Growth of Dehalococcoides spp. and increased abundance of reductive dehalogenase genes in anaerobic PCB-contaminated sediment microcosms

Polychlorinated biphenyls (PCBs) contaminate 19% of US Superfund sites and represent a serious risk to human and environmental health. One promising strategy to remediate PCB-contaminated sediments utilizes organohalide-respiring bacteria (OHRB) that dechlorinate PCBs.

However, functional genes that act as biomarkers for PCB dechlorination processes (i.e., reductive dehalogenase genes) are poorly understood. Here, we developed anaerobic sediment microcosms that harbor an OHRB community dominated by the genus Dehalococcoides. During the 430-day microcosm incubation, Dehalococcoides 16S rRNA sequences increased two orders of magnitude to 10⁷ copies/g of sediment, and at the same time, PCB118 decreased by as much as 70%. In addition, the OHRB community dechlorinated a range of penta- and tetra-chlorinated PCB congeners including PCBs 66, 70?+?74?+?76, 95, 90?+?101, and PCB110 without exogenous electron donor. We quantified candidate reductive dehalogenase (RDase) genes over a 430-day incubation period and found rd14, a reductive dehalogenase that belongs to Dehalococcoides mccartyi strain CG5, was enriched to 10⁷ copies/g of sediment. At the same time, pcbA5 was enriched to only 10⁵ copies/g of sediment. A survey for additional RDase genes revealed sequences similar to strain CG5’s rd4 and rd8. In addition to demonstrating the PCB dechlorination potential of native microbial communities in contaminated freshwater sediments, our results suggest candidate functional genes with previously unexplored potential could serve as biomarkers of PCB dechlorination processes.

     

Identification of a novel hydroxylated metabolite of 2,2′,3,5′,6-pentachlorobiphenyl formed in whole poplar plants

Polychlorinated biphenyls (PCBs) are a group of persistent organic pollutants consisting of 209 congeners. Oxidation of several PCB congeners to hydroxylated PCBs (OH-PCBs) in whole poplar plants has been reported before. Moreover, 2,2′,3,5′,6-pentachlorobiphenyl (PCB95), as a chiral congener, has been previously shown to be atropselectively taken up and transformed in whole poplar plants. The objective of this study was to determine if PCB95 is atropselectively metabolized to OH-PCBs in whole poplar plants. Two hydroxylated PCB95s were detected by high-performance liquid chromatography-mass spectrometry in the roots of whole poplar plants exposed to racemic PCB95 for 30 days. The major metabolite was confirmed to be 4′-hydroxy-2,2′,3,5′,6-pentachlorobiphenyl (4′-OH-PCB95) by gas chromatography-mass spectrometry (GC-MS) using an authentic reference standard. Enantioselective analysis showed that 4′-OH-PCB95 was formed atropselectively, with the atropisomer eluting second on the Nucleodex β-PM column (E2-4′-OH-PCB95) being slightly more abundant in the roots of whole poplar plants. Therefore, PCB95 can at least be metabolized into 4′-OH-PCB95 and another unknown hydroxylated PCB95 (as a minor metabolite) in whole poplar plants. Both atropisomers of 4′-OH-PCB95 are formed, but E2-4′-OH-PCB95 has greater atropisomeric enrichment in the roots of whole poplar plants. A comparison with mammalian biotransformation studies indicates a distinctively different metabolite profile of OH-PCB95 metabolites in whole poplar plants. Our observations suggest that biotransformation of chiral PCBs to OH-PCBs by plants may represent an important source of enantiomerically enriched OH-PCBs in the environment.     

Localization of hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX) and 2,4,6-trinitrotoluene (TNT) in poplar and switchgrass plants using phosphor imager autoradiography

Phosphor imager autoradiography is a technique for rapid, sensitive analysis of the localization of xenobiotics in plant tissues. Use of this technique is relatively new to research in the field of plant science, and the potential for enhancing visualization and understanding of plant uptake and transport of xenobiotics remains largely untapped. Phosphor imager autoradiography is used to investigate the uptake and translocation of the explosives 1,3,5-trinitro-1,3,5-triazine (RDX) and 2,4,6-trinitrotoluene within Populus deltoides × nigra DN34 (poplar) and Panicum vigratum Alamo (switchgrass). In both plant types, TNT and/or TNT-metabolites remain predominantly in root tissues while RDX and/or RDX-metabolites are readily translocated to leaf tissues. Phosphor imager autoradiography is further investigated for use in semi-quantitative analysis of uptake of TNT by switchgrass.     

How many samples are required? Evaluating a model for verification of carbon sequestration in a hybrid poplar buffer strip

A model is developed to predict annual and total above-ground carbon storage within a hybrid poplar stream buffer. The regression model predicts tree wet weight based upon circumference at breast-height (137 cm) with an r-square value of 0.9922. Carbon storage in above-ground biomass is estimated to be 3.57 to 3.71 metric tons per hectare, with a measured annual increment of 0.92 to 1.37 metric ton per hectare per year. The variability of carbon storage within this biological system, including soil organic matter, is explored, and the number of samples required to achieve a desired level of statistical certainty are predicted. As has been investigated previously for other biological systems (Garten and Wullschleger, 1999), the study shows that a prohibitively large number of samples must be taken in order to achieve high degrees of certainty about mean carbon storage values. The study also shows, however, that mean values with somewhat greater uncertainties can easily be achieved with much smaller sample sizes. Thus carbon sequestration verification might be accomplished cost-effectively if the degree of certainty required is not unrealistically high for highly variable natural systems.     

Microtox toxicity test: detoxification of TNT and RDX contaminated solutions by poplar tissue cultures

Poplar (Populus deltoidesxnigra DN34) tissue cultures removed 2,4,6-trinitrotoluene (TNT) from an aqueous solution in five days, reducing the toxicity of the solution from highly toxic Microtox EC value to that of the control. 1,3,5-Trinitro-1,3,5-triazacyclohexane (RDX) was taken up by the plant tissue cultures more slowly, but toxicity reduction of the solution was evident. The measurement of toxicity reduction of aqueous solutions containing TNT and RDX was performed using a novel methodology developed for use with the Microtox testing system. Radiolabeled TNT and RDX were used to confirm removal of explosives from hydroponic solutions containing plant tissue cultures and to verify that toxicity did not change in solutions where no plant cultures were present (positive controls). High Performance Liquid Chromatography (HPLC) and Liquid Scintillation Counter (LSC) measurements confirmed removal of TNT and RDX from solutions containing poplar plant tissue cultures and constancy of the plant-free controls. In addition, metabolites were identified in remediated solutions by HPLC, confirming the mechanism by which plants can remediate groundwater, surface water, and soil solutions.     

Benzo[a]pyrene degradation by Sphingomonas yanoikuyae JAR02

Batch experiments were conducted to characterize the degradation of benzo[a]pyrene, a representative high molecular weight (HMW) polycyclic aromatic hydrocarbon (PAH), by Sphingomonas yanoikuyae JAR02. Concentrations up to the solubility limit (1.2 microg l(-1)) of benzo[a]pyrene were completely removed from solution within 20 h when the bacterium was grown on salicylate. Additional experiments with [(14)C]7-benzo[a]pyrene demonstrated 3.8% mineralization over 7 days when salicylate was present is solution, and one major radio-labeled metabolite was observed that accounted for approximately 10% of the initial radio-label. Further characterization of the radio-labeled metabolite using HPLC/MS and HPLC/MS/MS identified radio-labeled pyrene-8-hydroxy-7-carboxylic acid and unlabeled pyrene-7-hydroxy-8-carboxylic acid as novel ring-cleavage metabolites, and a benzo[a]pyrene degradation pathway was proposed. Results indicate that biostimulation of HMW PAH degradation by salicylate, a water-soluble, non-toxic substrate, has significant potential for in situ bioremediation.     

DISSOLVED OXYGEN MODEL OF A SHORT DETENTION TIME RESERVOIR WITH ANAEROBIC HYPOLIMNION1

ABSTRACT: A vertical dissolved oxygen model was calibrated and verified using independent field data sets from 1972 and 1973 in Lake Lyndon B. Johnson (LBJ), a short detention time reservoir with anaerobic hypolimnion. Ammonia, carbonaceous BOD, conductivity, and temperature were also simulated as state variables. The conductivity results provided a check on the mass balance and the method of entering the inflows to the reservoir model. Emphasis was placed on calculating the sinks of dissolved oxygen in the hypolimnion which could be useful in management decisions.     

A methodology to estimate carbon storage and flux in forestland using existing forest and soils databases

Sequestration of carbon through expansion and management offorestland can assist in reducing greenhouse gas concentrationsin the atmosphere. Quantification of the amount of carbonpresents an ongoing challenge that calls for new approaches.These new approaches must seek to simplify the science-basedaccounting of carbon storage and flux, while adhering to generalprinciples of greenhous gas accounting. Quantifying change incarbon storage and carbon flux consists of two steps: developinga baseline of carbon storage, and measuring resulting storageand flux following a change of conditions. A methodology isproposed that accomplishes both steps, applicable to anaggregate-level analysis using the state of Iowa (U.S.A.) as a case study. The method combines existing databasesfrom the U.S. Forest Service (USFS) and U.S. Department ofAgriculture (USDA), and merges these with the methods of Birdsey(USDA, 1992, 1995; IPCC, 1997; EIIP, 1999) for partitioningcarbon stocks into storage pools. Forested ecosystems in the study area contain approximately 137.3 metric tons organiccarbon per hectare, or 114 million metric tons of carbon inaggregate. Of this total, 44.7 million tons are stored inbiomass tissue, and 69.2 million tons of carbon are contained insoils. Carbon flux due to forests in the state of Iowa isestimated to be a net annual sequestration (removal from theatmosphere) of 4.3 million metric tons of CO₂-equivalent,approximately 5% of the net annual CO₂-equivalentemissions from the state (Ney et al., 1996).