B as Pseudomonas citronellolis and Pseudomonas
aeruginosa by fatty acid analysis. IRRI-1C was identified as Klebsiella oxytoca using API 20 E.

Key Words: 16S rRNA, Congo red, Congo red decolorizing bacteria, consortia, monoculture, fatty
acid profile

INTRODUCTION NH2), which is a suspected carcinogen, phenyl (C6H5-

Dyes are usually aromatic and heterocyclic compounds
and are often recalcitrant, some of them being toxic
and even carcinogenic (Vyas and Molitoris 1995).
They include a broad spectrum of different chemical
structures, primarily based on substituted aromatic and
heterocyclic groups such as the aromatic amine (C6H5-
*Corresponding author: abuan@omsi.ph
ajbuan67@yahoo.com
CH2) and naphthyl (NO2-OH). Common to them is their
ability to absorb light in the visible region (Rajamohan
and Karthikeyan 2006). Direct dyes lack fastness during
washing and so they are popular for items which are less
likely to require fastness during washing like paper. A
direct diazo dye commonly used in the paper industry is
Congo red, which is intended primarily for the coloration
of paper products. It is a recalcitrant and a known
71
Philippine Journal of Science
Vol. 139 No. 1, June 2010
carcinogen because of the presence of the aromatic amine
group (Rajamohan and Karthikeyan 2006; van der Zee
2002; Cripps et al. 1990). Congo red is the sodium salt
of benzidinediazo-bis-1-naphtylamine-4-sulfonic acid
(formula: C32H22N6Na2O6S2; molecular weight: 696.66
g/mol). It is a secondary diazo dye.
Numerous studies have demonstrated the ability of
bacteria in monoculture to degrade dyes anaerobically
and aerobically (Banat et al. 1996). Dye decolorization
by bacteria has been observed under different conditions.
Reductive cleavage of the azo bond by a wide variety of
microorganisms, sediments, and sludges occurs readily
under anaerobic conditions (Field et al. 2003; Stolz
2001). The effectiveness of microbial decolorization
depends on the adaptability and the activity of selected
microorganisms. Many microorganisms are capable of
degrading azo dyes, including bacteria, fungi, and yeast
(Chen et al. 2003). A highly alkali thermostable Bacillus
sp. Strain SF isolated from the wastewater drain of a textile
finishing company was found capable of degrading an azo
dye (Maier et al. 2004). An azo dye-reducing, endospore
forming bacterium was isolated from textile industry
wastewater that can decolorize the azo dye Remazol Black
B (Meehan et al. 2001).
This study deals with the isolation of Congo Red
decolorizing bacteria (CRDB) singly and in consortium,
characterization and identification of Congo Red
decolorizing bacteria (CRDB) from the consortia and
determination of the toxicity of the decolorized culture
medium.
MATERIALS AND METHODS
Enrichment and isolation of Congo red decolorizing
bacteria
Samples used for isolation included sediment from
polluted canal receiving the effluents containing Congo
red and other dyes, soil from non-polluted source,
rhizosphere soil and roots of Imperata cylindrica
growing in the vicinity of the Congo red-polluted canal.
For the enrichment of Congo red decolorizing bacteria
(CRDB), the following liquid media were used: mineral
salts medium (Cohen-Bazire et al. 1957) supplemented
with 0.25% glucose, 0.1% ammonium sulfate and 1%
Congo red; mineral salts medium with 0.25% glucose,
0.1% ammonium sulfate, 0.05% potassium nitrate and
1% Congo red; mineral salts medium supplemented with
0.25% glucose, 0.1% yeast extract and 1% Congo red;
and mineral salts medium with 0.25% glucose, 0.1%
72
Jalandoni-Buan et al.: Congo Red Decolorizing
Bacteria Identification
yeast extract, 0.05% potassium nitrate and 1% Congo
red. For anaerobic condition, samples were enriched in
Venoject tubes with cotton plug and after inoculation
the cotton plug was replaced with rubber stopper and
incubated statically at 28º to 30ºC. For aerobic condition,
the media without potassium nitrate were used and cotton
plugged flasks containing the cultures were incubated
under shaking condition (150 rpm) at room temperature.
Putatively decolorizing cultures were further enriched by
transferring aliquots of the enriched cultures into fresh
media and processed similarly. The final enriched culture
was analyzed for Congo red and nitrate colorimetrically.
Potentially decolorizing cultures were plated onto same
medium for monoculture isolation. Pure cultures and
consortia were confirmed for decolorization activities
using the procedure as described above. Nitrate and
nitrite were analyzed spectrophotometrically (Gerhardt
et al. 1994).
Congo red decolorization analysis
Pure cultures isolated from the consortia were
tested for their ability to decolorize Congo red by
spectrophotometric analysis (Decena and Barraquio
2004). A 1.5 ml aliquot of the decolorized culture broth
was placed in Eppendorf tubes and centrifuged at 14,000
rpm for 3 minutes. The supernatant was recovered
and analyzed spectrophotometrically at a wavelength
corresponding to the maximum absorbance of the dye
which is 530 nm. The uninoculated medium was used
as control and the medium without dye was used as
blank. A standard curve of known concentrations (0 to
20 mM) of pure and crude Congo red was prepared with
the medium as diluent. The efficiency of the isolates to
degrade/decolorize Congo red (Decena and Barraquio
2004) was expressed as:
% Efficiency of degradation/decolorization
= Initial conc. of dye – residual conc. of dye x100
Initial conc. of dye
Toxicity of decolorized culture medium
Toxicity of decolorized culture medium was tested on
prokaryotic cells (Pseudomonas aeruginosa UPCC 1244
and Staphylococcus aureus UPCC 1143, Gram negative
and Gram positive bacteria, respectively) and eukaryotic
cells (Saccharomyces cerevisiae UPCC 2113, a yeast;
Oryza sativa NSIC RC112, rice; and Vigna radiata, mung
bean). Decolorized culture medium was centrifuged
at 10,000 rpm for 30 minutes at 4˚C and the resulting
supernatant was filter sterilized using 0.22 µm pore filter.
Philippine Journal of Science
Vol. 139 No. 1, June 2010
For the microbial toxicity test, semisolid nutrient
agar (NA) and Sabouraud dextrose agar (SDA) were
inoculated with bacteria and yeast, respectively and then
pour plated. Four treatments were tested: decolorized
medium on the 7 th day of incubation, decolorized
medium on the 14th day of incubation, NA/SDA (negative
control) and NA/SDA with 1% Congo red (positive
control). Aliquots from the four treatments were streaked
in three separate parallel lines on the inoculated NA and
SDA, and observed after 24 and 48 h incubation for
inhibition of growth of bacteria and yeast.
Rice and mungbeans were used for the plant toxicity test.
Rice seeds were manually dehulled then both rice and
mungbean seeds were surface sterilized using chemical
disinfection in 95% ethanol for five minutes followed by
5% sodium hypochlorite for 1 h. These were washed five
times with sterile tap water then germinated in tryptone-
glucose-yeast extract (TGYE) agar plates for two to three
days in the dark to check for bacterial contamination. Rice
seedlings with approximately 1-1.5 cm shoots and 2-4 cm
shoot/root lengths were used for the experiments.
The roots of selected rice and mungbean seedlings
were immersed overnight in the following treatments:
decolorized medium on the 7 th day of incubation,
decolorized medium on the 14th day of incubation, Fahreus
medium (Fahreus 1957) as negative control and 1%
Congo red solution (positive control). Seedlings were then
placed on top of sterile screens placed in tubes containing
0.4% agar and modified Fahreus medium. The tubes
were then covered with sterile transparent autoclavable
plastic laminates. The seedlings were incubated for 12h
in artificial (fluorescent) light and 12h darkness for a
maximum of 21 days or until the shoots have grown tall
enough that they bend when the cover is touched. After
incubation, the plants were carefully uprooted from the
media and the heights and dry weights were determined.
Heights were measured from the point separating the root
and shoot to the highest point or apex of the plant shoot.
Dry weights were determined by drying whole plants in
an oven at 50˚C until constant weight was obtained.
Characterization and identification of Congo red
decolorizing monocultures
The purity of the isolates was checked by streaking on
nutrient agar plates several times. They were also checked
for Congo red decolorization.
The 24-hour old cells were subjected to Gram staining
(Doetsch 1981), KOH (Arthi et al. 2003), catalase
(Gerhardt 1994) and modified oxidative/fermentative
(Holding and Colee 1971) tests.
Jalandoni-Buan et al.: Congo Red Decolorizing
Bacteria Identification
Motility of the isolates grown in nutrient agar slants for 24
h at 30ºC was observed using the hanging drop method
(Holding and Colee 1971). Similarly grown isolates were
also subjected to transmission electron microscopy (TEM)
at the Electron Microscopy Service Laboratory of the
National Institute of Molecular Biology and Biotechnology
(BIOTECH), University of the Philippines Los Baños.
To test for denitrification, the method of Holding and Colee
(1971), was modified using 10 times diluted nutrient broth in
semisolid agar (3g/L) with 1g/L KNO3 and stab inoculated
with the samples. Ten (10) times diluted nutrient broth with
1g/L KNO3 was also inoculated with the samples, then an
inverted Durham tube was placed inside the tube, covered
with rubber stopper, to trap the gas that was produced.
Chemosystematic tests using API 20 (Biomereux,
Scientific Biotech Specialties, Inc., 1007 Metrostar Ave.,
Makati City) and BIOLOG (Dakila Trading, Inc., 208 Pilar
St., Mandaluyong City) were done to further characterize
the isolates.
For genotypic characterization, the 24 h-old monocultures
grown at 30ºC in nutrient agar slants, were sent to
Microbial ID (MIDI), USA for fatty acid profiling and16S
rRNA sequencing. Only three (3) of the four (4) CRDB
isolated were sent to MIDI.
RESULTS AND DISCUSSION
Isolation of Congo red decolorizing bacteria
From the enriched samples, twenty (20) consortia were
observed to decolorize 1% Congo red in less than two
weeks with nitrate, organic compound, or oxygen (low
level) as electron acceptors but not with oxygen (high
level or under vigorous shaking). Four (4) denitrifying
and four (4) fermentative consortia were then selected
for monoculture isolation and quantification of Congo
red decolorization. From the eight (8) selected consortia,
IRRI-1 (fermentative) and S22 (denitrifying) were selected
for further studies based on their high decolorization
potential, their ability to effect decolorization in larger
vessels and their ability to maintain decolorization activity
after repeated transfers. Both IRRI-1 and S22 showed
almost complete decolorization of 1% Congo red in 7 d.
After decolorization, the members of the consortia were
plated out in nutrient agar plates to identify the decolorizing
member of each consortium. The best monocultures isolated,
based on their high decolorization potential, were SB13B
(fermentative, nitrate reducing), SB12D (fermentative),
IRRI-1C (fermentative) and S22B (denitrifying). Figure 1
shows Congo red decolorization of monocultures.
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Philippine Journal of Science Jalandoni-Buan et al.: Congo Red Decolorizing
Vol. 139 No. 1, June 2010 Bacteria Identification

Toxicity of decolorized culture medium

Figure 1. Congo red decolorization of monocultures IRRI-1C (A)
and S22-B (B) in big tubes after 14 days of incubation.
CTRL is the uninoculated medium (negative control).
The rate of decolorization of the monocultures after
purification was observed to be slower than that of the
consortia. The monocultures decolorized Congo red after
2 weeks of incubation (SB13B, SB12D and IRRI-1C) and
The growths of bacteria and yeast were not inhibited by
both the consortium-decolorized medium and monoculture
decolorized medium as shown by the absence of zones of
inhibition on the 24 to 48 h-old lawn of microorganisms.
This indicates that bacterial decolorization of Congo red
did not result in metabolic products more toxic than the
dye. The undegraded Congo red (positive control) also
did not affect the growth of the microbes. Thus, these
microbes have an inherent resistance to the toxic effects
of the undegraded Congo red. This inherent resistance to
diazo dye may not hold true, however, for all microbes
since some might still be sensitive to this toxic dye.
The effects of undegraded Congo red and the decolorized
culture medium on the plants, however, were different
from that on bacteria and yeast. Figure 2A shows that rice
seedlings soaked in denitrifying consortium-decolorized
medium obtained on the 14th day of incubation (T2) have
higher plant shoots compared with seedlings exposed to
decolorized filtrate on the 7th day of incubation (T1), in
Fahreus medium (T3), and 1% Congo red solution (T4).
Correspondingly, higher dry weight of rice seedlings also
resulted.
21.000

after one (1) month of incubation in the case of S22B. The

19.500

18.000

consortia, on the other hand, decolorized Congo red after

16.500

15.000
Height, cm

only a week of incubation (Table 1). Blackening of the 13.500

12.000

rubber stopper was observed in consortium S22 together 10.500

9.000

with rotten egg odor indicating the possible presence 7.500

6.000

of a sulfate reducing organism in the mixed bacterial 4.500

culture. The presence of sulfate reducing bacteria in the

3.000

1.500

consortium could have hastened the rate of decolorization

0.000
T1 T2 T3 T4

of the dye. Van der Zee et al. (2003) reported that the Treatment
sulphide produced by a sulfate reducing bacteria can Figure 2A. Effect of denitrifying consortium-decolorized medium on
growth of rice shoot. The bars are the averages of three
reduce azo bonds.
replicates from each treatment, and error bars indicate +
standard deviations from the means. Treatments used
were: T1 – Denitrifying consortium-decolorized medium
Table 1. Rate of decomposition by the microbial consortia and on 7th day of incubation, T2- Denitrifying consortium
resulting monocultures. decolorized medium on 14th day of incubation, T3-
Mean number of Fahreus medium (negative control) and T4 – 1% Congo
Congo red decolorizing red solution (positive control).
days for complete Efficiency of Congo
consortia and selected
decolorization red decolorization
monocultures
(n=3)
Overnight soaking of selected rice seedlings in nitrate
Consortium: IRRI-1 7 94.69% reducing monoculture decolorized medium obtained
Consortium: S22 7 93.20%
on the 7th day of incubation however, resulted in the
highest plant shoots compared with seedlings soaked in
Monoculture: IRRI-1C 14 92% decolorized medium derived on the 14th day of incubation,
Monoculture: S22B 30 96%
Fahreus medium and 1% Congo red solution. Rice
seedlings soaked in 1% Congo red solution resulted in
Monoculture: SB13B 14 97% the lowest shoot heights (Fig 2B) and lowest dry weights.
Monoculture: SB12D 14 96% The height of mung bean shoots exposed to denitrifying
n = no. of replications consortium-decolorized (Fig 2C) and nitrate-reducing

74
 Philippine Journal of Science Jalandoni-Buan et al.: Congo Red Decolorizing
Vol. 139 No. 1, June 2010 Bacteria Identification

16.500 18.000
16.500
15.000
15.000
13.500 13.500

Height, cm
12.000 12.000
10.500
10.000
9.000
Height, cm

9.000 7.500
7.500 6.000
4.500
6.000
3.000
4.500 1.500
3.000 0.000
T1 T2 T3 T4
1.500
Treatment
0.000
T1 T2 T3 T4
Figure 2D. Effect of nitrate-reducing monocultures-decolorized
Treatment
medium on growth of mung bean shoot. The bars are
Figure 2B. Effect of nitrate-reducing-decolorized medium on the averages of three replicates from each treatment,
growth of rice shoot. The bars are the averages of three and error bars indicate + standard deviations from
replicates from each treatment, and error bars indicate the means. Treatments used were: T1 – Nitrate-
+ standard deviations from the means. Treatments used reducing monoculture-decolorized medium on 7th day
were: T1 – Nitrate-reducing monoculture-decolorized of incubation, T2- Nitrate-reducing monoculture-
medium on 7th day of incubation, T2- Nitrate-reducing decolorized medium on 14 th day of incubation,
monoculture-decolorized medium on 14th day of T3- Fahreus medium (negative control) and T4 – 1%
incubation, T3- Fahreus medium (negative control) Congo red solution (positive control).
and T4 – 1% Congo red solution (positive control).

19.500 bacterial decolorization metabolites slightly toxic to both

18.000
16.500 rice and mung beans. Many bacteria reduce azo bonds to
15.000
13.500 the corresponding amines. Several aromatic amines can
be degraded under aerobic condition (van der Zee 2002).
Height, cm

12.000
10.500
9.000
7.500
6.000
4.500
3.000
1.500
0.000
T1 T2 T3 T4
Treatment
Figure 2C. Effect of denitrifying consortium-decolorized medium
on growth of mung bean shoot. The bars are the
averages of three replicates from each treatment,
and error bars indicate + standard deviations from
the means. Treatments used were: T1 – Denitrifying
consortium-decolorized medium on 7 th day of
incubation, T2- Denitrifying consortium decolorized
medium on 14 th day of incubation, T3- Fahreus
medium (negative control) and T4 – 1% Congo red
solution (positive control).
monocultures-decolorized medium (Fig 2D) obtained
on the 7th day and on the 14th day of incubation were
relatively similar to that of seedlings exposed to Fahreus
medium but higher than that of seedlings soaked in 1%
Congo red solution.
Based on the results of the toxicity of the decolorized
culture medium and Congo red towards plants, it can
be deduced that the undegraded Congo red, is toxic to
both rice and mung beans as evidenced by the stunting
of plant growth and low dry weights. The effects of
the denitrifying consortium decolorized medium and
nitrate reducing monoculture decolorized medium on
plant growth may be attributed to the presence of small
amounts of still undegraded Congo red and probably
The higher plant shoots over the negative control may be
attributed to the promotion of plant growth by the nutrients
formed during bacterial decolorization which were
imbibed by the seedlings. Therefore, the decolorization
of Congo red by the bacteria diminished the severe toxic
effect of the dye to the rice and mung bean plants.
Characterization of the Congo red decolorizing
bacteria
All the monocultures were Gram negative which were
confirmed by positive KOH string test. Under the light
microscope, SB13B, SB12D, and IRRI-1C were observed
to be rods that occur singly and in pairs while S22B was
short rods to cocci that occur singly and in pairs. Figure
3 showed TEM of SB13B as coccus. All the isolates
exhibited motility using the hanging drop method, while
under transmission electron microscope, only SB12D and
SB13B were observed to be flagellated indicating motility
(Fig.3A and B). Brownian movement due to the passive
motion of the cells being carried about by the currents
under the cover slip (Gerhardt et al. 1994) could have been
mistaken for the motility of IRRI-1C and S22B using the
hanging drop method, since both were not flagellated.
S22B produced gas in Cohen Bazire medium with Congo
red and potassium nitrate. Nitrate analysis on S22B
grown in nutrient broth with KNO3 was negative for both
nitrite and nitrate, indicating that nitrate was converted to
nitrogen gas. These results indicated that S22B is capable
of denitrification while degrading Congo red. SB13B also

75
Philippine Journal of Science Jalandoni-Buan et al.: Congo Red Decolorizing
Vol. 139 No. 1, June 2010 Bacteria Identification

A B

C D
Figure 3. Transmission electron micrographs of Congo red decolorizing bacteria showing their
morphology. Note the presence of flagella in (A) SB12D and (B) SB13B. No flagellation was
observed in (C) IRRI-1C and (D) S22B.

formed gas in the enriched medium but the bubble formed
was not due to denitrification but due to fermentation of
glucose present in the medium. Nitrate in the medium
was reduced only to nitrite.
Both S22B and SB13B decolorized Congo red in the
presence of nitrate as terminal electron acceptor. Congo
red decolorization occurred concurrently with nitrate
reduction at a ratio of 1.49 mM of nitrate consumed per
0.41 mM of Congo red degraded for SB13B and a ratio of
1.06 mM of nitrate consumed per 0.37 mM of Congo red
degraded for S22B (Table 2). In the study of Burland and
Edwards (1999), it was found that benzene biodegradation
can be linked to nitrate reduction via nitrite to nitrogen gas.
Identification of Congo red decolorizing bacteria
Strain SB13B was identified as Brevundimonas vesicularis
using Biolog. B. vesicularis is a non-lactose fermenting
bacilli which was previously named as Pseudomonas
vesiculare (Segers et al. 1994). Based on API 20 E, 16S
rRNA sequence and fatty acid analysis (Table 3), it was

Table 2. Amount of Congo red degraded and nitrate consumed by selected decolorizing and nitrate-reducing monocultures.
Congo red Congo red % Efficiency of Congo Nitrate Nitrate
Monocultures Conc. degraded red degradation/ Conc. Utilized
(mM) (mM)1 decolorization (mM) (mM)2

SB13B
0.4118
(putative assimilative 0.0254 94.19 1.6574 1.4918
nitrate reducing)

Control (Uninoculated 0.4372

- 3.1492 -
medium)

S22B
0.0245 0.3775 93.9 2.2707 1.0607
(denitrifying)

Control (Uninoculated
0.4020 - 3.3314 -
medium)
1
(mM)Congo red degraded = (mM) Uninoculated medium – (mM) decolorized medium
2
(mM)Nitrate utilized = (mM) Nitrate in uninoculated medium – (mM) Nitrate in decolorized medium

76
Philippine Journal of Science
Vol. 139 No. 1, June 2010
Table 3. Identity of monocultures based on biochemical and genotypic characteristics.
Monocultures API rapid kit
SB13B E. coli
SB12D Enterobacter cloacae
IRRI-1C Klebsiella oxytoca
S22B Alcaligenes xylosoxidans
identified as E. coli. There have been reports on the ability
of E. coli to reduce azo dyes (Banat et al. 1996). E.coli
UPCC 1195, which was used in this study as reference
strain, was able to decolorize Congo red under nitrate
reducing condition. SB13B was able to reduce nitrate to
a form that was either assimilated or dissimilated by the
cell. E. coli reduces nitrate to NH3 which is assimilated
by the cell (Robertson and Kuenen 1992).
Strain SB12D was identified as Enterobacter cloacae
using API 20 E. No definite identification was generated
using the Biolog system. Fatty acid analysis revealed
that it is similar to E. coli with a similarity index of
0.757 although it was also similar with E. cloacae with
an index of 0.635. The 16S rRNA sequencing revealed
that it is closely related to Enterobacter dissolvens
(Table 3) but is clustered with E. cloacae. Although
the four methods of identification revealed different
identities of SB12D (Table 3), these methods were in
agreement that it belonged to the genus Enterobacter.
Strain S22B was identified as Pseudomonas citronellolis
based on Biolog ID system and 16S rRNA sequence
analysis. Earlier work placed P. citronellolis in the P.
aeruginosa group by 16S rRNA (Anzai et al. 2000) Fatty
acid analysis revealed that it is similar with Pseudomonas
aeruginosa . However, using API 20 NE it was identified
as Alcaligenes xylosoxidans. A wild-type isolate
Pseudomonas luteola was able to decolorize reactive
black B while P. aeruginosa NBAR12 was capable of
decolorizing 12 different dyes (Yeh and Shang 2004).
Pseudomonas aeruginosa was also reported to decolorize
certain azo dyes (Banat et al. 1996). Many Pseudomonas
strains were also found to be capable of denitrification
(Robertson and Kuenen 1992).
Strain IRRI-1C was identified as Klebsiella oxytoca
using API 20 E while it was not identified using Biolog.
Two (2) consortia and four (4) monocultures isolated
from the consortia were evaluated for their Congo red
decolorization potential. The rate of decolorization by the
Jalandoni-Buan et al.: Congo Red Decolorizing
Bacteria Identification
BIOLOG Fatty Acid Analysis 16S rRNA sequence
Brevundimonas
E. coli E. coli
vesicularis
Enterobacter
No ID E. coli
dissolvens
No ID Not determined Not determined
Pseudomonas Pseudomonas Pseudomonas
citronellolis aeruginosa citronellolis
consortia was faster (1 week) than that of the monocultures
(2 weeks to 1 month). Toxicity tests of consortium- and
monoculture-decolorized media and undegraded Congo
red revealed that the test microorganisms used, bacteria
and yeast, were resistant to the toxic effects of Congo red.
Toxicity tests on plants, however, showed that undegraded
Congo red was toxic to both rice and mungbean seedlings
and that the toxic effect on plants was reduced by
decolorization. The monocultures SB13B and S22B
decolorized Congo red concurrently with nitrate reduction.
The 16S rRNA sequencing revealed identities of SB13B
as E. coli, SB12D as Enterobacter dissolvens and S22B as
Pseudomonas citronellolis. API 20E identified IRRI-1C
as Klebsiella oxytoca.
ACKNOWLEDGEMENT
The authors acknowledge the financial support from the
Natural Sciences Research Institute, College of Science,
University of the Philippines Diliman, Philippine Council
for Agriculture, Forestry and Natural Resources Research
and Development, the Philippine Council for Advanced
Science and Technology Research and Development and
the Philippine Society for Microbiology, Inc.
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