from sulfide minerals. These bacteria are gram negative, aerobic, and able to survive below pH 3.0 and temperature 25°C–35°C. Leaching bacteria are mostly belongs to proteobacteria (Acidithiobacillus sp., Acidiphilium sp., Acidiferrobacter sp., and Ferrovum sp.), Nitrospirae (Leptospirillum sp.), Firmicutes (Alicyclobacillus sp. and Sulfobacillus sp.), and Actinobacteria (Ferrimicrobium sp., Acidimicrobium sp., and Ferrithrix sp.) [68, 69]. But among all bioleaching bacteria, Thiobacillus sp. involved in solubilization of metal sulfide because it takes carbon dioxide from atmosphere for cellular synthesis, pulls their energy from oxidation of elemental sulfur, reduces sulfur compounds, and results in production of ferric ions and sulfuric acids, which are entangled in heavy metal extraction. The frequently used bacteria for bioleaching process are Acidothiobacillus thiooxidans, Acidothiobacillus ferrooxidans, and Leptospirillum ferrooxidans, and these are able to grow in high acidic condition (pH 1.5–3.0) [69].
Figure 3.3 Mechanism of bioleaching.
The oxidation of metal sulfide by Fe/S oxidizing bacteria is defined through two distinct pathways such as polysulfide and thiosulfate pathway [68, 69]. These mechanisms depend on metal sulfide reactivity with protons (acid solubility) [69]. In case of thiosulfate pathway, metals are acid-insoluble such as pyrite (FeS2), molybdenite (MoS2), and tungstenite (WS2), and Fe3+ ions occur through metal sulfide extraction. This reaction results the production of metal cations (M+) and thiosulfate that oxidizes to sulfuric acid. The production of sulfuric acid creates acidic condition so T. ferrooxidans and L. ferroxidans catalyze Fe3+ ions for recycling. In case of polysulfide pathway, metals are acid soluble such as sphalerite (ZnS), galena (PbS), arsenopyrite (FeAsS), chalcopyrite (CuFeS2), and hauerite (MnS2) through electron extraction by iron(III) ions and proton attack. In this mechanism, polysulfide is the main intermediate form and can be oxidized to sulfuric acid by using bacteria A. ferroxidans and A. thiooxidans [71]. In bioleaching process, maintenance of acidic condition is essential because the optimum action of Fe/S oxidizing bacteria and to retain metals constant in solution phase.
3.5.3 Biovolatilization
The transformation of metals by microbes into their volatile forms is known as bio-volatilization and contributes in the alteration of metal from soluble state to gaseous state. This biovolatilization process can remove metal from solid phase by utilizing microbes. Therefore, this process can be applied for both wastewater treatment and solid waste treatment. If the gas form of volatilized metals can trick from wastewater treatment method, they can be consequently recovered [72]. The metals commonly connected with their methylation and alkylation of biovolatilization method by microbes, whereas volatilization of mercury and arsenic may also be facilitated by their removal [72, 73]. Biovolatilization is a common method for mercury and arsenic in environment through which detoxification approaches applied on soil and water based on transformation of highly toxic compound to nontoxic or less toxic compound and highly volatile for removal of metals (Figure 3.4) [73].
Figure 3.4 Mechanism of biovolatilization.
In contaminated environment, bacteria developed resistance resulting due to the aforesaid mechanism which further leads to mercury detoxification. The reductase enzyme (Mercury(II)reductase) of the bacteria causes a reduction of Hg2+ to nontoxic Hg0, and hence, a diffusional loss of Hg0 from bacterial cell takes place. The mercuric reductase coded by merA gene is important for reduction of inorganic Hg while cytosolic mercuric lyase enzymes coded by merB gene breaks the C-Hg bond of most organomercury [69]. Earlier studies reported that bacteria involved in this mechanism and resistance to Hg such as Bacillus sp., Pseudomonas sp., Psychrobacter sp., Halmonas sp., Luteimonas sp., and Micrococcus sp. are isolated from highly polluted area [74]. The elemental mercury is highly volatile and the gas phase needs some special treatment to immobilize it. The Hg0 produced by volatilization and it is removed into gas phase by fast oxidative absorption process and recovered. This technique can be applied on soil, wastewater, and sediment [69].
The biovolatilization process also involve in arsenic removal from contaminated soil and water. In soil, arsenic could be converted into volatile byproducts and removed. Both aerobic and anaerobic microorganisms are involved in the evolution of volatile arsenicals. The volatilization of arsenic by microorganisms depends upon several factors like arsenic compound, concentration, and moisture of soil, organic materials, temperature, other similar components, growth of microbes, and ability of volatilization of arsenic. Biovolatilization of arsenic is by lessening of As(V) to As(III) with end product of TMAs. Currently, Escherichia coli have expressed arsenite S-adenosylmethionine methyltransferase gene (arsM), which is cloned from Rhodopseudomonas palustris and is capable to form methylate inorganic arsenic to TMA volatile form. In indigenous bacteria, arsM gene has capability to remove As through volatilization from soil. The strains express arsM gene in aquatic system such as Sphingomonas desiccabilis and Bacillus idriensis. The arsenic resistant bacteria can express arsM gene for biovolatilization of arsenic and these bacteria can engineered under laboratory condition to apply in aquatic and soil environment [73].
3.5.4 Bioimmobilization
Currently, bioimmobilization process is used in bioremediation, biodegradation, bio-control, pesticide use, and the manufacture of numerous compound products like antibiotics, enzyme or steroids, and amino acids. In this technique, metal can immobilized using microbial biomass by biosorption to cell walls or by extracellular substances and some common procedures are using for immobilization such as adsorption on exteriors, flocculation, cross connecting of cells, nanocoating, entrapment, covalent bonding to carriers, and encapsulation. The bacteria persuade immobilization mechanism to reduce the heavy metal concentration [69, 75]. The metabolism and intrinsic property of some bacteria associated with cell wall structure and the presence of extracellular polymeric substances are able to tolerate metal ions. Some other bacteria resist to metal by using resistance mechanisms such as active transport, efflux pump, intra- and extracellular sequestration, methylation, toxic chemical transfer to less toxic chemical through enzymatic transformation of redox reaction, and sensitivity reduction of cellular targets to metal (Figure 3.5) [76].
The heavy metals are reduced by using immobilization process. The Cr(VI) is reduced to Cr(III) by using both anaerobic and aerobic microorganisms. The presence of oxygen in aerobic condition and the reduction of Cr(VI) by microbes are generally catalyzed though soluble enzyme and lessening of Cr(VI) to Cr(III) by microbes as an eco-friendly method [77]. The bacterial strain such as E. coli, Pseudomonas putida, Desulfovibrio sp., Bacillus sp., Shewanella sp., Arthobacter sp., Microbacterium sp., and Cellulomonas sp., which reduce Cr(VI) isolated from contaminated area [78]. Arsenic compound used as an electron donor or accepter by microorganisms and possess the detoxification of arsenic, with pushes up to the membrane level of cells to eradicate As(III) from cells and metabolites of cell, finally As(V) removal arise [69]. Anaerobic bacteria are capable to reduce contaminated As(V) to As(III) and sulfate to elemental sulfur and precipitates in the form of arsenite sulfide [79]. Therefore sulfide precipitation is a useful mechanism for reduction of arsenic. The EPS of Chryseomonas luteola immobilized the metal ions such as cadmium, cobalt,
