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Microbial community composition of permeable reactive barriers: who is really doing the work?

PI: Jennifer Bowen, Univ. of Massachusetts - Boston, Joseph Vallino, Marine Biological Laboratory, Kenneth Foreman, Marine Biological Laboratory

Project Number:2012-R/RC-129

Proposal Summary

Objectives: Eutrophication of coastal waters is a growing environmental problem that can have catastrophic consequences for coastal ecosystems and the commercial pursuits that depend on them. Increasing supplies of anthropogenic nitrogen that originated as fertilizer runoff, atmospheric deposition, or septic waste induces eutrophication in these systems; finding ways to remediate these nitrogen supplies is a key focus of many municipalities in the region. One such mitigation strategy, the use of Permeable Reactive Barriers (PRBs) is currently being tested at the Waquoit Bay National Estuarine Research Reserve. The barriers are designed to remove nitrogen by promoting microbially mediated denitrification but there has been no examination of the microbial community that is responsible for the removal of the nitrogen and no attempt to understand how those microbial communities might respond to future environmental change. Since the barriers are designed to intercept groundwater flow prior to its entry into the coastal ocean they are located very near the seepage face of the estuary were fresh groundwater and seawater mix. In fact one of the two barriers in the Waquoit system already receives periodic inundation with salt water. As sea levels continue to rise, the response of the barrier microbial community to pulses of salt water will be a key determiner of their effectiveness. We propose to use cutting edge tools from molecular biology to examine how the barriers, and the periodic inundation of seawater, alter the structure and function of the microbial community. This information will help determine the best placement of the barriers to maximize their effectiveness.

Methodology: We will use a suite of tools from molecular biology to address these questions. First, we will use nearly continuous monitoring of salinity, temperature, and water table elevation within the wells to determine the frequency of saltwater inundation in the wells over lunar cycles. We will sample the network of wells in and around the barriers once in each season and we will also perform a high-resolution temporal sampling at spring and neap tides to document the dynamic nature of the zone surrounding the barriers. At each sampling time we will measure a suite of geochemical parameters (salinity, dissolved oxygen, pH, sulfate/chloride, dissolved Fe and Mn, sulfides, PO4, NO3, NH4, and total N) and we will filter groundwater for the later extraction of DNA and mRNA. Based on a detailed analysis of the geochemistry, a subset of the samples will be selected for further microbiological analysis. First, we will use high throughput pyrosequencing to examine how the barrier, and how the inundation of salt water to the barrier, alters the structure of the microbial community. Our primary focus, however, will be the analysis of nirS, a key gene in the denitrification pathway. We will use quantitative PCR to quantify this gene in the DNA collected from our samples. This will provide data on the overall capacity of the system to denitrify. We will also quantify the abundance of this gene in the mRNA, which quantifies those organisms that are actively synthesizing the enzyme involved in the process. Finally, we have developed a method to target functional genes, specifically nirS, for pyrosequencing, thus providing the deepest examination yet of the distribution and abundance of this gene in the environment. Based on our qPCR data we will select samples from both the DNA and mRNA for analysis by this new pyrosequencing method. Combining these various analyses will provide a comprehensive picture of the relationship between the microbial community and the geochemical processes that they perform. More importantly, it will allow us to examine how periodic inputs of salt water alter the capacity of the microbial community to effectively remove nitrogen and mitigate the eutrophication problem.

Rationale: The geochemical analysis to date suggest that the barriers are effective at removing land-derived nitrogen but the slight differences in location appear to have a profound effect on how the barriers function. There is some evidence to suggest that the barrier that receives inundation by salt water may facilitate a broader zone of effectiveness around the barrier because the addition of seawater to the barrier promotes a suite of different microbial pathways that result in differing biogeochemistry between the two barriers. To the extent that this zone of effectiveness actually enhances nitrogen removal, it has far reaching effects on the engineering of the barriers, which are generally located above the seawater/freshwater interface. Understanding how the microbial communities in the two barriers differ and under what environmental conditions the denitrifying bacteria thrive will help to guide future barrier construction, particular in the context of rising sea levels.

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