Optimization of textile effluent bacterial treatment and improvement of the process efficiency through SPB1 biosurfactant addition

Basically, in textile industries, 15% of the total textile dye employed for desizing, scouring, dyeing, printing and finishing remains un-reacted during the dyeing process in the textile industries. They are directly lost in the effluents which generate huge amounts of wastewater begetting great economic and environmental damage (Chockalingam et al. 2019; Lellis et al. 2019). These wastewaters, called textile effluent may involve a wide variety of compounds including inorganic elements especially heavy metals, polymers and organic products principally hydrocarbons, synthetic dyes and pigments (Chockalingam et al. 2019; Sghaier et al. 2019).

Notably, textile wastewaters are mainly composed of synthetic dyes. Belonging to their chemical nature, they are classified into azo, diazo, basic, acidic, disperse, reactive, metal-complex and anthraquinone-based dyes (Chockalingam et al. 2019; Sghaier et al. 2019). They are well recognized for their non-biodegradability as well as ecological toxicity (Khan & Malik 2018; Sghaier et al. 2019). They can trigger numerous direct and indirect noxious effects like the formation of tumors, cancers and allergies (Akhtar et al. 2018). In addition, they hindered the growth of plants algae, protozoan, bacteria and animals (Akhtar et al. 2018; Khan & Malik 2018; Lellis et al. 2019; Sghaier et al. 2019; Ardila-Leal et al. 2021; Al-Tohamy et al. 2022). Therefore, owing to their toxicity, several research works have been particularly oriented toward their treatment and elimination from the contaminated environment.

Numerous physicochemical methods were developed like adsorption on activated carbon, catalyzed chemical oxidation with hydrogen peroxide and coagulation–flocculation (Donkadokula et al. 2020; Jaafari et al. 2020). However, they remain quite expensive and can release toxic byproducts. Recently, biological methods including the use of microbial strain to decolorize dyes have been proven to be optimal replacements for physicochemical procedures. As no chemicals are invested, they are qualified as environmentally friendly and aroused much concern from both scientists and industrialists (Jamee & Siddique 2019; Sghaier et al. 2019). Applying microbial strain for color removal (CR) is qualified as an environmentally and economically sustainable approach. In this regard, numerous bacterial strains belonging mainly to Bacillus, Pseudomonas, Serratia, Acinetobacter and Aeromonas genera were investigated owing to their capacity to remove dyes (Zabłocka-Godlewska et al. 2018; Jamee & Siddique 2019; Sghaier et al. 2019). Aerobic and anaerobic processes were also discussed for dyes decolorization (Jamee & Siddique 2019; Sghaier et al. 2019; Shoukat et al. 2019). Furthermore, the simultaneous investment of numerous microbial strains offers multiple merits over the application of pure cultures in terms of the decolorization of dyes as they allow conjugal degradation (Sghaier et al. 2019; Al-Tohamy et al. 2022; Irshad et al. 2023).

However, the wide variety of dyes, the complexity of their chemical structures and their low solubility limit their bioavailability to microbes and therefore their susceptibility to biodegradation. In this line, surfactant additions may improve dyes solubility and therefore their biodecolorization (Rehman et al. 2020; Noor et al. 2022). Nevertheless, the high toxicity of surfactants towards aquatic and terrestrial organisms reduces the scope of their application in environmental technology (Han & Jung 2020; Kaczerewska et al. 2020). Microbial-derived surfactants can be the optimal substitutes.

It is to be noted that, during their growth on hydrophilic or hydrophobic substrates, various microorganisms synthesize secondary active metabolites called biosurfactants (BioS) (Mnif & Ghribi 2015a). Structurally, being composed of a hydrophobic tail (saturated or unsaturated fatty acids) and a hydrophilic head (polysaccharides, amino acids and peptides), these molecules had amphiphilic character (Mnif & Ghribi 2015a). Owing to the biochemical nature of the hydrophilic part, BioS are classified into six groups: glycolipids, lipopeptides, phospholipids, lipopolysaccharides, neutral lipids and polymeric surfactants (Mnif & Ghribi 2015a). Lipopeptides are perceived amongst the best-recognized BioS with a prominent structural variability and multiple functional activities. Structurally, they comprise a combination of a peptide moiety and a fatty acid chain. They represent a group of isoforms that vary in terms of the length of the fatty acid chain, the composition of the peptide moiety and the association between both parts (Mnif & Ghribi 2015b, 2015c). Fengycin, Iturin and Surfactin are considered among the most known lipopeptides BioS (Mnif & Ghribi 2015b, 2015c). BioS are treasured for their surface activity features, particularly their ability to decrease the surface and interfacial tension on the surface and interface, respectively. Furthermore, they are qualified by various functions, for example, the dispersing, foaming, emulsification/de-emulsification, solubilizing, mobilizing, viscosity reducers and pore-forming capacities. Additionally, they are talented in numerous biological activities including the antimicrobial, hemolytic, antiviral and antioxidant capacities enabling their application in multiple fields (Mnif & Ghribi 2015a; Nikolova & Gutierrez 2021). Furthermore, they are characterized by higher environmental biocompatibility, biodegradability, non-toxicity and higher performance upon severe salinity, pH and temperature (Mnif & Ghribi 2015a; Nikolova & Gutierrez 2021). Therefore, BioS are remarkably advantageous over chemical surfactants since they provide powerful opportunities as efficient and ecological substitutes (Mnif & Ghribi 2015a; Nikolova & Gutierrez 2021). From this perspective, they are proven to be potential adjuvant in cosmetics, food and pharmaceutical industry. Moreover, they are interesting candidates for the improvement of pollutant removal and for heavy metals sequestration in bioremediation domain (Mnif & Ghribi 2015a, 2015b). In this regard, a lipopeptide BioS derived from B. subtilis SPB1 was largely applied as an enhancer of diesel biodegradation (Mnif et al. 2014, 2015a, 2017) and dyes biodecolorization (Mnif et al. 2015b, 2015c, 2016). It consists of a mixture of Surfactin, Itruin and Fengycin isoforms that decrease the surface tension (ST) of water to about 34 mN/m with a critical micelle concentration (CMC) of 150 mg/L (Bouassida et al. 2018). It exhibits numerous biological activities including antimicrobial activities against multi-drug resistant bacteria and plants pathogenic fungi, insecticidal activity and anti-diabetic and anti-lipidemic capacities in alloxan-induced diabetic rats with low in vivo toxicity against male mice (Bouassida et al. 2018). Moreover, it is used as a best substitute of a synthetic emulsifier for food (Mnif et al. 2013) and detergent formulation (Bouassida et al. 2018).

In view of the importance of B. subtilis, SPB1 derived lipopeptides which whetted the widest concern and drew extensive attention and in order to broaden their wide spectrum of applications, we attempted in this research study to evaluate their potency to enhance textile effluent biotreatment. Therefore, we shall investigate the ability of a newly isolated bacterial consortium to decolorize textile effluent. Response surface methodology (RSM) would be applied as a statistical tool to enhance the biodecolorization rate. The three operating parameters would be optimized by a Box–Bhenken design including the initial pH value, the consortia density and the BioS percentage. CR and chemical oxygen demand (COD) decrease when monitored during the biotreatment. They were optimized by the RSM. Moreover, the mode of action of both selected bacteria was explored. This study was carried out in Tunisia 2 years ago.

Two bacterial strains, isolated through the enrichment technique from textile wastewater, were invested during this study for the treatment of textile effluent. They correspond to Citrobacter sedlakii RI11 (KJ865880) (Mnif et al. 2015b) and Aeromonas veronii GRI (KF964486) (Mnif et al. 2016).

Equal amounts of both selected strains were used for inoculation of the textile effluents during the treatment experiments. The cell pellets for the inoculation were prepared as described by Mnif et al. (2013, 2016). The treatment medium was incubated at 37 °C under static conditions. They were prepared according to the experiments illustrated in Table 2. Treatment efficiency was assessed by measuring the decreasing percentages of color intensity and COD of the effluent in the end of cultivation after been centrifuged at 10,000 rpm/min for 10 min.

A bacterial BioS derived from B. subtilis SPB1 (HQ392822) was investigated to enhance the biodecolorization efficiency. It was produced and extracted as portrayed in our previous work (Mnif et al. 2021a, 2021b, 2021c).

To optimize the decolorization process, a Box–Bhenken design was applied followed by the analysis of the results using the RSM. Hence, three operating parameters, namely pH, inoculum density and BioS concentration, were optimized. Biodecolorization assays were incubated at 37 °C for 3 days under static conditions. Each variable was evaluated at three coded levels (−1, 0 and +1) (Table 1). A total of 16 assays were performed (Table 1). The response values (̂y) recorded for each experiment correspond to the mean of the duplicates (Table 1). NemrodW Version 2007 software (LPRAI, Marseille, France) served to generate the design and for data analysis.

Student's test was conducted to assess the significance of the different coefficients. Subsequently, two-dimensional graphical representations as well as response surface charts were illustrated to examine the correlation between the individual parameters and to define the optimal conditions.

With a view to illuminate the mode of action of bacterial decolorization of the textile effluent, CR assays were applied to handling living and dead cells as well as the bacterial supernatant, as clarified in our prior research (Mnif et al. 2015b, 2015c, 2016). The two bacterial strains were grown separately in LB medium overnight at 37 °C and 150 rpm. To eradicate the remaining cells, the supernatant was filter sterilized using 0.2 μm membranes after centrifugation (10,000 rpm, 20 min). It served as extracellular enzyme preparation to estimate enzymatic dye biodegradation. Obtained biomass was suspended in 50 mM potassium phosphate buffer (pH 7.4) and was partitioned into two parts. The first part served as active cells and the second part served as an inert bio-sorbent material after being autoclaved. Both served to quantify CR after the implementation of the decolorization assay. All assays were conducted in duplicate with and without SPB1 lipopeptide BioS supplementation.

The studied effluent was gathered from a textile mill at the point of their discharge and stocked at 4 °C for 1 week at maximum until further use. In order to characterize the wastewater, multiple physicochemical parameters were analyzed. As enumerated in Table 1, they include the color intensity, pH, electrical conductivity (EC), dry matter (DM) content, dry volatile matter content (DVM), total solid material (TSM), total suspended solids (TSS), as well as the total volatile suspended solids (TVSS). Besides, we determine the biological oxygen demand after 5 days (BOD₅), COD, total Kjeldahl nitrogen and lipids content. They were determined as described by a standard method for the analysis of water and wastewater (APHA 1998). Results are summarized in Table 2.

Basically, the effluent characteristics need to be adequately monitored to explore the possibility of their discharge into the environment and to specify the best treatment method. From this perspective, in order to assess textile effluent wastewater properties and its biodegradability, physicochemical characterization was performed. As plotted in Table 2, the COD and BOD₅ levels of the effluent were of about 2,085 and 1,000 mg O₂/L, respectively. Its pH is about 8.5. All these characteristics are generally well above the discharge limit provided by Tunisian Standard (NT 106.02; 1989) (pH between 6.5 and 9; COD = 1,000 mg/L and BOD₅ = 400 mg O₂/L).

The ratio of the BOD₅/COD obtained departing from the results of about 0.48 near 0.5 indicated that the effluent contains a proportion of non-biodegradable organic matter but it can be classified as biodegradable (Makki & Khudhair 2018). In addition, this effluent presents high levels of DSM and nitrogen content exceeding the limits presented by the Tunisian standard (NT 106,02, 1989) (DSM = 400 mg/L; total nitrogen = 50 mg/L).

To conclude, the physicochemical characteristics of the collected textile effluent sample revealed a high load of pollution indicators. Moreover, the color of the effluent was black owing to a mixture of dyes invested during the dyeing procedure. Notably, dissolved compounds including dyes and pigments increase the color darkness of the polluted residue.

In order to improve textile effluent treatment through the selected consortia composed of both strains Aeromonas veronii GRI and Citrobacter sedlakii RI11, the experimental planning methodology was applied. According to previous reports and studies, three factors were optimized using a Box–Bhenken design (initial pH of the effluent, total inoculum size and BioS concentration). As reported in our previous work tackling orange methyl biodecolorization, initial pH value variability brings about a variation in the bacterial activity and consequently in its growth rate along with its CR performance (Mnif et al. 2015b, 2016). Indeed, bacteria were active over a particular scope of pH. Additionally, the inoculums size can have a great effect on the bioprocess efficiency. It was demonstrated that an initial inoculum density of about 0.5 supports the maximal decolorization of Methyl Orange by Aeromonas veronii GRI (Mnif et al. 2016). Accordingly, the level of SPB1 lipopeptide BioS can have a great impact on the biotreatment, as demonstrated in our previous published works handling Methyl Orange, Congo Red and Methyl Green biodecolorization (Mnif et al. 2015b, 2015c, 2016), as well as hydrocarbon biodegradation (Mnif et al. 2015a, 2017).

The sixteen experiments of the Box–Bhenken design onward with the respective predicted and observed responses are outlined in Table 1. Results stand for the average of two independent assays. Box–Bhenken design experiments revealed a wide variation in both responses, namely decolorization percentage quantified by the % of CR (39.13–93.78%) and COD abatement (41.58–86.31%). This variation reflected the adequacy of the selected variables and their levels. Furthermore, it ensured the significance of the optimization strategy to achieve higher responses. Responses were recorded after 3 days of aerobic incubation at static conditions.

In order to check the statistical significance of the regression equation, we applied the Fischer's test. Regarding the obtained results, ANOVA analysis elaborated in Table 3 indicated that both regression models were significant and the lack of fit was insignificant, suggesting that they are effective in terms of predicting responses. The fit of the model was assessed by specifying the coefficient of correlation R² displaying values of 0.948 and 0.978 for the decolorization % and COD abatement, respectively. As the value of R² was close to 1, the model would justify highly the variability between the experimental and the model predicted values (Mnif et al. 2021c). Adjusted R² values were about 0.807 and 0.879 for both studied responses reflecting an adequate adjustment of the quadratic model to the experimental data (Jaafari & Yaghmaeian 2019; Mnif et al. 2021c).

In order to evaluate the significance of each coefficient, a Student-test was carried out. Table 4 exhibits the Students’ distribution; the corresponding parameter estimate as well as p-value. The p-value allows the verification of the importance of each of the coefficients. Observed results are indicative that the three selected parameters affected significantly both responses. As far as the double and mutual interactions among the three factors are concerned, it is noteworthy that they are significant only for the decolorization increase.

In order to assess the performance of the bioprocess, the phytotoxicity of the textile effluent after treatment was evaluated (Mahmood et al. 2015; Mnif et al. 2015b, 2015c, 2016; Ayed et al. 2019). In this respect, the rise of seed germination was suggestive of the abolishment of a toxic effect of the effluent. Hence, phytotoxicity tests using different seeds, namely tomatoes, watercress, watermelon and lucerne revealed the efficiency of the treatment of the textile effluent. Treatment was conducted at the optimized conditions determined by the RSM study: pH 5 with an initial optical density of the newly selected bacterial consortium of about 0.4 and with the addition of 0.01% SPB1 BioS. At the same time, phytotoxicity was evaluated for textile effluent treated with Aeromonas veronii GRI and Citrobacter sedlakii RI11 at pH 5 without the addition of SPB1 BioS. Moreover, negative and positive controls were undertaken with water and non-treated textile effluent, respectively.

Generally, two different phenomena can occur during dye decolorization, either biodegradation of dyes compounds or biosorption namely the microbial accumulation of chemicals (Ngo & Tischler 2022; Pinheiro et al. 2022). Biodegradation involves enzymatic activities allowing the bioconversion of dyes to elemental compounds and/or to less toxic compounds (Ngo & Tischler 2022; Pinheiro et al. 2022). The biosorption basically occurs within the cell wall (Pinheiro et al. 2022). It differs according to the biomass type.

Thus, in order to elucidate the action mode of both strains during textile effluent treatment, we opted for handling effluent with each strain separately. Moreover, treatments were performed through the use of bacterial supernatant as well as living and dead cells. Results summarized in Table 5 indicate a high decolorization of the textile effluent when using the extracellular bacterial supernatant with a decolorization rate % near those obtained by living cells. This finding suggests that culture supernatant of the strains contains active enzymes in dye decolorization. Obtained results are not consistent with those reported in our previous work describing Congo Red removal by adsorption to the bacterial cells. In fact, a slight decolorization rate % was recorded by adsorption to dead cells, which ensures the occurrence of biodegradation phenomena of textile dyes by different enzymatic activities (Mnif et al. 2015b). Similarly, previous works revealed the occurrence of enzymatic biodegradation of Methyl Orange by Aeromonas veronii GRI (Mnif et al. 2016) and Malachite Green by Citrobacter sedlakii RI11 (Mnif et al. 2015c). In fact, different enzymes proved to be implicated in the biodegradation process of synthetic dyes. Among the most prominent ones, we state Reductase, Lignine Peroxidase, Manganese Peroxidase, Laccase and Tyrosinase (Jamee & Siddique 2019; Sghaier et al. 2019; Ngo & Tischler 2022; Pinheiro et al. 2022).

For organic dyes removal increase, surfactants and/or BioS can act according to three different ways, namely dyes solubilization, increasing the bacterial membrane permeability and enhancing the cell surface hydrophobicity. Regarding the first hypothesis, surfactants have the ability to solubilize dyes in aqueous media by incorporating them into their micelles structures (Kaczorek et al. 2018; Noor et al. 2022; Irshad et al. 2023). This can promote their bioavailability to biodegrading microorganisms and then their digestion. As for the second hypothesis, the enhancement of bacterial membrane permeability stimulates enzyme secretion and/or organic dyes diffusion via a liquefied membrane (Kaczorek et al. 2018). Concerning the third hypothesis, the increase of cell surface hydrophobicity promotes pollutants adhesion, facilitating, therefore, their diffusion to intracellular compartment or enhances their partitioning and bioavailability to microorganisms (Kaczorek et al. 2018).

In order to better and deeper explore the mechanism of action of SPB1 BioS in the course of the textile effluent treatment; we opted for studying the biodecolorization efficiency using intracellular and extracellular bacterial supernatant as well as living cells. The lipopeptide BioS was incorporated at its optimal level of about 0.01%, as determined in the first part of the work. Thus, we inferred that SPB1 BioS always improved biodecolorization efficiency in the different cases for both studied strains (Tables 6 and 7). Hence, it can be assumed that SPB1 BioS acts at three different levels to ameliorate biodegradation efficiency:

• - Either they can improve dyes solubility by their encapsulation into the hydrophobic core of the BioS micelles, increasing, therefore, their bioavailability to bacterial cells

• - Or they can improve the cell membrane hydrophobicity, facilitating dyes adhesion to membrane and biodegradation at the surface and/or diffusion to intracellular compartment and biodegradation

• - Or they can improve cells permeability facilitating accordingly the diffusion of dyes to the intracellular compartment for their assimilation by intracellular enzymes and/or facilitating extracellular enzymes secretion

• - Or they can activate the enzymes secretion and activities implicated in dye biodegradation.

Our obtained results go in good accordance with those obtained in our previous works reporting the biodecolorization of Malachite Green and Methyl Orange (Mnif et al. 2015b, 2016). SPB1 BioS addition permits a significant increase in CR. In this regard, Jadhav et al. (2011) confirmed the enhancement of the biodecolorization of a synthetic dye; Brown 3REL by Pseudomonas desmolyticum with the addition of 0.1% Rhamnolipid BioS. Moreover, the enhancement of Methyl Orange biotreatment by Aeromonas veronii GRI with the addition of 0.01% SPB1 BioS was correlated in our previous work (Mnif et al. 2016). However, for Malachite Green and Congo Red biodecolorization, 0.075% of SPB1 BioS addition improves significantly dyes treatment by Citrobacter sedlakii RI11 (Mnif et al. 2015b) and B. weihenstephanensis RI12 (Mnif et al. 2015c), respectively. Along the same line, prior works proved the relevance of SPB1 BioS in terms of the enhancement of diesel biodegradation in water and soil by diverse bacterial strains at 0.1% (Mnif et al. 2015a, 2017). Additionally, in situ BioS production improved diesel biodegradation by B. subtilis SPB1 (Mnif et al. 2014). Within this framework, previous studies demonstrated that chemical surfactants improve greatly hydrophobic dyes solubility, enhancing therefore their bioavailability for microorganisms and stimulating consequently their biodegradation (Noor et al. 2022; Irshad et al. 2023).

Moreover, the impact of pH on the decolorization effectiveness has been also addressed in previous studies. Basically, a moderate neutral pH supports dyes biodecolorization (Mnif et al. 2015b, 2016; Zin et al. 2020; Srivastava et al. 2022). However, as observed by Srivastava et al. (2022), bacterial strains were metabolically active and could efficiently decolorize dyes at a pH range of 6–9. For certain other species, they displayed optimum decolorization at both acidic and basic pH values as outlined by Coria-Oriundo et al. (2021). Other strains showed interesting biodecolorization efficiency at extreme basic pH values (Ayed et al. 2019). Regarding the biotreatment efficiency of the textile effluent quantified in the present work using CR and COD abatement measurement, the obtained results proved to be efficient and relevant as correlated by Ayed et al. (2020), Sur & Mukhopadhyay (2017) and Samuchiwal et al. (2021).

Notably, in situ BioS production and ex situ BioS addition enhance hydrophobic compounds biotreatment by improving their solubility, the surface area of contact and enzyme release. Other studies highlighted the improvement of enzymes activities by chemical surfactants addition into the reacting medium (Liu et al. 2017; Oliva-Taravilla et al. 2020; Wang et al. 2020). On the other side, having a dual hydrophobic-hydrophilic character, BioS can improve bacterial membrane permeability and facilitate therefore enzymes secretion (Jadhav et al. 2011; Kaczorek et al. 2018; Zdarta et al. 2020). According to Parthipan et al. (2017), BioS production by B. subtilis A1 was in correlation to the uptake of available hydrophobic substrates. Indeed, the produced BioS increase the substrates solubility, enhancing their digestion. During the degradation of crude oil, the cationic groups of the BioS captivate the negatively charged bacterial membrane increasing therefore the surface area of interaction (Parthipan et al. 2017). In fact, changes in cell wall structure or the nature of released extracellular substances can result in contact with surface-active compounds (Kaczorek et al. 2018). As they have the ability to alter significantly the cell surface biomorphology, hydrophobicity, surface functional groups as well as the electrokinetic potential, BioS may influence cells surface properties (Kaczorek et al. 2018). Thus, the modification of cell surface characteristics promotes their adhesiveness to pollutants or improves their partitioning and biological availability to microbes (Kaczorek et al. 2018). Furthermore, surfactant affects considerably the phospholipid membrane of the microbial cells. In fact, as argued by Otzen (2017), in vitro explorations revealed that these BioS highly accommodate phospholipid membranes by inducing leakage of phosphatidylcholine (POPC) vesicles dehydrating the membrane surface as well as making it more fluid. This is basically assisted through powerful interrelation between BioS and phospholipid molecules (Otzen 2017). Additionally, the penetration of surfactant molecules can lead to changes in cell membrane permeability increasing, therefore, the membrane permeability. As a matter of fact, the liquefied membrane may help biodegrading enzymes as well as other metabolites to escape from the cell (Kaczorek et al. 2018). All these factors can improve pollutants biotreatment through enzymatic biodegradation and/or biosorption.

Textile industry can be classified among the most polluting industries as it generates huge amounts of wastewaters composed of diverse organic and inorganic compounds commonly synthetic dyes, pigments, hydrocarbons and heavy metals (Ardila-Leal et al. 2021; Al-Tohamy et al. 2022). Being non-biodegradable and having the properties to bio-accumulate in living organisms, they beget serious problems of pollution and toxicity and disturb the ecological equilibrium (Akhtar et al. 2018; Ayed et al. 2019; Chockalingam et al. 2019; Lellis et al. 2019; Ardila-Leal et al. 2021; Al-Tohamy et al. 2022). From this perspective, a crucial demand for their environmental detoxification appeared. Recently, biological treatment methods using microorganisms appeared as the best alternative to physicochemical methods referring to their easy processing as well as their low cost (Pinheiro et al. 2022). They involved the use of microbial strains having the capability to decolorize dye and/or adsorb heavy metal either in pure culture or in consortia called bioremediation (Zabłocka-Godlewska et al. 2018; Ayed et al. 2019; Jamee & Siddique 2019). Basically, biological treatment or that of textile effluent allows the bioconversion of dyeing matters into biodegradable and less toxic compounds in contrast to physicochemical methods which generate byproducts (Jadhav et al. 2015; Khan & Malik 2018; Ayed et al. 2019). It is noteworthy that investing microbial consortia offered multiple and great advantages over the application of pure cultures in the scope of synthetic dyes degradation. Mixed microbial cultures have been applied extensively for raw-colored wastewater decolorization. In fact, owing to synergistic metabolic activities, mixed cultures display a great performance than pure cultures (Zabłocka-Godlewska et al. 2018; Ayed et al. 2019; Jamee & Siddique 2019). Thus, the association of different microorganisms helps in terms of the biodegradation and accelerates the treatment process (Zabłocka-Godlewska et al. 2018; Ayed et al. 2019; Jamee & Siddique 2019; Sghaier et al. 2019). Two hypotheses can corroborate the effectiveness and goodness of fit of mixed bacterial culture over pure one. Firstly, dye molecules can be attacked by individual strains at different positions. Secondly, in mixed processes, a degradation metabolite of a first strain can be utilized by a second strain promoting the dye metabolism.

Accordingly, in order to enhance the biodecolorization efficiency and ameliorate the treatment potency of textile wastewater, a consortium was constructed relying upon two newly identified strains having the ability to decolorize synthetic dyes. In order to polish the treatment process, biodecolorization efficiency was optimized using Box–Bhenken design. In addition, SPB1 BioS was evaluated as ameliorating agent of CR. In this regard, numerous research works have emphasized the impact of surfactants in terms of enhancing the apparent solubility of certain organic compounds and hydrocarbons and therefore increasing their digestion by microorganisms. Besides, chemical surfactants addition has been largely described as enhancers of the solubility and digestion of synthetic dyes (Lamichhane et al. 2017). However, having a chemical nature and being non-biodegradable, their addition could trigger toxicity to the environment as well as living organisms (Han & Jung 2020; Kaczerewska et al. 2020). Being biodegradable and non-toxic, microbial-derived emulsifiers or BioS, can be an optimal candidate for petrochemical derived surfactants in terms of ameliorating the bioavailability of hydrophobic contaminants since their application witnessed a blooming growth, recorded mainly in the industrial area.

In the present research work, an optimization of textile effluent biotreatment by two selected strains Aeromonas veronii GRI and Citrobacter sedlakii RI11 was undertaken. Physicochemical analyses of the explored effluent revealed an alkaline pH with a high content of suspended materials and COD and BOD₅ values of about 2,085 and 1,000 mg/L, respectively. A Box–Bhenken design allows the selection of the optimal operating conditions yielding a maximum of COD abatement and decolorization increase the efficiency of about 86 and 94%, respectively. This occurs particularly when inoculating the textile effluent at pH 5 with an initial optical density of 0.4 and with the addition of 0.01% SPB1 BioS. The evaluation of the action mode of both selected bacteria during textile effluent treatment revealed slight rates of decolorization by adsorption on dead cells. This finding suggests that enzymatic degradation of dyes is more important than biosorption. The investigation of the level of action of SPB1 BioS during textile effluent biotreatment allowed the supposing that SPB1 BioS can increase dyes solubility and consequently their bioavailability to bacterial cells. It can also increase bacterial cells permeability facilitating, therefore, dye diffusion in the intracellular compartment for their assimilation and/or for the activation of biodecolorization enzymes.

All procedures performed in studies involving animals were in accordance with the ethical standards of the institution or practice at which the studies were conducted.

Informed consent was obtained from all individual participants included in the study.

All the authors give the publisher the permission to publish the work.

The first author of the present manuscript Dr Inès Mnif realized and writes this paper. The second author Dr Mouna Bouassida helped in the redaction of the paper. Doctor Lamya Ayed and Professor Dhouha Ghribi helped in the elaboration of the plan of this work and corrected this paper.

The work is funded by the Ministry of Higher Education and Scientific Research.

All materials are available.

All relevant data are included in the paper.

The authors declare there is no conflict.