Project Abstract

Overcoming the Barriers to In-Situ PCB Bioremediation: Development and Testing of Field Ready Technologies for Sequential Anaerobic-Aerobic Treatment

Principal Investigators

James M. Tiedje, Tamara V. Tsoi and John F. Quensen, III
Michigan State University
E-mail: tiedjej@pilot.msu.edu

Summary

Previously we discovered, isolated and characterized aromatic ring dechlorinases that metabolize the products of PCB metabolism in bacteria. We have also demonstrated the prove of concept, namely that we can use these genes to construct a genetically modified organism that grows on certain important PCB congeners. We are now developing field-ready technologies that would allow employing the recombinant bacteria in the two-phase anaerobic-aerobic PCB bioremediation scheme.

Introduction

We initially focused on basic research addressing the fundamental barriers limiting PCB bioremediation since 20 years of research on this subject had not provided the solution, nor had alternative remediation technologies gained acceptance. Our general remediation strategy was to utilize sequential anaerobic-aerobic treatment. The barriers to application were the slow rate of anaerobic PCB dechlorination which usually yielded incompletely dechlorinated PCBs, the inability of wild type microbes to actually grow on important PCBs in nature and the means to couple these processes in a cost-effective in-situ remediation scheme. Hence, our approach was to study the possibility of enhancing anaerobic dechlorination of PCBs in sediments, use co-substrates to enhance aerobic co-metabolism of PCBs, isolate and characterize dechlorinase genes and design the recombinant organisms that would grow on important PCB congeners, and to use the obtained information to test PCB bioremediation in soil columns and to optimize the treatment conditions as necessary to arrive at a practical field scheme for PCB bioremediation.

Research Objectives

Current objectives are characterized by a shift from the discovery phase to the developmental phase, i.g. converting our discoveries into field technologies. They include:

demonstrate the feasibility of stimulating reductive dechlorination of PCBs in soils contaminated with several Aroclors as the first phase of a sequential anaerobic/aerobic biotreatment scheme;
construct, modify and optimize stable PCB-growing organisms and develop molecular tracking protocol for use in a field experiments;
develop inoculant technology for bioaugmentation of PCB contaminated soils and test its feasibility for a successful removal of PCBs in a reductively dechlorinated sediments.

Results Major accomplishments include:

isolation, expression and structural analysis of the novel ohb genes for the three-component ortho-halobenzoate 1,2-dioxygenase, a key genes for achieving growth on the problematic ortho-chlorinated PCBs.
discovery, isolation, expression and nucleotide sequence determination of the novel reductive ortho-dechlorination rod genes, the first reductive dechlorination genes yet discovered.
engineering the recombinant pathway for mineralization of ortho- and ortho+para-chlorinated PCB congeners via combined reductive ortho- and hydrolytic para-dechlorination of aromatic ring.
enhanced anaerobic PCB dechlorination by sequential inoculation with active sediment inocula and monitoring redox potential.

Future Directions

Although some of the engineered recombinant bacteria showed sufficient stability in flask experiments, we will follow their fate in microcosm and bioreactor environments. We will continue experiments on enhancing anaerobic reductive PCB dechlorination. We will focus on bioaugmentation problem to develop an inoculant preparation scheme and carrier for our consortium of genetically modified PCB degraders that will ensure long-term survival and facilitate dispersion in soil.