Free Suspended Cells Research Articles

Immobilized Microalgae using Alginate for Wastewater Treatment

Organic and inorganic substances are released into the environment because of domestic, agricultural, and industrial activities which contribute to the pollution of water bodies. Removal of these substances from wastewater using conventional treatment involves high energy cost for mechanical aeration to provide oxygen for aerobic digestion system. During this process, the aerobic bacteria rapidly consume the organic matter and convert it into single cell proteins, water, and carbon dioxide. Alternatively, this biological treatment step can be accomplished by growing microalgae in the wastewater. Chlorella vulgaris immobilized in calcium alginate was used to study the removal efficiency of main nutrients in wastewater such as ammonium and phosphate that act as an important factor in microalgae growth. The immobilized cells demonstrated higher percentage of ammonium and phosphate removal of 83% and 79% respectively, compared to free-suspended cells (76% and 56%). COD removal recorded was 89% and 83% for immobilized cells and free-suspended cells, respectively. The kinetics parameters of nutrients removal for immobilized C. vulgaris in synthetic wastewater were also determined. The specific ammonium removal rates (RA) and phosphate removal rates (RP) for Chlorella vulgaris in synthetic wastewater were 8.3 mg.L-1day-1 and 7.9 mg.L-1day-1, respectively. On the other hand, the kinetic coefficient for each nutrient removal determined were kA = 0.0462 L.mg-1 day-1 NH4 and kP = 0.0352 L.mg-1 day-1 PO43-. This study proves the application of immobilized microalgae cells is advantageous to the wastewater treatment efficiency. Furthermore, optimization on the immobilization process can be conducted to further improve the nutrients removal rates which potentially can be applied in the large-scale wastewater treatment process.

Enhanced bioremediation of crude oil polluted water by a hydrocarbon-degrading Bacillus strain immobilized on polyurethane foam

During the production and transportation of petroleum hydrocarbons, unsuitable operation and leakage may result in contamination of water and soil with petroleum hydrocarbons. Petroleum contamination causes significant marine environmental impacts and presents substantial hazards to human health. Bioremediation of contaminated water and soil is currently the effective and least harmful method of removing petroleum hydrocarbons from the environment. To improve the survival and retention of the bioremediation agents in the contaminated sites, microbial cells must be immobilized. It was demonstrated that immobilized microbial cells present advantages for degrading petroleum hydrocarbon pollutants compared to free suspended cells. In this study, the ability of a Bacillus strain (designed as Bacillus sp. VTVK15) to immobilize on PUF and to degrade crude oil was investigated. The immobilized Bacilllus strain had the highest number (5.38 ± 0.12 Í 108 CFU/g PUF) and a maximum attachment efficiency of 92% on PUF after 8 days. Analysis by GC-MS revealed that both free and immobilized cells of Bacillus sp. VTVK15 were able to degrade 65 and 90% of the hydrocarbons in 2% (v/v) crude oil tested after 14 days, respectively. The results suggest the potential of using PUF-immobilized Bacillus sp. VTVK15 to bioremediate petroleum hydrocarbons in an open marine environment.

Small bioreactor platform capsules provide persistent digestive biomass for continuous bioreactors operated under short hydraulic retention times

Bioaugmentation treatments in general and the use of encapsulated selected microbial cultures in particular are gaining significant attention as a promising approach for the treatment of wastewater containing phenols and other organic pollutants. Thus, the purpose of this study is to investigate the biodegradation performance of a Pseudomonas putida culture encapsulated in a unique 3-D capsule known as the small bioreactor platform (SBP). Batch and continuous bioreactors, bioaugmented with two different P. putida culture states, i.e., encapsulated and free suspended cells, were operated under different phenol loadings. During the batch experiments, the biodegradation rate of the suspended culture was equal to or higher than the encapsulated culture, except for the highest phenol concentration of 1000 mg/L, where the encapsulated bacterial culture exhibited a superior biodegradation rate (45 mg/L/h) relative to the free culture (16.7 mg/L/h). In addition, in the continuous bioreactor, at a hydraulic retention time (HRT) of up to 1 h, the encapsulated P. putida completely biodegraded the phenol in the influent with 50 mg/L phenol, showing a maximal biodegradation rate of 45 mg/L/h. However, decreasing the HRT to 0.5 and 0.25 h resulted in incomplete biodegradation, despite higher biodegradation rates (90 and 70 mg/L/h, respectively). Furthermore, a sharp decrease in the biodegradation process efficiency was observed when the suspended culture was released from the SBP capsules by cutting the microfiltration membrane of the capsules. Therefore, this study demonstrates that, compared with suspended cultures, SBP culture encapsulation achieves superior performance as a bioaugmentation strategy, especially in continuous bioreactors, such that it can be applied in the treatment of phenol rich wastewater.

A new Rhodococcus aetherivorans strain isolated from lubricant-contaminated soil as a prospective phenol-biodegrading agent

Microbe-based decontamination of phenol-polluted environments has significant advantages over physical and chemical approaches by being relatively cheaper and ensuring complete phenol degradation. There is a need to search for commercially prospective bacterial strains that are resistant to phenol and other co-pollutants, e.g. oil hydrocarbons, in contaminated environments, and able to carry out efficient phenol biodegradation at a variable range of concentrations. This research characterizes the phenol-biodegrading ability of a new actinobacteria strain isolated from a lubricant-contaminated soil environment. Phenotypic and phylogenetic analyses showed that the novel strain UCM Ac-603 belonged to the species Rhodococcus aetherivorans, and phenol degrading ability was quantitatively characterized for the first time. R. aetherivorans UCM Ac-603 tolerated and assimilated phenol (100% of supplied concentration) and various hydrocarbons (56.2–94.4%) as sole carbon sources. Additional nutrient supplementation was not required for degradation and this organism could grow at a phenol concentration of 500 mg L−1 without inhibition. Complete phenol assimilation occurred after 4 days at an initial concentration of 1750 mg L−1 for freely-suspended cells and at 2000 mg L−1 for vermiculite-immobilized cells: 99.9% assimilation of phenol was possible from a total concentration of 3000 mg L−1 supplied at daily fractional phenol additions of 750 mg L−1 over 4 days. In terms of phenol degradation rates, R. aetherivorans UCM Ac-602 showed efficient phenol degradation over a wide range of initial concentrations with the rates (e.g. 35.7 mg L−1 h−1 at 500 mg L−1 phenol, and 18.2 mg L−1 h−1 at 1750 mg L−1 phenol) significantly exceeding (1.2–5 times) reported data for almost all other phenol-assimilating bacteria. Such efficient phenol degradation ability compared to currently known strains and other beneficial characteristics of R. aetherivorans UCM Ac-602 suggest it is a promising candidate for bioremediation of phenol-contaminated environments.

The Potential of Using Grapefruit Peel as a Natural Support for Yeast Immobilization During Beer Fermentation

The potential use of grapefruit peel as support material for yeast immobilization during beer fermentation was evaluated. After conditioning, FTIR analysis revealed a higher quantity of methoxy (–OCH3) groups, suggesting that lignin is the major component of the support. Cell adhesion onto the conditioned support in 12°Plato laboratory malt wort was evaluated, observing a maximal cell adhesion (2.25 · 109 cells/gram of dried support) at 20 h of cultivation, remaining almost constant in the subsequent time points. Evaluations of the fermentative behaviour of the biocatalyst at 15±0.5 °C in a 14°Plato laboratory malt wort indicated good stability in terms of physical integrity (confirmed by SEM observation). The fermentation time was shortened to four days, and the rates of reducing sugar consumption and ethanol production were improved when compared to fermentations carried out with free suspended cells. These results show a promising potential of grapefruit peel as support material in beer fermentation.

Yeast Immobilization on Peanut Hulls for Potential Application in Beer Fermentation

The potential of using peanut hulls as support material for immobilization of yeast cells during beer fermentation was evaluated. After treatment of peanut hulls with 0.5% (w/v) NaOH, FTIR analysis revealed an important change on its chemical composition. For instance, the proportion of lignin decreased and as a result the adsorption of yeast cells onto the support material during fermentation was favoured. Cell adhesion onto the conditioned support in 8° Brix laboratory malt wort as fermentation medium was evaluated, observing a maximal cell adhesion (2.56×108 cells/g of dried support) at 20.5 h of cultivation. Evaluations of the performance of the biocatalyst in fermentations at 15±0.5°C in 14° Brix laboratory malt wort indicated a proper stability in terms of physical integrity (confirmed by SEM observation) and improved fermentation kinetics. The fermentation time was shortened to approximately four days and the rates of reducing sugar consumption and ethanol production were improved when compared to fermentations carried out with free suspended cells. These results show a promising potential of peanut hulls as support material in beer fermentation.

Semi-Continuous Reverse Membrane Bioreactor in Two-Stage Anaerobic Digestion of Citrus Waste.

The presence of an antimicrobial compound called D-Limonene in citrus waste inhibits methane production from such waste in anaerobic digestion. In this work, a two-stage anaerobic digestion method is developed using reverse membrane bioreactors (rMBRs) containing cells encased in hydrophilic membranes. The purpose of encasement is to retain a high cell concentration inside the bioreactor. The effectiveness of rMBRs in reducing cell washout is evaluated. Three different system configurations, comprising rMBRs, freely suspended cells (FCs), and a combination of both (abbreviated to rMBR–FCs), are incubated at three different organic loading rates (OLRs) each, namely 0.6, 1.2, and 3.6 g COD/(L cycle). Incubation lasts for eight feeding cycles at 55 °C. Methane yield and biogas composition results show that rMBRs perform better than rMBR–FCs and FCs at all three OLRs. Volatile fatty acid profiles and H2 production show that the reactors are working properly and no upset occurs. Additionally, a short digestion time of 4 days can be achieved using the rMBR configuration in this study.

Enhanced expression of catechol 1,2 dioxygenase gene in biofilm forming Pseudomonas mendocina EGD-AQ5 under increasing benzoate stress

In natural settings, microbes often associate closely with surfaces and interfaces called biofilms and adhere in an extracellular matrix. The biofilm provide diverse microenvironments and high resistance to environmental stress on the constituent cells that aid to generate cellular heterogeneity and phenotypic variability. The work presented here examines for the first time the capability of Pseudomonas mendocina EGD-AQ5 biofilm-associated cells, compared with that of planktonic cells to utilize toxic intermediate catechol under benzoate degradation conditions in liquid culture systems. Both planktonic and biofilm-associated cells utilized benzoate to the same extent but accumulated catechol differently. This differential catechol accumulation activity of the Pseudomonas mendocina EGD-AQ5 could be accredited to the cells adapted as a biofilm. Further experiments with transcriptome implicated to increased expression of genes like pelA (involved in biofilm formation) cat1,2D (ortho pathway) and decreased expression of cat2,3D (meta pathway) in biofilm harvested cells than free suspended cells. Whole genome sequence analysis of Pseudomonas mendocina EGD-AQ5 (Genbank Accession No AVQF00000000) deciphered catechol ortho and meta-pathways for benzoate utilization and pelABCDEFG as involved in biofilm formation. These results suggest that Pseudomonas mendocina EGD-AQ5 biofilms should facilitate bioaugmentation to be more resilient and efficient bioremediation technology than free suspended planktonic cells.

Biocarriers Improve Bioaugmentation Efficiency of a Rapid Sand Filter for the Treatment of 2,6-Dichlorobenzamide-Contaminated Drinking Water.

Aminobacter sp. MSH1 immobilized in an alginate matrix in porous stones was tested in a pilot system as an alternative inoculation strategy to the use of free suspended cells for biological removal of micropollutant concentrations of 2,6-dichlorobenzamide (BAM) in drinking water treatment plants (DWTPs). BAM removal rates and MSH1 cell numbers were recorded during operation and assessed with specific BAM degradation rates obtained in lab conditions using either freshly grown cells or starved cells to explain reactor performance. Both reactors inoculated with either suspended or immobilized cells showed immediate BAM removal under the threshold of 0.1 μg/L, but the duration of sufficient BAM removal was 2-fold (44 days) longer for immobilized cells. The longer sufficient BAM removal in case of immobilized cells compared to suspended cells was mainly explained by a lower initial loss of MSH1 cells at operational start due to volume replacement and shear. Overall loss of activity in the reactors though was due to starvation, and final removal rates did not differ between reactors inoculated with immobilized and suspended cells. Management of assimilable organic carbon, in addition to cell immobilization, appears crucial for guaranteeing long-term BAM degradation activity of MSH1 in DWTP units.

Bioremediation potential of Pseudomonas sp. isolated from a 2, 4, 6-trinitrotoluene contaminated site

Pseudomonas mosselii and Pseudomonas stutzeri isolated from TNT contaminated site were evaluated for their TNT degrading potential in aqueous phase. Free and immobilised cell cultures of P. mosselii and P. stutzeri were evaluated individually and in combination, by shake flask studies for degrading TNT in synthetic solutions and in actual waste waters respectively. All the flasks were incubated at temperature 37°C at 120 rpm for 48 hours in continuous mode. Biodegradation of TNT was monitored by carrying out qualitative and quantitative studies. Spectro-photometric measurements of the density of free suspended cells in the culture broth indicate the positive growth of the bacteria under toxic conditions of TNT contamination. The immobilised cells showed over all better performance as compared to free cells. The qualitative and quantitative analysis by HPLC revealed the 100% biodegradation of TNT to secondary metabolites of bacterial metabolism within 24 hours and 28 hours of incubation, by immobilised cells of P. mosselii in synthetic and in actual waste waters respectively.

Substrate interactions between 4-nitrophenol and 4-nitrotoluene during biodegradation of their mixture

Nitroaromatic compounds are toxic and rather recalcitrant pollutants of water and soil. Microbial degradation of the individual nitroaromatic compounds has already been well described in the literature. However, because several compounds often occur in the environment mutually influencing each other’s degradation, further research into the biodegradation of their mixtures is still needed. We investigated the degradation of a mixture of 4-nitrophenol (4-NP) and 4-nitrotoluene (4-NT) by a mixed microbial culture immobilized in a continuously operated packed-bed reactor (PBR) and by free suspended cells in shake flasks. Each compound was at first degraded separately, then in a mixture, and their degradation characteristics were compared. When treated separately, 4-NP and 4-NT were degraded with efficiency over 99 and 95%, respectively. When in a mixture, 4-NP was still completely removed from the media but the 4-NT removal efficiency dropped by a half and remained about the same during the whole experiment regardless the increasing 4-NP concentration at the PBR inlet. The shake flask experiments corroborated our finding that 4-NP negatively influenced 4-NT degradation by competitive inhibition and also indirectly on the adaptation level.

Biodegradation of tetradecane using Acinetobacter venetianus immobilized on bagasse

Immobilized cells present advantages for degrading organic pollutants compared to free suspended cells. However, the immobilization of cells with reference to agricultural waste is still limited. This study investigated using bagasse as a carrier to immobilize Acinetobacter venetianus (A. venetianus) for the degradation of n-alkane. The degradation of alkane based on both adsorption and biodegradation by immobilized A. venetianus was observed, where 93.3%, 77.7% and 24.0% of tetradecane (400mgL−1) were removed by the immobilized cells, free cells and bagasse after 36h incubation, respectively. Additionally, alkane (C10–C25) in diesel was effectively degraded by the immobilized cells, of which 800mgL−1 of alkane was completely degraded after 60h. Furthermore, results showed: (1) tetradecane served as the optimal carbon source at the concentration of 100mgL−1; and (2) cells immobilization reached its peak efficiency with the cells-to-bagasse ratio of 0.5mgcellg−1bagasse. Cells strongly immobilized on bagasse retained a high degradation rate of 77.6% after shaking at 1500rpm. Bagasse protected the cells from pollution stress including cadmium (Cd(II)) and phenanthrene. After 8 reuse cycles, immobilized cells became more stable and efficient. Bagasse-A. venetianus has potential for future application in remediating diesel contaminants.

Evaluation of different cell-immobilization strategies for simultaneous distillery wastewater treatment and electricity generation in microbial fuel cells

Cell-immobilization technology has long been considered as an important tool for microbial fuel cell (MFC) design. For the first time, a direct comparison between MFCs catalyzed by freely-suspended cells, surface-attached cells, and matrix-embedded cells for simultaneous electricity generation and distillery wastewater treatment in both batch and continuous operation modes was systematically performed. The results indicated that MFCs catalyzed by surface-attached cells and matrix-embedded cells had better chemical oxygen demand (COD) removal and power production efficiencies than those catalyzed by freely-suspended cells. The normalized energy recovery (NER), or power divided by the amount of COD consumed, was found to be decreasing with increasing COD concentration in all three different setups. Denaturing gradient gel electrophoresis (DGGE) analysis indicated that the structure of microbial community was influenced by the mode of operation, COD concentration and cell immobilization methods of the MFC. Nevertheless, surface-attached cells and matrix-embedded cells had a higher similarity than freely-suspended cells regardless of the mode of operation. The information obtained here will be an important reference for future design and application of MFCs for simultaneous wastewater treatment and power production.

Bioconversion of residual glycerol from biodiesel synthesis into 1,3-propanediol using immobilized cells of Klebsiella pneumoniae BLh-1

The bioconversion of residual glycerol obtained from biodiesel synthesis into 1,3-propanediol (1,3-PD) was carried out using Ca-alginate immobilized-cells cultures of Klebsiella pneumoniae BLh-1. The immobilization parameters bead size and cell concentration were determined for the best possible productivity and were found to be 3.4 mm and 100 mg mL−1, respectively. In 12 h-batch cultivations, immobilized cultures showed a 1,3-PD productivity of 1.85 g L−1 h−1, up from 1.22 g L−1 h−1 obtained in 16 h-batch cultivations using free-suspended cells. The possibility of reuse of the biocatalysts was tested in 5-cycles repeated batch cultures, with entrapped cells keeping their operational stability and biological activity, with 1,3-PD productivities as high as 4.48 g L−1 h−1. Other important metabolites such as 2,3-butanediol and ethanol were also produced (10.11 g L−1 and 3.71 g L−1, respectively) during the repeated batches. These results show the feasibility of using and further developing immobilized-cell systems of K. pneumoniae for the efficient bioprocessing of residual glycerol.

Biodegradation of nitroglycerin and ethylene glycol dinitrate by free and immobilized mixed cultures

Aerobic biodegradation of nitroglycerin (NG) and ethylene glycol dinitrate (EGDN), both as individual substrates and in their mixture, was tested using batch or fed-batch cultivation with free suspended cells enriched from a soil sample subjected to a long-term contamination with explosives. EGDN was degraded only in the presence of glycerol as a co-substrate whereas NG could serve as a sole carbon, energy and nitrogen source for growth, its degradation being only slightly boosted by either glycerol or pyruvate. NG was not sufficient as a co-substrate for microbial growth on EGDN; furthermore, the presence of EGDN inhibited the NG degradation. The growth inhibition by both NG and EGDN was alleviated by the addition of glycerol. At an optimum nitroglycerin concentration of 30 mg/L, a maximum specific degradation rate of 60.9 ± 1.8 mg/gdw/h was observed. The biodegradation of both pollutants occurred with a release of nitrite. A method was developed for growing substantial amounts of NG-degrading biomass in the presence of glycerol for its immobilization on expanded slate in a pot-scale packed-bed reactor. Preliminary reactor tests were conducted in a continuous operation mode yielding a 70–90% NG biodegradation up to a load of 20 mg/L/h, with a removal rate up to 16 mg/L/h.

Loofa (Luffa cylindrica) sponge: Review of development of the biomatrix as a tool for biotechnological applications

The review discusses the development of loofa sponge (Luffa cylindrica) as a biotechnological tool and the diversity of applications in which it has been successfully used since it was first reported as a matrix for the immobilization of microbiological cells in 1993. The fibro-vascular reticulated structure, made up of an open network of random lattices of small cross-sections coupled with very high porosity (79-93%), having very low density (0.02-0.04 g/cm(3) ), and high specific pore volume (21-29 cm(3) /g), has the characteristics of a carrier/scaffold well-suited for cell immobilization. This has been confirmed through the immobilization of cells of diverse types, including filamentous and microalgae, fungi, bacteria, yeasts, higher plants, and human and rat hepatocytes. The cells immobilized in loofa sponge have performed well and better than free suspended cells and those immobilized in conventionally used natural and synthetic polymeric materials for the production of ethanol, organic acids, enzymes, and secondary metabolites. The loofa-immobilized cell systems have been efficiently used for the treatment of wastewaters containing toxic metals, dyes, and chlorinated compounds, and the technology has been used to develop biofilms for the remediation of domestic and industrial wastewaters rich in inorganic and organic matter. In addition, three-dimensional loofa sponge scaffolds for hepatocyte culture have been suggested to have the potential for development into a bioartificial liver device. Loofa sponge is a cost-effective, eco-friendly, and easy to handle matrix that has been used successfully as a biotechnological tool in a variety of systems, purposes, and applications.

Early host-pathogen interactions in marine bivalves: Evidence that the alveolate parasite Perkinsus marinus infects through the oyster mantle during rejection of pseudofeces

Parasites have developed myriad strategies to reach and infect their specific hosts. One of the most common mechanisms for non-vector transmitted parasites to reach the internal host environment is by ingestion during feeding. In this study, we investigated the mechanisms of oyster host colonization by the alveolate Perkinsus marinus and focused on how oysters process infective waterborne P. marinus cells during feeding in order to determine the portal(s) of entry of this parasite to its host. We also compared the infectivity of freely-suspended cells of P. marinus with that of cells incorporated into marine aggregates to link changes in particle processing by the feeding organs with infection success and route. Finally, we evaluated the effect of oyster secretions (mucus) covering the feeding organs on P. marinus physiology because these host factors are involved in the processing of waterborne particles. The ensemble of results shows a unique mechanism for infection by which the parasite is mostly acquired during the feeding process, but not via ingestion. Rather, infection commonly occurs during the rejection of material as pseudofeces before reaching the mouth. The pseudofeces discharge area, a specialized area of the mantle where unwanted particles are accumulated for rejection as pseudofeces, showed significantly higher parasite loads than other host tissues including other parts of the mantle. Aggregated P. marinus cells caused significantly higher disease prevalence and infection intensities when compared to freely-suspended parasite cells. Mucus covering the mantle caused a quick and significant increase in parasite replication rates suggesting rapid impact on P. marinus physiology. A new model for P. marinus acquisition in oysters is proposed.

Aerobic Degradation of Nitrobenzene by Immobilization of <i>Streptomyces albidoflavus</i> in Polyurethane Foam

Streptomyces albidoflavus Z2 was immobilized in polyurethane foam and its ability to degrade nitrobenzene was investigated. Batch experiments showed that polyurethane-foam-immobilized cells (PFIC) possessed a higher degradation capacity. Compared to freely suspended cells (FSC), PFIC degraded nitrobenzene more efficiently under alkali and acid conditions, as well as higher temperature. Kinetic of nitrobenzene degradation by PFIC was well described by Andrews inhibition equation, which demonstrated that PFIC tolerated higher concentration of nitrobenzene than FSC and degraded nitrobenzene at a quicker speed. Moreover, in the presence of NaCl (≤7%, w/v), phenol (≤250 mg L-1) and aniline (100 mg L-1), respectively, PFIC exhibited better tolerance toward salinity and toxic chemicals than FSC. Therefore immobilization could be a promising method for treating nitrobenzene industrial wastewater.

Microbial Degradation of Nitrobenzene by Immobilized Cells of <i>Micrococcus luteus</i>

Micrococcus luteus Z3 was immobilized in polyurethane foam and its ability to degrade nitrobenzene was investigated. Batch experiments showed that polyurethane-foam-immobilized cells possessed a higher degradation capacity. Compared to freely suspended cells (FSC), immobilized cells degraded nitrobenzene more efficiently under alkali and acid conditions, as well as higher temperature. Kinetic studies revealed that higher concentrations of nitrobenzene were better tolerated and more quickly degraded by immobilized cells. Moreover, in the presence of NaCl (≤10%, w/v) immobilized cells exhibited better tolerance toward salinity. Therefore immobilization of Z3 in polyurethane foam could be a promising method for treating nitrobenzene industrial wastewater, especially with high salinity.

Lindane removal by pure and mixed cultures of immobilized actinobacteria

Lindane (γ-HCH) is an organochlorine insecticide that has been widely used in developing countries. It is known to persist in the environment and can cause serious health problems. One of the strategies adopted to remove lindane from the environment is bioremediation using microorganisms. Immobilized cells present advantages over free suspended cells, like their high degradation efficiency and protection against toxins. The aims of this work were: (1) To evaluate the ability of Streptomyces strains immobilized in four different matrices to remove lindane, (2) To select the support with optimum lindane removal by pure cultures, (3) To assay the selected support with consortia and (4) To evaluate the reusability of the immobilized cells. Four Streptomyces sp. strains had previously shown their ability to grow in the presence of lindane. Lindane removal by microorganisms immobilized was significantly higher than in free cells. Specifically immobilized cells in cloth sachets showed an improvement of around 25% in lindane removal compared to the abiotic control. Three strains showed significantly higher microbial growth when they were entrapped in silicone tubes. Strains immobilized in PVA-alginate demonstrated lowest growth. Mixed cultures immobilized inside cloth sachets showed no significant enhancement compared to pure cultures, reaching a maximum removal of 81% after 96h for consortium I, consisting of the four immobilized strains together. Nevertheless, the cells could be reused for two additional cycles of 96h each, obtaining a maximum removal efficiency of 71.5% when each of the four strains was immobilized in a separate bag (consortium III).