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Research Article Production of Extracellular Polymeric Substances by Halophilic Bacteria of Solar Salterns Jhuma Biswas and A. K. Paul Microbiology Laboratory, Department of Botany, University of Calcutta, 35 Ballygunge Circular Road, Kolkata 700 019, India Correspondence should be addressed to A. K. Paul; amalk [email protected] Received 13 May 2014; Accepted 15 July 2014; Published 6 August 2014 Academic Editor: Yinguang Chen Copyright © 2014 J. Biswas and A. K. Paul. is is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. Moderately halophilic aerobic bacteria were isolated from 31 soil and 18 water samples collected from multipond solar salterns of Gujarat, Orissa, and West Bengal, India. A total of 587 bacterial isolates with distinct morphological features were obtained from these samples following dilution and plating on MH agar medium supplemented with NaCl. e isolates were screened for growth associated extracellular polymeric substances (EPS) production in MY medium under batch culture. In all, 20 isolates were selected as potent ones producing more than 1 g/L of EPS. ese EPS producing isolates were characterized in detail for their morphological, physiological, and biochemical features and tentatively identified as members belonging to the genera Halomonas, Salinicoccus, Bacillus, Aidingimonas, Alteromonas, and Chromohalobacter. Apart from EPS production, these isolates also hold promise towards the production of various biomolecules of industrial importance. 1. Introduction Multipond solar salterns used for industrial production of marine salts by evaporation of sea water represent hyper- saline environments which are popular habitats for studying halophilic bacteria and have great potential towards indus- trial and biotechnological applications [1, 2]. e diversity of halophilic bacteria so far isolated and characterized is categorized into four different classes according to NaCl requirement for their growth and includes slight halophiles, moderate halophiles, extreme halophiles, and border line halophiles. e halotolerant bacteria on the other hand do not require NaCl for their growth but can tolerate a high salinity [3–5]. Halophilic diversity of solar salterns has been studied quite extensively across the globe and reviewed by several authors [6–8]. However, only very few studies have been made on the halophilic bacterial community in coastal solar salterns of India [9, 10], which deserves special attention for exploration and commercial exploitation of these microbial resources. Extracellular polymeric substances (EPS) are one of the industrially important compounds produced by a wide variety of marine microorganisms. Due to growing biotech- nological interest [11], production of bacterial EPS has become an attractive field of research. EPS is used as thick- eners, emulsifiers, and suspending agents in food, pharma- ceuticals, and petroleum industries. ey are also used as adhesives in detergents, textiles, papers, paints, and beverages industries. Moreover, EPS are used as metal removers and bioabsorbers in oil recovery, mining, and petroleum indus- tries [12]. During the course of extensive search for new strains pro- ducing extracellular polymeric substances (EPS) in their nat- ural hypersaline environments, large numbers of halophilic bacteria and archaea have been established as EPS pro- ducers. Most of them belong to the genera Haloferax, Haloarcula, Halococcus, Natronococcus, and Halobacterium [13, 14]. Nevertheless, the common halophilic EPS pro- ducing bacteria belong to the genus Halomonas, most importantly H. maura [8], H. eurihalina [15], H. ventosae, and H. anticariensis [16]. Exopolysaccharides synthesized by Halomonas strains unusually have high sulphate content and a significant amount of uronic acids determining their good gelifying properties. Moreover, recent reports have Hindawi Publishing Corporation Chinese Journal of Biology Volume 2014, Article ID 205731, 12 pages http://dx.doi.org/10.1155/2014/205731

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Page 1: Research Article Production of Extracellular …downloads.hindawi.com/archive/2014/205731.pdfResearch Article Production of Extracellular Polymeric Substances by Halophilic Bacteria

Research ArticleProduction of Extracellular Polymeric Substances by HalophilicBacteria of Solar Salterns

Jhuma Biswas and A K Paul

Microbiology Laboratory Department of Botany University of Calcutta 35 Ballygunge Circular Road Kolkata 700 019 India

Correspondence should be addressed to A K Paul amalk paulyahoocoin

Received 13 May 2014 Accepted 15 July 2014 Published 6 August 2014

Academic Editor Yinguang Chen

Copyright copy 2014 J Biswas and A K Paul This is an open access article distributed under the Creative Commons AttributionLicense which permits unrestricted use distribution and reproduction in any medium provided the original work is properlycited

Moderately halophilic aerobic bacteria were isolated from 31 soil and 18 water samples collected from multipond solar salterns ofGujarat Orissa and West Bengal India A total of 587 bacterial isolates with distinct morphological features were obtained fromthese samples following dilution and plating on MH agar medium supplemented with NaCl The isolates were screened for growthassociated extracellular polymeric substances (EPS) production inMYmedium under batch culture In all 20 isolates were selectedas potent ones producingmore than 1 gL of EPSThese EPS producing isolates were characterized in detail for their morphologicalphysiological and biochemical features and tentatively identified as members belonging to the genera Halomonas SalinicoccusBacillus Aidingimonas Alteromonas and Chromohalobacter Apart from EPS production these isolates also hold promise towardsthe production of various biomolecules of industrial importance

1 Introduction

Multipond solar salterns used for industrial production ofmarine salts by evaporation of sea water represent hyper-saline environments which are popular habitats for studyinghalophilic bacteria and have great potential towards indus-trial and biotechnological applications [1 2] The diversityof halophilic bacteria so far isolated and characterized iscategorized into four different classes according to NaClrequirement for their growth and includes slight halophilesmoderate halophiles extreme halophiles and border linehalophilesThehalotolerant bacteria on the other handdonotrequire NaCl for their growth but can tolerate a high salinity[3ndash5] Halophilic diversity of solar salterns has been studiedquite extensively across the globe and reviewed by severalauthors [6ndash8] However only very few studies have beenmade on the halophilic bacterial community in coastal solarsalterns of India [9 10] which deserves special attention forexploration and commercial exploitation of these microbialresources

Extracellular polymeric substances (EPS) are one ofthe industrially important compounds produced by a wide

variety of marine microorganisms Due to growing biotech-nological interest [11] production of bacterial EPS hasbecome an attractive field of research EPS is used as thick-eners emulsifiers and suspending agents in food pharma-ceuticals and petroleum industries They are also used asadhesives in detergents textiles papers paints and beveragesindustries Moreover EPS are used as metal removers andbioabsorbers in oil recovery mining and petroleum indus-tries [12]

During the course of extensive search for new strains pro-ducing extracellular polymeric substances (EPS) in their nat-ural hypersaline environments large numbers of halophilicbacteria and archaea have been established as EPS pro-ducers Most of them belong to the genera HaloferaxHaloarcula Halococcus Natronococcus and Halobacterium[13 14] Nevertheless the common halophilic EPS pro-ducing bacteria belong to the genus Halomonas mostimportantly H maura [8] H eurihalina [15] H ventosaeand H anticariensis [16] Exopolysaccharides synthesizedby Halomonas strains unusually have high sulphate contentand a significant amount of uronic acids determining theirgood gelifying properties Moreover recent reports have

Hindawi Publishing CorporationChinese Journal of BiologyVolume 2014 Article ID 205731 12 pageshttpdxdoiorg1011552014205731

2 Chinese Journal of Biology

also established that halophilic EPS producers belong to thegamma-proteobacteria (Idiomarina andAlteromonas) as wellas alpha-proteobacteria (Salipiger mucosus and Palleroniamarisminoris) [17 18]

This study is focused on the isolation of halophilicbacteria from water and soil samples of some selectedsolar salterns located in the states of Gujarat Orissa andWest Bengal India and evaluation of their EPS productionefficiency under laboratory conditions Attempts have alsobeen made on the tentative identification of some potent EPSproducing halophilic isolates based on their morphologicaland physiobiochemical features

2 Materials and Methods

21 Collection of Samples Soil and water samples frommultipond solar salterns situated along the coast of GujaratOrissa and West Bengal India were collected in sterilepolypropylene containers and stored at 4∘C until used forisolation of halophilic bacteria A total of 12 soil and 7water samples were collected from Jogrinar (23∘131015840N and69∘5910158405110158401015840 E) Khari Rohar (23∘410158401610158401015840N and 70∘910158403710158401015840 E)Kandla (22∘591015840N and 70∘131015840 E) and Albert Victor Port(21∘010158405810158401015840N and 71∘3210158405410158401015840 E) of Gujarat Similarly 10 soil and5 water samples were collected from Surala (19∘8410158407310158401015840N and84∘651015840 E) and Humma (19∘261015840N and 85∘51015840 E) the two majorsolar salterns of Orissa In West Bengal the sampling siteswere located at Dadanpatrabar (22∘261015840N and 87∘201015840 E) andBaksal (22∘11015840N and 87∘671015840 E) of East Midnapur and a total of9 soil and 6 water samples were collected

22 Isolation of Halophilic Bacteria Aerobic heterotrophicandhalophilic bacteria of soil andwater sampleswere isolatedby serial dilution and plating on MH agar medium [19]supplemented with different concentrations of NaCl Themedium contained (gL) yeast extract 10 protease peptone5 glucose 1 NaCl 100 MgCl

26H2O 7 MgSO

47H2O 96

CaCl22H2O 036 KCl 2 NaHCO

3006 and NaBr 0026

(pH 72) The plates were incubated at 37∘C for 3 to 5 daysand bacterial colonies with distinctmorphologywere isolatedin pure form and maintained on slopes of the same mediumTotal bacterial counts were expressed as colony forming units(cfu)mL and g of water and soil respectively

23 Screening of Halophilic Bacteria for EPS ProductionTo evaluate the EPS production capability the bacterialisolates were grown in MY medium [20] supplemented with5 NaCl for 12 days at 32∘C under continuous shaking(120 rpm) The medium contained (gL) NaCl 50 MgCl

2

6H2O 9 MgSO

47H2O13 CaCl

22H2O 02 KCl 13

NaHCO3 005 NaBr 015 FeCl

36H2O 0005 glucose 10

yeast extract 3 malt extract 3 and protease peptone 5(pH 72) The EPS from the growing culture was isolatedusing the method as described by Quesada et al [15] Theculture was centrifuged (at 10000timesg for 10min) the EPSin the supernatant was precipitated with chilled ethanol

and recovered by centrifugation (12000timesg for 10min) andwashed with chilled 70 ethanol The washed precipitatewas collected by centrifugation dissolved in known volumeof distilled water and used for quantification and chemicalanalysis

24 Chemical Analysis of EPS Total carbohydrate contentof the EPS was estimated following the method of Duboiset al [21] To 1 mL of dissolved EPS sample 05mL of 5phenol and 35mL of concentrated sulfuric acid were addedand incubated at 30ndash40∘C for 10ndash20 minutes in hot waterbath Absorbance was read at 490 nm and the amount ofcarbohydrate was determined from the calibration curveprepared using glucose as standard

Protein content of the EPS was determined followingthe Folin phenol method of Lowry et al [22] To 1mL ofEPS sample 5mL of alkaline solution and 05mL of Folinphenol reagent were added and incubated for 30 minutes atroom temperature in dark The absorbance was measured at670 nm and the concentration was read from the calibrationcurve prepared by using bovine serum albumin (BSA) as thestandard

25 Characterization and Identification of Selected BacterialIsolates The selected bacterial isolates were characterizedmorphologically and physiobiochemically following stan-dard microbiological methods as described by Gerhardt et al[23] To determine the antibiotic sensitivity pattern of theseisolates the antibiotic impregnated discs (Himedia 6mmdia) were placed on MH agar plates seeded with respectivebacterial isolates The plates were incubated for 24 h at32∘C and diameter of inhibition zones was measured to thenearest mm Production of acids from sugars by the bacterialisolates was tested on phenol redmedium supplemented with1 carbon source Characteristics of the bacterial isolateswere compared with those described in Bergeyrsquos Manual ofSystematic Bacteriology [24] and that of Mata et al [25] fordetermination of taxonomic identity

3 Results

31 Isolation of Halophilic Bacteria A total of 31 soil and18 water samples collected from 8 different sites spreadover the states of Gujarat Orissa and West Bengal Indiawere analyzed for the aerobic heterotrophic and halophilicbacteria following dilution and plating on MH agar mediumsupplemented with 5 10 and 15 NaCl The total bacterialpopulation of both soil and water samples as determinedby colony forming units varied considerably and declinedgradually irrespective of sampling sites with increasing NaClconcentration in the isolation medium (Tables 1 and 2)

A total of 587 halophilic and halotolerant hetero-trophic and aerobic bacterial isolates were obtainedin pure form The majority of these isolates (410)were derived from soil samples (Table 3) while177 were obtained from water samples (Table 4)Most of the isolates were Gram-negative motile aerobic

Chinese Journal of Biology 3

Table 1 Bacterial population of soil samples collected from solar salterns of Gujarat Orissa and West Bengal India

State Sampling sites Number of samplesBacterial count (cfug of soil times102)

NaCl () in medium5 10 15

Gujarat

Jogrinar 4 074ndash866 021ndash366 011ndash4Kandla Port 3 086ndash735 121ndash211 008ndash0Khari Rohar 3 239ndash768 012ndash178 047ndash966Victor Port 2 173ndash283 040ndash0 mdash

Orissa Humma 5 013ndash061 025ndash055 mdashSurala 5 1ndash366 007ndash023 mdash

West Bengal Baksal 4 2ndash390 020ndash040 020ndash0Dadanpatrabar 5 27ndash79 007ndash060 007ndash030

Table 2 Bacterial population of water samples collected from solar salterns of Gujarat Orissa and West Bengal India

States Sampling sites Number of samplesBacterial count (cfumL of water times102)

NaCl () in medium5 10 15

Gujarat

Jogrinar 2 028ndash556 172ndash0 040ndash0Kandla Port 2 053ndash540 016ndash11 066ndash0Khari Rohar 1 023ndash0 mdash mdashVictor Port 2 126ndash190 mdash mdash

Orissa Humma 3 023ndash23 mdash mdashSurala 2 027ndash120 083ndash0 mdash

West Bengal Baksal 3 110ndash136 013ndash06 010ndash037Dadanpatrabar 3 05ndash190 007ndash050 01-02

rods and produced white to cream colored circular smooth-edged colonies on MH agar In general the isolates werecapable of tolerating wide range of temperature and pH fortheir growth

32 Screening for EPS Production EPS producing abilityof these isolates was examined in batch culture using MYmedium supplemented with 5 NaCl and the soluble EPScontent of each of the isolates was evaluated in terms ofits carbohydrate content as determined by Dubois method[21] Among the 410 soil isolates majority (184) producedsoluble EPS ranging from 05 to 07 gL in terms of theircarbohydrate content while only 15 isolates produced morethan 1 gL of EPS (Table 5) On the contrary among the177 bacterial strains isolated from saline water samples only5 produced soluble EPS accounting for more than 1 gL ofcarbohydrate However majority (77) of them produced 05ndash07 gL of soluble EPS (Table 6)

In all 20 potent isolates producing gt1 gL of EPS werefurther allowed to grow in MY medium under continuousshaking (120 rpm) and the soluble EPS produced after 8days of growth was estimated following the method asdescribed above Kinetics of growth and EPS productionrevealed that EPS production by these isolates increasedwith biomass formation (Figure 1) and a few of the selectedisolates such as SUR202 SUR307 SUR310 JW307 and

JS904 appeared to be promising with an EPS yield of168ndash185 gL

33 Characterization and Identification of Selected BacterialIsolates Morphological and physiological studies revealed(Table 7) that these halophilic bacteria (with the exceptionof isolate SUR303) formed cream colored smooth colonieson MH agar medium Most of them were Gram-negativemotile rods only three isolates were Gram-positive one ofthem (isolate SUS303) was coccus while the isolate KW203was the only endospore former The isolates were capableof tolerating 15ndash20 NaCl in the medium and a pH of 5ndash11 Optimum growth was observed at a temperature of 32ndash37∘C but all were able to tolerate a temperature as high as40∘C

Analysis of biochemical characters (Table 8) showedthat the majority of these halophilic isolates gave negativeresponse to MR-VP tests and failed to produce extracellularenzymes like amylase cellulase pectinase inulinase gelati-nase lipase caseinase xylanase and urease None of theseisolates however were capable of producing H

2S lysine-

arginine- and ornithine decarboxylaseCarbon source utilization pattern (Table 9) of these

halophilic isolates varied considerably All the isolates wereable to utilize ribose fructose mannitol salicin cellobioseacetate benzoate and succinate and three of them (JS803

4 Chinese Journal of Biology

Table 3 Bacterial isolates obtained from soil samples of solar salterns of Gujarat Orissa and West Bengal India

States Sampling sites Number of samplesNumber of bacterial isolates

NaCl () in medium5 10 15

Gujarat

Jogrinar 4 44 23 4Kandla Port 3 19 12 4Khari Rohar 3 46 16 6Victor Port 2 20 5 mdash

Orissa Humma 5 48 14 mdashSurala 5 31 31 2

West Bengal Baksal 4 19 8 3Dadanpatrabar 5 29 16 10

Total 31 256 125 29

Table 4 Bacterial isolates obtained from water samples of solar salterns of Gujarat Orissa and West Bengal India

States Sampling sites Number of samplesNumber of bacterial isolates

NaCl () in medium5 10 15

Gujarat

Jogrinar 2 21 14 5Kandla Port 2 19 5 2Khari Rohar 1 1 3 mdashVictor Port 2 26 mdash 1

Orissa Humma 3 16 mdash mdashSurala 2 12 8 3

West Bengal Baksal 3 9 3 4Dadanpatrabar 3 17 4 4

Total 18 121 37 19

SUR301 and SUR307) appeared to be versatile in utilizingall 30 carbon sources Fermentation pattern of these strainsvaried remarkably the majority were unable to fermentmost of the carbon sources On the contrary fructosesorbitol and benzoate were fermented by most of the isolates(Table 9)

Sensitivity of these isolates to 22 different antibioticswas tested by disc-diffusion method (Table 10) and theantibiotic resistance index (ARI) was determined (Figure 2)Themajority of the isolates were sensitive to chloramphenicol(30 120583g) gentamycin (10 120583g) and norfloxacin (10 120583g) followedby ampicillin (10 120583g) Resistance to vancomycin (30120583g) fol-lowed by trimethoprim (30 120583g) was predominant amongstthe tested halophiles As judged by the ARI values the isolateSUR302 was the most resistant one (ARI = 065) followed bySUR301 KW203 and KW1805 while the lowest ARI (023)was indicated by the isolate JW307 which was most sensitiveto the tested antibiotics

The morphological physiological and biochemical char-acters including the carbon source utilization and fermen-tation patterns along with antibiotic susceptibility wereanalyzed and compared with the phenotypic characters ofhalophilic bacterial genera [5 18] so far reported According

to the phenotypic and biochemical characteristics 70 of theselected isolates (14) were tentatively identified asmembers ofthe genusHalomonasThe onlyGram-positive coccus (isolateSUR303) and the rod shaped endospore forming isolate(isolate KW203) were assigned to Salinicoccus and Bacillusrespectively Two of the isolates (KS1805 and SUR301) wereplaced in the genus Chromohalobacter while the remain-ing two (isolates JS504 and JS904) were included underAlteromonas and Aidingimonas respectively

4 Discussion

Halophiles have mainly been isolated from wide diversity ofenvironments such as saltern crystallizer ponds the DeadSea solar lakes and hypersaline lakes [26] Culture depen-dent diversity studies on halophiles have been made fromTunisian solar saltern [27 28] Tuzkoy salt mine [29] Sere-flikochisar Salt Lake [30] Kaldirim and Kayacik of Tuz Lake[31] Turkey Howz Soltan Lake Iran [32] and hypersalineenvironments in South Spain [33ndash36] In the Indian contextthe halophilic diversity studies have been restricted mainlyto the marine salterns of Bhavnagar [37ndash40] Lonar Lake[40] and Peninsular Coast [41 42]The present study reports

Chinese Journal of Biology 5

Table 5 Screening of halophilic bacterial isolates derived from soil samples for the production of extracellular polymeric substances

States Sampling sites Number of isolates Extracellular polymeric substances (gL)gt050 051ndash070 071ndash10 gt10

Gujarat

Jogrinar 71 18 42 7 4Kandla Port 35 13 13 7 2Khari Rohar 68 26 31 11 mdashVictor Port 25 20 3 2 mdash

Orissa Humma 62 22 37 3 mdashSurala 64 15 25 15 9

West Bengal Baksal 30 14 12 4 mdashDadanpatrabar 55 32 21 2 mdash

Total 410 160 184 51 15

Table 6 Screening of halophilic bacterial isolates derived from water samples for the production of extracellular polymeric substances

States Sampling sites Number of isolates Extracellular polymeric substances (gL)gt050 051ndash070 071ndash10 gt10

Gujarat

Jogrinar 40 26 9 3 2Kandla Port 26 13 11 1 1Khari Rohar 4 1 3 mdash mdashVictor Port 27 7 17 1 2

Orissa Humma 16 4 11 1 mdashSurala 23 8 6 9 mdash

West Bengal Baksal 16 7 6 3 mdashDadanpatrabar 25 8 14 3 mdash

Total 177 74 77 21 5

the distribution of halophilic and halotolerant bacterialcommunities in the inland multipond solar salterns spreadover the coasts of India Halophilic bacterial communitiesof 31 soil and 18 water samples from 8 different sites wereanalyzed by dilution and plating method (Tables 1 and 2) andprovide information regarding the availability anddiversity ofhalophilic bacteria in the solar saltern ponds Colony formingunits of soil and water samples revealed that soil samplesholdmore viable bacterial counts than water samples Ragha-van and Furtado [41] studied the occurrence of extremelyhalophilic archaea in sediments from the continental shelf ofwest coast of India and reported the presence of relatively lowaverage counts (7ndash5 times 103) of extreme halophiles in offshoresediments in contrast to the very high counts (105ndash109)of marine eubacteria Similar study by Joshi et al [40]reported that the total numbers of microorganisms in the soiland water samples were 102ndash106 cfug and 102ndash104 cfumLrespectively

A total of 587 halophilic and halotolerant bacterial strainswere isolated showing different degrees of NaCl tolerance(Tables 3 and 4) and supported the observations of Quesadaet al [35] andVentosa et al [34] However during the presentstudy we were unable to isolate extreme halophiles whichhave frequently been identified as the dominant phylotypes inhypersaline environments along with solar salterns of India

[39 41] Furthermore soil samples showed more bacterialdiversity than water ones which is in accordance with theobservations of Joshi et al [40]

During the course of screening of the moderatelyhalophilic isolates for EPS production (Tables 5 and 6) only20 isolates appeared to be promising with significant yield ofEPS (10ndash185 gL) (Figure 1) Joshi and coworkers [40] sim-ilarly screened 86 halophilic bacteria from Lonar Lake andreported Halomonas campisalis and Vagococcus carniphilusas potent EPS producers Similarly Nanjani and Soni [43]also isolated 73 halotolerant and halophilic bacteria from soilsamples of Veraval and Dwarka 23 of them produced EPSranging from 02 to 1060 gL In addition reports on EPSproduction by moderately halophilic bacteria of the genusHalomonas are not uncommon [8 15ndash17]

Attempts have been made to determine the taxonomicidentity of all 20 promising EPS producing isolates followingdetailed physiobiochemical characterization (Tables 7ndash9) andcomparison with Bergeyrsquos Manual of Systematic Bacteriology[24] and those of Mata et al [25]Themajority (14 isolates) ofthem were Gram-negative nonsporulating rods and capableof growing in 25ndash20NaCl similar to those ofHalomonas asreported by Quesada et al [36] Ghozlan et al [44] andMataet al [25] However the Gram-positive endospore formingisolate KW203 was assigned to the genus Bacillus

6 Chinese Journal of Biology

Table7Morph

ologicalandph

ysiologicalcharacteristicso

fthe

selected

EPSprod

ucinghaloph

ilicb

acteria

isolated

from

solarsalternso

fInd

ia

Character

Bacterialisolates

JS504

JS802

JS803

JS904

JW302

JW307

KS110

1KS

1805

KW203

VW301

Morph

ologicalcharacters

Colon

ymorph

olog

yCr

eamcirc

ular

Whitecirc

ular

Creamcirc

ular

Creamcirc

ular

Transparentcircular

Creamcirc

ular

Creamcirc

ular

Whitecirc

ular

Whitecirc

ular

Creamcirc

ular

Gram

nature

minus+

minusminus

minusminus

minusminus

+minus

Cellshape

Rod

Rod

Rod

Rod

Rod

Rod

Rod

Rod

Rod

Rod

Cell

arrang

ement

Sing

leSing

leSing

leSing

leSing

lechains

Sing

lechains

Sing

lechains

Sing

leSing

leclu

mped

Sing

lechains

Motility

++

++

++

minusminus

++

Pigm

entatio

nminus

minusminus

minusminus

minusminus

minusminus

minus

Endo

sporeformation

minusminus

minusminus

minusminus

minusminus

+minus

Physiologicalcharacters

pHrang

efor

grow

th5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

pHop

timum

forg

rowth

119

89

1111

118

89

Temprang

efor

grow

th(∘C)

27ndash4

027ndash4

027ndash4

027ndash4

027ndash4

027ndash4

022ndash4

027ndash4

027ndash4

022ndash4

0Tempop

timum

forg

rowth

(∘C)

3237

3732

3232

3232

3732

NaC

lrange

forg

rowth

()

0ndash15

0ndash20

0ndash15

0ndash20

0ndash15

0ndash20

0ndash20

25ndash15

25ndash15

0ndash20

NaC

loptim

umforg

rowth

()

510

105

105

575

105

Growth

onKingrsquosBmedium

++

++

++

++

++

Growth

onMcC

onky

agar

minusminus

minusminus

minusminus

minusminus

minusminus

Tentativeidentity

Alteromonas

Halom

onas

Halom

onas

Aidingim

onas

Halom

onas

Halom

onas

Halom

onas

Chromohalobacter

Bacillus

Halom

onas

Character

Bacterialisolates

VW402

SUR108

SUR3

01SU

R302

SUR3

03SU

R307

SUR3

09SU

R310

SUR2

02SU

R201

Morph

ologicalcharacters

Colon

ymorph

olog

yCr

eamcirc

ular

Creamcirc

ular

Whitegum

my

Transparentcircular

Flatreddish-pink

Creamcirc

ular

Creamcirc

ular

Creamcirc

ular

Creamcirc

ular

Creamcirc

ular

Gram

nature

minusminus

minusminus

+minus

minusminus

minusminus

Cellshape

Rod

Rod

Rod

Rod

Cocci

Rod

Rod

Rod

Rod

Rod

Cell

arrang

ement

Sing

leSing

leSing

leSing

leSing

leclu

mped

Sing

leSing

leSing

leSing

leSing

leMotility

+minus

minus+

minus+

minus+

minusminus

Pigm

entatio

nminus

minusminus

minusPink

minusminus

minusminus

minus

Endo

sporeformation

minusminus

minusminus

minusminus

minusminus

minusminus

Physiologicalcharacters

pHrang

efor

grow

th5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

pHop

timum

forg

rowth

97

77

117

77

79

Temprang

efor

grow

th(∘C)

27ndash4

027ndash4

027ndash4

032ndash4

532ndash4

522ndash4

032ndash4

022ndash4

522ndash4

532ndash4

5Tempop

timum

forg

rowth

(∘C)

3232

3732

3727

3227

2732

NaC

lrange

forg

rowth

()

0ndash15

0ndash15

25ndash15

25ndash15

25ndash15

0ndash20

0ndash20

0ndash20

0ndash20

0ndash15

NaC

loptim

umforg

rowth

()

525

575

525

7525

575

Growth

onKingrsquosBmedium

++

++

++

++

++

Growth

onMcC

onky

agar

minusminus

minusminus

minusminus

minusminus

minusminus

Tentativeidentity

Halom

onas

Halom

onas

Chromohalobacter

Halom

onas

Salin

icoccus

Halom

onas

Halom

onas

Halom

onas

Halom

onas

Halom

onas

Chinese Journal of Biology 7

Table8Biochemicalcharacteris

ticso

fthe

selected

EPSprod

ucinghaloph

ilicb

acteria

isolated

from

solarsalternso

fInd

ia

Character

Bacterialisolates

JS 504

JS 802

JS 803

JS 904

JW 302

JW 307

KS 1101

KS 1805

KW 203

VW 301

VW 402

SUR

108

SUR

301

SUR

302

SUR

303

SUR

307

SUR

309

SUR

310

SUR

202

SUR

201

Biochemicalcharacters

Voges-Proskauertest

minusminus

minusminus

minusminus

minusminus

+minus

minusminus

minusminus

minusminus

minusminus

minusminus

Citrateu

tilization

+minus

minusminus

minus+

minus+

minus+

++

+minus

minus+

++

++

Methylred

test

minusminus

minusminus

minusminus

minus+

+minus

minusminus

minusminus

minusminus

minusminus

minusminus

Prod

uctio

nof

Glutaminase

++

++

++

++

++

++

++

minus+

++

++

Asparaginase

++

++

++

++

++

++

++

minus+

++

++

Amylase

+minus

minusminus

minusminus

minusminus

+minus

minusminus

minusminus

minusminus

minusminus

minusminus

Cellulase

minusminus

minusminus

minusminus

minusminus

+minus

minusminus

minusminus

minusminus

minusminus

minusminus

Pectinase

+minus

minusminus

minusminus

minusminus

+minus

minusminus

minusminus

minusminus

minusminus

minusminus

Inulinase

+minus

minusminus

minusminus

minusminus

+minus

minusminus

minusminus

minusminus

minusminus

minusminus

Gelatinase

minusminus

minusminus

minusminus

minusminus

+minus

minusminus

minusminus

minusminus

minusminus

minusminus

Lipase

+minus

minusminus

minusminus

minus+

+minus

minusminus

+minus

minusminus

minusminus

minusminus

Xylanase

+minus

minusminus

minusminus

minusminus

+minus

minusminus

minusminus

minusminus

minusminus

minusminus

Caseinase

minusminus

minus+

minusminus

minusminus

minusminus

minusminus

minus+

minusminus

minusminus

minusminus

Urease

minusminus

minus+

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

Catalase

++

++

++

++

++

++

++

++

++

++

Nitrater

eductase

++

++

++

++

++

++

++

++

++

++

H2S

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

Lysin

edecarbo

xylase

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

Arginined

ecarbo

xylase

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

Ornith

ined

ecarbo

xylase

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

Indo

leminus

minusminus

minusminus

minusminus

+minus

minusminus

minus+

minus+

minusminus

minusminus

minus

Phenylalanined

eaminase

++

+minus

+minus

++

minus+

minus+

++

++

++

minus+

Tryptoph

andeam

inase

++

++

++

++

++

minusminus

++

++

++

+minus

8 Chinese Journal of Biology

Table9Ca

rbon

source

utilizatio

nandferm

entatio

nof

thes

elected

EPSprod

ucinghaloph

ilicb

acteria

isolated

from

samples

ofsolarsalternso

fInd

ia

Carbon

source

Bacterialisolates

JS 504

JS 802

JS 803

JS 904

JW 302

JW 307

KS 1101

KS 1805

KW 203

VW

301

VW

402

SUR

108

SUR

301

SUR

302

SUR

303

SUR

307

SUR

309

SUR

310

SUR

202

SUR

201

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

Glucose

++

++

+minusminus

++minus

+minus

+minus

++

++

+minus

+minus

+minus

+minusminus

++

++minus

+minus

+minusminusminusminusminus

Galactose

+minus

+minus

+minusminus

++minus

+minus

+minus

++

+minus

+minus

+minus

+minus

++minusminus

+minus

++

+minus

+minus

+minus

+minus

Arabino

seminusminusminusminus

+minusminusminusminusminus

+minus

+minus

++

++

+minus

+minus

+minus

+minusminus

++minus

+minus

+minusminusminus

+minus

+minus

Manno

se+minus

+minus

+minusminus

++minus

+minus

+minusminus

++

++minus

+minus

+minus

+minusminusminusminusminus

+minus

+minus

+minus

+minus

+minus

Rham

nose

+minus

+minus

+minus

+minusminusminusminusminusminusminus

++

+minusminusminus

+minus

+minus

++minus

++minus

+minusminusminusminusminusminusminus

+minus

Trehalose

+minus

+minus

+minusminus

+minus

+minusminusminusminus

+minus

++minusminusminusminus

+minus

+minusminus

++

++minusminusminusminusminusminusminus

+minus

Lactose

minusminusminusminus

+minusminusminus

+minus

+minus

+minus

+minus

+minus

+minus

+minusminusminus

+minus

++minusminus

+minusminusminusminusminus

+minus

+minus

Sucrose

+minus

+minus

+minus

++

++

+minus

+minus

+minus

++

+minus

+minusminusminus

+minus

++

+minus

+minus

+minus

+minusminusminus

+minus

Xylose

minusminusminusminus

+minus

+minusminusminus

+minus

++minus

+minusminus

+minus

+minus

+minus

+minusminus

+minus

++

+minusminusminusminus

+minusminusminus

Ribo

se+minus

++

++

++

+minus

+minus

+minus

++

++

+minus

+minus

+minus

++

++

++

+minus

+minus

+minus

+minus

+minus

Fructose

+minus

++

+minus

++

+minus

++

++

+minus

++

++

++

++

++

++

++

++

++

++

++

++

Maltose

minusminusminusminus

+minusminus

+minus

++minus

+minus

+minus

++

+minus

+minus

+minus

++minus

+minus

++minusminusminusminusminus

+minus

+minus

Glycine

minusminusminus

++minus

+minusminus

+minusminusminusminus

+minusminusminusminusminusminusminusminusminus

+minusminusminusminusminus

+minus

+minusminusminusminus

+minusminus

Glycerol

+minus

+minus

+minusminus

++

++minus

+minus

+minus

++

+minus

+minus

+minus

+minusminus

++

++minus

+minus

+minus

+minus

+minus

Adon

itol

+minus

+minus

+minus

+minusminusminus

+minus

+minus

+minus

+minus

+minus

+minusminusminus

+minus

++

+minus

+minus

+minus

+minusminusminusminusminus

Salicin

+minus

++

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

++

++

+minus

+minus

+minus

+minus

+minus

Meso-inosito

l+minusminusminus

+minus

+minus

+minus

+minus

++

+minus

++

+minus

+minusminusminus

+minusminus

++minus

+minus

+minus

+minus

+minus

+minus

Mannitol

+minus

+minus

+minus

++

+minus

+minus

+minus

+minus

++

+minus

+minus

+minus

+minus

+minus

++

+minus

+minus

+minus

+minus

+minus

Sorbito

l+

++

++minusminus

++

+minus

++

++

++

++

++

++

++

+minus

+minus

++

++minus

+minus

++

+minus

Raffino

seminusminus

++

++minusminus

+minus

+minus

+minus

+minus

++minusminusminusminusminusminusminusminus

++

++

+minus

+minus

+minus

+minus

+minus

Cellobiose

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

++

+minus

+minus

+minus

+minus

++

+minus

+minus

+minus

+minus

+minus

+minus

Melibioseminusminusminusminusminusminusminusminusminusminusminusminusminusminusminusminus

++minusminusminusminusminusminusminusminus

++minusminusminusminusminusminusminusminusminusminusminusminus

Na-acetate

+minus

+minus

+minus

+minus

++

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

++

++

+minus

+minus

+minus

+minus

+minus

Na-citrate

+minus

++

+minusminusminusminus

+minusminusminusminusminusminusminusminusminusminusminusminusminusminusminusminusminus

+minusminusminusminusminusminusminusminusminusminusminusminus

Na-fumarate

+minus

+minus

+minus

++

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minusminus

++minus

+minus

+minus

+minus

+minus

+minus

Na-form

ate

+minus

+minus

+minus

+minus

+minusminusminus

+minus

+minus

+minusminusminus

+minusminusminus

+minus

++minusminus

+minusminusminusminusminusminusminusminusminus

Na-benzoate

+minus

++

++

+minus

+minus

+minus

++

++

++

+minus

+minus

++

++

+minus

+minus

+minus

++

++

++

++

Na-malateminusminusminusminusminusminus

++

+minusminusminusminusminusminusminusminusminusminusminusminusminusminusminusminusminus

++

++minusminusminusminusminusminusminusminusminusminus

Na-prop

ionate

+minus

+minus

+minusminusminus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

++minus

++minus

+minus

+minus

+minus

+minus

Na-succinate

+minus

+minus

+minus

++

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

++

++

+minus

+minus

+minus

+minus

+minus

Uutilization

Fferm

entatio

n

Chinese Journal of Biology 9

Table10A

ntibiotic

sensitivitypatte

rnof

thes

electedEP

Sprod

ucinghaloph

ilicb

acteria

isolated

from

solarsalternso

fInd

ia

Antibiotics

Bacterialisolates

JS 504

JS 802

JS 803

JS 904

JW 302

JW 307

KS 1101

KS 1805

KW 203

VW 301

VW 402

SUR

108

SUR

301

SUR

302

SUR

303

SUR

307

SUR

309

SUR

310

SUR

202

SUR

201

Ampicillin(10120583

g)S

SR

SS

SS

RI

SS

SR

SS

SS

SS

SAmoxicillin

(30120583

g)S

SR

RS

SS

RS

SS

SR

RS

IR

SS

SBa

citracin

(10120583

g)S

SR

SS

SR

RS

IR

RR

RS

IR

RR

RCiprofl

oxacin

(30120583

g)I

II

RI

II

IR

II

II

IR

SR

II

ICh

lorte

tracyclin

e(30120583g)

SI

SS

IS

RR

SS

SS

RR

SS

II

IS

Chloramph

enicol(10120583

g)S

SS

SS

SS

SR

SS

SS

SS

SR

SS

SErythrom

ycin

(15120583

g)S

SI

SS

IS

II

II

II

SR

RI

IS

SFu

sidicacid

(30120583

g)I

RR

SS

IR

RS

RI

RR

RS

IR

II

RGentamycin

(10120583

g)S

SS

SS

SS

SR

SS

SS

SS

IS

SS

SKa

namycin

(30120583

g)I

IS

RS

II

SR

II

RI

RR

RI

RI

RMethicillin(5120583g)

SS

RS

IR

RR

RR

RR

RR

SR

RR

RR

Neomycin

(30120583

g)S

SI

RI

IR

II

IR

II

RR

RI

IR

RNorflo

xacin(30120583

g)S

SR

SS

SS

SI

SS

SS

SS

SS

SS

SNetillin

(10120583

g)R

SS

IR

SS

SR

RR

SS

RR

RR

RR

RNalidixicacid

(30120583

g)R

RS

RR

II

RR

II

RR

RR

SR

RR

RNovob

iosin

(30120583

g)I

RR

SR

SS

RS

SS

IR

SS

SI

SS

SPo

lymixin

B(50120583

g)R

RI

RI

IR

RR

RI

IR

RR

II

II

SPenicillinG(10120583

g)S

SR

IS

IR

RR

II

SR

RS

II

II

SStreptom

ycin

(10120583

g)S

SS

RS

RR

SR

RR

IS

RS

SR

II

RTetracyclin

e(30120583g)

RR

RR

RI

RR

SR

IR

RR

RS

RR

RI

Trim

etho

prim

e(30120583g)

SS

SS

IS

SI

SS

SS

IR

RS

SS

SS

Vancom

ycin

(30120583

g)R

RR

SR

RR

RS

RR

RR

RS

RR

RR

R

S=SensitiveR

=Re

sistantI

=Interm

ediate

10 Chinese Journal of Biology

0

1

2

3

4

Bacterial isolate

Gro

wth

OD

at540

nmto

tal E

PS (g

L)

OD at 540nmTotal EPS

JS504

JS802

JS803

JS904

JW302

JW307

KS1101

KS1805

KW203

VW301

VW402

SUR1

08

SUR3

01

SUR3

02

SUR3

03

SUR3

07

SUR3

09

SUR3

10

SUR2

02

SUR2

01

Figure 1 Screening of moderately halophilic bacteria isolatedfrom soil and water samples of solar salterns for production ofextracellular polymeric substances

0

02

04

06

08

1

Ant

ibio

tic re

sista

nce i

ndex

Bacterial isolate

JS504

JS802

JS803

JS904

JW302

JW307

KS1101

KS1805

KW203

SUR1

08

SUR2

01

SUR2

02

SUR3

01

SUR3

02

SUR3

03

SUR3

07

SUR3

09

SUR3

10

VW301

VW402

Figure 2 Antibiotic resistance index of the selected EPS producinghalophilic bacteria isolated from soil and water samples of solarsalterns of India

The isolates SUR301 and KS 1805 were tentatively iden-tified as Chromohalobacter based on the study by Arahal etal [45] and bear striking similarity with those of Chromo-halobacter sp isolated fromhypersaline soil sample of TriveniSangam Gujarat [38] The physiological morphological andbiochemical characteristics of isolate SUR303 were consistentwith the features of the genus Salinicoccus [10] while isolatesJS904 and JS504 were tentatively assigned to Aidingimonasand Alteromonas respectively

Finally sensitivity of these isolates to antibiotics (Table 10and Figure 2) also corroborates the findings ofMata et al [25]and Hedi et al [28]

5 Conclusion

It may be emphasized that study of these moderatelyhalophilic bacteria from coastal hypersaline solar salterns ofIndia with special attempt on screening for EPS productionhas led to the discovery of wide variety of halophilic speciesAlthough attention has been focused on the production

of EPS by these halophilic bacterial isolates their physio-biochemical features indicate that they may equally holdpotential towards production of various biomolecules ofindustrial interest

Conflict of Interests

It is declared by the authors that there is no conflict of interestsregarding the publication of this paper

Acknowledgment

This study was financially supported by grants from Uni-versity Grants Commission India (Sanction no F14-2(SC)2008 (SA-III) 31 March 2009) under the Scheme ofRajiv Gandhi National Fellowship

References

[1] E A Galinski and B J Tindall ldquoBiotechnological prospectsfor halophiles and halotolerant microorganismsrdquo in MolecularBiology and Biotechnology of Extremophiles R D Herbert andR J Sharp Eds Blackie London UK 1992

[2] R Margesin and F Schinner ldquoPotential of halotolerant andhalophilic microorganisms for biotechnologyrdquo Extremophilesvol 5 no 2 pp 73ndash83 2001

[3] F Rodriguez Valera F Ruiz Berraquero and A Ramos Cor-menzana ldquoCharacteristics of the heterotrophic bacterial pop-ulations in hypersaline environments of different salt concen-trationsrdquoMicrobial Ecology vol 7 no 3 pp 235ndash243 1981

[4] F Rodriguez-Valera ldquoCharacteristics and microbial ecol-ogy of hypersaline environmentsrdquo in Halophilic Bacteria FRodriguez-Valera Ed vol 1 pp 3ndash30 CRC Press Boca RatonFla USA 1988

[5] D J Kushner and M Kamekura ldquoPhysiology of halophiliceubacteriardquo inHalophilic Bacteria F Rodriguez-Valera Ed vol1 pp 109ndash138 CRC press Boca Raton Fla USA 1988

[6] M A Amoozegar F Malekzadeh and K AMalik ldquoProductionof amylase by newly isolated moderate halophile Halobacillussp strainMA-2rdquo Journal of Microbiological Methods vol 52 no3 pp 353ndash359 2003

[7] C O Jeon J Lim J Lee et al ldquoLentibacillus salarius spnov isolated from saline sediment in China and emendeddescription of the genus Lentibacillusrdquo International Journal ofSystematic and Evolutionary Microbiology vol 55 no 3 pp1339ndash1343 2005

[8] S Bouchotroch E Quesada A Del Moral I Llamas andV Bejar ldquoHalomonas maura sp nov a novel moderatelyhalophilic exopolysaccharide-producing bacteriumrdquo Interna-tional Journal of Systematic and Evolutionary Microbiology vol51 no 5 pp 1625ndash1632 2001

[9] A Biswas A Patra and A Paul ldquoProduction of poly-3-hydroxyalkanoic acids by a moderately halophilic bacteriumHalomonas marina HMA 103 isolated from solar saltern ofOrissa Indiardquo Acta Microbiologica et Immunologica Hungaricavol 56 no 2 pp 125ndash143 2009

[10] S Y Jayachandra S Anil Kumar D P Merley and MB Sulochana ldquoIsolation and characterization of extremehalophilic bacterium Salinicoccus sp JAS4 producing extra-cellular hydrolytic enzymesrdquo Recent Research in Science andTechnology vol 4 no 4 pp 46ndash49 2012

Chinese Journal of Biology 11

[11] I W Sutherland ldquoNovel and established applications of micro-bial polysaccharidesrdquo Trends in Biotechnology vol 16 no 1 pp41ndash46 1998

[12] MWMittelman andG G Geesey ldquoCopper-binding character-istics of exopolymers from a freshwater-sediment bacteriumrdquoApplied and EnvironmentalMicrobiology vol 49 no 4 pp 846ndash851 1985

[13] J Anton I Meseguer and F Rodriguez-Valera ldquoProductionof an extracellular polysaccharide by Haloferax mediterraneirdquoApplied Enviromental Microbiology vol 54 no 10 pp 2381ndash2386 1988

[14] H Parolis L A S Parolis I F Boan et al ldquoThe structure ofthe exopolysaccharide produced by the halophilic ArchaeonHaloferax mediterranei strain R4 (ATCC 33500)rdquo CarbohydrateResearch vol 295 pp 147ndash156 1996

[15] E Quesada V Bejar and C Calvo ldquoExopolysaccharide produc-tion by Volcaniella eurihalinardquo Experientia vol 49 no 12 pp1037ndash1041 1993

[16] J A Mata V Bejar I Llamas et al ldquoExopolysaccharides pro-duced by the recently described halophilic bacteria Halomonasventosae and Halomonas anticariensisrdquo Research in Microbiol-ogy vol 157 no 9 pp 827ndash835 2006

[17] I Llamas J A Mata R Tallon et al ldquoCharacterization ofthe exopolysaccharide produced by Salipiger mucosus A3119879 ahalophilic species belonging to the Alphaproteobacteria iso-lated on the Spanish Mediterranean seaboardrdquo Marine Drugsvol 8 no 8 pp 2240ndash2251 2010

[18] F Martınez-Checa E Quesada J Martınez-Canovas I Llamasand V Bejar ldquoPalleronia marisminoris gen nov sp nova moderately halophilic exopolysaccharide-producing bac-terium belonging to the rsquoAlphaproteobacteriarsquo isolated from asaline soilrdquo International Journal of Systematic and EvolutionaryMicrobiology vol 55 no 6 pp 2525ndash2530 2005

[19] A Ventosa M T Garcia M Kamekura H Onishi and FRuiz-Berraquero ldquoBacillus halophilus sp nov a moderatelyhalophilic Bacillus speciesrdquo Systematic and Applied Microbiol-ogy vol 12 no 2 pp 162ndash166 1989

[20] R A Moraine and P Rogovin ldquoKinetics of polysaccharide B1459 fermentationrdquoBiotechnology and Bioengineering vol 8 no4 pp 511ndash524 1966

[21] M Dubois K A Gilles J K Hamilton P A Rebers and FSmith ldquoColorimetric method for determination of sugars andrelated substancesrdquoAnalytical Chemistry vol 28 no 3 pp 350ndash356 1956

[22] O H Lowry N J Rosenbrough A L Farr and R J RandallldquoProtein measurement with the Folin phenol reagentrdquo TheJournal of Biological Chemistry vol 193 no 1 pp 265ndash275 1951

[23] P Gerhardt R G E Murray W A Wood and N R KriegMethods for General and Molecular Bacteriology AmericanSociety for Microbiology Washington DC USA 1994

[24] G M Garrity J A Bell and T G Lilburn ldquoTaxonomic outlineof the procaryotesrdquo inBergeyrsquosManual of Systematic BacteriologD R Boone and R W Castenholz Eds Springer New YorkNY USA 2nd edition 2003

[25] J A Mata J Martınez-Canovas E Quesada and V BejarldquoA detailed phenotypic characterisation of the type strains ofHalomonas speciesrdquo Systematic and Applied Microbiology vol25 no 3 pp 360ndash375 2002

[26] A Oren ldquoMolecular ecology of extremely halophilic archaeaand bacteriardquo FEMSMicrobiology Ecology vol 39 no 1 pp 1ndash72002

[27] H Baati R Amdouni N Gharsallah A Sghir and E AmmarldquoIsolation and characterization of moderately halophilic bacte-ria from tunisian solar salternrdquo Current Microbiology vol 60no 3 pp 157ndash161 2010

[28] A Hedi N Sadfi M Fardeau et al ldquoStudies on the biodiversityof halophilic microorganisms isolated from El-Djerid salt lake(Tunisia) under aerobic conditionsrdquo International Journal ofMicrobiology vol 2009 Article ID 731786 17 pages 2009

[29] M Birbir A Ogan B Calli and B Mertoglu ldquoEnzymecharacteristics of extremely halophilic archaeal community inTuzkoy Salt Mine Turkeyrdquo World Journal of Microbiology andBiotechnology vol 20 no 6 pp 613ndash621 2004

[30] M Birbir and C Sesal ldquoExtremely halophilic bacterial commu-nities in Sereflikochisar Salt Lake in Turkeyrdquo Turkish Journal ofBiology vol 27 no 7 pp 7ndash22 2003

[31] M Birbir B Calli B Mertoglu et al ldquoExtremely halophilicArchaea from Tuz Lake Turkey and the adjacent Kaldirim andKayacik salternsrdquoWorld Journal ofMicrobiology and Biotechnol-ogy vol 23 no 3 pp 309ndash316 2007

[32] R Rohban M A Amoozegar and A Ventosa ldquoScreeningand isolation of halophilic bacteria producing extracellularhydrolyses from Howz Soltan Lake Iranrdquo Journal of IndustrialMicrobiology and Biotechnology vol 36 no 3 pp 333ndash3402009

[33] M J Garabito M C Marquez and A Ventosa ldquoHalotolerantBacillus diversity in hypersaline environmentsrdquo Canadian Jour-nal of Microbiology vol 44 no 2 pp 95ndash102 1998

[34] A Ventosa A Ramos-Cormenzana and M Kocur ldquoModer-ately halophilic gram-positive cocci from hypersaline environ-mentsrdquo Systematic and Applied Microbiology vol 4 no 4 pp564ndash570 1983

[35] E Quesada A Ventosa F Rodriguez-Valera and A R Cor-menzana ldquoTypes and properties of some bacteria isolated fromhypersaline soilsrdquo Journal of Applied Bacteriology vol 53 no 2pp 155ndash161 1982

[36] E Quesada V Bejar M J Valderrama A Ventosa and AR Ramos Cormenzana ldquoIsolation and characterization ofmoderately halophilic nonmotile rods from different salinehabitatsrdquoMicrobiologia vol 1 no 1-2 pp 89ndash96 1985

[37] S R Dave and H B Desai ldquoMicrobial diversity at marinesalterns near Bhavnagar Gujarat Indiardquo Current Science vol60 no 4 pp 497ndash500 2006

[38] S Kumar R Karan S Kapoor S P Singh and S K KhareldquoScreening and isolation of halophilic bacteria producingindustrially important enzymesrdquo Brazilian Journal of Microbi-ology vol 43 no 4 pp 1595ndash1603 2012

[39] B P Dave and A Soni ldquoDiversity of halophilic archaea atsalt pans around Bhavnagar coast Gujaratrdquo Proceedings ofthe National Academy of Sciences India Section B BiologicalSciences vol 83 no 2 pp 225ndash232 2013

[40] A A Joshi P P Kanekar A S Kelkar et al ldquoCultivable bacterialdiversity of alkaline Lonar lake Indiardquo Microbial Ecology vol55 no 2 pp 163ndash172 2008

[41] T M Raghavan and I Furtado ldquoOccurrence of extremelyhalophilic Archaea in sediments from the continental shelf ofwest coast of Indiardquo Current Science vol 86 no 8 pp 1065ndash1067 2004

[42] S Vijayanand J Hemapriya J Selvin and S Kiran ldquoBiodiver-sity of extremely halophilic bacterial strains isolated from solarsalterns of Tuticorin Tamilnadu Indiardquo International Journal ofWater Resources and Environmental Sciences vol 1 no 1 pp 1ndash72012

12 Chinese Journal of Biology

[43] S G Nanjani and H P Soni ldquoIsolation and characterization ofextremely halotolerant and halophilic organisms from Dwarkaand Veravalrdquo Journal of Pharmacy and Biological Sciences vol2 no 2 pp 20ndash25 2012

[44] H Ghozlan H Deif R A Kandil and S Sabry ldquoBiodiversity ofmoderately halophilic bacteria in hypersaline habitats in EgyptrdquoJournal of General and Applied Microbiology vol 52 no 2 pp63ndash72 2006

[45] D R Arahal M T Garcıa W Ludwig K H Schleifer andA Ventosa ldquoTransfer ofHalomonas canadensis andHalomonasisraelensis to the genus Chromohalobacter as Chromohalobac-ter canadensis comb nov and Chromohalobacter israelensiscomb novrdquo International Journal of Systematic and EvolutionaryMicrobiology vol 51 no 4 pp 1443ndash1448 2001

Submit your manuscripts athttpwwwhindawicom

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Anatomy Research International

PeptidesInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporation httpwwwhindawicom

International Journal of

Volume 2014

Zoology

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Page 2: Research Article Production of Extracellular …downloads.hindawi.com/archive/2014/205731.pdfResearch Article Production of Extracellular Polymeric Substances by Halophilic Bacteria

2 Chinese Journal of Biology

also established that halophilic EPS producers belong to thegamma-proteobacteria (Idiomarina andAlteromonas) as wellas alpha-proteobacteria (Salipiger mucosus and Palleroniamarisminoris) [17 18]

This study is focused on the isolation of halophilicbacteria from water and soil samples of some selectedsolar salterns located in the states of Gujarat Orissa andWest Bengal India and evaluation of their EPS productionefficiency under laboratory conditions Attempts have alsobeen made on the tentative identification of some potent EPSproducing halophilic isolates based on their morphologicaland physiobiochemical features

2 Materials and Methods

21 Collection of Samples Soil and water samples frommultipond solar salterns situated along the coast of GujaratOrissa and West Bengal India were collected in sterilepolypropylene containers and stored at 4∘C until used forisolation of halophilic bacteria A total of 12 soil and 7water samples were collected from Jogrinar (23∘131015840N and69∘5910158405110158401015840 E) Khari Rohar (23∘410158401610158401015840N and 70∘910158403710158401015840 E)Kandla (22∘591015840N and 70∘131015840 E) and Albert Victor Port(21∘010158405810158401015840N and 71∘3210158405410158401015840 E) of Gujarat Similarly 10 soil and5 water samples were collected from Surala (19∘8410158407310158401015840N and84∘651015840 E) and Humma (19∘261015840N and 85∘51015840 E) the two majorsolar salterns of Orissa In West Bengal the sampling siteswere located at Dadanpatrabar (22∘261015840N and 87∘201015840 E) andBaksal (22∘11015840N and 87∘671015840 E) of East Midnapur and a total of9 soil and 6 water samples were collected

22 Isolation of Halophilic Bacteria Aerobic heterotrophicandhalophilic bacteria of soil andwater sampleswere isolatedby serial dilution and plating on MH agar medium [19]supplemented with different concentrations of NaCl Themedium contained (gL) yeast extract 10 protease peptone5 glucose 1 NaCl 100 MgCl

26H2O 7 MgSO

47H2O 96

CaCl22H2O 036 KCl 2 NaHCO

3006 and NaBr 0026

(pH 72) The plates were incubated at 37∘C for 3 to 5 daysand bacterial colonies with distinctmorphologywere isolatedin pure form and maintained on slopes of the same mediumTotal bacterial counts were expressed as colony forming units(cfu)mL and g of water and soil respectively

23 Screening of Halophilic Bacteria for EPS ProductionTo evaluate the EPS production capability the bacterialisolates were grown in MY medium [20] supplemented with5 NaCl for 12 days at 32∘C under continuous shaking(120 rpm) The medium contained (gL) NaCl 50 MgCl

2

6H2O 9 MgSO

47H2O13 CaCl

22H2O 02 KCl 13

NaHCO3 005 NaBr 015 FeCl

36H2O 0005 glucose 10

yeast extract 3 malt extract 3 and protease peptone 5(pH 72) The EPS from the growing culture was isolatedusing the method as described by Quesada et al [15] Theculture was centrifuged (at 10000timesg for 10min) the EPSin the supernatant was precipitated with chilled ethanol

and recovered by centrifugation (12000timesg for 10min) andwashed with chilled 70 ethanol The washed precipitatewas collected by centrifugation dissolved in known volumeof distilled water and used for quantification and chemicalanalysis

24 Chemical Analysis of EPS Total carbohydrate contentof the EPS was estimated following the method of Duboiset al [21] To 1 mL of dissolved EPS sample 05mL of 5phenol and 35mL of concentrated sulfuric acid were addedand incubated at 30ndash40∘C for 10ndash20 minutes in hot waterbath Absorbance was read at 490 nm and the amount ofcarbohydrate was determined from the calibration curveprepared using glucose as standard

Protein content of the EPS was determined followingthe Folin phenol method of Lowry et al [22] To 1mL ofEPS sample 5mL of alkaline solution and 05mL of Folinphenol reagent were added and incubated for 30 minutes atroom temperature in dark The absorbance was measured at670 nm and the concentration was read from the calibrationcurve prepared by using bovine serum albumin (BSA) as thestandard

25 Characterization and Identification of Selected BacterialIsolates The selected bacterial isolates were characterizedmorphologically and physiobiochemically following stan-dard microbiological methods as described by Gerhardt et al[23] To determine the antibiotic sensitivity pattern of theseisolates the antibiotic impregnated discs (Himedia 6mmdia) were placed on MH agar plates seeded with respectivebacterial isolates The plates were incubated for 24 h at32∘C and diameter of inhibition zones was measured to thenearest mm Production of acids from sugars by the bacterialisolates was tested on phenol redmedium supplemented with1 carbon source Characteristics of the bacterial isolateswere compared with those described in Bergeyrsquos Manual ofSystematic Bacteriology [24] and that of Mata et al [25] fordetermination of taxonomic identity

3 Results

31 Isolation of Halophilic Bacteria A total of 31 soil and18 water samples collected from 8 different sites spreadover the states of Gujarat Orissa and West Bengal Indiawere analyzed for the aerobic heterotrophic and halophilicbacteria following dilution and plating on MH agar mediumsupplemented with 5 10 and 15 NaCl The total bacterialpopulation of both soil and water samples as determinedby colony forming units varied considerably and declinedgradually irrespective of sampling sites with increasing NaClconcentration in the isolation medium (Tables 1 and 2)

A total of 587 halophilic and halotolerant hetero-trophic and aerobic bacterial isolates were obtainedin pure form The majority of these isolates (410)were derived from soil samples (Table 3) while177 were obtained from water samples (Table 4)Most of the isolates were Gram-negative motile aerobic

Chinese Journal of Biology 3

Table 1 Bacterial population of soil samples collected from solar salterns of Gujarat Orissa and West Bengal India

State Sampling sites Number of samplesBacterial count (cfug of soil times102)

NaCl () in medium5 10 15

Gujarat

Jogrinar 4 074ndash866 021ndash366 011ndash4Kandla Port 3 086ndash735 121ndash211 008ndash0Khari Rohar 3 239ndash768 012ndash178 047ndash966Victor Port 2 173ndash283 040ndash0 mdash

Orissa Humma 5 013ndash061 025ndash055 mdashSurala 5 1ndash366 007ndash023 mdash

West Bengal Baksal 4 2ndash390 020ndash040 020ndash0Dadanpatrabar 5 27ndash79 007ndash060 007ndash030

Table 2 Bacterial population of water samples collected from solar salterns of Gujarat Orissa and West Bengal India

States Sampling sites Number of samplesBacterial count (cfumL of water times102)

NaCl () in medium5 10 15

Gujarat

Jogrinar 2 028ndash556 172ndash0 040ndash0Kandla Port 2 053ndash540 016ndash11 066ndash0Khari Rohar 1 023ndash0 mdash mdashVictor Port 2 126ndash190 mdash mdash

Orissa Humma 3 023ndash23 mdash mdashSurala 2 027ndash120 083ndash0 mdash

West Bengal Baksal 3 110ndash136 013ndash06 010ndash037Dadanpatrabar 3 05ndash190 007ndash050 01-02

rods and produced white to cream colored circular smooth-edged colonies on MH agar In general the isolates werecapable of tolerating wide range of temperature and pH fortheir growth

32 Screening for EPS Production EPS producing abilityof these isolates was examined in batch culture using MYmedium supplemented with 5 NaCl and the soluble EPScontent of each of the isolates was evaluated in terms ofits carbohydrate content as determined by Dubois method[21] Among the 410 soil isolates majority (184) producedsoluble EPS ranging from 05 to 07 gL in terms of theircarbohydrate content while only 15 isolates produced morethan 1 gL of EPS (Table 5) On the contrary among the177 bacterial strains isolated from saline water samples only5 produced soluble EPS accounting for more than 1 gL ofcarbohydrate However majority (77) of them produced 05ndash07 gL of soluble EPS (Table 6)

In all 20 potent isolates producing gt1 gL of EPS werefurther allowed to grow in MY medium under continuousshaking (120 rpm) and the soluble EPS produced after 8days of growth was estimated following the method asdescribed above Kinetics of growth and EPS productionrevealed that EPS production by these isolates increasedwith biomass formation (Figure 1) and a few of the selectedisolates such as SUR202 SUR307 SUR310 JW307 and

JS904 appeared to be promising with an EPS yield of168ndash185 gL

33 Characterization and Identification of Selected BacterialIsolates Morphological and physiological studies revealed(Table 7) that these halophilic bacteria (with the exceptionof isolate SUR303) formed cream colored smooth colonieson MH agar medium Most of them were Gram-negativemotile rods only three isolates were Gram-positive one ofthem (isolate SUS303) was coccus while the isolate KW203was the only endospore former The isolates were capableof tolerating 15ndash20 NaCl in the medium and a pH of 5ndash11 Optimum growth was observed at a temperature of 32ndash37∘C but all were able to tolerate a temperature as high as40∘C

Analysis of biochemical characters (Table 8) showedthat the majority of these halophilic isolates gave negativeresponse to MR-VP tests and failed to produce extracellularenzymes like amylase cellulase pectinase inulinase gelati-nase lipase caseinase xylanase and urease None of theseisolates however were capable of producing H

2S lysine-

arginine- and ornithine decarboxylaseCarbon source utilization pattern (Table 9) of these

halophilic isolates varied considerably All the isolates wereable to utilize ribose fructose mannitol salicin cellobioseacetate benzoate and succinate and three of them (JS803

4 Chinese Journal of Biology

Table 3 Bacterial isolates obtained from soil samples of solar salterns of Gujarat Orissa and West Bengal India

States Sampling sites Number of samplesNumber of bacterial isolates

NaCl () in medium5 10 15

Gujarat

Jogrinar 4 44 23 4Kandla Port 3 19 12 4Khari Rohar 3 46 16 6Victor Port 2 20 5 mdash

Orissa Humma 5 48 14 mdashSurala 5 31 31 2

West Bengal Baksal 4 19 8 3Dadanpatrabar 5 29 16 10

Total 31 256 125 29

Table 4 Bacterial isolates obtained from water samples of solar salterns of Gujarat Orissa and West Bengal India

States Sampling sites Number of samplesNumber of bacterial isolates

NaCl () in medium5 10 15

Gujarat

Jogrinar 2 21 14 5Kandla Port 2 19 5 2Khari Rohar 1 1 3 mdashVictor Port 2 26 mdash 1

Orissa Humma 3 16 mdash mdashSurala 2 12 8 3

West Bengal Baksal 3 9 3 4Dadanpatrabar 3 17 4 4

Total 18 121 37 19

SUR301 and SUR307) appeared to be versatile in utilizingall 30 carbon sources Fermentation pattern of these strainsvaried remarkably the majority were unable to fermentmost of the carbon sources On the contrary fructosesorbitol and benzoate were fermented by most of the isolates(Table 9)

Sensitivity of these isolates to 22 different antibioticswas tested by disc-diffusion method (Table 10) and theantibiotic resistance index (ARI) was determined (Figure 2)Themajority of the isolates were sensitive to chloramphenicol(30 120583g) gentamycin (10 120583g) and norfloxacin (10 120583g) followedby ampicillin (10 120583g) Resistance to vancomycin (30120583g) fol-lowed by trimethoprim (30 120583g) was predominant amongstthe tested halophiles As judged by the ARI values the isolateSUR302 was the most resistant one (ARI = 065) followed bySUR301 KW203 and KW1805 while the lowest ARI (023)was indicated by the isolate JW307 which was most sensitiveto the tested antibiotics

The morphological physiological and biochemical char-acters including the carbon source utilization and fermen-tation patterns along with antibiotic susceptibility wereanalyzed and compared with the phenotypic characters ofhalophilic bacterial genera [5 18] so far reported According

to the phenotypic and biochemical characteristics 70 of theselected isolates (14) were tentatively identified asmembers ofthe genusHalomonasThe onlyGram-positive coccus (isolateSUR303) and the rod shaped endospore forming isolate(isolate KW203) were assigned to Salinicoccus and Bacillusrespectively Two of the isolates (KS1805 and SUR301) wereplaced in the genus Chromohalobacter while the remain-ing two (isolates JS504 and JS904) were included underAlteromonas and Aidingimonas respectively

4 Discussion

Halophiles have mainly been isolated from wide diversity ofenvironments such as saltern crystallizer ponds the DeadSea solar lakes and hypersaline lakes [26] Culture depen-dent diversity studies on halophiles have been made fromTunisian solar saltern [27 28] Tuzkoy salt mine [29] Sere-flikochisar Salt Lake [30] Kaldirim and Kayacik of Tuz Lake[31] Turkey Howz Soltan Lake Iran [32] and hypersalineenvironments in South Spain [33ndash36] In the Indian contextthe halophilic diversity studies have been restricted mainlyto the marine salterns of Bhavnagar [37ndash40] Lonar Lake[40] and Peninsular Coast [41 42]The present study reports

Chinese Journal of Biology 5

Table 5 Screening of halophilic bacterial isolates derived from soil samples for the production of extracellular polymeric substances

States Sampling sites Number of isolates Extracellular polymeric substances (gL)gt050 051ndash070 071ndash10 gt10

Gujarat

Jogrinar 71 18 42 7 4Kandla Port 35 13 13 7 2Khari Rohar 68 26 31 11 mdashVictor Port 25 20 3 2 mdash

Orissa Humma 62 22 37 3 mdashSurala 64 15 25 15 9

West Bengal Baksal 30 14 12 4 mdashDadanpatrabar 55 32 21 2 mdash

Total 410 160 184 51 15

Table 6 Screening of halophilic bacterial isolates derived from water samples for the production of extracellular polymeric substances

States Sampling sites Number of isolates Extracellular polymeric substances (gL)gt050 051ndash070 071ndash10 gt10

Gujarat

Jogrinar 40 26 9 3 2Kandla Port 26 13 11 1 1Khari Rohar 4 1 3 mdash mdashVictor Port 27 7 17 1 2

Orissa Humma 16 4 11 1 mdashSurala 23 8 6 9 mdash

West Bengal Baksal 16 7 6 3 mdashDadanpatrabar 25 8 14 3 mdash

Total 177 74 77 21 5

the distribution of halophilic and halotolerant bacterialcommunities in the inland multipond solar salterns spreadover the coasts of India Halophilic bacterial communitiesof 31 soil and 18 water samples from 8 different sites wereanalyzed by dilution and plating method (Tables 1 and 2) andprovide information regarding the availability anddiversity ofhalophilic bacteria in the solar saltern ponds Colony formingunits of soil and water samples revealed that soil samplesholdmore viable bacterial counts than water samples Ragha-van and Furtado [41] studied the occurrence of extremelyhalophilic archaea in sediments from the continental shelf ofwest coast of India and reported the presence of relatively lowaverage counts (7ndash5 times 103) of extreme halophiles in offshoresediments in contrast to the very high counts (105ndash109)of marine eubacteria Similar study by Joshi et al [40]reported that the total numbers of microorganisms in the soiland water samples were 102ndash106 cfug and 102ndash104 cfumLrespectively

A total of 587 halophilic and halotolerant bacterial strainswere isolated showing different degrees of NaCl tolerance(Tables 3 and 4) and supported the observations of Quesadaet al [35] andVentosa et al [34] However during the presentstudy we were unable to isolate extreme halophiles whichhave frequently been identified as the dominant phylotypes inhypersaline environments along with solar salterns of India

[39 41] Furthermore soil samples showed more bacterialdiversity than water ones which is in accordance with theobservations of Joshi et al [40]

During the course of screening of the moderatelyhalophilic isolates for EPS production (Tables 5 and 6) only20 isolates appeared to be promising with significant yield ofEPS (10ndash185 gL) (Figure 1) Joshi and coworkers [40] sim-ilarly screened 86 halophilic bacteria from Lonar Lake andreported Halomonas campisalis and Vagococcus carniphilusas potent EPS producers Similarly Nanjani and Soni [43]also isolated 73 halotolerant and halophilic bacteria from soilsamples of Veraval and Dwarka 23 of them produced EPSranging from 02 to 1060 gL In addition reports on EPSproduction by moderately halophilic bacteria of the genusHalomonas are not uncommon [8 15ndash17]

Attempts have been made to determine the taxonomicidentity of all 20 promising EPS producing isolates followingdetailed physiobiochemical characterization (Tables 7ndash9) andcomparison with Bergeyrsquos Manual of Systematic Bacteriology[24] and those of Mata et al [25]Themajority (14 isolates) ofthem were Gram-negative nonsporulating rods and capableof growing in 25ndash20NaCl similar to those ofHalomonas asreported by Quesada et al [36] Ghozlan et al [44] andMataet al [25] However the Gram-positive endospore formingisolate KW203 was assigned to the genus Bacillus

6 Chinese Journal of Biology

Table7Morph

ologicalandph

ysiologicalcharacteristicso

fthe

selected

EPSprod

ucinghaloph

ilicb

acteria

isolated

from

solarsalternso

fInd

ia

Character

Bacterialisolates

JS504

JS802

JS803

JS904

JW302

JW307

KS110

1KS

1805

KW203

VW301

Morph

ologicalcharacters

Colon

ymorph

olog

yCr

eamcirc

ular

Whitecirc

ular

Creamcirc

ular

Creamcirc

ular

Transparentcircular

Creamcirc

ular

Creamcirc

ular

Whitecirc

ular

Whitecirc

ular

Creamcirc

ular

Gram

nature

minus+

minusminus

minusminus

minusminus

+minus

Cellshape

Rod

Rod

Rod

Rod

Rod

Rod

Rod

Rod

Rod

Rod

Cell

arrang

ement

Sing

leSing

leSing

leSing

leSing

lechains

Sing

lechains

Sing

lechains

Sing

leSing

leclu

mped

Sing

lechains

Motility

++

++

++

minusminus

++

Pigm

entatio

nminus

minusminus

minusminus

minusminus

minusminus

minus

Endo

sporeformation

minusminus

minusminus

minusminus

minusminus

+minus

Physiologicalcharacters

pHrang

efor

grow

th5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

pHop

timum

forg

rowth

119

89

1111

118

89

Temprang

efor

grow

th(∘C)

27ndash4

027ndash4

027ndash4

027ndash4

027ndash4

027ndash4

022ndash4

027ndash4

027ndash4

022ndash4

0Tempop

timum

forg

rowth

(∘C)

3237

3732

3232

3232

3732

NaC

lrange

forg

rowth

()

0ndash15

0ndash20

0ndash15

0ndash20

0ndash15

0ndash20

0ndash20

25ndash15

25ndash15

0ndash20

NaC

loptim

umforg

rowth

()

510

105

105

575

105

Growth

onKingrsquosBmedium

++

++

++

++

++

Growth

onMcC

onky

agar

minusminus

minusminus

minusminus

minusminus

minusminus

Tentativeidentity

Alteromonas

Halom

onas

Halom

onas

Aidingim

onas

Halom

onas

Halom

onas

Halom

onas

Chromohalobacter

Bacillus

Halom

onas

Character

Bacterialisolates

VW402

SUR108

SUR3

01SU

R302

SUR3

03SU

R307

SUR3

09SU

R310

SUR2

02SU

R201

Morph

ologicalcharacters

Colon

ymorph

olog

yCr

eamcirc

ular

Creamcirc

ular

Whitegum

my

Transparentcircular

Flatreddish-pink

Creamcirc

ular

Creamcirc

ular

Creamcirc

ular

Creamcirc

ular

Creamcirc

ular

Gram

nature

minusminus

minusminus

+minus

minusminus

minusminus

Cellshape

Rod

Rod

Rod

Rod

Cocci

Rod

Rod

Rod

Rod

Rod

Cell

arrang

ement

Sing

leSing

leSing

leSing

leSing

leclu

mped

Sing

leSing

leSing

leSing

leSing

leMotility

+minus

minus+

minus+

minus+

minusminus

Pigm

entatio

nminus

minusminus

minusPink

minusminus

minusminus

minus

Endo

sporeformation

minusminus

minusminus

minusminus

minusminus

minusminus

Physiologicalcharacters

pHrang

efor

grow

th5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

pHop

timum

forg

rowth

97

77

117

77

79

Temprang

efor

grow

th(∘C)

27ndash4

027ndash4

027ndash4

032ndash4

532ndash4

522ndash4

032ndash4

022ndash4

522ndash4

532ndash4

5Tempop

timum

forg

rowth

(∘C)

3232

3732

3727

3227

2732

NaC

lrange

forg

rowth

()

0ndash15

0ndash15

25ndash15

25ndash15

25ndash15

0ndash20

0ndash20

0ndash20

0ndash20

0ndash15

NaC

loptim

umforg

rowth

()

525

575

525

7525

575

Growth

onKingrsquosBmedium

++

++

++

++

++

Growth

onMcC

onky

agar

minusminus

minusminus

minusminus

minusminus

minusminus

Tentativeidentity

Halom

onas

Halom

onas

Chromohalobacter

Halom

onas

Salin

icoccus

Halom

onas

Halom

onas

Halom

onas

Halom

onas

Halom

onas

Chinese Journal of Biology 7

Table8Biochemicalcharacteris

ticso

fthe

selected

EPSprod

ucinghaloph

ilicb

acteria

isolated

from

solarsalternso

fInd

ia

Character

Bacterialisolates

JS 504

JS 802

JS 803

JS 904

JW 302

JW 307

KS 1101

KS 1805

KW 203

VW 301

VW 402

SUR

108

SUR

301

SUR

302

SUR

303

SUR

307

SUR

309

SUR

310

SUR

202

SUR

201

Biochemicalcharacters

Voges-Proskauertest

minusminus

minusminus

minusminus

minusminus

+minus

minusminus

minusminus

minusminus

minusminus

minusminus

Citrateu

tilization

+minus

minusminus

minus+

minus+

minus+

++

+minus

minus+

++

++

Methylred

test

minusminus

minusminus

minusminus

minus+

+minus

minusminus

minusminus

minusminus

minusminus

minusminus

Prod

uctio

nof

Glutaminase

++

++

++

++

++

++

++

minus+

++

++

Asparaginase

++

++

++

++

++

++

++

minus+

++

++

Amylase

+minus

minusminus

minusminus

minusminus

+minus

minusminus

minusminus

minusminus

minusminus

minusminus

Cellulase

minusminus

minusminus

minusminus

minusminus

+minus

minusminus

minusminus

minusminus

minusminus

minusminus

Pectinase

+minus

minusminus

minusminus

minusminus

+minus

minusminus

minusminus

minusminus

minusminus

minusminus

Inulinase

+minus

minusminus

minusminus

minusminus

+minus

minusminus

minusminus

minusminus

minusminus

minusminus

Gelatinase

minusminus

minusminus

minusminus

minusminus

+minus

minusminus

minusminus

minusminus

minusminus

minusminus

Lipase

+minus

minusminus

minusminus

minus+

+minus

minusminus

+minus

minusminus

minusminus

minusminus

Xylanase

+minus

minusminus

minusminus

minusminus

+minus

minusminus

minusminus

minusminus

minusminus

minusminus

Caseinase

minusminus

minus+

minusminus

minusminus

minusminus

minusminus

minus+

minusminus

minusminus

minusminus

Urease

minusminus

minus+

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

Catalase

++

++

++

++

++

++

++

++

++

++

Nitrater

eductase

++

++

++

++

++

++

++

++

++

++

H2S

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

Lysin

edecarbo

xylase

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

Arginined

ecarbo

xylase

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

Ornith

ined

ecarbo

xylase

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

Indo

leminus

minusminus

minusminus

minusminus

+minus

minusminus

minus+

minus+

minusminus

minusminus

minus

Phenylalanined

eaminase

++

+minus

+minus

++

minus+

minus+

++

++

++

minus+

Tryptoph

andeam

inase

++

++

++

++

++

minusminus

++

++

++

+minus

8 Chinese Journal of Biology

Table9Ca

rbon

source

utilizatio

nandferm

entatio

nof

thes

elected

EPSprod

ucinghaloph

ilicb

acteria

isolated

from

samples

ofsolarsalternso

fInd

ia

Carbon

source

Bacterialisolates

JS 504

JS 802

JS 803

JS 904

JW 302

JW 307

KS 1101

KS 1805

KW 203

VW

301

VW

402

SUR

108

SUR

301

SUR

302

SUR

303

SUR

307

SUR

309

SUR

310

SUR

202

SUR

201

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

Glucose

++

++

+minusminus

++minus

+minus

+minus

++

++

+minus

+minus

+minus

+minusminus

++

++minus

+minus

+minusminusminusminusminus

Galactose

+minus

+minus

+minusminus

++minus

+minus

+minus

++

+minus

+minus

+minus

+minus

++minusminus

+minus

++

+minus

+minus

+minus

+minus

Arabino

seminusminusminusminus

+minusminusminusminusminus

+minus

+minus

++

++

+minus

+minus

+minus

+minusminus

++minus

+minus

+minusminusminus

+minus

+minus

Manno

se+minus

+minus

+minusminus

++minus

+minus

+minusminus

++

++minus

+minus

+minus

+minusminusminusminusminus

+minus

+minus

+minus

+minus

+minus

Rham

nose

+minus

+minus

+minus

+minusminusminusminusminusminusminus

++

+minusminusminus

+minus

+minus

++minus

++minus

+minusminusminusminusminusminusminus

+minus

Trehalose

+minus

+minus

+minusminus

+minus

+minusminusminusminus

+minus

++minusminusminusminus

+minus

+minusminus

++

++minusminusminusminusminusminusminus

+minus

Lactose

minusminusminusminus

+minusminusminus

+minus

+minus

+minus

+minus

+minus

+minus

+minusminusminus

+minus

++minusminus

+minusminusminusminusminus

+minus

+minus

Sucrose

+minus

+minus

+minus

++

++

+minus

+minus

+minus

++

+minus

+minusminusminus

+minus

++

+minus

+minus

+minus

+minusminusminus

+minus

Xylose

minusminusminusminus

+minus

+minusminusminus

+minus

++minus

+minusminus

+minus

+minus

+minus

+minusminus

+minus

++

+minusminusminusminus

+minusminusminus

Ribo

se+minus

++

++

++

+minus

+minus

+minus

++

++

+minus

+minus

+minus

++

++

++

+minus

+minus

+minus

+minus

+minus

Fructose

+minus

++

+minus

++

+minus

++

++

+minus

++

++

++

++

++

++

++

++

++

++

++

++

Maltose

minusminusminusminus

+minusminus

+minus

++minus

+minus

+minus

++

+minus

+minus

+minus

++minus

+minus

++minusminusminusminusminus

+minus

+minus

Glycine

minusminusminus

++minus

+minusminus

+minusminusminusminus

+minusminusminusminusminusminusminusminusminus

+minusminusminusminusminus

+minus

+minusminusminusminus

+minusminus

Glycerol

+minus

+minus

+minusminus

++

++minus

+minus

+minus

++

+minus

+minus

+minus

+minusminus

++

++minus

+minus

+minus

+minus

+minus

Adon

itol

+minus

+minus

+minus

+minusminusminus

+minus

+minus

+minus

+minus

+minus

+minusminusminus

+minus

++

+minus

+minus

+minus

+minusminusminusminusminus

Salicin

+minus

++

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

++

++

+minus

+minus

+minus

+minus

+minus

Meso-inosito

l+minusminusminus

+minus

+minus

+minus

+minus

++

+minus

++

+minus

+minusminusminus

+minusminus

++minus

+minus

+minus

+minus

+minus

+minus

Mannitol

+minus

+minus

+minus

++

+minus

+minus

+minus

+minus

++

+minus

+minus

+minus

+minus

+minus

++

+minus

+minus

+minus

+minus

+minus

Sorbito

l+

++

++minusminus

++

+minus

++

++

++

++

++

++

++

+minus

+minus

++

++minus

+minus

++

+minus

Raffino

seminusminus

++

++minusminus

+minus

+minus

+minus

+minus

++minusminusminusminusminusminusminusminus

++

++

+minus

+minus

+minus

+minus

+minus

Cellobiose

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

++

+minus

+minus

+minus

+minus

++

+minus

+minus

+minus

+minus

+minus

+minus

Melibioseminusminusminusminusminusminusminusminusminusminusminusminusminusminusminusminus

++minusminusminusminusminusminusminusminus

++minusminusminusminusminusminusminusminusminusminusminusminus

Na-acetate

+minus

+minus

+minus

+minus

++

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

++

++

+minus

+minus

+minus

+minus

+minus

Na-citrate

+minus

++

+minusminusminusminus

+minusminusminusminusminusminusminusminusminusminusminusminusminusminusminusminusminus

+minusminusminusminusminusminusminusminusminusminusminusminus

Na-fumarate

+minus

+minus

+minus

++

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minusminus

++minus

+minus

+minus

+minus

+minus

+minus

Na-form

ate

+minus

+minus

+minus

+minus

+minusminusminus

+minus

+minus

+minusminusminus

+minusminusminus

+minus

++minusminus

+minusminusminusminusminusminusminusminusminus

Na-benzoate

+minus

++

++

+minus

+minus

+minus

++

++

++

+minus

+minus

++

++

+minus

+minus

+minus

++

++

++

++

Na-malateminusminusminusminusminusminus

++

+minusminusminusminusminusminusminusminusminusminusminusminusminusminusminusminusminus

++

++minusminusminusminusminusminusminusminusminusminus

Na-prop

ionate

+minus

+minus

+minusminusminus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

++minus

++minus

+minus

+minus

+minus

+minus

Na-succinate

+minus

+minus

+minus

++

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

++

++

+minus

+minus

+minus

+minus

+minus

Uutilization

Fferm

entatio

n

Chinese Journal of Biology 9

Table10A

ntibiotic

sensitivitypatte

rnof

thes

electedEP

Sprod

ucinghaloph

ilicb

acteria

isolated

from

solarsalternso

fInd

ia

Antibiotics

Bacterialisolates

JS 504

JS 802

JS 803

JS 904

JW 302

JW 307

KS 1101

KS 1805

KW 203

VW 301

VW 402

SUR

108

SUR

301

SUR

302

SUR

303

SUR

307

SUR

309

SUR

310

SUR

202

SUR

201

Ampicillin(10120583

g)S

SR

SS

SS

RI

SS

SR

SS

SS

SS

SAmoxicillin

(30120583

g)S

SR

RS

SS

RS

SS

SR

RS

IR

SS

SBa

citracin

(10120583

g)S

SR

SS

SR

RS

IR

RR

RS

IR

RR

RCiprofl

oxacin

(30120583

g)I

II

RI

II

IR

II

II

IR

SR

II

ICh

lorte

tracyclin

e(30120583g)

SI

SS

IS

RR

SS

SS

RR

SS

II

IS

Chloramph

enicol(10120583

g)S

SS

SS

SS

SR

SS

SS

SS

SR

SS

SErythrom

ycin

(15120583

g)S

SI

SS

IS

II

II

II

SR

RI

IS

SFu

sidicacid

(30120583

g)I

RR

SS

IR

RS

RI

RR

RS

IR

II

RGentamycin

(10120583

g)S

SS

SS

SS

SR

SS

SS

SS

IS

SS

SKa

namycin

(30120583

g)I

IS

RS

II

SR

II

RI

RR

RI

RI

RMethicillin(5120583g)

SS

RS

IR

RR

RR

RR

RR

SR

RR

RR

Neomycin

(30120583

g)S

SI

RI

IR

II

IR

II

RR

RI

IR

RNorflo

xacin(30120583

g)S

SR

SS

SS

SI

SS

SS

SS

SS

SS

SNetillin

(10120583

g)R

SS

IR

SS

SR

RR

SS

RR

RR

RR

RNalidixicacid

(30120583

g)R

RS

RR

II

RR

II

RR

RR

SR

RR

RNovob

iosin

(30120583

g)I

RR

SR

SS

RS

SS

IR

SS

SI

SS

SPo

lymixin

B(50120583

g)R

RI

RI

IR

RR

RI

IR

RR

II

II

SPenicillinG(10120583

g)S

SR

IS

IR

RR

II

SR

RS

II

II

SStreptom

ycin

(10120583

g)S

SS

RS

RR

SR

RR

IS

RS

SR

II

RTetracyclin

e(30120583g)

RR

RR

RI

RR

SR

IR

RR

RS

RR

RI

Trim

etho

prim

e(30120583g)

SS

SS

IS

SI

SS

SS

IR

RS

SS

SS

Vancom

ycin

(30120583

g)R

RR

SR

RR

RS

RR

RR

RS

RR

RR

R

S=SensitiveR

=Re

sistantI

=Interm

ediate

10 Chinese Journal of Biology

0

1

2

3

4

Bacterial isolate

Gro

wth

OD

at540

nmto

tal E

PS (g

L)

OD at 540nmTotal EPS

JS504

JS802

JS803

JS904

JW302

JW307

KS1101

KS1805

KW203

VW301

VW402

SUR1

08

SUR3

01

SUR3

02

SUR3

03

SUR3

07

SUR3

09

SUR3

10

SUR2

02

SUR2

01

Figure 1 Screening of moderately halophilic bacteria isolatedfrom soil and water samples of solar salterns for production ofextracellular polymeric substances

0

02

04

06

08

1

Ant

ibio

tic re

sista

nce i

ndex

Bacterial isolate

JS504

JS802

JS803

JS904

JW302

JW307

KS1101

KS1805

KW203

SUR1

08

SUR2

01

SUR2

02

SUR3

01

SUR3

02

SUR3

03

SUR3

07

SUR3

09

SUR3

10

VW301

VW402

Figure 2 Antibiotic resistance index of the selected EPS producinghalophilic bacteria isolated from soil and water samples of solarsalterns of India

The isolates SUR301 and KS 1805 were tentatively iden-tified as Chromohalobacter based on the study by Arahal etal [45] and bear striking similarity with those of Chromo-halobacter sp isolated fromhypersaline soil sample of TriveniSangam Gujarat [38] The physiological morphological andbiochemical characteristics of isolate SUR303 were consistentwith the features of the genus Salinicoccus [10] while isolatesJS904 and JS504 were tentatively assigned to Aidingimonasand Alteromonas respectively

Finally sensitivity of these isolates to antibiotics (Table 10and Figure 2) also corroborates the findings ofMata et al [25]and Hedi et al [28]

5 Conclusion

It may be emphasized that study of these moderatelyhalophilic bacteria from coastal hypersaline solar salterns ofIndia with special attempt on screening for EPS productionhas led to the discovery of wide variety of halophilic speciesAlthough attention has been focused on the production

of EPS by these halophilic bacterial isolates their physio-biochemical features indicate that they may equally holdpotential towards production of various biomolecules ofindustrial interest

Conflict of Interests

It is declared by the authors that there is no conflict of interestsregarding the publication of this paper

Acknowledgment

This study was financially supported by grants from Uni-versity Grants Commission India (Sanction no F14-2(SC)2008 (SA-III) 31 March 2009) under the Scheme ofRajiv Gandhi National Fellowship

References

[1] E A Galinski and B J Tindall ldquoBiotechnological prospectsfor halophiles and halotolerant microorganismsrdquo in MolecularBiology and Biotechnology of Extremophiles R D Herbert andR J Sharp Eds Blackie London UK 1992

[2] R Margesin and F Schinner ldquoPotential of halotolerant andhalophilic microorganisms for biotechnologyrdquo Extremophilesvol 5 no 2 pp 73ndash83 2001

[3] F Rodriguez Valera F Ruiz Berraquero and A Ramos Cor-menzana ldquoCharacteristics of the heterotrophic bacterial pop-ulations in hypersaline environments of different salt concen-trationsrdquoMicrobial Ecology vol 7 no 3 pp 235ndash243 1981

[4] F Rodriguez-Valera ldquoCharacteristics and microbial ecol-ogy of hypersaline environmentsrdquo in Halophilic Bacteria FRodriguez-Valera Ed vol 1 pp 3ndash30 CRC Press Boca RatonFla USA 1988

[5] D J Kushner and M Kamekura ldquoPhysiology of halophiliceubacteriardquo inHalophilic Bacteria F Rodriguez-Valera Ed vol1 pp 109ndash138 CRC press Boca Raton Fla USA 1988

[6] M A Amoozegar F Malekzadeh and K AMalik ldquoProductionof amylase by newly isolated moderate halophile Halobacillussp strainMA-2rdquo Journal of Microbiological Methods vol 52 no3 pp 353ndash359 2003

[7] C O Jeon J Lim J Lee et al ldquoLentibacillus salarius spnov isolated from saline sediment in China and emendeddescription of the genus Lentibacillusrdquo International Journal ofSystematic and Evolutionary Microbiology vol 55 no 3 pp1339ndash1343 2005

[8] S Bouchotroch E Quesada A Del Moral I Llamas andV Bejar ldquoHalomonas maura sp nov a novel moderatelyhalophilic exopolysaccharide-producing bacteriumrdquo Interna-tional Journal of Systematic and Evolutionary Microbiology vol51 no 5 pp 1625ndash1632 2001

[9] A Biswas A Patra and A Paul ldquoProduction of poly-3-hydroxyalkanoic acids by a moderately halophilic bacteriumHalomonas marina HMA 103 isolated from solar saltern ofOrissa Indiardquo Acta Microbiologica et Immunologica Hungaricavol 56 no 2 pp 125ndash143 2009

[10] S Y Jayachandra S Anil Kumar D P Merley and MB Sulochana ldquoIsolation and characterization of extremehalophilic bacterium Salinicoccus sp JAS4 producing extra-cellular hydrolytic enzymesrdquo Recent Research in Science andTechnology vol 4 no 4 pp 46ndash49 2012

Chinese Journal of Biology 11

[11] I W Sutherland ldquoNovel and established applications of micro-bial polysaccharidesrdquo Trends in Biotechnology vol 16 no 1 pp41ndash46 1998

[12] MWMittelman andG G Geesey ldquoCopper-binding character-istics of exopolymers from a freshwater-sediment bacteriumrdquoApplied and EnvironmentalMicrobiology vol 49 no 4 pp 846ndash851 1985

[13] J Anton I Meseguer and F Rodriguez-Valera ldquoProductionof an extracellular polysaccharide by Haloferax mediterraneirdquoApplied Enviromental Microbiology vol 54 no 10 pp 2381ndash2386 1988

[14] H Parolis L A S Parolis I F Boan et al ldquoThe structure ofthe exopolysaccharide produced by the halophilic ArchaeonHaloferax mediterranei strain R4 (ATCC 33500)rdquo CarbohydrateResearch vol 295 pp 147ndash156 1996

[15] E Quesada V Bejar and C Calvo ldquoExopolysaccharide produc-tion by Volcaniella eurihalinardquo Experientia vol 49 no 12 pp1037ndash1041 1993

[16] J A Mata V Bejar I Llamas et al ldquoExopolysaccharides pro-duced by the recently described halophilic bacteria Halomonasventosae and Halomonas anticariensisrdquo Research in Microbiol-ogy vol 157 no 9 pp 827ndash835 2006

[17] I Llamas J A Mata R Tallon et al ldquoCharacterization ofthe exopolysaccharide produced by Salipiger mucosus A3119879 ahalophilic species belonging to the Alphaproteobacteria iso-lated on the Spanish Mediterranean seaboardrdquo Marine Drugsvol 8 no 8 pp 2240ndash2251 2010

[18] F Martınez-Checa E Quesada J Martınez-Canovas I Llamasand V Bejar ldquoPalleronia marisminoris gen nov sp nova moderately halophilic exopolysaccharide-producing bac-terium belonging to the rsquoAlphaproteobacteriarsquo isolated from asaline soilrdquo International Journal of Systematic and EvolutionaryMicrobiology vol 55 no 6 pp 2525ndash2530 2005

[19] A Ventosa M T Garcia M Kamekura H Onishi and FRuiz-Berraquero ldquoBacillus halophilus sp nov a moderatelyhalophilic Bacillus speciesrdquo Systematic and Applied Microbiol-ogy vol 12 no 2 pp 162ndash166 1989

[20] R A Moraine and P Rogovin ldquoKinetics of polysaccharide B1459 fermentationrdquoBiotechnology and Bioengineering vol 8 no4 pp 511ndash524 1966

[21] M Dubois K A Gilles J K Hamilton P A Rebers and FSmith ldquoColorimetric method for determination of sugars andrelated substancesrdquoAnalytical Chemistry vol 28 no 3 pp 350ndash356 1956

[22] O H Lowry N J Rosenbrough A L Farr and R J RandallldquoProtein measurement with the Folin phenol reagentrdquo TheJournal of Biological Chemistry vol 193 no 1 pp 265ndash275 1951

[23] P Gerhardt R G E Murray W A Wood and N R KriegMethods for General and Molecular Bacteriology AmericanSociety for Microbiology Washington DC USA 1994

[24] G M Garrity J A Bell and T G Lilburn ldquoTaxonomic outlineof the procaryotesrdquo inBergeyrsquosManual of Systematic BacteriologD R Boone and R W Castenholz Eds Springer New YorkNY USA 2nd edition 2003

[25] J A Mata J Martınez-Canovas E Quesada and V BejarldquoA detailed phenotypic characterisation of the type strains ofHalomonas speciesrdquo Systematic and Applied Microbiology vol25 no 3 pp 360ndash375 2002

[26] A Oren ldquoMolecular ecology of extremely halophilic archaeaand bacteriardquo FEMSMicrobiology Ecology vol 39 no 1 pp 1ndash72002

[27] H Baati R Amdouni N Gharsallah A Sghir and E AmmarldquoIsolation and characterization of moderately halophilic bacte-ria from tunisian solar salternrdquo Current Microbiology vol 60no 3 pp 157ndash161 2010

[28] A Hedi N Sadfi M Fardeau et al ldquoStudies on the biodiversityof halophilic microorganisms isolated from El-Djerid salt lake(Tunisia) under aerobic conditionsrdquo International Journal ofMicrobiology vol 2009 Article ID 731786 17 pages 2009

[29] M Birbir A Ogan B Calli and B Mertoglu ldquoEnzymecharacteristics of extremely halophilic archaeal community inTuzkoy Salt Mine Turkeyrdquo World Journal of Microbiology andBiotechnology vol 20 no 6 pp 613ndash621 2004

[30] M Birbir and C Sesal ldquoExtremely halophilic bacterial commu-nities in Sereflikochisar Salt Lake in Turkeyrdquo Turkish Journal ofBiology vol 27 no 7 pp 7ndash22 2003

[31] M Birbir B Calli B Mertoglu et al ldquoExtremely halophilicArchaea from Tuz Lake Turkey and the adjacent Kaldirim andKayacik salternsrdquoWorld Journal ofMicrobiology and Biotechnol-ogy vol 23 no 3 pp 309ndash316 2007

[32] R Rohban M A Amoozegar and A Ventosa ldquoScreeningand isolation of halophilic bacteria producing extracellularhydrolyses from Howz Soltan Lake Iranrdquo Journal of IndustrialMicrobiology and Biotechnology vol 36 no 3 pp 333ndash3402009

[33] M J Garabito M C Marquez and A Ventosa ldquoHalotolerantBacillus diversity in hypersaline environmentsrdquo Canadian Jour-nal of Microbiology vol 44 no 2 pp 95ndash102 1998

[34] A Ventosa A Ramos-Cormenzana and M Kocur ldquoModer-ately halophilic gram-positive cocci from hypersaline environ-mentsrdquo Systematic and Applied Microbiology vol 4 no 4 pp564ndash570 1983

[35] E Quesada A Ventosa F Rodriguez-Valera and A R Cor-menzana ldquoTypes and properties of some bacteria isolated fromhypersaline soilsrdquo Journal of Applied Bacteriology vol 53 no 2pp 155ndash161 1982

[36] E Quesada V Bejar M J Valderrama A Ventosa and AR Ramos Cormenzana ldquoIsolation and characterization ofmoderately halophilic nonmotile rods from different salinehabitatsrdquoMicrobiologia vol 1 no 1-2 pp 89ndash96 1985

[37] S R Dave and H B Desai ldquoMicrobial diversity at marinesalterns near Bhavnagar Gujarat Indiardquo Current Science vol60 no 4 pp 497ndash500 2006

[38] S Kumar R Karan S Kapoor S P Singh and S K KhareldquoScreening and isolation of halophilic bacteria producingindustrially important enzymesrdquo Brazilian Journal of Microbi-ology vol 43 no 4 pp 1595ndash1603 2012

[39] B P Dave and A Soni ldquoDiversity of halophilic archaea atsalt pans around Bhavnagar coast Gujaratrdquo Proceedings ofthe National Academy of Sciences India Section B BiologicalSciences vol 83 no 2 pp 225ndash232 2013

[40] A A Joshi P P Kanekar A S Kelkar et al ldquoCultivable bacterialdiversity of alkaline Lonar lake Indiardquo Microbial Ecology vol55 no 2 pp 163ndash172 2008

[41] T M Raghavan and I Furtado ldquoOccurrence of extremelyhalophilic Archaea in sediments from the continental shelf ofwest coast of Indiardquo Current Science vol 86 no 8 pp 1065ndash1067 2004

[42] S Vijayanand J Hemapriya J Selvin and S Kiran ldquoBiodiver-sity of extremely halophilic bacterial strains isolated from solarsalterns of Tuticorin Tamilnadu Indiardquo International Journal ofWater Resources and Environmental Sciences vol 1 no 1 pp 1ndash72012

12 Chinese Journal of Biology

[43] S G Nanjani and H P Soni ldquoIsolation and characterization ofextremely halotolerant and halophilic organisms from Dwarkaand Veravalrdquo Journal of Pharmacy and Biological Sciences vol2 no 2 pp 20ndash25 2012

[44] H Ghozlan H Deif R A Kandil and S Sabry ldquoBiodiversity ofmoderately halophilic bacteria in hypersaline habitats in EgyptrdquoJournal of General and Applied Microbiology vol 52 no 2 pp63ndash72 2006

[45] D R Arahal M T Garcıa W Ludwig K H Schleifer andA Ventosa ldquoTransfer ofHalomonas canadensis andHalomonasisraelensis to the genus Chromohalobacter as Chromohalobac-ter canadensis comb nov and Chromohalobacter israelensiscomb novrdquo International Journal of Systematic and EvolutionaryMicrobiology vol 51 no 4 pp 1443ndash1448 2001

Submit your manuscripts athttpwwwhindawicom

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Anatomy Research International

PeptidesInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporation httpwwwhindawicom

International Journal of

Volume 2014

Zoology

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Molecular Biology International

GenomicsInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

The Scientific World JournalHindawi Publishing Corporation httpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

BioinformaticsAdvances in

Marine BiologyJournal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Signal TransductionJournal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

BioMed Research International

Evolutionary BiologyInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Biochemistry Research International

ArchaeaHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

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Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

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Virolog y

Hindawi Publishing Corporationhttpwwwhindawicom

Nucleic AcidsJournal of

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Enzyme Research

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

International Journal of

Microbiology

Page 3: Research Article Production of Extracellular …downloads.hindawi.com/archive/2014/205731.pdfResearch Article Production of Extracellular Polymeric Substances by Halophilic Bacteria

Chinese Journal of Biology 3

Table 1 Bacterial population of soil samples collected from solar salterns of Gujarat Orissa and West Bengal India

State Sampling sites Number of samplesBacterial count (cfug of soil times102)

NaCl () in medium5 10 15

Gujarat

Jogrinar 4 074ndash866 021ndash366 011ndash4Kandla Port 3 086ndash735 121ndash211 008ndash0Khari Rohar 3 239ndash768 012ndash178 047ndash966Victor Port 2 173ndash283 040ndash0 mdash

Orissa Humma 5 013ndash061 025ndash055 mdashSurala 5 1ndash366 007ndash023 mdash

West Bengal Baksal 4 2ndash390 020ndash040 020ndash0Dadanpatrabar 5 27ndash79 007ndash060 007ndash030

Table 2 Bacterial population of water samples collected from solar salterns of Gujarat Orissa and West Bengal India

States Sampling sites Number of samplesBacterial count (cfumL of water times102)

NaCl () in medium5 10 15

Gujarat

Jogrinar 2 028ndash556 172ndash0 040ndash0Kandla Port 2 053ndash540 016ndash11 066ndash0Khari Rohar 1 023ndash0 mdash mdashVictor Port 2 126ndash190 mdash mdash

Orissa Humma 3 023ndash23 mdash mdashSurala 2 027ndash120 083ndash0 mdash

West Bengal Baksal 3 110ndash136 013ndash06 010ndash037Dadanpatrabar 3 05ndash190 007ndash050 01-02

rods and produced white to cream colored circular smooth-edged colonies on MH agar In general the isolates werecapable of tolerating wide range of temperature and pH fortheir growth

32 Screening for EPS Production EPS producing abilityof these isolates was examined in batch culture using MYmedium supplemented with 5 NaCl and the soluble EPScontent of each of the isolates was evaluated in terms ofits carbohydrate content as determined by Dubois method[21] Among the 410 soil isolates majority (184) producedsoluble EPS ranging from 05 to 07 gL in terms of theircarbohydrate content while only 15 isolates produced morethan 1 gL of EPS (Table 5) On the contrary among the177 bacterial strains isolated from saline water samples only5 produced soluble EPS accounting for more than 1 gL ofcarbohydrate However majority (77) of them produced 05ndash07 gL of soluble EPS (Table 6)

In all 20 potent isolates producing gt1 gL of EPS werefurther allowed to grow in MY medium under continuousshaking (120 rpm) and the soluble EPS produced after 8days of growth was estimated following the method asdescribed above Kinetics of growth and EPS productionrevealed that EPS production by these isolates increasedwith biomass formation (Figure 1) and a few of the selectedisolates such as SUR202 SUR307 SUR310 JW307 and

JS904 appeared to be promising with an EPS yield of168ndash185 gL

33 Characterization and Identification of Selected BacterialIsolates Morphological and physiological studies revealed(Table 7) that these halophilic bacteria (with the exceptionof isolate SUR303) formed cream colored smooth colonieson MH agar medium Most of them were Gram-negativemotile rods only three isolates were Gram-positive one ofthem (isolate SUS303) was coccus while the isolate KW203was the only endospore former The isolates were capableof tolerating 15ndash20 NaCl in the medium and a pH of 5ndash11 Optimum growth was observed at a temperature of 32ndash37∘C but all were able to tolerate a temperature as high as40∘C

Analysis of biochemical characters (Table 8) showedthat the majority of these halophilic isolates gave negativeresponse to MR-VP tests and failed to produce extracellularenzymes like amylase cellulase pectinase inulinase gelati-nase lipase caseinase xylanase and urease None of theseisolates however were capable of producing H

2S lysine-

arginine- and ornithine decarboxylaseCarbon source utilization pattern (Table 9) of these

halophilic isolates varied considerably All the isolates wereable to utilize ribose fructose mannitol salicin cellobioseacetate benzoate and succinate and three of them (JS803

4 Chinese Journal of Biology

Table 3 Bacterial isolates obtained from soil samples of solar salterns of Gujarat Orissa and West Bengal India

States Sampling sites Number of samplesNumber of bacterial isolates

NaCl () in medium5 10 15

Gujarat

Jogrinar 4 44 23 4Kandla Port 3 19 12 4Khari Rohar 3 46 16 6Victor Port 2 20 5 mdash

Orissa Humma 5 48 14 mdashSurala 5 31 31 2

West Bengal Baksal 4 19 8 3Dadanpatrabar 5 29 16 10

Total 31 256 125 29

Table 4 Bacterial isolates obtained from water samples of solar salterns of Gujarat Orissa and West Bengal India

States Sampling sites Number of samplesNumber of bacterial isolates

NaCl () in medium5 10 15

Gujarat

Jogrinar 2 21 14 5Kandla Port 2 19 5 2Khari Rohar 1 1 3 mdashVictor Port 2 26 mdash 1

Orissa Humma 3 16 mdash mdashSurala 2 12 8 3

West Bengal Baksal 3 9 3 4Dadanpatrabar 3 17 4 4

Total 18 121 37 19

SUR301 and SUR307) appeared to be versatile in utilizingall 30 carbon sources Fermentation pattern of these strainsvaried remarkably the majority were unable to fermentmost of the carbon sources On the contrary fructosesorbitol and benzoate were fermented by most of the isolates(Table 9)

Sensitivity of these isolates to 22 different antibioticswas tested by disc-diffusion method (Table 10) and theantibiotic resistance index (ARI) was determined (Figure 2)Themajority of the isolates were sensitive to chloramphenicol(30 120583g) gentamycin (10 120583g) and norfloxacin (10 120583g) followedby ampicillin (10 120583g) Resistance to vancomycin (30120583g) fol-lowed by trimethoprim (30 120583g) was predominant amongstthe tested halophiles As judged by the ARI values the isolateSUR302 was the most resistant one (ARI = 065) followed bySUR301 KW203 and KW1805 while the lowest ARI (023)was indicated by the isolate JW307 which was most sensitiveto the tested antibiotics

The morphological physiological and biochemical char-acters including the carbon source utilization and fermen-tation patterns along with antibiotic susceptibility wereanalyzed and compared with the phenotypic characters ofhalophilic bacterial genera [5 18] so far reported According

to the phenotypic and biochemical characteristics 70 of theselected isolates (14) were tentatively identified asmembers ofthe genusHalomonasThe onlyGram-positive coccus (isolateSUR303) and the rod shaped endospore forming isolate(isolate KW203) were assigned to Salinicoccus and Bacillusrespectively Two of the isolates (KS1805 and SUR301) wereplaced in the genus Chromohalobacter while the remain-ing two (isolates JS504 and JS904) were included underAlteromonas and Aidingimonas respectively

4 Discussion

Halophiles have mainly been isolated from wide diversity ofenvironments such as saltern crystallizer ponds the DeadSea solar lakes and hypersaline lakes [26] Culture depen-dent diversity studies on halophiles have been made fromTunisian solar saltern [27 28] Tuzkoy salt mine [29] Sere-flikochisar Salt Lake [30] Kaldirim and Kayacik of Tuz Lake[31] Turkey Howz Soltan Lake Iran [32] and hypersalineenvironments in South Spain [33ndash36] In the Indian contextthe halophilic diversity studies have been restricted mainlyto the marine salterns of Bhavnagar [37ndash40] Lonar Lake[40] and Peninsular Coast [41 42]The present study reports

Chinese Journal of Biology 5

Table 5 Screening of halophilic bacterial isolates derived from soil samples for the production of extracellular polymeric substances

States Sampling sites Number of isolates Extracellular polymeric substances (gL)gt050 051ndash070 071ndash10 gt10

Gujarat

Jogrinar 71 18 42 7 4Kandla Port 35 13 13 7 2Khari Rohar 68 26 31 11 mdashVictor Port 25 20 3 2 mdash

Orissa Humma 62 22 37 3 mdashSurala 64 15 25 15 9

West Bengal Baksal 30 14 12 4 mdashDadanpatrabar 55 32 21 2 mdash

Total 410 160 184 51 15

Table 6 Screening of halophilic bacterial isolates derived from water samples for the production of extracellular polymeric substances

States Sampling sites Number of isolates Extracellular polymeric substances (gL)gt050 051ndash070 071ndash10 gt10

Gujarat

Jogrinar 40 26 9 3 2Kandla Port 26 13 11 1 1Khari Rohar 4 1 3 mdash mdashVictor Port 27 7 17 1 2

Orissa Humma 16 4 11 1 mdashSurala 23 8 6 9 mdash

West Bengal Baksal 16 7 6 3 mdashDadanpatrabar 25 8 14 3 mdash

Total 177 74 77 21 5

the distribution of halophilic and halotolerant bacterialcommunities in the inland multipond solar salterns spreadover the coasts of India Halophilic bacterial communitiesof 31 soil and 18 water samples from 8 different sites wereanalyzed by dilution and plating method (Tables 1 and 2) andprovide information regarding the availability anddiversity ofhalophilic bacteria in the solar saltern ponds Colony formingunits of soil and water samples revealed that soil samplesholdmore viable bacterial counts than water samples Ragha-van and Furtado [41] studied the occurrence of extremelyhalophilic archaea in sediments from the continental shelf ofwest coast of India and reported the presence of relatively lowaverage counts (7ndash5 times 103) of extreme halophiles in offshoresediments in contrast to the very high counts (105ndash109)of marine eubacteria Similar study by Joshi et al [40]reported that the total numbers of microorganisms in the soiland water samples were 102ndash106 cfug and 102ndash104 cfumLrespectively

A total of 587 halophilic and halotolerant bacterial strainswere isolated showing different degrees of NaCl tolerance(Tables 3 and 4) and supported the observations of Quesadaet al [35] andVentosa et al [34] However during the presentstudy we were unable to isolate extreme halophiles whichhave frequently been identified as the dominant phylotypes inhypersaline environments along with solar salterns of India

[39 41] Furthermore soil samples showed more bacterialdiversity than water ones which is in accordance with theobservations of Joshi et al [40]

During the course of screening of the moderatelyhalophilic isolates for EPS production (Tables 5 and 6) only20 isolates appeared to be promising with significant yield ofEPS (10ndash185 gL) (Figure 1) Joshi and coworkers [40] sim-ilarly screened 86 halophilic bacteria from Lonar Lake andreported Halomonas campisalis and Vagococcus carniphilusas potent EPS producers Similarly Nanjani and Soni [43]also isolated 73 halotolerant and halophilic bacteria from soilsamples of Veraval and Dwarka 23 of them produced EPSranging from 02 to 1060 gL In addition reports on EPSproduction by moderately halophilic bacteria of the genusHalomonas are not uncommon [8 15ndash17]

Attempts have been made to determine the taxonomicidentity of all 20 promising EPS producing isolates followingdetailed physiobiochemical characterization (Tables 7ndash9) andcomparison with Bergeyrsquos Manual of Systematic Bacteriology[24] and those of Mata et al [25]Themajority (14 isolates) ofthem were Gram-negative nonsporulating rods and capableof growing in 25ndash20NaCl similar to those ofHalomonas asreported by Quesada et al [36] Ghozlan et al [44] andMataet al [25] However the Gram-positive endospore formingisolate KW203 was assigned to the genus Bacillus

6 Chinese Journal of Biology

Table7Morph

ologicalandph

ysiologicalcharacteristicso

fthe

selected

EPSprod

ucinghaloph

ilicb

acteria

isolated

from

solarsalternso

fInd

ia

Character

Bacterialisolates

JS504

JS802

JS803

JS904

JW302

JW307

KS110

1KS

1805

KW203

VW301

Morph

ologicalcharacters

Colon

ymorph

olog

yCr

eamcirc

ular

Whitecirc

ular

Creamcirc

ular

Creamcirc

ular

Transparentcircular

Creamcirc

ular

Creamcirc

ular

Whitecirc

ular

Whitecirc

ular

Creamcirc

ular

Gram

nature

minus+

minusminus

minusminus

minusminus

+minus

Cellshape

Rod

Rod

Rod

Rod

Rod

Rod

Rod

Rod

Rod

Rod

Cell

arrang

ement

Sing

leSing

leSing

leSing

leSing

lechains

Sing

lechains

Sing

lechains

Sing

leSing

leclu

mped

Sing

lechains

Motility

++

++

++

minusminus

++

Pigm

entatio

nminus

minusminus

minusminus

minusminus

minusminus

minus

Endo

sporeformation

minusminus

minusminus

minusminus

minusminus

+minus

Physiologicalcharacters

pHrang

efor

grow

th5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

pHop

timum

forg

rowth

119

89

1111

118

89

Temprang

efor

grow

th(∘C)

27ndash4

027ndash4

027ndash4

027ndash4

027ndash4

027ndash4

022ndash4

027ndash4

027ndash4

022ndash4

0Tempop

timum

forg

rowth

(∘C)

3237

3732

3232

3232

3732

NaC

lrange

forg

rowth

()

0ndash15

0ndash20

0ndash15

0ndash20

0ndash15

0ndash20

0ndash20

25ndash15

25ndash15

0ndash20

NaC

loptim

umforg

rowth

()

510

105

105

575

105

Growth

onKingrsquosBmedium

++

++

++

++

++

Growth

onMcC

onky

agar

minusminus

minusminus

minusminus

minusminus

minusminus

Tentativeidentity

Alteromonas

Halom

onas

Halom

onas

Aidingim

onas

Halom

onas

Halom

onas

Halom

onas

Chromohalobacter

Bacillus

Halom

onas

Character

Bacterialisolates

VW402

SUR108

SUR3

01SU

R302

SUR3

03SU

R307

SUR3

09SU

R310

SUR2

02SU

R201

Morph

ologicalcharacters

Colon

ymorph

olog

yCr

eamcirc

ular

Creamcirc

ular

Whitegum

my

Transparentcircular

Flatreddish-pink

Creamcirc

ular

Creamcirc

ular

Creamcirc

ular

Creamcirc

ular

Creamcirc

ular

Gram

nature

minusminus

minusminus

+minus

minusminus

minusminus

Cellshape

Rod

Rod

Rod

Rod

Cocci

Rod

Rod

Rod

Rod

Rod

Cell

arrang

ement

Sing

leSing

leSing

leSing

leSing

leclu

mped

Sing

leSing

leSing

leSing

leSing

leMotility

+minus

minus+

minus+

minus+

minusminus

Pigm

entatio

nminus

minusminus

minusPink

minusminus

minusminus

minus

Endo

sporeformation

minusminus

minusminus

minusminus

minusminus

minusminus

Physiologicalcharacters

pHrang

efor

grow

th5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

pHop

timum

forg

rowth

97

77

117

77

79

Temprang

efor

grow

th(∘C)

27ndash4

027ndash4

027ndash4

032ndash4

532ndash4

522ndash4

032ndash4

022ndash4

522ndash4

532ndash4

5Tempop

timum

forg

rowth

(∘C)

3232

3732

3727

3227

2732

NaC

lrange

forg

rowth

()

0ndash15

0ndash15

25ndash15

25ndash15

25ndash15

0ndash20

0ndash20

0ndash20

0ndash20

0ndash15

NaC

loptim

umforg

rowth

()

525

575

525

7525

575

Growth

onKingrsquosBmedium

++

++

++

++

++

Growth

onMcC

onky

agar

minusminus

minusminus

minusminus

minusminus

minusminus

Tentativeidentity

Halom

onas

Halom

onas

Chromohalobacter

Halom

onas

Salin

icoccus

Halom

onas

Halom

onas

Halom

onas

Halom

onas

Halom

onas

Chinese Journal of Biology 7

Table8Biochemicalcharacteris

ticso

fthe

selected

EPSprod

ucinghaloph

ilicb

acteria

isolated

from

solarsalternso

fInd

ia

Character

Bacterialisolates

JS 504

JS 802

JS 803

JS 904

JW 302

JW 307

KS 1101

KS 1805

KW 203

VW 301

VW 402

SUR

108

SUR

301

SUR

302

SUR

303

SUR

307

SUR

309

SUR

310

SUR

202

SUR

201

Biochemicalcharacters

Voges-Proskauertest

minusminus

minusminus

minusminus

minusminus

+minus

minusminus

minusminus

minusminus

minusminus

minusminus

Citrateu

tilization

+minus

minusminus

minus+

minus+

minus+

++

+minus

minus+

++

++

Methylred

test

minusminus

minusminus

minusminus

minus+

+minus

minusminus

minusminus

minusminus

minusminus

minusminus

Prod

uctio

nof

Glutaminase

++

++

++

++

++

++

++

minus+

++

++

Asparaginase

++

++

++

++

++

++

++

minus+

++

++

Amylase

+minus

minusminus

minusminus

minusminus

+minus

minusminus

minusminus

minusminus

minusminus

minusminus

Cellulase

minusminus

minusminus

minusminus

minusminus

+minus

minusminus

minusminus

minusminus

minusminus

minusminus

Pectinase

+minus

minusminus

minusminus

minusminus

+minus

minusminus

minusminus

minusminus

minusminus

minusminus

Inulinase

+minus

minusminus

minusminus

minusminus

+minus

minusminus

minusminus

minusminus

minusminus

minusminus

Gelatinase

minusminus

minusminus

minusminus

minusminus

+minus

minusminus

minusminus

minusminus

minusminus

minusminus

Lipase

+minus

minusminus

minusminus

minus+

+minus

minusminus

+minus

minusminus

minusminus

minusminus

Xylanase

+minus

minusminus

minusminus

minusminus

+minus

minusminus

minusminus

minusminus

minusminus

minusminus

Caseinase

minusminus

minus+

minusminus

minusminus

minusminus

minusminus

minus+

minusminus

minusminus

minusminus

Urease

minusminus

minus+

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

Catalase

++

++

++

++

++

++

++

++

++

++

Nitrater

eductase

++

++

++

++

++

++

++

++

++

++

H2S

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

Lysin

edecarbo

xylase

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

Arginined

ecarbo

xylase

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

Ornith

ined

ecarbo

xylase

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

Indo

leminus

minusminus

minusminus

minusminus

+minus

minusminus

minus+

minus+

minusminus

minusminus

minus

Phenylalanined

eaminase

++

+minus

+minus

++

minus+

minus+

++

++

++

minus+

Tryptoph

andeam

inase

++

++

++

++

++

minusminus

++

++

++

+minus

8 Chinese Journal of Biology

Table9Ca

rbon

source

utilizatio

nandferm

entatio

nof

thes

elected

EPSprod

ucinghaloph

ilicb

acteria

isolated

from

samples

ofsolarsalternso

fInd

ia

Carbon

source

Bacterialisolates

JS 504

JS 802

JS 803

JS 904

JW 302

JW 307

KS 1101

KS 1805

KW 203

VW

301

VW

402

SUR

108

SUR

301

SUR

302

SUR

303

SUR

307

SUR

309

SUR

310

SUR

202

SUR

201

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

Glucose

++

++

+minusminus

++minus

+minus

+minus

++

++

+minus

+minus

+minus

+minusminus

++

++minus

+minus

+minusminusminusminusminus

Galactose

+minus

+minus

+minusminus

++minus

+minus

+minus

++

+minus

+minus

+minus

+minus

++minusminus

+minus

++

+minus

+minus

+minus

+minus

Arabino

seminusminusminusminus

+minusminusminusminusminus

+minus

+minus

++

++

+minus

+minus

+minus

+minusminus

++minus

+minus

+minusminusminus

+minus

+minus

Manno

se+minus

+minus

+minusminus

++minus

+minus

+minusminus

++

++minus

+minus

+minus

+minusminusminusminusminus

+minus

+minus

+minus

+minus

+minus

Rham

nose

+minus

+minus

+minus

+minusminusminusminusminusminusminus

++

+minusminusminus

+minus

+minus

++minus

++minus

+minusminusminusminusminusminusminus

+minus

Trehalose

+minus

+minus

+minusminus

+minus

+minusminusminusminus

+minus

++minusminusminusminus

+minus

+minusminus

++

++minusminusminusminusminusminusminus

+minus

Lactose

minusminusminusminus

+minusminusminus

+minus

+minus

+minus

+minus

+minus

+minus

+minusminusminus

+minus

++minusminus

+minusminusminusminusminus

+minus

+minus

Sucrose

+minus

+minus

+minus

++

++

+minus

+minus

+minus

++

+minus

+minusminusminus

+minus

++

+minus

+minus

+minus

+minusminusminus

+minus

Xylose

minusminusminusminus

+minus

+minusminusminus

+minus

++minus

+minusminus

+minus

+minus

+minus

+minusminus

+minus

++

+minusminusminusminus

+minusminusminus

Ribo

se+minus

++

++

++

+minus

+minus

+minus

++

++

+minus

+minus

+minus

++

++

++

+minus

+minus

+minus

+minus

+minus

Fructose

+minus

++

+minus

++

+minus

++

++

+minus

++

++

++

++

++

++

++

++

++

++

++

++

Maltose

minusminusminusminus

+minusminus

+minus

++minus

+minus

+minus

++

+minus

+minus

+minus

++minus

+minus

++minusminusminusminusminus

+minus

+minus

Glycine

minusminusminus

++minus

+minusminus

+minusminusminusminus

+minusminusminusminusminusminusminusminusminus

+minusminusminusminusminus

+minus

+minusminusminusminus

+minusminus

Glycerol

+minus

+minus

+minusminus

++

++minus

+minus

+minus

++

+minus

+minus

+minus

+minusminus

++

++minus

+minus

+minus

+minus

+minus

Adon

itol

+minus

+minus

+minus

+minusminusminus

+minus

+minus

+minus

+minus

+minus

+minusminusminus

+minus

++

+minus

+minus

+minus

+minusminusminusminusminus

Salicin

+minus

++

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

++

++

+minus

+minus

+minus

+minus

+minus

Meso-inosito

l+minusminusminus

+minus

+minus

+minus

+minus

++

+minus

++

+minus

+minusminusminus

+minusminus

++minus

+minus

+minus

+minus

+minus

+minus

Mannitol

+minus

+minus

+minus

++

+minus

+minus

+minus

+minus

++

+minus

+minus

+minus

+minus

+minus

++

+minus

+minus

+minus

+minus

+minus

Sorbito

l+

++

++minusminus

++

+minus

++

++

++

++

++

++

++

+minus

+minus

++

++minus

+minus

++

+minus

Raffino

seminusminus

++

++minusminus

+minus

+minus

+minus

+minus

++minusminusminusminusminusminusminusminus

++

++

+minus

+minus

+minus

+minus

+minus

Cellobiose

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

++

+minus

+minus

+minus

+minus

++

+minus

+minus

+minus

+minus

+minus

+minus

Melibioseminusminusminusminusminusminusminusminusminusminusminusminusminusminusminusminus

++minusminusminusminusminusminusminusminus

++minusminusminusminusminusminusminusminusminusminusminusminus

Na-acetate

+minus

+minus

+minus

+minus

++

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

++

++

+minus

+minus

+minus

+minus

+minus

Na-citrate

+minus

++

+minusminusminusminus

+minusminusminusminusminusminusminusminusminusminusminusminusminusminusminusminusminus

+minusminusminusminusminusminusminusminusminusminusminusminus

Na-fumarate

+minus

+minus

+minus

++

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minusminus

++minus

+minus

+minus

+minus

+minus

+minus

Na-form

ate

+minus

+minus

+minus

+minus

+minusminusminus

+minus

+minus

+minusminusminus

+minusminusminus

+minus

++minusminus

+minusminusminusminusminusminusminusminusminus

Na-benzoate

+minus

++

++

+minus

+minus

+minus

++

++

++

+minus

+minus

++

++

+minus

+minus

+minus

++

++

++

++

Na-malateminusminusminusminusminusminus

++

+minusminusminusminusminusminusminusminusminusminusminusminusminusminusminusminusminus

++

++minusminusminusminusminusminusminusminusminusminus

Na-prop

ionate

+minus

+minus

+minusminusminus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

++minus

++minus

+minus

+minus

+minus

+minus

Na-succinate

+minus

+minus

+minus

++

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

++

++

+minus

+minus

+minus

+minus

+minus

Uutilization

Fferm

entatio

n

Chinese Journal of Biology 9

Table10A

ntibiotic

sensitivitypatte

rnof

thes

electedEP

Sprod

ucinghaloph

ilicb

acteria

isolated

from

solarsalternso

fInd

ia

Antibiotics

Bacterialisolates

JS 504

JS 802

JS 803

JS 904

JW 302

JW 307

KS 1101

KS 1805

KW 203

VW 301

VW 402

SUR

108

SUR

301

SUR

302

SUR

303

SUR

307

SUR

309

SUR

310

SUR

202

SUR

201

Ampicillin(10120583

g)S

SR

SS

SS

RI

SS

SR

SS

SS

SS

SAmoxicillin

(30120583

g)S

SR

RS

SS

RS

SS

SR

RS

IR

SS

SBa

citracin

(10120583

g)S

SR

SS

SR

RS

IR

RR

RS

IR

RR

RCiprofl

oxacin

(30120583

g)I

II

RI

II

IR

II

II

IR

SR

II

ICh

lorte

tracyclin

e(30120583g)

SI

SS

IS

RR

SS

SS

RR

SS

II

IS

Chloramph

enicol(10120583

g)S

SS

SS

SS

SR

SS

SS

SS

SR

SS

SErythrom

ycin

(15120583

g)S

SI

SS

IS

II

II

II

SR

RI

IS

SFu

sidicacid

(30120583

g)I

RR

SS

IR

RS

RI

RR

RS

IR

II

RGentamycin

(10120583

g)S

SS

SS

SS

SR

SS

SS

SS

IS

SS

SKa

namycin

(30120583

g)I

IS

RS

II

SR

II

RI

RR

RI

RI

RMethicillin(5120583g)

SS

RS

IR

RR

RR

RR

RR

SR

RR

RR

Neomycin

(30120583

g)S

SI

RI

IR

II

IR

II

RR

RI

IR

RNorflo

xacin(30120583

g)S

SR

SS

SS

SI

SS

SS

SS

SS

SS

SNetillin

(10120583

g)R

SS

IR

SS

SR

RR

SS

RR

RR

RR

RNalidixicacid

(30120583

g)R

RS

RR

II

RR

II

RR

RR

SR

RR

RNovob

iosin

(30120583

g)I

RR

SR

SS

RS

SS

IR

SS

SI

SS

SPo

lymixin

B(50120583

g)R

RI

RI

IR

RR

RI

IR

RR

II

II

SPenicillinG(10120583

g)S

SR

IS

IR

RR

II

SR

RS

II

II

SStreptom

ycin

(10120583

g)S

SS

RS

RR

SR

RR

IS

RS

SR

II

RTetracyclin

e(30120583g)

RR

RR

RI

RR

SR

IR

RR

RS

RR

RI

Trim

etho

prim

e(30120583g)

SS

SS

IS

SI

SS

SS

IR

RS

SS

SS

Vancom

ycin

(30120583

g)R

RR

SR

RR

RS

RR

RR

RS

RR

RR

R

S=SensitiveR

=Re

sistantI

=Interm

ediate

10 Chinese Journal of Biology

0

1

2

3

4

Bacterial isolate

Gro

wth

OD

at540

nmto

tal E

PS (g

L)

OD at 540nmTotal EPS

JS504

JS802

JS803

JS904

JW302

JW307

KS1101

KS1805

KW203

VW301

VW402

SUR1

08

SUR3

01

SUR3

02

SUR3

03

SUR3

07

SUR3

09

SUR3

10

SUR2

02

SUR2

01

Figure 1 Screening of moderately halophilic bacteria isolatedfrom soil and water samples of solar salterns for production ofextracellular polymeric substances

0

02

04

06

08

1

Ant

ibio

tic re

sista

nce i

ndex

Bacterial isolate

JS504

JS802

JS803

JS904

JW302

JW307

KS1101

KS1805

KW203

SUR1

08

SUR2

01

SUR2

02

SUR3

01

SUR3

02

SUR3

03

SUR3

07

SUR3

09

SUR3

10

VW301

VW402

Figure 2 Antibiotic resistance index of the selected EPS producinghalophilic bacteria isolated from soil and water samples of solarsalterns of India

The isolates SUR301 and KS 1805 were tentatively iden-tified as Chromohalobacter based on the study by Arahal etal [45] and bear striking similarity with those of Chromo-halobacter sp isolated fromhypersaline soil sample of TriveniSangam Gujarat [38] The physiological morphological andbiochemical characteristics of isolate SUR303 were consistentwith the features of the genus Salinicoccus [10] while isolatesJS904 and JS504 were tentatively assigned to Aidingimonasand Alteromonas respectively

Finally sensitivity of these isolates to antibiotics (Table 10and Figure 2) also corroborates the findings ofMata et al [25]and Hedi et al [28]

5 Conclusion

It may be emphasized that study of these moderatelyhalophilic bacteria from coastal hypersaline solar salterns ofIndia with special attempt on screening for EPS productionhas led to the discovery of wide variety of halophilic speciesAlthough attention has been focused on the production

of EPS by these halophilic bacterial isolates their physio-biochemical features indicate that they may equally holdpotential towards production of various biomolecules ofindustrial interest

Conflict of Interests

It is declared by the authors that there is no conflict of interestsregarding the publication of this paper

Acknowledgment

This study was financially supported by grants from Uni-versity Grants Commission India (Sanction no F14-2(SC)2008 (SA-III) 31 March 2009) under the Scheme ofRajiv Gandhi National Fellowship

References

[1] E A Galinski and B J Tindall ldquoBiotechnological prospectsfor halophiles and halotolerant microorganismsrdquo in MolecularBiology and Biotechnology of Extremophiles R D Herbert andR J Sharp Eds Blackie London UK 1992

[2] R Margesin and F Schinner ldquoPotential of halotolerant andhalophilic microorganisms for biotechnologyrdquo Extremophilesvol 5 no 2 pp 73ndash83 2001

[3] F Rodriguez Valera F Ruiz Berraquero and A Ramos Cor-menzana ldquoCharacteristics of the heterotrophic bacterial pop-ulations in hypersaline environments of different salt concen-trationsrdquoMicrobial Ecology vol 7 no 3 pp 235ndash243 1981

[4] F Rodriguez-Valera ldquoCharacteristics and microbial ecol-ogy of hypersaline environmentsrdquo in Halophilic Bacteria FRodriguez-Valera Ed vol 1 pp 3ndash30 CRC Press Boca RatonFla USA 1988

[5] D J Kushner and M Kamekura ldquoPhysiology of halophiliceubacteriardquo inHalophilic Bacteria F Rodriguez-Valera Ed vol1 pp 109ndash138 CRC press Boca Raton Fla USA 1988

[6] M A Amoozegar F Malekzadeh and K AMalik ldquoProductionof amylase by newly isolated moderate halophile Halobacillussp strainMA-2rdquo Journal of Microbiological Methods vol 52 no3 pp 353ndash359 2003

[7] C O Jeon J Lim J Lee et al ldquoLentibacillus salarius spnov isolated from saline sediment in China and emendeddescription of the genus Lentibacillusrdquo International Journal ofSystematic and Evolutionary Microbiology vol 55 no 3 pp1339ndash1343 2005

[8] S Bouchotroch E Quesada A Del Moral I Llamas andV Bejar ldquoHalomonas maura sp nov a novel moderatelyhalophilic exopolysaccharide-producing bacteriumrdquo Interna-tional Journal of Systematic and Evolutionary Microbiology vol51 no 5 pp 1625ndash1632 2001

[9] A Biswas A Patra and A Paul ldquoProduction of poly-3-hydroxyalkanoic acids by a moderately halophilic bacteriumHalomonas marina HMA 103 isolated from solar saltern ofOrissa Indiardquo Acta Microbiologica et Immunologica Hungaricavol 56 no 2 pp 125ndash143 2009

[10] S Y Jayachandra S Anil Kumar D P Merley and MB Sulochana ldquoIsolation and characterization of extremehalophilic bacterium Salinicoccus sp JAS4 producing extra-cellular hydrolytic enzymesrdquo Recent Research in Science andTechnology vol 4 no 4 pp 46ndash49 2012

Chinese Journal of Biology 11

[11] I W Sutherland ldquoNovel and established applications of micro-bial polysaccharidesrdquo Trends in Biotechnology vol 16 no 1 pp41ndash46 1998

[12] MWMittelman andG G Geesey ldquoCopper-binding character-istics of exopolymers from a freshwater-sediment bacteriumrdquoApplied and EnvironmentalMicrobiology vol 49 no 4 pp 846ndash851 1985

[13] J Anton I Meseguer and F Rodriguez-Valera ldquoProductionof an extracellular polysaccharide by Haloferax mediterraneirdquoApplied Enviromental Microbiology vol 54 no 10 pp 2381ndash2386 1988

[14] H Parolis L A S Parolis I F Boan et al ldquoThe structure ofthe exopolysaccharide produced by the halophilic ArchaeonHaloferax mediterranei strain R4 (ATCC 33500)rdquo CarbohydrateResearch vol 295 pp 147ndash156 1996

[15] E Quesada V Bejar and C Calvo ldquoExopolysaccharide produc-tion by Volcaniella eurihalinardquo Experientia vol 49 no 12 pp1037ndash1041 1993

[16] J A Mata V Bejar I Llamas et al ldquoExopolysaccharides pro-duced by the recently described halophilic bacteria Halomonasventosae and Halomonas anticariensisrdquo Research in Microbiol-ogy vol 157 no 9 pp 827ndash835 2006

[17] I Llamas J A Mata R Tallon et al ldquoCharacterization ofthe exopolysaccharide produced by Salipiger mucosus A3119879 ahalophilic species belonging to the Alphaproteobacteria iso-lated on the Spanish Mediterranean seaboardrdquo Marine Drugsvol 8 no 8 pp 2240ndash2251 2010

[18] F Martınez-Checa E Quesada J Martınez-Canovas I Llamasand V Bejar ldquoPalleronia marisminoris gen nov sp nova moderately halophilic exopolysaccharide-producing bac-terium belonging to the rsquoAlphaproteobacteriarsquo isolated from asaline soilrdquo International Journal of Systematic and EvolutionaryMicrobiology vol 55 no 6 pp 2525ndash2530 2005

[19] A Ventosa M T Garcia M Kamekura H Onishi and FRuiz-Berraquero ldquoBacillus halophilus sp nov a moderatelyhalophilic Bacillus speciesrdquo Systematic and Applied Microbiol-ogy vol 12 no 2 pp 162ndash166 1989

[20] R A Moraine and P Rogovin ldquoKinetics of polysaccharide B1459 fermentationrdquoBiotechnology and Bioengineering vol 8 no4 pp 511ndash524 1966

[21] M Dubois K A Gilles J K Hamilton P A Rebers and FSmith ldquoColorimetric method for determination of sugars andrelated substancesrdquoAnalytical Chemistry vol 28 no 3 pp 350ndash356 1956

[22] O H Lowry N J Rosenbrough A L Farr and R J RandallldquoProtein measurement with the Folin phenol reagentrdquo TheJournal of Biological Chemistry vol 193 no 1 pp 265ndash275 1951

[23] P Gerhardt R G E Murray W A Wood and N R KriegMethods for General and Molecular Bacteriology AmericanSociety for Microbiology Washington DC USA 1994

[24] G M Garrity J A Bell and T G Lilburn ldquoTaxonomic outlineof the procaryotesrdquo inBergeyrsquosManual of Systematic BacteriologD R Boone and R W Castenholz Eds Springer New YorkNY USA 2nd edition 2003

[25] J A Mata J Martınez-Canovas E Quesada and V BejarldquoA detailed phenotypic characterisation of the type strains ofHalomonas speciesrdquo Systematic and Applied Microbiology vol25 no 3 pp 360ndash375 2002

[26] A Oren ldquoMolecular ecology of extremely halophilic archaeaand bacteriardquo FEMSMicrobiology Ecology vol 39 no 1 pp 1ndash72002

[27] H Baati R Amdouni N Gharsallah A Sghir and E AmmarldquoIsolation and characterization of moderately halophilic bacte-ria from tunisian solar salternrdquo Current Microbiology vol 60no 3 pp 157ndash161 2010

[28] A Hedi N Sadfi M Fardeau et al ldquoStudies on the biodiversityof halophilic microorganisms isolated from El-Djerid salt lake(Tunisia) under aerobic conditionsrdquo International Journal ofMicrobiology vol 2009 Article ID 731786 17 pages 2009

[29] M Birbir A Ogan B Calli and B Mertoglu ldquoEnzymecharacteristics of extremely halophilic archaeal community inTuzkoy Salt Mine Turkeyrdquo World Journal of Microbiology andBiotechnology vol 20 no 6 pp 613ndash621 2004

[30] M Birbir and C Sesal ldquoExtremely halophilic bacterial commu-nities in Sereflikochisar Salt Lake in Turkeyrdquo Turkish Journal ofBiology vol 27 no 7 pp 7ndash22 2003

[31] M Birbir B Calli B Mertoglu et al ldquoExtremely halophilicArchaea from Tuz Lake Turkey and the adjacent Kaldirim andKayacik salternsrdquoWorld Journal ofMicrobiology and Biotechnol-ogy vol 23 no 3 pp 309ndash316 2007

[32] R Rohban M A Amoozegar and A Ventosa ldquoScreeningand isolation of halophilic bacteria producing extracellularhydrolyses from Howz Soltan Lake Iranrdquo Journal of IndustrialMicrobiology and Biotechnology vol 36 no 3 pp 333ndash3402009

[33] M J Garabito M C Marquez and A Ventosa ldquoHalotolerantBacillus diversity in hypersaline environmentsrdquo Canadian Jour-nal of Microbiology vol 44 no 2 pp 95ndash102 1998

[34] A Ventosa A Ramos-Cormenzana and M Kocur ldquoModer-ately halophilic gram-positive cocci from hypersaline environ-mentsrdquo Systematic and Applied Microbiology vol 4 no 4 pp564ndash570 1983

[35] E Quesada A Ventosa F Rodriguez-Valera and A R Cor-menzana ldquoTypes and properties of some bacteria isolated fromhypersaline soilsrdquo Journal of Applied Bacteriology vol 53 no 2pp 155ndash161 1982

[36] E Quesada V Bejar M J Valderrama A Ventosa and AR Ramos Cormenzana ldquoIsolation and characterization ofmoderately halophilic nonmotile rods from different salinehabitatsrdquoMicrobiologia vol 1 no 1-2 pp 89ndash96 1985

[37] S R Dave and H B Desai ldquoMicrobial diversity at marinesalterns near Bhavnagar Gujarat Indiardquo Current Science vol60 no 4 pp 497ndash500 2006

[38] S Kumar R Karan S Kapoor S P Singh and S K KhareldquoScreening and isolation of halophilic bacteria producingindustrially important enzymesrdquo Brazilian Journal of Microbi-ology vol 43 no 4 pp 1595ndash1603 2012

[39] B P Dave and A Soni ldquoDiversity of halophilic archaea atsalt pans around Bhavnagar coast Gujaratrdquo Proceedings ofthe National Academy of Sciences India Section B BiologicalSciences vol 83 no 2 pp 225ndash232 2013

[40] A A Joshi P P Kanekar A S Kelkar et al ldquoCultivable bacterialdiversity of alkaline Lonar lake Indiardquo Microbial Ecology vol55 no 2 pp 163ndash172 2008

[41] T M Raghavan and I Furtado ldquoOccurrence of extremelyhalophilic Archaea in sediments from the continental shelf ofwest coast of Indiardquo Current Science vol 86 no 8 pp 1065ndash1067 2004

[42] S Vijayanand J Hemapriya J Selvin and S Kiran ldquoBiodiver-sity of extremely halophilic bacterial strains isolated from solarsalterns of Tuticorin Tamilnadu Indiardquo International Journal ofWater Resources and Environmental Sciences vol 1 no 1 pp 1ndash72012

12 Chinese Journal of Biology

[43] S G Nanjani and H P Soni ldquoIsolation and characterization ofextremely halotolerant and halophilic organisms from Dwarkaand Veravalrdquo Journal of Pharmacy and Biological Sciences vol2 no 2 pp 20ndash25 2012

[44] H Ghozlan H Deif R A Kandil and S Sabry ldquoBiodiversity ofmoderately halophilic bacteria in hypersaline habitats in EgyptrdquoJournal of General and Applied Microbiology vol 52 no 2 pp63ndash72 2006

[45] D R Arahal M T Garcıa W Ludwig K H Schleifer andA Ventosa ldquoTransfer ofHalomonas canadensis andHalomonasisraelensis to the genus Chromohalobacter as Chromohalobac-ter canadensis comb nov and Chromohalobacter israelensiscomb novrdquo International Journal of Systematic and EvolutionaryMicrobiology vol 51 no 4 pp 1443ndash1448 2001

Submit your manuscripts athttpwwwhindawicom

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Anatomy Research International

PeptidesInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporation httpwwwhindawicom

International Journal of

Volume 2014

Zoology

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Molecular Biology International

GenomicsInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

The Scientific World JournalHindawi Publishing Corporation httpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

BioinformaticsAdvances in

Marine BiologyJournal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Signal TransductionJournal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

BioMed Research International

Evolutionary BiologyInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Biochemistry Research International

ArchaeaHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

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Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Advances in

Virolog y

Hindawi Publishing Corporationhttpwwwhindawicom

Nucleic AcidsJournal of

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Stem CellsInternational

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Enzyme Research

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

International Journal of

Microbiology

Page 4: Research Article Production of Extracellular …downloads.hindawi.com/archive/2014/205731.pdfResearch Article Production of Extracellular Polymeric Substances by Halophilic Bacteria

4 Chinese Journal of Biology

Table 3 Bacterial isolates obtained from soil samples of solar salterns of Gujarat Orissa and West Bengal India

States Sampling sites Number of samplesNumber of bacterial isolates

NaCl () in medium5 10 15

Gujarat

Jogrinar 4 44 23 4Kandla Port 3 19 12 4Khari Rohar 3 46 16 6Victor Port 2 20 5 mdash

Orissa Humma 5 48 14 mdashSurala 5 31 31 2

West Bengal Baksal 4 19 8 3Dadanpatrabar 5 29 16 10

Total 31 256 125 29

Table 4 Bacterial isolates obtained from water samples of solar salterns of Gujarat Orissa and West Bengal India

States Sampling sites Number of samplesNumber of bacterial isolates

NaCl () in medium5 10 15

Gujarat

Jogrinar 2 21 14 5Kandla Port 2 19 5 2Khari Rohar 1 1 3 mdashVictor Port 2 26 mdash 1

Orissa Humma 3 16 mdash mdashSurala 2 12 8 3

West Bengal Baksal 3 9 3 4Dadanpatrabar 3 17 4 4

Total 18 121 37 19

SUR301 and SUR307) appeared to be versatile in utilizingall 30 carbon sources Fermentation pattern of these strainsvaried remarkably the majority were unable to fermentmost of the carbon sources On the contrary fructosesorbitol and benzoate were fermented by most of the isolates(Table 9)

Sensitivity of these isolates to 22 different antibioticswas tested by disc-diffusion method (Table 10) and theantibiotic resistance index (ARI) was determined (Figure 2)Themajority of the isolates were sensitive to chloramphenicol(30 120583g) gentamycin (10 120583g) and norfloxacin (10 120583g) followedby ampicillin (10 120583g) Resistance to vancomycin (30120583g) fol-lowed by trimethoprim (30 120583g) was predominant amongstthe tested halophiles As judged by the ARI values the isolateSUR302 was the most resistant one (ARI = 065) followed bySUR301 KW203 and KW1805 while the lowest ARI (023)was indicated by the isolate JW307 which was most sensitiveto the tested antibiotics

The morphological physiological and biochemical char-acters including the carbon source utilization and fermen-tation patterns along with antibiotic susceptibility wereanalyzed and compared with the phenotypic characters ofhalophilic bacterial genera [5 18] so far reported According

to the phenotypic and biochemical characteristics 70 of theselected isolates (14) were tentatively identified asmembers ofthe genusHalomonasThe onlyGram-positive coccus (isolateSUR303) and the rod shaped endospore forming isolate(isolate KW203) were assigned to Salinicoccus and Bacillusrespectively Two of the isolates (KS1805 and SUR301) wereplaced in the genus Chromohalobacter while the remain-ing two (isolates JS504 and JS904) were included underAlteromonas and Aidingimonas respectively

4 Discussion

Halophiles have mainly been isolated from wide diversity ofenvironments such as saltern crystallizer ponds the DeadSea solar lakes and hypersaline lakes [26] Culture depen-dent diversity studies on halophiles have been made fromTunisian solar saltern [27 28] Tuzkoy salt mine [29] Sere-flikochisar Salt Lake [30] Kaldirim and Kayacik of Tuz Lake[31] Turkey Howz Soltan Lake Iran [32] and hypersalineenvironments in South Spain [33ndash36] In the Indian contextthe halophilic diversity studies have been restricted mainlyto the marine salterns of Bhavnagar [37ndash40] Lonar Lake[40] and Peninsular Coast [41 42]The present study reports

Chinese Journal of Biology 5

Table 5 Screening of halophilic bacterial isolates derived from soil samples for the production of extracellular polymeric substances

States Sampling sites Number of isolates Extracellular polymeric substances (gL)gt050 051ndash070 071ndash10 gt10

Gujarat

Jogrinar 71 18 42 7 4Kandla Port 35 13 13 7 2Khari Rohar 68 26 31 11 mdashVictor Port 25 20 3 2 mdash

Orissa Humma 62 22 37 3 mdashSurala 64 15 25 15 9

West Bengal Baksal 30 14 12 4 mdashDadanpatrabar 55 32 21 2 mdash

Total 410 160 184 51 15

Table 6 Screening of halophilic bacterial isolates derived from water samples for the production of extracellular polymeric substances

States Sampling sites Number of isolates Extracellular polymeric substances (gL)gt050 051ndash070 071ndash10 gt10

Gujarat

Jogrinar 40 26 9 3 2Kandla Port 26 13 11 1 1Khari Rohar 4 1 3 mdash mdashVictor Port 27 7 17 1 2

Orissa Humma 16 4 11 1 mdashSurala 23 8 6 9 mdash

West Bengal Baksal 16 7 6 3 mdashDadanpatrabar 25 8 14 3 mdash

Total 177 74 77 21 5

the distribution of halophilic and halotolerant bacterialcommunities in the inland multipond solar salterns spreadover the coasts of India Halophilic bacterial communitiesof 31 soil and 18 water samples from 8 different sites wereanalyzed by dilution and plating method (Tables 1 and 2) andprovide information regarding the availability anddiversity ofhalophilic bacteria in the solar saltern ponds Colony formingunits of soil and water samples revealed that soil samplesholdmore viable bacterial counts than water samples Ragha-van and Furtado [41] studied the occurrence of extremelyhalophilic archaea in sediments from the continental shelf ofwest coast of India and reported the presence of relatively lowaverage counts (7ndash5 times 103) of extreme halophiles in offshoresediments in contrast to the very high counts (105ndash109)of marine eubacteria Similar study by Joshi et al [40]reported that the total numbers of microorganisms in the soiland water samples were 102ndash106 cfug and 102ndash104 cfumLrespectively

A total of 587 halophilic and halotolerant bacterial strainswere isolated showing different degrees of NaCl tolerance(Tables 3 and 4) and supported the observations of Quesadaet al [35] andVentosa et al [34] However during the presentstudy we were unable to isolate extreme halophiles whichhave frequently been identified as the dominant phylotypes inhypersaline environments along with solar salterns of India

[39 41] Furthermore soil samples showed more bacterialdiversity than water ones which is in accordance with theobservations of Joshi et al [40]

During the course of screening of the moderatelyhalophilic isolates for EPS production (Tables 5 and 6) only20 isolates appeared to be promising with significant yield ofEPS (10ndash185 gL) (Figure 1) Joshi and coworkers [40] sim-ilarly screened 86 halophilic bacteria from Lonar Lake andreported Halomonas campisalis and Vagococcus carniphilusas potent EPS producers Similarly Nanjani and Soni [43]also isolated 73 halotolerant and halophilic bacteria from soilsamples of Veraval and Dwarka 23 of them produced EPSranging from 02 to 1060 gL In addition reports on EPSproduction by moderately halophilic bacteria of the genusHalomonas are not uncommon [8 15ndash17]

Attempts have been made to determine the taxonomicidentity of all 20 promising EPS producing isolates followingdetailed physiobiochemical characterization (Tables 7ndash9) andcomparison with Bergeyrsquos Manual of Systematic Bacteriology[24] and those of Mata et al [25]Themajority (14 isolates) ofthem were Gram-negative nonsporulating rods and capableof growing in 25ndash20NaCl similar to those ofHalomonas asreported by Quesada et al [36] Ghozlan et al [44] andMataet al [25] However the Gram-positive endospore formingisolate KW203 was assigned to the genus Bacillus

6 Chinese Journal of Biology

Table7Morph

ologicalandph

ysiologicalcharacteristicso

fthe

selected

EPSprod

ucinghaloph

ilicb

acteria

isolated

from

solarsalternso

fInd

ia

Character

Bacterialisolates

JS504

JS802

JS803

JS904

JW302

JW307

KS110

1KS

1805

KW203

VW301

Morph

ologicalcharacters

Colon

ymorph

olog

yCr

eamcirc

ular

Whitecirc

ular

Creamcirc

ular

Creamcirc

ular

Transparentcircular

Creamcirc

ular

Creamcirc

ular

Whitecirc

ular

Whitecirc

ular

Creamcirc

ular

Gram

nature

minus+

minusminus

minusminus

minusminus

+minus

Cellshape

Rod

Rod

Rod

Rod

Rod

Rod

Rod

Rod

Rod

Rod

Cell

arrang

ement

Sing

leSing

leSing

leSing

leSing

lechains

Sing

lechains

Sing

lechains

Sing

leSing

leclu

mped

Sing

lechains

Motility

++

++

++

minusminus

++

Pigm

entatio

nminus

minusminus

minusminus

minusminus

minusminus

minus

Endo

sporeformation

minusminus

minusminus

minusminus

minusminus

+minus

Physiologicalcharacters

pHrang

efor

grow

th5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

pHop

timum

forg

rowth

119

89

1111

118

89

Temprang

efor

grow

th(∘C)

27ndash4

027ndash4

027ndash4

027ndash4

027ndash4

027ndash4

022ndash4

027ndash4

027ndash4

022ndash4

0Tempop

timum

forg

rowth

(∘C)

3237

3732

3232

3232

3732

NaC

lrange

forg

rowth

()

0ndash15

0ndash20

0ndash15

0ndash20

0ndash15

0ndash20

0ndash20

25ndash15

25ndash15

0ndash20

NaC

loptim

umforg

rowth

()

510

105

105

575

105

Growth

onKingrsquosBmedium

++

++

++

++

++

Growth

onMcC

onky

agar

minusminus

minusminus

minusminus

minusminus

minusminus

Tentativeidentity

Alteromonas

Halom

onas

Halom

onas

Aidingim

onas

Halom

onas

Halom

onas

Halom

onas

Chromohalobacter

Bacillus

Halom

onas

Character

Bacterialisolates

VW402

SUR108

SUR3

01SU

R302

SUR3

03SU

R307

SUR3

09SU

R310

SUR2

02SU

R201

Morph

ologicalcharacters

Colon

ymorph

olog

yCr

eamcirc

ular

Creamcirc

ular

Whitegum

my

Transparentcircular

Flatreddish-pink

Creamcirc

ular

Creamcirc

ular

Creamcirc

ular

Creamcirc

ular

Creamcirc

ular

Gram

nature

minusminus

minusminus

+minus

minusminus

minusminus

Cellshape

Rod

Rod

Rod

Rod

Cocci

Rod

Rod

Rod

Rod

Rod

Cell

arrang

ement

Sing

leSing

leSing

leSing

leSing

leclu

mped

Sing

leSing

leSing

leSing

leSing

leMotility

+minus

minus+

minus+

minus+

minusminus

Pigm

entatio

nminus

minusminus

minusPink

minusminus

minusminus

minus

Endo

sporeformation

minusminus

minusminus

minusminus

minusminus

minusminus

Physiologicalcharacters

pHrang

efor

grow

th5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

pHop

timum

forg

rowth

97

77

117

77

79

Temprang

efor

grow

th(∘C)

27ndash4

027ndash4

027ndash4

032ndash4

532ndash4

522ndash4

032ndash4

022ndash4

522ndash4

532ndash4

5Tempop

timum

forg

rowth

(∘C)

3232

3732

3727

3227

2732

NaC

lrange

forg

rowth

()

0ndash15

0ndash15

25ndash15

25ndash15

25ndash15

0ndash20

0ndash20

0ndash20

0ndash20

0ndash15

NaC

loptim

umforg

rowth

()

525

575

525

7525

575

Growth

onKingrsquosBmedium

++

++

++

++

++

Growth

onMcC

onky

agar

minusminus

minusminus

minusminus

minusminus

minusminus

Tentativeidentity

Halom

onas

Halom

onas

Chromohalobacter

Halom

onas

Salin

icoccus

Halom

onas

Halom

onas

Halom

onas

Halom

onas

Halom

onas

Chinese Journal of Biology 7

Table8Biochemicalcharacteris

ticso

fthe

selected

EPSprod

ucinghaloph

ilicb

acteria

isolated

from

solarsalternso

fInd

ia

Character

Bacterialisolates

JS 504

JS 802

JS 803

JS 904

JW 302

JW 307

KS 1101

KS 1805

KW 203

VW 301

VW 402

SUR

108

SUR

301

SUR

302

SUR

303

SUR

307

SUR

309

SUR

310

SUR

202

SUR

201

Biochemicalcharacters

Voges-Proskauertest

minusminus

minusminus

minusminus

minusminus

+minus

minusminus

minusminus

minusminus

minusminus

minusminus

Citrateu

tilization

+minus

minusminus

minus+

minus+

minus+

++

+minus

minus+

++

++

Methylred

test

minusminus

minusminus

minusminus

minus+

+minus

minusminus

minusminus

minusminus

minusminus

minusminus

Prod

uctio

nof

Glutaminase

++

++

++

++

++

++

++

minus+

++

++

Asparaginase

++

++

++

++

++

++

++

minus+

++

++

Amylase

+minus

minusminus

minusminus

minusminus

+minus

minusminus

minusminus

minusminus

minusminus

minusminus

Cellulase

minusminus

minusminus

minusminus

minusminus

+minus

minusminus

minusminus

minusminus

minusminus

minusminus

Pectinase

+minus

minusminus

minusminus

minusminus

+minus

minusminus

minusminus

minusminus

minusminus

minusminus

Inulinase

+minus

minusminus

minusminus

minusminus

+minus

minusminus

minusminus

minusminus

minusminus

minusminus

Gelatinase

minusminus

minusminus

minusminus

minusminus

+minus

minusminus

minusminus

minusminus

minusminus

minusminus

Lipase

+minus

minusminus

minusminus

minus+

+minus

minusminus

+minus

minusminus

minusminus

minusminus

Xylanase

+minus

minusminus

minusminus

minusminus

+minus

minusminus

minusminus

minusminus

minusminus

minusminus

Caseinase

minusminus

minus+

minusminus

minusminus

minusminus

minusminus

minus+

minusminus

minusminus

minusminus

Urease

minusminus

minus+

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

Catalase

++

++

++

++

++

++

++

++

++

++

Nitrater

eductase

++

++

++

++

++

++

++

++

++

++

H2S

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

Lysin

edecarbo

xylase

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

Arginined

ecarbo

xylase

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

Ornith

ined

ecarbo

xylase

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

Indo

leminus

minusminus

minusminus

minusminus

+minus

minusminus

minus+

minus+

minusminus

minusminus

minus

Phenylalanined

eaminase

++

+minus

+minus

++

minus+

minus+

++

++

++

minus+

Tryptoph

andeam

inase

++

++

++

++

++

minusminus

++

++

++

+minus

8 Chinese Journal of Biology

Table9Ca

rbon

source

utilizatio

nandferm

entatio

nof

thes

elected

EPSprod

ucinghaloph

ilicb

acteria

isolated

from

samples

ofsolarsalternso

fInd

ia

Carbon

source

Bacterialisolates

JS 504

JS 802

JS 803

JS 904

JW 302

JW 307

KS 1101

KS 1805

KW 203

VW

301

VW

402

SUR

108

SUR

301

SUR

302

SUR

303

SUR

307

SUR

309

SUR

310

SUR

202

SUR

201

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

Glucose

++

++

+minusminus

++minus

+minus

+minus

++

++

+minus

+minus

+minus

+minusminus

++

++minus

+minus

+minusminusminusminusminus

Galactose

+minus

+minus

+minusminus

++minus

+minus

+minus

++

+minus

+minus

+minus

+minus

++minusminus

+minus

++

+minus

+minus

+minus

+minus

Arabino

seminusminusminusminus

+minusminusminusminusminus

+minus

+minus

++

++

+minus

+minus

+minus

+minusminus

++minus

+minus

+minusminusminus

+minus

+minus

Manno

se+minus

+minus

+minusminus

++minus

+minus

+minusminus

++

++minus

+minus

+minus

+minusminusminusminusminus

+minus

+minus

+minus

+minus

+minus

Rham

nose

+minus

+minus

+minus

+minusminusminusminusminusminusminus

++

+minusminusminus

+minus

+minus

++minus

++minus

+minusminusminusminusminusminusminus

+minus

Trehalose

+minus

+minus

+minusminus

+minus

+minusminusminusminus

+minus

++minusminusminusminus

+minus

+minusminus

++

++minusminusminusminusminusminusminus

+minus

Lactose

minusminusminusminus

+minusminusminus

+minus

+minus

+minus

+minus

+minus

+minus

+minusminusminus

+minus

++minusminus

+minusminusminusminusminus

+minus

+minus

Sucrose

+minus

+minus

+minus

++

++

+minus

+minus

+minus

++

+minus

+minusminusminus

+minus

++

+minus

+minus

+minus

+minusminusminus

+minus

Xylose

minusminusminusminus

+minus

+minusminusminus

+minus

++minus

+minusminus

+minus

+minus

+minus

+minusminus

+minus

++

+minusminusminusminus

+minusminusminus

Ribo

se+minus

++

++

++

+minus

+minus

+minus

++

++

+minus

+minus

+minus

++

++

++

+minus

+minus

+minus

+minus

+minus

Fructose

+minus

++

+minus

++

+minus

++

++

+minus

++

++

++

++

++

++

++

++

++

++

++

++

Maltose

minusminusminusminus

+minusminus

+minus

++minus

+minus

+minus

++

+minus

+minus

+minus

++minus

+minus

++minusminusminusminusminus

+minus

+minus

Glycine

minusminusminus

++minus

+minusminus

+minusminusminusminus

+minusminusminusminusminusminusminusminusminus

+minusminusminusminusminus

+minus

+minusminusminusminus

+minusminus

Glycerol

+minus

+minus

+minusminus

++

++minus

+minus

+minus

++

+minus

+minus

+minus

+minusminus

++

++minus

+minus

+minus

+minus

+minus

Adon

itol

+minus

+minus

+minus

+minusminusminus

+minus

+minus

+minus

+minus

+minus

+minusminusminus

+minus

++

+minus

+minus

+minus

+minusminusminusminusminus

Salicin

+minus

++

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

++

++

+minus

+minus

+minus

+minus

+minus

Meso-inosito

l+minusminusminus

+minus

+minus

+minus

+minus

++

+minus

++

+minus

+minusminusminus

+minusminus

++minus

+minus

+minus

+minus

+minus

+minus

Mannitol

+minus

+minus

+minus

++

+minus

+minus

+minus

+minus

++

+minus

+minus

+minus

+minus

+minus

++

+minus

+minus

+minus

+minus

+minus

Sorbito

l+

++

++minusminus

++

+minus

++

++

++

++

++

++

++

+minus

+minus

++

++minus

+minus

++

+minus

Raffino

seminusminus

++

++minusminus

+minus

+minus

+minus

+minus

++minusminusminusminusminusminusminusminus

++

++

+minus

+minus

+minus

+minus

+minus

Cellobiose

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

++

+minus

+minus

+minus

+minus

++

+minus

+minus

+minus

+minus

+minus

+minus

Melibioseminusminusminusminusminusminusminusminusminusminusminusminusminusminusminusminus

++minusminusminusminusminusminusminusminus

++minusminusminusminusminusminusminusminusminusminusminusminus

Na-acetate

+minus

+minus

+minus

+minus

++

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

++

++

+minus

+minus

+minus

+minus

+minus

Na-citrate

+minus

++

+minusminusminusminus

+minusminusminusminusminusminusminusminusminusminusminusminusminusminusminusminusminus

+minusminusminusminusminusminusminusminusminusminusminusminus

Na-fumarate

+minus

+minus

+minus

++

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minusminus

++minus

+minus

+minus

+minus

+minus

+minus

Na-form

ate

+minus

+minus

+minus

+minus

+minusminusminus

+minus

+minus

+minusminusminus

+minusminusminus

+minus

++minusminus

+minusminusminusminusminusminusminusminusminus

Na-benzoate

+minus

++

++

+minus

+minus

+minus

++

++

++

+minus

+minus

++

++

+minus

+minus

+minus

++

++

++

++

Na-malateminusminusminusminusminusminus

++

+minusminusminusminusminusminusminusminusminusminusminusminusminusminusminusminusminus

++

++minusminusminusminusminusminusminusminusminusminus

Na-prop

ionate

+minus

+minus

+minusminusminus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

++minus

++minus

+minus

+minus

+minus

+minus

Na-succinate

+minus

+minus

+minus

++

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

++

++

+minus

+minus

+minus

+minus

+minus

Uutilization

Fferm

entatio

n

Chinese Journal of Biology 9

Table10A

ntibiotic

sensitivitypatte

rnof

thes

electedEP

Sprod

ucinghaloph

ilicb

acteria

isolated

from

solarsalternso

fInd

ia

Antibiotics

Bacterialisolates

JS 504

JS 802

JS 803

JS 904

JW 302

JW 307

KS 1101

KS 1805

KW 203

VW 301

VW 402

SUR

108

SUR

301

SUR

302

SUR

303

SUR

307

SUR

309

SUR

310

SUR

202

SUR

201

Ampicillin(10120583

g)S

SR

SS

SS

RI

SS

SR

SS

SS

SS

SAmoxicillin

(30120583

g)S

SR

RS

SS

RS

SS

SR

RS

IR

SS

SBa

citracin

(10120583

g)S

SR

SS

SR

RS

IR

RR

RS

IR

RR

RCiprofl

oxacin

(30120583

g)I

II

RI

II

IR

II

II

IR

SR

II

ICh

lorte

tracyclin

e(30120583g)

SI

SS

IS

RR

SS

SS

RR

SS

II

IS

Chloramph

enicol(10120583

g)S

SS

SS

SS

SR

SS

SS

SS

SR

SS

SErythrom

ycin

(15120583

g)S

SI

SS

IS

II

II

II

SR

RI

IS

SFu

sidicacid

(30120583

g)I

RR

SS

IR

RS

RI

RR

RS

IR

II

RGentamycin

(10120583

g)S

SS

SS

SS

SR

SS

SS

SS

IS

SS

SKa

namycin

(30120583

g)I

IS

RS

II

SR

II

RI

RR

RI

RI

RMethicillin(5120583g)

SS

RS

IR

RR

RR

RR

RR

SR

RR

RR

Neomycin

(30120583

g)S

SI

RI

IR

II

IR

II

RR

RI

IR

RNorflo

xacin(30120583

g)S

SR

SS

SS

SI

SS

SS

SS

SS

SS

SNetillin

(10120583

g)R

SS

IR

SS

SR

RR

SS

RR

RR

RR

RNalidixicacid

(30120583

g)R

RS

RR

II

RR

II

RR

RR

SR

RR

RNovob

iosin

(30120583

g)I

RR

SR

SS

RS

SS

IR

SS

SI

SS

SPo

lymixin

B(50120583

g)R

RI

RI

IR

RR

RI

IR

RR

II

II

SPenicillinG(10120583

g)S

SR

IS

IR

RR

II

SR

RS

II

II

SStreptom

ycin

(10120583

g)S

SS

RS

RR

SR

RR

IS

RS

SR

II

RTetracyclin

e(30120583g)

RR

RR

RI

RR

SR

IR

RR

RS

RR

RI

Trim

etho

prim

e(30120583g)

SS

SS

IS

SI

SS

SS

IR

RS

SS

SS

Vancom

ycin

(30120583

g)R

RR

SR

RR

RS

RR

RR

RS

RR

RR

R

S=SensitiveR

=Re

sistantI

=Interm

ediate

10 Chinese Journal of Biology

0

1

2

3

4

Bacterial isolate

Gro

wth

OD

at540

nmto

tal E

PS (g

L)

OD at 540nmTotal EPS

JS504

JS802

JS803

JS904

JW302

JW307

KS1101

KS1805

KW203

VW301

VW402

SUR1

08

SUR3

01

SUR3

02

SUR3

03

SUR3

07

SUR3

09

SUR3

10

SUR2

02

SUR2

01

Figure 1 Screening of moderately halophilic bacteria isolatedfrom soil and water samples of solar salterns for production ofextracellular polymeric substances

0

02

04

06

08

1

Ant

ibio

tic re

sista

nce i

ndex

Bacterial isolate

JS504

JS802

JS803

JS904

JW302

JW307

KS1101

KS1805

KW203

SUR1

08

SUR2

01

SUR2

02

SUR3

01

SUR3

02

SUR3

03

SUR3

07

SUR3

09

SUR3

10

VW301

VW402

Figure 2 Antibiotic resistance index of the selected EPS producinghalophilic bacteria isolated from soil and water samples of solarsalterns of India

The isolates SUR301 and KS 1805 were tentatively iden-tified as Chromohalobacter based on the study by Arahal etal [45] and bear striking similarity with those of Chromo-halobacter sp isolated fromhypersaline soil sample of TriveniSangam Gujarat [38] The physiological morphological andbiochemical characteristics of isolate SUR303 were consistentwith the features of the genus Salinicoccus [10] while isolatesJS904 and JS504 were tentatively assigned to Aidingimonasand Alteromonas respectively

Finally sensitivity of these isolates to antibiotics (Table 10and Figure 2) also corroborates the findings ofMata et al [25]and Hedi et al [28]

5 Conclusion

It may be emphasized that study of these moderatelyhalophilic bacteria from coastal hypersaline solar salterns ofIndia with special attempt on screening for EPS productionhas led to the discovery of wide variety of halophilic speciesAlthough attention has been focused on the production

of EPS by these halophilic bacterial isolates their physio-biochemical features indicate that they may equally holdpotential towards production of various biomolecules ofindustrial interest

Conflict of Interests

It is declared by the authors that there is no conflict of interestsregarding the publication of this paper

Acknowledgment

This study was financially supported by grants from Uni-versity Grants Commission India (Sanction no F14-2(SC)2008 (SA-III) 31 March 2009) under the Scheme ofRajiv Gandhi National Fellowship

References

[1] E A Galinski and B J Tindall ldquoBiotechnological prospectsfor halophiles and halotolerant microorganismsrdquo in MolecularBiology and Biotechnology of Extremophiles R D Herbert andR J Sharp Eds Blackie London UK 1992

[2] R Margesin and F Schinner ldquoPotential of halotolerant andhalophilic microorganisms for biotechnologyrdquo Extremophilesvol 5 no 2 pp 73ndash83 2001

[3] F Rodriguez Valera F Ruiz Berraquero and A Ramos Cor-menzana ldquoCharacteristics of the heterotrophic bacterial pop-ulations in hypersaline environments of different salt concen-trationsrdquoMicrobial Ecology vol 7 no 3 pp 235ndash243 1981

[4] F Rodriguez-Valera ldquoCharacteristics and microbial ecol-ogy of hypersaline environmentsrdquo in Halophilic Bacteria FRodriguez-Valera Ed vol 1 pp 3ndash30 CRC Press Boca RatonFla USA 1988

[5] D J Kushner and M Kamekura ldquoPhysiology of halophiliceubacteriardquo inHalophilic Bacteria F Rodriguez-Valera Ed vol1 pp 109ndash138 CRC press Boca Raton Fla USA 1988

[6] M A Amoozegar F Malekzadeh and K AMalik ldquoProductionof amylase by newly isolated moderate halophile Halobacillussp strainMA-2rdquo Journal of Microbiological Methods vol 52 no3 pp 353ndash359 2003

[7] C O Jeon J Lim J Lee et al ldquoLentibacillus salarius spnov isolated from saline sediment in China and emendeddescription of the genus Lentibacillusrdquo International Journal ofSystematic and Evolutionary Microbiology vol 55 no 3 pp1339ndash1343 2005

[8] S Bouchotroch E Quesada A Del Moral I Llamas andV Bejar ldquoHalomonas maura sp nov a novel moderatelyhalophilic exopolysaccharide-producing bacteriumrdquo Interna-tional Journal of Systematic and Evolutionary Microbiology vol51 no 5 pp 1625ndash1632 2001

[9] A Biswas A Patra and A Paul ldquoProduction of poly-3-hydroxyalkanoic acids by a moderately halophilic bacteriumHalomonas marina HMA 103 isolated from solar saltern ofOrissa Indiardquo Acta Microbiologica et Immunologica Hungaricavol 56 no 2 pp 125ndash143 2009

[10] S Y Jayachandra S Anil Kumar D P Merley and MB Sulochana ldquoIsolation and characterization of extremehalophilic bacterium Salinicoccus sp JAS4 producing extra-cellular hydrolytic enzymesrdquo Recent Research in Science andTechnology vol 4 no 4 pp 46ndash49 2012

Chinese Journal of Biology 11

[11] I W Sutherland ldquoNovel and established applications of micro-bial polysaccharidesrdquo Trends in Biotechnology vol 16 no 1 pp41ndash46 1998

[12] MWMittelman andG G Geesey ldquoCopper-binding character-istics of exopolymers from a freshwater-sediment bacteriumrdquoApplied and EnvironmentalMicrobiology vol 49 no 4 pp 846ndash851 1985

[13] J Anton I Meseguer and F Rodriguez-Valera ldquoProductionof an extracellular polysaccharide by Haloferax mediterraneirdquoApplied Enviromental Microbiology vol 54 no 10 pp 2381ndash2386 1988

[14] H Parolis L A S Parolis I F Boan et al ldquoThe structure ofthe exopolysaccharide produced by the halophilic ArchaeonHaloferax mediterranei strain R4 (ATCC 33500)rdquo CarbohydrateResearch vol 295 pp 147ndash156 1996

[15] E Quesada V Bejar and C Calvo ldquoExopolysaccharide produc-tion by Volcaniella eurihalinardquo Experientia vol 49 no 12 pp1037ndash1041 1993

[16] J A Mata V Bejar I Llamas et al ldquoExopolysaccharides pro-duced by the recently described halophilic bacteria Halomonasventosae and Halomonas anticariensisrdquo Research in Microbiol-ogy vol 157 no 9 pp 827ndash835 2006

[17] I Llamas J A Mata R Tallon et al ldquoCharacterization ofthe exopolysaccharide produced by Salipiger mucosus A3119879 ahalophilic species belonging to the Alphaproteobacteria iso-lated on the Spanish Mediterranean seaboardrdquo Marine Drugsvol 8 no 8 pp 2240ndash2251 2010

[18] F Martınez-Checa E Quesada J Martınez-Canovas I Llamasand V Bejar ldquoPalleronia marisminoris gen nov sp nova moderately halophilic exopolysaccharide-producing bac-terium belonging to the rsquoAlphaproteobacteriarsquo isolated from asaline soilrdquo International Journal of Systematic and EvolutionaryMicrobiology vol 55 no 6 pp 2525ndash2530 2005

[19] A Ventosa M T Garcia M Kamekura H Onishi and FRuiz-Berraquero ldquoBacillus halophilus sp nov a moderatelyhalophilic Bacillus speciesrdquo Systematic and Applied Microbiol-ogy vol 12 no 2 pp 162ndash166 1989

[20] R A Moraine and P Rogovin ldquoKinetics of polysaccharide B1459 fermentationrdquoBiotechnology and Bioengineering vol 8 no4 pp 511ndash524 1966

[21] M Dubois K A Gilles J K Hamilton P A Rebers and FSmith ldquoColorimetric method for determination of sugars andrelated substancesrdquoAnalytical Chemistry vol 28 no 3 pp 350ndash356 1956

[22] O H Lowry N J Rosenbrough A L Farr and R J RandallldquoProtein measurement with the Folin phenol reagentrdquo TheJournal of Biological Chemistry vol 193 no 1 pp 265ndash275 1951

[23] P Gerhardt R G E Murray W A Wood and N R KriegMethods for General and Molecular Bacteriology AmericanSociety for Microbiology Washington DC USA 1994

[24] G M Garrity J A Bell and T G Lilburn ldquoTaxonomic outlineof the procaryotesrdquo inBergeyrsquosManual of Systematic BacteriologD R Boone and R W Castenholz Eds Springer New YorkNY USA 2nd edition 2003

[25] J A Mata J Martınez-Canovas E Quesada and V BejarldquoA detailed phenotypic characterisation of the type strains ofHalomonas speciesrdquo Systematic and Applied Microbiology vol25 no 3 pp 360ndash375 2002

[26] A Oren ldquoMolecular ecology of extremely halophilic archaeaand bacteriardquo FEMSMicrobiology Ecology vol 39 no 1 pp 1ndash72002

[27] H Baati R Amdouni N Gharsallah A Sghir and E AmmarldquoIsolation and characterization of moderately halophilic bacte-ria from tunisian solar salternrdquo Current Microbiology vol 60no 3 pp 157ndash161 2010

[28] A Hedi N Sadfi M Fardeau et al ldquoStudies on the biodiversityof halophilic microorganisms isolated from El-Djerid salt lake(Tunisia) under aerobic conditionsrdquo International Journal ofMicrobiology vol 2009 Article ID 731786 17 pages 2009

[29] M Birbir A Ogan B Calli and B Mertoglu ldquoEnzymecharacteristics of extremely halophilic archaeal community inTuzkoy Salt Mine Turkeyrdquo World Journal of Microbiology andBiotechnology vol 20 no 6 pp 613ndash621 2004

[30] M Birbir and C Sesal ldquoExtremely halophilic bacterial commu-nities in Sereflikochisar Salt Lake in Turkeyrdquo Turkish Journal ofBiology vol 27 no 7 pp 7ndash22 2003

[31] M Birbir B Calli B Mertoglu et al ldquoExtremely halophilicArchaea from Tuz Lake Turkey and the adjacent Kaldirim andKayacik salternsrdquoWorld Journal ofMicrobiology and Biotechnol-ogy vol 23 no 3 pp 309ndash316 2007

[32] R Rohban M A Amoozegar and A Ventosa ldquoScreeningand isolation of halophilic bacteria producing extracellularhydrolyses from Howz Soltan Lake Iranrdquo Journal of IndustrialMicrobiology and Biotechnology vol 36 no 3 pp 333ndash3402009

[33] M J Garabito M C Marquez and A Ventosa ldquoHalotolerantBacillus diversity in hypersaline environmentsrdquo Canadian Jour-nal of Microbiology vol 44 no 2 pp 95ndash102 1998

[34] A Ventosa A Ramos-Cormenzana and M Kocur ldquoModer-ately halophilic gram-positive cocci from hypersaline environ-mentsrdquo Systematic and Applied Microbiology vol 4 no 4 pp564ndash570 1983

[35] E Quesada A Ventosa F Rodriguez-Valera and A R Cor-menzana ldquoTypes and properties of some bacteria isolated fromhypersaline soilsrdquo Journal of Applied Bacteriology vol 53 no 2pp 155ndash161 1982

[36] E Quesada V Bejar M J Valderrama A Ventosa and AR Ramos Cormenzana ldquoIsolation and characterization ofmoderately halophilic nonmotile rods from different salinehabitatsrdquoMicrobiologia vol 1 no 1-2 pp 89ndash96 1985

[37] S R Dave and H B Desai ldquoMicrobial diversity at marinesalterns near Bhavnagar Gujarat Indiardquo Current Science vol60 no 4 pp 497ndash500 2006

[38] S Kumar R Karan S Kapoor S P Singh and S K KhareldquoScreening and isolation of halophilic bacteria producingindustrially important enzymesrdquo Brazilian Journal of Microbi-ology vol 43 no 4 pp 1595ndash1603 2012

[39] B P Dave and A Soni ldquoDiversity of halophilic archaea atsalt pans around Bhavnagar coast Gujaratrdquo Proceedings ofthe National Academy of Sciences India Section B BiologicalSciences vol 83 no 2 pp 225ndash232 2013

[40] A A Joshi P P Kanekar A S Kelkar et al ldquoCultivable bacterialdiversity of alkaline Lonar lake Indiardquo Microbial Ecology vol55 no 2 pp 163ndash172 2008

[41] T M Raghavan and I Furtado ldquoOccurrence of extremelyhalophilic Archaea in sediments from the continental shelf ofwest coast of Indiardquo Current Science vol 86 no 8 pp 1065ndash1067 2004

[42] S Vijayanand J Hemapriya J Selvin and S Kiran ldquoBiodiver-sity of extremely halophilic bacterial strains isolated from solarsalterns of Tuticorin Tamilnadu Indiardquo International Journal ofWater Resources and Environmental Sciences vol 1 no 1 pp 1ndash72012

12 Chinese Journal of Biology

[43] S G Nanjani and H P Soni ldquoIsolation and characterization ofextremely halotolerant and halophilic organisms from Dwarkaand Veravalrdquo Journal of Pharmacy and Biological Sciences vol2 no 2 pp 20ndash25 2012

[44] H Ghozlan H Deif R A Kandil and S Sabry ldquoBiodiversity ofmoderately halophilic bacteria in hypersaline habitats in EgyptrdquoJournal of General and Applied Microbiology vol 52 no 2 pp63ndash72 2006

[45] D R Arahal M T Garcıa W Ludwig K H Schleifer andA Ventosa ldquoTransfer ofHalomonas canadensis andHalomonasisraelensis to the genus Chromohalobacter as Chromohalobac-ter canadensis comb nov and Chromohalobacter israelensiscomb novrdquo International Journal of Systematic and EvolutionaryMicrobiology vol 51 no 4 pp 1443ndash1448 2001

Submit your manuscripts athttpwwwhindawicom

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Anatomy Research International

PeptidesInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporation httpwwwhindawicom

International Journal of

Volume 2014

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Microbiology

Page 5: Research Article Production of Extracellular …downloads.hindawi.com/archive/2014/205731.pdfResearch Article Production of Extracellular Polymeric Substances by Halophilic Bacteria

Chinese Journal of Biology 5

Table 5 Screening of halophilic bacterial isolates derived from soil samples for the production of extracellular polymeric substances

States Sampling sites Number of isolates Extracellular polymeric substances (gL)gt050 051ndash070 071ndash10 gt10

Gujarat

Jogrinar 71 18 42 7 4Kandla Port 35 13 13 7 2Khari Rohar 68 26 31 11 mdashVictor Port 25 20 3 2 mdash

Orissa Humma 62 22 37 3 mdashSurala 64 15 25 15 9

West Bengal Baksal 30 14 12 4 mdashDadanpatrabar 55 32 21 2 mdash

Total 410 160 184 51 15

Table 6 Screening of halophilic bacterial isolates derived from water samples for the production of extracellular polymeric substances

States Sampling sites Number of isolates Extracellular polymeric substances (gL)gt050 051ndash070 071ndash10 gt10

Gujarat

Jogrinar 40 26 9 3 2Kandla Port 26 13 11 1 1Khari Rohar 4 1 3 mdash mdashVictor Port 27 7 17 1 2

Orissa Humma 16 4 11 1 mdashSurala 23 8 6 9 mdash

West Bengal Baksal 16 7 6 3 mdashDadanpatrabar 25 8 14 3 mdash

Total 177 74 77 21 5

the distribution of halophilic and halotolerant bacterialcommunities in the inland multipond solar salterns spreadover the coasts of India Halophilic bacterial communitiesof 31 soil and 18 water samples from 8 different sites wereanalyzed by dilution and plating method (Tables 1 and 2) andprovide information regarding the availability anddiversity ofhalophilic bacteria in the solar saltern ponds Colony formingunits of soil and water samples revealed that soil samplesholdmore viable bacterial counts than water samples Ragha-van and Furtado [41] studied the occurrence of extremelyhalophilic archaea in sediments from the continental shelf ofwest coast of India and reported the presence of relatively lowaverage counts (7ndash5 times 103) of extreme halophiles in offshoresediments in contrast to the very high counts (105ndash109)of marine eubacteria Similar study by Joshi et al [40]reported that the total numbers of microorganisms in the soiland water samples were 102ndash106 cfug and 102ndash104 cfumLrespectively

A total of 587 halophilic and halotolerant bacterial strainswere isolated showing different degrees of NaCl tolerance(Tables 3 and 4) and supported the observations of Quesadaet al [35] andVentosa et al [34] However during the presentstudy we were unable to isolate extreme halophiles whichhave frequently been identified as the dominant phylotypes inhypersaline environments along with solar salterns of India

[39 41] Furthermore soil samples showed more bacterialdiversity than water ones which is in accordance with theobservations of Joshi et al [40]

During the course of screening of the moderatelyhalophilic isolates for EPS production (Tables 5 and 6) only20 isolates appeared to be promising with significant yield ofEPS (10ndash185 gL) (Figure 1) Joshi and coworkers [40] sim-ilarly screened 86 halophilic bacteria from Lonar Lake andreported Halomonas campisalis and Vagococcus carniphilusas potent EPS producers Similarly Nanjani and Soni [43]also isolated 73 halotolerant and halophilic bacteria from soilsamples of Veraval and Dwarka 23 of them produced EPSranging from 02 to 1060 gL In addition reports on EPSproduction by moderately halophilic bacteria of the genusHalomonas are not uncommon [8 15ndash17]

Attempts have been made to determine the taxonomicidentity of all 20 promising EPS producing isolates followingdetailed physiobiochemical characterization (Tables 7ndash9) andcomparison with Bergeyrsquos Manual of Systematic Bacteriology[24] and those of Mata et al [25]Themajority (14 isolates) ofthem were Gram-negative nonsporulating rods and capableof growing in 25ndash20NaCl similar to those ofHalomonas asreported by Quesada et al [36] Ghozlan et al [44] andMataet al [25] However the Gram-positive endospore formingisolate KW203 was assigned to the genus Bacillus

6 Chinese Journal of Biology

Table7Morph

ologicalandph

ysiologicalcharacteristicso

fthe

selected

EPSprod

ucinghaloph

ilicb

acteria

isolated

from

solarsalternso

fInd

ia

Character

Bacterialisolates

JS504

JS802

JS803

JS904

JW302

JW307

KS110

1KS

1805

KW203

VW301

Morph

ologicalcharacters

Colon

ymorph

olog

yCr

eamcirc

ular

Whitecirc

ular

Creamcirc

ular

Creamcirc

ular

Transparentcircular

Creamcirc

ular

Creamcirc

ular

Whitecirc

ular

Whitecirc

ular

Creamcirc

ular

Gram

nature

minus+

minusminus

minusminus

minusminus

+minus

Cellshape

Rod

Rod

Rod

Rod

Rod

Rod

Rod

Rod

Rod

Rod

Cell

arrang

ement

Sing

leSing

leSing

leSing

leSing

lechains

Sing

lechains

Sing

lechains

Sing

leSing

leclu

mped

Sing

lechains

Motility

++

++

++

minusminus

++

Pigm

entatio

nminus

minusminus

minusminus

minusminus

minusminus

minus

Endo

sporeformation

minusminus

minusminus

minusminus

minusminus

+minus

Physiologicalcharacters

pHrang

efor

grow

th5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

pHop

timum

forg

rowth

119

89

1111

118

89

Temprang

efor

grow

th(∘C)

27ndash4

027ndash4

027ndash4

027ndash4

027ndash4

027ndash4

022ndash4

027ndash4

027ndash4

022ndash4

0Tempop

timum

forg

rowth

(∘C)

3237

3732

3232

3232

3732

NaC

lrange

forg

rowth

()

0ndash15

0ndash20

0ndash15

0ndash20

0ndash15

0ndash20

0ndash20

25ndash15

25ndash15

0ndash20

NaC

loptim

umforg

rowth

()

510

105

105

575

105

Growth

onKingrsquosBmedium

++

++

++

++

++

Growth

onMcC

onky

agar

minusminus

minusminus

minusminus

minusminus

minusminus

Tentativeidentity

Alteromonas

Halom

onas

Halom

onas

Aidingim

onas

Halom

onas

Halom

onas

Halom

onas

Chromohalobacter

Bacillus

Halom

onas

Character

Bacterialisolates

VW402

SUR108

SUR3

01SU

R302

SUR3

03SU

R307

SUR3

09SU

R310

SUR2

02SU

R201

Morph

ologicalcharacters

Colon

ymorph

olog

yCr

eamcirc

ular

Creamcirc

ular

Whitegum

my

Transparentcircular

Flatreddish-pink

Creamcirc

ular

Creamcirc

ular

Creamcirc

ular

Creamcirc

ular

Creamcirc

ular

Gram

nature

minusminus

minusminus

+minus

minusminus

minusminus

Cellshape

Rod

Rod

Rod

Rod

Cocci

Rod

Rod

Rod

Rod

Rod

Cell

arrang

ement

Sing

leSing

leSing

leSing

leSing

leclu

mped

Sing

leSing

leSing

leSing

leSing

leMotility

+minus

minus+

minus+

minus+

minusminus

Pigm

entatio

nminus

minusminus

minusPink

minusminus

minusminus

minus

Endo

sporeformation

minusminus

minusminus

minusminus

minusminus

minusminus

Physiologicalcharacters

pHrang

efor

grow

th5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

pHop

timum

forg

rowth

97

77

117

77

79

Temprang

efor

grow

th(∘C)

27ndash4

027ndash4

027ndash4

032ndash4

532ndash4

522ndash4

032ndash4

022ndash4

522ndash4

532ndash4

5Tempop

timum

forg

rowth

(∘C)

3232

3732

3727

3227

2732

NaC

lrange

forg

rowth

()

0ndash15

0ndash15

25ndash15

25ndash15

25ndash15

0ndash20

0ndash20

0ndash20

0ndash20

0ndash15

NaC

loptim

umforg

rowth

()

525

575

525

7525

575

Growth

onKingrsquosBmedium

++

++

++

++

++

Growth

onMcC

onky

agar

minusminus

minusminus

minusminus

minusminus

minusminus

Tentativeidentity

Halom

onas

Halom

onas

Chromohalobacter

Halom

onas

Salin

icoccus

Halom

onas

Halom

onas

Halom

onas

Halom

onas

Halom

onas

Chinese Journal of Biology 7

Table8Biochemicalcharacteris

ticso

fthe

selected

EPSprod

ucinghaloph

ilicb

acteria

isolated

from

solarsalternso

fInd

ia

Character

Bacterialisolates

JS 504

JS 802

JS 803

JS 904

JW 302

JW 307

KS 1101

KS 1805

KW 203

VW 301

VW 402

SUR

108

SUR

301

SUR

302

SUR

303

SUR

307

SUR

309

SUR

310

SUR

202

SUR

201

Biochemicalcharacters

Voges-Proskauertest

minusminus

minusminus

minusminus

minusminus

+minus

minusminus

minusminus

minusminus

minusminus

minusminus

Citrateu

tilization

+minus

minusminus

minus+

minus+

minus+

++

+minus

minus+

++

++

Methylred

test

minusminus

minusminus

minusminus

minus+

+minus

minusminus

minusminus

minusminus

minusminus

minusminus

Prod

uctio

nof

Glutaminase

++

++

++

++

++

++

++

minus+

++

++

Asparaginase

++

++

++

++

++

++

++

minus+

++

++

Amylase

+minus

minusminus

minusminus

minusminus

+minus

minusminus

minusminus

minusminus

minusminus

minusminus

Cellulase

minusminus

minusminus

minusminus

minusminus

+minus

minusminus

minusminus

minusminus

minusminus

minusminus

Pectinase

+minus

minusminus

minusminus

minusminus

+minus

minusminus

minusminus

minusminus

minusminus

minusminus

Inulinase

+minus

minusminus

minusminus

minusminus

+minus

minusminus

minusminus

minusminus

minusminus

minusminus

Gelatinase

minusminus

minusminus

minusminus

minusminus

+minus

minusminus

minusminus

minusminus

minusminus

minusminus

Lipase

+minus

minusminus

minusminus

minus+

+minus

minusminus

+minus

minusminus

minusminus

minusminus

Xylanase

+minus

minusminus

minusminus

minusminus

+minus

minusminus

minusminus

minusminus

minusminus

minusminus

Caseinase

minusminus

minus+

minusminus

minusminus

minusminus

minusminus

minus+

minusminus

minusminus

minusminus

Urease

minusminus

minus+

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

Catalase

++

++

++

++

++

++

++

++

++

++

Nitrater

eductase

++

++

++

++

++

++

++

++

++

++

H2S

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

Lysin

edecarbo

xylase

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

Arginined

ecarbo

xylase

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

Ornith

ined

ecarbo

xylase

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

Indo

leminus

minusminus

minusminus

minusminus

+minus

minusminus

minus+

minus+

minusminus

minusminus

minus

Phenylalanined

eaminase

++

+minus

+minus

++

minus+

minus+

++

++

++

minus+

Tryptoph

andeam

inase

++

++

++

++

++

minusminus

++

++

++

+minus

8 Chinese Journal of Biology

Table9Ca

rbon

source

utilizatio

nandferm

entatio

nof

thes

elected

EPSprod

ucinghaloph

ilicb

acteria

isolated

from

samples

ofsolarsalternso

fInd

ia

Carbon

source

Bacterialisolates

JS 504

JS 802

JS 803

JS 904

JW 302

JW 307

KS 1101

KS 1805

KW 203

VW

301

VW

402

SUR

108

SUR

301

SUR

302

SUR

303

SUR

307

SUR

309

SUR

310

SUR

202

SUR

201

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

Glucose

++

++

+minusminus

++minus

+minus

+minus

++

++

+minus

+minus

+minus

+minusminus

++

++minus

+minus

+minusminusminusminusminus

Galactose

+minus

+minus

+minusminus

++minus

+minus

+minus

++

+minus

+minus

+minus

+minus

++minusminus

+minus

++

+minus

+minus

+minus

+minus

Arabino

seminusminusminusminus

+minusminusminusminusminus

+minus

+minus

++

++

+minus

+minus

+minus

+minusminus

++minus

+minus

+minusminusminus

+minus

+minus

Manno

se+minus

+minus

+minusminus

++minus

+minus

+minusminus

++

++minus

+minus

+minus

+minusminusminusminusminus

+minus

+minus

+minus

+minus

+minus

Rham

nose

+minus

+minus

+minus

+minusminusminusminusminusminusminus

++

+minusminusminus

+minus

+minus

++minus

++minus

+minusminusminusminusminusminusminus

+minus

Trehalose

+minus

+minus

+minusminus

+minus

+minusminusminusminus

+minus

++minusminusminusminus

+minus

+minusminus

++

++minusminusminusminusminusminusminus

+minus

Lactose

minusminusminusminus

+minusminusminus

+minus

+minus

+minus

+minus

+minus

+minus

+minusminusminus

+minus

++minusminus

+minusminusminusminusminus

+minus

+minus

Sucrose

+minus

+minus

+minus

++

++

+minus

+minus

+minus

++

+minus

+minusminusminus

+minus

++

+minus

+minus

+minus

+minusminusminus

+minus

Xylose

minusminusminusminus

+minus

+minusminusminus

+minus

++minus

+minusminus

+minus

+minus

+minus

+minusminus

+minus

++

+minusminusminusminus

+minusminusminus

Ribo

se+minus

++

++

++

+minus

+minus

+minus

++

++

+minus

+minus

+minus

++

++

++

+minus

+minus

+minus

+minus

+minus

Fructose

+minus

++

+minus

++

+minus

++

++

+minus

++

++

++

++

++

++

++

++

++

++

++

++

Maltose

minusminusminusminus

+minusminus

+minus

++minus

+minus

+minus

++

+minus

+minus

+minus

++minus

+minus

++minusminusminusminusminus

+minus

+minus

Glycine

minusminusminus

++minus

+minusminus

+minusminusminusminus

+minusminusminusminusminusminusminusminusminus

+minusminusminusminusminus

+minus

+minusminusminusminus

+minusminus

Glycerol

+minus

+minus

+minusminus

++

++minus

+minus

+minus

++

+minus

+minus

+minus

+minusminus

++

++minus

+minus

+minus

+minus

+minus

Adon

itol

+minus

+minus

+minus

+minusminusminus

+minus

+minus

+minus

+minus

+minus

+minusminusminus

+minus

++

+minus

+minus

+minus

+minusminusminusminusminus

Salicin

+minus

++

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

++

++

+minus

+minus

+minus

+minus

+minus

Meso-inosito

l+minusminusminus

+minus

+minus

+minus

+minus

++

+minus

++

+minus

+minusminusminus

+minusminus

++minus

+minus

+minus

+minus

+minus

+minus

Mannitol

+minus

+minus

+minus

++

+minus

+minus

+minus

+minus

++

+minus

+minus

+minus

+minus

+minus

++

+minus

+minus

+minus

+minus

+minus

Sorbito

l+

++

++minusminus

++

+minus

++

++

++

++

++

++

++

+minus

+minus

++

++minus

+minus

++

+minus

Raffino

seminusminus

++

++minusminus

+minus

+minus

+minus

+minus

++minusminusminusminusminusminusminusminus

++

++

+minus

+minus

+minus

+minus

+minus

Cellobiose

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

++

+minus

+minus

+minus

+minus

++

+minus

+minus

+minus

+minus

+minus

+minus

Melibioseminusminusminusminusminusminusminusminusminusminusminusminusminusminusminusminus

++minusminusminusminusminusminusminusminus

++minusminusminusminusminusminusminusminusminusminusminusminus

Na-acetate

+minus

+minus

+minus

+minus

++

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

++

++

+minus

+minus

+minus

+minus

+minus

Na-citrate

+minus

++

+minusminusminusminus

+minusminusminusminusminusminusminusminusminusminusminusminusminusminusminusminusminus

+minusminusminusminusminusminusminusminusminusminusminusminus

Na-fumarate

+minus

+minus

+minus

++

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minusminus

++minus

+minus

+minus

+minus

+minus

+minus

Na-form

ate

+minus

+minus

+minus

+minus

+minusminusminus

+minus

+minus

+minusminusminus

+minusminusminus

+minus

++minusminus

+minusminusminusminusminusminusminusminusminus

Na-benzoate

+minus

++

++

+minus

+minus

+minus

++

++

++

+minus

+minus

++

++

+minus

+minus

+minus

++

++

++

++

Na-malateminusminusminusminusminusminus

++

+minusminusminusminusminusminusminusminusminusminusminusminusminusminusminusminusminus

++

++minusminusminusminusminusminusminusminusminusminus

Na-prop

ionate

+minus

+minus

+minusminusminus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

++minus

++minus

+minus

+minus

+minus

+minus

Na-succinate

+minus

+minus

+minus

++

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

++

++

+minus

+minus

+minus

+minus

+minus

Uutilization

Fferm

entatio

n

Chinese Journal of Biology 9

Table10A

ntibiotic

sensitivitypatte

rnof

thes

electedEP

Sprod

ucinghaloph

ilicb

acteria

isolated

from

solarsalternso

fInd

ia

Antibiotics

Bacterialisolates

JS 504

JS 802

JS 803

JS 904

JW 302

JW 307

KS 1101

KS 1805

KW 203

VW 301

VW 402

SUR

108

SUR

301

SUR

302

SUR

303

SUR

307

SUR

309

SUR

310

SUR

202

SUR

201

Ampicillin(10120583

g)S

SR

SS

SS

RI

SS

SR

SS

SS

SS

SAmoxicillin

(30120583

g)S

SR

RS

SS

RS

SS

SR

RS

IR

SS

SBa

citracin

(10120583

g)S

SR

SS

SR

RS

IR

RR

RS

IR

RR

RCiprofl

oxacin

(30120583

g)I

II

RI

II

IR

II

II

IR

SR

II

ICh

lorte

tracyclin

e(30120583g)

SI

SS

IS

RR

SS

SS

RR

SS

II

IS

Chloramph

enicol(10120583

g)S

SS

SS

SS

SR

SS

SS

SS

SR

SS

SErythrom

ycin

(15120583

g)S

SI

SS

IS

II

II

II

SR

RI

IS

SFu

sidicacid

(30120583

g)I

RR

SS

IR

RS

RI

RR

RS

IR

II

RGentamycin

(10120583

g)S

SS

SS

SS

SR

SS

SS

SS

IS

SS

SKa

namycin

(30120583

g)I

IS

RS

II

SR

II

RI

RR

RI

RI

RMethicillin(5120583g)

SS

RS

IR

RR

RR

RR

RR

SR

RR

RR

Neomycin

(30120583

g)S

SI

RI

IR

II

IR

II

RR

RI

IR

RNorflo

xacin(30120583

g)S

SR

SS

SS

SI

SS

SS

SS

SS

SS

SNetillin

(10120583

g)R

SS

IR

SS

SR

RR

SS

RR

RR

RR

RNalidixicacid

(30120583

g)R

RS

RR

II

RR

II

RR

RR

SR

RR

RNovob

iosin

(30120583

g)I

RR

SR

SS

RS

SS

IR

SS

SI

SS

SPo

lymixin

B(50120583

g)R

RI

RI

IR

RR

RI

IR

RR

II

II

SPenicillinG(10120583

g)S

SR

IS

IR

RR

II

SR

RS

II

II

SStreptom

ycin

(10120583

g)S

SS

RS

RR

SR

RR

IS

RS

SR

II

RTetracyclin

e(30120583g)

RR

RR

RI

RR

SR

IR

RR

RS

RR

RI

Trim

etho

prim

e(30120583g)

SS

SS

IS

SI

SS

SS

IR

RS

SS

SS

Vancom

ycin

(30120583

g)R

RR

SR

RR

RS

RR

RR

RS

RR

RR

R

S=SensitiveR

=Re

sistantI

=Interm

ediate

10 Chinese Journal of Biology

0

1

2

3

4

Bacterial isolate

Gro

wth

OD

at540

nmto

tal E

PS (g

L)

OD at 540nmTotal EPS

JS504

JS802

JS803

JS904

JW302

JW307

KS1101

KS1805

KW203

VW301

VW402

SUR1

08

SUR3

01

SUR3

02

SUR3

03

SUR3

07

SUR3

09

SUR3

10

SUR2

02

SUR2

01

Figure 1 Screening of moderately halophilic bacteria isolatedfrom soil and water samples of solar salterns for production ofextracellular polymeric substances

0

02

04

06

08

1

Ant

ibio

tic re

sista

nce i

ndex

Bacterial isolate

JS504

JS802

JS803

JS904

JW302

JW307

KS1101

KS1805

KW203

SUR1

08

SUR2

01

SUR2

02

SUR3

01

SUR3

02

SUR3

03

SUR3

07

SUR3

09

SUR3

10

VW301

VW402

Figure 2 Antibiotic resistance index of the selected EPS producinghalophilic bacteria isolated from soil and water samples of solarsalterns of India

The isolates SUR301 and KS 1805 were tentatively iden-tified as Chromohalobacter based on the study by Arahal etal [45] and bear striking similarity with those of Chromo-halobacter sp isolated fromhypersaline soil sample of TriveniSangam Gujarat [38] The physiological morphological andbiochemical characteristics of isolate SUR303 were consistentwith the features of the genus Salinicoccus [10] while isolatesJS904 and JS504 were tentatively assigned to Aidingimonasand Alteromonas respectively

Finally sensitivity of these isolates to antibiotics (Table 10and Figure 2) also corroborates the findings ofMata et al [25]and Hedi et al [28]

5 Conclusion

It may be emphasized that study of these moderatelyhalophilic bacteria from coastal hypersaline solar salterns ofIndia with special attempt on screening for EPS productionhas led to the discovery of wide variety of halophilic speciesAlthough attention has been focused on the production

of EPS by these halophilic bacterial isolates their physio-biochemical features indicate that they may equally holdpotential towards production of various biomolecules ofindustrial interest

Conflict of Interests

It is declared by the authors that there is no conflict of interestsregarding the publication of this paper

Acknowledgment

This study was financially supported by grants from Uni-versity Grants Commission India (Sanction no F14-2(SC)2008 (SA-III) 31 March 2009) under the Scheme ofRajiv Gandhi National Fellowship

References

[1] E A Galinski and B J Tindall ldquoBiotechnological prospectsfor halophiles and halotolerant microorganismsrdquo in MolecularBiology and Biotechnology of Extremophiles R D Herbert andR J Sharp Eds Blackie London UK 1992

[2] R Margesin and F Schinner ldquoPotential of halotolerant andhalophilic microorganisms for biotechnologyrdquo Extremophilesvol 5 no 2 pp 73ndash83 2001

[3] F Rodriguez Valera F Ruiz Berraquero and A Ramos Cor-menzana ldquoCharacteristics of the heterotrophic bacterial pop-ulations in hypersaline environments of different salt concen-trationsrdquoMicrobial Ecology vol 7 no 3 pp 235ndash243 1981

[4] F Rodriguez-Valera ldquoCharacteristics and microbial ecol-ogy of hypersaline environmentsrdquo in Halophilic Bacteria FRodriguez-Valera Ed vol 1 pp 3ndash30 CRC Press Boca RatonFla USA 1988

[5] D J Kushner and M Kamekura ldquoPhysiology of halophiliceubacteriardquo inHalophilic Bacteria F Rodriguez-Valera Ed vol1 pp 109ndash138 CRC press Boca Raton Fla USA 1988

[6] M A Amoozegar F Malekzadeh and K AMalik ldquoProductionof amylase by newly isolated moderate halophile Halobacillussp strainMA-2rdquo Journal of Microbiological Methods vol 52 no3 pp 353ndash359 2003

[7] C O Jeon J Lim J Lee et al ldquoLentibacillus salarius spnov isolated from saline sediment in China and emendeddescription of the genus Lentibacillusrdquo International Journal ofSystematic and Evolutionary Microbiology vol 55 no 3 pp1339ndash1343 2005

[8] S Bouchotroch E Quesada A Del Moral I Llamas andV Bejar ldquoHalomonas maura sp nov a novel moderatelyhalophilic exopolysaccharide-producing bacteriumrdquo Interna-tional Journal of Systematic and Evolutionary Microbiology vol51 no 5 pp 1625ndash1632 2001

[9] A Biswas A Patra and A Paul ldquoProduction of poly-3-hydroxyalkanoic acids by a moderately halophilic bacteriumHalomonas marina HMA 103 isolated from solar saltern ofOrissa Indiardquo Acta Microbiologica et Immunologica Hungaricavol 56 no 2 pp 125ndash143 2009

[10] S Y Jayachandra S Anil Kumar D P Merley and MB Sulochana ldquoIsolation and characterization of extremehalophilic bacterium Salinicoccus sp JAS4 producing extra-cellular hydrolytic enzymesrdquo Recent Research in Science andTechnology vol 4 no 4 pp 46ndash49 2012

Chinese Journal of Biology 11

[11] I W Sutherland ldquoNovel and established applications of micro-bial polysaccharidesrdquo Trends in Biotechnology vol 16 no 1 pp41ndash46 1998

[12] MWMittelman andG G Geesey ldquoCopper-binding character-istics of exopolymers from a freshwater-sediment bacteriumrdquoApplied and EnvironmentalMicrobiology vol 49 no 4 pp 846ndash851 1985

[13] J Anton I Meseguer and F Rodriguez-Valera ldquoProductionof an extracellular polysaccharide by Haloferax mediterraneirdquoApplied Enviromental Microbiology vol 54 no 10 pp 2381ndash2386 1988

[14] H Parolis L A S Parolis I F Boan et al ldquoThe structure ofthe exopolysaccharide produced by the halophilic ArchaeonHaloferax mediterranei strain R4 (ATCC 33500)rdquo CarbohydrateResearch vol 295 pp 147ndash156 1996

[15] E Quesada V Bejar and C Calvo ldquoExopolysaccharide produc-tion by Volcaniella eurihalinardquo Experientia vol 49 no 12 pp1037ndash1041 1993

[16] J A Mata V Bejar I Llamas et al ldquoExopolysaccharides pro-duced by the recently described halophilic bacteria Halomonasventosae and Halomonas anticariensisrdquo Research in Microbiol-ogy vol 157 no 9 pp 827ndash835 2006

[17] I Llamas J A Mata R Tallon et al ldquoCharacterization ofthe exopolysaccharide produced by Salipiger mucosus A3119879 ahalophilic species belonging to the Alphaproteobacteria iso-lated on the Spanish Mediterranean seaboardrdquo Marine Drugsvol 8 no 8 pp 2240ndash2251 2010

[18] F Martınez-Checa E Quesada J Martınez-Canovas I Llamasand V Bejar ldquoPalleronia marisminoris gen nov sp nova moderately halophilic exopolysaccharide-producing bac-terium belonging to the rsquoAlphaproteobacteriarsquo isolated from asaline soilrdquo International Journal of Systematic and EvolutionaryMicrobiology vol 55 no 6 pp 2525ndash2530 2005

[19] A Ventosa M T Garcia M Kamekura H Onishi and FRuiz-Berraquero ldquoBacillus halophilus sp nov a moderatelyhalophilic Bacillus speciesrdquo Systematic and Applied Microbiol-ogy vol 12 no 2 pp 162ndash166 1989

[20] R A Moraine and P Rogovin ldquoKinetics of polysaccharide B1459 fermentationrdquoBiotechnology and Bioengineering vol 8 no4 pp 511ndash524 1966

[21] M Dubois K A Gilles J K Hamilton P A Rebers and FSmith ldquoColorimetric method for determination of sugars andrelated substancesrdquoAnalytical Chemistry vol 28 no 3 pp 350ndash356 1956

[22] O H Lowry N J Rosenbrough A L Farr and R J RandallldquoProtein measurement with the Folin phenol reagentrdquo TheJournal of Biological Chemistry vol 193 no 1 pp 265ndash275 1951

[23] P Gerhardt R G E Murray W A Wood and N R KriegMethods for General and Molecular Bacteriology AmericanSociety for Microbiology Washington DC USA 1994

[24] G M Garrity J A Bell and T G Lilburn ldquoTaxonomic outlineof the procaryotesrdquo inBergeyrsquosManual of Systematic BacteriologD R Boone and R W Castenholz Eds Springer New YorkNY USA 2nd edition 2003

[25] J A Mata J Martınez-Canovas E Quesada and V BejarldquoA detailed phenotypic characterisation of the type strains ofHalomonas speciesrdquo Systematic and Applied Microbiology vol25 no 3 pp 360ndash375 2002

[26] A Oren ldquoMolecular ecology of extremely halophilic archaeaand bacteriardquo FEMSMicrobiology Ecology vol 39 no 1 pp 1ndash72002

[27] H Baati R Amdouni N Gharsallah A Sghir and E AmmarldquoIsolation and characterization of moderately halophilic bacte-ria from tunisian solar salternrdquo Current Microbiology vol 60no 3 pp 157ndash161 2010

[28] A Hedi N Sadfi M Fardeau et al ldquoStudies on the biodiversityof halophilic microorganisms isolated from El-Djerid salt lake(Tunisia) under aerobic conditionsrdquo International Journal ofMicrobiology vol 2009 Article ID 731786 17 pages 2009

[29] M Birbir A Ogan B Calli and B Mertoglu ldquoEnzymecharacteristics of extremely halophilic archaeal community inTuzkoy Salt Mine Turkeyrdquo World Journal of Microbiology andBiotechnology vol 20 no 6 pp 613ndash621 2004

[30] M Birbir and C Sesal ldquoExtremely halophilic bacterial commu-nities in Sereflikochisar Salt Lake in Turkeyrdquo Turkish Journal ofBiology vol 27 no 7 pp 7ndash22 2003

[31] M Birbir B Calli B Mertoglu et al ldquoExtremely halophilicArchaea from Tuz Lake Turkey and the adjacent Kaldirim andKayacik salternsrdquoWorld Journal ofMicrobiology and Biotechnol-ogy vol 23 no 3 pp 309ndash316 2007

[32] R Rohban M A Amoozegar and A Ventosa ldquoScreeningand isolation of halophilic bacteria producing extracellularhydrolyses from Howz Soltan Lake Iranrdquo Journal of IndustrialMicrobiology and Biotechnology vol 36 no 3 pp 333ndash3402009

[33] M J Garabito M C Marquez and A Ventosa ldquoHalotolerantBacillus diversity in hypersaline environmentsrdquo Canadian Jour-nal of Microbiology vol 44 no 2 pp 95ndash102 1998

[34] A Ventosa A Ramos-Cormenzana and M Kocur ldquoModer-ately halophilic gram-positive cocci from hypersaline environ-mentsrdquo Systematic and Applied Microbiology vol 4 no 4 pp564ndash570 1983

[35] E Quesada A Ventosa F Rodriguez-Valera and A R Cor-menzana ldquoTypes and properties of some bacteria isolated fromhypersaline soilsrdquo Journal of Applied Bacteriology vol 53 no 2pp 155ndash161 1982

[36] E Quesada V Bejar M J Valderrama A Ventosa and AR Ramos Cormenzana ldquoIsolation and characterization ofmoderately halophilic nonmotile rods from different salinehabitatsrdquoMicrobiologia vol 1 no 1-2 pp 89ndash96 1985

[37] S R Dave and H B Desai ldquoMicrobial diversity at marinesalterns near Bhavnagar Gujarat Indiardquo Current Science vol60 no 4 pp 497ndash500 2006

[38] S Kumar R Karan S Kapoor S P Singh and S K KhareldquoScreening and isolation of halophilic bacteria producingindustrially important enzymesrdquo Brazilian Journal of Microbi-ology vol 43 no 4 pp 1595ndash1603 2012

[39] B P Dave and A Soni ldquoDiversity of halophilic archaea atsalt pans around Bhavnagar coast Gujaratrdquo Proceedings ofthe National Academy of Sciences India Section B BiologicalSciences vol 83 no 2 pp 225ndash232 2013

[40] A A Joshi P P Kanekar A S Kelkar et al ldquoCultivable bacterialdiversity of alkaline Lonar lake Indiardquo Microbial Ecology vol55 no 2 pp 163ndash172 2008

[41] T M Raghavan and I Furtado ldquoOccurrence of extremelyhalophilic Archaea in sediments from the continental shelf ofwest coast of Indiardquo Current Science vol 86 no 8 pp 1065ndash1067 2004

[42] S Vijayanand J Hemapriya J Selvin and S Kiran ldquoBiodiver-sity of extremely halophilic bacterial strains isolated from solarsalterns of Tuticorin Tamilnadu Indiardquo International Journal ofWater Resources and Environmental Sciences vol 1 no 1 pp 1ndash72012

12 Chinese Journal of Biology

[43] S G Nanjani and H P Soni ldquoIsolation and characterization ofextremely halotolerant and halophilic organisms from Dwarkaand Veravalrdquo Journal of Pharmacy and Biological Sciences vol2 no 2 pp 20ndash25 2012

[44] H Ghozlan H Deif R A Kandil and S Sabry ldquoBiodiversity ofmoderately halophilic bacteria in hypersaline habitats in EgyptrdquoJournal of General and Applied Microbiology vol 52 no 2 pp63ndash72 2006

[45] D R Arahal M T Garcıa W Ludwig K H Schleifer andA Ventosa ldquoTransfer ofHalomonas canadensis andHalomonasisraelensis to the genus Chromohalobacter as Chromohalobac-ter canadensis comb nov and Chromohalobacter israelensiscomb novrdquo International Journal of Systematic and EvolutionaryMicrobiology vol 51 no 4 pp 1443ndash1448 2001

Submit your manuscripts athttpwwwhindawicom

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Anatomy Research International

PeptidesInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporation httpwwwhindawicom

International Journal of

Volume 2014

Zoology

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Molecular Biology International

GenomicsInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

The Scientific World JournalHindawi Publishing Corporation httpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

BioinformaticsAdvances in

Marine BiologyJournal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Signal TransductionJournal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

BioMed Research International

Evolutionary BiologyInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Biochemistry Research International

ArchaeaHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

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Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Advances in

Virolog y

Hindawi Publishing Corporationhttpwwwhindawicom

Nucleic AcidsJournal of

Volume 2014

Stem CellsInternational

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Enzyme Research

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

International Journal of

Microbiology

Page 6: Research Article Production of Extracellular …downloads.hindawi.com/archive/2014/205731.pdfResearch Article Production of Extracellular Polymeric Substances by Halophilic Bacteria

6 Chinese Journal of Biology

Table7Morph

ologicalandph

ysiologicalcharacteristicso

fthe

selected

EPSprod

ucinghaloph

ilicb

acteria

isolated

from

solarsalternso

fInd

ia

Character

Bacterialisolates

JS504

JS802

JS803

JS904

JW302

JW307

KS110

1KS

1805

KW203

VW301

Morph

ologicalcharacters

Colon

ymorph

olog

yCr

eamcirc

ular

Whitecirc

ular

Creamcirc

ular

Creamcirc

ular

Transparentcircular

Creamcirc

ular

Creamcirc

ular

Whitecirc

ular

Whitecirc

ular

Creamcirc

ular

Gram

nature

minus+

minusminus

minusminus

minusminus

+minus

Cellshape

Rod

Rod

Rod

Rod

Rod

Rod

Rod

Rod

Rod

Rod

Cell

arrang

ement

Sing

leSing

leSing

leSing

leSing

lechains

Sing

lechains

Sing

lechains

Sing

leSing

leclu

mped

Sing

lechains

Motility

++

++

++

minusminus

++

Pigm

entatio

nminus

minusminus

minusminus

minusminus

minusminus

minus

Endo

sporeformation

minusminus

minusminus

minusminus

minusminus

+minus

Physiologicalcharacters

pHrang

efor

grow

th5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

pHop

timum

forg

rowth

119

89

1111

118

89

Temprang

efor

grow

th(∘C)

27ndash4

027ndash4

027ndash4

027ndash4

027ndash4

027ndash4

022ndash4

027ndash4

027ndash4

022ndash4

0Tempop

timum

forg

rowth

(∘C)

3237

3732

3232

3232

3732

NaC

lrange

forg

rowth

()

0ndash15

0ndash20

0ndash15

0ndash20

0ndash15

0ndash20

0ndash20

25ndash15

25ndash15

0ndash20

NaC

loptim

umforg

rowth

()

510

105

105

575

105

Growth

onKingrsquosBmedium

++

++

++

++

++

Growth

onMcC

onky

agar

minusminus

minusminus

minusminus

minusminus

minusminus

Tentativeidentity

Alteromonas

Halom

onas

Halom

onas

Aidingim

onas

Halom

onas

Halom

onas

Halom

onas

Chromohalobacter

Bacillus

Halom

onas

Character

Bacterialisolates

VW402

SUR108

SUR3

01SU

R302

SUR3

03SU

R307

SUR3

09SU

R310

SUR2

02SU

R201

Morph

ologicalcharacters

Colon

ymorph

olog

yCr

eamcirc

ular

Creamcirc

ular

Whitegum

my

Transparentcircular

Flatreddish-pink

Creamcirc

ular

Creamcirc

ular

Creamcirc

ular

Creamcirc

ular

Creamcirc

ular

Gram

nature

minusminus

minusminus

+minus

minusminus

minusminus

Cellshape

Rod

Rod

Rod

Rod

Cocci

Rod

Rod

Rod

Rod

Rod

Cell

arrang

ement

Sing

leSing

leSing

leSing

leSing

leclu

mped

Sing

leSing

leSing

leSing

leSing

leMotility

+minus

minus+

minus+

minus+

minusminus

Pigm

entatio

nminus

minusminus

minusPink

minusminus

minusminus

minus

Endo

sporeformation

minusminus

minusminus

minusminus

minusminus

minusminus

Physiologicalcharacters

pHrang

efor

grow

th5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

5ndash11

pHop

timum

forg

rowth

97

77

117

77

79

Temprang

efor

grow

th(∘C)

27ndash4

027ndash4

027ndash4

032ndash4

532ndash4

522ndash4

032ndash4

022ndash4

522ndash4

532ndash4

5Tempop

timum

forg

rowth

(∘C)

3232

3732

3727

3227

2732

NaC

lrange

forg

rowth

()

0ndash15

0ndash15

25ndash15

25ndash15

25ndash15

0ndash20

0ndash20

0ndash20

0ndash20

0ndash15

NaC

loptim

umforg

rowth

()

525

575

525

7525

575

Growth

onKingrsquosBmedium

++

++

++

++

++

Growth

onMcC

onky

agar

minusminus

minusminus

minusminus

minusminus

minusminus

Tentativeidentity

Halom

onas

Halom

onas

Chromohalobacter

Halom

onas

Salin

icoccus

Halom

onas

Halom

onas

Halom

onas

Halom

onas

Halom

onas

Chinese Journal of Biology 7

Table8Biochemicalcharacteris

ticso

fthe

selected

EPSprod

ucinghaloph

ilicb

acteria

isolated

from

solarsalternso

fInd

ia

Character

Bacterialisolates

JS 504

JS 802

JS 803

JS 904

JW 302

JW 307

KS 1101

KS 1805

KW 203

VW 301

VW 402

SUR

108

SUR

301

SUR

302

SUR

303

SUR

307

SUR

309

SUR

310

SUR

202

SUR

201

Biochemicalcharacters

Voges-Proskauertest

minusminus

minusminus

minusminus

minusminus

+minus

minusminus

minusminus

minusminus

minusminus

minusminus

Citrateu

tilization

+minus

minusminus

minus+

minus+

minus+

++

+minus

minus+

++

++

Methylred

test

minusminus

minusminus

minusminus

minus+

+minus

minusminus

minusminus

minusminus

minusminus

minusminus

Prod

uctio

nof

Glutaminase

++

++

++

++

++

++

++

minus+

++

++

Asparaginase

++

++

++

++

++

++

++

minus+

++

++

Amylase

+minus

minusminus

minusminus

minusminus

+minus

minusminus

minusminus

minusminus

minusminus

minusminus

Cellulase

minusminus

minusminus

minusminus

minusminus

+minus

minusminus

minusminus

minusminus

minusminus

minusminus

Pectinase

+minus

minusminus

minusminus

minusminus

+minus

minusminus

minusminus

minusminus

minusminus

minusminus

Inulinase

+minus

minusminus

minusminus

minusminus

+minus

minusminus

minusminus

minusminus

minusminus

minusminus

Gelatinase

minusminus

minusminus

minusminus

minusminus

+minus

minusminus

minusminus

minusminus

minusminus

minusminus

Lipase

+minus

minusminus

minusminus

minus+

+minus

minusminus

+minus

minusminus

minusminus

minusminus

Xylanase

+minus

minusminus

minusminus

minusminus

+minus

minusminus

minusminus

minusminus

minusminus

minusminus

Caseinase

minusminus

minus+

minusminus

minusminus

minusminus

minusminus

minus+

minusminus

minusminus

minusminus

Urease

minusminus

minus+

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

Catalase

++

++

++

++

++

++

++

++

++

++

Nitrater

eductase

++

++

++

++

++

++

++

++

++

++

H2S

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

Lysin

edecarbo

xylase

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

Arginined

ecarbo

xylase

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

Ornith

ined

ecarbo

xylase

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

Indo

leminus

minusminus

minusminus

minusminus

+minus

minusminus

minus+

minus+

minusminus

minusminus

minus

Phenylalanined

eaminase

++

+minus

+minus

++

minus+

minus+

++

++

++

minus+

Tryptoph

andeam

inase

++

++

++

++

++

minusminus

++

++

++

+minus

8 Chinese Journal of Biology

Table9Ca

rbon

source

utilizatio

nandferm

entatio

nof

thes

elected

EPSprod

ucinghaloph

ilicb

acteria

isolated

from

samples

ofsolarsalternso

fInd

ia

Carbon

source

Bacterialisolates

JS 504

JS 802

JS 803

JS 904

JW 302

JW 307

KS 1101

KS 1805

KW 203

VW

301

VW

402

SUR

108

SUR

301

SUR

302

SUR

303

SUR

307

SUR

309

SUR

310

SUR

202

SUR

201

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

Glucose

++

++

+minusminus

++minus

+minus

+minus

++

++

+minus

+minus

+minus

+minusminus

++

++minus

+minus

+minusminusminusminusminus

Galactose

+minus

+minus

+minusminus

++minus

+minus

+minus

++

+minus

+minus

+minus

+minus

++minusminus

+minus

++

+minus

+minus

+minus

+minus

Arabino

seminusminusminusminus

+minusminusminusminusminus

+minus

+minus

++

++

+minus

+minus

+minus

+minusminus

++minus

+minus

+minusminusminus

+minus

+minus

Manno

se+minus

+minus

+minusminus

++minus

+minus

+minusminus

++

++minus

+minus

+minus

+minusminusminusminusminus

+minus

+minus

+minus

+minus

+minus

Rham

nose

+minus

+minus

+minus

+minusminusminusminusminusminusminus

++

+minusminusminus

+minus

+minus

++minus

++minus

+minusminusminusminusminusminusminus

+minus

Trehalose

+minus

+minus

+minusminus

+minus

+minusminusminusminus

+minus

++minusminusminusminus

+minus

+minusminus

++

++minusminusminusminusminusminusminus

+minus

Lactose

minusminusminusminus

+minusminusminus

+minus

+minus

+minus

+minus

+minus

+minus

+minusminusminus

+minus

++minusminus

+minusminusminusminusminus

+minus

+minus

Sucrose

+minus

+minus

+minus

++

++

+minus

+minus

+minus

++

+minus

+minusminusminus

+minus

++

+minus

+minus

+minus

+minusminusminus

+minus

Xylose

minusminusminusminus

+minus

+minusminusminus

+minus

++minus

+minusminus

+minus

+minus

+minus

+minusminus

+minus

++

+minusminusminusminus

+minusminusminus

Ribo

se+minus

++

++

++

+minus

+minus

+minus

++

++

+minus

+minus

+minus

++

++

++

+minus

+minus

+minus

+minus

+minus

Fructose

+minus

++

+minus

++

+minus

++

++

+minus

++

++

++

++

++

++

++

++

++

++

++

++

Maltose

minusminusminusminus

+minusminus

+minus

++minus

+minus

+minus

++

+minus

+minus

+minus

++minus

+minus

++minusminusminusminusminus

+minus

+minus

Glycine

minusminusminus

++minus

+minusminus

+minusminusminusminus

+minusminusminusminusminusminusminusminusminus

+minusminusminusminusminus

+minus

+minusminusminusminus

+minusminus

Glycerol

+minus

+minus

+minusminus

++

++minus

+minus

+minus

++

+minus

+minus

+minus

+minusminus

++

++minus

+minus

+minus

+minus

+minus

Adon

itol

+minus

+minus

+minus

+minusminusminus

+minus

+minus

+minus

+minus

+minus

+minusminusminus

+minus

++

+minus

+minus

+minus

+minusminusminusminusminus

Salicin

+minus

++

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

++

++

+minus

+minus

+minus

+minus

+minus

Meso-inosito

l+minusminusminus

+minus

+minus

+minus

+minus

++

+minus

++

+minus

+minusminusminus

+minusminus

++minus

+minus

+minus

+minus

+minus

+minus

Mannitol

+minus

+minus

+minus

++

+minus

+minus

+minus

+minus

++

+minus

+minus

+minus

+minus

+minus

++

+minus

+minus

+minus

+minus

+minus

Sorbito

l+

++

++minusminus

++

+minus

++

++

++

++

++

++

++

+minus

+minus

++

++minus

+minus

++

+minus

Raffino

seminusminus

++

++minusminus

+minus

+minus

+minus

+minus

++minusminusminusminusminusminusminusminus

++

++

+minus

+minus

+minus

+minus

+minus

Cellobiose

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

++

+minus

+minus

+minus

+minus

++

+minus

+minus

+minus

+minus

+minus

+minus

Melibioseminusminusminusminusminusminusminusminusminusminusminusminusminusminusminusminus

++minusminusminusminusminusminusminusminus

++minusminusminusminusminusminusminusminusminusminusminusminus

Na-acetate

+minus

+minus

+minus

+minus

++

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

++

++

+minus

+minus

+minus

+minus

+minus

Na-citrate

+minus

++

+minusminusminusminus

+minusminusminusminusminusminusminusminusminusminusminusminusminusminusminusminusminus

+minusminusminusminusminusminusminusminusminusminusminusminus

Na-fumarate

+minus

+minus

+minus

++

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minusminus

++minus

+minus

+minus

+minus

+minus

+minus

Na-form

ate

+minus

+minus

+minus

+minus

+minusminusminus

+minus

+minus

+minusminusminus

+minusminusminus

+minus

++minusminus

+minusminusminusminusminusminusminusminusminus

Na-benzoate

+minus

++

++

+minus

+minus

+minus

++

++

++

+minus

+minus

++

++

+minus

+minus

+minus

++

++

++

++

Na-malateminusminusminusminusminusminus

++

+minusminusminusminusminusminusminusminusminusminusminusminusminusminusminusminusminus

++

++minusminusminusminusminusminusminusminusminusminus

Na-prop

ionate

+minus

+minus

+minusminusminus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

++minus

++minus

+minus

+minus

+minus

+minus

Na-succinate

+minus

+minus

+minus

++

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

++

++

+minus

+minus

+minus

+minus

+minus

Uutilization

Fferm

entatio

n

Chinese Journal of Biology 9

Table10A

ntibiotic

sensitivitypatte

rnof

thes

electedEP

Sprod

ucinghaloph

ilicb

acteria

isolated

from

solarsalternso

fInd

ia

Antibiotics

Bacterialisolates

JS 504

JS 802

JS 803

JS 904

JW 302

JW 307

KS 1101

KS 1805

KW 203

VW 301

VW 402

SUR

108

SUR

301

SUR

302

SUR

303

SUR

307

SUR

309

SUR

310

SUR

202

SUR

201

Ampicillin(10120583

g)S

SR

SS

SS

RI

SS

SR

SS

SS

SS

SAmoxicillin

(30120583

g)S

SR

RS

SS

RS

SS

SR

RS

IR

SS

SBa

citracin

(10120583

g)S

SR

SS

SR

RS

IR

RR

RS

IR

RR

RCiprofl

oxacin

(30120583

g)I

II

RI

II

IR

II

II

IR

SR

II

ICh

lorte

tracyclin

e(30120583g)

SI

SS

IS

RR

SS

SS

RR

SS

II

IS

Chloramph

enicol(10120583

g)S

SS

SS

SS

SR

SS

SS

SS

SR

SS

SErythrom

ycin

(15120583

g)S

SI

SS

IS

II

II

II

SR

RI

IS

SFu

sidicacid

(30120583

g)I

RR

SS

IR

RS

RI

RR

RS

IR

II

RGentamycin

(10120583

g)S

SS

SS

SS

SR

SS

SS

SS

IS

SS

SKa

namycin

(30120583

g)I

IS

RS

II

SR

II

RI

RR

RI

RI

RMethicillin(5120583g)

SS

RS

IR

RR

RR

RR

RR

SR

RR

RR

Neomycin

(30120583

g)S

SI

RI

IR

II

IR

II

RR

RI

IR

RNorflo

xacin(30120583

g)S

SR

SS

SS

SI

SS

SS

SS

SS

SS

SNetillin

(10120583

g)R

SS

IR

SS

SR

RR

SS

RR

RR

RR

RNalidixicacid

(30120583

g)R

RS

RR

II

RR

II

RR

RR

SR

RR

RNovob

iosin

(30120583

g)I

RR

SR

SS

RS

SS

IR

SS

SI

SS

SPo

lymixin

B(50120583

g)R

RI

RI

IR

RR

RI

IR

RR

II

II

SPenicillinG(10120583

g)S

SR

IS

IR

RR

II

SR

RS

II

II

SStreptom

ycin

(10120583

g)S

SS

RS

RR

SR

RR

IS

RS

SR

II

RTetracyclin

e(30120583g)

RR

RR

RI

RR

SR

IR

RR

RS

RR

RI

Trim

etho

prim

e(30120583g)

SS

SS

IS

SI

SS

SS

IR

RS

SS

SS

Vancom

ycin

(30120583

g)R

RR

SR

RR

RS

RR

RR

RS

RR

RR

R

S=SensitiveR

=Re

sistantI

=Interm

ediate

10 Chinese Journal of Biology

0

1

2

3

4

Bacterial isolate

Gro

wth

OD

at540

nmto

tal E

PS (g

L)

OD at 540nmTotal EPS

JS504

JS802

JS803

JS904

JW302

JW307

KS1101

KS1805

KW203

VW301

VW402

SUR1

08

SUR3

01

SUR3

02

SUR3

03

SUR3

07

SUR3

09

SUR3

10

SUR2

02

SUR2

01

Figure 1 Screening of moderately halophilic bacteria isolatedfrom soil and water samples of solar salterns for production ofextracellular polymeric substances

0

02

04

06

08

1

Ant

ibio

tic re

sista

nce i

ndex

Bacterial isolate

JS504

JS802

JS803

JS904

JW302

JW307

KS1101

KS1805

KW203

SUR1

08

SUR2

01

SUR2

02

SUR3

01

SUR3

02

SUR3

03

SUR3

07

SUR3

09

SUR3

10

VW301

VW402

Figure 2 Antibiotic resistance index of the selected EPS producinghalophilic bacteria isolated from soil and water samples of solarsalterns of India

The isolates SUR301 and KS 1805 were tentatively iden-tified as Chromohalobacter based on the study by Arahal etal [45] and bear striking similarity with those of Chromo-halobacter sp isolated fromhypersaline soil sample of TriveniSangam Gujarat [38] The physiological morphological andbiochemical characteristics of isolate SUR303 were consistentwith the features of the genus Salinicoccus [10] while isolatesJS904 and JS504 were tentatively assigned to Aidingimonasand Alteromonas respectively

Finally sensitivity of these isolates to antibiotics (Table 10and Figure 2) also corroborates the findings ofMata et al [25]and Hedi et al [28]

5 Conclusion

It may be emphasized that study of these moderatelyhalophilic bacteria from coastal hypersaline solar salterns ofIndia with special attempt on screening for EPS productionhas led to the discovery of wide variety of halophilic speciesAlthough attention has been focused on the production

of EPS by these halophilic bacterial isolates their physio-biochemical features indicate that they may equally holdpotential towards production of various biomolecules ofindustrial interest

Conflict of Interests

It is declared by the authors that there is no conflict of interestsregarding the publication of this paper

Acknowledgment

This study was financially supported by grants from Uni-versity Grants Commission India (Sanction no F14-2(SC)2008 (SA-III) 31 March 2009) under the Scheme ofRajiv Gandhi National Fellowship

References

[1] E A Galinski and B J Tindall ldquoBiotechnological prospectsfor halophiles and halotolerant microorganismsrdquo in MolecularBiology and Biotechnology of Extremophiles R D Herbert andR J Sharp Eds Blackie London UK 1992

[2] R Margesin and F Schinner ldquoPotential of halotolerant andhalophilic microorganisms for biotechnologyrdquo Extremophilesvol 5 no 2 pp 73ndash83 2001

[3] F Rodriguez Valera F Ruiz Berraquero and A Ramos Cor-menzana ldquoCharacteristics of the heterotrophic bacterial pop-ulations in hypersaline environments of different salt concen-trationsrdquoMicrobial Ecology vol 7 no 3 pp 235ndash243 1981

[4] F Rodriguez-Valera ldquoCharacteristics and microbial ecol-ogy of hypersaline environmentsrdquo in Halophilic Bacteria FRodriguez-Valera Ed vol 1 pp 3ndash30 CRC Press Boca RatonFla USA 1988

[5] D J Kushner and M Kamekura ldquoPhysiology of halophiliceubacteriardquo inHalophilic Bacteria F Rodriguez-Valera Ed vol1 pp 109ndash138 CRC press Boca Raton Fla USA 1988

[6] M A Amoozegar F Malekzadeh and K AMalik ldquoProductionof amylase by newly isolated moderate halophile Halobacillussp strainMA-2rdquo Journal of Microbiological Methods vol 52 no3 pp 353ndash359 2003

[7] C O Jeon J Lim J Lee et al ldquoLentibacillus salarius spnov isolated from saline sediment in China and emendeddescription of the genus Lentibacillusrdquo International Journal ofSystematic and Evolutionary Microbiology vol 55 no 3 pp1339ndash1343 2005

[8] S Bouchotroch E Quesada A Del Moral I Llamas andV Bejar ldquoHalomonas maura sp nov a novel moderatelyhalophilic exopolysaccharide-producing bacteriumrdquo Interna-tional Journal of Systematic and Evolutionary Microbiology vol51 no 5 pp 1625ndash1632 2001

[9] A Biswas A Patra and A Paul ldquoProduction of poly-3-hydroxyalkanoic acids by a moderately halophilic bacteriumHalomonas marina HMA 103 isolated from solar saltern ofOrissa Indiardquo Acta Microbiologica et Immunologica Hungaricavol 56 no 2 pp 125ndash143 2009

[10] S Y Jayachandra S Anil Kumar D P Merley and MB Sulochana ldquoIsolation and characterization of extremehalophilic bacterium Salinicoccus sp JAS4 producing extra-cellular hydrolytic enzymesrdquo Recent Research in Science andTechnology vol 4 no 4 pp 46ndash49 2012

Chinese Journal of Biology 11

[11] I W Sutherland ldquoNovel and established applications of micro-bial polysaccharidesrdquo Trends in Biotechnology vol 16 no 1 pp41ndash46 1998

[12] MWMittelman andG G Geesey ldquoCopper-binding character-istics of exopolymers from a freshwater-sediment bacteriumrdquoApplied and EnvironmentalMicrobiology vol 49 no 4 pp 846ndash851 1985

[13] J Anton I Meseguer and F Rodriguez-Valera ldquoProductionof an extracellular polysaccharide by Haloferax mediterraneirdquoApplied Enviromental Microbiology vol 54 no 10 pp 2381ndash2386 1988

[14] H Parolis L A S Parolis I F Boan et al ldquoThe structure ofthe exopolysaccharide produced by the halophilic ArchaeonHaloferax mediterranei strain R4 (ATCC 33500)rdquo CarbohydrateResearch vol 295 pp 147ndash156 1996

[15] E Quesada V Bejar and C Calvo ldquoExopolysaccharide produc-tion by Volcaniella eurihalinardquo Experientia vol 49 no 12 pp1037ndash1041 1993

[16] J A Mata V Bejar I Llamas et al ldquoExopolysaccharides pro-duced by the recently described halophilic bacteria Halomonasventosae and Halomonas anticariensisrdquo Research in Microbiol-ogy vol 157 no 9 pp 827ndash835 2006

[17] I Llamas J A Mata R Tallon et al ldquoCharacterization ofthe exopolysaccharide produced by Salipiger mucosus A3119879 ahalophilic species belonging to the Alphaproteobacteria iso-lated on the Spanish Mediterranean seaboardrdquo Marine Drugsvol 8 no 8 pp 2240ndash2251 2010

[18] F Martınez-Checa E Quesada J Martınez-Canovas I Llamasand V Bejar ldquoPalleronia marisminoris gen nov sp nova moderately halophilic exopolysaccharide-producing bac-terium belonging to the rsquoAlphaproteobacteriarsquo isolated from asaline soilrdquo International Journal of Systematic and EvolutionaryMicrobiology vol 55 no 6 pp 2525ndash2530 2005

[19] A Ventosa M T Garcia M Kamekura H Onishi and FRuiz-Berraquero ldquoBacillus halophilus sp nov a moderatelyhalophilic Bacillus speciesrdquo Systematic and Applied Microbiol-ogy vol 12 no 2 pp 162ndash166 1989

[20] R A Moraine and P Rogovin ldquoKinetics of polysaccharide B1459 fermentationrdquoBiotechnology and Bioengineering vol 8 no4 pp 511ndash524 1966

[21] M Dubois K A Gilles J K Hamilton P A Rebers and FSmith ldquoColorimetric method for determination of sugars andrelated substancesrdquoAnalytical Chemistry vol 28 no 3 pp 350ndash356 1956

[22] O H Lowry N J Rosenbrough A L Farr and R J RandallldquoProtein measurement with the Folin phenol reagentrdquo TheJournal of Biological Chemistry vol 193 no 1 pp 265ndash275 1951

[23] P Gerhardt R G E Murray W A Wood and N R KriegMethods for General and Molecular Bacteriology AmericanSociety for Microbiology Washington DC USA 1994

[24] G M Garrity J A Bell and T G Lilburn ldquoTaxonomic outlineof the procaryotesrdquo inBergeyrsquosManual of Systematic BacteriologD R Boone and R W Castenholz Eds Springer New YorkNY USA 2nd edition 2003

[25] J A Mata J Martınez-Canovas E Quesada and V BejarldquoA detailed phenotypic characterisation of the type strains ofHalomonas speciesrdquo Systematic and Applied Microbiology vol25 no 3 pp 360ndash375 2002

[26] A Oren ldquoMolecular ecology of extremely halophilic archaeaand bacteriardquo FEMSMicrobiology Ecology vol 39 no 1 pp 1ndash72002

[27] H Baati R Amdouni N Gharsallah A Sghir and E AmmarldquoIsolation and characterization of moderately halophilic bacte-ria from tunisian solar salternrdquo Current Microbiology vol 60no 3 pp 157ndash161 2010

[28] A Hedi N Sadfi M Fardeau et al ldquoStudies on the biodiversityof halophilic microorganisms isolated from El-Djerid salt lake(Tunisia) under aerobic conditionsrdquo International Journal ofMicrobiology vol 2009 Article ID 731786 17 pages 2009

[29] M Birbir A Ogan B Calli and B Mertoglu ldquoEnzymecharacteristics of extremely halophilic archaeal community inTuzkoy Salt Mine Turkeyrdquo World Journal of Microbiology andBiotechnology vol 20 no 6 pp 613ndash621 2004

[30] M Birbir and C Sesal ldquoExtremely halophilic bacterial commu-nities in Sereflikochisar Salt Lake in Turkeyrdquo Turkish Journal ofBiology vol 27 no 7 pp 7ndash22 2003

[31] M Birbir B Calli B Mertoglu et al ldquoExtremely halophilicArchaea from Tuz Lake Turkey and the adjacent Kaldirim andKayacik salternsrdquoWorld Journal ofMicrobiology and Biotechnol-ogy vol 23 no 3 pp 309ndash316 2007

[32] R Rohban M A Amoozegar and A Ventosa ldquoScreeningand isolation of halophilic bacteria producing extracellularhydrolyses from Howz Soltan Lake Iranrdquo Journal of IndustrialMicrobiology and Biotechnology vol 36 no 3 pp 333ndash3402009

[33] M J Garabito M C Marquez and A Ventosa ldquoHalotolerantBacillus diversity in hypersaline environmentsrdquo Canadian Jour-nal of Microbiology vol 44 no 2 pp 95ndash102 1998

[34] A Ventosa A Ramos-Cormenzana and M Kocur ldquoModer-ately halophilic gram-positive cocci from hypersaline environ-mentsrdquo Systematic and Applied Microbiology vol 4 no 4 pp564ndash570 1983

[35] E Quesada A Ventosa F Rodriguez-Valera and A R Cor-menzana ldquoTypes and properties of some bacteria isolated fromhypersaline soilsrdquo Journal of Applied Bacteriology vol 53 no 2pp 155ndash161 1982

[36] E Quesada V Bejar M J Valderrama A Ventosa and AR Ramos Cormenzana ldquoIsolation and characterization ofmoderately halophilic nonmotile rods from different salinehabitatsrdquoMicrobiologia vol 1 no 1-2 pp 89ndash96 1985

[37] S R Dave and H B Desai ldquoMicrobial diversity at marinesalterns near Bhavnagar Gujarat Indiardquo Current Science vol60 no 4 pp 497ndash500 2006

[38] S Kumar R Karan S Kapoor S P Singh and S K KhareldquoScreening and isolation of halophilic bacteria producingindustrially important enzymesrdquo Brazilian Journal of Microbi-ology vol 43 no 4 pp 1595ndash1603 2012

[39] B P Dave and A Soni ldquoDiversity of halophilic archaea atsalt pans around Bhavnagar coast Gujaratrdquo Proceedings ofthe National Academy of Sciences India Section B BiologicalSciences vol 83 no 2 pp 225ndash232 2013

[40] A A Joshi P P Kanekar A S Kelkar et al ldquoCultivable bacterialdiversity of alkaline Lonar lake Indiardquo Microbial Ecology vol55 no 2 pp 163ndash172 2008

[41] T M Raghavan and I Furtado ldquoOccurrence of extremelyhalophilic Archaea in sediments from the continental shelf ofwest coast of Indiardquo Current Science vol 86 no 8 pp 1065ndash1067 2004

[42] S Vijayanand J Hemapriya J Selvin and S Kiran ldquoBiodiver-sity of extremely halophilic bacterial strains isolated from solarsalterns of Tuticorin Tamilnadu Indiardquo International Journal ofWater Resources and Environmental Sciences vol 1 no 1 pp 1ndash72012

12 Chinese Journal of Biology

[43] S G Nanjani and H P Soni ldquoIsolation and characterization ofextremely halotolerant and halophilic organisms from Dwarkaand Veravalrdquo Journal of Pharmacy and Biological Sciences vol2 no 2 pp 20ndash25 2012

[44] H Ghozlan H Deif R A Kandil and S Sabry ldquoBiodiversity ofmoderately halophilic bacteria in hypersaline habitats in EgyptrdquoJournal of General and Applied Microbiology vol 52 no 2 pp63ndash72 2006

[45] D R Arahal M T Garcıa W Ludwig K H Schleifer andA Ventosa ldquoTransfer ofHalomonas canadensis andHalomonasisraelensis to the genus Chromohalobacter as Chromohalobac-ter canadensis comb nov and Chromohalobacter israelensiscomb novrdquo International Journal of Systematic and EvolutionaryMicrobiology vol 51 no 4 pp 1443ndash1448 2001

Submit your manuscripts athttpwwwhindawicom

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Anatomy Research International

PeptidesInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporation httpwwwhindawicom

International Journal of

Volume 2014

Zoology

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Molecular Biology International

GenomicsInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

The Scientific World JournalHindawi Publishing Corporation httpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

BioinformaticsAdvances in

Marine BiologyJournal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Signal TransductionJournal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

BioMed Research International

Evolutionary BiologyInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Biochemistry Research International

ArchaeaHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Genetics Research International

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Advances in

Virolog y

Hindawi Publishing Corporationhttpwwwhindawicom

Nucleic AcidsJournal of

Volume 2014

Stem CellsInternational

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

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Enzyme Research

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

International Journal of

Microbiology

Page 7: Research Article Production of Extracellular …downloads.hindawi.com/archive/2014/205731.pdfResearch Article Production of Extracellular Polymeric Substances by Halophilic Bacteria

Chinese Journal of Biology 7

Table8Biochemicalcharacteris

ticso

fthe

selected

EPSprod

ucinghaloph

ilicb

acteria

isolated

from

solarsalternso

fInd

ia

Character

Bacterialisolates

JS 504

JS 802

JS 803

JS 904

JW 302

JW 307

KS 1101

KS 1805

KW 203

VW 301

VW 402

SUR

108

SUR

301

SUR

302

SUR

303

SUR

307

SUR

309

SUR

310

SUR

202

SUR

201

Biochemicalcharacters

Voges-Proskauertest

minusminus

minusminus

minusminus

minusminus

+minus

minusminus

minusminus

minusminus

minusminus

minusminus

Citrateu

tilization

+minus

minusminus

minus+

minus+

minus+

++

+minus

minus+

++

++

Methylred

test

minusminus

minusminus

minusminus

minus+

+minus

minusminus

minusminus

minusminus

minusminus

minusminus

Prod

uctio

nof

Glutaminase

++

++

++

++

++

++

++

minus+

++

++

Asparaginase

++

++

++

++

++

++

++

minus+

++

++

Amylase

+minus

minusminus

minusminus

minusminus

+minus

minusminus

minusminus

minusminus

minusminus

minusminus

Cellulase

minusminus

minusminus

minusminus

minusminus

+minus

minusminus

minusminus

minusminus

minusminus

minusminus

Pectinase

+minus

minusminus

minusminus

minusminus

+minus

minusminus

minusminus

minusminus

minusminus

minusminus

Inulinase

+minus

minusminus

minusminus

minusminus

+minus

minusminus

minusminus

minusminus

minusminus

minusminus

Gelatinase

minusminus

minusminus

minusminus

minusminus

+minus

minusminus

minusminus

minusminus

minusminus

minusminus

Lipase

+minus

minusminus

minusminus

minus+

+minus

minusminus

+minus

minusminus

minusminus

minusminus

Xylanase

+minus

minusminus

minusminus

minusminus

+minus

minusminus

minusminus

minusminus

minusminus

minusminus

Caseinase

minusminus

minus+

minusminus

minusminus

minusminus

minusminus

minus+

minusminus

minusminus

minusminus

Urease

minusminus

minus+

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

Catalase

++

++

++

++

++

++

++

++

++

++

Nitrater

eductase

++

++

++

++

++

++

++

++

++

++

H2S

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

Lysin

edecarbo

xylase

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

Arginined

ecarbo

xylase

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

Ornith

ined

ecarbo

xylase

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

minusminus

Indo

leminus

minusminus

minusminus

minusminus

+minus

minusminus

minus+

minus+

minusminus

minusminus

minus

Phenylalanined

eaminase

++

+minus

+minus

++

minus+

minus+

++

++

++

minus+

Tryptoph

andeam

inase

++

++

++

++

++

minusminus

++

++

++

+minus

8 Chinese Journal of Biology

Table9Ca

rbon

source

utilizatio

nandferm

entatio

nof

thes

elected

EPSprod

ucinghaloph

ilicb

acteria

isolated

from

samples

ofsolarsalternso

fInd

ia

Carbon

source

Bacterialisolates

JS 504

JS 802

JS 803

JS 904

JW 302

JW 307

KS 1101

KS 1805

KW 203

VW

301

VW

402

SUR

108

SUR

301

SUR

302

SUR

303

SUR

307

SUR

309

SUR

310

SUR

202

SUR

201

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

Glucose

++

++

+minusminus

++minus

+minus

+minus

++

++

+minus

+minus

+minus

+minusminus

++

++minus

+minus

+minusminusminusminusminus

Galactose

+minus

+minus

+minusminus

++minus

+minus

+minus

++

+minus

+minus

+minus

+minus

++minusminus

+minus

++

+minus

+minus

+minus

+minus

Arabino

seminusminusminusminus

+minusminusminusminusminus

+minus

+minus

++

++

+minus

+minus

+minus

+minusminus

++minus

+minus

+minusminusminus

+minus

+minus

Manno

se+minus

+minus

+minusminus

++minus

+minus

+minusminus

++

++minus

+minus

+minus

+minusminusminusminusminus

+minus

+minus

+minus

+minus

+minus

Rham

nose

+minus

+minus

+minus

+minusminusminusminusminusminusminus

++

+minusminusminus

+minus

+minus

++minus

++minus

+minusminusminusminusminusminusminus

+minus

Trehalose

+minus

+minus

+minusminus

+minus

+minusminusminusminus

+minus

++minusminusminusminus

+minus

+minusminus

++

++minusminusminusminusminusminusminus

+minus

Lactose

minusminusminusminus

+minusminusminus

+minus

+minus

+minus

+minus

+minus

+minus

+minusminusminus

+minus

++minusminus

+minusminusminusminusminus

+minus

+minus

Sucrose

+minus

+minus

+minus

++

++

+minus

+minus

+minus

++

+minus

+minusminusminus

+minus

++

+minus

+minus

+minus

+minusminusminus

+minus

Xylose

minusminusminusminus

+minus

+minusminusminus

+minus

++minus

+minusminus

+minus

+minus

+minus

+minusminus

+minus

++

+minusminusminusminus

+minusminusminus

Ribo

se+minus

++

++

++

+minus

+minus

+minus

++

++

+minus

+minus

+minus

++

++

++

+minus

+minus

+minus

+minus

+minus

Fructose

+minus

++

+minus

++

+minus

++

++

+minus

++

++

++

++

++

++

++

++

++

++

++

++

Maltose

minusminusminusminus

+minusminus

+minus

++minus

+minus

+minus

++

+minus

+minus

+minus

++minus

+minus

++minusminusminusminusminus

+minus

+minus

Glycine

minusminusminus

++minus

+minusminus

+minusminusminusminus

+minusminusminusminusminusminusminusminusminus

+minusminusminusminusminus

+minus

+minusminusminusminus

+minusminus

Glycerol

+minus

+minus

+minusminus

++

++minus

+minus

+minus

++

+minus

+minus

+minus

+minusminus

++

++minus

+minus

+minus

+minus

+minus

Adon

itol

+minus

+minus

+minus

+minusminusminus

+minus

+minus

+minus

+minus

+minus

+minusminusminus

+minus

++

+minus

+minus

+minus

+minusminusminusminusminus

Salicin

+minus

++

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

++

++

+minus

+minus

+minus

+minus

+minus

Meso-inosito

l+minusminusminus

+minus

+minus

+minus

+minus

++

+minus

++

+minus

+minusminusminus

+minusminus

++minus

+minus

+minus

+minus

+minus

+minus

Mannitol

+minus

+minus

+minus

++

+minus

+minus

+minus

+minus

++

+minus

+minus

+minus

+minus

+minus

++

+minus

+minus

+minus

+minus

+minus

Sorbito

l+

++

++minusminus

++

+minus

++

++

++

++

++

++

++

+minus

+minus

++

++minus

+minus

++

+minus

Raffino

seminusminus

++

++minusminus

+minus

+minus

+minus

+minus

++minusminusminusminusminusminusminusminus

++

++

+minus

+minus

+minus

+minus

+minus

Cellobiose

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

++

+minus

+minus

+minus

+minus

++

+minus

+minus

+minus

+minus

+minus

+minus

Melibioseminusminusminusminusminusminusminusminusminusminusminusminusminusminusminusminus

++minusminusminusminusminusminusminusminus

++minusminusminusminusminusminusminusminusminusminusminusminus

Na-acetate

+minus

+minus

+minus

+minus

++

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

++

++

+minus

+minus

+minus

+minus

+minus

Na-citrate

+minus

++

+minusminusminusminus

+minusminusminusminusminusminusminusminusminusminusminusminusminusminusminusminusminus

+minusminusminusminusminusminusminusminusminusminusminusminus

Na-fumarate

+minus

+minus

+minus

++

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minusminus

++minus

+minus

+minus

+minus

+minus

+minus

Na-form

ate

+minus

+minus

+minus

+minus

+minusminusminus

+minus

+minus

+minusminusminus

+minusminusminus

+minus

++minusminus

+minusminusminusminusminusminusminusminusminus

Na-benzoate

+minus

++

++

+minus

+minus

+minus

++

++

++

+minus

+minus

++

++

+minus

+minus

+minus

++

++

++

++

Na-malateminusminusminusminusminusminus

++

+minusminusminusminusminusminusminusminusminusminusminusminusminusminusminusminusminus

++

++minusminusminusminusminusminusminusminusminusminus

Na-prop

ionate

+minus

+minus

+minusminusminus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

++minus

++minus

+minus

+minus

+minus

+minus

Na-succinate

+minus

+minus

+minus

++

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

++

++

+minus

+minus

+minus

+minus

+minus

Uutilization

Fferm

entatio

n

Chinese Journal of Biology 9

Table10A

ntibiotic

sensitivitypatte

rnof

thes

electedEP

Sprod

ucinghaloph

ilicb

acteria

isolated

from

solarsalternso

fInd

ia

Antibiotics

Bacterialisolates

JS 504

JS 802

JS 803

JS 904

JW 302

JW 307

KS 1101

KS 1805

KW 203

VW 301

VW 402

SUR

108

SUR

301

SUR

302

SUR

303

SUR

307

SUR

309

SUR

310

SUR

202

SUR

201

Ampicillin(10120583

g)S

SR

SS

SS

RI

SS

SR

SS

SS

SS

SAmoxicillin

(30120583

g)S

SR

RS

SS

RS

SS

SR

RS

IR

SS

SBa

citracin

(10120583

g)S

SR

SS

SR

RS

IR

RR

RS

IR

RR

RCiprofl

oxacin

(30120583

g)I

II

RI

II

IR

II

II

IR

SR

II

ICh

lorte

tracyclin

e(30120583g)

SI

SS

IS

RR

SS

SS

RR

SS

II

IS

Chloramph

enicol(10120583

g)S

SS

SS

SS

SR

SS

SS

SS

SR

SS

SErythrom

ycin

(15120583

g)S

SI

SS

IS

II

II

II

SR

RI

IS

SFu

sidicacid

(30120583

g)I

RR

SS

IR

RS

RI

RR

RS

IR

II

RGentamycin

(10120583

g)S

SS

SS

SS

SR

SS

SS

SS

IS

SS

SKa

namycin

(30120583

g)I

IS

RS

II

SR

II

RI

RR

RI

RI

RMethicillin(5120583g)

SS

RS

IR

RR

RR

RR

RR

SR

RR

RR

Neomycin

(30120583

g)S

SI

RI

IR

II

IR

II

RR

RI

IR

RNorflo

xacin(30120583

g)S

SR

SS

SS

SI

SS

SS

SS

SS

SS

SNetillin

(10120583

g)R

SS

IR

SS

SR

RR

SS

RR

RR

RR

RNalidixicacid

(30120583

g)R

RS

RR

II

RR

II

RR

RR

SR

RR

RNovob

iosin

(30120583

g)I

RR

SR

SS

RS

SS

IR

SS

SI

SS

SPo

lymixin

B(50120583

g)R

RI

RI

IR

RR

RI

IR

RR

II

II

SPenicillinG(10120583

g)S

SR

IS

IR

RR

II

SR

RS

II

II

SStreptom

ycin

(10120583

g)S

SS

RS

RR

SR

RR

IS

RS

SR

II

RTetracyclin

e(30120583g)

RR

RR

RI

RR

SR

IR

RR

RS

RR

RI

Trim

etho

prim

e(30120583g)

SS

SS

IS

SI

SS

SS

IR

RS

SS

SS

Vancom

ycin

(30120583

g)R

RR

SR

RR

RS

RR

RR

RS

RR

RR

R

S=SensitiveR

=Re

sistantI

=Interm

ediate

10 Chinese Journal of Biology

0

1

2

3

4

Bacterial isolate

Gro

wth

OD

at540

nmto

tal E

PS (g

L)

OD at 540nmTotal EPS

JS504

JS802

JS803

JS904

JW302

JW307

KS1101

KS1805

KW203

VW301

VW402

SUR1

08

SUR3

01

SUR3

02

SUR3

03

SUR3

07

SUR3

09

SUR3

10

SUR2

02

SUR2

01

Figure 1 Screening of moderately halophilic bacteria isolatedfrom soil and water samples of solar salterns for production ofextracellular polymeric substances

0

02

04

06

08

1

Ant

ibio

tic re

sista

nce i

ndex

Bacterial isolate

JS504

JS802

JS803

JS904

JW302

JW307

KS1101

KS1805

KW203

SUR1

08

SUR2

01

SUR2

02

SUR3

01

SUR3

02

SUR3

03

SUR3

07

SUR3

09

SUR3

10

VW301

VW402

Figure 2 Antibiotic resistance index of the selected EPS producinghalophilic bacteria isolated from soil and water samples of solarsalterns of India

The isolates SUR301 and KS 1805 were tentatively iden-tified as Chromohalobacter based on the study by Arahal etal [45] and bear striking similarity with those of Chromo-halobacter sp isolated fromhypersaline soil sample of TriveniSangam Gujarat [38] The physiological morphological andbiochemical characteristics of isolate SUR303 were consistentwith the features of the genus Salinicoccus [10] while isolatesJS904 and JS504 were tentatively assigned to Aidingimonasand Alteromonas respectively

Finally sensitivity of these isolates to antibiotics (Table 10and Figure 2) also corroborates the findings ofMata et al [25]and Hedi et al [28]

5 Conclusion

It may be emphasized that study of these moderatelyhalophilic bacteria from coastal hypersaline solar salterns ofIndia with special attempt on screening for EPS productionhas led to the discovery of wide variety of halophilic speciesAlthough attention has been focused on the production

of EPS by these halophilic bacterial isolates their physio-biochemical features indicate that they may equally holdpotential towards production of various biomolecules ofindustrial interest

Conflict of Interests

It is declared by the authors that there is no conflict of interestsregarding the publication of this paper

Acknowledgment

This study was financially supported by grants from Uni-versity Grants Commission India (Sanction no F14-2(SC)2008 (SA-III) 31 March 2009) under the Scheme ofRajiv Gandhi National Fellowship

References

[1] E A Galinski and B J Tindall ldquoBiotechnological prospectsfor halophiles and halotolerant microorganismsrdquo in MolecularBiology and Biotechnology of Extremophiles R D Herbert andR J Sharp Eds Blackie London UK 1992

[2] R Margesin and F Schinner ldquoPotential of halotolerant andhalophilic microorganisms for biotechnologyrdquo Extremophilesvol 5 no 2 pp 73ndash83 2001

[3] F Rodriguez Valera F Ruiz Berraquero and A Ramos Cor-menzana ldquoCharacteristics of the heterotrophic bacterial pop-ulations in hypersaline environments of different salt concen-trationsrdquoMicrobial Ecology vol 7 no 3 pp 235ndash243 1981

[4] F Rodriguez-Valera ldquoCharacteristics and microbial ecol-ogy of hypersaline environmentsrdquo in Halophilic Bacteria FRodriguez-Valera Ed vol 1 pp 3ndash30 CRC Press Boca RatonFla USA 1988

[5] D J Kushner and M Kamekura ldquoPhysiology of halophiliceubacteriardquo inHalophilic Bacteria F Rodriguez-Valera Ed vol1 pp 109ndash138 CRC press Boca Raton Fla USA 1988

[6] M A Amoozegar F Malekzadeh and K AMalik ldquoProductionof amylase by newly isolated moderate halophile Halobacillussp strainMA-2rdquo Journal of Microbiological Methods vol 52 no3 pp 353ndash359 2003

[7] C O Jeon J Lim J Lee et al ldquoLentibacillus salarius spnov isolated from saline sediment in China and emendeddescription of the genus Lentibacillusrdquo International Journal ofSystematic and Evolutionary Microbiology vol 55 no 3 pp1339ndash1343 2005

[8] S Bouchotroch E Quesada A Del Moral I Llamas andV Bejar ldquoHalomonas maura sp nov a novel moderatelyhalophilic exopolysaccharide-producing bacteriumrdquo Interna-tional Journal of Systematic and Evolutionary Microbiology vol51 no 5 pp 1625ndash1632 2001

[9] A Biswas A Patra and A Paul ldquoProduction of poly-3-hydroxyalkanoic acids by a moderately halophilic bacteriumHalomonas marina HMA 103 isolated from solar saltern ofOrissa Indiardquo Acta Microbiologica et Immunologica Hungaricavol 56 no 2 pp 125ndash143 2009

[10] S Y Jayachandra S Anil Kumar D P Merley and MB Sulochana ldquoIsolation and characterization of extremehalophilic bacterium Salinicoccus sp JAS4 producing extra-cellular hydrolytic enzymesrdquo Recent Research in Science andTechnology vol 4 no 4 pp 46ndash49 2012

Chinese Journal of Biology 11

[11] I W Sutherland ldquoNovel and established applications of micro-bial polysaccharidesrdquo Trends in Biotechnology vol 16 no 1 pp41ndash46 1998

[12] MWMittelman andG G Geesey ldquoCopper-binding character-istics of exopolymers from a freshwater-sediment bacteriumrdquoApplied and EnvironmentalMicrobiology vol 49 no 4 pp 846ndash851 1985

[13] J Anton I Meseguer and F Rodriguez-Valera ldquoProductionof an extracellular polysaccharide by Haloferax mediterraneirdquoApplied Enviromental Microbiology vol 54 no 10 pp 2381ndash2386 1988

[14] H Parolis L A S Parolis I F Boan et al ldquoThe structure ofthe exopolysaccharide produced by the halophilic ArchaeonHaloferax mediterranei strain R4 (ATCC 33500)rdquo CarbohydrateResearch vol 295 pp 147ndash156 1996

[15] E Quesada V Bejar and C Calvo ldquoExopolysaccharide produc-tion by Volcaniella eurihalinardquo Experientia vol 49 no 12 pp1037ndash1041 1993

[16] J A Mata V Bejar I Llamas et al ldquoExopolysaccharides pro-duced by the recently described halophilic bacteria Halomonasventosae and Halomonas anticariensisrdquo Research in Microbiol-ogy vol 157 no 9 pp 827ndash835 2006

[17] I Llamas J A Mata R Tallon et al ldquoCharacterization ofthe exopolysaccharide produced by Salipiger mucosus A3119879 ahalophilic species belonging to the Alphaproteobacteria iso-lated on the Spanish Mediterranean seaboardrdquo Marine Drugsvol 8 no 8 pp 2240ndash2251 2010

[18] F Martınez-Checa E Quesada J Martınez-Canovas I Llamasand V Bejar ldquoPalleronia marisminoris gen nov sp nova moderately halophilic exopolysaccharide-producing bac-terium belonging to the rsquoAlphaproteobacteriarsquo isolated from asaline soilrdquo International Journal of Systematic and EvolutionaryMicrobiology vol 55 no 6 pp 2525ndash2530 2005

[19] A Ventosa M T Garcia M Kamekura H Onishi and FRuiz-Berraquero ldquoBacillus halophilus sp nov a moderatelyhalophilic Bacillus speciesrdquo Systematic and Applied Microbiol-ogy vol 12 no 2 pp 162ndash166 1989

[20] R A Moraine and P Rogovin ldquoKinetics of polysaccharide B1459 fermentationrdquoBiotechnology and Bioengineering vol 8 no4 pp 511ndash524 1966

[21] M Dubois K A Gilles J K Hamilton P A Rebers and FSmith ldquoColorimetric method for determination of sugars andrelated substancesrdquoAnalytical Chemistry vol 28 no 3 pp 350ndash356 1956

[22] O H Lowry N J Rosenbrough A L Farr and R J RandallldquoProtein measurement with the Folin phenol reagentrdquo TheJournal of Biological Chemistry vol 193 no 1 pp 265ndash275 1951

[23] P Gerhardt R G E Murray W A Wood and N R KriegMethods for General and Molecular Bacteriology AmericanSociety for Microbiology Washington DC USA 1994

[24] G M Garrity J A Bell and T G Lilburn ldquoTaxonomic outlineof the procaryotesrdquo inBergeyrsquosManual of Systematic BacteriologD R Boone and R W Castenholz Eds Springer New YorkNY USA 2nd edition 2003

[25] J A Mata J Martınez-Canovas E Quesada and V BejarldquoA detailed phenotypic characterisation of the type strains ofHalomonas speciesrdquo Systematic and Applied Microbiology vol25 no 3 pp 360ndash375 2002

[26] A Oren ldquoMolecular ecology of extremely halophilic archaeaand bacteriardquo FEMSMicrobiology Ecology vol 39 no 1 pp 1ndash72002

[27] H Baati R Amdouni N Gharsallah A Sghir and E AmmarldquoIsolation and characterization of moderately halophilic bacte-ria from tunisian solar salternrdquo Current Microbiology vol 60no 3 pp 157ndash161 2010

[28] A Hedi N Sadfi M Fardeau et al ldquoStudies on the biodiversityof halophilic microorganisms isolated from El-Djerid salt lake(Tunisia) under aerobic conditionsrdquo International Journal ofMicrobiology vol 2009 Article ID 731786 17 pages 2009

[29] M Birbir A Ogan B Calli and B Mertoglu ldquoEnzymecharacteristics of extremely halophilic archaeal community inTuzkoy Salt Mine Turkeyrdquo World Journal of Microbiology andBiotechnology vol 20 no 6 pp 613ndash621 2004

[30] M Birbir and C Sesal ldquoExtremely halophilic bacterial commu-nities in Sereflikochisar Salt Lake in Turkeyrdquo Turkish Journal ofBiology vol 27 no 7 pp 7ndash22 2003

[31] M Birbir B Calli B Mertoglu et al ldquoExtremely halophilicArchaea from Tuz Lake Turkey and the adjacent Kaldirim andKayacik salternsrdquoWorld Journal ofMicrobiology and Biotechnol-ogy vol 23 no 3 pp 309ndash316 2007

[32] R Rohban M A Amoozegar and A Ventosa ldquoScreeningand isolation of halophilic bacteria producing extracellularhydrolyses from Howz Soltan Lake Iranrdquo Journal of IndustrialMicrobiology and Biotechnology vol 36 no 3 pp 333ndash3402009

[33] M J Garabito M C Marquez and A Ventosa ldquoHalotolerantBacillus diversity in hypersaline environmentsrdquo Canadian Jour-nal of Microbiology vol 44 no 2 pp 95ndash102 1998

[34] A Ventosa A Ramos-Cormenzana and M Kocur ldquoModer-ately halophilic gram-positive cocci from hypersaline environ-mentsrdquo Systematic and Applied Microbiology vol 4 no 4 pp564ndash570 1983

[35] E Quesada A Ventosa F Rodriguez-Valera and A R Cor-menzana ldquoTypes and properties of some bacteria isolated fromhypersaline soilsrdquo Journal of Applied Bacteriology vol 53 no 2pp 155ndash161 1982

[36] E Quesada V Bejar M J Valderrama A Ventosa and AR Ramos Cormenzana ldquoIsolation and characterization ofmoderately halophilic nonmotile rods from different salinehabitatsrdquoMicrobiologia vol 1 no 1-2 pp 89ndash96 1985

[37] S R Dave and H B Desai ldquoMicrobial diversity at marinesalterns near Bhavnagar Gujarat Indiardquo Current Science vol60 no 4 pp 497ndash500 2006

[38] S Kumar R Karan S Kapoor S P Singh and S K KhareldquoScreening and isolation of halophilic bacteria producingindustrially important enzymesrdquo Brazilian Journal of Microbi-ology vol 43 no 4 pp 1595ndash1603 2012

[39] B P Dave and A Soni ldquoDiversity of halophilic archaea atsalt pans around Bhavnagar coast Gujaratrdquo Proceedings ofthe National Academy of Sciences India Section B BiologicalSciences vol 83 no 2 pp 225ndash232 2013

[40] A A Joshi P P Kanekar A S Kelkar et al ldquoCultivable bacterialdiversity of alkaline Lonar lake Indiardquo Microbial Ecology vol55 no 2 pp 163ndash172 2008

[41] T M Raghavan and I Furtado ldquoOccurrence of extremelyhalophilic Archaea in sediments from the continental shelf ofwest coast of Indiardquo Current Science vol 86 no 8 pp 1065ndash1067 2004

[42] S Vijayanand J Hemapriya J Selvin and S Kiran ldquoBiodiver-sity of extremely halophilic bacterial strains isolated from solarsalterns of Tuticorin Tamilnadu Indiardquo International Journal ofWater Resources and Environmental Sciences vol 1 no 1 pp 1ndash72012

12 Chinese Journal of Biology

[43] S G Nanjani and H P Soni ldquoIsolation and characterization ofextremely halotolerant and halophilic organisms from Dwarkaand Veravalrdquo Journal of Pharmacy and Biological Sciences vol2 no 2 pp 20ndash25 2012

[44] H Ghozlan H Deif R A Kandil and S Sabry ldquoBiodiversity ofmoderately halophilic bacteria in hypersaline habitats in EgyptrdquoJournal of General and Applied Microbiology vol 52 no 2 pp63ndash72 2006

[45] D R Arahal M T Garcıa W Ludwig K H Schleifer andA Ventosa ldquoTransfer ofHalomonas canadensis andHalomonasisraelensis to the genus Chromohalobacter as Chromohalobac-ter canadensis comb nov and Chromohalobacter israelensiscomb novrdquo International Journal of Systematic and EvolutionaryMicrobiology vol 51 no 4 pp 1443ndash1448 2001

Submit your manuscripts athttpwwwhindawicom

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Anatomy Research International

PeptidesInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporation httpwwwhindawicom

International Journal of

Volume 2014

Zoology

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Molecular Biology International

GenomicsInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

The Scientific World JournalHindawi Publishing Corporation httpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

BioinformaticsAdvances in

Marine BiologyJournal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Signal TransductionJournal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

BioMed Research International

Evolutionary BiologyInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Biochemistry Research International

ArchaeaHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Genetics Research International

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Advances in

Virolog y

Hindawi Publishing Corporationhttpwwwhindawicom

Nucleic AcidsJournal of

Volume 2014

Stem CellsInternational

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

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Enzyme Research

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

International Journal of

Microbiology

Page 8: Research Article Production of Extracellular …downloads.hindawi.com/archive/2014/205731.pdfResearch Article Production of Extracellular Polymeric Substances by Halophilic Bacteria

8 Chinese Journal of Biology

Table9Ca

rbon

source

utilizatio

nandferm

entatio

nof

thes

elected

EPSprod

ucinghaloph

ilicb

acteria

isolated

from

samples

ofsolarsalternso

fInd

ia

Carbon

source

Bacterialisolates

JS 504

JS 802

JS 803

JS 904

JW 302

JW 307

KS 1101

KS 1805

KW 203

VW

301

VW

402

SUR

108

SUR

301

SUR

302

SUR

303

SUR

307

SUR

309

SUR

310

SUR

202

SUR

201

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

UF

Glucose

++

++

+minusminus

++minus

+minus

+minus

++

++

+minus

+minus

+minus

+minusminus

++

++minus

+minus

+minusminusminusminusminus

Galactose

+minus

+minus

+minusminus

++minus

+minus

+minus

++

+minus

+minus

+minus

+minus

++minusminus

+minus

++

+minus

+minus

+minus

+minus

Arabino

seminusminusminusminus

+minusminusminusminusminus

+minus

+minus

++

++

+minus

+minus

+minus

+minusminus

++minus

+minus

+minusminusminus

+minus

+minus

Manno

se+minus

+minus

+minusminus

++minus

+minus

+minusminus

++

++minus

+minus

+minus

+minusminusminusminusminus

+minus

+minus

+minus

+minus

+minus

Rham

nose

+minus

+minus

+minus

+minusminusminusminusminusminusminus

++

+minusminusminus

+minus

+minus

++minus

++minus

+minusminusminusminusminusminusminus

+minus

Trehalose

+minus

+minus

+minusminus

+minus

+minusminusminusminus

+minus

++minusminusminusminus

+minus

+minusminus

++

++minusminusminusminusminusminusminus

+minus

Lactose

minusminusminusminus

+minusminusminus

+minus

+minus

+minus

+minus

+minus

+minus

+minusminusminus

+minus

++minusminus

+minusminusminusminusminus

+minus

+minus

Sucrose

+minus

+minus

+minus

++

++

+minus

+minus

+minus

++

+minus

+minusminusminus

+minus

++

+minus

+minus

+minus

+minusminusminus

+minus

Xylose

minusminusminusminus

+minus

+minusminusminus

+minus

++minus

+minusminus

+minus

+minus

+minus

+minusminus

+minus

++

+minusminusminusminus

+minusminusminus

Ribo

se+minus

++

++

++

+minus

+minus

+minus

++

++

+minus

+minus

+minus

++

++

++

+minus

+minus

+minus

+minus

+minus

Fructose

+minus

++

+minus

++

+minus

++

++

+minus

++

++

++

++

++

++

++

++

++

++

++

++

Maltose

minusminusminusminus

+minusminus

+minus

++minus

+minus

+minus

++

+minus

+minus

+minus

++minus

+minus

++minusminusminusminusminus

+minus

+minus

Glycine

minusminusminus

++minus

+minusminus

+minusminusminusminus

+minusminusminusminusminusminusminusminusminus

+minusminusminusminusminus

+minus

+minusminusminusminus

+minusminus

Glycerol

+minus

+minus

+minusminus

++

++minus

+minus

+minus

++

+minus

+minus

+minus

+minusminus

++

++minus

+minus

+minus

+minus

+minus

Adon

itol

+minus

+minus

+minus

+minusminusminus

+minus

+minus

+minus

+minus

+minus

+minusminusminus

+minus

++

+minus

+minus

+minus

+minusminusminusminusminus

Salicin

+minus

++

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

++

++

+minus

+minus

+minus

+minus

+minus

Meso-inosito

l+minusminusminus

+minus

+minus

+minus

+minus

++

+minus

++

+minus

+minusminusminus

+minusminus

++minus

+minus

+minus

+minus

+minus

+minus

Mannitol

+minus

+minus

+minus

++

+minus

+minus

+minus

+minus

++

+minus

+minus

+minus

+minus

+minus

++

+minus

+minus

+minus

+minus

+minus

Sorbito

l+

++

++minusminus

++

+minus

++

++

++

++

++

++

++

+minus

+minus

++

++minus

+minus

++

+minus

Raffino

seminusminus

++

++minusminus

+minus

+minus

+minus

+minus

++minusminusminusminusminusminusminusminus

++

++

+minus

+minus

+minus

+minus

+minus

Cellobiose

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

++

+minus

+minus

+minus

+minus

++

+minus

+minus

+minus

+minus

+minus

+minus

Melibioseminusminusminusminusminusminusminusminusminusminusminusminusminusminusminusminus

++minusminusminusminusminusminusminusminus

++minusminusminusminusminusminusminusminusminusminusminusminus

Na-acetate

+minus

+minus

+minus

+minus

++

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

++

++

+minus

+minus

+minus

+minus

+minus

Na-citrate

+minus

++

+minusminusminusminus

+minusminusminusminusminusminusminusminusminusminusminusminusminusminusminusminusminus

+minusminusminusminusminusminusminusminusminusminusminusminus

Na-fumarate

+minus

+minus

+minus

++

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minusminus

++minus

+minus

+minus

+minus

+minus

+minus

Na-form

ate

+minus

+minus

+minus

+minus

+minusminusminus

+minus

+minus

+minusminusminus

+minusminusminus

+minus

++minusminus

+minusminusminusminusminusminusminusminusminus

Na-benzoate

+minus

++

++

+minus

+minus

+minus

++

++

++

+minus

+minus

++

++

+minus

+minus

+minus

++

++

++

++

Na-malateminusminusminusminusminusminus

++

+minusminusminusminusminusminusminusminusminusminusminusminusminusminusminusminusminus

++

++minusminusminusminusminusminusminusminusminusminus

Na-prop

ionate

+minus

+minus

+minusminusminus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

++minus

++minus

+minus

+minus

+minus

+minus

Na-succinate

+minus

+minus

+minus

++

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

+minus

++

++

+minus

+minus

+minus

+minus

+minus

Uutilization

Fferm

entatio

n

Chinese Journal of Biology 9

Table10A

ntibiotic

sensitivitypatte

rnof

thes

electedEP

Sprod

ucinghaloph

ilicb

acteria

isolated

from

solarsalternso

fInd

ia

Antibiotics

Bacterialisolates

JS 504

JS 802

JS 803

JS 904

JW 302

JW 307

KS 1101

KS 1805

KW 203

VW 301

VW 402

SUR

108

SUR

301

SUR

302

SUR

303

SUR

307

SUR

309

SUR

310

SUR

202

SUR

201

Ampicillin(10120583

g)S

SR

SS

SS

RI

SS

SR

SS

SS

SS

SAmoxicillin

(30120583

g)S

SR

RS

SS

RS

SS

SR

RS

IR

SS

SBa

citracin

(10120583

g)S

SR

SS

SR

RS

IR

RR

RS

IR

RR

RCiprofl

oxacin

(30120583

g)I

II

RI

II

IR

II

II

IR

SR

II

ICh

lorte

tracyclin

e(30120583g)

SI

SS

IS

RR

SS

SS

RR

SS

II

IS

Chloramph

enicol(10120583

g)S

SS

SS

SS

SR

SS

SS

SS

SR

SS

SErythrom

ycin

(15120583

g)S

SI

SS

IS

II

II

II

SR

RI

IS

SFu

sidicacid

(30120583

g)I

RR

SS

IR

RS

RI

RR

RS

IR

II

RGentamycin

(10120583

g)S

SS

SS

SS

SR

SS

SS

SS

IS

SS

SKa

namycin

(30120583

g)I

IS

RS

II

SR

II

RI

RR

RI

RI

RMethicillin(5120583g)

SS

RS

IR

RR

RR

RR

RR

SR

RR

RR

Neomycin

(30120583

g)S

SI

RI

IR

II

IR

II

RR

RI

IR

RNorflo

xacin(30120583

g)S

SR

SS

SS

SI

SS

SS

SS

SS

SS

SNetillin

(10120583

g)R

SS

IR

SS

SR

RR

SS

RR

RR

RR

RNalidixicacid

(30120583

g)R

RS

RR

II

RR

II

RR

RR

SR

RR

RNovob

iosin

(30120583

g)I

RR

SR

SS

RS

SS

IR

SS

SI

SS

SPo

lymixin

B(50120583

g)R

RI

RI

IR

RR

RI

IR

RR

II

II

SPenicillinG(10120583

g)S

SR

IS

IR

RR

II

SR

RS

II

II

SStreptom

ycin

(10120583

g)S

SS

RS

RR

SR

RR

IS

RS

SR

II

RTetracyclin

e(30120583g)

RR

RR

RI

RR

SR

IR

RR

RS

RR

RI

Trim

etho

prim

e(30120583g)

SS

SS

IS

SI

SS

SS

IR

RS

SS

SS

Vancom

ycin

(30120583

g)R

RR

SR

RR

RS

RR

RR

RS

RR

RR

R

S=SensitiveR

=Re

sistantI

=Interm

ediate

10 Chinese Journal of Biology

0

1

2

3

4

Bacterial isolate

Gro

wth

OD

at540

nmto

tal E

PS (g

L)

OD at 540nmTotal EPS

JS504

JS802

JS803

JS904

JW302

JW307

KS1101

KS1805

KW203

VW301

VW402

SUR1

08

SUR3

01

SUR3

02

SUR3

03

SUR3

07

SUR3

09

SUR3

10

SUR2

02

SUR2

01

Figure 1 Screening of moderately halophilic bacteria isolatedfrom soil and water samples of solar salterns for production ofextracellular polymeric substances

0

02

04

06

08

1

Ant

ibio

tic re

sista

nce i

ndex

Bacterial isolate

JS504

JS802

JS803

JS904

JW302

JW307

KS1101

KS1805

KW203

SUR1

08

SUR2

01

SUR2

02

SUR3

01

SUR3

02

SUR3

03

SUR3

07

SUR3

09

SUR3

10

VW301

VW402

Figure 2 Antibiotic resistance index of the selected EPS producinghalophilic bacteria isolated from soil and water samples of solarsalterns of India

The isolates SUR301 and KS 1805 were tentatively iden-tified as Chromohalobacter based on the study by Arahal etal [45] and bear striking similarity with those of Chromo-halobacter sp isolated fromhypersaline soil sample of TriveniSangam Gujarat [38] The physiological morphological andbiochemical characteristics of isolate SUR303 were consistentwith the features of the genus Salinicoccus [10] while isolatesJS904 and JS504 were tentatively assigned to Aidingimonasand Alteromonas respectively

Finally sensitivity of these isolates to antibiotics (Table 10and Figure 2) also corroborates the findings ofMata et al [25]and Hedi et al [28]

5 Conclusion

It may be emphasized that study of these moderatelyhalophilic bacteria from coastal hypersaline solar salterns ofIndia with special attempt on screening for EPS productionhas led to the discovery of wide variety of halophilic speciesAlthough attention has been focused on the production

of EPS by these halophilic bacterial isolates their physio-biochemical features indicate that they may equally holdpotential towards production of various biomolecules ofindustrial interest

Conflict of Interests

It is declared by the authors that there is no conflict of interestsregarding the publication of this paper

Acknowledgment

This study was financially supported by grants from Uni-versity Grants Commission India (Sanction no F14-2(SC)2008 (SA-III) 31 March 2009) under the Scheme ofRajiv Gandhi National Fellowship

References

[1] E A Galinski and B J Tindall ldquoBiotechnological prospectsfor halophiles and halotolerant microorganismsrdquo in MolecularBiology and Biotechnology of Extremophiles R D Herbert andR J Sharp Eds Blackie London UK 1992

[2] R Margesin and F Schinner ldquoPotential of halotolerant andhalophilic microorganisms for biotechnologyrdquo Extremophilesvol 5 no 2 pp 73ndash83 2001

[3] F Rodriguez Valera F Ruiz Berraquero and A Ramos Cor-menzana ldquoCharacteristics of the heterotrophic bacterial pop-ulations in hypersaline environments of different salt concen-trationsrdquoMicrobial Ecology vol 7 no 3 pp 235ndash243 1981

[4] F Rodriguez-Valera ldquoCharacteristics and microbial ecol-ogy of hypersaline environmentsrdquo in Halophilic Bacteria FRodriguez-Valera Ed vol 1 pp 3ndash30 CRC Press Boca RatonFla USA 1988

[5] D J Kushner and M Kamekura ldquoPhysiology of halophiliceubacteriardquo inHalophilic Bacteria F Rodriguez-Valera Ed vol1 pp 109ndash138 CRC press Boca Raton Fla USA 1988

[6] M A Amoozegar F Malekzadeh and K AMalik ldquoProductionof amylase by newly isolated moderate halophile Halobacillussp strainMA-2rdquo Journal of Microbiological Methods vol 52 no3 pp 353ndash359 2003

[7] C O Jeon J Lim J Lee et al ldquoLentibacillus salarius spnov isolated from saline sediment in China and emendeddescription of the genus Lentibacillusrdquo International Journal ofSystematic and Evolutionary Microbiology vol 55 no 3 pp1339ndash1343 2005

[8] S Bouchotroch E Quesada A Del Moral I Llamas andV Bejar ldquoHalomonas maura sp nov a novel moderatelyhalophilic exopolysaccharide-producing bacteriumrdquo Interna-tional Journal of Systematic and Evolutionary Microbiology vol51 no 5 pp 1625ndash1632 2001

[9] A Biswas A Patra and A Paul ldquoProduction of poly-3-hydroxyalkanoic acids by a moderately halophilic bacteriumHalomonas marina HMA 103 isolated from solar saltern ofOrissa Indiardquo Acta Microbiologica et Immunologica Hungaricavol 56 no 2 pp 125ndash143 2009

[10] S Y Jayachandra S Anil Kumar D P Merley and MB Sulochana ldquoIsolation and characterization of extremehalophilic bacterium Salinicoccus sp JAS4 producing extra-cellular hydrolytic enzymesrdquo Recent Research in Science andTechnology vol 4 no 4 pp 46ndash49 2012

Chinese Journal of Biology 11

[11] I W Sutherland ldquoNovel and established applications of micro-bial polysaccharidesrdquo Trends in Biotechnology vol 16 no 1 pp41ndash46 1998

[12] MWMittelman andG G Geesey ldquoCopper-binding character-istics of exopolymers from a freshwater-sediment bacteriumrdquoApplied and EnvironmentalMicrobiology vol 49 no 4 pp 846ndash851 1985

[13] J Anton I Meseguer and F Rodriguez-Valera ldquoProductionof an extracellular polysaccharide by Haloferax mediterraneirdquoApplied Enviromental Microbiology vol 54 no 10 pp 2381ndash2386 1988

[14] H Parolis L A S Parolis I F Boan et al ldquoThe structure ofthe exopolysaccharide produced by the halophilic ArchaeonHaloferax mediterranei strain R4 (ATCC 33500)rdquo CarbohydrateResearch vol 295 pp 147ndash156 1996

[15] E Quesada V Bejar and C Calvo ldquoExopolysaccharide produc-tion by Volcaniella eurihalinardquo Experientia vol 49 no 12 pp1037ndash1041 1993

[16] J A Mata V Bejar I Llamas et al ldquoExopolysaccharides pro-duced by the recently described halophilic bacteria Halomonasventosae and Halomonas anticariensisrdquo Research in Microbiol-ogy vol 157 no 9 pp 827ndash835 2006

[17] I Llamas J A Mata R Tallon et al ldquoCharacterization ofthe exopolysaccharide produced by Salipiger mucosus A3119879 ahalophilic species belonging to the Alphaproteobacteria iso-lated on the Spanish Mediterranean seaboardrdquo Marine Drugsvol 8 no 8 pp 2240ndash2251 2010

[18] F Martınez-Checa E Quesada J Martınez-Canovas I Llamasand V Bejar ldquoPalleronia marisminoris gen nov sp nova moderately halophilic exopolysaccharide-producing bac-terium belonging to the rsquoAlphaproteobacteriarsquo isolated from asaline soilrdquo International Journal of Systematic and EvolutionaryMicrobiology vol 55 no 6 pp 2525ndash2530 2005

[19] A Ventosa M T Garcia M Kamekura H Onishi and FRuiz-Berraquero ldquoBacillus halophilus sp nov a moderatelyhalophilic Bacillus speciesrdquo Systematic and Applied Microbiol-ogy vol 12 no 2 pp 162ndash166 1989

[20] R A Moraine and P Rogovin ldquoKinetics of polysaccharide B1459 fermentationrdquoBiotechnology and Bioengineering vol 8 no4 pp 511ndash524 1966

[21] M Dubois K A Gilles J K Hamilton P A Rebers and FSmith ldquoColorimetric method for determination of sugars andrelated substancesrdquoAnalytical Chemistry vol 28 no 3 pp 350ndash356 1956

[22] O H Lowry N J Rosenbrough A L Farr and R J RandallldquoProtein measurement with the Folin phenol reagentrdquo TheJournal of Biological Chemistry vol 193 no 1 pp 265ndash275 1951

[23] P Gerhardt R G E Murray W A Wood and N R KriegMethods for General and Molecular Bacteriology AmericanSociety for Microbiology Washington DC USA 1994

[24] G M Garrity J A Bell and T G Lilburn ldquoTaxonomic outlineof the procaryotesrdquo inBergeyrsquosManual of Systematic BacteriologD R Boone and R W Castenholz Eds Springer New YorkNY USA 2nd edition 2003

[25] J A Mata J Martınez-Canovas E Quesada and V BejarldquoA detailed phenotypic characterisation of the type strains ofHalomonas speciesrdquo Systematic and Applied Microbiology vol25 no 3 pp 360ndash375 2002

[26] A Oren ldquoMolecular ecology of extremely halophilic archaeaand bacteriardquo FEMSMicrobiology Ecology vol 39 no 1 pp 1ndash72002

[27] H Baati R Amdouni N Gharsallah A Sghir and E AmmarldquoIsolation and characterization of moderately halophilic bacte-ria from tunisian solar salternrdquo Current Microbiology vol 60no 3 pp 157ndash161 2010

[28] A Hedi N Sadfi M Fardeau et al ldquoStudies on the biodiversityof halophilic microorganisms isolated from El-Djerid salt lake(Tunisia) under aerobic conditionsrdquo International Journal ofMicrobiology vol 2009 Article ID 731786 17 pages 2009

[29] M Birbir A Ogan B Calli and B Mertoglu ldquoEnzymecharacteristics of extremely halophilic archaeal community inTuzkoy Salt Mine Turkeyrdquo World Journal of Microbiology andBiotechnology vol 20 no 6 pp 613ndash621 2004

[30] M Birbir and C Sesal ldquoExtremely halophilic bacterial commu-nities in Sereflikochisar Salt Lake in Turkeyrdquo Turkish Journal ofBiology vol 27 no 7 pp 7ndash22 2003

[31] M Birbir B Calli B Mertoglu et al ldquoExtremely halophilicArchaea from Tuz Lake Turkey and the adjacent Kaldirim andKayacik salternsrdquoWorld Journal ofMicrobiology and Biotechnol-ogy vol 23 no 3 pp 309ndash316 2007

[32] R Rohban M A Amoozegar and A Ventosa ldquoScreeningand isolation of halophilic bacteria producing extracellularhydrolyses from Howz Soltan Lake Iranrdquo Journal of IndustrialMicrobiology and Biotechnology vol 36 no 3 pp 333ndash3402009

[33] M J Garabito M C Marquez and A Ventosa ldquoHalotolerantBacillus diversity in hypersaline environmentsrdquo Canadian Jour-nal of Microbiology vol 44 no 2 pp 95ndash102 1998

[34] A Ventosa A Ramos-Cormenzana and M Kocur ldquoModer-ately halophilic gram-positive cocci from hypersaline environ-mentsrdquo Systematic and Applied Microbiology vol 4 no 4 pp564ndash570 1983

[35] E Quesada A Ventosa F Rodriguez-Valera and A R Cor-menzana ldquoTypes and properties of some bacteria isolated fromhypersaline soilsrdquo Journal of Applied Bacteriology vol 53 no 2pp 155ndash161 1982

[36] E Quesada V Bejar M J Valderrama A Ventosa and AR Ramos Cormenzana ldquoIsolation and characterization ofmoderately halophilic nonmotile rods from different salinehabitatsrdquoMicrobiologia vol 1 no 1-2 pp 89ndash96 1985

[37] S R Dave and H B Desai ldquoMicrobial diversity at marinesalterns near Bhavnagar Gujarat Indiardquo Current Science vol60 no 4 pp 497ndash500 2006

[38] S Kumar R Karan S Kapoor S P Singh and S K KhareldquoScreening and isolation of halophilic bacteria producingindustrially important enzymesrdquo Brazilian Journal of Microbi-ology vol 43 no 4 pp 1595ndash1603 2012

[39] B P Dave and A Soni ldquoDiversity of halophilic archaea atsalt pans around Bhavnagar coast Gujaratrdquo Proceedings ofthe National Academy of Sciences India Section B BiologicalSciences vol 83 no 2 pp 225ndash232 2013

[40] A A Joshi P P Kanekar A S Kelkar et al ldquoCultivable bacterialdiversity of alkaline Lonar lake Indiardquo Microbial Ecology vol55 no 2 pp 163ndash172 2008

[41] T M Raghavan and I Furtado ldquoOccurrence of extremelyhalophilic Archaea in sediments from the continental shelf ofwest coast of Indiardquo Current Science vol 86 no 8 pp 1065ndash1067 2004

[42] S Vijayanand J Hemapriya J Selvin and S Kiran ldquoBiodiver-sity of extremely halophilic bacterial strains isolated from solarsalterns of Tuticorin Tamilnadu Indiardquo International Journal ofWater Resources and Environmental Sciences vol 1 no 1 pp 1ndash72012

12 Chinese Journal of Biology

[43] S G Nanjani and H P Soni ldquoIsolation and characterization ofextremely halotolerant and halophilic organisms from Dwarkaand Veravalrdquo Journal of Pharmacy and Biological Sciences vol2 no 2 pp 20ndash25 2012

[44] H Ghozlan H Deif R A Kandil and S Sabry ldquoBiodiversity ofmoderately halophilic bacteria in hypersaline habitats in EgyptrdquoJournal of General and Applied Microbiology vol 52 no 2 pp63ndash72 2006

[45] D R Arahal M T Garcıa W Ludwig K H Schleifer andA Ventosa ldquoTransfer ofHalomonas canadensis andHalomonasisraelensis to the genus Chromohalobacter as Chromohalobac-ter canadensis comb nov and Chromohalobacter israelensiscomb novrdquo International Journal of Systematic and EvolutionaryMicrobiology vol 51 no 4 pp 1443ndash1448 2001

Submit your manuscripts athttpwwwhindawicom

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Anatomy Research International

PeptidesInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporation httpwwwhindawicom

International Journal of

Volume 2014

Zoology

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Molecular Biology International

GenomicsInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

The Scientific World JournalHindawi Publishing Corporation httpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

BioinformaticsAdvances in

Marine BiologyJournal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Signal TransductionJournal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

BioMed Research International

Evolutionary BiologyInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Biochemistry Research International

ArchaeaHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Genetics Research International

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Advances in

Virolog y

Hindawi Publishing Corporationhttpwwwhindawicom

Nucleic AcidsJournal of

Volume 2014

Stem CellsInternational

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

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Enzyme Research

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

International Journal of

Microbiology

Page 9: Research Article Production of Extracellular …downloads.hindawi.com/archive/2014/205731.pdfResearch Article Production of Extracellular Polymeric Substances by Halophilic Bacteria

Chinese Journal of Biology 9

Table10A

ntibiotic

sensitivitypatte

rnof

thes

electedEP

Sprod

ucinghaloph

ilicb

acteria

isolated

from

solarsalternso

fInd

ia

Antibiotics

Bacterialisolates

JS 504

JS 802

JS 803

JS 904

JW 302

JW 307

KS 1101

KS 1805

KW 203

VW 301

VW 402

SUR

108

SUR

301

SUR

302

SUR

303

SUR

307

SUR

309

SUR

310

SUR

202

SUR

201

Ampicillin(10120583

g)S

SR

SS

SS

RI

SS

SR

SS

SS

SS

SAmoxicillin

(30120583

g)S

SR

RS

SS

RS

SS

SR

RS

IR

SS

SBa

citracin

(10120583

g)S

SR

SS

SR

RS

IR

RR

RS

IR

RR

RCiprofl

oxacin

(30120583

g)I

II

RI

II

IR

II

II

IR

SR

II

ICh

lorte

tracyclin

e(30120583g)

SI

SS

IS

RR

SS

SS

RR

SS

II

IS

Chloramph

enicol(10120583

g)S

SS

SS

SS

SR

SS

SS

SS

SR

SS

SErythrom

ycin

(15120583

g)S

SI

SS

IS

II

II

II

SR

RI

IS

SFu

sidicacid

(30120583

g)I

RR

SS

IR

RS

RI

RR

RS

IR

II

RGentamycin

(10120583

g)S

SS

SS

SS

SR

SS

SS

SS

IS

SS

SKa

namycin

(30120583

g)I

IS

RS

II

SR

II

RI

RR

RI

RI

RMethicillin(5120583g)

SS

RS

IR

RR

RR

RR

RR

SR

RR

RR

Neomycin

(30120583

g)S

SI

RI

IR

II

IR

II

RR

RI

IR

RNorflo

xacin(30120583

g)S

SR

SS

SS

SI

SS

SS

SS

SS

SS

SNetillin

(10120583

g)R

SS

IR

SS

SR

RR

SS

RR

RR

RR

RNalidixicacid

(30120583

g)R

RS

RR

II

RR

II

RR

RR

SR

RR

RNovob

iosin

(30120583

g)I

RR

SR

SS

RS

SS

IR

SS

SI

SS

SPo

lymixin

B(50120583

g)R

RI

RI

IR

RR

RI

IR

RR

II

II

SPenicillinG(10120583

g)S

SR

IS

IR

RR

II

SR

RS

II

II

SStreptom

ycin

(10120583

g)S

SS

RS

RR

SR

RR

IS

RS

SR

II

RTetracyclin

e(30120583g)

RR

RR

RI

RR

SR

IR

RR

RS

RR

RI

Trim

etho

prim

e(30120583g)

SS

SS

IS

SI

SS

SS

IR

RS

SS

SS

Vancom

ycin

(30120583

g)R

RR

SR

RR

RS

RR

RR

RS

RR

RR

R

S=SensitiveR

=Re

sistantI

=Interm

ediate

10 Chinese Journal of Biology

0

1

2

3

4

Bacterial isolate

Gro

wth

OD

at540

nmto

tal E

PS (g

L)

OD at 540nmTotal EPS

JS504

JS802

JS803

JS904

JW302

JW307

KS1101

KS1805

KW203

VW301

VW402

SUR1

08

SUR3

01

SUR3

02

SUR3

03

SUR3

07

SUR3

09

SUR3

10

SUR2

02

SUR2

01

Figure 1 Screening of moderately halophilic bacteria isolatedfrom soil and water samples of solar salterns for production ofextracellular polymeric substances

0

02

04

06

08

1

Ant

ibio

tic re

sista

nce i

ndex

Bacterial isolate

JS504

JS802

JS803

JS904

JW302

JW307

KS1101

KS1805

KW203

SUR1

08

SUR2

01

SUR2

02

SUR3

01

SUR3

02

SUR3

03

SUR3

07

SUR3

09

SUR3

10

VW301

VW402

Figure 2 Antibiotic resistance index of the selected EPS producinghalophilic bacteria isolated from soil and water samples of solarsalterns of India

The isolates SUR301 and KS 1805 were tentatively iden-tified as Chromohalobacter based on the study by Arahal etal [45] and bear striking similarity with those of Chromo-halobacter sp isolated fromhypersaline soil sample of TriveniSangam Gujarat [38] The physiological morphological andbiochemical characteristics of isolate SUR303 were consistentwith the features of the genus Salinicoccus [10] while isolatesJS904 and JS504 were tentatively assigned to Aidingimonasand Alteromonas respectively

Finally sensitivity of these isolates to antibiotics (Table 10and Figure 2) also corroborates the findings ofMata et al [25]and Hedi et al [28]

5 Conclusion

It may be emphasized that study of these moderatelyhalophilic bacteria from coastal hypersaline solar salterns ofIndia with special attempt on screening for EPS productionhas led to the discovery of wide variety of halophilic speciesAlthough attention has been focused on the production

of EPS by these halophilic bacterial isolates their physio-biochemical features indicate that they may equally holdpotential towards production of various biomolecules ofindustrial interest

Conflict of Interests

It is declared by the authors that there is no conflict of interestsregarding the publication of this paper

Acknowledgment

This study was financially supported by grants from Uni-versity Grants Commission India (Sanction no F14-2(SC)2008 (SA-III) 31 March 2009) under the Scheme ofRajiv Gandhi National Fellowship

References

[1] E A Galinski and B J Tindall ldquoBiotechnological prospectsfor halophiles and halotolerant microorganismsrdquo in MolecularBiology and Biotechnology of Extremophiles R D Herbert andR J Sharp Eds Blackie London UK 1992

[2] R Margesin and F Schinner ldquoPotential of halotolerant andhalophilic microorganisms for biotechnologyrdquo Extremophilesvol 5 no 2 pp 73ndash83 2001

[3] F Rodriguez Valera F Ruiz Berraquero and A Ramos Cor-menzana ldquoCharacteristics of the heterotrophic bacterial pop-ulations in hypersaline environments of different salt concen-trationsrdquoMicrobial Ecology vol 7 no 3 pp 235ndash243 1981

[4] F Rodriguez-Valera ldquoCharacteristics and microbial ecol-ogy of hypersaline environmentsrdquo in Halophilic Bacteria FRodriguez-Valera Ed vol 1 pp 3ndash30 CRC Press Boca RatonFla USA 1988

[5] D J Kushner and M Kamekura ldquoPhysiology of halophiliceubacteriardquo inHalophilic Bacteria F Rodriguez-Valera Ed vol1 pp 109ndash138 CRC press Boca Raton Fla USA 1988

[6] M A Amoozegar F Malekzadeh and K AMalik ldquoProductionof amylase by newly isolated moderate halophile Halobacillussp strainMA-2rdquo Journal of Microbiological Methods vol 52 no3 pp 353ndash359 2003

[7] C O Jeon J Lim J Lee et al ldquoLentibacillus salarius spnov isolated from saline sediment in China and emendeddescription of the genus Lentibacillusrdquo International Journal ofSystematic and Evolutionary Microbiology vol 55 no 3 pp1339ndash1343 2005

[8] S Bouchotroch E Quesada A Del Moral I Llamas andV Bejar ldquoHalomonas maura sp nov a novel moderatelyhalophilic exopolysaccharide-producing bacteriumrdquo Interna-tional Journal of Systematic and Evolutionary Microbiology vol51 no 5 pp 1625ndash1632 2001

[9] A Biswas A Patra and A Paul ldquoProduction of poly-3-hydroxyalkanoic acids by a moderately halophilic bacteriumHalomonas marina HMA 103 isolated from solar saltern ofOrissa Indiardquo Acta Microbiologica et Immunologica Hungaricavol 56 no 2 pp 125ndash143 2009

[10] S Y Jayachandra S Anil Kumar D P Merley and MB Sulochana ldquoIsolation and characterization of extremehalophilic bacterium Salinicoccus sp JAS4 producing extra-cellular hydrolytic enzymesrdquo Recent Research in Science andTechnology vol 4 no 4 pp 46ndash49 2012

Chinese Journal of Biology 11

[11] I W Sutherland ldquoNovel and established applications of micro-bial polysaccharidesrdquo Trends in Biotechnology vol 16 no 1 pp41ndash46 1998

[12] MWMittelman andG G Geesey ldquoCopper-binding character-istics of exopolymers from a freshwater-sediment bacteriumrdquoApplied and EnvironmentalMicrobiology vol 49 no 4 pp 846ndash851 1985

[13] J Anton I Meseguer and F Rodriguez-Valera ldquoProductionof an extracellular polysaccharide by Haloferax mediterraneirdquoApplied Enviromental Microbiology vol 54 no 10 pp 2381ndash2386 1988

[14] H Parolis L A S Parolis I F Boan et al ldquoThe structure ofthe exopolysaccharide produced by the halophilic ArchaeonHaloferax mediterranei strain R4 (ATCC 33500)rdquo CarbohydrateResearch vol 295 pp 147ndash156 1996

[15] E Quesada V Bejar and C Calvo ldquoExopolysaccharide produc-tion by Volcaniella eurihalinardquo Experientia vol 49 no 12 pp1037ndash1041 1993

[16] J A Mata V Bejar I Llamas et al ldquoExopolysaccharides pro-duced by the recently described halophilic bacteria Halomonasventosae and Halomonas anticariensisrdquo Research in Microbiol-ogy vol 157 no 9 pp 827ndash835 2006

[17] I Llamas J A Mata R Tallon et al ldquoCharacterization ofthe exopolysaccharide produced by Salipiger mucosus A3119879 ahalophilic species belonging to the Alphaproteobacteria iso-lated on the Spanish Mediterranean seaboardrdquo Marine Drugsvol 8 no 8 pp 2240ndash2251 2010

[18] F Martınez-Checa E Quesada J Martınez-Canovas I Llamasand V Bejar ldquoPalleronia marisminoris gen nov sp nova moderately halophilic exopolysaccharide-producing bac-terium belonging to the rsquoAlphaproteobacteriarsquo isolated from asaline soilrdquo International Journal of Systematic and EvolutionaryMicrobiology vol 55 no 6 pp 2525ndash2530 2005

[19] A Ventosa M T Garcia M Kamekura H Onishi and FRuiz-Berraquero ldquoBacillus halophilus sp nov a moderatelyhalophilic Bacillus speciesrdquo Systematic and Applied Microbiol-ogy vol 12 no 2 pp 162ndash166 1989

[20] R A Moraine and P Rogovin ldquoKinetics of polysaccharide B1459 fermentationrdquoBiotechnology and Bioengineering vol 8 no4 pp 511ndash524 1966

[21] M Dubois K A Gilles J K Hamilton P A Rebers and FSmith ldquoColorimetric method for determination of sugars andrelated substancesrdquoAnalytical Chemistry vol 28 no 3 pp 350ndash356 1956

[22] O H Lowry N J Rosenbrough A L Farr and R J RandallldquoProtein measurement with the Folin phenol reagentrdquo TheJournal of Biological Chemistry vol 193 no 1 pp 265ndash275 1951

[23] P Gerhardt R G E Murray W A Wood and N R KriegMethods for General and Molecular Bacteriology AmericanSociety for Microbiology Washington DC USA 1994

[24] G M Garrity J A Bell and T G Lilburn ldquoTaxonomic outlineof the procaryotesrdquo inBergeyrsquosManual of Systematic BacteriologD R Boone and R W Castenholz Eds Springer New YorkNY USA 2nd edition 2003

[25] J A Mata J Martınez-Canovas E Quesada and V BejarldquoA detailed phenotypic characterisation of the type strains ofHalomonas speciesrdquo Systematic and Applied Microbiology vol25 no 3 pp 360ndash375 2002

[26] A Oren ldquoMolecular ecology of extremely halophilic archaeaand bacteriardquo FEMSMicrobiology Ecology vol 39 no 1 pp 1ndash72002

[27] H Baati R Amdouni N Gharsallah A Sghir and E AmmarldquoIsolation and characterization of moderately halophilic bacte-ria from tunisian solar salternrdquo Current Microbiology vol 60no 3 pp 157ndash161 2010

[28] A Hedi N Sadfi M Fardeau et al ldquoStudies on the biodiversityof halophilic microorganisms isolated from El-Djerid salt lake(Tunisia) under aerobic conditionsrdquo International Journal ofMicrobiology vol 2009 Article ID 731786 17 pages 2009

[29] M Birbir A Ogan B Calli and B Mertoglu ldquoEnzymecharacteristics of extremely halophilic archaeal community inTuzkoy Salt Mine Turkeyrdquo World Journal of Microbiology andBiotechnology vol 20 no 6 pp 613ndash621 2004

[30] M Birbir and C Sesal ldquoExtremely halophilic bacterial commu-nities in Sereflikochisar Salt Lake in Turkeyrdquo Turkish Journal ofBiology vol 27 no 7 pp 7ndash22 2003

[31] M Birbir B Calli B Mertoglu et al ldquoExtremely halophilicArchaea from Tuz Lake Turkey and the adjacent Kaldirim andKayacik salternsrdquoWorld Journal ofMicrobiology and Biotechnol-ogy vol 23 no 3 pp 309ndash316 2007

[32] R Rohban M A Amoozegar and A Ventosa ldquoScreeningand isolation of halophilic bacteria producing extracellularhydrolyses from Howz Soltan Lake Iranrdquo Journal of IndustrialMicrobiology and Biotechnology vol 36 no 3 pp 333ndash3402009

[33] M J Garabito M C Marquez and A Ventosa ldquoHalotolerantBacillus diversity in hypersaline environmentsrdquo Canadian Jour-nal of Microbiology vol 44 no 2 pp 95ndash102 1998

[34] A Ventosa A Ramos-Cormenzana and M Kocur ldquoModer-ately halophilic gram-positive cocci from hypersaline environ-mentsrdquo Systematic and Applied Microbiology vol 4 no 4 pp564ndash570 1983

[35] E Quesada A Ventosa F Rodriguez-Valera and A R Cor-menzana ldquoTypes and properties of some bacteria isolated fromhypersaline soilsrdquo Journal of Applied Bacteriology vol 53 no 2pp 155ndash161 1982

[36] E Quesada V Bejar M J Valderrama A Ventosa and AR Ramos Cormenzana ldquoIsolation and characterization ofmoderately halophilic nonmotile rods from different salinehabitatsrdquoMicrobiologia vol 1 no 1-2 pp 89ndash96 1985

[37] S R Dave and H B Desai ldquoMicrobial diversity at marinesalterns near Bhavnagar Gujarat Indiardquo Current Science vol60 no 4 pp 497ndash500 2006

[38] S Kumar R Karan S Kapoor S P Singh and S K KhareldquoScreening and isolation of halophilic bacteria producingindustrially important enzymesrdquo Brazilian Journal of Microbi-ology vol 43 no 4 pp 1595ndash1603 2012

[39] B P Dave and A Soni ldquoDiversity of halophilic archaea atsalt pans around Bhavnagar coast Gujaratrdquo Proceedings ofthe National Academy of Sciences India Section B BiologicalSciences vol 83 no 2 pp 225ndash232 2013

[40] A A Joshi P P Kanekar A S Kelkar et al ldquoCultivable bacterialdiversity of alkaline Lonar lake Indiardquo Microbial Ecology vol55 no 2 pp 163ndash172 2008

[41] T M Raghavan and I Furtado ldquoOccurrence of extremelyhalophilic Archaea in sediments from the continental shelf ofwest coast of Indiardquo Current Science vol 86 no 8 pp 1065ndash1067 2004

[42] S Vijayanand J Hemapriya J Selvin and S Kiran ldquoBiodiver-sity of extremely halophilic bacterial strains isolated from solarsalterns of Tuticorin Tamilnadu Indiardquo International Journal ofWater Resources and Environmental Sciences vol 1 no 1 pp 1ndash72012

12 Chinese Journal of Biology

[43] S G Nanjani and H P Soni ldquoIsolation and characterization ofextremely halotolerant and halophilic organisms from Dwarkaand Veravalrdquo Journal of Pharmacy and Biological Sciences vol2 no 2 pp 20ndash25 2012

[44] H Ghozlan H Deif R A Kandil and S Sabry ldquoBiodiversity ofmoderately halophilic bacteria in hypersaline habitats in EgyptrdquoJournal of General and Applied Microbiology vol 52 no 2 pp63ndash72 2006

[45] D R Arahal M T Garcıa W Ludwig K H Schleifer andA Ventosa ldquoTransfer ofHalomonas canadensis andHalomonasisraelensis to the genus Chromohalobacter as Chromohalobac-ter canadensis comb nov and Chromohalobacter israelensiscomb novrdquo International Journal of Systematic and EvolutionaryMicrobiology vol 51 no 4 pp 1443ndash1448 2001

Submit your manuscripts athttpwwwhindawicom

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Anatomy Research International

PeptidesInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporation httpwwwhindawicom

International Journal of

Volume 2014

Zoology

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Molecular Biology International

GenomicsInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

The Scientific World JournalHindawi Publishing Corporation httpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

BioinformaticsAdvances in

Marine BiologyJournal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Signal TransductionJournal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

BioMed Research International

Evolutionary BiologyInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Biochemistry Research International

ArchaeaHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Genetics Research International

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Advances in

Virolog y

Hindawi Publishing Corporationhttpwwwhindawicom

Nucleic AcidsJournal of

Volume 2014

Stem CellsInternational

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Enzyme Research

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

International Journal of

Microbiology

Page 10: Research Article Production of Extracellular …downloads.hindawi.com/archive/2014/205731.pdfResearch Article Production of Extracellular Polymeric Substances by Halophilic Bacteria

10 Chinese Journal of Biology

0

1

2

3

4

Bacterial isolate

Gro

wth

OD

at540

nmto

tal E

PS (g

L)

OD at 540nmTotal EPS

JS504

JS802

JS803

JS904

JW302

JW307

KS1101

KS1805

KW203

VW301

VW402

SUR1

08

SUR3

01

SUR3

02

SUR3

03

SUR3

07

SUR3

09

SUR3

10

SUR2

02

SUR2

01

Figure 1 Screening of moderately halophilic bacteria isolatedfrom soil and water samples of solar salterns for production ofextracellular polymeric substances

0

02

04

06

08

1

Ant

ibio

tic re

sista

nce i

ndex

Bacterial isolate

JS504

JS802

JS803

JS904

JW302

JW307

KS1101

KS1805

KW203

SUR1

08

SUR2

01

SUR2

02

SUR3

01

SUR3

02

SUR3

03

SUR3

07

SUR3

09

SUR3

10

VW301

VW402

Figure 2 Antibiotic resistance index of the selected EPS producinghalophilic bacteria isolated from soil and water samples of solarsalterns of India

The isolates SUR301 and KS 1805 were tentatively iden-tified as Chromohalobacter based on the study by Arahal etal [45] and bear striking similarity with those of Chromo-halobacter sp isolated fromhypersaline soil sample of TriveniSangam Gujarat [38] The physiological morphological andbiochemical characteristics of isolate SUR303 were consistentwith the features of the genus Salinicoccus [10] while isolatesJS904 and JS504 were tentatively assigned to Aidingimonasand Alteromonas respectively

Finally sensitivity of these isolates to antibiotics (Table 10and Figure 2) also corroborates the findings ofMata et al [25]and Hedi et al [28]

5 Conclusion

It may be emphasized that study of these moderatelyhalophilic bacteria from coastal hypersaline solar salterns ofIndia with special attempt on screening for EPS productionhas led to the discovery of wide variety of halophilic speciesAlthough attention has been focused on the production

of EPS by these halophilic bacterial isolates their physio-biochemical features indicate that they may equally holdpotential towards production of various biomolecules ofindustrial interest

Conflict of Interests

It is declared by the authors that there is no conflict of interestsregarding the publication of this paper

Acknowledgment

This study was financially supported by grants from Uni-versity Grants Commission India (Sanction no F14-2(SC)2008 (SA-III) 31 March 2009) under the Scheme ofRajiv Gandhi National Fellowship

References

[1] E A Galinski and B J Tindall ldquoBiotechnological prospectsfor halophiles and halotolerant microorganismsrdquo in MolecularBiology and Biotechnology of Extremophiles R D Herbert andR J Sharp Eds Blackie London UK 1992

[2] R Margesin and F Schinner ldquoPotential of halotolerant andhalophilic microorganisms for biotechnologyrdquo Extremophilesvol 5 no 2 pp 73ndash83 2001

[3] F Rodriguez Valera F Ruiz Berraquero and A Ramos Cor-menzana ldquoCharacteristics of the heterotrophic bacterial pop-ulations in hypersaline environments of different salt concen-trationsrdquoMicrobial Ecology vol 7 no 3 pp 235ndash243 1981

[4] F Rodriguez-Valera ldquoCharacteristics and microbial ecol-ogy of hypersaline environmentsrdquo in Halophilic Bacteria FRodriguez-Valera Ed vol 1 pp 3ndash30 CRC Press Boca RatonFla USA 1988

[5] D J Kushner and M Kamekura ldquoPhysiology of halophiliceubacteriardquo inHalophilic Bacteria F Rodriguez-Valera Ed vol1 pp 109ndash138 CRC press Boca Raton Fla USA 1988

[6] M A Amoozegar F Malekzadeh and K AMalik ldquoProductionof amylase by newly isolated moderate halophile Halobacillussp strainMA-2rdquo Journal of Microbiological Methods vol 52 no3 pp 353ndash359 2003

[7] C O Jeon J Lim J Lee et al ldquoLentibacillus salarius spnov isolated from saline sediment in China and emendeddescription of the genus Lentibacillusrdquo International Journal ofSystematic and Evolutionary Microbiology vol 55 no 3 pp1339ndash1343 2005

[8] S Bouchotroch E Quesada A Del Moral I Llamas andV Bejar ldquoHalomonas maura sp nov a novel moderatelyhalophilic exopolysaccharide-producing bacteriumrdquo Interna-tional Journal of Systematic and Evolutionary Microbiology vol51 no 5 pp 1625ndash1632 2001

[9] A Biswas A Patra and A Paul ldquoProduction of poly-3-hydroxyalkanoic acids by a moderately halophilic bacteriumHalomonas marina HMA 103 isolated from solar saltern ofOrissa Indiardquo Acta Microbiologica et Immunologica Hungaricavol 56 no 2 pp 125ndash143 2009

[10] S Y Jayachandra S Anil Kumar D P Merley and MB Sulochana ldquoIsolation and characterization of extremehalophilic bacterium Salinicoccus sp JAS4 producing extra-cellular hydrolytic enzymesrdquo Recent Research in Science andTechnology vol 4 no 4 pp 46ndash49 2012

Chinese Journal of Biology 11

[11] I W Sutherland ldquoNovel and established applications of micro-bial polysaccharidesrdquo Trends in Biotechnology vol 16 no 1 pp41ndash46 1998

[12] MWMittelman andG G Geesey ldquoCopper-binding character-istics of exopolymers from a freshwater-sediment bacteriumrdquoApplied and EnvironmentalMicrobiology vol 49 no 4 pp 846ndash851 1985

[13] J Anton I Meseguer and F Rodriguez-Valera ldquoProductionof an extracellular polysaccharide by Haloferax mediterraneirdquoApplied Enviromental Microbiology vol 54 no 10 pp 2381ndash2386 1988

[14] H Parolis L A S Parolis I F Boan et al ldquoThe structure ofthe exopolysaccharide produced by the halophilic ArchaeonHaloferax mediterranei strain R4 (ATCC 33500)rdquo CarbohydrateResearch vol 295 pp 147ndash156 1996

[15] E Quesada V Bejar and C Calvo ldquoExopolysaccharide produc-tion by Volcaniella eurihalinardquo Experientia vol 49 no 12 pp1037ndash1041 1993

[16] J A Mata V Bejar I Llamas et al ldquoExopolysaccharides pro-duced by the recently described halophilic bacteria Halomonasventosae and Halomonas anticariensisrdquo Research in Microbiol-ogy vol 157 no 9 pp 827ndash835 2006

[17] I Llamas J A Mata R Tallon et al ldquoCharacterization ofthe exopolysaccharide produced by Salipiger mucosus A3119879 ahalophilic species belonging to the Alphaproteobacteria iso-lated on the Spanish Mediterranean seaboardrdquo Marine Drugsvol 8 no 8 pp 2240ndash2251 2010

[18] F Martınez-Checa E Quesada J Martınez-Canovas I Llamasand V Bejar ldquoPalleronia marisminoris gen nov sp nova moderately halophilic exopolysaccharide-producing bac-terium belonging to the rsquoAlphaproteobacteriarsquo isolated from asaline soilrdquo International Journal of Systematic and EvolutionaryMicrobiology vol 55 no 6 pp 2525ndash2530 2005

[19] A Ventosa M T Garcia M Kamekura H Onishi and FRuiz-Berraquero ldquoBacillus halophilus sp nov a moderatelyhalophilic Bacillus speciesrdquo Systematic and Applied Microbiol-ogy vol 12 no 2 pp 162ndash166 1989

[20] R A Moraine and P Rogovin ldquoKinetics of polysaccharide B1459 fermentationrdquoBiotechnology and Bioengineering vol 8 no4 pp 511ndash524 1966

[21] M Dubois K A Gilles J K Hamilton P A Rebers and FSmith ldquoColorimetric method for determination of sugars andrelated substancesrdquoAnalytical Chemistry vol 28 no 3 pp 350ndash356 1956

[22] O H Lowry N J Rosenbrough A L Farr and R J RandallldquoProtein measurement with the Folin phenol reagentrdquo TheJournal of Biological Chemistry vol 193 no 1 pp 265ndash275 1951

[23] P Gerhardt R G E Murray W A Wood and N R KriegMethods for General and Molecular Bacteriology AmericanSociety for Microbiology Washington DC USA 1994

[24] G M Garrity J A Bell and T G Lilburn ldquoTaxonomic outlineof the procaryotesrdquo inBergeyrsquosManual of Systematic BacteriologD R Boone and R W Castenholz Eds Springer New YorkNY USA 2nd edition 2003

[25] J A Mata J Martınez-Canovas E Quesada and V BejarldquoA detailed phenotypic characterisation of the type strains ofHalomonas speciesrdquo Systematic and Applied Microbiology vol25 no 3 pp 360ndash375 2002

[26] A Oren ldquoMolecular ecology of extremely halophilic archaeaand bacteriardquo FEMSMicrobiology Ecology vol 39 no 1 pp 1ndash72002

[27] H Baati R Amdouni N Gharsallah A Sghir and E AmmarldquoIsolation and characterization of moderately halophilic bacte-ria from tunisian solar salternrdquo Current Microbiology vol 60no 3 pp 157ndash161 2010

[28] A Hedi N Sadfi M Fardeau et al ldquoStudies on the biodiversityof halophilic microorganisms isolated from El-Djerid salt lake(Tunisia) under aerobic conditionsrdquo International Journal ofMicrobiology vol 2009 Article ID 731786 17 pages 2009

[29] M Birbir A Ogan B Calli and B Mertoglu ldquoEnzymecharacteristics of extremely halophilic archaeal community inTuzkoy Salt Mine Turkeyrdquo World Journal of Microbiology andBiotechnology vol 20 no 6 pp 613ndash621 2004

[30] M Birbir and C Sesal ldquoExtremely halophilic bacterial commu-nities in Sereflikochisar Salt Lake in Turkeyrdquo Turkish Journal ofBiology vol 27 no 7 pp 7ndash22 2003

[31] M Birbir B Calli B Mertoglu et al ldquoExtremely halophilicArchaea from Tuz Lake Turkey and the adjacent Kaldirim andKayacik salternsrdquoWorld Journal ofMicrobiology and Biotechnol-ogy vol 23 no 3 pp 309ndash316 2007

[32] R Rohban M A Amoozegar and A Ventosa ldquoScreeningand isolation of halophilic bacteria producing extracellularhydrolyses from Howz Soltan Lake Iranrdquo Journal of IndustrialMicrobiology and Biotechnology vol 36 no 3 pp 333ndash3402009

[33] M J Garabito M C Marquez and A Ventosa ldquoHalotolerantBacillus diversity in hypersaline environmentsrdquo Canadian Jour-nal of Microbiology vol 44 no 2 pp 95ndash102 1998

[34] A Ventosa A Ramos-Cormenzana and M Kocur ldquoModer-ately halophilic gram-positive cocci from hypersaline environ-mentsrdquo Systematic and Applied Microbiology vol 4 no 4 pp564ndash570 1983

[35] E Quesada A Ventosa F Rodriguez-Valera and A R Cor-menzana ldquoTypes and properties of some bacteria isolated fromhypersaline soilsrdquo Journal of Applied Bacteriology vol 53 no 2pp 155ndash161 1982

[36] E Quesada V Bejar M J Valderrama A Ventosa and AR Ramos Cormenzana ldquoIsolation and characterization ofmoderately halophilic nonmotile rods from different salinehabitatsrdquoMicrobiologia vol 1 no 1-2 pp 89ndash96 1985

[37] S R Dave and H B Desai ldquoMicrobial diversity at marinesalterns near Bhavnagar Gujarat Indiardquo Current Science vol60 no 4 pp 497ndash500 2006

[38] S Kumar R Karan S Kapoor S P Singh and S K KhareldquoScreening and isolation of halophilic bacteria producingindustrially important enzymesrdquo Brazilian Journal of Microbi-ology vol 43 no 4 pp 1595ndash1603 2012

[39] B P Dave and A Soni ldquoDiversity of halophilic archaea atsalt pans around Bhavnagar coast Gujaratrdquo Proceedings ofthe National Academy of Sciences India Section B BiologicalSciences vol 83 no 2 pp 225ndash232 2013

[40] A A Joshi P P Kanekar A S Kelkar et al ldquoCultivable bacterialdiversity of alkaline Lonar lake Indiardquo Microbial Ecology vol55 no 2 pp 163ndash172 2008

[41] T M Raghavan and I Furtado ldquoOccurrence of extremelyhalophilic Archaea in sediments from the continental shelf ofwest coast of Indiardquo Current Science vol 86 no 8 pp 1065ndash1067 2004

[42] S Vijayanand J Hemapriya J Selvin and S Kiran ldquoBiodiver-sity of extremely halophilic bacterial strains isolated from solarsalterns of Tuticorin Tamilnadu Indiardquo International Journal ofWater Resources and Environmental Sciences vol 1 no 1 pp 1ndash72012

12 Chinese Journal of Biology

[43] S G Nanjani and H P Soni ldquoIsolation and characterization ofextremely halotolerant and halophilic organisms from Dwarkaand Veravalrdquo Journal of Pharmacy and Biological Sciences vol2 no 2 pp 20ndash25 2012

[44] H Ghozlan H Deif R A Kandil and S Sabry ldquoBiodiversity ofmoderately halophilic bacteria in hypersaline habitats in EgyptrdquoJournal of General and Applied Microbiology vol 52 no 2 pp63ndash72 2006

[45] D R Arahal M T Garcıa W Ludwig K H Schleifer andA Ventosa ldquoTransfer ofHalomonas canadensis andHalomonasisraelensis to the genus Chromohalobacter as Chromohalobac-ter canadensis comb nov and Chromohalobacter israelensiscomb novrdquo International Journal of Systematic and EvolutionaryMicrobiology vol 51 no 4 pp 1443ndash1448 2001

Submit your manuscripts athttpwwwhindawicom

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Anatomy Research International

PeptidesInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporation httpwwwhindawicom

International Journal of

Volume 2014

Zoology

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Molecular Biology International

GenomicsInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

The Scientific World JournalHindawi Publishing Corporation httpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

BioinformaticsAdvances in

Marine BiologyJournal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Signal TransductionJournal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

BioMed Research International

Evolutionary BiologyInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Biochemistry Research International

ArchaeaHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Genetics Research International

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Advances in

Virolog y

Hindawi Publishing Corporationhttpwwwhindawicom

Nucleic AcidsJournal of

Volume 2014

Stem CellsInternational

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Enzyme Research

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

International Journal of

Microbiology

Page 11: Research Article Production of Extracellular …downloads.hindawi.com/archive/2014/205731.pdfResearch Article Production of Extracellular Polymeric Substances by Halophilic Bacteria

Chinese Journal of Biology 11

[11] I W Sutherland ldquoNovel and established applications of micro-bial polysaccharidesrdquo Trends in Biotechnology vol 16 no 1 pp41ndash46 1998

[12] MWMittelman andG G Geesey ldquoCopper-binding character-istics of exopolymers from a freshwater-sediment bacteriumrdquoApplied and EnvironmentalMicrobiology vol 49 no 4 pp 846ndash851 1985

[13] J Anton I Meseguer and F Rodriguez-Valera ldquoProductionof an extracellular polysaccharide by Haloferax mediterraneirdquoApplied Enviromental Microbiology vol 54 no 10 pp 2381ndash2386 1988

[14] H Parolis L A S Parolis I F Boan et al ldquoThe structure ofthe exopolysaccharide produced by the halophilic ArchaeonHaloferax mediterranei strain R4 (ATCC 33500)rdquo CarbohydrateResearch vol 295 pp 147ndash156 1996

[15] E Quesada V Bejar and C Calvo ldquoExopolysaccharide produc-tion by Volcaniella eurihalinardquo Experientia vol 49 no 12 pp1037ndash1041 1993

[16] J A Mata V Bejar I Llamas et al ldquoExopolysaccharides pro-duced by the recently described halophilic bacteria Halomonasventosae and Halomonas anticariensisrdquo Research in Microbiol-ogy vol 157 no 9 pp 827ndash835 2006

[17] I Llamas J A Mata R Tallon et al ldquoCharacterization ofthe exopolysaccharide produced by Salipiger mucosus A3119879 ahalophilic species belonging to the Alphaproteobacteria iso-lated on the Spanish Mediterranean seaboardrdquo Marine Drugsvol 8 no 8 pp 2240ndash2251 2010

[18] F Martınez-Checa E Quesada J Martınez-Canovas I Llamasand V Bejar ldquoPalleronia marisminoris gen nov sp nova moderately halophilic exopolysaccharide-producing bac-terium belonging to the rsquoAlphaproteobacteriarsquo isolated from asaline soilrdquo International Journal of Systematic and EvolutionaryMicrobiology vol 55 no 6 pp 2525ndash2530 2005

[19] A Ventosa M T Garcia M Kamekura H Onishi and FRuiz-Berraquero ldquoBacillus halophilus sp nov a moderatelyhalophilic Bacillus speciesrdquo Systematic and Applied Microbiol-ogy vol 12 no 2 pp 162ndash166 1989

[20] R A Moraine and P Rogovin ldquoKinetics of polysaccharide B1459 fermentationrdquoBiotechnology and Bioengineering vol 8 no4 pp 511ndash524 1966

[21] M Dubois K A Gilles J K Hamilton P A Rebers and FSmith ldquoColorimetric method for determination of sugars andrelated substancesrdquoAnalytical Chemistry vol 28 no 3 pp 350ndash356 1956

[22] O H Lowry N J Rosenbrough A L Farr and R J RandallldquoProtein measurement with the Folin phenol reagentrdquo TheJournal of Biological Chemistry vol 193 no 1 pp 265ndash275 1951

[23] P Gerhardt R G E Murray W A Wood and N R KriegMethods for General and Molecular Bacteriology AmericanSociety for Microbiology Washington DC USA 1994

[24] G M Garrity J A Bell and T G Lilburn ldquoTaxonomic outlineof the procaryotesrdquo inBergeyrsquosManual of Systematic BacteriologD R Boone and R W Castenholz Eds Springer New YorkNY USA 2nd edition 2003

[25] J A Mata J Martınez-Canovas E Quesada and V BejarldquoA detailed phenotypic characterisation of the type strains ofHalomonas speciesrdquo Systematic and Applied Microbiology vol25 no 3 pp 360ndash375 2002

[26] A Oren ldquoMolecular ecology of extremely halophilic archaeaand bacteriardquo FEMSMicrobiology Ecology vol 39 no 1 pp 1ndash72002

[27] H Baati R Amdouni N Gharsallah A Sghir and E AmmarldquoIsolation and characterization of moderately halophilic bacte-ria from tunisian solar salternrdquo Current Microbiology vol 60no 3 pp 157ndash161 2010

[28] A Hedi N Sadfi M Fardeau et al ldquoStudies on the biodiversityof halophilic microorganisms isolated from El-Djerid salt lake(Tunisia) under aerobic conditionsrdquo International Journal ofMicrobiology vol 2009 Article ID 731786 17 pages 2009

[29] M Birbir A Ogan B Calli and B Mertoglu ldquoEnzymecharacteristics of extremely halophilic archaeal community inTuzkoy Salt Mine Turkeyrdquo World Journal of Microbiology andBiotechnology vol 20 no 6 pp 613ndash621 2004

[30] M Birbir and C Sesal ldquoExtremely halophilic bacterial commu-nities in Sereflikochisar Salt Lake in Turkeyrdquo Turkish Journal ofBiology vol 27 no 7 pp 7ndash22 2003

[31] M Birbir B Calli B Mertoglu et al ldquoExtremely halophilicArchaea from Tuz Lake Turkey and the adjacent Kaldirim andKayacik salternsrdquoWorld Journal ofMicrobiology and Biotechnol-ogy vol 23 no 3 pp 309ndash316 2007

[32] R Rohban M A Amoozegar and A Ventosa ldquoScreeningand isolation of halophilic bacteria producing extracellularhydrolyses from Howz Soltan Lake Iranrdquo Journal of IndustrialMicrobiology and Biotechnology vol 36 no 3 pp 333ndash3402009

[33] M J Garabito M C Marquez and A Ventosa ldquoHalotolerantBacillus diversity in hypersaline environmentsrdquo Canadian Jour-nal of Microbiology vol 44 no 2 pp 95ndash102 1998

[34] A Ventosa A Ramos-Cormenzana and M Kocur ldquoModer-ately halophilic gram-positive cocci from hypersaline environ-mentsrdquo Systematic and Applied Microbiology vol 4 no 4 pp564ndash570 1983

[35] E Quesada A Ventosa F Rodriguez-Valera and A R Cor-menzana ldquoTypes and properties of some bacteria isolated fromhypersaline soilsrdquo Journal of Applied Bacteriology vol 53 no 2pp 155ndash161 1982

[36] E Quesada V Bejar M J Valderrama A Ventosa and AR Ramos Cormenzana ldquoIsolation and characterization ofmoderately halophilic nonmotile rods from different salinehabitatsrdquoMicrobiologia vol 1 no 1-2 pp 89ndash96 1985

[37] S R Dave and H B Desai ldquoMicrobial diversity at marinesalterns near Bhavnagar Gujarat Indiardquo Current Science vol60 no 4 pp 497ndash500 2006

[38] S Kumar R Karan S Kapoor S P Singh and S K KhareldquoScreening and isolation of halophilic bacteria producingindustrially important enzymesrdquo Brazilian Journal of Microbi-ology vol 43 no 4 pp 1595ndash1603 2012

[39] B P Dave and A Soni ldquoDiversity of halophilic archaea atsalt pans around Bhavnagar coast Gujaratrdquo Proceedings ofthe National Academy of Sciences India Section B BiologicalSciences vol 83 no 2 pp 225ndash232 2013

[40] A A Joshi P P Kanekar A S Kelkar et al ldquoCultivable bacterialdiversity of alkaline Lonar lake Indiardquo Microbial Ecology vol55 no 2 pp 163ndash172 2008

[41] T M Raghavan and I Furtado ldquoOccurrence of extremelyhalophilic Archaea in sediments from the continental shelf ofwest coast of Indiardquo Current Science vol 86 no 8 pp 1065ndash1067 2004

[42] S Vijayanand J Hemapriya J Selvin and S Kiran ldquoBiodiver-sity of extremely halophilic bacterial strains isolated from solarsalterns of Tuticorin Tamilnadu Indiardquo International Journal ofWater Resources and Environmental Sciences vol 1 no 1 pp 1ndash72012

12 Chinese Journal of Biology

[43] S G Nanjani and H P Soni ldquoIsolation and characterization ofextremely halotolerant and halophilic organisms from Dwarkaand Veravalrdquo Journal of Pharmacy and Biological Sciences vol2 no 2 pp 20ndash25 2012

[44] H Ghozlan H Deif R A Kandil and S Sabry ldquoBiodiversity ofmoderately halophilic bacteria in hypersaline habitats in EgyptrdquoJournal of General and Applied Microbiology vol 52 no 2 pp63ndash72 2006

[45] D R Arahal M T Garcıa W Ludwig K H Schleifer andA Ventosa ldquoTransfer ofHalomonas canadensis andHalomonasisraelensis to the genus Chromohalobacter as Chromohalobac-ter canadensis comb nov and Chromohalobacter israelensiscomb novrdquo International Journal of Systematic and EvolutionaryMicrobiology vol 51 no 4 pp 1443ndash1448 2001

Submit your manuscripts athttpwwwhindawicom

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Anatomy Research International

PeptidesInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporation httpwwwhindawicom

International Journal of

Volume 2014

Zoology

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Molecular Biology International

GenomicsInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

The Scientific World JournalHindawi Publishing Corporation httpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

BioinformaticsAdvances in

Marine BiologyJournal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Signal TransductionJournal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

BioMed Research International

Evolutionary BiologyInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Biochemistry Research International

ArchaeaHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Genetics Research International

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Advances in

Virolog y

Hindawi Publishing Corporationhttpwwwhindawicom

Nucleic AcidsJournal of

Volume 2014

Stem CellsInternational

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Enzyme Research

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

International Journal of

Microbiology

Page 12: Research Article Production of Extracellular …downloads.hindawi.com/archive/2014/205731.pdfResearch Article Production of Extracellular Polymeric Substances by Halophilic Bacteria

12 Chinese Journal of Biology

[43] S G Nanjani and H P Soni ldquoIsolation and characterization ofextremely halotolerant and halophilic organisms from Dwarkaand Veravalrdquo Journal of Pharmacy and Biological Sciences vol2 no 2 pp 20ndash25 2012

[44] H Ghozlan H Deif R A Kandil and S Sabry ldquoBiodiversity ofmoderately halophilic bacteria in hypersaline habitats in EgyptrdquoJournal of General and Applied Microbiology vol 52 no 2 pp63ndash72 2006

[45] D R Arahal M T Garcıa W Ludwig K H Schleifer andA Ventosa ldquoTransfer ofHalomonas canadensis andHalomonasisraelensis to the genus Chromohalobacter as Chromohalobac-ter canadensis comb nov and Chromohalobacter israelensiscomb novrdquo International Journal of Systematic and EvolutionaryMicrobiology vol 51 no 4 pp 1443ndash1448 2001

Submit your manuscripts athttpwwwhindawicom

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Anatomy Research International

PeptidesInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporation httpwwwhindawicom

International Journal of

Volume 2014

Zoology

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Molecular Biology International

GenomicsInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

The Scientific World JournalHindawi Publishing Corporation httpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

BioinformaticsAdvances in

Marine BiologyJournal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Signal TransductionJournal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

BioMed Research International

Evolutionary BiologyInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Biochemistry Research International

ArchaeaHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Genetics Research International

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Advances in

Virolog y

Hindawi Publishing Corporationhttpwwwhindawicom

Nucleic AcidsJournal of

Volume 2014

Stem CellsInternational

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Enzyme Research

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

International Journal of

Microbiology

Page 13: Research Article Production of Extracellular …downloads.hindawi.com/archive/2014/205731.pdfResearch Article Production of Extracellular Polymeric Substances by Halophilic Bacteria

Submit your manuscripts athttpwwwhindawicom

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Anatomy Research International

PeptidesInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporation httpwwwhindawicom

International Journal of

Volume 2014

Zoology

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Molecular Biology International

GenomicsInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

The Scientific World JournalHindawi Publishing Corporation httpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

BioinformaticsAdvances in

Marine BiologyJournal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Signal TransductionJournal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

BioMed Research International

Evolutionary BiologyInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Biochemistry Research International

ArchaeaHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Genetics Research International

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Advances in

Virolog y

Hindawi Publishing Corporationhttpwwwhindawicom

Nucleic AcidsJournal of

Volume 2014

Stem CellsInternational

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Enzyme Research

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

International Journal of

Microbiology


SCREENING OF HALOPHILIC OR SALT TOLERANT LICHEN …

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Genome Sequence of an Extremely Halophilic Archaeon

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Crystallography of halophilic ribosome: the isolation of