Characterization of Fe(III)-reducing enrichment cultures and isolation of Fe(III)-reducing bacteria from the Savannah River site, South Carolina

The Savannah River site, South Carolina (SRS), has been subjected to heavy metal and radionuclide contamination. Dissimilatory Fe(III)-reducing bacteria, by reducing insoluble Fe(III) to soluble Fe(II), may enhance contaminant mobility through subsurface environments. In order to investigate populations of the indigenous iron-reducing microbes from the SRS, duplicate enrichment cultures were initiated using a 10% inoculum of 7 sediment/soil samples, and serial dilutions were made into Wolfe's minimal salts media amended with 50 mM Fe(III) floc, 10 mM acetate and 0.01% yeast extract. Terminal restriction fragment length polymorphism (T-RFLP) analysis was used to generate fingerprints of the cultures and track changes in the microbial communities through the dilutions. Cluster analysis determined the relatedness of individual fingerprints. Initial enrichment cultures exhibited complex fingerprints consisting of many individual T-RF peaks, and demonstrated low similarity between sites. After four serial dilutions the fingerprints were less complex and clustered at higher similarities. Several individual T-RF peaks became dominant in a majority of the fingerprints. Cloning and sequence analysis revealed the presence of microbes closely related to Clostridium and Bacillus species and to known iron reducers such as Geobacter species and Pantoea agglomerans. Several Fe(III)-reducing isolates related to Aeromonas, Bacillus and Clostridium species were obtained.