Microbial functional gene diversity with a shift of subsurface redox conditions during In Situ uranium reduction

Abstract

To better understand the microbial functional diversity changes with subsurface redox conditions during in situ uranium bioremediation, key functional genes were studied with GeoChip, a comprehensive functional gene microarray, in field experiments at a uranium mill tailings remedial action (UMTRA) site (Rifle, CO). The results indicated that functional microbial communities altered with a shift in the dominant metabolic process, as documented by hierarchical cluster and ordination analyses of all detected functional genes. The abundance of dsrAB genes (dissimilatory sulfite reductase genes) and methane generation-related mcr genes (methyl coenzyme M reductase coding genes) increased when redox conditions shifted from Fe-reducing to sulfate-reducing conditions. The cytochrome genes detected were primarily from Geobacter sp. and decreased with lower subsurface redox conditions. Statistical analysis of environmental parameters and functional genes indicated that acetate, U(VI), and redox potential (E(h)) were the most significant geochemical variables linked to microbial functional gene structures, and changes in microbial functional diversity were strongly related to the dominant terminal electron-accepting process following acetate addition. The study indicates that the microbial functional genes clearly reflect the in situ redox conditions and the dominant microbial processes, which in turn influence uranium bioreduction. Microbial functional genes thus could be very useful for tracking microbial community structure and dynamics during bioremediation.

Fig 1

Layout of the experimental plots at the Old Rifle uranium mill tailings site. Each plot had 5 injection wells (open circles) perpendicular to groundwater flow, 4 monitoring wells (filled circles) down-gradient of acetate injection, and 1 monitoring well positioned up-gradient of the injection wells (filled triangle). The 2004 experimental plot was maintained under Fe-reducing conditions, and the 2005 experimental plot was driven to sulfate-reducing conditions by using different durations of biostimulation.

Fig 2

Hierarchical cluster analysis of all functional genes detected (a). Genes that were present in at least three time points were used for cluster analysis. Results were generated in CLUSTER and visualized using TREEVIEW. Red indicates signal intensities above background, while black indicates signal intensities below background. Brighter red coloring indicates higher signal intensities. A total of 6 major groups were observed (b). The numbers equal groupings found among the hybridization patterns.

Fig 3

Functional gene abundance in the background well (B05), the Fe-reducing well (M16), and the shift from Fe-reducing to sulfate-reducing wells (M21 and M24). Abbreviations: Org, organic contaminant degradation; Nred, nitrogen reduction; Nit, nitrification; Nfix, nitrogen fixation; methane, methane generation; Met, metal reduction and resistance; DSR, sulfate reduction; Cfix, carbon fixation; Cdeg, carbon degradation.

Fig 4

Hierarchical clustering of c-type cytochrome genes. Red indicates signal intensities above background, while black indicates signal intensities below background. Brighter red coloring indicates higher signal intensities. The bar of colors below the sample names indicates the average signal intensities of each sample. Mantel test results indicated significant correlations between the functional gene patterns and the U(VI) concentration (P < 0.05).

Fig 5

Canonical correspondence analysis (CCA) of total functional genes and geochemical data.