Full Length Research Paper - IJSR · PDF fileChatterjee et al. Isolation and Characterization...

International Journal of Scientific Research in Environmental Sciences, 4(1), pp. 0001-0011, 2016

Available online at http://www.ijsrpub.com/ijsres

ISSN: 2322-4983; ©2016; Author(s) retain the copyright of this article

http://dx.doi.org/10.12983/ijsres-2016-p0001-0011

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Full Length Research Paper

Isolation and Characterization of Arsenite Tolerant Bacterial Strains from

Contaminated Water of West Bengal, India

Sabyasachi Chatterjee1*, Yogesh Bhai Patel1, Sajjan Rajpoot1, Sonam Rani1, Naba Kumar Mondal2*

1Department of Biotechnology, The University of Burdwan, Burdwan, West Bengal, India

2Department of Environmental Science, The University of Burdwan, Burdwan, West Bengal, India

*Corresponding author: [email protected], [email protected]; Cell: +919434545694, Fax: (0342) 2634200

Received 22 November 2015; Accepted 24 January 2016

Abstract. Arsenic contamination in groundwater of West Bengal, India is a serious environmental issue which is likely cause

diseases to the population dwelling in those areas. However, many studies shown that microorganisms have developed some

mechanism to resist themselves from arsenic compound. The objective of this study was to isolate arsenite resistant bacterial

strains from arsenic contaminated groundwater collected from West Bengal and to find the arsenic bioremediation efficiency

of most effective isolated strains. We cultured, identified and characterized two arsenite resistant bacterial strains (named as

ADSY K and ADSY R). Isolates were subjected for various biochemical test, co-resistant test with other heavy metals and

growth pattern observed in different growth parameters. These strains were closely related to various species of Bacillus and

Pseudomonas based on their 16S rRNA gene sequences and phylogenetic tree. Extracellular protein (soluble protein) profiling

through SDS-PAGE shown up and down regulation in presence of arsenite. Total arsenite estimation was done in different cell

extract as a bioremediation approach. The bacterial isolates (ADSY K and ADSY R) can be exploited for bioremediation of

arsenic containing wastes.

Keywords: Arsenite, Groundwater, Bacteria, Bioremediation, 16S rRNA gene sequence, Phylogenetic tree

1. INTRODUCTION

Exposure to heavy metals such as lead, cadmium,

mercury and arsenic is a major health hazard (Selvi et

al., 2014). Among these, arsenic (As) is a well known

human carcinogen and it is widely distributed in food,

water, soils, and air (Liao et al., 2011). It is a

ubiquitous element of both natural and anthropogenic

origin and is often responsible for contaminating

water supplies. However, Arsenic contamination is

most severe and unprecedented across a 0.173 million

square kilometer-geographical region in West Bengal,

India, where 36 million people are at risk for As

exposure (BGS and DPHE, 2001). Arsenic is

frequently found in nature as trivalent arseniteAs(III)

and pentavalent arsenate As(V) . Although both

As(III) and As(V) are toxic, As(III) is relatively more

toxic than As(V) (Munawar et al., 2012). At present,

even though As-rich groundwater has not been used

for drinking, it is still extensively used for irrigation,

aquaculture and industrial purposes (Kar et al., 2013).

Agricultural soil acts as a principal sink of As through

irrigation of cropland, and most of the arsenical

residues have low solubility and low volatility,

generally accumulating in the top soil layers. Topsoil

thus contaminated with As may have influence on the

entry of As into the food chain (Das et al., 2013).

Acute exposure to high levels of inorganic arsenic by

humans can be fatal while acute exposure to lower

levels can result in vomiting, decreased production of

red and white blood cells, abnormal heart rhythm, and

damage to blood vessels (Selvi et al., 2014). Due to

its ability to induce chromosomal aberration during

DNA replication, As is considered as a human

carcinogen and a potential mutagenic agent (Wang et

al., 2001). Moreover, previous literature also

highlighted that arsenic can interferes with the DNA

repair system, signal transduction pathways and

inhibits many enzymatic activities and also damages

respiratory, digestive, and circulatory as well as

nervous system (Rehman et al., 2010).

Therefore, removal of arsenic from environment is

of great significance to local agriculture and the

population. There are many conventional methods

such as chemical precipitation, chemical oxidation or

reduction, ion exchange, filtration, electrochemical

treatment; reverse osmosis, membrane technologies

and evaporation recovery are available for removal of

heavy metals from industrial effluent. However, all

the above mentioned processes may be ineffective or

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Chatterjee et al.

Isolation and Characterization of Arsenite Tolerant Bacterial Strains from Contaminated Water of West Bengal, India

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extremely expensive especially when the metals in

solution are in the higher range (1-100 mg/L).

Therefore, it is extremely important to develop an

innovative, low cost and eco-friendly method for

removal of toxic heavy metal ions from the water and

wastewater (Lou and Chin, 2008). A wide variety of

microorganisms are capable of growth in the presence

of heavy metal ions and tolerates high concentrations

(Gaballa and Helmann, 2003; Rehman et al., 2007).

Very recently Pal et al. (2014) reported one gram-

positive, nonpigmented, rod-shaped fluoride-tolerant

bacterial strain which tolerated more than 1500 ppm

of fluoride in brain-heart infusion agar medium.

However, arsenic resistant bacterial strain from West

Bengal is very rare in previous literature. Major

portion of West Bengal is arsenic affected. From this

back drop, present study was conducted to isolate and

characterize the arsenic tolerant bacterial strain from

arsenic contaminated water of West Bengal.

1. Materials and Methods

2.1 Site Description, Water Sampling and

Chemical Analysis

Water samples (0–15 cm) were collected from the

rice-growing areas of the Malda(Kaliachowk) and

Mushirdabad (Raghunathgang) Districts. The tubewell

water contaminated with arsenic and it exceed the

WHO permissible limits (0.01 mg/L). The

physicochemical properties of the groundwater

including pH (Jackson, 1967), conductivity, available

N (Subbiah and Asija, 1956), K (Brown and Warncke,

1934) and P (Olsen, 1954) were determined using

standard protocols. Total As (Sparks et al. 2006) and

NaHCO3-extractable As (Johnson and Barnard, 1979)

levels were determined using Spectrophotometer

(SDDC method).

2.2. Enrichment and Isolation of Arsenic Resistant

Bacteria

Samples were collected from arsenic prone area

[Raghunathganj, SAIL (Durgapur)] of west Bengal.

Special care was taken during sample collection. For

long term preservation, the samples were stored at

4°C. Serial dilution techniques were used for the

isolation of arsenic resistance bacteria. 10-3 dilution of

the sample was selected as inoculums. Different

concentrations of arsenic trioxide (0.1-10 mM) were

used to isolate arsenic resistant bacteria.

Approximately 108 isolates were found in the plate

containing 0.5mM arsenic trioxide after 48 hours of

incubation at 37°C. Among all, two dominant colonies

were further selected for future experimental analyses

and named as ADSY K,ADSY R respectively.

Arsenic-contaminated water (1 ml) was suspended in

Luria Bertani (LB) medium supplemented with 1 mM

of As(III) and incubated at 30°C for 48 hours

(Kinegam et al., 2008). Approximately 0.1mL of

enriched culture was plated on 1mM of As(III)

containing LB agar and incubated at 30°C for 24 hrs.

After incubation different colonies were formed on

the plates. Two dominant colonies were chosen for

further experimental analyses and named as ADSYK

and ADSYR.

2.3. Biochemical Analysis

The two dominant isolated colonies were subjected

for several biochemical analyses to characterize the

nature of the strains following “Bergey’s Manual of

Determinative Bacteriology”. The amylase test,

protease test, oxidase test, VP test, indole test and

catalase test were performed by using standard

protocols.

2.4. Scanning Electron Micrograph Study

Observation of ADSY R and ADSY R strain both in

arsenite treated and untreated condition was done

under Scanning Electron Microscope (SEM) at the

Centre of Scanning Electron Microscopy of Burdwan

University on HITACHI-S-530 operating at an

accelerating voltage of 20 kV.

2.5. Growth Parameters

2.5.1 Effect of Temperature on the Isolates

The isolated microorganisms were grown in LB broth

at different temperatures: 10℃, 20℃, 30℃, 37℃ and

40℃ for 24 hours on an orbital shaker at 120 rpm.

After incubation period, growth was monitored in

spectrophotometer (UV-Vis 1700 Pharmaspec,

Shimadzu) at 620 nm.

2.5.2. Effect of pH on the Isolates


at different pH: 5.0-8.0 for 24 hours at 37℃ on an

orbital shaker at 120 rpm. After incubation period,

growth was monitored in spectrophotometer (UV-Vis

1700 Pharmaspec, Shimadzu) at 620 nm.


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Fig. 1a: Phylogenetic tree for ADSYK

Fig. 1b: Phylogenetic tree for ADSY R

Fig. 1c: Virtual Restriction sites of 16S rDNA sequence of ADSY K strain

Chatterjee et al.


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2.5.3. Effect of Osmotic Pressure on the Isolates


at different salt (NaCl) concentration: 1M, 2M, 3M,

4M and 5M for 24 hours at 370C on an orbital shaker

at 120 rpm. After incubation dry weight of biomass

was measured.

2.6. Co-resistance of Isolates

Co resistance property to the other heavy metals were

also checked by growing them in LB broth

supplemented with 0.1mM, 0.5mM and 1mM of lead

(Pb) and mercury (Hg) and incubated at 37˚C for 24

hours on an orbital shaker at 120 rpm. After

incubation dry weight of biomass was measured.

2.7. Agar Cup Assay Method for Antibiotic

Sensitivity

Agar cup assay was followed to determine the

antibiotic sensitivity of the isolates. Chloramphenicol,

ampicillin, and tetracycline at various concentrations

were added to the cups of the agar plate. Arsenic

resistant isolates were spread throughout the plate.

After 24 hours of incubation, zone of clearance was

measured.

Fig. 2: Agarose Gel Electrophoresis

2.8. Protein Extraction and Its Profiling by SDS-

PAGE

To compare the extracellular proteins expression in

the stressed i.e. Arsenite and normal condition

(control), proteins were extracted and SDS-PAGE was

done. For protein isolation, 48 hours incubated pure.

Cultures [ADSYR and ADSYK] respectively at 37°C,

30°C were used to harvest the cells at 5000 rpm for 10

minutes. The supernatant contains soluble proteins

which were then dialyzed and separated by Sodium

Dodecyl Sulfate-Poly-Acryl amide gel electrophoresis

(SDS-PAGE).


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Fig. 3: SDS-PAGE of extracellular protein of ADSY K and ADSY R

Table 1: Physical and Chemical Properties of Water Sample

Parameters Tested water quality WHO recommended

value

ISI recommended value

pH 7.68 ± 0.21 6.5 t0 8.5 6.5 to 8.5

Conductivity(S/cm) 740 ± 1.05 600 -

TSS (mg/L) 30.0 ± 0.41

TDS (mg/L) 205 ± 0.11 500 500

DO (mg/L) 6.10 ± 0.03

COD (mg/L) 2.11 ± 0.71 10.0 -

Nitrate (mg/L) 3.11 ± 0.31 40.0 45

Chloride (mg/L) 11.75 ± 0.19 250

Sodium (mg/L) 2.5 ± 0.11 200 200

Potassium (mg/L)

7.32 ± 0.01 50 50

Phosphate (mg/L) 1.72 ± 0.21 0.02

Arsenic (mg/L) 4.51 0.05 0.05

Iron (mg/L) 0.163 0.30 0.30

2.9. Genomic DNA Extraction and Sequencing

The genomic DNA were extracted by standard

method with some modifications (Pitcher et al. 1989)

and confirmed through 0.8% Agarose Gel

Electrophoresis. The extracted DNA samples were

used to amplify the 16S rRNA gene using Universal

primer (27F and 1492R) and Taq polymerase in PCR

(Thermo HBSP02220) and further sequencing was

done at Xcelris genomics (ABI 3730xl genetic

analyzer).

Chatterjee et al.


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Table 2: Biochemical test of the isolated bacteria

Biochemical test reagents sample

ADSY R

sample

ADSY K

Amylase Test

Starch Agar, Gram’s iodine +ve +ve

Catalase Test 3% hydrogen peroxide +ve +ve

VP Test

MR-VP Broth, Baritt’s

reagent -ve -ve

Protease Test Gelatin agar medium -ve -ve

Indole Test

Tryptophan water, kovacs

reagent -ve -ve

Oxidase Test Glucose Tryptopan media -ve -ve

2.10. Bioinformatics Study

The 16S rRNA gene sequences were used for

determining the isolated strains and to draw the

phylogenetic tree of the isolated strain through

bioinformatics analysis. The sequences were also used

to find out the particular restriction sites by using

NEB cutter V2.0 (http://nc2.neb.com/NEBcutter2/).

For similar sequence search, standard nucleotide

BLAST (https://blast.ncbi.nlm.nih.gov) was

performed and few similar sequences with low “e

value” were selected for multiple sequence alignment

(MSA) by using the CLUSTAL W tool

(http://www.ebi.ac.uk/Tools/msa/clustalw2/ ). Ten

multiply aligned sequences were further used for

phylogenetic tree construction using the MEGA 6

tools.

2.11. Arsenic Estimation from the Cell Culture

For total arsenite concentration in the cell culture was

estimated spectrophotometrically using (SDDC)

method ((PERKINELMER, FTIR, Model-RX1

Spectrometer, USA). three different cell suspensions

were prepared for arsenic measurement which were

cell supernatant, cell wash and cell lysate. On the

other hand, total arsenic content in water sample was

measured spectrophotometrically using

Silverdiethyldithiocarbamate (SDDC) method

(PERKINELMER, FTIR, Model-RX1 Spectrometer,

USA).

2. RESULTS AND DISCUSSIONS

3.1. Water Physical and Chemical Properties

Water containing elevated concentration of metal is

potential source of those metal tolerant bacteria. It is

because the environmental condition promotes

adaptation of those isolates in such environment

(Clausen et al., 2000). The physicochemical

characterization of experimental water is shown in

Table 1. From the Table 1, it is shown that pH ranges

from 7.39 to 7.76 and conductivity varies between

0.72 to 0.75 S/cm, respectively. Conductivity of

water sample was found in the range of 655-789

S/cm, which is much higher than the WHO

standards. Electrical conductivity is considered to be a

rapid and good measure of dissolved solids.

Conductivity is an important criterion in determining

the suitability of water for irrigation. Chemical

oxygen demand is a valuable water quality parameter.

COD is a measure of the oxygen equivalent of the

organic matter in a water sample that is susceptible to

oxidation by a strong chemical oxidant, such as

dichromate. It is an index of organic content of water

because the most common substance oxidized by

dissolve oxygen in water is organic matter having

biological origin i.e. dead plant and animal wastes

(Singh, 2002). COD values convey the amount of

dissolved oxidisable organic matter including the non-

biodegradable matters present in it. The value of COD

was found in the range of 1.8 mg/l to 2.5 mg/l. Its

value is much lower than the permissible limit

http://nc2.neb.com/NEBcutter2/

https://blast.ncbi.nlm.nih.gov/

http://www.ebi.ac.uk/Tools/msa/clustalw2/


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prescribed by WHO. However, other parameters such

as available phosphorus, nitrogen, sodium, potassium,

chloride, TSS, TDS, and DO clearly revealed that the

water is moderate quality. The results of the study

revealed that all bacterial isolates did not have the

same degree of tolerance to As toxicity. This might be

due to developing of As tolerance and resistant ability

of the inherent individual soil microorganisms (Smith

et al., 1998). Exposure of indigenous bacteria to

gradient of As concentrations during enrichment for

isolation might have developed metal resistance

systems for protecting sensitive cellular components

(Pattamaporn et al., 2008).

Fig. 4: Scanning Electron Micrograph of isolated bacteria

3.2. Biochemical Analysis and Co-Resistant

Activity of Isolated Bacteria

From the various biochemical analyses it was found

that the isolated strain ADSYR and ADSYK showed

positive results for catalase test but negative for

protease and oxidase test (Table 2). Optimum

temperature has been found at 37˚C for ADSYR and

30˚C for ADSYK whereas optimum pH was found to

be 6.0 and 8.0 for ADSYR and ADSYK, respectively.

Both the isolates ADSY R and ADSY K were also

found co-resistant in lead (Pb) and mercury (Hg) in all

the tested concentrations (Table 3).

3.3. SEM Study of Isolated Bacteria

From the SEM study, it has been observed that the

ADSY bacterial samples are rod shaped bacterium

and their cell size increased when they were grown in

presence of arsenite (Fig. 4a & Fig-4b).Enlargement

of the cell size may have ultimately lead to bursting of

the cell because of variation in osmotic pressure

between the cytosol and the outer environment. It had

been reported earlier that SEM analysis of

Pseudomonas aeruginosa strain MCCB 102 showed

an increase in cell size due to Cd and lead

accumulation in the cell wall and along the external

cell surfaces (Zolgharnein et al., 2010). The effect of

metal on cell morphology was also demonstrated by

transmission electron microscopy analysis of

P. putida strain 62BN which showed an increase in

size of the cells grown in the presence of Cd and also

showed intracellular and periplasmic accumulation of

metal in the cells (Rani et al., 2009).

3.4. Genomic and Proteomic Study

Genomic study has revealed that the presence of high

molecular weight genomic DNA (Approx. 23kb) (Fig.

2) in ADSY R and ADSY K. But proteomic study of

ADSY R and ADSY K confirmed the phenomenon of

time specific regulation of protein expression. From

figure 3, it can be observed that some of genes

became up regulated in stress condition (As treatment)

and showed protein bands in spots 5, 6, and 7 whereas

some of them became down regulated in same

condition and has not expressed as in corresponding

spots of 1, 2, and 3 in treated condition in ADSY R. In

case of ADSY K that the lower expression of a protein

in spot 4 has been observed. So, ultimately it can be

inferred from this analysis that protein profile of

ADSY R and ADSY K were different in stress

condition. This is perhaps due to the nullifying the

stress of arsenic. Almost similar results reported by

Chatterjee et al.


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Bachate et al. (2009) in their study where they

highlighted that Phylogenetically diverse arsenic-

resistant bacteria present in agricultural soils of

Bangladesh is capable of reducing arsenate to arsenite

under aerobic conditions.

Fig. 5: Level of arsenic in supernatant, cell wash and cell lysate of isolated bacteria

3.5. Bioinformatics Study

From the bioinformatics analysis using BLAST tool, it

has been found that isolates ADSY K (Fig.1a) and

ADSY R (Fig.1b ) has sequence homology with

Pseudomonas sp. and Bacillus sp. respectively. In

multiple sequence alignment, ADSY R has maximum

alignment with most of strain of Bacillus megaterium

and Bacillus aryabhatti, while ADSY K (Fig.1c,

Fig.1d) has maximum sequence alignment with

Pseudomonasaeruginosaand bears most of their

morphological characteristics and property.

3.6. Antibiotic Sensitivity

Antibiotic sensitivity test results are given in Table 4.

From the Table 4 it is clear that ADSY K is much

more antibiotic resistant than ADSY R in lower

concentration. However, in higher dose ADSY R

showed much more antibiotic resistance against all the

tested antibiotics (Chloramphenicol, Ampicillin and

Tetracycline). Almost similar results were reported by

Dey et al. (2015).

3.7. Arsenic Concentration in Cell Culture

Level of arsenic was estimated from supernatant, cell

wash and cell lysate of bacterial isolate ADSY K and

ADSY R. Results revealed that bacterial isolate

ADSY K accumulates higher level of total arsenite in

both supernatant and cell wash (Fig. 5). However, in

cell lysate the bacterial isolate ADSY R accumulate

higher level of arsenite compared to the isolate ADSY

K (Fig. 5). Almost similar bioaccumulation of arsenic

from culture media by bacterial isolate strain AGH-21

was reported by Majumder et al. (2013). Their

isolated strain showed highest sequence similarity

(98%) with Bacillus sp. Apart from the arsenic

resistant bacteria, literature also cited the Cd(II),

Zn(II), Ni(II), Ag(I), Cu(II) resistant bacteria also

(Pepi et al., 2007; Moore et al., 2005).

Most arsenic resistant bacteria are separated from

arsenic-rich environments. In natural environments,

the number of arsenite resistant bacteria is less than

arsenate resistant bacteria (Jackson et al., 2005).

Among the isolated resistant strains from

contaminated water, two strains demonstrated

dramatic resistance to arsenite. These arsenite-

resistant strains were probably Pseudomonas sp. and

Bacillus sp. However, these two species showed

unequal bioremediation efficiency. Abu-shnab et al.

(2003) showed that in a contaminated soil, about

11.1% of isolated bacteria were resistant to As with

the MIC of 20 mM/L. High levels of soil- metal

concentration can lead to achieving such a high MIC

in resistant strains.


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Table 3: Biochemical tests result positive (+)ve and (-)ve indicates efficient growth and no response respectively. Sl

.

N

o.

Bacterial

Isolates

Growth

Parameters

Observed value

1 ADSY R NaCl Conc.

(M)

1M 2M 3M 4M 5M

OD at 620 nm 0.04 0.02 0.01 0.01 0.00

Co-resistance

to Pb

0.1mM 0.5mM 1mM

Dry weight

mass 0.04 0.03 0.01

Co-resistance

to Hg

0.1mM 0.5mM 1mM

Dry weight

mass 0.02 0.02 0.00

Temperature

(0C)

10 20 30 37 40

OD at 620 nm 0.04 0.05 0.19 0.17 0.28

pH 5.0 5.5 6.0 6.5 7.0 7.5 8.0 8.5 9.0

OD at 620 nm 0.1 0.70 0.77 1.37 0.74 0.93 0.77 0.54 0.56

.

2 ADSY K NaCl Conc.

(M)

1M 2M 3M 4M 5M

OD at 620 nm 0.07 0.05 0.04 0.02 0.01

Co-resistance

to Pb

0.1mM 0.5mM 1mM

Dry weight

mass 0.05 0.01 0.00

Co-resistance

to Hg

0.1mM 0.5mM 1mM

Dry weight

mass 0.05 0.01 0.00

Temperature

(0C)

10 20 30 37 40

OD at 620 nm 0.02 0.05 0.14 0.15 0.16

pH 5.0 5.5 6.0 6.5 7.0 7.5 8.0 8.5 9.0

OD at 620 nm 0.01 0.52 1.30 0.91 0.73 1.15 0.71 0.94 1.05

Table 4: Growth Parameters of ADSY R and ADSY K Bacterial Strain ADSY R ADSY K

Antibiotic Concentration (µg/ml) 200 400 600 800 1000 200 400 600 800 1000

Antibiotics Used

(Zone of Clearance)

Chloramphenicol 0.0 1.1 1.7 2.0 2.5 2.7 2.3 2.1 1.8 1.2 Ampicillin 0.0 1.1 1.3 1.7 2.0 0.0 0.0 0.0 0.0 0.0

Tetracycline 0.0 1.0 1.6 2.5 2.7 1.5 1.2 1.5 1.3 1.1

Also Chitpirom et al. (2009), in Thailand, isolated

arsenic-resistant bacteria from tannery effluent and

agricultural soils that were belonged to Klebsiella,

Pseudomonas, Comamonas and Enterobacter with the

MIC of 40 mM (arsenite) and 400 mM (arsenate).

Pepi et al. (2007) isolated 3 arsenic resistant genera

(Aeromonas, Bacillus and Pseudomonas) from

contaminated sediments with the MIC of 16.66 mM

(arsenite) and 133.47 mM (arsenate). They also

concluded that these bacteria are suitable for arsenic

bioremediation in contaminated sediments. In a study

by Luis et al. (2006) in Spain with the aim of

biological removal of arsenic,

Corynebacteriumglutamicum with over 60 mM

arsenite resistance identified as one of the most

tolerant species to arsenic. These results are in

agreement with our findings but our isolates could

tolerate the higher concentration of arsenite that was

related to high level of arsenite in water.

4. CONCLUSIONS

The naturally occurring arsenic-resistant isolates are

more environmentally acceptable and safe for

detoxification of arsenic. Hence, isolation of such

arsenic-resistant species has considerable ecological

advantage. However, characterization of arsenic

metabolizing genes is required for their successful

exploitation in in situ arsenic bioremediation. The

present study concentrated only on the assay of water

Chatterjee et al.


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for identification of microbes, and it is recommended

that a further study be conducted to find out the

contribution of microorganisms in the enhancement of

the arsenic level in the study area. In order to fully

appreciate the arsenic remediation potential of these

two selected isolates, further studies should be

focused with the biomass of these two isolates as

adsorbing material, optimizing the bioadsorption

conditions, the possible recycling of this bioadsorbing

material, and optimizing of the adsorbing and

desorbing conditions.

Acknowledgements

Authors gratefully acknowledge to the staff members

of both the Department of Biotechnology and

Department of Environmental Science for their

unconditional academic help. Moreover authors also

like to extend their gratitude to all senior professors of

the University of Burdwan, Burdwan.

REFERENCES

Abu-Shanab RAI, Delorme TA, Angle JS, Chaney

RL, Ghanem K, Moawad H, Ghozlan HA

(2003). Phenotypic characterization of microbes

in rhizosphere of Alyssum murale. Int. J.

Phyto., 5:367-380.

Bachate SP, Cavalca L, Andreoni V, (2009). Arsenic-

resistant bacteria isolated from agricultural soils

of Bangladesh and characterization of arsenate-

reducing strains. J. Appl. Microbiol. 107: 145–

156.

Chitpirom K, Akaracharanya A, Tanasupawat S,

Leepipatpibooim N, Woong KK (2009).

Isolation and characterization of arsenic

resistant bacteria from tannery wastes and

agricultural soils in Thailand. J. Ecol. Environ.

Microbiol. 59(4):649-656.

Das S, Jean J-S, Kar S, Liu CC (2013). Changes in

bacterial community structure and abundance in

agricultural soils under varying levels of arsenic

contamination, Geomicrob. J., 30:635–644.

Dey U, Das K, Roy P, Chatterjee SN, Mondal NK

(2015). Searching of Microbial Agent for

Bioremediation of Arsenic. Int. J. Ext. Res.,

5:60-64.

Gaballa A, Helmann JD (2003). Bacillus subtilis

CPx-type ATPases: characterization of Cd, Zn,

Co and Cu efflux systems. Biometals 16:497-

505.

Liao VH-C, Chu Yu-Ju, Su Yu-Chen, Hsiao S-Y,

Wei C-C, Liu C-W., Liao C-M, Shen W-C,

Chang Fi-John (2011). Arsenite-oxidizing and

arsenate-reducing bacteria associated with

arsenic-rich groundwater in Taiwan. J.

Contamin. Hydroly. 123:20–29.

Lou JC, Lin YC (2008). Assessing the feasibility of

wastewater recycling and treatment efficiency

of wastewater treatment units. Environ Monit

Assess 137:471-479.

Jackson CR, Dugas SL, Harrison KG (2005).

Enumeration and characterization of arsenat-

resistant bacteria in arsenic free soils. J. Soil

Biol. Biochem., 37(12):2319-2322.

Jackson ML (1967). Soil Chemical analysis. Prentice

Hall of India, Pvt. Ltd., New Delhi : 498.

Kinegam S, Yingprasertchai T, Tanasupawat S,

Leepipatpiboom N, Akaracharanya A, Kim K-

W (2008). Isolation and characterization of

arsenite-oxidizing bacteria from arsenic-

contaminated soil in Thailand. World Journal of

Microbiology and Biotechnology 24:3091-96.

Kar S, Das S, Jean J-S, Chakraborty S, Liu C-C

(2013). Arsenic in the water–soil–plant system

and the potential health risks in the coastal part

of Chianan Plain, Southwestern Taiwan. J.

Asian. Earth Sci., 77:295–302.

Luis M, Ordonez E, Letek M, Gil J (2006).

Corynebacteriumglutamicumas a model

bacterium for bioremediation of arsenic. Int. J.

Microbiol., 9: 207-215.

Munawar S, Susann V, Kamrun Z, Anne_CS, Chad

S, Jana S, Michael (2012). New

clusters of arsenite oxidase and unusual bacterial

groups in enrichments from arsenic-

contaminated soil. Archiv. Microbiol.

194(7):623-635.

Majumder A, Ghosh S, Saha N, Kole SC, Sarkar S

(2013). Arsenic accumulating bacteria isolated

from soil from possible application in

bioremediation. J. Eviron. Bio., 34: 841-846.

Moore CM, Gaballa A, Hui M, Ye RW, Helmann JD

(2005). Genetic and

physiological responses of Bacillus subtilis to metal

ion stress. Mol Microbiol 9:207-215.

Olsen SR, Cole CV, Watambe FS, Dean LA (1954).

Estimation of available

phosphorus in soil by extraction with NaHCO3,

U.S.D.A. Ciraza (Quoted from, Method of soil

Analysis, C.A. Black 2 nd ed.) 1165 Am. Soc.

Agron., Inc. Medision Wisconsin, USA. (1035).

Pal KC, Mondal NK, Chatterjee S, Ghosh TS, Datta

JK (2014). Characterization of fluoride tolerant

halophilic Bacillus flexus NM25(HQ875778)

isolated from fluoride affected soil in Birbhum

district, West Bengal, India. Environ. Assess.

186:699-709.

Pitcher DG, Saunders A, Owe RJ (1989). Rapid

extraction of bacterial genomic DNA with


11

guanidiumthiocyanate. Lett. Appl. Microbiol.,

8: 151–156.

Pattamaporn A, Poonsuk P, Vorasan S (2008).

Isolation of As-tolerant bacteria from As-

contaminated soil. Songklanakarian Sci.

Technol. 30:95-102.

Pepi M, Volterrani M, Renzi M, Marvasi M,

Gasperini S, Franchi E, Focardi SE (2007).

Arsenic-resistant bacteria isolated from

contaminated sediments of the Orbetello

Lagoon, Italy, and their characterization. J.

Appl. Microbiol., 103(6): 2299-2308.

Rani R, Souche Y, Goel R (2009). Comparative

assessment of in-situ bioremediation potential

of cadmium resistant Pseudomonas putida 62

BN and alkalophilic Pseudomonas monteilli

97AN strains on soyabean. Int. Biodeterio.

Biodeg. 63:62–66.

Rehman A, Butt AS, Hasnain S (2010). Isolation and

characterization of arsenite oxidizing

Pseudomonas lubricans and its potential use in

bioremediation of wastewater. African J.

Biotechnol., 9:1493-1498.

Rehman A, Ali A, Muneer B, Shakoori A (2007).

Resistance and biosorption of mercury by

bacteria isolated from industrial effluents. Pak J

Zool 39:137-150.

Smith AH, Goycolea M, Haque R, Biggs ML (1998).

Marked increase in bladder and lung cancer

mortality in a region of northern Chile due to

As in drinking water. Ame. J. Epidemiol.,

147:660-669.

Selvi MS, Sasikumar S, Gomathi S, Rjkumar P,

Sasikumar P, Sadasivam SG (2014). Isolation

and characterization of arsenic resistant bacteria

from agricultural soil, and their potential for

arsenic bioremediation. Int. J. Agril. Pol. Res.,

2(11):393-405.

Singh R, Adholeya A (2002). Biodiversity of AMF

and agricultural potential II: The impact of

agromic practices. Mycorrhiza News, 13(4):22 -

24.

Wang TS, Hsu TY, Chung CH, Wang AS, Bau DT,

Jan KY (2001). Arsenite induces oxidative

DNA adducts and DNA-protein crosslinks in

mammalian cells. Free Radical Biol. Med.,

31:321-330.

Zolgharnein H, Karami K, Mazaheri AM, Dadolahi

SA (2010). Investigation of heavy metals

biosorption on Pseudomonas aeruginosa strain

MCCB 102 isolated from the persian gulf.

Asian J. Biotechnol., 2:99–109.

Chatterjee et al.


Dr. Sabyasachi Chatterjee

Research Area: Bioremediation of heavy metals, Phytoremediation, Medicinal Plants, Enzymology of

beneficial microbes, Arctic microbes.

Ph.D. in Botany, 1st class 1st in M.Sc Microbiology, presently faculty of the Biotechnology

Department, The University of Burdwan from 2007, lecturer in the field of Microbiology in Asansol

Girls’ College from 2002-2007.One of the CO-PI in a DST (SERC) sanctioned project on Arctic

microflora and production of their industrial enzymes. Reviewer for Bioremediation Journal (Taylor &

Francis), Journal of Medicine and Medical Science, African Journal of Microbiology (Academic

Publisher), Agricultural Science Research Journal. Editorial Board member for the Journal Trends in

Life Science, Trends in Parasitology Research, Journal of Medicinal and Aromatic plants (OMICS Group), Academia Journal

of Biotechnology, European Journal of Medicinal Plants, Annual Review and Research in Biology, Ecology and

Environmental Sciences, International Journal of Microbiology Research, Research in Biology. Published 24 research papers,

review articles in different International and National Journals and in Books. Life member of Association of Microbiologist of

India.

Yogesh Bhai Patel awarded with Master degree in Biorechnology (2012) from Dept. of Biotechnology,

The University of Burdwan. He has actively worked on current study as a part of his M.Sc. Project work

and remains exporing the possibility of efficient Bioremediation. He is currently working as Scientist in

R & D (Bioanalytical), Biocon Research Limited, where he engages with developing & validiating

Pharmacokinetic and Immunogenicity assays for the MAb based biotherapeutics.

Mr. SajjanRajpoot, I have received Bachelor of Technology (B.Tech) degree in Biotechnology from

EIILM University, Sikkim in year 2012. I then qualified JNU-CBEE(Jawaharlal Nehru University-

Combined Biotechnology Entrance Examination) session 2013-2015 and selected for DBT, Govt. of

India supported M.Sc. Biotechnology program in Department of Biotechnology, The University of

Burdwan, Burdwan, West Bengal. I did 6 months of in-house dissertation workon arsenic and received

the Master of Science (M.Sc.) degree in Biotechnology from Department of Biotechnology, The

University of Burdwan, Burdwanin 2015. In year 2015, I qualified GATE BT and also selected for West

Bengal Biotech Development Corporation’s (WBBDC) RISE program for internship at IIT-KGP,

Kharagpur. The area of my research interest is Microbial and parasitic diseases, Molecular Biology of

cell and Immunology.

Ms. Sonam Ran, I Pursued B.Sc. Biotechnology (session 2009-12) from G.L.A College, N.P.U,

Daltonganj, Jharkhand, and M.Sc.Biotechnology (through JNU-CBEE session 2013-15) ofDBT, Govt.

of India supported program from The University of Burdwan, Burdwan, West Bengal. Worked on

Arsenic Bioremediation during the course of M.Sc. as the dissertation project. After M.Sc. course,

selected for West Bengal Biotech Development Corporation’s (WBBDC) RISE program for internship

at BOSE Institute, Kolkata Areas of research apart from Ecotoxicology are study of cell signaling,

recognition and response in toxic environment.

Dr Naba Kumar Mondal presently holding the position as Assistant professor in the department of

Environmental Science, The University of Burdwan, India. Dr Mondal has experience more than 16

years of teaching and research in both Education and Environmental Science (masters degree). His

research interest includes: Pure Science:Adsorption Chemistry, Nutrient dynamics, indoor pollution, soil

Chemistry, Plant Physiology, Social Science: corporal punishment, development of teaching

methodology, noise and its impact on school children etc. Dr Mondal also published more than 170

research papers in reputed International and National Journals and four (06) Ph.D. scholars (upto

January’ 2015) and has been serving as an guest Editor and reviewer in many prestigious International

Journals.

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