Phylogenetic Analysis of Microbial Communities in Contaminated Nearshore Marine Sediments

EPA Grant Number: R826108
Title: Phylogenetic Analysis of Microbial Communities in Contaminated Nearshore Marine Sediments
Investigators: Herwig, Russell P.
Institution: University of Washington
EPA Project Officer: Hahn, Intaek
Project Period: September 1, 1997 through August 31, 2000 (Extended to September 30, 2001)
Project Amount: $370,446
RFA: Exploratory Research - Environmental Biology (1997) RFA Text |  Recipients Lists
Research Category: Biology/Life Sciences , Aquatic Ecosystems


Little is known about the composition of microbial communities that live in contaminated nearshore marine sediments and how the composition in contaminated sediments compares to that found in clean reference sites. In recent years my laboratory has developed protocols for the analysis of the microbial communities in marine sediments. With support from the EPA we will continue to develop our protocols and examine the microbial composition of (a) marine sediments contaminated with creosote, a contaminant composed largely of polycyclic aromatic hydrocarbons (PAHs); (b) clean reference marine sites; (c) and a contaminated site that was capped with a layer of clean sediment. The primary objective of our research is to develop and test protocols for performing "large-scale" analyses on the composition of the microbial communities in nearshore marine sediments from Puget Sound and other marine sites around the United States using 16S rRNA phylogenetic techniques. This objective will be pursued so the following hypotheses can be addressed during our investigation and in the future:
Hypothesis 1. The composition of the microbial communities in contaminated marine sediments is different that that found in clean reference sediments.
Hypothesis 2. Indicator species of microorganisms, that may or may not be able to degrade pollutants, are present in contaminated marine sediments.
Hypothesis 3. The compositions of the microbial communities change or evolve in sediment sites that are recovering or undergoing intrinsic bioremediation.


To begin our study, samples will be collected from at least four sites in Puget Sound, Washington: two sites that are contaminated with PAHs introduced by a former wood treatment facility and two sites with control reference sediments. Samples may also be collected a "sediment cap" that was placed over PAH-contaminated sediments. Microbial DNA will be extracted and purified from sediment samples using a protocol developed in my laboratory. The 16S rDNA from sediment microorganisms will be amplified with universal primers. We will develop a protocol for screening and clustering 500 to 1,000 16S rDNA clones from each sediment sample by restriction fragment length analysis using capillary gel electrophoresis or conventional gel electrophoresis, and representative 16S rDNA clones will be sequenced on an automated sequencer. A phylogenetic analysis of the representative clones will be performed using the maximum likelihood method. To examine the representation of the 16S rDNA clones within the sediment community, "nested" 16S rDNA oligonucleotide probes will be designed to enumerate the most numerous species and major phylogenetic groups within the various sediment samples.

Expected Results:

The composition of the microbial communities from contaminated and reference nearshore marine sediments will be described. Protocols will be developed to allow for the screening of a large number of clones that are representative of the 16S rRNA from environmental microorganisms. A better understanding about the impact of environmental pollution on the structure of the microbial community in marine sediments.

Supplemental Keywords:

PAH, PNA, bioremediation, phylogenetics, microbial ecology, microbiology, RNA, Northwest, Washington, WA, Puget Sound, Region 10, Eagle Harbor., Scientific Discipline, Toxics, Geographic Area, Waste, Water, Contaminated Sediments, State, HAPS, Environmental Microbiology, Ecology and Ecosystems, Ecological Risk Assessment, Biology, contaminated marine sediment, contaminant transport, contaminated sediment, Washington (WA), microbial pollution, aquatic ecosystems, phylogenic analysis, ecology assessment models, indicator species, marine ecosytems

Progress and Final Reports:

  • 1998
  • 1999 Progress Report
  • 2000 Progress Report
  • 2001
  • Final