Rhizosphere/Soil Microorganisms in the Phytoremediation of Biphenyl and Dioxane

Abstract

Biphenyl and dioxane biodegradation by poplar and willow rhizosphere microorganisms was studied in a phytoremediation test-plot contaminated with biphenyl and dioxane. A dioxane-degrading consortium enriched from the contaminated rhizosphere soil did not use dioxane as the sole source of carbon and energy, but did co-metabolize dioxane in the presence of tetrahydrofuran (THF). An isolate obtained on agar plates containing basal salts and glucose grew on glucose and co-metabolically degraded dioxane after THF degradation. The rate and extent of dioxane degradation by this particular isolate increased with increasing THF concentration. This isolate was subsequently identified as a Flavobacterium by 16S rDNA sequencing. This is the first report of a dioxane-degrading Flavobacterium which is phylogenetically distinct from any previously identified dioxane degrader. Rhizosphere microorganisms in the phytoremediation test-plot were capable of degrading biphenyl in the presence of the terminal electron acceptors (TEAs) nitrate, sulfate or carbon dioxide. TEAs (sulfate and carbon dioxide), nutrients in basal salts medium (BSM) or fertilizer enhanced biphenyl degradation. Although root exudates appeared to enhance biphenyl degradation slightly, it was not statistically significant (p > 0.10). A fungus enriched and isolated from the rhizosphere soil, was found to degrade biphenyl under anaerobic conditions only. The fungus was identified by a primer pair ITS4 and ITS1F as Pseudallescheria boydii. The microbial community in the rhizosphere of the poplar and willow trees was sampled from different soil locations in the test-plot over a 5-year period. The denaturing gradient gel electrophoresis (DGGE) results indicated that soil type had a significant impact on microbial community composition with a more diverse microbial population in native soil samples than in engineered soil samples. Although tree type had less influence on microbial diversity, diversity did decrease with time in the engineered soil of the willow rhizosphere. There was no significant influence of soil depth on the microbial community. Three aerobic biphenyl-degrading consortia were enriched from different rhizosphere soils, and the major microorganisms found in the enriched consortia were identified by 16S DNA sequencing as being in the family of Flavobacteriaceae, Alcaligenaceae, and Mycobacteriaceae. Direct polymerase chain reaction (DPCR) without prior DNA extraction and DGGE yielded the same results as PCR assays using extracted DNA in the analysis of microbial populations in dioxane- and biphenyl-degrading consortia. Therefore, the combination of DPCR and DGGE has the potential for the fast analysis of the microbial populations of environmental and clinical samples with limited diversity.