Raising globalization, urbanization and rapid development of industrialization have been causes different kinds of environmental pollutions.Among various industries, the textile dying industries discharge large volume of waste water after dyeing process (Zollinger, 1987). It is estimated that around 10 -15% of the dyes are lost in the effluent during the dyeing processes (Kadiret al., 2009). Dyes are classified to the basis of chemical structure of chromophore there are 20 -30 different groups of dyes like azo (monoazo, diazo, triazo, polyazo),anthraquinone, phthalocyanine and triarylmethane dyes are the most important groups (Chen et al., 2011). The majority of industrial important azo dyes belong to the following classes: acid dyes, basic dyes, direct dyes, Disperse dyes, mordant dyes, reactive dyes and solvent dyes (Wanyonyi et al., 2017).Dyes make up an abundant class of organic compounds characterized by the presence of unsaturated groups (chromophores) such as –C=C-, ─N═N─ and ─CH═, which are responsible for colors (Molinariet al., 2004).
Around 10,000 different dyes with an annual production of more than 7 ×105 metric tons worldwide are commercially available and are extensively used in textile processing, paper, printing, pharmaceutical, food and other industries (McMullan et al., 2001; Siddeeg et al., 2020). The textile effluents containing of various toxic substances namely suspended detergents, surfactants, carcinogenic amines, formaldehyde and organochloride pesticides associated with dyes (Jadhavet al.,2010). During the fabrication process approximately 10% of the dyes have not fixed with fibers, it released into the environment (Asadet al.,2007).The release of untreated effluents from the dyeing industries causes a major threat to the environment and reduces sun light penetration of the stream. These effluents are toxic to fish and mammalian life and to inhibit the activity and growth of microorganisms. It alters the pH, increases the biological oxygen demand (BOD) and chemical oxygen demand (COD) and gives the rivers intense colorations. It also affects the soil fertility and plant growth (Afrad et al., 2020). In humans the azo dyes are capable of producing intestinal cancers, cerebral and skeletal abnormalities in foetuses(Saha and Rao, 2020). Moreover, disposal of textile waste water on an open land contaminates the subsoil water, so that drinking water gives color as well as bad taste. Dissolved substances in industrial effluents alter the chemical and biological status of the soil and water, which may affect growth and productivity of plants and alters the color and quality of the water bodies has been proved to be hazardous to aquatic organisms (Lade et al., 2015). Toxic compounds from dye effluent get into aquatic organisms and ultimately reach man through food chain to cause various physiological disorders like hypertension, skin cancer, sporadic fever, renal damage and cramps etc. They possess toxicity like lethal effect, genotoxicity, mutagenicity, and carcinogenicity to plants and (Puvaneswariet al.,2006).
There are several physical and chemical treatment methods are used for dyes like adsorption, sedimentation, floatation, coagulation, polymerization, flocculation, reverse osmosis, ultra-filtration, ionization, radiation, reduction, oxidation, electrolysis and ion exchange are extensively used. These methods are not environmentally friendly and produces enormous amount of secondary pollutants and are cost-effective (Roy et al., 2018).Biological treatment is often the most economical alternative compared to physical and chemical processes. Biological methods like stabilization, aerated lagoons, trickling filter, activated sludge, anaerobic digestion and bio-augmentation etc. The biological methods have lower cost but it having large land spaceandlesseffective(Sheam et al., 2020).The removal of textile carcinogenic pollutants is the major task of the bioremediation. The ability of microorganisms to decolorize and metabolize dyes has long been known and the use of bioremediation based technologies for treating textile waste water has attracted interest (McMullanet al., 2001). Several microorganisms have been found to decolorize and mineralize azo dyes. Many microorganisms belonging to different taxonomic group of bacteria, fungi, actinomycetes and algae have been reported for their ability to decolourizeazo dyes (parmar, 2014; Hassan etal.,2013;Zabinet al., 2011,). Biodegradation systems of color removal through the use of bacteria have been shown to be highly effective.
The bacterial metabolism of azo dyes is initiated by a reductive cleavage of the azo bond in most cases, which results in the formation of amines. Azoreductase enzymes responsible for decolourization of azo dyes are purified from several bacterial strains. These reductive processes have been studied in some aerobic bacteria, which grow on azo compounds. Bioreactor is a engineered device, usually a vessel, used to direct the activity of a biological catalyst to achieve a desired chemical transformation. Basic principles of process control, fermentation monitoring, dissolved oxygen, PH,temperature, monitoring, subtract (Glucose). In current research aimed to focus on isolation and characterization of effluent decoloring bacteria from the textile effluent wastewater. Two bacterial strains were screened for potential degradation of effluents industrial dye, after that 15 days incubation. The decolorized by-product was performed phytotoxic study that revealed toxicity assessment of the end by-product.
- Materials and methods
2.1. Sample collection
Textile dye effluent waste water was collected from in Edapadi, Salem district, Tamil Nadu, India. Effluent was collected using sterile container and aseptically transported to the laboratory and stored at 4ºC for further use.
2.2. Isolation of dye degrading bacteria
Collected effluent was mixed with sterile 100mL Bushnell Hass broth and incubated at 37ºC for 7 days. 1 mL of the above broth sample was serially diluted and spread on nutrient agar plates for the isolation of dye degrading bacterial strains and incubated at 37ºC for 24 h. Afterthatincubation period, fifteen bacterial strains were isolated and stored at 4ºC. The selected bacterial strains were noted as KADB01 to KADB15.
2.3. Identification of bacteria strain
The isolated bacterial strains were identified by following staining and biochemical methods of Gram staining, spore staining, motility, oxidase, indole, methyl red voges-prosjauer, citrate utilization, urease and carbohydrate fermentation test. The results further were compared with manual of systematic bacteriology.
2.4. Screening of dye degrading bacterial strain
The isolated bacterial strains were screening for the selection of efficient dye degradation activity. In briefly described, LB agar medium was prepared and directly added into the different concentration of azo dye solution under the ambient temperature and properly mixed. After that process immediately poured into the petri plates and allowed to solidification. Then respective isolated bacterial strains were streaked on the plates and incubated at 37ºC for 24 h. The efficiency of bacterial dye degradations were observed based on the zone of inhibition (Kabeer et al., 2019).
2.5. Bioreactor study
Bioreactor is an engineered device, usually a vessel, used to direct the activity of a biological catalyst to achieve a desired chemical transformation. Bioreactor of 4L capacity was designed for lab scale study for the treatment of dye contaminated water. The bioreactor was filled with 4L of the dye contaminate water to which 1% of bacterial consortium was added. Temperature was maintained at room temperature and continuously aerated with constant agitation using stirrer at 250 rpm. The samples were collected fromvarious parameters such as pH, CFU and Optical density was measured by UV spectrophotometer. The percentage of dye degradation was calculated by following formula.
% degradation = Initial absorbance value-final absorbance value/Initial absorbance valuex100
2.6. Phytotoxicity assay
The soil was taken in a 250ml of sterile plastic cups and 4 Vignaradiata seeds were placed into each cup at 1.5 cm depth. All the cups were irrigated regularly with 5 ml of untreated as negative control and bioreactor treated dye effluent. The seeds were allowed to germinate and seed germination was assessed on every day of the experiment. At the end of 10th day shoot length and size of the leave was measured. Control set was carried out using distilled water to replace the textile dye effluent at same time.
- Results and discussion
Industrial dye contaminated water sample was collectedand physicochemical parameters analyzed such as color, oder, PH,
temperature, OD value were observed and shown in Table 1. The dye degrading bacterial strains were isolated from dye effluent sample by using spread plate technique. The totally fifteen bacterial strains were isolated and biochemical characterized, the results were revealed in the Table 2. Similar report revealed,
textilewastewater showed abnormal coloration of black turquoise blue colored. The pH determined the collected test sample was slightly acidic to neutral (Bhatia et al., 2018).
3.1. Screening of dye degradation bacteria
Luria Bertani Agar medium incorporated with azo dye and the bacterial isolates were inoculated. The maximum zone of clearance of the isolatedstrains were taken as an indicator of the dye degradation bacteria its shows in Table 3, the colony size and zone of clearing size were measured in millimeters. Several researchers have been reported, similar results as related to zone of clearing on the agar plate, its indication of the dye degradation ability of the selected bacteria (Shazia et al., 2017).
3.2. Construction of consortium
The Bacillussp and Pseudomonassp were used for the construction of consortium due to the synergistic effects with each other.
3.3. Dye decolourization
The dye decolourization by the bacterial consortium was detected by the following optical density value of the bioreactor
Table 4 Dye effluent decolourization measurement by UV spectrophotometer
|UV Spectroscopic OD values of the treated sample
treated dye effluent shows in Table4.Our bacterial consortium degraded 38% percentages of dye within fifteen days of treatment. Similar research was conducted by Lalnunhlimi et al.,(2016) for the decolorization of mixture dyes using of Bacillus cereus has showed 93.37% of highest decolorization within 5 days. Another related study was reported byKarim et al. (2018), the decolorization experiment using of five frequently employed textile azo dyesby different bacterial isolates strains of EK13 (41%), EK7 (41%) and EK6 (42%) for incubation after 6 days in Bushnell-Haas (BH) broth culture medium (Lalnunhlimi et al., 2016).
The microbial treated dye effluents was applied to study the phytotoxicity of green gram (Vigna radiate) plant such as germination capability, shoot length and root length. The result was compared with control tape water it’s shown in Table 5.In comparison of similar research reported by Nouren et al., 2017, using of yellow 4 dye by-product for phytotoxicity experiment. Zea mays seeds treated with degraded by-product of test samples and untreated DY4 dye samples. The germination percentage was calculated based on the root and shoot lengths of germinated seeds were compared with control samples. The result suggested 50% of negatively affected then compared to treated samples, which is confirmed exhibit degradation revealed in the test samples (Karim et al., 2018).
The textile industries are produced large amount of wastewater effluents which hazardous to ecosystem, because of their higher concentration of dyes mixtures. Although bacterial decolorization or degradation of textile effluent is a challenging process, but nowadays many researchers have been reported microbial mediated degradation of textile effluent. In the current research study, consortium of two bacterial strains such as Bacillussp and Pseudomonassp were potential effluent decolorization was screened through the plate assay method. The consortium of two bacterial strains was observed maximum growth at 37°C and pH 7.0. The construction of consortium with two bacterial isolates is able to decolor of industrial effluent dye up to 38% for 15 days incubation. The seeds germination was revealed less toxic effect in treated samples and also enhancing of plant growth. These results suggested the potential applicable of these two bacterial strains for the treatment of wastewater in the future.
Funding: The authors received no specific funding for this work.
Conflicts of Interest: None
- Zollinger, H. Synthesis, Properties and Application of Organic Dye and Pigments. Color Chemistry, VCH New York, 1987 pp.92-102.
- Kadir, T., Zuhal, T. Decolorization of direct dye in textile wastewater by ozonization in a semi-batch bubble column reactor. Desalination, 242(2009) 256-263.
- Chen, Z., Jin, X., Chen, Z., Megharaj, M., Naidu, R. Removal of methyl orange from aqueous solution using bentonite-supported nanoscale zero-valent iron. J. Colloid Interface Sci. 361 (2011) 601–663.
- Wanyonyi, W. C., Onyari, J. M., Shiundu, P. M., Mulaa, F. J. Biodegradation and detoxification of malachite green dye using novel enzymes from bacillus cereus strain km201428: kinetic and metabolite analysis. Energy Procedia 119 (2017) 38–51.
- Molinari, R., Pirillo, F., Falco, M., Loddo, V., Palmisano, L. Photocatalytic degradation of dyes by using a membrane reactor. Chemical Engineering Process 43 (2004)1103-1114.
- McMullan, G., Meehan, C., Conneely, A., Kirby, N., Robinson, T., Nigam, P., Banat, I.M., Marchant, R., and Smyth, W.F. Microbial decolorization and degradation of textile dyes. Appl Microbial Biotechnol. 561 (2001) 81-87.
- Siddeeg, S. M., Tahoon, M. A., Mnif, W., Ben Rebah, F. Iron oxide/chitosan magnetic nanocomposite immobilized manganese peroxidase for decolorization of textile wastewater. Processes 8 (2020) 5.
- Jadhav, J. P., S. S. Phugare, R. S. Dhanve and S. B. Jadhav. Rapid biodegradation and decolorization of direct orange 39 (orange TGLL) by an isolated bacterium Pseudomonas aeruginosastrain BCH. Biodegradation, 21 (2010) 453–463.
- Asad, S., Amoosegear M. A., Pourbabaee, A. A., Sarboloukki, M. N., Dastgheib, M. Decolonization of textile azo dyes by newly isolation halophilic and halotolerant bacteria. BioresourTechnol 98 (2007) 2082-2088.
- Afrad, M. S. I., Monir, M. B., Haque, M. E., Barau, A., Haque, M. M. Impact of industrial effluent on water, soil and rice production in Bangladesh: a case of Turag river bank. J. Environ. Health Sci. Eng. 18(2020) https://doi.org/10.1007/s40201-020-00506-8.
- Saha, P., Rao, K. Biotransformation of Reactive Orange 16 by alkaliphilic bacterium Bacillus flexus VITSP6 and toxicity assessment of biotransformed metabolites. Int. J. Environ. Sci. Technol. 17 (2020) 99–114.
- Lade, H., Kadam, A., Paul, D., Govindwar, S. Biodegradation and detoxification of textile azo dyes by bacterial consortium under sequential microaerophilic/aerobic processes. EXCLI J. 14 (2015) 158. doi:10.17179/excli2014-642.
- Puvaneswari, N., Muthukrishnan, J., Gunasekaren, P. Toxicity assessment and microbial degradation of azo dyes. Indian Journal of Experimental ,vol.44 (2006)618-626.
- Roy, D. C., Sahan, A. K., Sikdar, B., Rahman, M., Roy, A. K., Prodhan, Z. H., Tang, S. S. Biodegradation of crystal violet dye by bacteria isolated from textile industry effluents. PeerJ 6 (2018) e5015
- Sheam, M., Haque, Z., Nain, Z. Towards the antimicrobial therapeutic and invasive properties of MikaniamicranthaKnuth : a brief overview. J. Adv. Biotechnol. Exp. Ther. 3 (2020) 92–101.
- Parmar, P. R. Decolorization of acridine red dye by the fungi Aspergillus species. Journal of Scientific and Innovative Research, 3(2014) 454-459.
- Hassan, M. M., Alam, M. Z. and Anwar, M. N. Biodegradation of textile Azo dyes In Effluent. International Research Journal of Biological Sciences, 2(2013):27-31.
- Zabin, K., Bagewadi, Amithkumar, G., Vernekar, Aishwarya, Y., Patil, Abhijit, A., Limaye, Vandana,M., Jain. Biodegradation of industrially important textile dyes by actinomycetes isolated from activated sludge. Research Article, Biotechnol. Bioinf .Bioeng, 1(2011):351-360.
- Kabeer, F. A., John, N., Abdulla, M. H. Biodegradation of malachite green by a newly isolated Bacillus vietnamensissp . MSB17 from continental slope of the Eastern Arabian Sea: enzyme analysis, degradation pathway and toxicity studies. Bioremediat. J. 0 (2019) 1–9. https://doi.org/10.1080/10889868.2019.1671790.
- Bhatia, D., Sharma, N.R., Kanwar, R., Singh, J. Physicochemical assessment of industrial textile effluents of Punjab (India), Appl. Water Sci 8, (2018) 83. https://doi.org/10.1007/s13201-018-0728-4.
- Shazia, N., Haq, N. B., Munawar, I., Ismat , B., Shagufta, K., and Yusra, S. By-product identification and phytotoxicity of biodegraded Direct Yellow 4 dye, Chemosphere, 167 (2017) 474-484.
- Lalnunhlimi, S., Krishnaswamy, V. Decolorization of azo dyes (Direct Blue 151 and Direct Red 31) by moderately alkaliphilic bacterial consortium, Brazilian Journal of Microbiology, 47 (2016) 39-46.
Karim, M.E., Dhar, K., Hossain, M.T.,.Decolorization of textile reactive dyes by
bacterial monoculture and consortium screened from textile dyeing effluent. J. Genet.
Eng. Biotechnol, 16 (2018) 375-380. doi:10.1016/j.jgeb.2018.02.005.