Reflection on Molecular Approaches Influencing State-of-the-Art Bioremediation Design: Culturing to Microbial Community Fingerprinting to Omics

Abstract

Bioremediation is generally viewed as a cost effective and sustainable technology because it relies on microbes to transform pollutants into benign compounds. Advances in molecular biological analyses allow unprecedented microbial detection and are increasingly incorporated into bioremediation. Throughout history, state-of-the-art techniques have informed bioremediation strategies. However, the insights those techniques provided were not as in depth as those provided by recently developed omics tools. Advances in next generation sequencing (NGS) have now placed metagenomics and metatranscriptomics within reach of environmental engineers. As NGS costs decrease, metagenomics and metatranscriptomics have become increasingly feasible options to rapidly scan sites for specific degradative functions and identify microorganisms important in pollutant degradation. These omic techniques are capable of revolutionizing biological treatment in environmental engineering by allowing highly sensitive characterization of previously uncultured microorganisms. Omics enables the discovery of novel microorganisms for use in bioaugmentation and supports systematic optimization of biostimulation strategies. This review describes the omics journey from roots in biology and medicine to its current status in environmental engineering including potential future directions in commercial application.

Figure 1

Coding function of the cell illustrated by a simplified cross section of a generic bacterium (a). Most bacteria contain at least two important features to aid in the genetic process-the chromosome and the ribosome. The chromosome consists of DNA and is where transcription takes place (b). DNA consists of adenine (A), thymine (T), cytosine (C) and guanine (G) bases. These bases form bonds with the bases on the opposite strand: A always pairs with T and C always pairs with G. When the microorganism needs to carry out a function, RNA polymerase transcribes the section needed into a single-stranded RNA transcript (and substitutes uracil (U) for T). The RNA transcript then directs the ribosome to assemble amino acids into proteins (c).

Figure 2

Molecular tools used to inform studies in bioremediation and wastewater treatment throughout the past few decades according to advanced searches of the Web of Science database. Query topics for each decade and technology were as follows: TS=(Cloning AND bioremediation) OR TS=(cloning AND wastewater AND treatment); TS=(DGGE AND bioremediation) OR TS=(DGGE AND wastewater AND treatment); TS=((quantitative PCR) AND bioremediation) OR TS=((quantitative PCR) AND wastewater AND treatment) OR TS=((reverse transcript* quantitative PCR) AND bioremediation) OR TS=((reverse transcript* quantitative PCR) AND wastewater AND treatment); TS=((ion torrent) AND bioremediation) OR TS=((Ion torrent) AND wastewater AND treatment) OR TS=((Illumina) AND bioremediation) OR TS=((Illumina) AND wastewater AND treatment) OR TS=((nanopore) AND bioremediation) OR TS=((nanopore) AND wastewater AND treatment) OR TS=((pacific biosciences) AND bioremediation) OR TS=((pacific biosciences) AND wastewater AND treatment).

Figure 3

Chronology of Key Discoveries in the Field of Microbiology and Molecular Biology Relevant to Bioremediation.

Figure 4

Genetic cloning overview. Genetic cloning consists of three steps. First, the unculturable DNA fragment is attached to a well-studied host organism’s circular DNA (a). Then, the altered DNA is inserted into the host organism (b). Finally, the host organism multiplies, copying the unculturable fragment (c).

Figure 5

Sanger Sequencing. Sanger sequencing is a common method used to sequence DNA, one strand at a time. First, the strands must be separated (a). Then, a primer marks the place where DNA polymerase begins adding nucleotides (b). Four sequencing reactions happen, one for each base A,T,C, and G. In the G reaction (c), DNA polymerase incorporates dATP, dCTP, dGTP, and dTTP until it adds a chain-inhibiting ddGTP to complement the C base, stopping the process. Here, a ddGTP is incorporated to complement the 7^th and 8^th base, inferring that the 7^th and 8^th bases in the sequence are Cs. (d) Incorporating these chain-inhibiting bases creates fragments of different sizes from each of the four different reactions. These different fragments are then sorted by length and their complementary bases inferred (the G reaction fragments tell the location of the Cs, the As give T locations, etc.).

Figure 6

Polymerase Chain Reaction