(86c) Biomolecular And Statistical Comparison Of Two Field-Scale Sulfate-Reducing Bioreactors Treating Acid Mine Drainage

The microbial communities of two field-scale sulfate-reducing bioreactors treating acid mine drainage (AMD) were compared using biomolecular tools and multivariate statistical analyses. The two bioreactors, Luttrell and Peerless Jenny King (PJK), were highly suitable for this study because of their similar geographic locations and substrate compositions. The main differences between the two bioreactors were the quality of influent AMD, the configuration of the bioreactors, and the degree of exposure to aerobic conditions.

The microbial communities of the two bioreactors were found to be functionally similar. Clone libraries of the 16S rRNA gene revealed the presence of the three groups of bacteria typically associated with compost-based sulfate-reducing remediation systems (cellulose degraders, fermenters, and sulfate-reducing bacteria (SRB)) at both sites. In addition to the three main groups, a high diversity of species covering a broad phylogeny were also detected at both sites, including denitrifiers, rhizosphere microorganisms, acetogenic bacteria, metal-reducing species, and AMD-generating bacteria.

A narrowed analysis scope of the SRB community was performed by targeting the adenosine 5'-phosphosulfate reductase (apsA) gene. Clone libraries of the apsA gene revealed that the Luttrell site was dominated by uncultured SRB most closely related to Desulfovibrio spp., while the PJK site was dominated by Thiobacillus denitrificans and other uncultured Thiobacillus spp. T. denitrificans is a facultative aerobe known to use oxygen or nitrate as electron acceptors, and all Thiobacillus spp. can oxidize inorganic Fe(II) and sulfur compounds, which would effectively reverse the desired AMD remediation process. The fractions of two genera of SRB, Desulfovibrio and Desulfobacterium, were also higher at Luttrell compared to PJK as determined by real-time quantitative polymerase chain reaction (Q-PCR).

Principle component analysis (PCoA) and significance testing based on the phylogenetic alignment of 16S rRNA gene sequences yielded no obvious clustering or separation between the two field sites. No significant differences between the overall communities of the two sites (p-value = 0.07) were obtained based on the 16S rRNA gene sequences. PCoA of the SRB community, based on the apsA gene sequences, revealed a strongly bimodal clustering between the two sites, with all sampling locations from the Luttrell field site grouped tightly, while the sampling locations from the PJK bioreactors were grouped separately. This separation was clearly along the first principal coordinate, which explained 82% of the variance in the data. Further analysis indicated significant differences between the two bioreactors (p-value = 0.00) based on the apsA gene sequences.

This study is the first comparison of two field-scale bioreactors treating AMD with both inter- and intra-reactor differences statistically analyzed, and further develops the use of biomolecular tools for the study of microbially driven remediation systems.