16S rRNA Restriction Fragment Length Polymorphism Analysis of Bacterial Diversity as a Biomarker of Ecological Health in Polluted Sediments from New Bedford Harbor, Massachusetts, USA

A polymerase chain reaction (PCR)-based method was developed to compare bacterial diversity among environmental sites with varying degrees of anthropogenic impact. New Bedford Harbor, MA, a US Environmental Protection Agency-designated Superfund hazardous waste site, was studied to assess changes in bacterial diversity resulting from long-term inputs of organic and inorganic pollutants. Total DNA was extracted from surficial sediments sampled from four sites along a transect of decreasing contamination (Upper and Lower Acushnet Estuary, New Bedford Harbor, and Buzzards Bay, respectively). Oligonnucleotide primers specific to conserved regions of the 16S rRNA gene were used to PCR-amplify sequences from DNA extracts. Restriction fragment length polymorphism (RFLP) analysis resulted in generation of a number of unique operational taxonomic units (OTUs). Cluster analysis of fragment pattern data using the computer program RESTSITE® allowed for bacterial diversity estimations, which, in agreement with standard culture techniques, showed higher bacterial diversity in New Bedford Harbor sediments, relative to Buzzards Bay. By employing bacterial diversity as a sensitive indicator of environmental stress, the method has wide applicability to many environments for the assessment of anthropogenic impact on aquatic ecosystems.