Combination of slurry-bioreactors and actinobacteria consortia as strategy to bioremediate chlordane-contaminated soils.

Soil contamination caused by pesticides poses a significant environmental challenge, and addressing it requires effective solutions. Bioremediation, combining the utilization of slurry-bioreactors and microbial consortia, emerges as an appropiated strategy to tackle this issue. Therefore, this research evaluated the chlordane (CLD) removal efficiency by a Streptomyces consortium through bioaugmentation of polluted soils, and slurry-bioreactors. For that, a Streptomyces defined consortium with CLD removal abilities was inoculated in soil microcosms and soil-slurry bioreactors (SB), with (SB-TSB) and without stimulation (SB-water). In soil, CLD presence has no negative effect on consortium growth. This was supported by comparing its duplication time (7.48 ± 0.14 h) with the obtained in the biotic control (7.45 ± 0.04 h). Furthermore, 17% of pesticide removal by microbial action was detected in the treated microcosms. In SB, the microbial development was not affected by the pesticide presence. In SB-TSB, the microbial growth was higher than in SB-water. This was supported by its lesser duplication time (7.27 ± 0.17 h) with respect to the non-stimulated systems (10.88 ± 0.29 h). However, SB-water showed the highest CLD removal ability (34.8%), with a concomitant increase in the chloride ion release. In the phytotoxicity test, the vigor index showed that the bioremediation in SB-water did not exert adverse effects greater than those generated by the CLD. Indeed, the root length increased after the treatment. These findings demonstrate the versatility of the Streptomyces consortium to remediate solid and semi-solid matrices impacted with pesticides, and the advantage of using bioaugmented SB to enhance the pollutants removal and accelerating the clean-up time required.

Applied of actinobacteria consortia-based bioremediation to restore co-contaminated systems.

Global industrialization and natural resources extraction have left cocktails of environmental pollutants. Thus, this work focuses on developing a defined actinobacteria consortium able to restore systems co-contaminated with pollutants occurring in Argentinian environments. In this context, five actinobacteria were tested in solid medium to evaluate antagonistic interactions and tolerance against lindane (LIN), Reactive Black B-V (RBV), phenanthrene (Ph) and Cr(VI). The strains showed absence of antagonism, and most of them tolerated the presence of individual pollutants and their mixtures, except Micromonospora sp. A10. Thus, a quadruple consortium constituted by Streptomyces sp. A5, M7, MC1, and Amycolatopsis tucumanensis DSM 45259T, was tested in liquid systems with individual contaminants. The best microbial growth was observed in the presence of RBV and the lowest on Cr(VI). Removals detected were 83.3%, 65.0% and 52.4% for Ph, RBV and LIN, respectively, with absence of Cr(VI) dissipation. Consequently, the consortium performance was tested against the organic mixture, and a microbial growth similar to the biotic control and a LIN removal increase (61.2%) were observed. Moreover, the four actinobacteria of the consortium survived the mixture bioremediation process. These results demonstrate the potential of the defined actinobacteria consortium as a tool to restore environments co-contaminated with organic pollutants.

Actinobacteria bioaugmentation and substrate evaluation for biobeds useful for the treatment of atrazine residues in agricultural fields.

Biopurification systems (BPS) or biobeds are bioprophylaxis systems to prevent pesticide point-source contamination, whose efficiency relies mostly on the pesticide removal capacity of the biomixture, the majority component of a BPS. The adaptation of the components of the biomixtures to local availabilities is a key aspect to ensure the sustainability of the system. In this work, the removal of atrazine (ATZ) was evaluated in biomixtures formulated with three sugarcane by-products as alternative lignocellulosic substrates. Based on the capacity of actinobacteria to tolerate and degrade diverse pesticides, the effect of biomixtures bioaugmentation with actinobacteria was evaluated as a strategy to enhance the depuration capacity of biobeds. Also, the effect of ATZ and/or the bioaugmentation on microbial developments and enzymatic activities were studied. The biomixtures formulated with bagasse, filter cake, or harvest residue, reached pesticide removal values of 37-41% at 28 d of incubation, with t1/2 between 37.9 ± 0.4 d and 52.3 ± 0.4 d. The bioaugmentation with Streptomyces sp. M7 accelerated the dissipation of the pesticide in the biomixtures, reducing ATZ t1/2 3-fold regarding the controls, and achieving up to 72% of ATZ removal. Atrazine did not exert a clear effect on microbial developments, although most of the microbial counts were less in the contaminated biomixtures at the end of the assay. The bioaugmentation improved the development of the microbiota in general, specially actinobacteria and fungi, regarding the non-bioaugmented systems. The inoculation with Streptomyces sp. M7 enhanced acid phosphatase activity and/or reversed a possible effect of the pesticide over this enzymatic activity.

Glycerol as a substrate for actinobacteria of biotechnological interest: Advantages and perspectives in circular economy systems.

Actinobacteria represent a ubiquitous group of microorganisms widely distributed in ecosystems. They have diverse physiological and metabolic properties, including the production of extracellular enzymes and a variety of secondary bioactive metabolites, such as antibiotics, immunosuppressants, and other compounds of industrial interest. Therefore, actinobacteria have been used for biotechnological purposes for more than three decades. The development of a biotechnological process requires the evaluation of its cost/benefit ratio, including the search for economic and efficient substrates for microorganisms development. Biodiesel is a clean, renewable, quality and economically viable source of energy, which also contributes to the conservation of the environment. Crude glycerol is the main by-product of biodiesel production and has many properties, so it has a commercial value that can be used to finance the biofuel production process. Actinobacteria can use glycerol as a source of carbon and energy, either pure o crude. A circular economy system aims to eliminate waste and pollution, keep products and materials in use, and regenerate natural systems. Although these principles are not yet met, some approaches are being made in this direction; the transformation of crude glycerol by actinobacteria is a process with great potential to be scaled on an industrial level. This review discusses the reports on glycerol as a promising source of carbon and energy for obtaining biomass and high-added value products by actinobacteria. Also, the factors influencing the biomass and secondary metabolites production in bioreactors are analyzed, and the tools available to overcome those that generate the main problems are discussed.

Use of glycerol for the production of actinobacteria with well-known bioremediation abilities.

In recent years, there has been an increasing interest in the remediation of contaminated environments, and a suitable solution is in situ bioremediation. To achieve this, large-scale bacterial biomass production should be sustainable, using economic culture media. The main aim of this study was to optimize the physicochemical conditions for the biomass production of an actinobacterium with well-known bioremediation ability using inexpensive substrates and to scale-up its production in a bioreactor. For this, the growth of four strains of actinobacteria were evaluated in minimal medium with glucose and glycerol as carbon and energy sources. In addition, l-asparagine and ammonium sulfate were assayed as alternative nitrogen sources. The strain Streptomyces sp. A5 showed the highest biomass production in shake-flasks culture using glycerol and ammonium sulfate as carbon and nitrogen sources, respectively. Factorial designs with five factors (glycerol concentration, inoculum size, pH, temperature, and agitation) were employed to optimize the biomass production of Streptomyces sp. A5. The maximum biomass production was obtained using 5 g L-1 of glycerol, 0.25 µL of inoculum, pH 7, 30 °C and 200 rpm. Finally, the production was successfully scaled to a 2 L stirred tank bioreactor. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s13205-020-02588-5.

Bioremediation of lindane-contaminated soils by combining of bioaugmentation and biostimulation: Effective scaling-up from microcosms to mesocosms.

The scaling-up of lindane-contaminated soils bioremediation from microcosms to mesocosms bioaugmentated with an actinobacteria quadruple culture and biostimulated with sugarcane filter cake (SCFC) was surveyed. Mesocosms of silty loam soil, clayey soil, and sandy soil were polluted with the pesticide, bioaugmented with the mixed culture, biostimulated with adequate amounts of 0.5 mm SCFC particles, and assessed during 63 days maintaining environmental parameters with minimal intervention. Samples were taken to determine residual lindane, heterotrophic microorganisms, enzymatic activities, and bioremediation effectiveness using ecotoxicity tests with Raphanus sativus, Lactuca sativa, and Lycopersicon esculentum. The bioaugmentation and biostimulation of the three soils improved lindane removal, microbial counts, and enzymatic activities, and reduced pesticide T1/2, regarding the values obtained in non-bioremediated controls. The removal process was significantly affected by the soil type, and the highest pesticide dissipation (82.6%) was detected in bioremediated sandy soil. Ecotoxicity tests confirmed the bioremediation success through a rise in the vigor index of seedlings compared to non-treated soils (R. sativus: 12-22%; L. sativa: 12-20%; L. esculentum: 30-45%). Finally, scanning electron microscopy corroborated soil colonization by actinobacteria. Successful scaling-up of the combined application of an actinobacteria quadruple culture and SCFC as an appropriate strategy for restoring lindane-polluted soils at mesocosms-scale was confirmed.

Enhanced bioremediation of lindane-contaminated soils through microbial bioaugmentation assisted by biostimulation with sugarcane filter cake.

Lindane is a toxic and persistent organochlorine pesticide, whose extensive use generated its accumulation in different environmental matrices. Bioremediation is a promising technology that can be used combining bioaugmentation and biostimulation processes to soil restoration. The aim of the present work was to determine the conditions of maximum lindane removal by bioaugmentation with an actinobacteria consortium and biostimulation with sugarcane filter cake (SCFC). The assays were carried out on lindane-contaminated silty loam (SLS), clayey (CS), and sandy (SS) soils. Through complete factorial designs, the effects of three abiotic factors (moisture content, proportion and size of SCFC particles) were evaluated on lindane removal. In addition, a response optimizer determined the optimal conditions for pesticide removal in bioaugmented and biostimulated soils, in the range of levels studied for each factor. In these conditions, bioaugmentation of biostimulated soils increased the pesticide removal (SLS: 61.4%, CS: 70.8%, SS: 86.3%), heterotrophic microbial counts, and soil enzymatic activities, and decreased lindane T1/2, regarding the non-bioaugmented biostimulated controls, after 14 days of assay. The values of these parameters confirmed the efficiency of the bioremediation process. Finally, the viability of the four strains was demonstrated at the end of the assay. The results indicate that the simultaneous application of bioaugmentation with the actinobacteria consortium and biostimulation with SCFC constitutes a promising tool for restoring soils contaminated with lindane, by using the optimal conditions obtained through the factorial designs.

Coupling of bioaugmentation and biostimulation to improve lindane removal from different soil types.

Lindane is an organochlorine pesticide that, due to its persistence in the environment, is still detected in different matrices. Bioremediation using actinobacteria consortia proved to be promising for the restoration of contaminated soils. Another alternative to remove xenobiotics is to use agricultural residues, which stimulates microbial activity, increasing its capacity to degrade organic pollutants. The present work studies the coupling of sugarcane bagasse biostimulation and bioaugmentation with the actinobacteria consortium composed of Streptomyces sp. A2, A5, A11 and M7 on lindane removal in different soil types. In this sense, factorial designs with three factors (proportion and size of sugarcane bagasse particles, and moisture content) were employed. A response optimizer identified the combination of factors levels that jointly allowed obtaining the maximum lindane removal in the evaluated conditions. In the optimal conditions, the effect of the bioremediation process on soil microbiota was studied by evaluating different parameters. The highest lindane removal percentages were detected in biostimulated microcosms bioaugmented with the microbial consortium, which were accompanied by a decrease in lindane half-life respect to the controls. Also, the bioaugmentation of biostimulated microcosms increased the microbial counts and enhanced soil enzymatic activities, corroborating the bioremediation process efficiency. The survival of the four actinobacteria at the end of the assay confirmed the ability of all Streptomyces strains to colonize amended soils. Bioremediation by simultaneous application of biostimulation with sugarcane bagasse and bioaugmentation with the actinobacteria consortium, in the optimized conditions, represents an efficient strategy to restore lindane contaminated soils.

Lindane dissipation in a biomixture: Effect of soil properties and bioaugmentation.

The biomixture is the major constituent of a biopurification system and one of the most important factors in its efficiency; hence the selection of the components is crucial to ensure the efficient pesticides removal. Besides, bioaugmentation is an interesting approach for the optimization of these systems. A mixed culture of the fungus Trametes versicolor SGNG1 and the actinobacteria Streptomyces sp. A2, A5, A11, and M7, was designed to inoculate the biomixtures, based on previously demonstrated ligninolytic and pesticide-degrading activities and the absence of antagonism among the strains. The presence of lindane and/or the inoculum in the biomixtures had no significant effect on the development of culturable microorganisms regardless the soil type. The consortium improved lindane dissipation achieving 81-87% of removal at 66 d of incubation in the different biomixtures, decreasing lindane half-life to an average of 24 d, i.e. 6-fold less than t1/2 of lindane in soils. However, after recontamination, only the bioaugmented biomixture of silty loam soil enhanced lindane dissipation and decreased the t1/2 compared to non-bioaugmented. The biomixture formulated with silty loam soil, sugarcane bagasse, and peat, inoculated with a fungal-actinobacterial consortium, could be appropriate for the treatment of agroindustrial effluents contaminated with organochlorine pesticides in biopurification systems.

Cr(VI) and lindane removal by Streptomyces M7 is improved by maize root exudates.

Environmental mixed pollution by both organic and inorganic compounds are detected worldwide. Phytoremediation techniques have been proposed as ecofriendly methods for cleaning up polluted sites. Several studies have demonstrated enhanced dissipation of contaminants at the root-soil interface through an increase in microbial activity caused by the release of plant root exudates (REs). The aim of this study was to evaluate the effectiveness for Cr(VI) and lindane removal by Streptomyces M7 cultured in a co-contaminated system in presence of maize REs. Our results showed when REs were added to the contaminated minimal medium (MM) as the only carbon source, microbial removal of Cr(VI) and lindane increased significantly in comparison to contaminant removal obtained in MM with glucose 1 g L-1 . The maximum removal of 91% of lindane and 49.5% of Cr(VI) were obtained in the co-contaminated system. Moreover, Streptomyces M7 showed plant growth promoting traits which could improve plant performance in contaminated soils. The results presented in this study provide evidence that maize REs improved growth of Streptomyces M7 when REs were used as a carbon source in comparison to glucose. Consequently, lindane and Cr(VI) removal was considerably enhanced making evident the phytoremediation potential of the actinobacteria-plant partnership.

Integral use of sugarcane vinasse for biomass production of actinobacteria: Potential application in soil remediation.

The use of living actinobacteria biomass to clean up contaminated soils is an attractive biotechnology approach. However, biomass generation from cheap feedstock is the first step to ensure process sustainability. The present work reports the ability of four actinobacteria, Streptomyces sp. M7, MC1, A5, and Amycolatopsis tucumanensis, to generate biomass from sugarcane vinasse. Optimal vinasse concentration to obtain the required biomass (more than 0.4 g L-1) was 20% for all strains, either grown individually or as mixed cultures. However, the biomass fraction recovered from first vinasse was discarded as it retained trace metals present in the effluent. Fractions recovered from three consecutive cycles of vinasse re-use obtained by mixing equal amounts of biomass from single cultures or produced as a mixed culture were evaluated to clean up contaminated soil with lindane and chromium. In all cases, the decrease in pesticide was about 50% after 14 d of incubation. However, chromium removal was statistically different depending on the preparation methodology of the inoculum. While the combined actinobacteria biomass recovered from their respective single cultures removed about 85% of the chromium, the mixed culture biomass removed more than 95%. At the end of the reused vinasse cycle, the mixed culture removed more than 70% of the biological oxygen demand suggesting a proportional reduction in the effluent toxicity. These results represent the first integral approach to address a problematic of multiple contaminations, concerning pesticides, heavy metals and a regionally important effluent like vinasse.

Removal of a mixture of pesticides by a Streptomyces consortium: Influence of different soil systems.

Although the use of organochlorine pesticides (OPs) is restricted or banned in most countries, they continue posing environmental and health concerns, so it is imperative to develop methods for removing them from the environment. This work is aimed to investigate the simultaneous removal of three OPs (lindane, chlordane and methoxychlor) from diverse types of systems by employing a native Streptomyces consortium. In liquid systems, a satisfactory microbial growth was observed accompanied by removal of lindane (40.4%), methoxychlor (99.5%) and chlordane (99.8%). In sterile soil microcosms, the consortium was able to grow without significant differences in the different textured soils (clay silty loam, sandy and loam), both contaminated or not contaminated with the OPs-mixture. The Streptomyces consortium was able to remove all the OPs in sterile soil microcosm (removal order: clay silty loam > loam > sandy). So, clay silty loam soil (CSLS) was selected for next assays. In non-sterile CSLS microcosms, chlordane removal was only about 5%, nonetheless, higher rates was observed for lindane (11%) and methoxychlor (20%). In CSLS slurries, the consortium exhibited similar growth levels, in the presence of or in the absence of the OPs-mixture. Not all pesticides were removed in the same way; the order of pesticide dissipation was: methoxychlor (26%)>lindane (12.5%)>chlordane (10%). The outlines of microbial growth and pesticides removal provide information about using actinobacteria consortium as strategies for bioremediation of OPs-mixture in diverse soil systems. Texture of soils and assay conditions (sterility, slurry formulation) were determining factors influencing the removal of each pesticide of the mixture.

Selection of an actinobacteria mixed culture for chlordane remediation. Pesticide effects on microbial morphology and bioemulsifier production.

Chlordane bioremediation using actinobacteria mixed culture is an attractive clean-up technique. Their ability to produce bioemulsifiers could increase the bioavailability of this pesticide. In order to select a defined actinobacteria mixed culture for chlordane remediation, compatibility assays were performed among six Streptomyces strains. The strains did not show growth inhibition, and they were assayed for chlordane removal, either as pure or as mixed cultures. In pure cultures, all of the strains showed specific dechlorination activity (1.42-24.20 EU mg(-1)) and chlordane removal abilities (91.3-95.5%). The specific dechlorination activity was mainly improved with cultures of three or four microorganisms. The mixed culture consisting of Streptomyces sp. A2-A5-A13 was selected. Their ability to produce bioemulsifiers in the presence of glucose or chlordane was tested, but no significant differences were observed (p > 0.05). However, the stability of the emulsions formed was linked to the carbon source used. Only in chlordane presence the emulsions retained 100% of their initial height. Finally, the selected consortium showed a high degree of sporulation in the pesticide presence. This is the first study on the effects that chlordane exerts on microbe morphology and emulsifier production for a defined mixed culture of Streptomyces with ability to remediate the pesticide.

Enhanced removal of a pesticides mixture by single cultures and consortia of free and immobilized Streptomyces strains.

Pesticides are normally used to control specific pests and to increase the productivity in crops; as a result, soils are contaminated with mixtures of pesticides. In this work, the ability of Streptomyces strains (either as pure or mixed cultures) to remove pentachlorophenol and chlorpyrifos was studied. The antagonism among the strains and their tolerance to the toxic mixture was evaluated. Results revealed that the strains did not have any antagonistic effects and showed tolerance against the pesticides mixture. In fact, the growth of mixed cultures was significantly higher than in pure cultures. Moreover, a pure culture (Streptomyces sp. A5) and a quadruple culture had the highest pentachlorophenol removal percentages (10.6% and 10.1%, resp.), while Streptomyces sp. M7 presented the best chlorpyrifos removal (99.2%). Mixed culture of all Streptomyces spp. when assayed either as free or immobilized cells showed chlorpyrifos removal percentages of 40.17% and 71.05%, respectively, and for pentachlorophenol 5.24% and 14.72%, respectively, suggesting better removal of both pesticides by using immobilized cells. These results reveal that environments contaminated with mixtures of xenobiotics could be successfully cleaned up by using either free or immobilized cultures of Streptomyces, through in situ or ex situ remediation techniques.

Bacterial bio-resources for remediation of hexachlorocyclohexane.

In the last few decades, highly toxic organic compounds like the organochlorine pesticide (OP) hexachlorocyclohexane (HCH) have been released into the environment. All HCH isomers are acutely toxic to mammals. Although nowadays its use is restricted or completely banned in most countries, it continues posing serious environmental and health concerns. Since HCH toxicity is well known, it is imperative to develop methods to remove it from the environment. Bioremediation technologies, which use microorganisms and/or plants to degrade toxic contaminants, have become the focus of interest. Microorganisms play a significant role in the transformation and degradation of xenobiotic compounds. Many Gram-negative bacteria have been reported to have metabolic abilities to attack HCH. For instance, several Sphingomonas strains have been reported to degrade the pesticide. On the other hand, among Gram-positive microorganisms, actinobacteria have a great potential for biodegradation of organic and inorganic toxic compounds. This review compiles and updates the information available on bacterial removal of HCH, particularly by Streptomyces strains, a prolific genus of actinobacteria. A brief account on the persistence and deleterious effects of these pollutant chemical is also given.

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 96 h for consortium I, consisting of the four immobilized strains together. Nevertheless, the cells could be reused for two additional cycles of 96 h each, obtaining a maximum removal efficiency of 71.5% when each of the four strains was immobilized in a separate bag (consortium III).