Article Index

1.1. Bioremediation of Ecosystems Contaminated With Heavy Metals

All metals, in spite of whether they are essential or non-essential, can exhibit toxic effects at elevated concentrations. Once a pollutant finds entry into a living organism, it may exhibit an injurious action. The effect of the pollutant is therefore a function of its concentration at the site of its action. Metal toxicity becomes more severe in acidic medium, nutrient-deficient ecosystem and poor physical conditions.

The remediation can be attempted through conventional remedial measures such as land filling and leaching, excavation and burial or soil washing. An extensive use of solidwaste landfills for disposal of municipal and industrial wastes as well as inappropriate use of agro-chemicals has generated a huge amount of leachate causing groundwater pollution, and the potential for groundwater contamination by leachate has necessitated for the invention of novel engineering designs for landfills. Remediation of heavy metals-polluted ecosystems could be carried out using physicochemical processes such as ion exchange, precipitation, reverse osmosis, evaporation and chemical reduction. However, due to problems such as membrane fouling, high costs, high energy requirement and low removal efficiency, these processes show little relevance in industries. In general, technical applicability, cost-effectiveness and plant simplicity are the key factors in selecting the most suitable treatment method to remove heavy metals (such as Cu, Ar, Pb and Zn) and cyanide from contaminated ecosystem. However, the latest technologies like photocatalytic reduction, surfactant-based membranes, liquid membranes and surface complexation are more efficient for heavy metals removal from contaminated ecosystems.

1.1.1. Microbe-Based Clean Up System (Microbial Bioremediation)

Microorganisms uptake heavy metals actively (bioaccumulation) and/or passively (adsorption). The microbial cell walls, which mainly consist of polysaccharides, lipids and proteins, offer many functional groups that can bind heavy metal ions, and these include carboxylate, hydroxyl, amino and phosphate groups. Among various microbe-mediated methods, the biosorption process seems to be more feasible for large scale application compared to the bioaccumulation process, because microbes will require addition of nutrients for their active uptake of heavy metals, which increases the biological oxygen demand or chemical oxygen demand in the waste. Further, it is very difficult to maintain a healthy population of microorganisms due to heavy metal toxicity and other environmental factors. Fungi of the genera Penicillium, Aspergillus and Rhizopus are potential microbial agents for the removal of heavy metals from aqueous solutions. Endophytic bacteria that are known to be beneficial to plants also enhance the ability of host plants accumulating higher levels of heavy metals.

Microorganisms are ubiqutious in heavy metal-contaminated environments and can easily convert heavy metals into non-toxic forms. In bioremediation processes, microorganisms mineralize the organic contaminants to end-products such as CO2 and H2O, or to metabolic intermediates which are used as primary substrates for cell growth. Different mechanisms of bioremediation are known, including biosorption, metal-microbe interactions, bioaccumulation, biomineralisation, biotransformation and bioleaching. Microorganisms are capable of dissolving metals and reducing or oxidizing transition metals. Different methods by which microbes restore the environment are oxidizing, binding, immobilizing, volatizing and transformation of heavy metals. Bioremediation can be made successful in a particular location by the designer microbe approach, and by understanding the mechanism controlling growth and activity of microorganisms in the contaminated sites, their metabolic capabilities and their response to environmental changes.

Microbial bioremediation by adsorption

Heavy metals can be biosorbed by microbes at binding sites present in cellular structure without the involvement of energy. Among the various reactive compounds associated with bacterial cell walls, the extracellular polymeric substances (high-molecular weight compounds secreted into their environment) are of particular importance and are well known to have significant effects on acid-base properties and metal adsorption. Secreted extracellular polymeric substances have a great ability to complex heavy metals through various mechanisms including proton exchange and micro-precipitation of metals.

Microbial bioremediation by physio-bio-chemical mechanism

Biosorption is the process which involves higher affinity of a biosorbent towards sorbate (metal ions), continued until equilibrium is established between the two components. Saccharomyces cerevisiae acts as a biosorbent for the removal of Zn (II) and Cd (II) through the ion exchange mechanism. Cunninghamella elegans is a promising sorbent against heavy metals released by textile wastewater. Fungi are potential biocatalysts to access heavy metals and transform them into less toxic compounds. Some fungi such as Klebsiella oxytoca, Allescheriella sp., Stachybotrys sp., Phlebia sp., Pleurotus pulmonarius, Botryosphaeria rhodina have metal binding potential. Pb (II) contaminated soils can be remediated by fungal species like A. parasitica and Cephalosporium aphidicola with biosorption process. Hg resistant fungi (Hymenoscyphus ericae, Neocosmospora vasinfecta and Verticillium terrestre) were able to biotransform a Hg (II) state to a nontoxic state. Many of the contaminants are hydrophobic, and they are taken up by microbes through the secretion of some biosurfactant and direct cell-contaminant association. Biosurfactants form stronger ionic bonds with metals and form complexes before being desorbed from soil matrix to water phase due to low interfacial tension.

Bioremediation may also involve aerobic or anaerobic microbial activities. Aerobic degradation often involves introduction of oxygen atoms into the reactions mediated by monooxygenases, dioxygenases, hydroxylases, oxidative dehalogenases, or chemically reactive oxygen atoms generated by enzymes such as ligninases or peroxidases. Anaerobic degradations of contaminants involve initial activation reactions followed by oxidative catabolism mediated by anoxic electron acceptors. The immobilization technique is used to reduce the mobilization of heavy metals from contaminated sites by changing the physical or chemical state of the toxic metals. Solidification treatment involves mixing of chemical agents at the contaminated sites or precipitation of hydroxides. In the contaminated sites, microbes mobilize the heavy metals by leaching, chelation, methylation and redox tansformation of toxic metals. It is not possible to destroy heavy metals completely, but the process transforms their oxidation state or organic complex, making them water-soluble, precipitated and less toxic. In the bioremediation of contaminated environments, microbes use heavy metals and trace elements as terminal electron acceptors or reduce them through the detoxification mechanism. Microorganisms remove heavy metals through the mechanisms which they employ to derive energy from metals redox reactions, to deal with toxic metal through enzymatic and non-enzymatic processes. Two main mechanisms for development of resistance in bacteria are detoxification (transformation of the toxic metal state and making it unavailable) and active efflux pumping of the toxic metal from cells. The basic redox (oxidation and reduction) reaction takes place in the soil between toxic metals and microorganisms; microorganisms act as an oxidizing agent for heavy metals and cause them to lose electrons, which are accepted by alternative electron acceptors (nitrate, sulphate and ferric oxides).

In aerobic conditions, oxygen acts as an electron acceptor, while in anaerobic conditions microbes oxidize organic contaminants by reducing electron acceptors. The microorganism takes energy for growth by oxidizing the organic compound with Fe (III) or Mn (IV) as an electron acceptor. Anaerobic degradation of organic contamination is stimulated with the higher availability of Fe (III) for microbial reduction. Biodegradation of chlorines from contaminants takes place through reductive dechlorination, where contaminants as chlorinated solvents act as an electron acceptors in respiration. Microorganisms reduce the state of metals and change their solubility, like the Geobacter (anaerobic respiration bacterial species found in anaerobic conditions in soils and aquatic sediment), and reduce the Uranium soluble state (U6+) to insoluble state (U4+). Different defense systems (exclusion, compartmentalization, complex formation and synthesis of binding protein and peptides) reduce the stress developed by toxic metals. These metal binding protein transcription factors are known to mediate in hormone and redox signaling process in the context of toxic metal (Cd, Zn, Hg, Cu, Au, Ag, Co, Ni and Bi) exposure.

1.1.2. Phytoremediation of Heavy Metals

Phytoremediation is an eco-friendly in situ remediation technology driven by solar energy. Plants and associated microorganisms can be used for removal of heavy metals partially or completely remediate selected contaminants from soil, sludge, sediments, wastewater and ground water. In the phytoremediation of heavy metals, the initial step is phytoextraction, the uptake of heavy metal contaminants from soil or water by plant roots and their translocation to and accumulation in biomass. Translocation of metals to shoots is an important biochemical process and is desirable in an effective phytoextraction. The next important process of phytoremediation is phytofiltration, which includes rhizofiltration, blastofiltration or caulofiltration. In this, the metals are absorbed or adsorbed and thus their movement in soil and underground water is minimized. In addition to the above process, phytostabilization or phytoimmobilization reduces the mobility and bioavailability of metals in the environment. Plants perform the immobilization of heavy metals in soils by sorption through roots, precipitation, complex formation or metal valence reduction in the rhizosphere. Some of the heavy metals such as Hg and Se, absorbed by plants from polluted soils, get converted into volatile forms and subsequently released into the atmosphere by phytovolatilization process. This process does not remove the metals completely but rather transfers them from one medium (soil or water) to another (atmosphere) from which they can reenter soil and water.

Removal of heavy metals through phytoremediation, especially hyperaccumulators to degrade and detoxify contaminants receives wide attention due to its efficacy and cost efficiency. The criteria used for hyperaccumulation varies according the metal. Hyperaccumulator plants exhibit higher heavy metal tolerance and accumulating abilities compared to other plants. Difficulty in finding heavy metal hyperaccumulators, slow growth and lower biomass yield limit the use of hyperaccumulators. This makes the process time-consuming and therefore not feasible for rapidly contaminated sites or sewage treatments. Rhizospheric microorganisms such as Arbuscular mycorrhizal fungi and plant growth-promoting rhizobacteria, playing important roles in plant growth and/or metal tolerance via different mechanisms, are beneficial for the design of a phytoremediation plan to select appropriate multifunctional microbial combinations from the rhizosphere. It is likely that remediation role of rhizosphere is the main part of phytoremediation and removal of contaminants is achieved by the combined activity of plants and microorganisms. The main reason for the enhanced removal of metals in the rhizosphere is likely the increase in the number and metabolic activities of microorganisms. In the rhizospheric degradation process, the metal toxicity to plants can be reduced by the use of plant growth-promoting bacteria, free-living soil microorganisms that exert beneficial effects on plant growth. In this process, plants can stimulate microbial activity about 10–100 times by the secretion of exudates which contain carbohydrates, amino acids, flavonoids etc. In return, the rhizosphere bacteria facilitate the generation of larger roots helping to enhance plant survival.