Superfund Research Program

Research Support Core B: Environmental Molecular Analysis Core

Project Leader: James M. Tiedje
Grant Number: P42ES004911
Funding Period: 2006-2021

Project-Specific Links

Final Progress Reports

Year:   2020  2012 

New molecular tools (PCR primers) were designed to target genes specific to enzymes that degrade dioxins and dibenzofurans. These primers are similar to others that we developed previously, except they are more comprehensive and are designed to target functional components of the enzyme. The new primers were successfully used to identify genes encoding dioxin degrading oxygenases in a variety of pristine and industrially contaminated environmental samples. These new tools will enable us to identify novel dioxin degrading genes and characterize the ability of the encoded enzymes to destroy this class of toxic compounds.

One question crucial for environmental remediation is how the microbial community reacts to external stimuli. Dr. Zylstra and his research team approached this question by analyzing bacterial and biodegradative gene diversity among non-enriched sediments and sediment enrichments. The researchers generated tens of thousands of sequences of PCR products from sediments and corresponding enrichment cultures for 16S rRNA genes and functional genes encoding enzymes for anaerobic and aerobic aromatic ring cleavage (BamA anaerobic ring cleavage and the aerobic enzymes catechol 1,2-dioxygenase and catechol 2,3-dioxygenase). The researchers performed taxonomic classification and clustering for the 16S rRNA gene datasets using customized RDP programs including the RDP Classifier and RDP mcClust. The research team examined the functional gene sequences using ordination analysis (Non-metric Multidimensional Scaling and Principal Component Analysis) using customized Java programs and R scripts, and estimated beta diversity between sites using RDP implementations of the Chao-corrected Jaccard and Sorensen indices. The researchers found that non-enriched samples are similar, but that the enrichments grouped by sampling location.

Dr. Zylstra and his research teamanalyzed Sphingomonas wittichii strain RW1 transcriptomes to understand gene expression responses to three culture conditions: clay-Cesium (SAP), dibenzo-p-dioxin (DD) and dibenzofuran (DF). The research team discovered that DD and DF share similar suppressive effects on a number of genes involved in cell motility, intracellular trafficking, signal transduction, and amino acid metabolism. Both DD and DF induce the dbfB gene in the upper pathway for DD/DF degradation and a set of genes in catabolic routes for degrading aliphatic compounds. The effects of DD and DF were different on genes involved in transcription regulation, signal transduction, inorganic ion transport, and efflux. A large number of genes with unknown functions were found to be up-regulated by DD compared to DF. Interference with cell-to-cell and cell-to-environment interactions such as motility and chemotaxis was observed under all three conditions (DD, DF, and SAP). Most notably, DD was found to be more strongly associated with cytotoxicity and induction of stress responses than DF.