Pyrosequence Analysis of Unamplified and Whole Genome Amplified DNA from Hydrocarbon-Contaminated Groundwater

Pyrosequence data was used to analyze the composition and metabolic potential of a metagenome from a hydrocarbon-contaminated site. Unamplified and whole genome amplified (WGA) sequence data was compared from this source. According to MG-RAST, an additional 2,742,252 bp of DNA was obtained with the WGA, indicating that WGA has the ability to generate a large amount of DNA from a small amount of starting sample. However, it was observed that WGA introduced a bias with respect to the distribution of the amplified DNA and the types of microbial populations that were accessed from the metagenome. The dominant order in the WGA metagenome was Flavobacteriales, whereas the unamplified metagenome was dominated by Actinomycetales as determined by RDPII and CARMA databases. According to the SEED database, the subsystems shown to be present for the individual metagenomes were associated with the metabolic potential that was expected to be present in the contaminated groundwater, such as the metabolism of aromatic compounds. A higher percentage (4.4) of genes associated with the metabolism of aromatic compounds was identified in the unamplified metagenome when compared to the WGA metagenome (0.66%). This could be attributed to the increased number of hydrocarbon degrading bacteria that had been accessed from this metagenome (Mycobacteria, Nocardia, Brevibacteria, Clavibacter, Rubrobacter, and Rhodoccocus). Therefore, it was possible to relate the taxonomic groups accessed to the contamination profile of the metagenome. By collating the sequencing data obtained pre- and post-amplification, this study provided insight regarding the survival strategies of microbial communities inhabiting contaminated environments.