Brugia Malayi - Clark Science Center - Smith College

Exploring the Diversity of Sulfate-Reducing Bacteria Diversity in the
Avery Brook Beaver Pond Sediments
Rachel Kaminsky
Beavers influence ecosystem dynamics by building dams that retain sediment and organic material. This creates wetlands
and modifies nutrient cycling, affecting the character of the water and materials transported downstream. Beaver ponds are
biogeochemical hotspots that contain highly unexplored microbial activity and diversity. 1 They contain sulfate-reducing bacteria
(SRBs), which reduces sulfate, using it as the final electron acceptor in respiration. 2 This reduced sulfate facilitates the methylation
of mercury, which is a bioaccumulative environmental toxicant.
We conducted a study of the SRB community located in the sediments in a system of beaver ponds in the Avery Brook
stream system located in Conway, MA. This subcatchment forms part of the Mill River watershed and drains into the
Northampton Reservoir, the main source of drinking water for Northampton. 3 This summer, we focused on distinguishing how
the SRB community diverges between multiple sites within one pond.
The methods used for sampling the sediments involved the use of a foam board to float out towards the middle of the pond
(where we hypothesized more SRB would be located due to the more anoxic nature) and using a five foot-long straw to collect
sediment from the sediment-water interface. A previous method involved collecting sediment cores and segmenting off the top
three centimeters of the core. Genomic DNA was extracted from the samples using the PowerSoil ® DNA Isolation Kit from
MoBio Laboratories, Inc. Denaturing gradient gel electrophoresis (DGGE) was performed on bacterial dsrB sequences that were
amplified using PCR. PCR products of the same length can be separated by DGGE based on nucleotide composition. This
approach is useful for creating a genetic profile of environmental communities and can also be used to identify abundant taxa,
which is very difficult using other methods given the hyper-diverse nature of sediment environments. 4 Bands of interest were
excised from the gel and sequenced. Bacterial dsrA sequences were also amplified using PCR, and cloned into competent cells.
A few clones from each sampling site were chosen for sequencing. The sequences derived from DGGE and cloned DNA were
entered into the BLAST database to determine their identities. Cloning products were then miniprepped and sent to Penn State
for sequencing. In addition to molecular techniques, the geochemistry of the beaver ponds is being studied by observing the
levels of metals, ions, pH and dissolved oxygen and carbon.
Preliminary data from Sara Sirois’ honors thesis as well as BLAST sequences obtained this summer and last summer show
that there is a wide variety of sulfate reducing bacteria, many of which are novel and therefore not identifiable at the species level
when compared to BLAST. The majority of dsrA sequences were derived from members of the Deltaproteobacteria and the
Firmicutes. At this point, only one pond has been studied. Further collection of data from more ponds will answer questions
regarding the community divergence between different ponds. Plans for the future include isolating both SSU and dsrA RNA
in order to identify active taxa. We will continue to use DGGE to generate a more comprehensive profile of the pond in this
study as well as others in the Avery Brook series. This work will be continued in the 2012-13 school year as an honors project.
(Supported by the Howard Hughes Medical Institute)
Advisors: Robert Merritt, Laura Katz, Robert Newton and Steven Williams
References:
1 Naiman RJ, Johnston CA, Kelley JC. Alteration of north American streams by beaver. Bioscience 1988 Dec; 38(11): 753-62.
2 Stahl DA, Fishbain S, Klein M, Baker BJ, Wagner M. Origins and diversification of sulfate-respiring microorganisms. Antonie van Leeuwenhoek. 2002; 81:189-195.
3 Fletcher L. The geochemistry of the Avery Brook subcatchment in the Mill River watershed, Conway, MA. Smith.
4 Teske A, Wawer C, Muyzer G, Ramsing NB. Distribution of Sulfate-Reducing Bacteria in a stratified Fjord (Mariager Fjord, Denmark) as evaluated by Most-
Probable Number Counts and Denaturing Gradient Gel Electrophoresis of PCR-Amplified Ribosomal DNA Fragments. Appl. Enviro. Microbiol. 1996; 62(4):
1405-1415.
2012
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