In the previous section, microbial remediation was studied. Bioremediation is a waste management technique to neutralize pollutants from a contaminated site. This is done with the use of organisms which can be microorganisms, fungi, plants or algae. They either metabolize pollutants to less toxic forms, assimilate or immobilize. Bioremediation works in two ways. First is enhanced growth of suitable organisms at the site. Other way is by introducing specialized microbes at the contaminated site to degrade contaminants. Naturally occurring organisms possess genes with remediating properties. However, using genetic engineering techniques is advantageous to transfer these genes to other organisms (Joutey et al., 2013). Such techniques help increase the bioremediation efficiency.
Need for genetic engineering in bioremediation
The natural attenuation or intrinsic bioremediation process of environmental cleanup are efficient. However, they are slow in restoring the perfect environmental balance after contamination (Lendvay et al., 2003). Large scale dumping of pollutants due to industrialization is taking place everywhere today. Furthermore, The rate of natural attenuation processes are not fast enough to restore balance in the environment. Therefore, there is a need to find efficient methods, with low costs and capable of highly accelerated remediation.
Creating an efficient bioremediation process for environmental clean-up
Efficient bioremediation process depends on environmental factors. It also depends on organism’s growth characteristics and substrate needs. Therefore, genetic advantage of one organism can be transferred to another capable of accelerated growth and low substrate need. Genetic engineering is used to construct new strains of organisms, like microbes, plants or algae (Kulshreshtha, 2013). For microbes, these are known as Genetically Engineered Microorganisms (GEM). They have unique characteristics as compared to the wild type strains. They possess broad spectrum catabolic potential for bioremediation of xenobiotics (Hassani et al., 2014).
Using genetically engineered microorganisms for bioremediation is environmental friendly. It is an effective and economical clean-up technique for the remediation of pollutants. Furthermore, these techniques are also advantageous over traditional methods. This is because they increase molecular biodiversity and improve chemical selectivity of enzymes (Han, 2005). Genetic modification techniques have often resulted in a wide variety of current and promising applications for bioremediation process.
Target processes of genetic engineering in microbial remediation
In order to improve bioremediation efficiency, different molecular processes within a microbial cell can be targeted using genetic engineering. This can include introducing a new gene or upregulating the expression of an existing gene. The latter is done using regulator sequences. These genes are selected based on a number of factors. These include:
- the goal of bioremediation,
- the target pollutant,
- the organism and
- the molecular process needed for remediating.
The figure below shows different molecular targets that can be considered in all organisms which depend upon the goal of bioremediation.
There are four main processes for enhancing bioremediation efficiency and capacity (Joutey et al., 2013):
- Pathway construction, modification and regulation: To develop new molecular pathways for remediating a pollutant molecule.
- Modifying enzyme specificity: To modify enzymes and improve their pollutant binding capacities.
- Bioprocess (biological systems) development, monitoring and control: To develop microbial systems that can be commercially applied.
- Biosensor development: To use microbial cells for chemical testing and end point analysis of pollutants.
Table below shows examples of several engineered bacteria capable of expressing metal remediating molecules.
|No.||Metal||Bacterial Species||Gene Expressed||Removal Efficiency|
|1||Arsenic||E.coli||Metalloregulatory protein ArsR||100.|
|2||Chromium(VI)||Methylococcus capsulatus||CrR protein||100.|
|4||Nickel||P.fluorescens||Phytochelatin synthase (PCS)||80.|
Genetically modified microbes capable of heavy metal remediation (Dixit et al., 2015)
Target areas of genetic engineering in phytoremediation
Plants are capable of uptake of pollutants. However, they don’t have the necessary mechanism for metabolising or mineralising them but this is where phytoremediation fills the gap. The term ‘phytoremediation’ refers to using plants and their associated microbes to clean up toxic substances in the environment (Enviromental Protection Agency, 2010). Plants can achieve this through phytoextraction, phytovolatilization, detoxification and sequestration (Joutey et al., 2013). Those suitable for phytoextraction possess the following qualities:
- high capacities for absorption,
- root or shoot translocation,
- detoxification of the metals or non-metals to be extracted and
- a high biomass and preferably a rapid growth rate (Verbruggen, LeDuc and Vanek, 2009).
Using genetic engineering capable of uptake, transport or metabolism of pollutants can be introduced in plants. As a result, such genetically modified plants can then show higher efficiencies in remediation. Therefore, relevant genes from bacterial sources can be genetically engineered into candidate plants. This will enable successful metabolizing of pollutants (Jafari et al., 2013).
Genetic manipulation in plants
Genetic manipulation in plants has aimed to improve accumulation, tolerance and detoxification capacities of high biomass and rapidly growing plants. As a result, optimisation of phytoextraction process takes place (Hassani et al., 2014). With respect to heavy metal phytoremediation, metallothionein (MT) genes have been cloned. They have also been introduced into several plant species. Phytochelatins are short and cysteine-rich peptides. They offer many advantages over metallothioneins due to their structural characteristics and higher metal binding capacity. For example, transfer of human MT-2 gene in tobacco or rapeseed resulted in plants with enhances Cadmium tolerance. The target areas also include metal transporters known to alter metal tolerance and accumulation in plants. (Verbruggen, LeDuc and Vanek, 2009).
A unique source of phytoremediation technology is also the properties of plant growth promoting rhizobacteria. They provide several mechanisms of increased phytoremediation to the plant. Image 2 below shows the different mechanisms possessed by the rhizobacteria microbes applicable in phytoremediation.
Role of CRISPR-Cas9 in bioremediation technology
A wide range of genetic engineering tools exist. They are capable of efficiently introducing new foreign sequences and genes into a target organism. Of these, the Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)-Cas9 system is a highly attractive tool. CRISPR are short segments of prokaryotic DNA and they contain short, repetitive base sequences. These segments in combination with the Cas9 enzyme are capable of targeting highly specific sequence of DNA for gene insertion (Didovyk et al., 2016). Also, they help to introduce a wide range of genes in different candidate organisms not explored before. This is because of the versatility of the gene editing tool. This can enable transferring remediating traits to organisms possessing certain unique qualities. Moreover, CRISPR-Cas9 system can also allow for high efficient mode of genetic engineering within a wide range of species.
A popular way of increasing the efficacy of bioremediation is through the use of genetic engineering. This will produce microorganisms capable of degrading specific contaminants. Another way is to enhance such processes in the native organisms. However, researchers need to access and predict the consequences before releasing a GMO into the environment. In order to determine how released GMOs are affecting the environment, it is necessary to be able to detect and enumerate them in complex samples. In addition to the GMO itself, it is useful to track the recombinant DNA with which the GMO has been engineered. This will help monitor potential loss of these genes and their possible horizontal transfer to other microorganisms.
- Didovyk, A. et al. (2016) ‘Transcriptional regulation with CRISPR-Cas9: Principles, advances, and applications’, Current Opinion in Biotechnology. Elsevier Ltd, 40, pp. 177–184. doi: 10.1016/j.copbio.2016.06.003.
- Dixit, R. et al. (2015) ‘Bioremediation of heavy metals from soil and aquatic environment: An overview of principles and criteria of fundamental processes’, Sustainability (Switzerland), 7(2), pp. 2189–2212. doi: 10.3390/su7022189.
- Enviromental Protection Agency (2000) ‘Introduction to Phytoremediation’, U.S. Environmental Protection Agency, (February), pp. 1–72. doi: EPA/600/R-99/107.
- Han, L. (2005) ‘Genetically Modified Microorganisms: Development and Applications’, in Parekh, S. R. (ed.) The GMO Handbook: Genetically Modified Animals, Microbes, and Plants in Biotechnology. Totowa, NJ: Humana Press Inc., pp. 29–51.
- Hassani, A. H. et al. (2014) ‘Phytoremediation of Soils Contaminated with Heavy Metals Resulting from Acidic Sludge of Eshtehard Industrial Town using Native Pasture Plants’, Journal of Environment and Earth Science, 4(19), pp. 87–94.
- Jafari, M. et al. (2013) ‘Bioremediation and Genetically Modified Organisms’, in Fungi as Bioremediators. Springer Berlin Heidelberg, pp. 433–451. Available at: https://www.researchgate.net/profile/Morad_Jafari/publication/258979391_Bioremediation_and_Genetically_Modified_Organisms/links/5639e9dc08aed5314d239f83/Bioremediation-and-Genetically-Modified-Organisms.pdf.
- Joutey, N. T. et al. (2013) ‘Biodegradation: Involved Microorganisms and Genetically Engineered Microorganisms’, in Chamy, I. and Rosenkranz, F. (eds) Biodegradation- Life of Science. InTech, pp. 289–320. doi: 10.5772/56194.
- Kulshreshtha, S. (2013) ‘Genetically Engineered Microorganisms: A Problem Solving Approach for Bioremediation’, Journal of Bioremediation and Biodegradation, 4(4), pp. 1–2. doi: 10.4172/2155-6199.1000e133.
- Lendvay, J. . et al. (2003) ‘Bioreactive barriers: a comparison of bioaugmentation and biostimulation for chlorinated solvent remediation’, Environmental Science & Technology, 37(7), pp. 1422–1431.
- Verbruggen, N., LeDuc, D. and Vanek, T. (2009) ‘Potential of Plant Genetic Engineering for Phytoremediation of Toxic Trace Elements’, in Phytotechnologies Solutions for Sustainable Land Management. EOLSS Publishers, pp. 1–24. Available at: http://www.eolss.net/Sample-Chapters/C09/E6-199-12-00.pdf.