Profiling In Situ Microbial Community Structure with an Amplification Microarray

Abstract

The objectives of this study were to unify amplification, labeling, and microarray hybridization chemistries within a single, closed microfluidic chamber (an amplification microarray) and verify technology performance on a series of groundwater samples from an in situ field experiment designed to compare U(VI) mobility under conditions of various alkalinities (as HCO 3 ؊ ) during stimulated microbial activity accompanying acetate amendment. Analytical limits of detection were between 2 and 200 cell equivalents of purified DNA. Amplification microarray signatures were well correlated with 16S rRNA-targeted quantitative PCR results and hybridization microarray signatures. The succession of the microbial community was evident with and consistent between the two microarray platforms. Amplification microarray analysis of acetate-treated groundwater showed elevated levels of iron-reducing bacteria (Flexibacter, Geobacter, Rhodoferax, and Shewanella) relative to the average background profile, as expected. Identical molecular signatures were evident in the transect treated with acetate plus NaHCO 3 , but at much lower signal intensities and with a much more rapid decline (to nondetection). Azoarcus, Thaurea, and Methylobacterium were responsive in the acetate-only transect but not in the presence of bicarbonate. Observed differences in microbial community composition or response to bicarbonate amendment likely had an effect on measured rates of U reduction, with higher rates probable in the part of the field experiment that was amended with bicarbonate. The simplification in microarray-based work flow is a significant technological advance toward entirely closed-amplicon microarray-based tests and is generally extensible to any number of environmental monitoring applications.

Bicarbonate impact on U(VI) bioreduction in a shallow alluvial aquifer

“…3.7 Microbiology. Relative changes in Plot C microbiology with time are shown in Figure 8, and provide more extensive data than the amplification microarray data reported previously for this experiment (Chandler et al 2013). Relative to the CU01 background (i.e., unimpacted by acetate or bicarbonate) signatures, the bloom of metal-reducing bacteria in all wells was dominated by Geobacter and Pelobacter 16S rRNA gene signatures, as expected.…”

“…Bicarbonate-acetate and acetate-only treatments were compared microbiologically by 16S rRNA gene analysis of the groundwater. A 16S rRNA-targeted gel element 'amplification' microarray (combining amplification, labeling and hybridization in a single closed microfluidic chamber) method was used, which targets 24 genera of dissimilatory metal-, sulfate-and nitrate-reducers (Chandler et al 2013). The link between phylogeny and function in the microarray targets has been established via culture-based methods (see Andersen et al (2010) and references therein).…”

“…Here we emphasize the conservative tracer, biogeochemistry, microbiology, and U(VI) reduction results from the bicarbonate-acetate and acetate-only parts of the Super 8 field experiment. Other studies conducted as part of the same experiment are reported elsewhere (Alessi et al, 2014;Bao et al, 2014;Chandler et al, 2013;Handley et al, 2014;Holmes et al, 2013a;Holmes et al, 2013b;Shiel et al, 2013). These studies investigated selected aspects of the experiment, including reactive transport modeling, microbiology of selected wells, gene expression, U isotopic shifts during bioreduction, and predation by protists on bacteria during biostimulation.…”

Applications of Functional Gene Microarrays for Profiling Microbial Communities

“…An amplification array was developed to allow for amplification, labeling, and hybridization to occur in a single reaction vessel (142). This array was developed as a way to perform microarray hybridization in the field with minimal equipment and rapid turnaround time.…”

Influence of bicarbonate on the abundance of microbial communities capable of reducing U(vi) in groundwater

“…3 illustrates that the compositions of microbial communities whose abundance is more than 1%. 44 Microbial communities, including Bacillus, Brevundimonas, Carnobacterium, Enterococcus, Lactococcus, Lysinibacillus, Solibacillus, Spirochaeta, and Sporosarcina, were detected in the preliminary sediment. Among these microbial communities, only Brevundimonas was reported to have the ability of reducing nitrate, 45 which indicated that functionalized indigenous microbial communities cannot be stimulated without adding electron donor.…”