Describe how microbes can act as agents of bioremediation and discuss the reasons for successes and failures in their application

Background

Bioremediation is defined as a process that employs living organisms, mainly micro-organisms and plants in order to degrade or reduce or to detoxify waste products and other pollutants. Principal players of bioremediation are the micro-organisms, bacteria. Bacteria are best suited for the task of contamination destruction due of their special bacterial enzymes that enable them to make use of the environmental contaminants as food. The use of these environmental contaminants as their food helps to execute the process of bioremediation. The process is mainly executed by contacts contaminants (Hazen, 2018). In-situ process of bio-remediation is regarded as an extension of the purpose that the micro-organisms have served for the nature inception: the breakdown of the complex by-products of human, animals and plants waste so that life can continue to grow from one generation to the other. In the absence of the activity of the micro-organisms, the earth would be buried under the wastes and the important nutrients that are indispensible for the continuation of life will be buried up inside the debris (Hazen, 2018).

Whether the micro-organisms will be successful in degrading the man-made contaminants in the sub-surface mainly depends on three different factors: type of the organisms, the nature of the contaminants and the geological and chemical conditions at the site of contamination. Adams et al. (2015) are of the opinion that the principal goals of bioremediation is to stimulate microorganisms with the help the nutrients and other chemicals that will enable them to destroy the contaminants. The overall operation of bioremediation mainly rely on the strength of the micro-organisms that are native to the site of contamination and thus encouraging them to work by supplying them with standard level of nutrients and other group of chemicals that are important for their metabolism (Hazen, 2018). Thus, at present the process of bioremediation is limited by the capabilities of the native microbes. However, the researchers are making active initiatives for investigating the ways for augmenting the contaminating sites with the help of non-native microbes, mainly genetically engineered microbes that are specifically designed in order to degrade the contaminants at the concerned site. Regardless of whether the microbes are native to the site of contamination or are introduced as fresh, a proper understanding of the how the micro-organisms destroys the contaminants are critical for proper understanding of the process of bioremediation. The nature of microbial process that is used for the cleanup of the contaminants dictate the nature of the nutritional supplements important for the process of bioremediation. Furthermore, the by-products generated as a result of bioremediation indicates whether the process is successful in degrading the microbes (Adams et al., 2015).

The following scientific case study assignment aims towards identification of the how the micro-organisms can act as an agent for successful execution of the process of bioremediation. The case study assignment will also discuss the underlying reasons behind the success and failures in their application. The essay will begin with the findings of the already published research articles based on the process microbial bioremediation along with its success and failures. At the end scientific case study assignment will provide an overall discussion of the findings from the articles. The essay will end with recommendation in order to refine the process of bio-remediation along with the implication of practice. The essay will thus help in understanding the importance of microbes in bioremediation, the role of bioremediation in environment and how bioremediation can be refine further by modulating the role of microbes.

Findings

Basic Process of Bioremediation

The study conducted by Dzionek, Wojcieszyńska and Guzik (2016) highlighted that the process of bioremediation is used for commercial cleanup with a limited range of contaminants, mainly the hydrocarbons that are found in gasoline. Hydrocarbons are degraded through bioremediation because, hydrocarbons act as an important source of carbon. Carbons are the principal building blocks of the bacterial cell. They also provide electrons from which the organisms can extract energy. For example, the microbial cells extract energy from the contaminants via breaking the chemical bonds and transferring the electrons from contaminants to oxygen, an electron acceptor. This is known as aerobic bio-remediation. The reduction of the oxygen upon the uptake of the electron leads to the generation of water and the carbon is oxidised to carbon dioxide. The presence of water and carbon dioxide further helps to promote the growth of bacteria (Dzionek, Wojcieszyńska & Guzik, 2016). 

Figure: Carbon as a rich source of energy and building blocks of cell

(Source: Varjani, 2017)

Varjani (2017) are of the opinion that apart from the transformation of the contaminants by the use of the aerobic respiration, the bacterial microorganism also employs several variations over the basic process for the conduction of bioremediation. This process has evolved over time and enables the microorganism to thrive under unusual environment like in the underground and to degrade the toxic contaminants that are harmful for the nature. In order words, it can be said that apart from the utilization of the oxygen, micro-organisms can also break the contaminants under the absence of oxygen and this is known as anaerobic breakdown of the contaminants in the absence of oxygen. For example, the study conducted by Dzionek, Wojcieszyńska and Guzik (2016). highlighted that few of the micro-organisms is successful in employing soluble uranium (U6+) as the principal electron acceptor and reducing it to insoluble uranium (U4+). Under such circumstances, the organisms mainly use the uranium to precipitate and thereby helping to decrease the concentration of uranium in the ground water and restricting its mobility. Varjani (2017) stated that in addition to the organisms that make use of the inorganic chemical as the main electron acceptors for the conduction of the anaerobic respiration, other micro-organisms also make use inorganic molecules as electron donors. For example, inorganic electron donors are ammonium (NH4+), reduced state of iron (Fe2+), nitrite (NO2-), reduced state of manganese (Mn2+) and H2S. The oxidation of these inorganic molecule lead to the transfer of the electrons from to an electron acceptor (mainly oxygen) in order to generate energy for synthesis of cells. In the majority of the cases, bacteria make use of inorganic molecule as their primary electron donor and obtain their carbon the atmospheric CO2 (Varjani, 2017).

The critical analysis of the process of bioremediation

Bioremediation and removals of the hydrocarbons

The study conducted by Varjani (2017) is based on the background that petroleum hydrocarbon pollutants are the persistent organic pollutants in the environment. Cleaning up this pollutants from the environment is a challenge. Thus, the environmentalists are making use of the process of bioremediation as one of the principal method for restoration of the petroleum hydrocarbon polluted environment. The microbes with natural microbial bio-degradation activity are the main target for the process of degradation of petroleum hydrocarbons. Microorganisms that make use of the petroleum hydrocarbons are distributed ubiquitously within the environment. These microorganisms naturally bio-degrade pollutants and thus helping to remove them from the environment. Effective removal of the petroleum hydrocarbon pollutants from the environment is done under the application of the bio-remediation activity of the oleophilic microorganisms. The process is known as oil spill bioremediation. These microorganisms are eco-friendly and the overall process is cost-effective (Varjani, 2017). Microorganisms either catabolize petroleum hydrocarbons pollutants for extracting energy or assimilating them into a cell biomass. The microorganisms utilize hydrocarbons in three possible ways. First is the phototrophic, anoxygenic, chemotrophic, aerobic and chemotrophic anaerobic (Varjani, 2017).

Figure: Different by-products of petroleum bio-remediation

(Source: Varjani, 2017)

Figure: Possible pathway for the degradation of the petroleum hydrocarbons by bioremediation

(Source: Varjani, 2017)

Different microbes in bioremediation and their success

Wu et al. (2017) highlighted in the study that apart from oleophilic microorganisms, several other micro-organism also take part in the process of bioremediation of hydrocarbon degradation in the soil that are polluted with petroleum. The results of the Biolog (MT2) MicroPlates assay showed linear correlations between the total petroleum hydrocarbons (TPH) and alkane degradation rate. However, there is a lack of significant association between polycyclic aromatic hydrocarbon degradation rate and PAH population and role of microorganisms in degradation of hydrocarbons. The capability of the degradation of the petroleum hydrocarbons is mainly measured by the use of most probable number (MPN). Increase in the MPN indicates the increase in the bio-degradation activity of the microorganism. On the basis of the MPN activity score, it can be said that the process of bioremediation by the micro-organism mainly depends on the presence of certain microbial enzymes (Wu et al., 2017). The presence of these enzymes ensures successful bioremediation activity. For example, Abbasian et al. (2015) showed that the breakdown of the petroleum hydrocarbon compound like C1-C8 alkanes, C1-C5 (halogenated) alkanes, cycloalkanes and alkenes is mainly done by the action of enzymes, monooxygenases. Bacterial like Methylocystis, Methylomonas, Methylocella, Methylobacter, Methylococcus, Geobacillus hermodenitrificans, Methylomirabilis oxyfera posses these enzymes and thereby helping to conduct the bio-remediation process involving alkenes and cyclo-alkanes. The petroleum hydrocarbons like C5-C16 alkanes, alkyl benzenes and cycloalkanes are regarded under the action of hydroxylases enzymes. Bacteria like Pseudomonas, Burkholderia, Rhodococcus, Mycobacterium are a rich source of hydroxylases enzyme and thereby helping to conduct the process of degradation of these hydrocarbon compounds that belong to the category of alkyl benzenes.

The study conducted by Siles and Margesin (2018) helped to gain insights about the microbial communities that are mediating the process of bioremediation of hydrocarbon contaminated soil under an Alpine former military site. The study mainly aimed towards highlighting the characteristics of the microorganisms that help in improving the rate of decontamination. Siles and Margesin (2018) characteristed bacterial, fungal and archaeal communities in the domain of abundance (by the use of quantitative PCR) and under the light of the taxonomic diversity and structure by the use of the Illumina amplicon sequencing during the conduction of the process of bioremediation in long-term hydrocarbon contaminated soil for a tenure of 15 weeks of biostimulation. The biostimulation was given by the use of the inorganic NPK fertilization and subsequent comparison of biostimulation by external agent was done with the help of natural attenuation under optimal temperature (10 to 20 degree centigrade). The analysis of the results highlighted that though these is a considerable amount of the total petroleum hydrocarbon (TPH) loss under the process of the natural attenuation, the removal of TPH was significantly higher in the with the use of the inorganic NPK fertilizers at an increased rate of temperature. The authors are of the opinion that the use of the NPK fertilizers help to activate the microorganisms at an exponential rate and thus assisting the indigenous soil bacterial to perform better in the process of bioremediation. The same results were not reflected in case of archaeal and fungal communities of the soil at that site. This is because, the in-organic NPK fertilizer was ineffective in stimulating their biostimulation role. The study thus showed that the use of the NPK inorganic fertilizer (nitrogen, phosphorus, and potassium) help in stimulating soil bacteria of Gammaproteobacteria and Bacteroidia classes and thus helping to increase their overall activity in the process of bioremediation. The study helped in highlighting an important fact underlying the process of bioremediation (Siles & Margesin, 2018). The study showed that presence of the nitrogen, phosphorus and potassium help to promote successful outcome in the process of bioremediation. Such that the soils that lack adequate supply of nitrogenous bacteria or nitrosomonas enzymes, external supply of inorganic fertilizer helps to promote the successful outcome of the process of bioremediation.

Mikkonen et al. (2019) stated that that in order to increase the overall success of the process of bioremediation, genetic modification must be made in the genome of the bacterial residing in the ground water. The genetic modification must be done in situ by increasing the rate of synthesis of the pentachlorophenol hydroxylase gene (pcpB). The increase in the expression of pcpB genes help to promote overall aerobic incubation of the microsomes. This help in the remission of the chlorophenol-pollutents (pentachlorophenol and tetrachlorophenol) present in the ground water and thus helping in the process of water purification.

Microbial melanins and their reasons for success behind the process of bioremediation

Melanins are defined as special multifunctional pigments. It is mainly found in the skin of humans. It is also present in animals, fungi and bacteria (Solano, 2014). They are regarded as structurally complex biomolecules and is mainly extracted from the oxidation of indolic or phenolic compunds that mainly polymerizes in an ordered planer layers that clusters in an disordered configurations of macromoleculaes (Solano, 2014). In biology the melanin holds prime importance for providing proper protection and adaptation to different layers of the mechanical stressors like the adaptation of the extreme climatic condition, high level of humidity, toxicity produce by different level of pollutants. The ecology of some of the melanotic environments is important as it helps the bacteria to thrive under extreme weather conditions. Thus Gustavsson et al. (2016) is of the opinion those microorganisms that are rich in melanine are effective in taking part in the process of the bioremediation. Apart from the bacteria, fungi that contain melanine are also effective in taking part in the process of bioremediation with help of the assimilatory metabolic properties of the polluatnts (Cordero, Vij & Casadevall, 2017).

Failures and other limitations in the process of bioremediation

Numerous attempts for the successful bioremediation of the polycyclic aromatic hydrocarbon (PAH) contaminated sites failed miserably in the past. However, the reasons behind this failure were not well understood (Kästner & Miltner, 2018). Rein et al. (2016) used an improved model for conducting an integrated assessment for the mass transfer, biodegration and residual concentration for predicting the success of the overall process of remediation actions. Rein et al. (2016) first provided the growth parameters for Mycobacterium rutilum and Mycobacterium pallens growing over the pyrene (PYR) or phenanthrene (PHE). PYR degraded the PAH completely under the investigation concentration. The results also highlighted that maximum metabolic rates and the growth rate are similar for both the bacteria under both the substrates. However, Mycobacterium species were not superior in the degradation of PHE. Under real-world, circumstances of simulations including the diffusive flux for the microbial cells indicated that the process of bioaugmentation has only limited and short lived effect on the process of bioaugmentation. The study also showed that increasing sorption shifts the remaining PAH to absorb or to sequester the PAH pool. The last observation showed that mobilizing by solvents or surfactants resulted in a significant rise of the sequestered PHA and co-metabolization by compost addition. These contribute to a significant reduction of PAH due to the presence of active biomass in the compost (Rein et al., 2016). Thus, the overall study helped in understanding of the process of co-metabolization and how it decrease the efficacy of bacteria towards conduction of the process of bioremediation. Varjani (2017) stated that co-metabolism is a special case of secondary transformation. Secondary transformation can be defined as a process in which the bacteria transform the contaminants however; the transformation reaction fails to generate any positive effects to the cell. An important special case of secondary utilization is known as co-metabolism. In co-metabolism the transformation of the contaminant is defined as an incidental reaction catalyzed by the enzymes that are involved in the normal metabolism of cell or in special process of detoxification. For example, in oxidation of methane some bacteria fortuitously degrade chlorinated solves which is otherwise unable to attack. When the micro-organism oxidize the methane, it leads to the generation of certain enzymes that incidentally destroys the chlorinated solvents even when the solvent itself fail to support process of microbial growth. Co-metabolism of the chlorinated solvents under the action of the primary substrates like phenol, toluene and methane produce a negative environment for the bacteria and thus leading to bacterial death and thereby limiting the process of bioremediation.

Discussion

Environment pollution caused by several contaminants like the petroleum products is an alarming environmental issue. The environmental pollution that is occurring throughout the world as a result of petroleum is reported in the form of oil spill. The release of large amount of oil either directly or indirectly leads to huge environmental pollution along with negative environmental impact. Bioremediation (biostimulation and bioaugmentation) is defined as a natural yet eco-friendly process that helps to get rid of the environmental pollutants under the action of the microbial organisms (Dellagnezze, Gomes and de Oliveira, 2018).

Bacterial and its enzymes for bioremediation

The findings of the scientific case study assignment highlighted that microbial organism conduct the process of bioremediation either by the use of the aerobic or anaerobic respiration. The main microorganism that was found to conduct the process of bioremediation for the degradation of the petroleum byproducts includes oleophilic micro-organisms. The further analysis of the scientific articles highlighted that presence of certain enzymes in the bacteria help to increase their power of degrading the petroleum byproducts like C5-C16 alkanes, alkyl benzenes and cycloalkanes. The enzymes identified were hydroxylases and mono-oxygenases that are required to promote the bioremediation properties of bacteria. In special alpine lands, the process of bioremediation is slow due to the unavailability of the nitrogen, potassium and sodium in the soil. In the absence of these micronutrients, the bacteria fail to perform effective in the process of bioremediation and thus delaying the clearance of contaminates or toxic contaminates of the soil. The critical analysis of the articles highlighted that external supply of the inorganic fertilizer help to increase the bioremediation properties of the microbes and thus promoting the effective clearance of the toxic contaminants. Li et al. (2016) stated that external application of the inorganic fertilizer lead to the increase in the overall salinity of the social. These decrease the overall fertility of the social. Li et al. (2016) also highlighted in their study that for prolong period of time, increase in the soil salinity leads to decrease in the overall activity microbes to conduct the process of bioremediation. Such change in the microenvironment of the soil cast an irreversible change in the microbial mechanism making the bacterial cell incapable to conduct the process of bioremediation even under adequate supply of NKP fertilizers.

Bioremediation and genetic modifications

The success of the bioremediation by bioremediation can be increased by making genetic modification in the genome of the microorganism. For example, increase in the expression of the pentachlorophenol hydroxylase gene helps to increase the bioremediation properties of the bacteria and thereby helping to increase the capacity of bioremediation of pentachlorophenol and tetrachlorophenol. However, Partovinia and Rasekh (2018) are of the opinion that the gene manipulation of the micro-organism in order to increase their strength of bioremediation is associated with several disadvantages first is it is an expensive process and is time consuming and requires large scale approach in order to bring a comprehensive change. The recent studies are being conducted to generate two component regulatory systems (TCRSs) in order to mediate the cellular response by coupling sensing and regulatory mechanisms. The histidine kinase activity of the biosensors is helpful in regulating the expression of the effector gene after it is being phosphorylated by histidine kinase. Thus these attributes of the bacterial TCRSs can be effectively genetically engineered in order to design microbial systems for the process of bio-remediation. It is proposed that such system can have potential impact on the biorefinery (Ravikumar et al., 2017).

Melanin and bioremediation

The analysis of the literary papers also highlighted that melanine is an important constituents in the microbial systems that help the microbes in order to conduct the process of bioremediation. The melanine itself has special properties to act effectively under the stressors environment and these stressors properties of the melanine present in the microbes is utilised in the process of bioremediation. However, Cordero, Vij and Casadevall (2017) are of the opinion that recent research has been undertaken in order to highlight the radioactive properties of melanine. The anti-ionizing radiation present in the melanine is effective in counter acting the harmful effects of the ionizing radiation. Inonizing radiation is harmful for the humans as it leads to the generation of the reactive oxygen species within the body, this leads to the generation of the cellular stress and at the same time increase the vulnerability of developing cancer. Thus the skin with high deposition of melanine is less likely to become affected with the ionizing ration of the u.v rays and thus helping to prevent serious health related problems. This discovery is significant as it helped to show that the melanine containing bacteria is effective in thriving in extreme weather condition like in humid or in draught condition where there are high levels of sunrays. This capability to thrive in high temperature condition is effective in using it as an agent for bioremediation in extreme weather condition (Cordero, Vij & Casadevall, 2017).

Co-metabolism and bioremediation

The analysis of the review also highlighted that one of the limiters’ for the conduction of the process of bioremediation is the mechanism of the co-metabolism. Under this process, the chlorinated solvents that are produced by the breakdown of the primary substrates by the phenol, toluene and methane generate a negative environment for the bacterial to thrive in the soil and thereby leading to complete eradication of the bacterial colony. However, Pérez-de-Mora, A., Zila, McMaster and Edwards (2014) made use of enhanced in situ anaerobic bioremediation (EISB) and bioaugmentation for the treatment of the trichloroethane (TCE)-impacted groundwater at the fractured carbonated rock. The analysis of the study highlighted that changes in the dechlorinating and nondechlorinating populations during the process of biostimulation does not lead to decrease in the overall bacterial colony in the bedrock environment. During the process of biostimulation with the help of ethanol, the overall concentration of cis-dischloroethene (cDCE) and vinyl chloride (VC) increases under the bedrock environment. This production of VC and cDCE help in the enrichment of the vcrA (VC reductive dehalogenase) and thereby helping to increase the bioremediation and fermenting capacity of the bacteria under the bedrock environment. Under the light of the comparative study, it can be highlighted that the use of the microorganism for the chlorinated contaminants different on the basis of the use of media or the environment over which the bioremediation is occurring. 

Conclusion

Thus from the above discussion, it can be concluded that bioremediation is an important process that helps in the removal of the toxic pollutants present in the environment like the hydrocarbons and other chlorinated compounds under the action of the microbes. The microbes like bacteria and fungi play an important role in the process of conduction of bioremediation. The microorganism conduct aerobic and anaerobic metabolism in order to breakdown or oxidised the bio-pollutants. The main activities used by the microorganism include melanine, the process of co-metabolism and by the process of conduction of the genetic modifications. However, co-metabolism resists the process of bioremediation by increasing the salinity of the soils and creating an adverse environment for thriving of the microbes. In such cases fungi containing melanine or other fungi that are capable to thriving under extreme climatic conditions can be used to conduct the process of bioremediation.

Recommendation

Alternative techniques for the promotion of bioremediation by microorganisms

Immobilization approaches

Partovinia and Rasekh (2018) recommended that in order to bring a cost-effective change in the microbial mechanism effective use of the immobilization technique can be proved to be helpful under the harsh environments for example the environment that are high on deposition of the hydrocarbons. The use of the immobilization environment is advantageous in comparison to the genetic approach of the gene manipulation or the use of the external supply of the inorganic fertilizers. Immobilization process is cost-effective and ensures easier separation of the microorganism from the degraded byproducts. It also helps in the reutilization of the microorganism unlike the process of external supply of inorganic microorganism where the microorganisms die due to increase in the level of salinity of soli. The immobilization process also helps to increase the pH tolerance of the soil and helps the bacteria to conduct the process of bioremediation under extreme temperature conditions. The study of Li et al. (2016) also highlighted that immobilized consortium help to increase the overall bioremediation activity of the microbes.

Use of fungi for bioremediation

In order to promote effective bioremediation of the chlorinated microorganism Marco-Urrea, Garcia-Romera and Aranda (2015) a review study. The review highlighted that for effective bioremediation of the chlorinated toxic contaminants and polycyclic aromatic hydrocarbons, non-ligninolytic fungi can be regarded as an important alternative to bacteria. One of the non-ligninolytic fungi that has evolved as a potential bioremediation agent in the study is white-rot fungi. This particular fungi is important agent for the process of conducting bioremediation process because of the presence of unspecific oxidative enzymes. White-rot fungi that belong to the genus of Ascomycota and Zygomycota phylum have the capability to undergo enzymatic transformation of the environmental pollutants due to presence of the non-specific oxidative enzyme. This enzymatic transformation helps the fungi to overcome the some of the limitations experienced by the bacteria like incapability to thrive under extreme pH, drought condition or other adverse environmental condition. The presence of the  such enzymatic capacity helps them to successfully conduct the process of bioremediation of the chlorinated hydrocarbons and polycyclic aromatic hydrocarbons. However, there is little knowledge in the domain of degradation pathways and the enzymatic mechanisms used for the fungi in order to conduct the bioremediation process. Further research is required to be conducted in this filed in order to highlight the underlying enzymatic mechanism of fungi in conducting the bioremediation of chlorinated hydrocarbons (Marco-Urrea, Garcia-Romera & Aranda, 2015).

Use of nanotechnology

Bhandari (2018) is of the opinion that the use of nanotechnology is the new emerging field in the domain of effective bioremediation. The overall results of the use of nanotechnology for the use of the bioremediation is cost effective and is fast in comparison to the other conventional approaches for the process of bioremediation like the sue of naturally microorganisms. Nanotechnology with their enhanced surface area, transport properties and other special sequestering characteristics is prove to be effective in conducting fat and productive bioremediation process. At present research is being conducted at the nano scale by the use of the zero-valent iron and other carbon nanotubes and other nano fibres in order to found new process of remediation for a different variety of contaminants like the hydrocarbons, chlorinated compounds, heavy metals and other organic compounds.

Implementation

The implementation of the process of bioremediation can be done by genetic engineers and biological engineers. The role of the engineers will be employ cost-effective measures of the designing of the biomarkers in order to bring the genetic modifications in the bacteria or the other microbes and thereby helping to gain a cost effective bioremediation process. Moreover, nanotechnologists will also be involved in order to made special nano-particles for the promotion of the bio-remediation process. Varjani (2017) are of the opinion that proper participation of scientists from different fileds will be helpful in bringing change in the process of bioremediation, making it cost-effective. Time will require at least one year in order erect proper experiment.

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