ARSH AGARWAL, ALL ISON BRADFORD, KERRY CHENG, RAMITA DEWAN, ENRIQUE DISLA , ADDISON GOODLEY, NATHAN L IM,

L ISA L IU, LUCAS PLACE, RAEVATHI RAMADORAI , JA ISHRI SHANKAR , MICHAEL

WELLEN, DIANE YE , EDWARD YU

MENTOR: DR . DAVE T ILLEYL IBRARIAN: ROBERT KACKLEY

GEMSTONE PROGRAM03/18 /2011

SWAMPSuperior Wetlands Against Malicious Pollutants

Research Problem

Agricultural runoff, especially in the spring, leads to high nitrate levels in the Chesapeake Bay Watershed

Causes harmful algal blooms Result: Dead zones due to depletion of oxygen and

nutrients vital to aquatic wildlife Dead zone: low oxygen area of water

Research Problem – Significance of Project

Affects fishing industry, seafood consumers, environmental groups, residents of the Chesapeake Bay Watershed

Health of the Chesapeake Bay is vital for maintaining biodiversity

Overview of Project

Goal: to build a wetland that optimally removes nitrates from the Chesapeake Bay and its surrounding waters

How? With a constructed wetland! Mostly greenhouse-based experiment in 3 phases Emulate conditions of the Tuckahoe Creek within the

greenhouseQuestions to answer through literature:

Where does the agricultural runoff come from? What plants can we use to remove the nitrates? Can we affect the rate of nitrate removal? How? With what?

Literature Review – Agricultural Runoff

One of the largest sources of pollution into the Bay Main sources: fertilizer and manure Plants only absorb up to 18% of nitrogen from fertilizer Up to 35% of nitrogen fertilizer washes into coastal waters and

their surrounding bodies of water Nitrates come mostly from chicken manure in agricultural

runoffEutrophication causes harmful algal blooms

Eutrophication: steep increase in nutrient concentration in neighboring bodies of water

Algal blooms lead to dead zonesConstructed wetlands

Can remove up to 80% of inflowing nitrates

Literature Review – River Selection

Big picture: Chesapeake Bay Not ideal for accessibility, too large a body of water for us

to study in such a short timeChoptank River – largest eastern tributary of the

Bay 70% of nitrogen input is from agricultural runoff Still not very accessible for a large group of students with

limited funds and transportationTuckahoe Creek

Tuckahoe sub-basin represents 34% of Choptank Watershed

More accessible for our team

The Nitrogen Cycle

Image from: www.fao.org

Literature Review – Plant Selection

Criteria for plant selection Non-invasive Native to the Chesapeake Bay Watershed Biofuel-capable

Literature Review – Plant Selection

Cattail (Typha latifolia) Very commonly researched wetland plant Especially viable as a biofuel

Soft-stem Bulrush (Schoenoplectus validus) More effective at denitrification than

other comparable plant species. Study: Schoenoplectus is responsible for

90% of all nitrate removal in experimental treatments

Switchgrass (Panicum virgatum) One of the most common, effective

nitrate-removing plants in the Chesapeake Bay area

Literature Review – Biofuels

Why biofuels? To accommodate changing energy and environmental needs Secondary data analysis

Cattail Potential ethanol source Can be harvested for cellulose

Switchgrass One hectare plot of switchgrass yielded up to 21.0 dry megagrams of

biomassSoft-stem bulrush

In one study, out of 20 wetland species, soft-stem bulrush ranked second in energy output per unit mass

Cross-referenced list of Chesapeake Bay native, non-invasive plants with list of biofuel-capable plants Selected plants seemed to be the best options for research

Literature Review – Organic Factors

Why? Increase statistical significance of differences in nitrate

removal Three carbon-based factors

Glucose Increases nitrate removal rates in artificial wetlands

Sawdust Study compared glucose & sawdust glucose ranked first,

sawdust ranked second Wheat straw

Increases nitrate removal rate for 7 days, then decreases in effectiveness

Methodology – Experimental Design & Setup

Take several samples at Tuckahoe Creek Mostly in spring highest nitrate concentration Use highest value of collected samples in greenhouse

environment Samples include water and soil

Soil samples are necessary to inoculate the greenhouse soil Inoculating soil will allow Tuckahoe-native bacteria to grow in

our greenhouse environmentExtraneous variables?

Realistically, we cannot emulate all elements of the Tuckahoe Creek in the greenhouse.

Nitrate concentration, soil composition, & temperature are three elements that we can realistically control

Methodology – Experimental Design & Setup (Phase 1)

Goal: find most effective organic factor Use single plant species (cattail) In each microcosm, place one or a combination of organic factors

Each microcosm will contain potting soil, top soil, soil from the Tuckahoe Creek (for inoculation), and the experimental variable Inoculating greenhouse soil with Tuckahoe soil will allow Tuckahoe-

native bacteria to grow in our greenhouse environment Collect effluent from each microcosm and pour it back over the

microcosm once a day for 7 days Measure nitrate concentration of the effluent at the end of the

week. Determine which factor or combination of factors per

experimental unit most effectively increases nitrate uptake Experimental unit is one bucket

Methodology – Example Diagram of Setup for Phase 1

Note: Phase 2 will look similar, but with different combinations in each bucket – the combinations will be of different plants, same organic factor

Methodology – Experimental Design & Setup (Phase 2)

Goal: find most effective plant or combination of plants using the organic factor determined in phase 1 Use multiple plant species Place each combination in a microcosm

Each microcosm will contain potting soil, top soil, soil from the Tuckahoe Creek (for inoculation), and the experimental variable

Collect effluent from each microcosm and pour back over the microcosm once a day for 7 days Standard water analysis will determine water quality

Determine which plant or plants (experimental unit) most effectively removes nitrates from water Experimental unit is one bucket

Methodology – Experimental Design & Setup (Phase 3)

Goal: apply the results of Phases 1 & 2 to a larger, more wetland-like setting Use the best factor and best combinations of plants Place them in a larger setting (i.e. a mini constructed

wetland within the greenhouse) Run experiment for 7 days, flowing water through this

larger-scale wetland environment Measure effluent once a day for 7 days to determine

nitrate removal efficiencyPending results of 1&2 depends on time

Methodology – Data Collection

Data Collection Effluent collected every day for 7 day trial Standard water analysis

Includes our variables, plus other details about water quality

Mostly within greenhouse Some data collection in the field (Tuckahoe) for samples

and testing of environment Six 1-week long trials

7 replicates of each microcosm per trial Total of 42 data points (can assume normal distribution)

Methodology - Data Analysis

Data Analysis Phase 1: Two-factor ANOVA

2 levels 4 treatments

Phase 2: Single factor ANOVA, Tukey’s Studentized Range 1 level 8 treatments

Statistical Analysis Software (SAS) to perform calculations

Current Progress

Finishing experimental setup and design Ironing out the fine details of water

collection/measurement/etcApplying for grants

Bill James, ACCIAC, Library (submitted), Sea Grant, HHMI Ongoing literature reviewTuckahoe Creek visits

Soil samples: early March Water samples: late April/early May

This is when nitrate concentration is highestGreenhouse space

Guaranteed space in the UMD greenhouse until May 2012

References Anderson, D., & Glibert, P., & Burkholder J. (2002). Harmful algal blooms and eutrophication: Nutrient

sources, composition, and consequences. Coastal and Estuarine Research Federation, 24(4), 704-726.  Burgin, A., Groffman, P., & Lewis, D. (2010). Factors regulating denitrification in a riparian wetland. Soil

Sci. Soc. Am. J., 74(5), 1826-1833. doi: 10.2136/sssaj2009.0463 Fraser, L. H., Carty, S. M., & Steer, D. (2004). A test of four plant species to reduce total nitrogen and

total phosphorus from soil leachate in subsurface wetland microcosms. Bioresource Technology, 94(2), 185-192. 

Hien, T. (2010). Influence of different substrates in wetland soils on denitrification. Water, Air, and Soil Pollution, June 2010, 1-12. doi:10.1007/s11270-010-0498-6

Gray, K. & Serivedhin, T. (2006). Factors affecting denitrification rates in experimental wetlands: Field and laboratory studies. Ecological Engineering, 26, 167-181.

Ines, M., Soares, M., & Abeliovich, A. (1998). Wheat straw as substrate for water denitrification. Water Research. 32(12), 3790-3794.

Karrh, R., Romano, W., Raves-Golden, R., Tango, P., Garrison, S., Michael, B., Karrh, L. (2007). Maryland tributary strategy Choptank River basin summary report for 1985-2005 Data. Annapolis, MD: Maryland Department of Natural Resources.

Rogers, K., Breen, P., & Chick, A. (1991). Nitrogen removal in experimental wetland treatment systems: Evidence for the role of aquatic plants. Research Journal of the Water Pollution Control Federation, 63(7), 9.

Staver, L. W., Staver, K. W., & Stevenson, J. C. (1996). Nutrient inputs to the Choptank river estuary: Implications for watershed management. Estuaries, 19(2), 342-358.

Wright, L., & Turhollow, A. (2010). Switchgrass selection as a “model” bioenergy crop: A history of the process. Biomass and Bioenergy, 34(6), 851-868. doi:10.1016/j.biombioe.2010.01.030

Zedler, J. B. (2003). Wetlands at your service: reducing impacts of agriculture at the watershed scale. Frontiers in Ecology and the Environment, 1(2), 65-72.

Zhang, B., Shahbazi, A., Wang, L., Diallo, O., & Whitmore, A. (2010). Hot-water pretreatment of cattails for extraction of cellulose. Journal of Industrial Microbiology & Biotechnology, 1-6. doi: 10.1007/s10295-010-0847-x

Thank you!

Many thanks to... Dr. Dave Tilley Dr. Bruce James Brandon Winfrey Dr. Wallace Dr. Thomas Courtenay Barrett Gemstone Program & Staff Robert Kackley

Any questions?