Microbial mediated decolorization of textile dye: A bioremediation approach

Introduction

Rapid industrialization has substantially contributed to economic development and employment generation worldwide. However, it has also led to the discharge of large quantities of industrial effluents into the environment. Among various industries, the textile industry is recognized as one of the major contributors to environmental pollution due to extensive use of synthetic dyes and chemicals during dyeing, printing and finishing processes (Sudarshan et al., 2023) [14] . Approximately 80% of the dyes produced globally are utilized in the textile industry and nearly 10-30% of these dyes are released into the environment through wastewater generated during processing. The discharge of this untreated dye-containing effluent poses serious threats to the soil and aquatic ecosystem. Textile effluents are characterized by high chemical oxygen demand (COD), biological oxygen demand (BOD), and suspended solids (Bhatia et al., 2018) [4] . Azo dyes constitute the largest and widely used class of textile dyes. These dyes are characterized by one or more azo bonds linked to aromatic structures that confer high color intensity and chemical stability (Ranga et al., 2019) [12] . Furthermore, under reductive conditions, azo dyes can be transformed into aromatic amines that are toxic, mutagenic and potentially carcinogenic, posing serious ecological and public health risks (Aquino et al., 2014, Khatri et al., 2018) [2,7] . The environmental and agricultural impact of azo dye contamination have received increasing attention in recent years. The discharge of dye-containing effluents into agricultural land alters the physicochemical properties of soil by affecting porosity, nutrient availability and microbial diversity. The intense coloration of contaminated water bodies reduces light penetration, thereby affecting photosynthetic activity in aquatic ecosystems ((Hassan & Carr, 2018) [6] . In agricultural ecosystems, azo dyes can interfere with soil microbial communities responsible for nutrient cycling and organic matter decomposition ultimately leading to reduced soil fertility. Such disturbances often result in poor seed germination, reduced root and shoot growth and decreased crop productivity. To mitigate the adverse effects of textile dye pollution, several physicochemical treatment methods have been developed and implemented as well (Robinson et al., 2001). These methods include coagulation, flocculation, adsorption, membrane filtration and oxidation processes (Mishra and Maiti 2018) [11,13] . Although these technologies can effectively remove color from waste water, they are often associated with significant limitations such as high operational and maintenance costs, generation of secondary sludge, reduced effectiveness against structurally complex dyes etc. These limitations emphasize the need for more sustainable, economical and environmentally compatible treatment approaches. Biological remediation has emerged as a promising alternative for the treatment of dye-contaminated environments. Microorganisms possess diverse metabolic activities that enable them to transform and detoxify complex pollutants. Among various microbial groups, bacteria have attracted considerable attention because of their rapid growth rates, adaptability to diverse environmental conditions and ability to produce extracellular and intracellular enzymes involved in dye degradation (Das et al., 2023) [5] . Enzymes such as laccases, peroxidases, reductases and oxygenases play a crucial role in breaking azo bonds and in degrading aromatic intermediates into less toxic compounds. In addition to their biodegradation potential, several bacterial species possess plant growth promoting attributes including phosphate solubilization, zinc solubilization, phytohormone production etc. (Mathur et al., 2023) [10] Thus, bacterial bioremediation offers a dual advantage of environmental detoxification and enhancement of agricultural sustainability (Bera & Tank, 2021) [3] . In view of these concerns, the present study was undertaken to isolate and characterize indigenous bacteria from textile dye contaminated soil and to evaluate their potential for the textile dye as well as effluent decolourization.

Materials and methods

Sample collection and Bacterial Isolation

Soil and effluent samples contaminated with textile dye were collected from textile industrial sites in Panipat. The collected samples were brought to laboratory and processed for bacterial isolation. For the isolation of dye-tolerant bacteria, serial dilutions of the samples were prepared and inoculated onto mineral salt medium (MSM) supplemented with coractive blue dye (10mg/L). The inoculated plates were incubated under favourable growth conditions. The bacterial colonies exhibiting growth on the dye-containing medium were considered dye-tolerant and were selected for further studies. Pure cultures were obtained by repeated streaking on MSM plates.

Screening of bacterial isolates for dye decolourization

Bacterial isolates were screened for their ability to decolorize the dye at different concentrations (10, 20, 30 and 40 mg L⁻¹). The decolorization efficiency was assessed by measuring the absorbance of the dye solution at of dye by measuring absorbance at 620 nm using UV-Visible Spectrophotometer. Samples were collected at regular intervals, and the percent decolorization was calculated using the following formula: Decolorization (%): (A0 -At) / A0 × 100 where (A0) is the initial absorbance of the dye solution and (At) is the absorbance at time (t)

Decolorization of textile effluent by selected bacterial isolates:

The decolorization potential of selected bacterial isolates was evaluated using textile effluent at different concentrations (50, 75 and 100%). Each isolate was inoculated into the effluent containing medium and incubated under suitable conditions. Decolorization efficiency was determined by measuring the reduction in absorbance of the effluent at 620 nm using UV-Visible Spectrophotometer. The percentage decolorization was calculated using the standard formula.

Optimization of culture conditions (temperature, pH and aeration) for decolorization of textile effluent by potent bacterial isolates:

The promising bacterial isolates, selected on the basis of decolourization performance, were subjected to optimization studies to determine the effect of various culture conditions on textile effluent decolorization. The effect of temperature (25, 30, 35 and 40 °C), pH (6, 7, 8, 9, and 10) and aeration (static and shaking) was studied using 50% effluent. Decolorization efficiency was assessed spectrophotometrically. The optimum conditions were determined based upon the maximum percentage decolorization achieved.

Phytotoxicity analysis

Phytotoxic studies of untreated and bacterial treated effluent was evaluated using seeds of on seed of Mungbean and Chickpea. Healthy and surface sterilized seeds were used for the study. Seed germination percentage was recorded after incubation (24-72 h). The phytotoxic effect of the effluent was assessed by comparing the germination of treated and untreated effluent with that of control.

Effect of treated as well as untreated textile effluent on wheat crop under pot-house conditions

A pot-house experiment was conducted to evaluate the effect of treated as well as untreated textile effluent o the growth and performance of wheat (Triticum aestivum L.). the experiment consisted of seven as shown in Table 1.

Statistical analysis

The experimental data was analysed by the application of completely randomised design (CRD) using Online Statistical Analysis Tools (OPSTAT) software.

Results and discussion

Soil and effluent samples contaminated with textile dye collected from textile industrial sites in Panipat were used for isolation of bacteria. 34 bacterial isolates were retrieved from six samples (soil, sludge and effluent) collected from different sites (Table 2). Primary screening of these isolates was done at 10mg/L of dye on the basis of change in colour (Fig 22 .). Secondary screening using different concentrations (10, 20, 30 and 40 mg/L) of coractive blue dye was performed for selected isolates obtained after primary screening (Table 3 andFig Dry Soil (white) Dumping 7 6.

Textile Effluent ETP (Industry) 3 The table 4 presents the percentage decolourization achieved by different bacterial isolates (TCB4, TCB8, CB11, TCB17, TCB21, TCB26, TCB30, and TCB33) over a period of 24, 48, 72, and 96 hours at three different concentrations (10, 20, and 30 mg L⁻¹). The results are expressed as mean ± standard deviation, indicating the reproducibility of the experiments. At the lowest concentration (10 mg L⁻¹), all isolates exhibited an increase in decolourization efficiency with increasing incubation time (Fig 3). Among the isolates, TCB17 demonstrated the highest decolourization activity, increasing from 20.7% at 24 h to 44.5% at 96 h. Similarly, TCB8 showed 40.7% decolourization after 96 h. TCB30 also showed decolourization, 38.0% at 96 h. When the concentration was increased to 20 mg L⁻¹ (Fig 4 ), the decolourization efficiency of all isolates decreased compared with the 10 mg L⁻¹ treatment. Several isolates showed no measurable decolourization at 24 h (represented by "-"), suggesting an adaptation or lag phase before active degradation. Thus, increasing the dye concentration reduced the decolourization capability of the isolates.At the highest concentration (30 mg L⁻¹), decolourization efficiencies declined significantly, indicating an inhibitory effect of the higher dye concentration on microbial activity. Textile effluent (75%) was decolorized up to 16, 28 and 17% by isolates TCB8, TCB17 and TCB30 respectively. Potent bacterial isolate TCB17 showed maximum decolorization (36%) of textile effluent (50%) at optimized temperature 35 0 C and pH 8.0 (Fig. 6) under static conditions (Fig. 7) after 120h of incubation. Untreated (Raw) textile effluent showed 8% and 12% germination of chickpea and mungbean seeds whereas treated effluent (100%) showed 15% and 21% germination of chickpea and mungbean after 72h, respectively (Table 5).

Textile effluent (50%) treated with TCB 17 showed 40%, and 62% seed germination in chickpea and mungbean respectively. The effect of treated and untreated effluent on the seed germination characteristics of chickpea and mungbean was studied. The parameters evaluated include germination percentage, plumule length, and radicle length (Table 6). It has been observed that treated effluent had a significantly more favorable effect on seed germination and early seedling growth than untreated effluent. In chickpea, seeds irrigated with treated effluent showed germination percentage of 40.2%. Similarly, the plumule length increased from 0.24 mm under untreated effluent to 2.01 mm with treated effluent. The radicle length also increased from 0.51 mm in untreated effluent to 2.36 mm following treatment. These findings suggest that the treatment process effectively reduced the phytotoxic effects of the effluent, thus, promoting better seed germination. A similar trend was observed in mungbean, which showed greater tolerance and response to treated effluent. The germination percentage was 62.6% under treated effluent, compared to only 24.4% with untreated effluent. The plumule length increased from 1.21 mm in untreated effluent to 3.07 mm with treated effluent, while the radicle length increased from 1.34 mm to 4.17 mm, respectively. Treated effluent significantly enhanced germination percentage, plumule growth, and radicle development in both chickpea and mungbean compared with untreated effluent. Among the two crops, mungbean exhibited higher germination and better seedling growth than chickpea under both treated and untreated conditions, suggesting greater tolerance to residual contaminants. The results demonstrate microbial treatment of the textile effluent resulted in improved plant height, biomass accumulation, and grain production as compared to untreated effluent. Furthermore, dilution of the treated effluent (50%) proved to be the most suitable among the effluent treatments, suggesting that a combination of biological treatment and appropriate dilution can reduce the harmful impact of textile wastewater on wheat cultivation. These findings highlight the importance of treating textile effluent before its agricultural reuse and indicate that treated effluent, particularly at lower concentrations, has greater potential for safe irrigation than untreated wastewater.

Conclusion

The research findings highlight that the indigenous bacterial isolate TCB17 proves to be a potential candidate for ecofriendly bioremediation of textile effluent. The study highlights the potential of bacterial treatment as a sustainable and cost-effective alternative to the conventional methods for reducing dye pollution and facilitating the safe reuse of textile wastewater in agriculture.

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