Microbial communities and their enzymes involved in biodegradation of herbicides and dioxins

In the Vietnam war from 1961 to 1971, the United States military defoliated trees and plants using approximately 80 million liters of the herbicides 2,4-dichlorophenoxyacetic acid (2,4-D) and 2,4,5-trichlorophenoxyacetic acid (2,4,5-T), the last of which appeared to be contaminated with trace amounts of the highly toxic dioxin congener 2,3,7,8-tetrachlorodibenzo-p-dioxin (2,3,7,8-TCDD). Until recently, Bien Hoa airbase in South Vietnam was a highly contaminated hotspot where soils and sediments in the vicinity of the site were polluted with high concentrations of herbicides and dioxins due to large spills. The aim of the studies was to correlate the composition and dynamics of microbial communities originating from soils contaminated with herbicides and dioxins at Bien Hoa airbase in Vietnam to the biodegradation of 2,3,7,8-TCDD, 2,4-D and 2,4,5-T in order to obtain a better understanding of the role of key bacteria participating in their biodegradation, and to get insight into the best conditions for bioremediation. Our general approach was i) to enrich microbial communities from dioxin and herbicide polluted soil in Vietnam under defined conditions and to follow their community compositions over time using high throughput 16S rRNA gene profiling using the Illumina platform, and ii) to measure the degradation of 2,3,7,8-TCDD or the herbicides 2,4-D and 2,4,5-T using the DR-CALUX® bioassay or LC-MS/MS in parallel. A more specific approach was the isolation and characterization of bacteria and fungi that were dominant in some of the enrichments and which showed breakdown of 2,3,7,8-TCDD or the herbicides 2,4-D and 2,4,5-T when grown in isolation. The inspection of the genomes of some of the isolated bacterial species provided a more detailed insight in their genetic and metabolic potential to degrade these xenobiotics. Chapter 2 determined the correlation of the microbial community structure potentially contributing to 2,3,7,8-TCDD biodegradation. We also observed that bacterial genera such as Bordetella and Sphingomonas were present at high relative abundance during biodegradation of 2,3,7,8-TCDD. We inferred that these genera may play an important role in 2,3,7,8-TCDD biodegradation. Chapter 3 presents a study on the degradation of 2,4-D and 2,4,5-T by the white rot fungus (WRF) Rigidoporus sp. FMD21 and demonstrated the role played by intracellular and extracellular enzymes such as CYPs and laccases in these herbicide degradation pathways. In addition, two newly identified filamentous fungi belonging to genera Verticillium and Fusarium and isolated from heavy herbicide and dioxin-contaminated soil at Bien Hoa airbase were able to degrade 2,4-D and 2,4,5-T. Chapter 4 focuses on 2,4-D and 2,4,5-T biodegradation by bacterial cultures enriched from soil contaminated with dioxins and herbicides as collected from Bien Hoa airbase in Vietnam. It correlates changes in bacterial community structure with the degradation rate of these herbicides. Chapter 5 investigates possibly important genes involved in the degradation pathway of 2,4-D and 2,4,5-T by analysing the genomes of 4 bacteria. These bacteria were isolated from enrichment cultures with herbicide-contaminated soil collected from Bien Hoa airbase as described in chapter 4. The PhD thesis shows that the biodegradation of xenobiotics like 2,3,7,8-TCDD and the herbicides 2,4-D and 2,4,5-T may be improved by the addition of nutrients to promote the growth of organisms that are important for biodegradation, be it by metabolism or cometabolism. However, care has to be taken with the addition of extra carbon and Gibbs energy sources as this may lead to outcompetition of the specialist degraders by species growing much better on these extra sources, which may result in competition for oxygen, for vitamins or for other essential nutrients. My studies suggest that keeping a carbon starvation condition during bioremediation may well result in an efficient, low-cost and environmentally friendly remediation process for dioxin- and herbicide-contaminated soils.