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Available online at http:/ /www.urpjournals.com International Journal of Environmental Biology Universal Research

Available online at http://www.urpjournals.com

International Journal of Environmental Biology

Universal Research Publications. All rights reserved

Universal Research Publications. All rights reserved ISSN 2277 – 386X Original Article Biodegradation

ISSN 2277386X Original Article Biodegradation potential of some bacterial strains isolated from sewage water

Sankar Narayan Sinha* and Dipak Paul Environmental Microbiology Research Laboratory, Department of Botany, University of Kalyani, Kalyani 741235, West Bengal, India *Corresponding author email: sinhasn62@yahoo.co.in

Received 20 March 2014; accepted 28 March 2014

Abstract

Bacteria are well known for their scavenging activity. They are capable of producing a wide variety of enzymes that can degrade complex organic compounds. The present study aims at the isolation of different bacterial strains that are capable of removing organic matters like protein, carbohydrates etc. and alter BOD and COD values of river water. Highest removal of protein and carbohydrate was brought about by Micrococcus luteus SML-1 followed by SML-4 and SML-5. Micrococcus luteus SML-1 and SML-4 removed maximum COD, while maximum BOD was removed by Staphylococcus aureus SSA-1 isolate.

© 2014 Universal Research Publications. All rights reserved Keywords: Biodegradation, Sewage, Micrococcus luteus, Staphylococcus aureus

1. Introduction Water plays an important role in the life of human beings. It is most vital and important resource of our planet. Water is being deteriorated by environmental pollution and also by some other factors [1]. In the last few decades, limitless urbanization has caused a serious pollution problem due to the disposal of sewage and industrial effluents to water bodies. Sewage is waste matter resulting from the discharge into the sewers of human excreta and wastewater originating from a community and its industries [2]. It has a high content of both inorganic and organic matter, as well as high densities of living biota which include pathogenic, commensal and environmental bacteria [3]. In India it is reported that about 70% of the available water is polluted. The chief source of pollution is identified as sewage constituting 84 to 92 percent of the waste water [4]. River and aquatic ecosystems continually received huge amount of sewage and industrial effluents, changing the status of different water quality parameters such as dissolved oxygen, BOD, COD etc [5]. In recent decades the threat due to pollution has become a matter of serious concern. As the quantity of untreated sewage is enormous, it accumulates in the vicinity of human dwellings, leading to uncontrolled decomposition of organic content [6]. Several physico-chemical methods of treatment of sewage and industrial effluents are there to minimize the pollution load of the aquatic water bodies such as adsorption, chlorination, coagulation, flocculation, ozonation, solvent extraction, membrane process and

biological degradation [7,8]. However, these methods suffer from the drawbacks of either being costly and/or might generate secondary pollutants, which shows toxicity than the parent ones [9,10]. Accordingly, biological methods are generally preferred for being more economical and environmentally friendly [11,12]. Bioremediation provides an alternative to chemical treatments. Bioremediation uses naturally occurring microorganisms to degrade various types of wastes. Contaminants are often potential energy sources for microorganisms. Microorganisms survive in contaminated habitat because they are metabolically capable of utilizing its resources and therefore highly suitable for reduction of pollutant load of an effluent as microorganisms are capable of oxidizing the organic and inorganic constituents [13,14]. Thus, the present study was aimed to isolate and identify bacteria from the sewage effluent that could produce extracellular protease enzyme to enhance bioremediation process. 2. Materials and methods 2.1 Sampling Sewage water samples were collected aseptically in a pre- sterilized bottle from municipal area of Jagaddal, North 24 parganas, West Bengal and transported immediately to the laboratory. 2.2 Isolation, characterization and identification of protease producing bacteria Heterotrophic bacteria were isolated by dilution plate technique on nutrient agar. The bacterial colonies were

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isolated by repeated streaking on fresh nutrient agar medium at room temperature and maintained at 4°C on slants of nutrient agar containing 1% gelatine, which act as an inducer for production of proteolytic enzymes. Identification of protease producing bacterial strains was performed by morphological, biochemical and physiological features comparing with standard references [15]. Physiological, morphological and biochemical tests of the bacterial isolates were carried out for their identification as per the procedures outlined in Bergey’s Manual of Systemic Bacteriology [16]. 2.3 Study of biodegradation potential The biodegradation potentials of isolated bacterial strains were determined using the method proposed by Kumari et al [17]. The collected water samples were separately inoculated on sterile nutrient agar plates under aseptic condition. The obtained culture is further sub-cultured by streak plate method. The inoculum was taken from the pure culture grown on nutrient agar slants and inoculated into each test tube containing 5 ml of nutrient broth. The tubes

were then incubated at 37°C for 24 h. 5 ml of this uniform suspension of each strain was inoculated as initial inoculum into each 1000 ml Erlenmeyer flask containing 500 ml of sterilized sample. Samples were incubated for 2, 4, 6, 8 and 10 days under room temperature. After desired incubation period the samples were analyzed for their physico- chemical parameters, total carbohydrate and protein using standard methods [18,19,20]. 3. Results and discussion Physiological, morphological and biochemical characterization of isolated bacterial strains were presented in Table 1. Based on these characters, the bacterial isolates were identified as Micrococcus luteus and Staphylococcus aureus though molecular analysis based on 16S rRNA gene sequencing are necessary for final characterization. Maximum carbohydrate (30%) and protein (36%) was found to be removed by Micrococcus luteus (SML-1) on 2 days, followed by SML-4 (carbohydrate- 28% and protein- 33%) and SML-5 (carbohydrate- 24% and protein- 28%) (Figure 1 and 2).

SML-5 (carbohydrate- 24% and protein- 28%) (Figure 1 and 2). Figure 1: Biodegradation potential of Micrococcus

Figure 1: Biodegradation potential of Micrococcus luteus (SML-1, SML-4, SML-5) and Staphylococcus aureus (SSA-1) in carbohydrate removal.

and Staphylococcus aureus (SSA-1) in carbohydrate removal. Figure 2: Biodegradation potential of Micrococcus luteus

Figure 2: Biodegradation potential of Micrococcus luteus (SML-1, SML-4, SML-5) and Staphylococcus aureus (SSA-1) in protein removal.

Waste water containing high BOD and COD are responsible for a heavy depletion of oxygen levels in the particular sector of the water bodies [21,22]. Therefore, these effluents needed elimination of BOD and COD through proper treatment methods before discharge. Maximal removal of COD (14%) by SML-1 on 4 days and 13% COD removed by SML-4 on 4 days were noted.

Staphylococcus aureus (SSA-1) showed maximum removal of BOD (24%) on 6 days, while this bacterium removed maximum carbohydrate (22%) on 6 days, protein (18%) on 2 days and COD (11%) on 6 days (Figure 3 and 4). High BOD and COD reduction in the treatment of different effluents by bacterial species was reported [23,24,25]. Bioremediation is a microorganism mediated transforma- -tion or degradation of contaminants into nonhazardousor

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Figure 3: Biodegradation potential of Micrococcus luteus (SML-1, SML-4, SML-5) and Staphylococcus aureus (SSA-1) in

Figure 3: Biodegradation potential of Micrococcus luteus (SML-1, SML-4, SML-5) and Staphylococcus aureus (SSA-1) in BOD removal

SML-5) and Staphylococcus aureus (SSA-1) in BOD removal Figure 4: Biodegradation potential of Micrococcus luteus

Figure 4: Biodegradation potential of Micrococcus luteus (SML-1, SML-4, SML-5) and Staphylococcus aureus (SSA-1) in COD removal

Table 1: Physiological, morphological and biochemical characterization of isolated bacterial strains

   

Bacterial isolates

 

Test

Micrococcus luteus

Micrococcus luteus

Micrococcus luteus

Staphylococcus aureus

SML-1

SML-4

SML-5

SSA-1

 

Colony characteristics

Form

Circular

Circular

Circular

Circular

Elevation

Convex

Convex

Convex

Raised

Margin

Entire

Entire

Entire

Entire

Colour

Yellow

Yellow

Yellow

White

Cell shape

Coccus

Coccus

Coccus

Coccus

Gram reaction

+

+

+

+

Coagulase

-

-

-

+

Catalase

+

+

+

+

Oxidase

+

+

+

-

Urease

+

+

+

+

 

IMViC test

Indole

-

-

-

-

Methyl red

-

-

-

+

Voges-

       

Proskauer

-

-

-

-

Citrate

+

+

+

-

H 2 S production

-

-

-

-

NO 3 - reduction

-

-

-

+

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International Journal of Environmental Biology 2014; 4(2): 107-111

 

Gelatine

         

liquefaction

+

+

+

+

 

Starch hydrolysis

 

-

-

-

-

 

Casein hydrolysis

 

+

+

+

 
 

Lipid hydrolysis

 

-

-

-

+

 

Hugh-Leiffson

 

O

O

O

O

(O/F) reaction

   

Utilization of carbon source

 
 

Glucose

-

-

-

+

 

Fructose

+

+

+

+

 

Sucrose

-

-

-

+

 

Lactose

-

-

-

+

 

Maltose

-

-

-

+

 

Mannose

+

+

+

+

 

Trehalose

+

+

+

+

 

Raffinose

-

-

-

-

 

Cellobiose

-

-

-

-

 

Xylose

-

-

-

-

 

Galactose

-

-

-

+

 

Ribose

-

-

-

+

 

Mannitol

-

-

-

+

 

Sorbitol

-

-

-

+

+

indicates presence or positive reaction; - indicates absence or negative reaction;

 

O

= Oxidation; F= Fermentation

 

less-hazardous substances. Bioremediation converts the organic matter into CO 2 and water. Microorganisms are capable of producing a wide variety of enzymes that can degrade complex organic compounds [26, 27]. The nature of the break down products depends upon both the microorganisms present and the environmental circumstances [28]. There are reports of bioremediation of industrial and domestic waste effluents by microorganisms [29]. The mechanism underline biological treatment is the decomposition of finely dispersed matter, colloidal and dissolved substances by metabolism of aerobic microorganisms. In sewage treatment, the additions of selected microorganisms improve the treatment efficiency of BOD, COD, detergent, oil and grease [30, 31]. The present study indicates the application of Micrococcus and Staphylococcus to removed contaminants from sewage water that is continually polluting the river water. 4. Conclusion The results of this study revealed that the Micrococcus luteus (SML-1, SML-4, SML-5) and Staphylococcus aureus (SSA-1), isolated from the sewage effluent are efficient enough to degrade the components in the sewage and therefore could effectively be used for the treatment of sewage water. This biodegradation study certainly will be helpful to some extent for making a pollution free environment. Acknowledgement The authors are thankful to University of Kalyani, West Bengal, India for providing necessary facilities for doing this research. Authors acknowledge the financial support received under the grant from DST PURSE, New Delhi, India for this study. References 1. P.S. Moschella, R.P.W.M. Laane, S. Back, H. Behrendt, G. Bendoricchio, S. Georgiou, P. M. J.

Herman, H. Lindeboom, M.S. Skourtous, P. Tett, M. Voss, W. Windhorst, Group report: methodologies to support implementation of the water framework directive, In: J.E. Vermaat, L.M. Bouwer, W. Salomons, R.K. Turner (eds), Managing European coasts: past, present and future. Berlin, 2005, pp.

137152.

2. L. Guardabassi, A. Dalsgaard, Occurrence and fate of antibiotic resistant bacteria in sewage, Danish Environmental Protection Agency, Danish Ministry of the Environment. Environmental Project No. 722, 2002, pp. 1-59.

3. H.A. Elliott, Land application of municipal sewage sludge, J. Soil Water Conserv. 41(1) (1986) 5-10.

4. A.K. Agarwal, G.S. Rajwar, Physico-chemical and microbiological study of Tehri Dam Reservoir Garhwal Himalaya, India. J. Am. Sci. 6(6) (2010) 65-

71.

5. R.M. Atlas, R. Bartha, Microbial Ecology:

Fundamentals and Applications, Addison-Wesley Publishing Company, 1981, pp. 560.

6. K.V. Ramana, L. Singh, Microbial degradation of organic wastes at low temperatures Defence Sci. J. 50(4) (2000) 371-382.

7. G. Tchobanoglous, F.L. Burton, H.D. Stensel, Wastewater Engineering: Treatment and Reuse, 4 th ed., Metcalf & Eddy Inc., New York, 2003.

8. S.S. Adav, D.J. Lee, N.Q. Ren, Biodegradation of pyridine using aerobic granules in the presence of phenol, Water Res. 41(13) (2007) 2903-2910.

9. H. Kurihara, Water quality of reusing waste water, J. Jpn. Sewage Works Assoc. 27 (1990) 38-41.

10. D. Mazumder, B. Roy, Low cost options for treatment and reuse of municipal wastewater, Indian J. Environ. Prot. 20(7) (2000) 529-532.

110

International Journal of Environmental Biology 2014; 4(2): 107-111

11.

G.M. Evans, J.C. Furlong, Environmental

T.O. Sogbanmu, A.A. Otitoloju, Efficacy and

effluents and their possible impacts on quality of

Biotechnology: Theory and Application, John Wiley

underground water, Soil Environ. 25(1) (2006) 64-69.

12.

& Sons, West Sussex, England, 2003.

23. A. Kumar, B.S. Bisht, V.D. Joshi, Bioremediation potential of five distinctive microorganisms to

24. S. Shruthi, M.P. Raghavendra, H.S. Swarna Smitha,

bioremediation enhancement potential of four dispersants approved for oil spill control in Nigeria, J. Bioremed. Biodegrad. 3(2) (2012) 1-5.

effluent of municipality waste, The IUP J. Life Sci. 5(1) (2011) 69-76.

13.

D.R. Lovley, Cleaning up with genomics: applying

K.

Girish, Bioremediation of rubber processing

molecular biology to bioremediation Nature Rev. 1(1) (2003) 3544.

industry effluent by Pseudomonas sp., Int. J. Res. Environ. Sci. Technol. 2(2) (2012) 27-30.

14.

T. Subramani, D. Haribalaji, Biodegradation of tannery effluent and designing the reactor for clarifier and activated sludge process, Int. J. Mod. Eng. Res. 2(3) (2012) 774-781.

25. A.A Telke, S.W. Kim, S.P. Govindwar, Significant reduction in toxicity, BOD, and COD of textile dyes and textile industry effluent by a novel bacterium Pseudomonas sp. LBC1, Folia Microbiol (Praha),

15.

C.H. Collins, P.M. Lyne, Microbiological methods,

57(2) (2012) 115-122.

Butterworth and Co., London, 1980.

26. L. D. Manh, Bioremediation of vegetable oil and

16.

J.G. Holt, N.R. Kreig, P.H. Sneath, J.T. Staley, S.T. Williams, Bergey's Manual of Determinative Bacteriology, 9 th edition, Williams and Wilkins, Baltimore, USA, 1994.

grease from polluted wastewater in dairy factory, VNU J. Sci. Nat. Sci. Technol. 24 (2008) 56-62.

27. T. Verma, V. Baiswar, Isolation and characterization of extracellular thermoalkaline protease producing

28. T. Schenk, R. Muller, F. Lingens, Mechanism of

29. H. Li, P. Li, T. Hua, Y. Zhang, X. Xiong, Z. Gong,

30. K.K. Chin, S.S. Ong, L.H. Poh, H.L. Kway, Waste

17.

S.

Kumari, A. Mehta, S. Shukla, J. John, P. Mehta,

Bacillus cereus isolated from tannery effluent, Int. J.

Biodegradation potential of the selected bacterial strains isolated from Sagar Lake, Asian J. Exp. Sci.

22(1) (2008) 43-48.

Eng. Sci. 2(7) (2013) 23-29.

enzymatic dehalogenation of pentachlorophenol by

18.

O.H. Lowry, N.J. Rosebrough, A.L. Farr, R.J. Randall, Protein measurement with the folin phenol reagent, J. Biol. Chem. 193 (1951) 265275.

Arthrobacter sp. strain ATCC 33790, J. Bacteriol. 172(12) (1990) 7272-7274.

19.

M.

DuBois, K.A. Gilles, J.K. Hamilton, P.A. Rebers,

Bioremediation of contaminated surface water by

F.

Smith, Colorimetric method for determination of

immobilized Micrococcus Roseus, Environ. Technol.

20.

sugars and related substances, Anal. Chem. 28(3) (1956) 350-356.

APHA, Standard Methods for the Examination of

26(8) (2005) 931939.

water treatment with bacterial augmentation, Water

Water and Waste Water, 21 st edition, American Public Health Association, Washington DC, 2005.

Sci.

Technol. 33(8) (1996) 17-22.

31. P. Dhall, R. Kumar, A. Kumar, Biodegradation of

21.

A.

Kumar, Environmental chemistry, Wiley Eastern

sewage wastewater using autochthonous bacteria, The

Limited, New Delhi, India, 1989.

Scientific World J. Article ID 861903 (2012) 1-8.

22.

M. Tariq, M Ali, Z. Shah, characteristics of industrial

Source of support: Nil; Conflict of interest: None declared

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International Journal of Environmental Biology 2014; 4(2): 107-111