Lake Pontchartrain Basin Research Program: Overview...... 1 Administration ...... 1 Letter from the Director ...... 1 Science Advisory Committee...... 3 Mission Statement ...... 4 Target Areas...... 4 Information Transfer & Outreach ...... 4 Publications & Presentations ...... 5 Training Graduate Students...... 7 Project Titles & PIs for Phases 1-5...... 8 Establishment of Baseline Concentrations and Elucidation of Environmental Processes Controlling the Bioavailability and Bioaccumulation of Mercury and Other Toxic Metals in the Lake Maurepas Basin (Phase 1: Field Study)...... 11 Growth Response and Tissue Accumulation Trends of Herbaceous Wetland Plant Species Exposed to Elevated Aqueous Mercury Levels ...... 21 Establishment of Baseline Concentrations and Elucidation of Environmental Processes Controlling the Bioavailability and Bioaccumulation of Mercury and Other Toxic Metals in the Lake Maurepas Basin (Phase 2: Greenhouse Investigation)...... 34 Viability of Mitigation in the Lake Maurepas and Manchac Swamp Region ...... 39 Hydraulic Conductivity and Vulnerability to Xylem Cavitation of Baldcypress along a Salinity Gradient as Indicators for Restoration Success...... 44 Salinity as a Stressor of the Freshwater Turtle, Trachemys scripta, in the Lake Pontchartrain Basin ...... 52 Genetic Variation between Lake Pontchartrain and Mississippi River Basin Fishes ...... 56 Western Lake Pontchartrain Basin Research Program Education Outreach Component: Phase 4...... 64 De-energizing Hurricanes with Cypress/Tupelo Buffers: A Plan to Restore the Repressed Swamps of the Lake Pontchartrain Basin by Using Point and Nonpoint Freshwater Sources ...... 67 Mitigating the Spread of Zebra Mussels into Wetlands from Mississippi River Diversions ...... 78 Development of an Index of Biological Integrity for the Lake Pontchartrain Basin Wetlands...... 83 Determining the Potential for Algal Bloom in Lake Maurepas: Effects of Changing Nutrient Load from Freshwater Diversion and Changes in Human Population...... 89 Information Transfer and Outreach Program for the Lake Pontchartrain Basin Research Program ...... 99

Program Overview

Lake Pontchartrain Basin Research Program: Overview

ADMINISTRATION William N. Norton, Ph.D. Lacy Landrum, Ph.D. Director Assistant Director SLU Box 10736 SLU Box 10585 Southeastern Louisiana University Southeastern Louisiana University Hammond, LA 70402 Hammond, LA 70402 O: 985.549.2923 O: 985.549.2268 F: 985.549.3851 F: 985.549.3851 [email protected] [email protected]

LETTER FROM THE DIRECTOR The Lake Pontchartrain Basin Research Program (PBRP) was established in 2001 as an interdisciplinary and inter-institutional program with research and education/outreach activities supported by funds from the EPA. PBRP is guided by an external Science Advisory Committee (SAC) comprised of 12 highly qualified and respected individuals who represent academia, federal and state agencies, the local community, and the private sector. Since its inception, PBRP has awarded over four million dollars to 44 investigators to support their environmental investigations of the Lake Pontchartrain Basin and for education/outreach activities that complement the program’s research.

PBRP has a very active and productive Information Transfer and Outreach Program, ensuring that the knowledge garnered through the program’s projects is disseminated widely to technical professionals in the regulatory agencies and to community leaders, as well as to citizen stakeholders and the news media. Several Research Reports that focus on program supported research projects and their results, as well as a Pontchartrain Basin Update entitled “Understanding the Environmental Impacts of Cypress Mulch” have been published and released. A pamphlet entitled “Notable Accomplishments of the Lake Pontchartrain Basin Research Program” has been distributed to the Louisiana Congressional Delegation.

Twelve research projects currently funded through PBRP at a total value of approximately one million dollars have been either completed or are in their final year of activity. The projects focus on diverse subjects, including an effort to determine whether genetic variation exists between Lake Maurepas and Mississippi River Basin fishes. Such information is critical for scientists to predict the effects of fresh water diversion projects. Additional studies focus on the bioavailability and bioaccumulation of mercury and other toxic metals in the Lake Maurepas Basin; the degree to which salinity induces stress on the endocrine system of an economically important species of turtle; the establishment of a physiological indicator of

PBRP Annual Report | 2009 1 Program Overview

restoration success for baldcypress after its exposure to high salinity; and the determination of potential algal blooms in Lake Maurepas as a consequence of nutrient loading from freshwater diversion projects. PBRP is also funding research projects that are attempting to determine methods for mitigating the spread of zebra mussels into the wetlands from diversions of the Mississippi River; to develop an Index of Biological Integrity for Lake Pontchartrain Basin wetlands; and to design a restoration plan for the swamps of the basin by using point and non-point freshwater sources.

PBRP also supports projects that pertain to the impact of human behavior on the Lake Pontchartrain Basin, such as the one designed to develop a “white paper,” a how-to manual, outreach workshops, and a website for mitigation banking in the Manchac Swamp. The education and outreach programs of PBRP are structured to emphasize hands-on and interdisciplinary educational experiences for both K-12 teachers and their students. The workshop-oriented activities introduce participants to the basin’s ecology, emphasizing the important link between the region’s ecology and its cultural and economic vitality.

William N. Norton, Ph.D. Professor of Biological Sciences Director of the Lake Pontchartrain Basin Research Program

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SCIENCE ADVISORY COMMITTEE PBRP is guided by an external Science Advisory Committee (SAC) comprised of 12 individuals representing academia, federal and state agencies, the local community, and the private sector. The SAC members advise the program director and, through a peer review system, critique submitted proposals for merit and compatibility with the PBRP mission. SAC recommends proposals worthy of funding to the director who then finalizes the ranking and dispersion of funds to the principal investigators (PIs). The PIs submit annual progress reports to the director who delivers those reports to the SAC members and to the EPA Project Officer for their review. SAC meets biannually to review proposals for each funding cycle and to review the progress of funded projects.

Member Expertise

Mr. Carleton Dufrechou Wetland restoration and Committee Chair community involvement Lake Pontchartrain Basin Foundation Dr. Dale Manty Hazardous substances EPA, Washington DC Dr. Robert Reimers Bioremediation Health Sciences Center Tulane University Dr. Kenneth Teague Wetland restoration EPA, Region 6 Mr. Dan Llewellyn Wetland ecology Louisiana Department of Natural Resources Dr. Len Bahr Environmental policy and Coastal Activities Division in the Louisiana Governor’s Office regulation Dr. Fred Kopfler Coastal restoration EPA, Region 4, EPA Gulf of Mexico Program Mr. Gordon Austin Treatment of sewage and Sewerage & Water Board of New Orleans waste water Dr. Mike Livingston Environmental toxicology Sobran Environmental Consultants Dr. Marilyn Kilgen Environmental microbiology Head of Biology Department, Nicholls State University Dr. David Constant Professor and Assistant Director of EPA HSRC Environmental engineering Louisiana State University Mr. Bill Hawkins Environmental toxicology Executive Director GCRL, University of Mississippi

PBRP Annual Report | 2009 3 Program Overview

MISSION STATEMENT The mission of PBRP is to determine the ecological stresses, including those associated with human behavior, on the Lake Pontchartrain Basin ecosystem and to provide scientific information to decision-makers and stakeholders on the methods and policies to stabilize, sustain, and/or enhance its environmental and economic recovery in a manner that is harmonious with the Comprehensive Management Plan for the restoration of the Lake Pontchartrain Basin and the Louisiana Coastal Area.

TARGET AREAS PBRP has established central themes or target areas for investigators to address. Funded projects in Phase V were designed to accomplish the following: 1. Identify the various environmental factors, including biotic and abiotic stressors that both positively and negatively impact the Lake Pontchartrain Basin and determine the extent to which they affect that ecosystem. 2. Determine the social, economic, and governmental factors that must be considered to achieve environmental recovery and sustainability of the Lake Pontchartrain Basin ecosystem. 3. Employ established and effective environmental models to determine the impact of specific variables of the ecosystem. Of particular interest are models that may be used to investigate various parameters associated with proposed freshwater diversion projects, such as the analysis of water quality, the deposition of particles, and the flow rates. 4. Design projects that will provide pertinent information regarding freshwater diversion programs. Examples of relevant topics include the proliferation of algal blooms; pollutant loading; nutrient loading; the impact of toxins such as benzene, heavy metals, herbicides, and pesticides; an analysis of the socioeconomic impacts; and the effect on threatened and endangered species. 5. Address factors specifically associated with Katrina/Rita-induced demographic changes, especially as they relate to the north shore of Lake Pontchartrain. Examples include environmental stressors induced by significant population increases, pollutant loading, land use, and wetland loss. Of particular importance are the effects of development and urbanization in the Pontchartrain watershed on the water quality in the estuaries.

INFORMATION TRANSFER & OUTREACH All proposals submitted to PBRP must have an information transfer and outreach component. Funded PIs are expected to address their progress in this effort throughout the project. As federal and state agencies search for ways to manage the complex ecosystems of southeastern Louisiana, information transfer is vital to communicating the most scientifically sound methods, the most current data, and insightful analysis that connects the data with short- and long-term outcomes. The stakeholders and policymakers use this information to make meaningful and appropriate decisions on how to manage our wetlands, including flood protection.

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RECENT PUBLICATIONS & PRESENTATIONS The investigators have actively published and presented the results of their studies to further their outreach efforts. The following list describes the formal publications and presentations, but many of these investigators have attended local and national meetings where they guide informal discussions of their findings and future research areas.

William Font and Sarah Brock (formerly Sarah Temple) recently learned their findings will be published in the July 2009 issue of Comparative Parasitology. Their article is titled “Helminths of the Western Mosquitofish (Gambusia affinis) in Bayou Traverse, Louisiana, U.S.A.”

Brian Crother, Clifford Fontenot, Tiffany Schriever and Joe Ramspott have published their findings in the March 2009 issue of Wetlands. Their article, entitled “Effects of Hurricanes Ivan, Katrina, and Rita on a Southeastern Louisiana Herpetofauna,” explains that herpetofauna diversity decreased after each hurricane along with drastic decreases in the overall number of amphibians while the number of reptiles varied with habitat.

Mark Hester and Jonathan Willis have presented their findings twice. “Establishment of baseline concentrations and elucidation of environmental processes controlling the bioavailability and bioaccumulation of mercury and other toxic metals in the Lake Maurepas Basin” was first presented at the Society of Wetland Scientists South Central South Atlantic Joint Chapter Meeting in Tuscaloosa, Alabama, in October 2008. The second presentation (same title) was at the 29th Meeting of the Society of Environmental Toxicology and Chemistry North America in Tampa, Florida, in November 2008. They are submitting the greenhouse findings of their study along with their hydroponics study to peer-reviewed journals this summer.

Kyle Piller and Lisa Cordes have presented their findings at three conferences. First, they presented “Genetic variation between Lake Pontchartrain and Mississippi River basin fishes” at the Louisiana/Mississippi Chapter of American Fisheries Society in Natchez, Mississippi in 2006. Their second presentation (same title) was in 2006 at SEEC in Tuscaloosa, Alabama. Also in 2006, they presented (same title) at the American Society of Ichthyologists and Herpetologists in New Orleans, Louisiana. They plan to submit their findings to a peer-reviewed journal by the end of the summer.

Gary Shaffer and his team have disseminated their findings through three publication venues. First, they have a journal article, entitled “Through droughts and hurricanes: tree mortality, forest structure, and biomass production in a coastal swamp targeted for restoration in the Mississippi River Deltaic,” in Forest Ecology and Management in 2008 (co-authors: Susanne Hoeppner and Thais Perkins). Second, in 2007, Gary Shaffer and John Day published a white paper, “Use of freshwater resources to restore Baldcypress-Water Tupelo swamps in the upper Lake Pontchartrain Basin,” for the Louisiana Department of Wildlife and Fisheries. And finally, their findings will be published this year in a special issue of the Journal of Coastal Research. The article is titled “Degradation of Baldcypress-Water Tupelo swamp to marsh and open water in southeastern Louisiana, USA: an irreversible trajectory? (co-authors: William Bernard Wood, S.S. Hoeppner, T. Perkins, Jason Zoller, and Demetra Kandalepas).

Volker Stiller has presented his findings through two posters and has one manuscript in press. His first poster, entitled “The effects of salinity on xylem cavitation in one-year-old Baldcypress

PBRP Annual Report | 2009 5 Program Overview

(Taxodium distichum) seedlings,” was presented at the Joint Congress of the American Society of Plant Biologists (ASPB), the American Fern Society (AFS), the American Society of Plant Taxonomists (ASPT) and the Botanical Society of America (BSA) in Chicago, Illinois in July 2007. His second poster, entitled “Drought and salinity affect wood density and vulnerability to xylem cavitation of Baldcypress (Taxodium distichum) seedlings,” was presented at the 93rd Annual Meeting of the Ecological Society of America (ESA) in Milwaukee, Wisconsin in August 2008, and then again at the 2009 meeting of the Louisiana Academy of Science in Hammond, Louisiana. His manuscript, “Soil salinity and drought alter wood density and vulnerability to xylem cavitation of Baldcypress (Taxodium distichum (L.) Rich.) seedlings,” has been accepted by Environmental and Experimental Botany.

Phillip Voegel and his team have presented five posters and given several invited lectures. They also have two manuscripts detailing their findings that they will submit this year, the first to Estuaries and Coasts and the other to Chemistry and Ecology. The poster presentations are as follows: Silcio, Kellie, Ricky Risley, and P.D. Voegel. “Changes in nutrient levels in Lake Maurepas following Hurricane Katrina.” 64th Southwest Regional Meeting of the American Chemical Society, Little Rock, AR, October 2008. Stricks, Jessica D. and P.D. Voegel. “Monitoring the effects of freshwater diversion from the Mississippi River into Lake Pontchartrain through the Bonnet Carré Spillway.” 64th Southwest Regional Meeting of the American Chemical Society, Little Rock, AR, October 2008. Silcio, Kellie and P.D. Voegel. “Nutrient levels in Lake Maurepas before and after Hurricane Katrina.” 235th National ACS Meeting, New Orleans, LA, April 2008. Silcio, Kellie, Kristy Ball, R. Risley, and P.D. Voegel. “Comparison of current nutrient levels in Lake Maurepas to pre-Katrina levels.” Pittsburgh Conference, New Orleans, LA, March 2008. Silcio, Kellie, K. Ball, and P.D. Voegel. “Assessing the impact of Hurricane Katrina on nutrients and algal growth in Lake Maurepas.” 34th FACSS Meeting, Memphis, TN, October 2007.

Of the 12 projects funded in Phases 4 and 5, six projects are complete. For three projects, the data collection is complete, and data analysis should conclude this fall. So we anticipate more publications and presentations as the data collection and analyses conclude for the remaining six projects.

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TRAINING GRADUATE STUDENTS An important goal of PBRP is to train a new generation of young scientists who will continue to investigate environmental problems. The following graduate students, working under major professors, have been involved in PBRP projects. Each graduate student received $16,000, meaning the program provided a total of $338,000 for graduate student financial support.

Graduate Student Major Professor Projects for Phase 1 Megan Collins William Font David Fox Phil Stouffer Ellen Geho Paul Keddy Brett Henry Ann Cheek Demetra Kandalepas Paul Keddy Timothy Menzel Paul Keddy Sarah Temple William Font Projects for Phases 2, 3, 4 Lisa Cordes Kyle Piller Todd Hymel Brian Crother Jessica Klopf Penny Shockett Eddie Koch Gary Shaffer Leonard McCauley Gary Shaffer Erica Perrer Penny Shockett Joe Ramspott Brian Crother Roxanne Rudowicz Colin Jackson Tiffany Schriever Brian Crother Jack Siegrist Paul Keddy Spencer Varnado Gary Shaffer Jason Zoller Gary Shaffer Projects for Phase 5 Colby Morgan William Font Chris Lundberg Gary Shaffer

PBRP Annual Report | 2009 7 Program Overview

PROJECT TITLES & PIS FOR PHASES 1-5 The following table describes the projects funded during phases 1-5. The project title and budget are listed along with the researchers and their expertise.

Project Title Budget Principal Investigator & PI Expertise ($US) Research Associates Phase 1 Effects of Multiple Stressors on 95,000 Paul Keddy, PI Wetland ecology Marshes and Swamps Southeastern Louisiana U Ecosystem Health and 197,701 Gary Shaffer, PI Wetland ecology Restoration Needs for Swamps Southeastern Louisiana U Constraints on Plant 187,011 Mark Hester, PI Plant physiology Establishment and Community U of New Orleans (now at Composition U of Louisiana, Lafayette) Vegetation and Bird 124,519 Phil Stouffer, PI Ornithology Communities Louisiana State U The Fish Parasite Community 131,960 William Font, PI Parasitology Southeastern Louisiana U Effects of Contaminants 121,425 Ann Cheek, PI Endocrinology and Southeastern Louisiana U physiology Teacher Workshops and In- 97,196 Debbie Dardis, PI Science education Service Training Southeastern Louisiana U Public Outreach and 23,412 Robert Moreau, PI Environmental Environmental History Southeastern Louisiana U studies Phases 2 & 3 Contingent Valuation of the 30,000 Jay Johnson Environmental Western Lake Pontchartrain Southeastern Louisiana U economics Basin Ecosystem Amphibian and Reptile 96,000 Brian Crother Herpetology Monitoring in the Pontchartrain- Southeastern Louisiana U Maurepas Region Restoring Biological Diversity to 260,000 Paul Keddy Wetland ecology Wetlands of the Greater Southeastern Louisiana U Manchac Region The Historical Transformation of 65,000 Samuel Hyde History the Manchac Basin Ecosystem: Southeastern Louisiana U Ecological Degradation at the Hands of Man Organic Matter Processing in 221,000 Colin Jackson Microbiology Western Lake Pontchartrain Southeastern Louisiana U Basin Wetlands now at U of Mississippi

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A Whole-System Approach for 340,000 Gary Shaffer Wetland ecology Restoring the Wetlands of the Southeastern Louisiana U Western Lake Pontchartrain Basin Outreach Component for 54,000 Robert Moreau Environmental Southeastern Louisiana Southeastern Louisiana U studies University’s Western Lake Pontchartrain Basin Research Program Western Lake Pontchartrain 155,000 Debbie Dardis Science education Basin Research Program: Southeastern Louisiana U Education Outreach Program Are Polycyclic Aromatic 100,000 Penny Shockett Immunology Hydrocarbons Stressors for Southeastern Louisiana U Lymphocyte Development or Activation in Frog Populations in Bayou Trepagnier? Genetic Variation between Lake 160,000 Kyle Piller Ichthyology Maurepas and Mississippi River Southeastern Louisiana U Basin Fishes Heavy Metal Contaminants: A 75,000 Maury Howard Environmental Study Investigating the Southeastern Louisiana U chemistry Occurrence, Distribution, and Species of Trace Metal Inputs to Western Lake Pontchartrain Phase 4 1. Establishment of Baseline 68,000 Mark Hester Wetland ecology Concentrations and Elucidation U of Louisiana, Lafayette of Environmental Processes Controlling the Bioavailability and Bioaccumulation of Mercury and Other Toxic Metals in the Lake Maurepas Basin 2. Growth Response and Tissue [This “Growth” project was an offshoot of the Accumulation Trends of “Establishment” project and was conducted with no Herbaceous Wetland Plant additional funding; its research results are included as an Species Exposed to appendix to the “Establishment” project.] Elevated Aqueous Mercury Levels Development of White Paper, 77,148 Robert Moreau Environmental How-To Manual, Outreach Southeastern Louisiana U studies Workshops, and Website for Mitigation Banking in Manchac Swamp

PBRP Annual Report | 2009 9 Program Overview

Hydraulic Conductivity and 77,500 Volker Stiller Plant physiology Vulnerability to Xylem Southeastern Louisiana U Cavitation of Baldcypress (Taxodium distichuml) along a Salinity Gradient as Indicators for Restoration Success Salinity as a Stressor of the 37,500 Roldan Valverde Animal physiology Freshwater Turtle (Trachemys scripta) in the Lake Pontchartrain Basin Genetic Variation between Lake 160,000 Kyle Piller Ichthyology Maurepas and Mississippi River Southeastern Louisiana U Basin Fishes Western Lake Pontchartrain 15,000 Debbie Dardis Science education Basin Research Program: Southeastern Louisiana U Education Outreach Program Phase 5 De-energizing Storms with 136,318 Gary Shaffer Wetland ecology Cypress/Tupelo Buffers: A Plan Southeastern Louisiana U to Restore the Repressed Swamps of the Lake Pontchartrain Basin by Using Point and Non-Point Freshwater Sources Mitigating the Spread of Zebra 88,492 William Font Parisitology Mussels into Wetlands from Southeastern Louisiana U Mississippi River Diversions Development of an Index of 145,817 Janice Bossart Entomology and Biological Integrity for Lake Southeastern Louisiana U ecology Pontchartrain Basin Wetlands Colin Jackson Microbiology U of Mississippi Determining the Potential for 87,512 Philip Voegel Chemistry Algal Bloom in Lake Maurepas: Southeastern Louisiana U Effects of Changing Nutrient Load from Freshwater Diversion and Changes in Human Population Information Transfer and 72,179 Robert Moreau Environmental Outreach for the Lake Southeastern Louisiana U studies Pontchartrain Basin Research Lacy Landrum Environmental Program Southeastern Louisiana U communication

The remainder of this report is comprised of the investigators’ reports. For phase 4 projects, the investigators have provided a final report of their findings. For phase 5 projects, the investigators have provided a progress report.

10 PBRP Annual Report | 2009 Bioavailability and Bioaccumulation of Mercury—Field Study

Establishment of Baseline Concentrations and Elucidation of Environmental Processes Controlling the Bioavailability and Bioaccumulation of Mercury and Other Toxic Metals in the Lake Maurepas Basin (Phase 1: Field Study)

Mark W. Hester1, Assaf A. Abdelghani2, Kyle R. Piller3, and Jonathan M. Willis1 1 Department of Biology; University of Louisiana, Lafayette 2 Department of Environmental Health Sciences; Tulane University 3 Department of Biological Sciences; Southeastern Louisiana University

ABSTRACT Mercury is a toxicant of global concern due to its widespread distribution and its methylated form that is highly bioavailable and neurotoxic. Because the methylation of mercury is primarily accomplished by sulfate-reducing bacteria prevalent in wetlands and aquatic sediments, these habitat types require intense scrutiny to protect the health of humans and ecosystems from unanticipated risks. We examined mercury in the soils and vegetation of the Maurepas wetlands along with other relevant environmental parameters and found no elevated concentrations of mercury or methyl mercury in soils or total mercury in vegetation.

BACKGROUND Despite recent research in the Lake Pontchartrain Basin, a major data gap remains regarding baseline concentrations and behavior of toxic metals within the region’s wetlands. The sediments, water, and biota of Lake Maurepas and Lake Pontchartrain, the major open waterbodies of the Lake Pontchartrain Basin, have been thoroughly characterized for the concentration and activity of toxic metals (Manheim and Hayes 2000, Delaune et al. 2008), but only recently has data of this nature become available for the soils and biota of the associated wetlands. This lack of data is especially worrisome in light of metal cycling processes, such as mercury methylation, documented in other large wetlands such as the Florida Everglades. Given how the fringing wetlands contribute to the health and sustainability of this region, it is critical that the concentrations and activities of toxic metals in these areas be examined. This data is imperative for the optimal engineering of many restoration strategies within these systems (e.g., river diversions, spoil bank gapping, dredge placement) that could significantly alter soil characteristics and result in increased bioavailability of toxic metals. Events associated with Hurricanes Katrina and Gustav may have introduced inorganic contaminants, such as lead and mercury, into the Lake Pontchartrain system. Recently, data on total and methyl mercury levels in the wetland sediments along the northern portion of Lake Maurepas have become available (Hall et al. 2008, Yu et al. 2008), but relatively little information is available for the southern wetlands of Lake Maurepas where wetland restoration is proposed.

The concentrations of metals in the bottom sediments, waters, and fishes of both Lake Maurepas and Lake Pontchartrain have been well documented (Manheim and Hayes 2000, Delaune et al. 2008). The Louisiana Department of Environmental Quality also maintains a

PBRP Annual Report | 2009 11 Bioavailability and Bioaccumulation of Mercury—Field Study

statewide monitoring system for mercury in waterway sediments and fish, including a number of sites within the Lake Pontchartrain Basin. Numerous water bodies within the Lake Pontchartrain Basin are included on the Louisiana Department of Environmental Quality fish consumption and swimming advisory list, including the Blind River, Tickfaw River, Amite River, and their associated streams and tributaries. Becnel et al. (2004) examined mercury concentrations in tree rings from various tree species and lichens in the Lake Maurepas Basin but suggested that these mercury concentrations were likely more indicative of airborne mercury concentrations than aqueous sources. Despite these evaluations of toxic metals in Lake Maurepas and Lake Pontchartrain bottom sediment and waters, fewer studies examine concentrations and availabilities of toxic metals within the surrounding wetland systems (Hall et al. 2008, Yu et al. 2008). Methyl mercury concentrations may be of concern given the number of studies documenting the capacity of wetlands to transform less toxic species of mercury into the more toxic methyl mercury form. Many of the wetlands surrounding Lake Maurepas have only limited hydrological exchange with Lake Maurepas and other waterways within it, and they experience soil redox potentials indicative of sulfate reduction. Thus, it is possible that the wetlands surrounding southern Lake Maurepas represent a potential reservoir of methyl mercury, which may become available if the hydrology of this wetland system is altered to allow exchange with Lake Maurepas.

OBJECTIVES For this study, we focused on elucidating the concentrations of mercury and methyl mercury in various environmental compartments. Specifically, by evaluating these data and other recently published data relevant to the area, we wanted to understand the potential risks of altered hydrology of the Lake Maurepas Basin to adjacent waters. We also wanted to investigate the role, if any, of aboveground and belowground vegetation in mercury cycling.

Hypotheses

1. Ho: Concentration of total mercury in abiotic (soil) and biotic (aboveground and belowground plant tissue) compartments in the Lake Maurepas wetlands is typical of uncontaminated wetlands.

2. Ho: Soil nitrate and sulfate status are primary controlling factors that poise surficial soil redox status and thereby methyl mercury concentrations.

MATERIALS & METHODS Experimental Design: A field study was implemented by selecting six sites throughout the Lake Maurepas wetlands that included major wetland vegetative habitat types. Four permanent plots were established within each of these sites, yielding a total of 24 plots. Pore-water pH, pore-water salinity, pore-water nutrients (NO3-NO2-N, NH4-N, SO4-S), soil organic matter, soil redox potential (1 and 10 cm depths), soil total Hg, soil methyl Hg, and aboveground/ belowground plant tissue Hg, Cd, Cr, Cu, Pb and Zn were determined seasonally at each plot. To investigate the statistical relationships, we created a randomized-block design with four blocks, i.e., each plot was treated as a block, yielding a 24 experimental plot experimental design. This design was entered into SAS 9.1 using the general linear model procedures, and the data were analyzed with repeated measures.

12 PBRP Annual Report | 2009 Bioavailability and Bioaccumulation of Mercury—Field Study

Field Site Selection: The six field sites were selected in the Lake Maurepas wetland system to assess the effects of wetland habitats dominated by various species on mercury cycling. A subset of these sites—sites at Blind River, Reserve Relief, Tobe Canal, and Turtle Cove—represent the gradient of proposed influence of the Maurepas diversion project as determined from previous hydrologic modeling studies. This gradient occurs along the south shore of Lake Maurepas in a roughly southwest to northeast direction, and it represents an inverse gradient of salinity and sulfate concentration, which are important factors in the bioavailability of many metals, and are among critical factors controlling mercury methylation. The final two sites are located along the northern portion of Lake Maurepas with the Amite Rive site close to a major tributary and with the Joyce WMA site in a wetland receiving tertiary treated wastewater. These sites were chosen to provide further spatial and environmental information.

Soil Redox Potential Characterization: Soil redox potential was determined at 1 and 10 cm depths using three bright, Pt soil redox electrodes per depth and a calomel reference electrode.

Soil Pore-Water Characterization: Where available, soil pore-waters (composite sample 15 cm in depth) were collected using acid-washed soil sippers. Immediately after collection of samples, one 3 ml aliquot of pore-water was placed into an equal volume of antioxidant buffer (SAOB reagent, ThermoOrion Corporation) and analyzed for total dissolved sulfides using an Orion ion-selective electrode (Orion Research Inc.) upon returning to the lab, but within 24 hours. A second aliquot for nutrient determination was expunged into a sample bottle and immediately placed on ice for transport back to the laboratory. Nutrient analysis was accomplished following EPA methods 350.1 (ammonia), 353.2 (nitrate-nitrite), 375.4 (sulfate) and 1620 (total phosphorus). A third aliquot was expunged into a sample bottle for onsite determination of pore-water pH, conductivity, and salinity using hand held meters (Orion, YSI 30) and approved EPA methods (APHA 1998).

Bulk Soil Characterization: Soil cores were collected to a depth of 15 cm, using a 7.62-cm diameter thin-wall aluminum soil corer, and placed into clean polyethylene bags. These soil cores were processed for the determination of total and methyl mercury following the methods outlined in EPA (2002) for total mercury and those in Bloom (1989) for methyl mercury. An additional soil core was collected from each plot and placed into a preweighed, polyethylene bag for the determination of organic matter upon returning to the lab (Soil Analysis and Plant Council 1999).

Plant Metal Uptake Accumulation: For the field study, plant cover was assessed through visual estimation in permanent plots. Samples of both aboveground and belowground biomass were collected into clean polyethylene bags and, upon returning to the lab, rinsed with deionized water and either dried in a lab oven to a constant weight at 65° C (for ICP metal determination) or blotted dry with kim wipes (total mercury determination). Thereafter, dry tissue samples were homogenized with stainless steel cutting tools, digested in 5-ml of trace metal grade nitric acid in a block digestor, and subjected to ICP-OES spectrophotometry for the determination of Cd, Cu, Cr, Pb, and Zn (see APHA 1998). Wet tissue samples were homogenized with stainless steel cutting tools, digested for a minimum of 12 hours in trace metal grade sulfuric and nitric acid at 100° C, oxidized with bromine chloride for a minimum of 12 hours, and then analyzed for total mercury concentration (see EPA method 1631 appendix for details).

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RESULTS Total Mercury Characterization: Significant effects were detected in total mercury concentrations in Lake Maurepas wetland soils in regard to season (F=2.80, P=0.0479), site (F=3.57, P=0.0071) and the interaction thereof (F=3.04, P=0.0012). However, all values reported for total soil mercury fall within a range that would be considered background soil levels for the United States (Eisner 2006) and also within the range reported for Lake Maurepas wetlands by Yu et al. (2008). Thus, these statistically significant effects, which likely result from microscale environmental differences, do not appear to be of concern from a contaminant standpoint. Significant effects were also detected in total mercury concentrations in belowground plant tissue in regard to season (F= 4.51, P= 0.0170), and the interaction of season and site (F=3.09, P=0.0051), but not the main effect of site (F=1.46, P=0.2236). Similarly for aboveground plant tissue, no significant effect was detected for site or season, but a significant interaction of site and season was detected (F=4.67, P=0.0002). As with total soil mercury concentration, all total belowground and aboveground tissue mercury concentrations fell within a range considered background for wetland plant species. The statistically significant differences in belowground plant tissue concentrations may reflect local scale phenomenon not captured by the experimental design. Importantly, all the plant tissue mercury concentrations reported are well within ranges that suggest no environmental contamination has occurred and no undue environmental concern is warranted.

Nutrients: Pore-water nitrate-nitrite-N concentrations were significantly higher in the Amite River site than in other sites (F=22.16, P<0.0001), particularly in the spring sampling, which resulted in a significant interaction (F=4.58, P<0.0001). Pore-water ammonium-N concentrations were significantly higher in the Blind River site than in other sites (F=29.27, P<0.001) and were significantly higher in spring than other sampling periods (F=8.10, P<0.001). A significant interaction of site and season was detected, likely resulting from the Joyce, Tobe Canal, and Turtle Cove sites having greater pore-water ammonium-N in the spring than in other seasons; whereas, the Blind River sites contained elevated pore-water concentrations for the study duration.

General Pore-Water Characteristics: A significant interaction of season and site was detected (F=4.967, P=0.003), likely a result of the spring and summer pore-water salinities being higher than the pore-water salinities for all sites other than the Amite River and Reserve Canal sites. A significant effect of site was also detected in regard to pore-water salinity, which likely results from Tobe Canal and Turtle Cove being more saline than other sites. No significant effect of season or interaction of season and site was detected for pH; however, an overall effect of site was detected, with Turtle Cove pore-water being more acidic than other sites.

General Soil Characteristics: A significant interaction of season and site was detected for both surface (F=13.283, P<0.001) and deep (F=34.117, P<0.001) soil redox potential, with Reserve Canal, Tobe Canal, and Turtle Cove being more reduced in summer than the other field sites. A significant effect of season was also detected, for both surface (F=59.948, P<0.001) and deep (F=111.160, P<0.001) soil redox potential, with soils being much less reduced in winter than the other seasons. No significant effect of season or interaction of season and site was detected for soil organic matter. However, an overall effect of site was detected, with Turtle Cove having less organic matter than other sites.

14 PBRP Annual Report | 2009 Bioavailability and Bioaccumulation of Mercury—Field Study

140 Spring Summer Fall Winter

120

100

40 Total Sediment Mercury (ng/g)Total Sediment Mercury 20

0 Amite Blind Joyce Reserve Tobe Turtle River River WMA Canal Canal Cove

Site 1.2 Spring Summer Fall Winter

1.0

0.8

0.6

0.4

Sediment Methyl Mercury (ng/g) Mercury Methyl Sediment 0.2

0.0 Amite Blind Joyce Reserve Tobe Turtle River River WMA Canal Canal Cove Site Figure 1. Effect of site and season on total sediment mercury (top fig., mean +/- s.e.) and sediment methyl mercury (bottom fig., mean +/- s.e.)

PBRP Annual Report | 2009 15 Bioavailability and Bioaccumulation of Mercury—Field Study

Table 1. Concentrations of relevant pore-water constituents and total mercury in aboveground and belowground plant tissue. Values are means with standard error in parentheses. Site Pore-water Pore-water Pore-water Aboveground Belowground NO3-N (mg/L) NH4-N (mg/L) SO4-S (mg/L) Plant Tissue Plant Tissue Total mercury Total mercury (ng/g) (ng/g) Spring Amite River 1.81 (0.12) 0.07 (0.1) 2.7 (1.8) 10.3 (1.6) 48.4 (26.1) Blind River 0.23 (0.01) 22.8 (1.8) 1.3 (1.1) 23.3 (13.6) 36.9 (12.5) Joyce WMA 0.67 (0.03) 25.3 (9.5) 1.3 (1.1) 6.3 (3.1) 16.7 (7.4) Reserve Relief 0.12 (0.07) 3.2 (1.5) 13.3 (2.9) 6.5 (2.9) 15.4 (6.4) Tobe Canal 0.58 (0.06) 6.2 (1.7) 68.1 (52.8) 15.1 (4.5) 25.8 (5.5) Turtle Cove 0.15 (0.13) 11.8 (1.9) 118.7 (59.3) 12.6 (5.4) 28.1 (7.2) Summer Amite River 0.85 (0.34) 3.1 (3.1) 9.3 (1.8) 10.5 (0.2) 15.4 (0.9) Blind River 0.21 (0.01) 14.2 (4.9) 1.3 (0.7) 16.1 (6.6) 12.9 (2.8) Joyce WMA 0.72 (0.01) 6.4 (0.1) 4.7 (3.7) 8.4 (4.1) 12.7 (2.7) Reserve Relief 0.52 (0.05) 0.7 (0.2) 26 (15.9) 19.3 (8.7) 21.4 (7) Tobe Canal 0.89 (0.01) 0.1 (0.1) 48.1 (38.2) 44.4 (12.5) 63.7 (8.7) Turtle Cove 0.37 (0.03) 5.1 (0.9) 107.3 (43.7) 27.6 (11) 46.9 (10.9) Fall Amite River 1.03 (0.41) 4.3 (1.3) 10.1 (3.5) 3.7 (1.8) 37.6 (27.2) Blind River 0.27 (0.02) 16.1 (5.7) 1.3 (1.3) 25.9 (16.2) 95.6 (33.9) Joyce WMA 0.52 (0.01) 9.8 (0.1) 3.3 (3.3) 2.5 (1.7) 29.2 (16.4) Reserve Relief 0.41 (0.08) 4.1 (1) 0.7 (0.7) 0.1 (0.1) 0.2 (0.1) Tobe Canal 0.34 (0.09) 0.4 (0.1) 74.1 (1.2) 1 (1) 4 (3.2) Turtle Cove 0.24 (0.02) 2.7 (0.1) 178.7 (2.9) 1.8 (0.7) 24.3 (5.4) Winter Amite River 0.49 (0.08) 6.1 (0.1) 4.7 (2.4) 16.7 (2.8) 92.2 (50.2) Blind River 0.18 (0.01) 21.2 (0.6) 5.3 (2.9) 1.9 (1.9) 2.3 (0.9) Joyce WMA 0.43 (0.07) 10.2 (0.5) 6.1 (4.1) 8 (3.5) 8.3 (3.1) Reserve Relief 0.37 (0.21) 0.6 (0.1) 20.7 (10.7) 0.6 (0.1) 24.6 (12.1) Tobe Canal 0.81 (0.13) 0.1 (0.1) 70.7 (0.7) 0.7 (0.1) 9.7 (3.9) Turtle Cove 0.34 (0.09) 3.3 (0.5) 204.7 (2.4) 8.4 (2.4) 13.2 (5.4)

16 PBRP Annual Report | 2009 Bioavailability and Bioaccumulation of Mercury—Field Study

DISCUSSION This assessment of mercury levels in the Lake Maurepas wetlands generally indicates that in all examined partitions total mercury concentrations are within a range considered to be representative of an uncontaminated wetland, thereby safe for local users (Eisler 2006), and also below the listed NOAA threshold effects level. Concentrations of total mercury reported in this study are within the range reported for Louisiana sediments and soils presented by other researchers, such as O’Rourke et al. (2001) 0.07-0.12 mg/kg, Dupre et al. (1999) below detection to 0.250 µg/g, Kongchum et al. (2006) 78 -240 µg/g, Delaune et al. (2008) 10.6-177 µg/g, and Yu et al. (2008) 8.7-288.9 µg/g. These levels of mercury likely reflect the lack of elevated atmospheric deposition of mercury and the absence of major industrial sources of mercury in the Lake Maurepas area. Methyl mercury levels were determined in surficial soils, which are frequently the major of site of mercury methylation in aquatic systems, and were also found to be within a range typical of an uncontaminated wetland (Eisler 2006). Interestingly, sediment methyl mercury concentrations determined in this study are slightly lower than those reported by Yu et al. (2008), which may reflect differences in hydrology prior to sampling. Water levels were unusually low during this study, with wetland soils moist, but not saturated during each sampling event; whereas, Yu et al. (2008) reported all soils either saturated or flooded. Also, it should be noted that the sediment methyl mercury concentrations reported by Yu et al. (2008) for many of their sampling locations around Lake Maurepas were similar to the values reported in this study. However, no sites sampled for this study exhibited the extreme elevated methyl mercury levels that were seen in four of sampling locations described in Yu et al. (2008), which likely reflects the difference in sampling scheme. Similarly, Hall et al. (2008), investigated surface and pore- water total and methyl mercury in several Louisiana wetlands, including the Blind River area, and found higher levels of methyl mercury in freshwater wetlands compared to adjacent surface waters, suggesting that the wetlands may function as a net source of methyl mercury to these open water bodies. Discussion of these findings highlights the need for multiple-year investigations as well as the need for a thorough understanding of the local environment, e.g., hydrology, to appropriately frame interpretations.

Total mercury levels in aboveground plant tissue determined in this study were similar to values detected in other studies of uncontaminated wetlands, such as Moore et al. (1995) 4 to 160 ng/g and Rencz et al. (2004), 5 to 58 ng/g. Total mercury in vegetation for this study were also similar to that recorded for the understory of boreal upland forest in Canada as reported by Hall and St. Louis (2004), 5 to 58 ng/g, and Mailman and Bodaly (2005) 4 to 52 ng/g, and mosses in the arctic Landers et al. (1995) 0.02–0.112 ug/g. In general, it does not appear that either aboveground or belowground vegetation components represent substantial total mercury reservoirs in the Lake Maurepas wetlands.

All wetland soil and soil pore-water variables evaluated were generally similar across sites, with the exception of the Turtle Cove, which had a higher pore-water salinity and lower sediment organic matter content reflecting its more brackish nature. Pore-water variables determined for the other five sites in this study—including salinity, pH, dissolved nitrate-nitrite, dissolved ammonia, and dissolved sulfate—were similar to the results other researchers have found for the Lake Maurepas wetlands (e.g., Hall et al. 2008) and also fall within the range reported for other oligohaline wetlands in Louisiana. It should be noted that sediment redox potential was higher for all sampling periods in this study than in other studies of the Lake Maurepas wetlands, likely a result of water levels in this system being unusually low during this study period. The soil

PBRP Annual Report | 2009 17 Bioavailability and Bioaccumulation of Mercury—Field Study

redox potentials determined during this study are generally corroborated by the relatively higher pore-water concentrations of ammonium, a reduced nitrogen compound, with the essential absence of sulfides, the main product of microbial sulfate reduction (Delaune et al. 2002). The ratio of oxidized to reduced nitrogen and sulfur compounds, in conjunction with the soil redox data, indicate that at the time these areas in the Lake Maurepas wetlands were sampled, dissimilatory nitrate reduction was the primary microbial metabolic pathway in use for these soils (Delaune et al. 2002).

In summary, mercury and methyl mercury levels in the various environmental compartments measured within the Lake Maurepas wetlands appear to be within the range of a normal uncontaminated wetland. Interestingly, other recently published data concerning mercury and methyl mercury concentrations in these wetlands report similar levels of total mercury in sediments, but higher levels of methyl mercury. This apparent contradiction may merely reflect the local hydrology at the time of sampling for different studies. Soil characteristics such as the large amount of organic matter with associated reduced sulfur functional groups may provide some substantial capacity to render newly deposited mercury unavailable for microbial activity, thus providing additional protection (see Skyllberg et al. 2003). However, experimental manipulation of soil mercury levels in a controlled setting coupled with an estimate of microbial bioavailability would be necessary to test this hypothesis. No consistent seasonal variation was detected in methyl mercury levels or in related sediment characteristics although, again, this was likely due to the abnormally mild winter and low water levels throughout the study. Results from this study, as well as Yu et al. (2008) and Hall et al. (2008), indicate that methyl mercury levels in the Lake Maurepas wetlands are either equivalent to or elevated in comparison to those of Lake Maurepas itself on average. However, sites with highly elevated levels of methyl mercury in the wetlands were completely absent from this study and rare (2 out of 35) in the study by Yu et al. (2008). Hall et al. (2008) generally found that freshwater and brackish wetlands in the Lake Pontchartrain Basin (e.g., Blind River and Bayou Lacombe) have elevated surface water methyl mercury levels compared with the surface waters of Pass Manchac and Lake Pontchartrain. This finding suggests that the surrounding wetlands are likely a source of methyl mercury to adjacent lake waters, which is considered typical for wetland-surface water systems (St. Louis et al. 1994).

The results of these various research projects do not preclude the use of wetland restoration projects that increase hydrologic connectivity between Lake Maurepas and adjacent wetlands; however, they do underscore the need for long-term, directed monitoring to ensure public safety. Currently, this monitoring could be accomplished efficiently through the existing Louisiana Department of Environmental Quality game fish mercury monitoring program. Game fish in appropriate Lake Maurepas tributaries should be an excellent indicator of elevated mercury levels resulting from restoration programs, as it should be directly indicative of mercury that may bioaccumulate and, with sufficient replication, be less sensitive to local microenvironments.

TECHNOLOGY TRANSFER This research aimed to determine levels of total mercury and methyl mercury in the Lake Maurepas wetlands and to elucidate factors controlling mercury cycling and bioaccumulation in this environment. Because mercury contamination has been problematic in many wetland systems that do not have a direct industrial input of this contaminant (e.g., the Everglades),

18 PBRP Annual Report | 2009 Bioavailability and Bioaccumulation of Mercury—Field Study determining the current levels and seasonal variation of mercury and methyl mercury in the Lake Maurepas wetlands is important for local users of these resources. Further, this data will enable local managers to make informed decisions about restoration strategies using hydrologic alteration (e.g., spoil bank gapping, river diversions), a strategy that could release currently isolated mercury into Lake Maurepas itself. Finally, by focusing on the cycling of mercury in wetland soils and vegetation, bioaccumulation risks and possibilities for sequestration can be addressed, potentially providing tools for not only local managers, but for other mangers working in similar environments in the southeastern United States.

Sustainability Questions 1. What are the current concentrations of methyl and total mercury in the Lake Maurepas wetland soils and how do they vary seasonally? 2. Do the dominant herbaceous plant species of the Lake Maurepas wetlands bioaccumulate mercury, and if so, in what portion of the plant does it tend to be located? 3. How do edaphic conditions and the potential alteration thereof affect mercury cycling?

Hypotheses 1. The concentration of mercury or methyl mercury in the soils of the Lake Maurepas wetlands soils are expected to be typical of an uncomtaminated wetland with methyl mercury levels being lowest in the winter season due to reduced microbial activity. 2. It is anticipated that all of the plant species tested will bioaccumulate mercury to some extent and will tend to store more mercury in root material. 3. Anoxic soil conditions with moderate levels of available sulfate and labile carbon are expected to lead to maximum methyl mercury production, with increased levels of nitrate tending to ameliorate this production.

This research suggests that the soils of the Lake Maurepas wetlands have typical levels of total mercury and methyl mercury and that Lake Maurepas vegetation has typical levels of total mercury in aboveground and belowground partitions. No distinct trends were detected in soil methyl mercury concentrations with season, likely due to the mild winter. Thus restoration efforts that alter local hydrology (e.g., diversions of river water, spoil bank gapping) will not likely result in increased mercury levels in adjacent waters.

The research conducted for this project indicates that the soils and vegetation of the Lake Maurepas wetlands have typical, background levels of mercury. Altering the local hydrology, which will occur with most restoration efforts, should not concern local users because these actions will not increase mercury levels. Agencies impacted by this research include the Louisiana Department of Natural Resources, the Louisiana Department of Environmental Quality, and the Environmental Protection Agency. Specific parishes that will benefit from this research are those with the plant species studied, including St. John the Baptist, Tangipahoa, and Livingston. Stakeholders directly benefiting from this research include hunters, shrimpers, fishermen, and recreational users of Lake Pontchartrain.

PBRP Annual Report | 2009 19 Bioavailability and Bioaccumulation of Mercury—Field Study

REFERENCES Becnel, J., C. Falgeust, T. Cavalier, K. Gauthreaux, F. Landry, M. Blanchard, M.J. Beck, and J.N. Beck. 2004. Correlation of mercury concentrations in tree core and lichen samples in southeastern Louisiana. Microchemical Journal 78:205-210. Delaune, R.D., R.P. Gambrell, A. Jugsujinda, I. Devai, A. Hou. 2008. Total Hg, methyl Hg and other toxic heavy metals in a northern Gulf of Mexico estuary: Louisiana Pontchartrain Basin. Journal of Environmental Science and Health, Part A Toxic/Hazardous Substances and Environmental Engineering 43:1006-1015. Dupre, T.P., T.J. Granier, S. Keife, R. Marino, S. O’Rourke, C. Partridge, D.L. Schultz, K. Mandhare and J.N. Beck. 1999. Variation of mercury concentration in fish taken from Lake Boeuf, southeastern Louisiana. Microchemical Journal 61: 156-164. Eisler, R. 2006. Mercury hazards to living organisms. Taylor and Francis Group, Boca Raton, FL, USA. Hall, B.D. and V. St. Louis. 2004. Methylmercury and total mercury in plant litter decomposing in upland forests and flooded landscapes. Environmental Science and Technology 38:5010-5021. Hall, B.D., G.R. Aiken, D.P. Krabbenhoft, M. Marvin-DiPasquale, C.M. Swarzenskif, E.T. Korthals, and M.R. Winfrey. 2008. Seasonal and spatial variations in mercury methylation and demethylation in an oligotrophic lake. Applied and Environmental Microbiology 53: 2397-2404. Landers, D.H., J. Ford, C. Gubala, M. Monetti, B.K. Lasorsa, and J. Martinson. 1995. Mercury in vegetation and lake sediments from the U.S. Arctic. Water, Air, and Soil Pollution 80:1573-2932. Louisiana Department of Environmental Quality. 2003. Resource guide to understanding mercury in Louisiana’s environment. Louisiana Department of Environmental Quality, Baton Rouge, LA, USA. Mailman, M. and R.A. Bodaly. 2005. Total mercury, methyl mercury, and carbon in fresh and burned plants and soil in Northwestern Ontario. Environmental Pollution 138:161-166. Manheim, F.T. and L. Hayes. 2000. Lake Pontchartrain Basin: bottom sediments and related natural resources. USGS Electronic Professional Paper 1634. Moore, T.R., J.L. Bubier, A. Heyes, and J. Flett. 1995. Methyl and total mercury in boreal wetland plants, experimental lakes area, Northwestern Ontario. Journal of Environmental Quality 24:845-850. O’Rourke, S., K. Gauthreaux, C.O. Noble, J. Sneddon and J.N. Beck. 2001. Mercury in sediments collected at the Sabine National Wildlife Refuge Marsh reclamation site in southwest Louisiana. Microchemical Journal 70:1-5. Rencz, A.N., N.J. O’Driscoll, G.E.M. Hall, T. Peron, K.Telmer, and N.M. Burgess. 2004. Spatial variation and correlations of mercury levels in the terrestrial and aquatic components of a wetland dominated ecosystem: Kejimkujik Park, Nova Scotia, Canada 143:271-288. Skyllberg. U, J. Qian, W. Frech, K. Xia, and W.F. Bleam. 2003. Distribution of mercury, methyl mercury and organic sulphur species in soil, soil solution and stream of a boreal forest catchment. Biogeochemistry 64: 53-76. Yu K, R.D. Delaune, I. Devai, R. Tao, and A. Jugsujinda. 2008. Total and methyl mercury in wetland soils and sediments of Louisiana’s Pontchartrain Basin (USA). Journal of Environmental Science and Health. Part A, Toxic/Hazardous Substances and Environmental Engineering 43:1657-1662.

20 PBRP Annual Report | 2009 Bioavailability and Bioaccumulation of Mercury—Field Study

APPENDIX After the field study, a hydroponics study was conducted to examine the utility of several species common to the Lake Maurepas wetlands for phytoremediation applications. This hydroponics study is separate from the greenhouse investigation (see p. 34) funded by the PBRP phase 2 portion of this research. Growth Response and Tissue Accumulation Trends of Herbaceous Wetland Plant Species Exposed to Elevated Aqueous Mercury Levels

ABSTRACT For this study, we investigated how elevated aqueous mercury levels (0, 2 and 4 ppm) impacted the growth status and mercury tissue concentrations of Eleocharis parvula, Saururus cernuus, Juncus

effuses, Typha latifolia, and Panicum hemitomon. Both short-term (net CO2 assimilation) and long- term (biomass) indicators of plant growth status suggest that Eleocharis parvula, Saururus cernuus, and Juncus effuses were relatively unaffected by elevated mercury levels; whereas, Typha latifolia and Panicum hemitomon were somewhat affected by such levels. All species demonstrated elevated tissue concentrations of mercury at elevated levels, with Eleocharis parvula, Panicum hemitomon, and Typha latifolia generally having the greatest tissue concentration of mercury. However, the species investigated in this study demonstrated lower levels of mercury accumulation into tissues when compared with similar investigations of other aquatic plants, suggesting that the studied species are not optimal for phytoremediation efforts.

INTRODUCTION Phytoremediation of contaminants in aquatic ecosystems has been proposed as a means of reducing various environmental pollutants including industrial organics (Cunningham et al. 1997), pesticides (Xia et al. 2001), petroleum products (Newman et al. 1998), explosives residue (Hughes et al. 1997), and metals (Weis and Weis 2004). Specifically, the phytoremediation of metals using wetland plants has received the most attention (Rai 2008, Weis and Weis 2004, Williams 2002). Recently, investigations into the phytoremediation of aqueous and soil-borne mercury have intensified (Moreno et al. 2004 and 2008, Skinner et al. 2007, Su et al. 2008), likely due to the highly toxic and bioaccumulative nature of this metal (Morel 1998). Several variables unique to individual contaminated sites affect the utility of phytoremediation (Weis and Weis 2004), including edaphic characteristics (e.g., pH, redox potential, organic matter content), the phytoremedation approach (e.g., phytoextraction versus phytostabilization), and the nature and concentration of the contaminant in question (EPA 2001, Otte and Jacob 2006, Sparks 2003). Thus, based on site characteristics and the desired approach, appropriate vegetation can theoretically be selected for optimal site restoration efforts (Sparks 2003). Yet further research on the capacity of wetland plant species to accumulate and sequester contaminants, particularly metals, would be informative.

The use of wetland plants to reduce mercury levels in various wetland environments has been examined for a number of years (e.g., Valiela et al. 1981, Sundberg-Jones and Hassan 2007). Recent research on the uptake of mercury by various aquatic species has indicated that

PBRP Annual Report | 2009 21 Bioavailability and Bioaccumulation of Mercury—Field Study

substantial phytoremediation of mercury-contaminated waters may be possible (Kamal et al. 2004, Skinner et al. 2007). Species that show promise as phytoremediating agents for mercury contamination of aquatic environments include Azolla carolinia (Bennicelli et al. 2004), Myriophyllum spicata, Ludwigia peploides, and Mentha aquatica (Kamal et al. 2004), as well as Eichornia crassipes, Pistia stratiotes, Scirpus tabernaemontani, and Colocasia esculenta (Skinner et al. 2007). Understanding the uptake and partitioning of inorganic mercury in wetland vegetation under different scenarios of elevated mercury loadings has implications for management of habitats. For example, King et al. (2002) determined variable removal of mercury from source waters in a wetland planted with Scirpus californicus and that mercury not sequestered by the vegetation became methylated. Incorporation of mercury into vegetative tissue where methylation appears to occur sparingly is an important, if temporary, reservoir of inorganic mercury unavailable for methylation by microbial organisms (Weis and Weis 2004). However, this mercury may still be incorporated into the food chain, even if in a form less toxic and less prone to bioaccumulation. Substantial interspecific variation in inorganic mercury uptake by and toxicity to wetland vegetation has been documented. Descriptions of tolerance to inorganic mercury, as well as patterns of partitioning into aboveground and belowground vegetation for individual species, are needed to better understand mercury cycling in wetland environments. This information will enable the proper management of natural wetlands subjected to elevated mercury inputs as well as guide the continued refinement of phytoremediation technology. The objectives of this research are to determine interspecific differences in mercury uptake and localization, as well as the physiological response and tolerance to elevated mercury levels by common herbaceous fresh marsh plants.

MATERIALS & METHODS A randomized factorial design was used to investigate the mercury uptake of several plant species. The design included five species (Eleocharis parvula, Saururus cernuus, Juncus effuses, Typha latifolia, and Panicum hemitomon) x three mercury levels (0-μg ml-1 2-μg ml-1 4-μg ml-1) x four true replicates (60 experimental units). The experiment was established as a greenhouse container study using 1-gallon nursery pots placed within 2-gallon buckets as reservoirs. Healthy specimens of the above plant species were collected from the Joyce WMA, Louisiana, placed into 1-gallon nursery pots, and transported back to the University of New Orleans greenhouse facility. Upon returning to the greenhouse facility, plant roots were rinsed of soil material, placed into acid-washed 1-gallon nursery pots with acid-washed sand, and flooded to 2.5 cm above the sand surface using type III deionized water to which nutrients were added to equal 25% Hoaglands solution. Plants were allowed to acclimate for two weeks prior to the initiation of the study. Treatments were randomly assigned, and aqueous solutions of mercuric chloride (or type III deionized water for the control) were added to the surface water of appropriate vessels, after which pots were agitated in reservoirs by rapidly lifting them up and down several times to

ensure mixing of treatment solutions throughout the sand matrix. After two months, net CO2 assimilation rate, aboveground tissue total mercury, belowground tissue total mercury, residual mercury in pore-water, and biomass partitioning were determined.

Net CO2 assimilation rate was determined on the two youngest, fully-expanded leaves for each experimental unit and then averaged using a LI-Cor 6400 Photosystem with leaf chamber light -2 -1 intensity set to 1,500 umol m s , leaf chamber CO2 set to 370 ppm, and leaf chamber relative humidity maintained between 20 to 60%. At the conclusion of the study, aboveground and

22 PBRP Annual Report | 2009 Bioavailability and Bioaccumulation of Mercury—Field Study

belowground tissue was clipped with stainless steel scissors, rinsed with type III de-ionized water, placed into paper bags, and dried to a constant weight. Subsets of tissue were homogenized with a Wiley Mill and digested using a 1:3 mixture of trace-metal grade nitric and hydrochloric acid (i.e., aqua regia) in Teflon digestion vessels (see appendix to EPA method 1631, digestion 2 for details and caveats). Total mercury analysis was accomplished following EPA method 1631 (Cold Vapor Atomic Fluorescence) using a four-unit bubbler setup in conjunction with a Brooksrand amalgation control module and a Model III atomic fluorescence spectrophotometer. Statistical relationships were elucidated using the appropriate general linear model procedures of PC-SAS 8.0 (SAS 2004).

RESULTS Tissue and Pore-Water Mercury Concentrations: Species differed significantly in their total aboveground tissue concentrations of mercury (fig. 2; F=2.919, P=0.34), with Eleocharis parvula, Panicum hemitomon, and Typha latifolia tending to have the greatest concentration and Juncus effuses and Saururus cernuus tending to have the lowest concentration. Mercury treatment level had a significant effect on total aboveground tissue mercury concentration, with greater concentrations of mercury accumulating into aboveground tissue with higher mercury levels (fig. 2; F=3.243, P<0.001). A significant interaction of species and mercury level was detected, resulting from Eleocharis parvula, Panicum hemitomon, and Typha latifolia having total aboveground mercury concentrations that were substantially elevated at 4 ppm compared to 0 and 2 ppm; whereas, Juncus effuses and Saururus cernuus had elevated aboveground total mercury tissue concentrations at both 2 and 4 ppm (fig. 2; F=3.050, P<=0.047).

Belowground tissue concentrations of mercury displayed similar trends with Eleocharis parvula, Panicum hemitomon, and Typha latifolia displaying the greatest concentrations and Juncus effuses and Saururus cernuus the least (fig. 2; F=3.30, P=0.0495). As was found with the aboveground tissue analyses, mercury level significantly increased total belowground tissue mercury concentration (fig. 2; F=22.096, P<0.001). A significant interaction of species and mercury level was detected, resulting from elevated belowground tissue mercury concentrations in Eleocharis parvula, Panicum hemitomon and Typha latifolia in the 2 ppm treatment compared with Juncus effuses and Saururus cernuus, which had minimal accumulation in this treatment (fig. 2; F=3.05, P=0.0372). For all species, accumulation of mercury into belowground tissues was significantly greater than into aboveground tissues. Final pore-water concentrations were substantially reduced in the 4 ppm mercury loading for all species and in the 2 ppm mercury loading in all species except Panicum hemitomon (fig. 3). However, reductions in final pore-water concentrations in the 2 ppm mercury loading generally tended to be more variable than the reductions in the 4 ppm mercury loading (fig. 3). Importantly, when the mass balance of total mercury added to each experimental unit (14.6 mg and 7.3 mg for 4 and 2 ppm treatments, respectively) versus the amount of mercury absorbed into plant tissue is considered, vegetation appears to play a minimal role compared to other effects (e.g., volatilization and binding to solids) in reducing total mercury load (tab. 2).

PBRP Annual Report | 2009 23 Bioavailability and Bioaccumulation of Mercury—Field Study

350 0 ppm 2 ppm 300 4 ppm

250

200

150

100

Aboveground Tissue Hg (ng/g) Tissue Hg Aboveground 50

0 Eleocharis Juncus Panicum Saururus Typha parvula effusus hemitomon cernuus latifolia Species 20,000 0 ppm 18,000 2 ppm 16,000 4 ppm 14,000 12,000 10,000 8,000 6,000 4,000 2,000 Belowground Tissue Hg (ng/g) Tissue Hg Belowground 0 Eleocharis Juncus Panicum Saururus Typha parvula effusus hemitomon cernuus latifolia

Species

Figure 2. The effect of species and mercury level on aboveground plant tissue mercury concentration (top fig.) and belowground plant tissue mercury concentration (bottom fig., mean +/-se)

24 PBRP Annual Report | 2009 Bioavailability and Bioaccumulation of Mercury—Field Study

100 4 ppm 90 80 70 60

ewater Hg (%) Hg ewater 50 40 30 20 10

Reduction in por in Reduction 0 Eleocharis Juncus Panicum Saururus Typha parvula effusus hemitomon cernuus latifolia Species 100 2 ppm 90 80 70 60 50 40 30 20 10

Reduction in porewater Hg (%) Hg porewater in Reduction 0 Eleocharis Juncus Panicum Saururus Typha parvula effusus hemitomon cernuus latifolia Species

Figure 3. The effect of species on reduction in pore water concentration (mean +/-se)

PBRP Annual Report | 2009 25

Table 2. Aqueous mercury reduction through plant tissue accumulation (%) and relevant metrics

Treatment Species Belowground Belowground Aboveground Aboveground Total Total aqueous Aqueous mercury Tissue Biomass (g) Tissue Biomass (g) mercury mercury (µg) reduction through mercury mercury (µg/g) accumulated plant tissue (µg/g) into biomass accumulation (%) (µg) Eleocharis 2 ppm parvula 7.21 36.7 0.01 8.4 264.8 7,300 3.63 Juncus 2 ppm effusus 0.14 27.5 0.07 7.3 4.3 7,300 0.06 Panicum 2 ppm hemitomon 7.32 45.2 0.04 19.2 331.9 7,300 4.55 Saururus 2 ppm cernuus 0.15 24.1 0.07 4.8 4.0 7,300 0.05 Typha 2 ppm latifolia 9.64 32.2 0.02 8.0 311.2 7,300 4.26

Eleocharis 4 ppm parvula 16.23 26.5 0.26 4.4 431.0 14,600 2.95 Juncus 4 ppm effusus 4.26 37.1 0.08 8.6 158.4 14,600 1.08 Panicum 4 ppm hemitomon 18.23 33.3 0.11 10.6 608.0 14,600 4.16 Saururus 4 ppm cernuus 4.46 23.6 0.08 5.4 105.5 14,600 0.72 Typha 4 ppm latifolia 13.98 55.6 0.17 16.0 779.3 14,600 5.34

Bioavailability and Bioaccumulation of Mercury—Field Study

Photosynthetic Response: A significant effect of species on net CO2 assimilation rate was detected (fig. 4; F=77.596, P<0.001), with Juncus effuses, Saururus cernuus, and Typha latifolia having greater net CO2 assimilation rates than Eleocharis parvula and Panicum hemitomon overall. However, Typha latifolia and especially Panicum hemitomon, displayed significant depressions in CO2 assimilation rate when exposed to the highest mercury levels (fig. 4). Interestingly, no significant effects of mercury level, or the interaction of mercury level and species, were detected. A significant effect of species on stomatal conductance was detected, with Eleocharis parvula displaying much greater stomatal conductance than the other species (fig. 4; F=10.832, P<0.001).

Biomass Response: A significant effect of species on aboveground biomass was detected (fig. 5; F=4.954, P=0.002). The aboveground biomass production of Panicum hemitomon and Typha latifolia was greater than that of Juncus effuses, Saururus cernuus, and Eleocharis parvula although no significant effect of mercury level on biomass production was evident. A marginally significant interaction of species and mercury level was detected (fig. 5; F=1.970, P=0.073), in which Eleocharis parvula produced less aboveground biomass at higher mercury levels while all other species had equivalent or greater aboveground biomass at the higher levels. A significant effect of species on belowground biomass was detected that closely mirrored the aboveground biomass response (fig. 5; F=3.447, P=0.015), but the belowground production of both Panicum hemitomon and Eleocharis parvula were depressed at the highest levels of mercury. No significant effect of mercury level or interaction of mercury level and species was detected for belowground biomass. Total biomass demonstrated trends consistent with the aboveground and belowground metrics (F=4.749, P=0.003), with no significant effect of mercury level or the interaction of mercury level and species.

DISCUSSION All species investigated were able to survive and sustain reasonable levels of metabolic function, allowing for growth at elevated aqueous mercury concentrations (up to 4 µg ml-1) while accumulating substantial amounts of mercury in belowground tissues. However, although extremely high reductions in pore-water mercury concentrations were noted by the end of the study (up to 97%), accumulation into vegetative tissue appears to play a relatively small role. Interestingly, interspecific differences in mercury accumulated into tissues were found, with Eleocharis parvula, Panicum hemitomon, and Typha latifolia, generally having greater concentrations of mercury in their belowground tissues than Juncus effusus and Sarurus cernuus. Inherent differences between species in growth characteristics relevant to phytoremediation, such as biomass production and stomatal conductance, were also elucidated.

PBRP Annual Report | 2009 27 Bioavailability and Bioaccumulation of Mercury—Field Study

16 0 ppm 14 2 ppm 4 ppm 12 )

-1 10 s -2 8 Assimilation Assimilation 2

(umol m (umol 6 Net CO 4

0 Eleocharis Juncus Panicum Saururus Typha latifolia parvula effusus hemitomon cernuus Species

0.80 0 ppm 0.70 2 ppm 4 ppm 0.60

) 0.50 -1 s -2 0.40

(mol m (mol 0.30

Stomatal Conductance Conductance Stomatal 0.20

0.10

0.00 Eleocharis Juncus Panicum Saururus Typha latifolia parvula effusus hemitomon cernuus Species

Figure 4. The effect of species and mercury level on net CO2 assimilation (top fig.) and stomatal conductance (bottom fig.)

28 PBRP Annual Report | 2009 Bioavailability and Bioaccumulation of Mercury—Field Study 80 0 ppm 70 2 ppm 4 ppm 60

20 Aboveground Biomass (g) Biomass (g) Aboveground 10

0 Eleocharis Juncus Panicum Saururus Typha parvula effusus hemitomon cernuus latifolia Species 80 0 ppm 70 2 ppm 4 ppm 60

20 Belowground Biomass (g) Biomass (g) Belowground 10

0 Eleocharis Juncus Panicum Saururus Typha parvula effusus hemitomon cernuus latifolia Species

Figure 5. The effect of species and mercury level on aboveground biomass (top fig.) and belowground biomass (bottom fig., mean +/-se)

PBRP Annual Report | 2009 29 Bioavailability and Bioaccumulation of Mercury—Field Study

The net CO2 assimilation rates of Typha latifolia and particularly Panicum hemitomon were decreased from the controls at 4 ppm mercury loadings, but not at 2 ppm mercury loadings.

The net CO2 assimilation rates of the remaining species were not significantly decreased from the controls at either mercury loading, and, in fact, for Eleocharis parvula a stimulation of

net CO2 assimilation at the 4 ppm mercury loading was discerned. As net CO2 assimilation is an instantaneous indicator of current growth status, these results indicate that other than Typha latifolia and Panicum hemitomon, these species are capable of maintaining normal metabolic growth functions at highly-elevated mercury loadings after two months. The

decrease in net CO2 assimilation rate for Typha latifolia and Panicum hemitomon occurred only at the 4 ppm mercury loading and was modest, suggesting that these plants would survive and possibly even expand at this elevated loading, but at a slower rate than under control conditions. Stomatal conductance was highest for Eleocharis parvula and generally similar for the remaining species. Although transpiration is known to be an important component of mercury removal through phytovolatilization (Moreno et al. 2008), it does not appear to be a primary driver in this study as reductions in aqueous mercury were not correlated with stomatal conductance rates.

Total biomass was generally comparable between the control and elevated mercury treatments with the exception of Typha latifolia and Panicum hemitomon. This finding contrasts that of Du et al. (2005) who reported biomass production by Oryza sativa was decreased when plants were exposed to aqueous mercury loads as low as 0.5 ppm. As was noted with

net CO2 assimilation rates, the decrease in total biomass for these species are relatively small, indicating these plant would likely be able to survive at these elevated mercury loadings, but would grow at a reduced rate compared with control conditions. Typha latifolia and Panicum hemitomon produced the greatest total biomass overall, with the remaining species having fairly similar total biomass. This production, in combination with the concentrations of mercury detected in the tissue of Typha latifolia and Panicum hemitomon, suggests that of the species examined these may be preferable for use in a phytoremediation setting.

All plants in control treatments were found to have mercury concentrations typical of plants growing in uncontaminated wetlands (Mailman and Bodaly 2005, Sparks 2003). Eleocharis parvula followed by Typha latifolia had the greatest concentration of mercury in aboveground plant tissue, indicating that of the species investigated, these would be preferable for a phytoextraction approach where aboveground tissues are harvested and belowground tissues are left in place. In the 2 ppm treatment, final pore-water mercury levels were highly variable, suggesting differential removal of mercury, although species differences in physiological response (e.g., stomtal conductance, vegetative uptake of mercury) did not appear to be driving this variation. Pore-water mercury levels in the 4 ppm treatment after two months were fairly consistent regardless of species or interspecific differences in uptake, indicating that using any of these species for phytoremediation of aqueous mercury contamination at similar levels would likely have equitable results.

Because similar degrees of reduction in final pore-water mercury concentration occurred both in experimental units planted with Sarurus cernuus (low tissue mercury concentrations and biomass) and experimental units planted with Typha latifolia (higher levels of tissue mercury concentrations and biomass), some other mechanism of mercury removal may have occurred. This supposition is reinforced by the extremely small percentage of mercury absorbed into plant tissue compared to the initial mercury load. Volatilization of mercury

30 PBRP Annual Report | 2009 Bioavailability and Bioaccumulation of Mercury—Field Study

directly from experimental water or adherence of mercury to some component of the experimental units could be factors in this apparent discrepancy (Skinner et al. 2007).

Skinner et al. (2007) conducted a similar experiment in which Eichornia crassipes, Pistia stratiotes, Scirpus tabernaemontani, and Colocasia esculenta were subjected to hydroponic mercury loadings of 0, 0.5, and 2 ppm. The reductions in final pore-water mercury concentrations in this study are lower than in Skinner et al. (2007); however, final tissue mercury concentrations for the two emergent species (Scirpus tabernaemontani and Colocasia esculenta) for the Skinner et al. study were generally similar to those in this study (except for Sarurus cernuus). Skinner et al. (2007) reported that the floating aquatic species (Eichornia crassipes and Pistia stratiotes) accumulated considerably more mercury into their aboveground tissues than the emergent species (Scirpus tabernaemontani and Colocasia esculenta). Similarly, Molisani et al. (2006), in a survey of plant tissue mercury concentrations in Brazil, found that free-floating vegetation tended to have greater tissue mercury concentrations than emergent species.

Elevated accumulation of mercury into the tissues by floating aquatic plants has also been documented in a number of other hydroponic mercury dosage studies (e.g., Eichornia crassipes, Chigbo et al. 1982; Salvinia natans, Sen and Mondal 1987; Lemna minor, Choi et al. 1989). In a hydroponic study by Moreno et al. (2008), Brassica juncea was found to accumulate much greater tissue concentrations of mercury than plants in this study when subjected to similar aqueous mercury levels. Similarly, Oryza sativa was found to accumulate tissue mercury concentrations nearly two orders of magnitude higher than species in this study when subjected to similar mercury loads (Du et al. 2004). An investigation by Rai and Tripathi (2009) also reported high levels of aqueous mercury reduction in experimental vessels containing Azolla pinnata and Vallisneria spiralis (up to 94% and 84%, respectively) when vessels received mercury loadings similar to those used in this study.

Results from this study indicate that all species investigated have some capacity to reduce aqueous mercury loads in a hydroponic setting. However, aqueous mercury reduction through uptake into vegetative tissue appears to be less than that presented in other studies using different species (Du et al. 2004, Skinner et al. 2007, Moreno et al. 2008). As has been documented for a number of species, translocation of mercury from belowground to aboveground plant partitions was minimal, suggesting these plants would be effective for sequestering limited amounts of mercury in belowground material. Nonetheless, when compared to previous studies, other species, particularly free-floating species, may be more efficient in aqueous mercury removal than the species investigated herein.

CONCLUSIONS This study indicates that Eleocharis parvula, Typha latifolia, and Panicum hemitomon all accumulate mercury into their belowground tissues under elevated loadings to a greater extent than Sarurus cernuus and Juncus effuses, which demonstrated more modest concentrations of tissue mercury. Overall these levels are lower than have been reported for other species during screening experiments for the phytoremediation of mercury. This study emphasizes the importance of determining both biomass production and final tissue concentration to fully understand the ability of a plant species to remove aqueous mercury.

PBRP Annual Report | 2009 31 Bioavailability and Bioaccumulation of Mercury—Field Study

REFERENCES Bennicelli, R., A. Banach, K. Szajnocha, and J. Ostrowski. 2004. The ability of Azolla caroliniana to remove heavy metals (Hg(II), Cr(III), Cr(VI)) from municipal wastewater. Chemosphere 55:141-146. Choi, D.S., J.W. Robinson, and S.C. Mo. 1989. Uptake of mercury from aqueous solution by duckweed: The effects of pH, copper and humic acid. Journal of Environmental Science & Health, Part A: Environmental Science & Engineering 27:135-146. Chigbo FE, R.W. Smith and F.L. Shore. 1982. Uptake of arsenic, cadmium, lead and mercury from polluted waters by the water hyacinth. Environmental Pollution, Series A 27:31-36. Cunningham, S.D., J.R. Shann, and D.E. Crowley. 1997. Phytoremediation of contaminated water and soil. Pages 2-19. in E.L. Kruger, T.A. Anderson, and J.R. Coats, editors. Phytoremediation of soil and water contaminants. ACS Symposium Series No. 664. American Chemical Society, Washington, DC, USA. Du, X., Y.G. Zhua, W.J. Liua, and X.S. Zhao. 2005. Uptake of mercury (Hg) by seedlings of rice (Oryza sativa L.) grown in solution culture and interactions with arsenate uptake. Environmental and Experimental Botany 54:1-7. EPA. 2001. A citizen’s guide to phytoremediation. U.S. Environmental Protection Agency, Office of Solid Waste and Emergency Response. EPA-542-F-01-002. Hughes, J.B., J. Shanks, M. Vanderford, J. Lauritzen, and R. Bhadra. 1997. Transformation of TNT by aquatic plants and plant tissue cultures. Environmental Science and Technology 31:266-271. Hutchinson, S.L., M.K. Banks and A.P. Schwab. Phytoremediation of aged petroleum sludge: Effect of inorganic fertilizer. Journal of Environmental Quality 30:395-403. Kamal, M., A.E. Ghalya, N. Mahmouda, and R. Cote. 2004. Phytoaccumulation of heavy metals by aquatic plants. Environment International 29 1029-1039. King, J.K., M.S. Harmon, T.T. Fu, and J.B. Gladden. 2002. Mercury removal, methylmercury formation, and sulfate-reducing bacteria profiles in wetland mesocosms. Chemosphere 46:859-870. Lindau, C.W., R.D. Delaune, and I. Devai. 2003. Rate of turnover and attenuation of crude oil added to a Louisiana Sagittaria lancifolia freshwater marsh soil. Spill Science and Technology Bulletin 8:445-450. Mailman, M. and R.A. Bodaly. 2005. Total mercury, methyl mercury, and carbon in fresh and burned plants and soil in Northwestern Ontario. Environmental Pollution 138:161- 166. Molisani, M., M.R. Rocha, W. Machado, R.C. Barreto, and D.L. Lacerda. Mercury contents in aquatic macrophytes from two reservoirs in the Paraíba do Sul: Guandú river system, SE Brazil. Brazilian Journal of Biology 66:101-107. Morel, F.M.M., A.M.L. Kraepiel, and M. Amyot. 1998. The chemical cycle and bioaccumulation of mercury. Annual Review of Ecology and Systematics 29:543-566. Moreno, F.N., C.W.N. Anderson, R.B. Stewart, and B.H. Robinson. 2004. Phytoremediation of mercury-contaminated mine tailings by induced plant-mercury accumulation. Environmental Practice 6:165-175. Moreno, F.N., C.W.N. Anderson, R.B. Stewart, and B.H. Robinson. 2008. Phytofiltration of mercury-contaminated water: volatilisation and plant-accumulation aspects. Environmental and Experimental Botany 62:78-85.

32 PBRP Annual Report | 2009 Bioavailability and Bioaccumulation of Mercury—Field Study

Newman, L.A., S.L. Doty, K.L. Gery, P.E. Heilman, I. Muiznieks, T.Q. Shang, S.T. Siemieniec, S.E. Strand, X. Wang, A.M. Wilson, M.P. Gordon. 1998. Phytoremediation of organic contaminants: a review of phytoremediation research at the University of Washington. Soil and Sediment Contamination 7:531-542. Otte, L.M. and D.L. Jacob. 2006. Constructed wetlands for phytoremediation: rhizofiltration, phytostabilisation and phytoextraction. Pages 57-67 in M. Mackova, D. Dowling, T. Macek. Phytoremediation and rhizoremediation. Springer, Dordrecht, The Netherlands. Rai, P.K. 2008. Heavy metal pollution in aquatic ecosystems and its phytoremediation using wetland plants: an ecosustainable approach. International Journal of Phytoremediation 10:133-160. Rai, P.K. and B.D. Tripathi. 2009. Comparative assessment of Azolla pinnata and Vallisneria spiralis in Hg removal from G.B. Pant Sagar of Singrauli Industrial region, India. Environmental Monitoring and Assessment 148:75-84. Sen, A.K. and N.G. Mondal. 1987. Salvinia natans as the scavenger of Hg(II). Water, Air, and Soil Pollution 34:439-446. Skinner, K., N. Wright, and E. Porter-Goff. 2007. Mercury uptake and accumulation by four species of aquatic plants. Environmental Pollution 145:234-237. Sparks, D.L. 2003. Environmental soil chemistry. 2nd edition. Academic Press, Amsterdam, The Netherlands. Su, Y., F.X. Han, J. Chen, B.B.M. Sridhar, D.L. Monts. 2008. Phytoextraction and accumulation of mercury in three plant species: Indian mustard (Brassica juncea), beard grass (Polypogon monospeliensis), and Chinese brake fern (Pteris vittata). International Journal of Phytoremediation 10:547-560. Sundberg-Jones, S. and S.M. Hassan. 2007. Macrophyte sorption and bioconcentration of elements in a pilot constructed wetland for flue gas desulfurization wastewater treatment. Water Air and Soil Pollution 183:187-200. Valiela, I., J.M. Teal, and R.J., Breteler. 1981. Bioavailability of mercury in several northeastern U.S. Spartina ecosystems. Estuarine, Coastal, and Shelf Science 12:155-166. Weis, J.S. and P. Weis. 2004. Metal uptake, transport and release by wetland plants: implications for phytoremediation and restoration. Environment International 30:685-700. Williams, J.B. 2002. Phytoremediation in wetland ecosystems: progress, problems, and potential. Critical Reviews in Plant Sciences 21:607-636. Xia, H., L. Wu, and Q. Tao. 2001. Water hyacinth accelerating the degradation of malathion in aqueous solution. China Environmental Science 21:553-555.

PBRP Annual Report | 2009 33 Bioavailability and Bioaccumulation of Mercury—Greenhouse Investigation

ABSTRACT Mercury is a toxicant of global concern due to its widespread distribution and its methylated form that is highly bioavailable and neurotoxic. The methylation of mercury is accomplished primarily by sulfate-reducing bacteria prevalent in wetlands and aquatic sediments, so these habitats require scrutiny to protect the human and ecosystem health from unanticipated risks. To this end, we created a wetland mesocosm and documented how mercury cycling, with an elevated loading rate, affected the mercury concentrations in the soil, water, and plant species. This understanding of mercury concentration is critical when responding to mercury contamination events as well as when evaluating potential phytoaccumulation applications.

PRIMARY OBJECTIVES The careful elucidation of mercury cycling with typical wetland soil conditions and plant species under elevated mercury loadings can inform two broad areas of applied research. Firstly, this information can benefit local habitat managers during a mercury contamination event. Secondly, the plant species used in this study can be evaluated for phytoremediation applications. The specific aims of this study are as follows: 1. To evaluate the potential transfer of mercury between native sediments, local vegetation, and a common invertebrate. 2. To assess interspecific differences demonstrated by tidal freshwater wetland vegetation in the bioaccumulation and translocation of inorganic mercury.

RESULTS TO DATE Findings thus far suggest that the three plant genera assessed (Polygonum, Pontederia, and Sagittaria) are relatively tolerant to elevated mercury loading rates in regard to their

photosynthetic status, which is indicated by negligible impacts to their net CO2assimilation rate and chlorophyll fluorescence. This tolerance likely results from the minimal levels of mercury that were translocated into aboveground tissue, even under the elevated mercury loading treatments. Importantly, analyses for all three species indicate that although a

34 PBRP Annual Report | 2009 Bioavailability and Bioaccumulation of Mercury—Greenhouse Investigation

substantial amount of mercury is accumulated into belowground tissues, a relatively small proportion of this mercury is translocated into the aboveground tissues. Sequential extraction of soils indicates that addition of inorganic mercury in an aqueous dissolved form was primarily recoverable in sulfide-bound and residual fractions rather than in the organic or water-soluble fraction. This indication is important as it suggests that a large portion of the mercury added to the mesocosms was rapidly sequestered into either belowground plant material or less microbially-available components of the soil matrix (e.g., precipitated as mercury-sulfides). These preliminary findings imply that the three plant species could be used to rehabilitate mercury-contaminated sediments where phytostabilization is the primary goal. The completion of total mercury analyses of sequential soil extractions and invertebrate tissue digestions by the end of March 2009 should clarify the feasibility of using these species for phytostbilization purposes.

TECHNOLOGY TRANSFER Project Goal: The primary objective of this research is to evaluate the cycling of mercury through soils, plants, and waters of an oligohaline wetland, typical of the southern United States under ambient and elevated mercury loadings. This research will simultaneously enable local mangers to assess risks to local habitat in the event of elevated mercury loadings as well as elucidate the capacity of local wetland plant species for phytoaccumulation applications.

Sustainability Questions 1. What are the impacts to the photosynthetic apparatus of several common wetland plant species to elevated mercury loadings? 2. Do these common wetland plant species bioaccumulate mercury, and if so, in what portion of the plant does it tend to be located? 3. How is the bioaccumulation of mercury by a common invertebrate affected by mercury concentration, plant species, and edaphic conditions?

Hypotheses Under elevated mercury loadings all plant species will experience impacts to the

photosynthetic apparatus as evidenced by reduced net CO2 assimilation and chlorophyll fluorescence. 1. It is anticipated that all of the plant species tested will bioaccumulate mercury into belowground components under elevated mercury loadings with minimal transfer to aboveground components. 2. Invertebrate mercury bioaccumulation is expected to be positively correlated with mercury concentrations in both ambient water and plant tissues and to a lesser extent with soil mercury concentrations.

PBRP Annual Report | 2009 35 Bioavailability and Bioaccumulation of Mercury—Greenhouse Investigation 200 control 2.5 ppm 180 5.0 ppm

160 ) /g g (n 140 g l H ta 120 o T e su 100 is T d n u 80 o gr ve o 60 b A 40 Aboveground Tissue Total Hg (ng/g) 20 (ng/g) Hg Total Tissue Aboveground (ng/g) Hg Total Tissue Aboveground

0 Polygonum punctatum Pontederia cordata Sagittaria lancifolia Species

8,000 control

2.5 ppm 7,000 5.0 ppm ) /g 6,000 g (n g H l 5,000 ta o T e su 4,000 is T d n u o 3,000 gr w lo e B 2,000 Belowground Tissue Total Hg (ng/g) Belowground Tissue Total 1,000

0 Polygonum punctatum Pontederia cordata Sagittaria lancifolia Species

Figure 1: The effect of species and mercury level on aboveground plant tissue total mercury (top fig.) and belowground plant tissue total mercury (bottom fig.), averaged across salinity level (mean +/- se).

36 PBRP Annual Report | 2009 Bioavailability and Bioaccumulation of Mercury—Greenhouse Investigation 14 control 2.5 ppm 5.0 ppm 12 ) 1 ‐ s 2

‐ 10

(umol

6 assimilation 2 CO 4 Net

0 Polygonum punctatum Pontederia cordata Sagittaria lancifolia Species

0.860 control

0.840 2.5 ppm 5.0 ppm ) 0.820 m /F Fv ( 0.800 ce n e sc re 0.780 lu l F yl h p 0.760 ro lo h C

Chlorophyll Fluorescence (Fv/Fm) 0.740

0.720

0.700 Polygonum punctatum Pontederia cordata Sagittaria lancifolia Species

Figure 2. The effect of mercury loading rate and plant species, averaged across salinity level on aboveground plant tissue mercury (top fig.) and belowground plant tissue mercury (bottom fig.) (mean +/- se).

PBRP Annual Report | 2009 37 Bioavailability and Bioaccumulation of Mercury—Greenhouse Investigation

Management Recommendations: This research will enable management officials to better assess risks presented by mercury contamination because the research provides data about the uptake of mercury by wetland plant species and an invertebrate tissue under elevated mercury loading scenarios in realistic wetland soils. Further, the capacity of these wetland plant species to be used in a phytoremediation context is assessed. Data thus far is promising in regard to using these species for phytostabilization where in situ stabilization rather than contaminant removal is optimal. Completion of total mercury analyses of sequential soil extractions and of invertebrate tissue digestions, anticipated at the end of this month, will hopefully further corroborate these findings.

Affected Parties: Agencies impacted by this research include the Louisiana Department of Natural Resources, the Louisiana Department of Environmental Quality, and the Environmental Protection Agency. Specific parishes that will benefit from this research are those with the plant species studied, including St. John the Baptist, Tangipahoa, and Livingston. Stakeholders directly benefiting from this research include hunters, shrimpers, fishermen, and recreational users of Lake Pontchartrain.

Summary Statement: The research conducted for this project is designed to provide key data that will allow for informed managers and stakeholders to safeguard the resources available in the Lake Maurepas Basin. Specifically, the findings will provide local mangers with information to assess the severity of habitat impacts in the event of local mercury contamination and elucidate the potential application of local plant species for phytoremediation purposes. Preliminary results indicate that elevated levels of mercury introduced into these simulated wetland systems is sequestered into belowground plant material and into the less biologically available portions of soil (e.g., insoluble mercury- sulfide compounds). This finding suggests that these species may be useful for phytostabilization approaches with mercury contaminated sediments.

38 PBRP Annual Report | 2009 Viability of Mitigation in Lake Maurepas and Manchac Swamp

Viability of Mitigation in the Lake Maurepas and Manchac Swamp Region

Robert Moreau1, Richard Campanella2, Lacy Landrum3, Randy Myers4, Gary Shaffer5, and William Bernard Wood5 1 Turtle Cove Environmental Research Station; Southeastern Louisiana University 2 Center for Bioenvironmental Research; Tulane and Xavier Universities 3 Lake Pontchartrain Basin Research Program; Southeastern Louisiana University 4 Independent Biologist 5 Department of Biological Sciences; Southeastern Louisiana University

ABSTRACT The purpose of this four-year project is to study the viability of wetland mitigation in Manchac/Maurepas Swamp in southeast Louisiana. This project has two aims: 1) to determine viable—or potentially viable—mitigation sites in the Maurepas/Manchac Swamp Region by defining what qualifies as a viable mitigation site and 2) to identify the hurdles that might impede mitigation success and to identify solutions for overcoming those hurdles. LIDAR data was analyzed and compared with Thematic Mapping (TM) and ground truthing of vegetation and elevation variables. Preliminary results show that LIDAR is more accurate than TM for identifying degraded wetland sites (100% of the time). Identification of relic (74%) and sustainable (79%) sites were less accurate but still beneficial. There are many competing programs to mitigation, mostly under the annual Farm Bill, but prices for mitigation sites have risen dramatically in the past several years.

OBJECTIVES & SITE DETAILS The overall goal of this project is to create a series of tools that will help stakeholders identify viable mitigation sites in the Manchac/Maurepas Swamp. Specific objectives include the following: 1. Develop a White Paper to provide mitigation information to local, state, and federal agency representatives involved in mitigation in the region. The white paper will describe a new technique to map the viability of suitable mitigation sites within the study area. The new technique to be tested uses LIDAR data of tree canopy height as well as information and maps of key factors, such as a) ownership of large tracts of lands (shown as private vs. public), b) suitability of sites for mitigation (based on LIDAR produced tree canopy data, fresh water inputs including potential waste water effluent sources in the same area, elevation, and salinity data), and other geographic and demographic data. 2. Develop a Short How-To Manual to translate the white paper information to landowners, developers, and the general public. 3. Deliver a mitigation workshop at the end of the process, and at a location near the study site, that promotes these and other educational materials to the stakeholders

PBRP Annual Report | 2009 39 Viability of Mitigation in Lake Maurepas and Manchac Swamp

addressed in the study (e.g., agency representatives, landowners, developers, nonprofit groups, and the general public). 4. Develop a website to serve as an information clearinghouse for the whitepaper, manual, workshop materials, and other education materials related to mitigation.

The study site was defined as the 2ft contour around the Manchac/Maurepas swamps, consisting of a terrestrial area approximately 460.5 sq miles (294,735 acres). Establishment of the 2ft contour for the study site is appropriate because it divides the land neatly between undeveloped marsh and swamp habitat from more developed agricultural sites. The designation is also cadastrally appropriate.

RESULTS The project began in February 2006, a delayed start due to the storms of 2005, and pertinent information learned thus far has been helpful in fine-tuning the remainder of the study. For instance, it was determined early on that viable mitigation sites in the study area would be few and far between due to low elevations and high salinity, underlying reasons that the cypress forests of the Lake Maurepas WMA are mostly in a “relic” state. However, potential sources of fresh water from small sewage effluent sites surrounding the study site, sites above the two foot contour, could be a compensating factor. Therefore these sources and their locations will be placed in the GIS model along with the LIDAR data on tree canopy height, land ownership, and other relevant variables. LIDAR data processed during the past year has been, for the most part, beneficial because it accurately describes tree canopy height and density. LIDAR is especially accurate in designated quads; however, LIDAR is less accurate when mapping multiple quads, and some of the LIDAR data is questionable because of obvious differences between the model and what is really there.

Key findings and results learned to date that have helped establish, fine-tune, and in some cases re-direct the research are organized into three categories: 1) Start-up and literature review results, 2) Specific results of LIDAR mapping, and 3) Broad-scale results of LIDAR mapping as compared to Thematic Mapping.

Start-Up and Literature Review Results Growth rates along the north shore in St. Tammany Parish have been approximately five times that of the national average, and habitat developed there is mainly marsh, wetland forest, and scrub brush wetland (making the Manchac/Maurepas Swamp a natural area for comparable mitigation). Growth rates in St. Tammany Parish are even higher since the storms of 2005 (rates of 40% growth though 2007). The population growth rates (seen as a proxy for development) on the north shore, particularly in St. Tammany Parish, point toward the need to have comparable mitigation sites available. The Manchac/Maurepas Swamp is appropriate from that standpoint if, of course, appropriate and suitable mitigation sites can be identified, approved for mitigation, and are sustainable.

Ownership of land has a significant impact on mitigation potential. In general, public lands may be more difficult to designate as mitigation sites, mainly because such lands are already assigned some sort of management status and may not be appropriate to “upgrade” into mitigation because they are already being conserved or preserved in specific ways under an

40 PBRP Annual Report | 2009 Viability of Mitigation in Lake Maurepas and Manchac Swamp

existing Farm Bill or other program. Land ownership designation in the study site is currently broken down into private lands (69%) and public lands (31%), and that ratio is continually changing in a positive direction for public lands (i.e., LDWF has in the past year purchased more land for inclusion in their Maurepas WMA). Regarding public land ownership, one interesting development is the capability of public agencies, like LDWF, to purchase land for public use with mitigation funds, thereby having the land also be “preserved” as a mitigation site through the mitigation process. For example, the purchased land, if in good shape environmentally, may provide a large mitigation ratio and may be used to offset development. So the land is purchased as an already completed mitigation project, but it has a higher environmental value than traditional degraded land that has to be mitigated.Public lands may also be more difficult to put into mitigation than private lands because of political issues involving price and fund allocation within the agency, as well as the competition that then results between private and public landowners in the broader mitigation arena. The value of public lands in the mitigation arena should not be discarded, but rather reviewed on a case-by-case basis.

Impediments to mitigation in the study site include the following: appropriate land suitability (areas where trees have a chance to grow), landowner ignorance of mitigation, complexity of mitigation guidelines, access to appropriate sites, and, perhaps most importantly, competing landowner choices. Private landowners may choose from many government-sponsored programs that compete with mitigation, such as the Conservation Reserve Program (CRP), Wildlife Habitat Improvement Program (WHIP), forthcoming Carbon Sequestration Programs, and others. Most of these programs are sponsored through annual Farm Bill Policy programming and do not allow for “double dipping,” or having land placed under more than one designated category which might include mitigation. Another competing issue to mitigation is landowners’ right and ability to use their land on the “open market” for various uses, including leasing (hunting and fishing or mineral rights), outright sale of land, and other market-based types of activities, although some of these may allow for simultaneous mitigation as well. One example of permissible double-dipping is leasing the mineral rights of land while also designating that same land as a mitigation site.

Currently, there are only two mitigation sites in the Maurepas Swamp area of the study site, and these sites have historically averaged about $10,000 per acre. One site is a private bank that entails 1,000 acres of cypress replanting; the other is a WLF site purchased in an already “preserved” (mitigated) state of about 2,500 acres. On the Manchac side, there is one site, managed by Southeastern Louisiana University, but with limited success rates for cypress replanting, due to high salinities and low elevations, and with limited acres (180) in play. The lack of more sites suggests that a major impediment to mitigation on the Manchac side is site unsuitability (high salinities/low elevations) and the low probability for long-term success of cypress plantings. But preliminary results from recent research (Day and Shaffer 2008) has shown that proper amounts of waste water effluent delivered to areas—even those areas of low(er) elevation and high(er) salinities—can aid in development and growth of young cypress trees. This has prompted the researchers to include those areas as data points in the GIS mapping exercise. Overall, mitigation sites in south Louisiana have seen dramatic increases in per acre prices; in late 2008 prices were as high as $40,000 per acre (a 400% increase since this study began in 2006). In fact, mitigation sites in other ecosystems, such as Pine Savannahs and Bottom Land Hardwood Forests, have seen similar increases in price and value.

PBRP Annual Report | 2009 41 Viability of Mitigation in Lake Maurepas and Manchac Swamp

Specific Results of LIDAR Mapping Certain LIDAR variables (SDheight, Mground, CVground), with some groundtruthing variables added (TUPstem and OTHERstem), have provided excellent separation of Degraded areas (100% of time) within our sample study site. Under LIDAR mapping, separation of Relic (74%) and Sustainable (79%) areas are not as clean as Degraded (100%) areas, but neither is it clean in reality (difficult to assess under any method).

There are a few areas of the LIDAR generated maps that do not correlate with groundtruthing data. These areas are currently undergoing a review by Richard Campanella, Gary Shaffer, and Bernard Wood. A preliminary assessment of these areas by Richard Campanella concludes that raw LIDAR data is extremely accurate in localized areas. However, when large tracts are arranged, discrete discrepancies that are unaccounted for may be prevalent and may limit large-scale mapping potential, mainly due to unknown discrepancies between tracts.

Results of LIDAR Mapping as Compared to Thematic Mapping LIDAR Mapping (measuring tree and terrain height) and Thematic Mapping (measuring spectral reflectance) complement each other and both should be used, along with groundtruthing, to accurately map swamp and marsh habitats. LIDAR exhibits several benefits over TM. First, LIDAR’s fine resolution, with 5m pixels for the terrain elevation and 1-2m pixels for vegetation height, captures levels of detail that TM, with 30m resolution, cannot discern. Also LIDAR plus ground truthing bears extremely high correlations (r = .95 and above) with groundtruthed forest inventory data much higher than TM’s correlations. LIDAR is less weather dependent because, as an active system, it penetrates cloud cover while passive sensors are at the mercy of the weather.

However, LIDAR data for Louisiana (captured in 1999-2000) is seven years old now, and much natural and anthropogenic change has occurred over that time period. New LIDAR data is expensive to capture (must hire it out through private firms); whereas, TM or other remotely sensed space-borne spectral data are more economical. Satellite LIDAR data exists, but it has a much coarser resolution than data gathered through LIDAR fly by. LIDAR may be most suitable for localized mapping given the accuracy of data sets in this project. However, some sectors of adjoining data sets have unaccountable and large discrepancies (in excess of 20ft in some cases). These problems may limit large scale mapping with LIDAR alone. So, in most cases, LIDAR plus some groundtruthing is a method that accurately maps S, D, and R with a higher degree of resolution than Thematic Mapping (TM) in localized sectors. Final analysis of the statistical relationships between the field data and the LIDAR data, including those areas that do not correlate well at all, will inform us as to what extent we may be able to use the LIDAR dataset available for the entire basin as a valuable base map for directing mitigation banking decisions.

FUTURE PLANS During the final year of this project, we will accomplish the following tasks: 1. Determine the reason (if possible) that LIDAR data in some cases does not correlate at all with groundtruthing variables (several of these instances are under study now by Campanella, Shaffer and Wood);

42 PBRP Annual Report | 2009 Viability of Mitigation in Lake Maurepas and Manchac Swamp

2. Mesh and refine LIDAR/Ground Truthing relationships in a way that provides the most accurate and comprehensive measurements of tree canopy and ground elevations (Campanella, Shaffer and Wood); 3. Incorporate above relationships into overall GIS model, which will also be refined by obtaining up-to-date land ownership data, potential waste water effluent sources, and data from other variables that will be used in the model (Campanella, Myers, Shaffer and Wood); 4. Complete drafts of White Paper, How-To Manual and Mitigation Website (Moreau, Campanella, Shaffer, Wood, Myers and Landrum); 5. Design and conduct mitigation workshop in local area, including compilations of participant information (Moreau, Campanella, Shaffer, Wood, Myers and Landrum); 6. Compile lists of designated agencies/entities/persons for distribution of the materials and for inclusion on website as contacts (Moreau, Myers and Landrum); 7. Determine the extent by which this model (LIDAR) can accurately map other areas of the Lake Pontchartrain Basin ecosystem (Campanella, Shaffer, Wood and Myers).

TECHNOLOGY TRANSFER This project is, in itself, an exercise in technology transfer. Information learned in the research of mitigation potential in the Manchac/Maurepas Swamps will be disseminated to a wide group of decision-makers, including natural resource agency personnel, landowners, financial interest entities, community and environmental groups, local politicians, and other interested members of the public. Information will be transferred through several project deliverables, including journals (for the white paper), a website, workshops, and online and print versions of the how-to manual.

This project is important from a restoration and sustainability standpoint in that it will provide tools for identifying viable mitigation, and hence restoration, sites in an area in need of such restoration. The project gives policymakers accurate and timely information about where viable mitigation sites might exist as development further encroaches on the north shore communities. Local, available, and suitable sites for mitigation is of paramount importance right now in our region as development for both housing and infrastructure (including levees for hurricane protection) continues to increase. It is now widely known that the U.S. Army Corps of Engineers (USACE) is now actively pursing mitigation sites as they begin planning the development of enhanced levee systems throughout southeastern Louisiana because many of these new levee sites will require mitigation.

Agencies impacted by this information include LDWF, LDEQ, LDNR, USACE, NRCS, and USEPA, among others. Besides landowners and developers, various stakeholders, such as hunters, fishermen, and wildlife enthusiasts, need to be aware of appropriate mitigation alternatives. All parishes within the Lake Pontchartrain Basin will be impacted and thus will benefit from this information. It is hoped that this research will aid stakeholders as they decide which mitigation activities are appropriate and affordable for the region.

PBRP Annual Report | 2009 43 Hydraulic Conductivity and Vulnerability to Xylem Cavitation of Baldcypress

Hydraulic Conductivity and Vulnerability to Xylem Cavitation of Baldcypress (Taxodium distichum) along a Salinity Gradient as Indicators for Restoration Success

Volker Stiller Department of Biological Sciences; Southeastern Louisiana University

ABSTRACT In the past, considerable efforts have been undertaken to restore baldcypress trees around Lake Pontchartrain. Vast amounts of cypress trees have been planted with varying rates of success. This project aimed to support future restoration efforts of baldcypress by investigating the role of plant hydraulics on restoration success. This investigation had two specific goals: 1) to evaluate the amount of “drought” stress young baldcypress trees are subjected to, due to increased salinity in their habitat, and 2) to evaluate if the xylem of baldcypress possesses the inherent plasticity to acclimate to increased salinity. The answers to these two questions provide more detailed parameters for deciding when and where to plant cypress trees.

PRIMARY OBJECTIVE & METHODOLOGY This project aimed to support future restoration efforts of baldcypress by investigating the role of plant hydraulics on restoration success. This investigation had two specific goals: a) to evaluate the amount of “drought” stress young baldcypress trees are subjected to, due to increased salinity in their habitat, and b) to evaluate if the xylem of baldcypress possesses the inherent plasticity to acclimate to increased salinity. The answers to these two questions provide more detailed parameters for deciding when and where to plant cypress trees.

This project achieved its goals through four simple experiments. The first experiment evaluated the vulnerability of baldcypress to xylem dysfunction by establishing so-called “vulnerability curves.” These curves show the percent loss of hydraulic conductivity in relation to negative xylem pressure and are a fundamental parameter to understand a species’ ability to grow in a specific environment. In two field experiments, the in situ hydraulic conductivity and the amount of xylem cavitation in baldcypress was measured along a salinity gradient. In addition to these field studies, a greenhouse study was conducted to evaluate the ability of cypress trees to acclimate to increased salinity. Baldcypress was grown in the Southeastern greenhouse, and the salinity of the irrigation water was gradually increased from 0ppt to 5ppt. Transpiration and growth measured throughout the experiment detected possible acclimation to salinity.

RESULTS FOR 2006 – 2007 Field Experiment (summer 2006): Two field sites were selected for field measurements. The control site was located on the Tchefuncte River (Fairview-Riverside State Park,

44 PBRP Annual Report | 2009 Hydraulic Conductivity and Vulnerability to Xylem Cavitation of Baldcypress

30°24’42”N, 90°08’35”W), the elevated salinity site (ES-site) was located on the North Shore of Lake Pontchartrain (Fontainebleau State Park, 30°20’20”N, 90°02’55”W). In both sites, leaf water potential as well as native xylem embolism was measured on 6-8 plants, and stem segments were collected and brought to the lab for vulnerability curve measurements. Salinity at the ES-site was 5.9 ppt and was 2.3 ppt at the control site. This relatively high salinity at the control site was due to low river water levels and to measurements taken at noon shortly after a high tide. Midday leaf water potentials were significantly different at both sites (–1.62±0.12 MPa at the control site and –2.12±0.34 MPa at the ES-site).

Vulnerability curves measured on plants from both sites were not significantly different (50% loss of hydraulic conductivity at –2.75 MPa and –2.50 MPa for control and ES-site, respectively). In situ xylem embolism was negligible at both sites and agreed well with what the vulnerability curves predicted. In situ hydraulic conductivity of the collected stem segments was not different at both sites.

Greenhouse Experiment (summer 2006 and spring/summer 2007): Preliminary experiments showed that greenhouse-grown baldcypress seedlings were very vulnerable to xylem cavitation (50% loss of hydraulic conductivity at –1.59 MPa xylem pressure for one- year old stems) and that new growth was even more vulnerable (50% loss at –1.23 MPa). During the summer of 2006, approximately 50 one-year-old, bare-root seedlings were grown under ambient conditions in the greenhouse of Southeastern Louisiana University. After plants were established, they were split into two groups and irrigated twice daily, half with salt water and the other half (control) with fresh water. The concentration of the salt solution was gradually raised over an 8-week period from 2ppt to 8ppt and was then kept constant at 8ppt for the remainder of the experiment (10 weeks). Throughout the growing season, transpiration rates, stomatal conductances, and relative diameter growth rates were monitored. At the end of the growing season, plants were harvested, and vulnerability curves and wood densities were measured on current year stem segments.

Salinity reduced transpiration rates and stomatal conductance approximately by half (E = 2.5±0.8 and 4.7±1.0 mmol s-1 s-2; gS = 215±64 and 436±127 mmol s-1 s-2 for salt and control plants, respectively. Mean ± SD, n = 5). Reduced transpiration rates were mirrored by lower relative diameter growth rates in salt-treated plants (RGR = 0.09±0.03 mm/mm and 0.15±0.07 mm/mm for salt and control plants, respectively. Mean ± SD, n = 9). At the end of the growing season (late October to early November), 6-10 treatment and control plants were harvested, and xylem vulnerability to cavitation was measured. Vulnerability curves showed that the P50 of salt-treated plants was significantly more negative (–2.57 MPa) compared to control plants (–2.25 MPa; n = 9, p<0.001, F-test). Wood density of salt-treated plants was greater (0.33±0.02 g/cm3) compared to control plants (0.29±0.02 g/cm3; n = 9, p<0.05). Although P50, RGR, and wood density were measured on the same plants, wood density was correlated only with RGR (p<0.05) but not with P50.

RESULTS FOR 2008 During the spring of 2007, I decided to repeat the greenhouse experiment to obtain a more robust data set and to implement an additional drought stress treatment, which should determine if the effects of salt and drought treatments are comparable.

PBRP Annual Report | 2009 45 Hydraulic Conductivity and Vulnerability to Xylem Cavitation of Baldcypress

In the Southeastern greenhouse, 100 one-year-old, bare-root baldcypress seedlings were grown under ambient conditions. During the first 8 weeks of seedling establishment, all plants were irrigated frequently to avoid drought stress. After plants were established, they were divided into three groups: 1) control plants were watered twice daily, 2) drought plants were watered once per week, and 3) plants subjected to a salinity treatment were irrigated twice daily with a salt solution. The concentration of the salt solution was gradually raised over a 4-week period from 2 to 6 ppt and was then kept constant at approximately 6 ppt for the remainder of the experiment. To avoid salt build-up in the soil, plants of the salinity treatment were heavily watered with fresh water once per week.

Basal stem diameters were measured at the beginning of the treatments (May), in the middle of the growing season (August), and at the end of the growing season when plants were harvested for hydraulic measurements (November). Relative diameter growth rates were

calculated for the first and second half of the growing season (RGR May-August and RGR August- November, respectively), as well as for the entire season (RGR May-November). In the middle of the growing season and at the height of the weekly drought stress, CO2 assimilation rate (A), transpiration rate (E), and stomatal conductance (gS) of young, fully expanded leaves of 11 representative plants from each treatment were measured using a LICOR LI-6400 photosynthesis system. Midday leaf water potential was measured with a Scholander-type pressure chamber on comparable leaves of the same plants. Soil was sampled, and soil water potentials were measured. Soil-to-leaf hydraulic conductance of the

plants (kL) was calculated as: kL = E / (ψ soil – ψ leaf ) where E is the transpiration rate of the plants and ψ is the water potential of the soil or the leaves.

At the end of the growing season, all treatments were irrigated with fresh water twice daily for one week to allow stressed plants to recover and to minimize native embolism. Ten representative plants from each treatment were harvested, and their vulnerability to xylem cavitation (vulnerability curves) was measured on current-year stem segments. After vulnerability measurements were complete, the wood density of the stem segments was determined. The results of my measurements are summarized in tables 1 and 2.

Table 1. Basal stem diameters and relative daily diameter growth rates of one-year-old baldcypress seedlings. Different letters underneath the values indicate homogeneous sets of means determined with a Tukey’s HSD test after a one-way ANOVA tested significant at the 0.05 level. Sample sizes were n = 19 for salt treatment and n = 25 for drought and control plants. Stem diameter Salt Drought Control

May (D May), mm 12.0±0.45 11.5±0.29 11.2±0.38 a b a a

August (D August), mm 15.5±0.47 14.1±0.39 17.3±0.58 c d b c d

November (D November), mm 16.3±0.55 17.2±0.48 25.6±0.72 c d d e

46 PBRP Annual Report | 2009 Hydraulic Conductivity and Vulnerability to Xylem Cavitation of Baldcypress

Relative diameter growth rate -3 -3 -3 May—Aug (RGR May-August), (3.5±0.3) x 10 (2.6±0.3) x 10 (6.6±0.5) x 10 -1 -1 mm mm d a a b c -3 -3 -3 Aug—Nov (RGR August-November), (0.6±0.1) x 10 (2.3±0.1) x 10 (5.1±0.2) x 10 -1 -1 mm mm d e a b d -3 -3 -3 May—Nov (RGR May-November), (2.0±0.1) x 10 (2.8±0.2) x 10 (7.3±0.4) x 10 -1 -1 mm mm d b a b c

Table 2. Gas exchange parameter (A, E, gS), water potential (ψ leaf, ψ soil), soil-to-leaf hydraulic conductance (kL), and average and specific leaf area (AL, SLA) of baldcypress seedlings. Stem- specific hydraulic conductivity (KS native), wood density (ρ w), and P50 of present-year stem segments used in hydraulic measurements. Values are mean ± SE. Sample sizes for A, E, gS, ψ leaf and kL were n = 11 for all three treatments. Sample sizes for ρ w, DS, KS native, and P50 were n = 10 for salt and drought treatments and n = 9 for control plants. AL and SLA sample sizes were n = 13, 21 and 21 for salt, drought, and control plants, respectively. As ψ soil was measured on pooled soil samples, no statistics are given for this parameter. Different letters (in a row) indicate means are significantly different at the 0.05 level, Tukey’s HSD following one-way ANOVA. Italicized letters indicate significant differences at the 0.06 level. Salt Drought Control

-1 -2 CO2 assimilation rate (A), μmol s m 4.6±0.76 a 2.5±0.52 a 9.2±0.46 b Transpiration rate (E), mmol s-1 m-2 3.9±0.42 a 2.5±0.23 a 8.7±0.73 b -1 -2 Stomatal conductance (gS), mmol s m 84±9.4 a 54±5.2 a 191±22.9 b

Midday leaf water potential (ψ leaf), MPa -1.8±0.05 a -2.0±0.05 b -1.3±0.03 c

Soil water potential, pooled (ψ soil), MPa -0.93 -0.64 -0.34

Soil-to-leaf hydraulic conductance (kL), 4.9±0.69 a 1.9±0.20 b 8.8±0.84 c mmol s-1 m-2 MPa-1

2 Average leaf area per leaf (AL), cm 12.0±1.6 a 6.4±0.5 b 14.7±0.9 a Specific leaf area (SLA), m2 kg-1 13.3±0.4 a 11.3±0.3 b 14.0±0.3 a

Stem-specific hydraulic conductivity 1.01±0.10 a 0.49±0.06 b 1.51±0.08 c -1 -1 -1 (KS native), kg m s MPa -3 Wood density (ρ w), g cm 0.31±0.01 a 0.34±0.003 b 0.27±0.01 c

P50, MPa -2.50±0.08 a -2.88±0.07 b -2.01±0.04 c

Gas exchange rates of stressed plants were reduced by approximately 50% (salt) and 70% (drought), resulting in a 50-60% reduction in diameter growth for both treatments. Stem- specific hydraulic conductivity (KS native) of stressed plants was 33% (salt) and 66% (drought) lower than controls, and we observed a strong positive correlation between KS native and gas exchange (fig. 1).

PBRP Annual Report | 2009 47 Hydraulic Conductivity and Vulnerability to Xylem Cavitation of Baldcypress

10 control salt

) drought

-2 8 m -1 6 mol s μ

( 4 A

2 a 0 300

250 ) -2

m 200 -1

150 (mmol s (mmol

S 100 g

50 b 0 12

10 ) -2

m 8 -1

(mmol s (mmol 4 E

2 c 0 0.0 0.5 1.0 1.5 2.0 K (kg m-1 s-1 MPa-1) S native Figure 1. The relationship between (a) CO2 assimilation rate (A), (b) stomatal conductance (gS) and (c) transpiration rate (E) and native hydraulic conductivity (KS native). Mean values from table 1. Error bars show the upper and lower boundaries of the 95% confidence intervals. Sample sizes for A, gS and E were n = 11 for all treatments. Sample sizes for Knative were n = 10 for salt and drought treatments and n = 9 for control plants.

In addition, I found a strong relationship between CO2 assimilation rate (A) and the soil-to- leaf hydraulic conductance (kL). The relationship was identical for all treatments, suggesting that the moderate salt stress (as well as drought) did not affect the photosynthetic biochemistry of leaves, but rather reduced A via stomatal closure (fig. 2).

48 PBRP Annual Report | 2009 Hydraulic Conductivity and Vulnerability to Xylem Cavitation of Baldcypress

10 ) -2 r2 = 0.73 m 8 -1

r2 = 0.76 mol s mol 6

μ 2

( r = 0.79 A 4 control salt 2 drought

0 0246810121416 k (mmol s-1 m-2 MPa-1) L

Figure 2. The relationship between soil-to-leaf hydraulic conductance (kL) and CO2 assimilation rate (A) in Taxodium distichum. Correlations for all treatments were highly significant (P < 0.01). The r2 values within the figure are for linear regressions (solid lines). In addition to the regression analysis, I fitted an exponential rise to a maximum function (dashed line) to the pooled data [y = -0.82+16.46*(1-exp (-0.103x), r2 = 0.87].

Lower KS native of stressed plants was associated with increased wood density and greater resistance to xylem cavitation. Vulnerability curves of all 3 treatments are shown in figure 3.

Xylem pressures causing 50% loss of hydraulic conductivity (P50) were –2.88±0.07 MPa (drought), –2.50±0.08 MPa (salt), and –2.01±0.04 MPa (control). P50s were strongly correlated with wood density (r = –0.71, P < 0.01, fig. 4) and KS native (r = 0.74, P < 0.01, fig. 4, insert).

PBRP Annual Report | 2009 49 Hydraulic Conductivity and Vulnerability to Xylem Cavitation of Baldcypress

100 control salt 80 drought

PLC (%) PLC 40

-5 -4 -3 -2 -1 0

xylem pressure (MPa) Figure 3. Vulnerability curves of current-year stems in Taxodium distichum. Curves show percentage loss of hydraulic conductivity (PLC) with decreasing xylem pressure for drought (n = 10), salt (n = 10) and control plants (n = 9). Means and SE (error bars are partly covered by symbols) are shown for greater clarity; however, Weibull functions were fitted to the raw data of each treatment (solid line = control, dotted line = salt, dashed line = drought).

-1.6

-1.8 control salt -2.0 drought

-2.2 -1 -1 -1 KS native (kg m s MPa ) 0.20.40.60.81.01.21.41.61.8 -2.4 -1.6 (MPa) -1.8 50 P -2.6 -2.0 -2.2 -2.4

-2.8 (MPa)

-2.6 50 P -2.8 -3.0 -3.0 -3.2 -3.2 0.10 0.15 0.20 0.25 0.30 0.35 0.40 wood density, ρ (g cm-3) W Figure 4. The relationship between P50 and wood density (ρ w) of current-year stem xylem in 2 Taxodium distichum (r = 0.50) and between P50 and stem-specific native hydraulic conductivity (KS native, insert). Error bars indicate 1SE and are partly covered by symbols (n = 10 for salt and drought treatments and n = 9 for control plants, r = 0.74, P < 0.01).

50 PBRP Annual Report | 2009 Hydraulic Conductivity and Vulnerability to Xylem Cavitation of Baldcypress

The results of this study show that baldcypress seedlings react to moderate soil salinity and drought stress similarly. Both treatments reduced the plants’ gas exchange, diameter growth rates, and hydraulic conductivity while increasing wood density. These results indicate that stressed plants partitioned their biomass in a way that strengthened their xylem and reduced vulnerability to xylem cavitation. Hence, these seedlings could be better suited to be planted in environments with elevated soil salinity. Drought had an even more pronounced effect on cavitation vulnerability than salinity, suggesting that nurseries could produce “stress- acclimated” seedlings simply by reducing irrigation amounts rather than contaminating soils in their nursery beds with salt applications. Whether stress-acclimated seedlings truly have lower mortality rates in saline environments needs to be evaluated in future field studies.

TECHNOLOGY TRANSFER This study aimed to support future restoration efforts of baldcypress by investigating the role of plant hydraulics on restoration success. This investigation had two specific goals: 1) to evaluate the amount of “drought” stress young baldcypress trees are subjected to, due to increased salinity in their habitat, and 2) to evaluate if the xylem of baldcypress possesses the inherent plasticity to acclimate to increased salinity. The answers to these two questions provide more detailed parameters for deciding when and where to plant cypress trees.

Our results revealed that midday water potentials under saline field conditions are significantly lower than “regular” baldcypress seedlings (obtained as Louisiana Department of Agriculture nursery stock) can tolerate. However, our greenhouse study revealed that seedlings can acclimate and those acclimated should be better suited to be planted in saline environments. Especially important is the result that salt and drought treatments have a comparable effect on the plants because nurseries could produce “stress-acclimated” seedlings simply by reducing irrigation amounts rather than contaminating soils in their nursery beds with salt applications.

Hence, the results from this study not only provide new insights into the limitations of water transport in baldcypress, but also lead to a better understanding of the hydraulic constraints on restoration efforts. These results should be of interest to all groups and management agencies involved with the restoration of baldcypress.

Furthermore, results from the greenhouse “acclimation study” should help develop possible preconditioning treatments for cypress trees in nurseries. Such preconditioning treatments could lead to hardier cypress trees that would be better suited to withstand the transplantation shock when planted in brackish or saline habitats and could be a valuable tool for nurseries providing the plant material for restoration projects.

PBRP Annual Report | 2009 51 Salinity as a Stressor of the Freshwater Turtle

Salinity as a Stressor of the Freshwater Turtle, Trachemys scripta, in the Lake Pontchartrain Basin

Roldán Valverde Department of Biological Sciences; Southeastern Louisiana University

ABSTRACT The Lake Pontchartrain System, constituted by Lakes Pontchartrain and Maurepas, are under increasing influence of salt water intrusion from the Gulf of Mexico. This influx of saltwater is thought to place stressful physiological constrains on the biota of the basin, particularly of freshwater species. Research efforts during the first year of the grant have focused on solving logistic aspects of field and lab work, such as developing an effective method to trap turtles and establishing the best set up to expose turtles to various salinities in the lab and testing them for stress. During the second year we focused on trapping turtles and conducting the salinity exposure experiments of wild caught turtles. Currently, laboratory salinity exposure trials have been completed for lab raised turtles. Trapping and salinity exposure trials continue with wild turtles. We have measured corticosterone, the stress hormone, in many of the samples and our efforts still continue.

PRIMARY OBJECTIVE The primary objective of this study is to demonstrate that the red-eared slider freshwater turtle is a good physiological model to serve as an environmental sentinel with regard to salinity in the Lake Pontchartrain Basin. This objective will be accomplished by testing the following hypotheses. 1. Red-eared slider turtles living at higher salinity exhibit a hyperactive adrenal gland. This hypothesis is being tested using animals captured in the field. A hyperactive adrenal gland would suggest that the turtle lives in a stressful environment. 2. Increases in environmental salinity induce an endocrine stress response in the red-eared slider turtle. This hypothesis is being tested in the laboratory using captive-raised animals. 3. The red-eared slider turtle is more abundant in less saline environments within the Lake Pontchartrain Basin. This hypothesis predicts that red-eared slider is more abundant in a Lake Maurepas’ tributary where salinity is lowest within the Lake Pontchartrain Basin.

RESULTS 1. Red-eared slider turtles living at higher salinity exhibit a hyperactive adrenal gland. To test our first hypothesis, during the past year, we have continued with our trapping of wild turtles. Unfortunately, last spring the Bonnet Carré Spillway was open,

52 PBRP Annual Report | 2009 Salinity as a Stressor of the Freshwater Turtle which significantly lowered the salinity of the Lake. This prevented us from finalizing our trapping efforts. We are currently planning to return to Slidell during the month of April 2009, to trap the last ten turtles and finalize our field experiments. After this, we plan to make the final measurements of corticosterone and to then analyze the data and write a final report. This project will constitute the main thesis of one of my graduate students, Diana Solis.

2. Increases in environmental salinity induce an endocrine stress response in the red- eared slider turtle. During this past year, we have also made progress testing our second hypothesis concerning the endocrine stress response. We have succeeded at developing a reliable method to collect blood from the turtles. This breakthrough has allowed us to complete all our exposure experiment with lab-raised turtles. These experiments included testing the effect of various exposure times (12 days, 1 month, 3 months) on the stress system of the turtles to determine which exposure time will show a physiological response. We are currently in the process of validating the corticosterone assay kit to measure stress hormone in all samples collected. Our goal is to have the assay validated and all samples analyzed by the end of the summer in 2009.

3. The red-eared slider turtle is more abundant in less saline environments within the Lake Pontchartrain Basin. We had originally planned to address our third hypothesis by deploying several basking platforms at key locations representing the salinity gradient of the Lake Pontchartrain Basin. However, after testing several platforms at the three originally proposed sites, we decided to discontinue our efforts because the turtles did not use the platforms as actively as we expected. Thus, the platforms appear inadequate to undertake this part of the study. In addition, the weak construction of the platforms (Styrofoam) led the structures to suffer damages, mostly caused by wind.

Instead of this approach, we have recently developed a new method to generate a relative index of abundance. The methodology consists of determining a trapping per unit time effort. The method will be based on the number of traps deployed and the number of turtles trapped at every site with regard to trapping time (i.e., how long the traps were set to capture a given number of turtles). Theoretically, the number of turtles trapped per unit of time and number of traps is a direct function of turtle abundance, where the index is expected to be very high in areas where turtles are more abundant and then to fall in greater numbers in the traps. Because we are limited to trapping turtles in the Slidell area, we do not have data to report yet.

FUTURE PLANS Since the beginning of the project we have overcome several drawbacks. First, we identified the best areas on the west side of Lake Maurepas to trap turtles. Second, we have developed and refined the blood sampling technique that we have successfully used to complete the salinity exposure experiments and will use to complete the experiments with turtles captured in the wild. Third, the lack of a graduate student supported by this grant has been compensated by hiring two very competent undergraduate students: Leslie Francks, a senior who graduated last May, and Judd Thompson, a senior currently working with the project. Both of these students are certified in the use of University boats through the Turtle Cove

PBRP Annual Report | 2009 53 Salinity as a Stressor of the Freshwater Turtle

boat safety program and have been heavily involved in the field and the laboratory components of the project. In addition, last spring I gave this project to my graduate student, Diana Solis, as her thesis project. She expects to conclude the field and lab components this spring to graduate in August of 2009. Another undergraduate, Melissa Juno, a junior, has been dedicated at helping with the maintenance of the turtles in the vivarium facilities. All students are now trained on field techniques regarding turtle capture as well as lab techniques involving turtle handling, blood sampling and processing, and are currently training on the conduction of hormonal measurements in the samples taken from the multiple experiments. We plan to have completed all hormonal measurements by the end of spring 2009. Soon after that we plan to publish our results in a scientific journal.

TECHNOLOGY TRANSFER The overall goal of this study is to demonstrate that the red-eared slider freshwater is an excellent animal model for studying the physiological effects of elevated salinity due to saltwater intrusion into the Lake Pontchartrain basin. This turtle works well as a model because its species is abundant in the basin, is not adapted to salt water, and is easy to manipulate.

Specifically, we aim to answer the following questions: 1. Does salinity induce a neuroendocrine stress response in freshwater vertebrates? Our preliminary data indicates that this is the case. 2. Does salinity limit the abundance and distribution of freshwater vertebrates in the basin? Our preliminary data indicates that this is the case. 3. Can the data generated by the study provide managers with a dynamic tool to assess the impact of increased saltwater intrusion in the Lake Pontchartrain Basin? Our preliminary data indicates that this is the case.

We hypothesized that salinity is an environmental stressor to the freshwater turtle. Our preliminary data indicate that this is the case. As such, salinity triggers a neuroendocrine stress response to mediate the adaptation of this vertebrate to a challenging environment. Indeed, our preliminary data indicates that turtles exposed to salinities above normal physiological salinity (0.9% or 9%) tend to respond by not producing corticosterone, the stress hormone. This blunted response may be due to the Na+ retaining effect of corticosterone in vertebrates. It would be detrimental for a freshwater vertebrate to retain Na+ in the face of already elevated environmental Na+ concentration.

Because salinity is stressful to the turtle, salinity is expected to impact the abundance and distribution of the turtle in the basin, as well as that of other freshwater vertebrates. We have indeed observed that there are many more turtles in western Lake Pontchartrain basin than in the Slidell area, judging by the preliminary index of capture that we have generated.

The hypotheses of this study have predictive and, as such, management value. For instance, the study predicts that the sustained decreases or increases of salinity in the basin will impact the physiology of freshwater vertebrates. This finding is particularly relevant to water diversion projects as they may need short-term biological indicators to evaluate the effect of increasing freshwater input in the basin. Also, it may help environmental biologists assess the

54 PBRP Annual Report | 2009 Salinity as a Stressor of the Freshwater Turtle

damage to the basin’s biota by sustained saltwater intrusion as exacerbated by land loss driven by subsidence (long-term) and seasonal storms (short-term).

The scope of this study is expected to be of interest to local, state, and federal agencies in charge of protecting the environment such as Louisiana Department of Environmental Quality, the Louisiana Department of Wildlife and Fisheries, the Coastal Restoration and Management Division of the Louisiana Department of Natural Resources, the US Geological Survey, and the Environmental Protection Agency. The results of the study will be important to all parishes found in southeastern and coastal Louisiana that are affected by saltwater intrusion such as New Orleans, St. Tammany, Tangipahoa, St. Bernard, and Jefferson. Because this study focuses on how salinity affects a species of high economic importance to Louisiana, turtle farmers will benefit the most from the results. However, because the study considers the physiological mechanistic impact of saltwater intrusion on the turtle, the results are relevant to the entire basin as the abundance and distribution of many other freshwater vertebrates may be limited by similar mechanisms.

The value of the study’s results can be enhanced significantly if repeated after the Mississippi River diversion projects begin. Our study establishes a baseline for vertebrates in the basin sensitive to salt water. If this study is repeated after diversions begin, then we can trace how the diversions impact these vertebrates. In essence, our hypothesis predicts that diverting Mississippi River waters into the Lake Pontchartrain Basin will induce a significant, sustained decrease in salinity throughout the basin. This decrease, in turn, will allow the proliferation of species sensitive to salt water, those which will benefit from salt water intrusion. Specifically, the results of this project may allow managers to determine the optimal amount of freshwater that may be diverted into Lake Maurepas to provide a suitable living environment for freshwater species. Because we expect that high salinity will increase corticosterone, the main stress hormone, in turtles, future research should determine exactly how increasing or decreasing salinity levels affect these turtles.

PBRP Annual Report | 2009 55 Genetic Variation between Lake Pontchartrain and Mississippi River Basin Fishes

Genetic Variation between Lake Pontchartrain and Mississippi River Basin Fishes

Kyle R. Piller and Lisa M. Cordes Southeastern Louisiana University; Department of Biological Sciences

ABSTRACT The Lake Pontchartrain Basin harbors a distinctive and ever changing fish community that will likely change in the near future due to the construction of an interbasin canal that will connect the Mississippi River and Lake Pontchartrain Basins. Although the influx of water is aimed at restoring the wetlands around Lake Maurepas by providing freshwater to the system, it also may negatively impact the ichthyofauna of basin through homogenization of genetically distinctive stocks of fishes. The objective of this project is to examine genetic differentiation between Lake Maurepas and Mississippi River Basin fish populations of bluegill (Lepomis macrochirus) using microsatellite markers.

OVERVIEW & OBJECTIVES Historically, the prevailing view in fisheries science was that conspecific populations, stocks, and/or strains could be freely transferred between lakes, rivers, and even drainage basins without detriment to the natives. However, this was most often conducted without examining whether there were distinctive morphological or molecular differences among populations. Recent studies that have incorporated molecular data have shown that species often are composed of genetically distinct stocks that have survived and evolved in isolation from other conspecific stocks and maintain unique features, life-histories, and physiological adaptations. Therefore, from a management perspective, it is critical to identify unique populations prior to stocking or environmental manipulation. Unfortunately, these types of studies have rarely been conducted a priori.

Conceptually, the idea of what constitutes a population seems simple and straightforward, however, empirically this is not the case. Waples and Gaggiotti (2006) summarized commonly used definitions of a “population” under ecological, evolutionary, and statistical paradigms and concluded that none of the working definitions were truly operational. Historically, the criteria that have been used to define a “population” simply have been based on 1) locality such as drainage basin, and/or 2) slight differences in morphology, and/or distinctive aspects of life-history. In the fisheries literature, “stocks” and “strains” have routinely been used in place of “population.” Ihssen et al. (1981) defined a stock as an intraspecific group of randomly mating individuals with temporal or spatial integrity. Phillip et al. (1993) expanded on this definition and suggested that a stock is defined as a genetically distinct group of fish unique to a particular body of water or basin that has spatial, temporal, and behavioral integrity from other randomly breeding groups of the same species. No matter what definitions are used to identify unique biological units, it is important to have sound operational methodologies to allow researchers to address and better diagnosis

56 PBRP Annual Report | 2009 Genetic Variation between Lake Pontchartrain and Mississippi River Basin Fishes

distinct populations, stocks, or strains. Molecular techniques offer the most promising aspect to address these types of issues.

Disruption of distinct stocks either through intentional or unintentional avenues can have detrimental effects on the genetic integrity of native fish populations through decreased levels of fitness and lower survivability in offspring derived by hybridized stocks (Allendorf 1991, Epifanio et al. 2001, Hindar et al. 1991, Krueger and May 1991, Launey et al. 2006). Ecological and genetic effects of fish stocking, in theory, may cause native species to 1) be eliminated 2) have changes in growth and survival, 3) be genetically changed 4) alter community structure, 5) exhibit a combination of the above, or 6) exhibit no detectable changes (Moyle 1986, Krueger and May 1991).

The primary objective of this study is to address genetic differentiation between Lake Maurepas and Mississippi River Basin fish populations prior to construction of the Hope Canal Diversion. The examination of multiple genetic markers from bluegill (Lepmois macroshirus) will assess whether Lake Maurepas and Mississippi River Basin bluegill populations are distinctive from one another prior to mixing of these stocks. Since fishes comprise a major component of this system, collection of a baseline data set on the genetic variability and distinctiveness of these populations is critical to the restoration and long-term sustainability of this resource.

RESULTS & DISCUSSION Data from 207 individual bluegills (Lepomis macrochirus) were collected from five locations in the Lake Pontchartrain and Mississippi River basins (tab. 1).

Table 1. Sampling localities, abbreviations, and number of specimens collected Site Abbreviation GPS Basin Species N Pass Manchac PM 30.265N 90.399W Maurepas L. macrochirus 40 Carthage Bluff CB 30.308N 90.588W Maurepas L. macrochirus 48 Blind River BR 30.212N 90.594W Maurepas L. macrochirus 31 Devil's Swamp DS 30.317N 90.423W Mississippi L. macrochirus 28 Atchafalaya AtR 30.513N 91.719W Mississippi L. macrochirus 60

Two multiplex reactions, three loci in each, were optimized using previously published primer sets (fig. 1) (Colburne et al. 1996, Neff et al. 1999). Data from these six loci (Lma20, 117, 121, 102, 87, and 21) were collected from 207 individuals.

The results suggest that the loci chosen in this study are highly variable and do provide informative information regarding gene flow within and among both drainage basins. Across the six loci, the number of alleles per locus ranges from 20-36 (tab. 2). Locus Lma20 is the most variable locus, with 36 alleles, whereas Lma87 is the least variable with 20 alleles recovered from the 207 individuals that were genotyped.

PBRP Annual Report | 2009 57 Genetic Variation between Lake Pontchartrain and Mississippi River Basin Fishes

Lma102

Lma87

Lma21

Figure 1. Bluegill multiplex showing three microsatellite loci (Lma102, Lma87, and Lma21) each labeled with a different fluorescent dye

Table 2. Summary of genetic variation within Lepomis macrochirus by locus

Locus Total # alleles HE HO Allele size range LMA102 27 0.864 0.817 77-141 LMA87 20 0.823 0.734 108-170 LMA21 35 0.919 0.908 144-236 LMA20 36 0.932 0.938 67-147 LMA117 27 0.874 0.914 127-189 LMA121 21 0.887 0.945 138-200 LMAR10 34 0.938 1 168-344 Mean 28.57 0.891 0.894

Genotypic data has been gathered from five populations. The Devil’s Swamp population (Mississippi River Basin) is the most variable, averaging 21.29 alleles per locus. The Blind River (Lake Pontchartrain Basin) is the least variable, averaging only 15.71 alleles per locus (tab. 3). The levels of variability for these microsatellite loci are much higher compared to the levels of variability observed among blue catfish from similar locations.

This study focused on determining the genetic structure of bluegill, Lepomis macrochirus, within and among the Lake Pontchartrain and Mississippi River Basins, in an effort to understand the potential genetic impacts of an impending freshwater diversion. This study is somewhat novel because it examined genetic structure of these fishes prior to construction of the freshwater diversion.

58 PBRP Annual Report | 2009 Genetic Variation between Lake Pontchartrain and Mississippi River Basin Fishes

Table 3. Summary of genetic variation by population

Site Total # alleles NA HE HO PM 124 17.71 0.9082 0.9036 CB 130 18.57 0.9045 0.9077 BR 115 15.71 0.8668 0.8596 DS 110 21.29 0.8777 0.869 AtR 149 16.43 0.8979 0.9286 L. macrochirus mean 125.6 17.94 0.891 0.8937

The results from multiple analyses suggest there is no structuring within or among the Lake Pontchartrain and Mississippi River basins, resulting in limited genetic differentiation of bluegill and a high degree of gene flow. Most of the statistical analyses (AMOVA in tab. 4, Heterozygote Excess in tab. 5, FCA in fig. 2) converged on similar results, with the exception of the pairwise FSTstatistics (tab. 6). Although FST values were low, after a Bonferroni type correction for multiple tests, these values suggested significant genetic differentiation between some population pairs of bluegill. Bluegill from the Carthage Bluff sampling location were not genetically differentiated from the other two sample locations within the Lake Pontchartrain Basin. The Devil’s Swamp and Atchafalaya River “populations” of bluegill within the Mississippi River Basin also were not genetically distinct from each other.

After further statistical analyses congruently suggested a lack of genetic structure, it is likely that the significance of the FST values is inflated. Hedrick (1999) showed that highly polymorphic loci, such as microsatellites, can confound some statistical analyses such as pairwise differentiation tests (FST). He suggested that the statistical power provided by multiple microsatellite loci may result in statistical significance, when in fact there is no biological significance (Hedrick 1999). FST values have a strong correlation with heterozygosity; it has been shown that lower heterozygosities (<0.1) decrease the median value of the FST (Beaumont and Nichols 1996). This study exhibited high levels of heterozygosities (ranging from 0.412-0.938), which would not decrease the efficiency of FST statistical analyses. Low levels of FST values are consistent with low genetic differentiation and high levels of gene flow. Concordance of all other analyses conducted in this study suggests that the low, by statistically significant FST values, is not the result of biological phenomena.

PBRP Annual Report | 2009 59 Genetic Variation between Lake Pontchartrain and Mississippi River Basin Fishes

Table 4: Analysis of molecular variance (AMOVA) in bluegill populations in the Lake Pontchartrain and Mississippi River Basins Sum of Variance Percentage of Source of variation d.f. squares components variation Among Basins 1 12.472 0.04164 1.32 Among pops within basins 3 11.528 0.00960 0.30 Within populations 409 1266.677 3.09701 98.37 Total 413 1290.676 3.14825

Table 5: Hardy Weinberg exact tests of heterozygote excess

Species P-value S.E. Lepomis macrochirus 0.8334 0.0257 *Exact P-values by the Markov chain method

Figure 2: Factorial Correspondence analysis of bluegill showing a lack of differentiation based on multilocus genotypes. Grey and white symbols represent Mississippi River Basin populations, and blue, yellow, and purple represent Lake Pontchartrain Basin populations.

60 PBRP Annual Report | 2009 Genetic Variation between Lake Pontchartrain and Mississippi River Basin Fishes

Table 6. Pairwise FST values bluegill in the Lake Pontchartrain and Mississippi River Basins PM CB BL AR DS PM - CB 0.0007 - BL 0.0098 0.0029 - AR 0.0159 0.0186 0.017 - DS 0.0169 0.0149 0.0133 0.0026 * All Fst values not significant following Bonferroni correction are highlighted in bold

The bluegill populations within the Lake Pontchartrain and Mississippi River Basins are not spatially isolated. There was a lack of significant correlation of genetic distance (FST) and geographic distance (km) between populations bluegill (r = -0.4336, p≤0.989) (fig. 3). High levels genetic variation within individual populations corroborates the lack of genetic structure, as recovered in the other analyses.

Figure 3. Pairwise comparisons of genetic distance versus geographic distance of bluegill in the Lake Pontchartrain and Mississippi River Basins. (* Z = -42.8677, r = -0.4336, one-sided p <= 0.9890 from 1000 randomizations)

One possible reason for low levels of population differentiation may be low levels of variability within the markers studied. However, high number of alleles per locus, along with high levels of heterozygosity suggests high variability within the markers examined in this

PBRP Annual Report | 2009 61 Genetic Variation between Lake Pontchartrain and Mississippi River Basin Fishes

study. Amos and Harwood (1998) suggest that high levels of genetic variability are healthy and enable species to cope with threats such as disease, predation, and environmental change. The high levels of variability recovered in this study that would detect population structure if any were present.

High levels of gene flow have been shown to limit the amount of genetic differentiation among populations. One artificial factor contributing to the high levels of gene flow is the Bonnet Carré Spillway, which serves as a corridor for dispersal of fishes between the Mississippi River and Lake Pontchartrain basins. Historical connections of Lake Maurepas and the Mississippi River, and multiple levee breeches in the 1800s may have facilitated gene flow and inhibiting differentiation between basins.

In summary, multiple genetic analyses suggest that gene flow among populations within the basins is high. The lack of differentiation between bluegill populations in the Lake Pontchartrain and Mississippi River basins suggests that construction of the Hope Canal Diversion will likely have little genetic impacts (i.e., reduction of fitness) on bluegill populations. However, future studies examining genetic structure of other species and experimental crosses of population examining fitness differences of the cross would be informative.

TECHNOLOGY TRANSFER Freshwater diversions are routinely being offered as a valuable tool for wetland restoration, however, the potential genetic impacts that this type of restoration project has on native species has not been thoroughly investigated. This is particularly the case in Louisiana, which has many on-going or proposed freshwater diversion projects. This present study focuses on the evaluation of this restoration technique and is aimed at assessing the potential impacts that it has on genetic diversity and biodiversity. Not only will quantitative baseline genetic data be beneficial for this particular diversion project, but more importantly, this data will serve as a model study for any future wetland restoration diversion projects that are aimed at preserving biodiversity and genetic integrity while restoring coastal and wetland habitats. The data from this EPA project (bluegill), and the PI’s previous recently completed EPA project on blue catfish also should provide a comprehensive overview of the impacts of life- history on genetic differentiation for two species with varying fecundities, migratory abilities, and sensitivities to fishing pressure. This data is critical and will be useful to policy makers including state and federal agencies, including the U.S. Army Corps of Engineers and the Louisiana Department of Wildlife and Fisheries, because these two agencies are the principal players in wetland restoration.

REFERENCES Allendorf, F.W. 1991. Ecological and genetic effects of fish introductions: synthesis and recommendations. Canadian Journal of Fisheries and Aquatic Sciences 48:178-181.

Amos, W. and J. Harwood. 1998. Factors affecting levels of genetic diversity in natural populations. Philosophical Transactions of the Royal Society of London, B 353:177-186.

62 PBRP Annual Report | 2009 Genetic Variation between Lake Pontchartrain and Mississippi River Basin Fishes

Beaumont, M.A., and R.A. Nichols. 1996. Evaluating loci for use in the genetic analysis of population structure. Proceedings of the Royal Society B: Biological Sciences 263:1619-1626.

Colbourne, J.K., B.D. Neff, J.M. Wright, and M.R. Gross. 1996. DNA fingerprinting of bluegill sunfish (Lepomis macrochirus) using (GT)n microsatellites and its potential for assessment of mating success. Canadian Journal of Fisheries and Aquatic Sciences 53:342- 349.

Epifanio, J., F. Utter, and D. Philipp. 2001. The effects of fish stocking on genetic level biodiversity. Pages 23-64. in J. Nalbone and E.L. Michael, editors. Taking stock of our future. Great Lakes United Conference Proceedings. Great Lakes United, Buffalo, NY, USA.

Hedrick, P.W. 1999. Perspective: Highly variable loci and their interpretation in evolution and conservation. Evolution 53:313-318.

Hindar, K., N. Ryman, and F. Utter. 1991. Genetic effects of cultured fish on natural fish populations. Canadian Journal of Fisheries and Aquatic Sciences 48:945–957. Ihssen, P.E., H.E. Booke, J.M. Casselman, J.M. McGlade, N.R. Payne, and F.M. Utter. 1981. Stock identification: materials and methods. Canadian Journal of Fisheries and Aquatic Sciences 38:1838-1855.

Krueger, C.C., and B. May. 1991. Ecological and genetic effects of salmonid introductions in North America. Canadian Journal of Fisheries and Aquatic Sciences 48:66–77.

Launey, S., J. Morin, S. Minery, and J. Laroche. 2006. Microsatellite genetic variation reveals extensive introgression between wild and introduced stocks, and a new evolutionary unit in French pike Esox lucius L. Journal of Fish Biology 68:193-216.

Moyle, P.B. 1986. Fish introductions into North America: patterns and ecological impact. in H. Mooney and J.A. Drake, editors. Biological invasions in North America. Springer-Verlag, New York, NY, USA.

Neff, B.D., P. Fu, and M.R. Gross. 1999. Microsatellite evolution in sunfish (Centrarchidae). Canadian Journal of Fisheries and Aquatic Sciences 56:1198-1205.

Philipp, D.P., J.E. Epifanio, and M.J. Jennings. 1993. Conservation genetics and current stocking practices: Are they compatible? Fisheries 18:14-16.

Waples, R.S. and O. Gaggiotti. 2006. What is a population? An empirical evaluation of some genetic methods for identifying the number of gene pools and their degree of connectivity. Molecular Ecology 15:1419-1439.

PBRP Annual Report | 2009 63 Lake Pontchartrain Basin Research Program: Education Outreach

Western Lake Pontchartrain Basin Research Program Education Outreach Component: Phase 4

Deborah Dardis Department of Biological Sciences; Southeastern Louisiana University

ABSTRACT Phase 4 of the educational outreach focused on providing three interdisciplinary workshops with classroom and field educational experiences to K-12 teachers and community members. Thirty-two people participated in activities that were designed to introduce the ecology of the basin and southeast Louisiana wetlands, to discuss the current research on habitat restoration and sustainability in the Manchac area, and to emphasize the connection between the region’s ecology and its cultural and economic vitality. A written survey using a 4-point Likert scale was administered at the end of each workshop to measure participant satisfaction. The average score for the overall workshop experience was 3.7, proving the workshops successful.

PRIMARY OBJECTIVE The primary objective of this project was to increase public awareness of the deterioration of southeast Louisiana’s wetlands; the contribution of human activity to this deterioration; the economic, cultural, and social ramifications of the ecosystem’s demise; and the current research performed by local universities on habitat restoration and sustainability.

METHODS & ACTIVITIES To reach the primary objective of increasing public awareness, four workshops were scheduled for K-12 teachers and members of the community; however, after a large number of teachers withdrew for various reasons, we combined two scheduled workshops, offering a total of three throughout the summer months of 2008 (June 19, June 27, July 11). Teachers from surrounding parishes were invited to participate and to bring a ‘visitor’ from the community. Twelve middle-school teachers (from Orleans, St. Tammany, and Jefferson parishes) and approximately 20 community visitors (from various parishes including Tangipahoa, St. Tammany, and East Baton Rouge parishes) participated in the workshops.

Each workshop consisted of a full day, 8:00AM– 4:00PM, of interdisciplinary instruction and activities that included classroom lectures and canoeing and boating explorations of different types of wetlands. After a brief overview of the day and information about the Lake Pontchartrain area, participants canoed through a mixed wetlands forest and a marsh. Then, they returned to the classroom at Galva Canal to debrief.

The next block of activities involved presentations about the history and purpose of the wetlands in southeast Louisiana. After these presentations, participants completed a watershed and estuary handout with detailed information about the rivers, lakes, bayous, and

64 PBRP Annual Report | 2009 Lake Pontchartrain Basin Research Program: Education Outreach

land masses in southeast Louisiana, which is all information they can translate to student projects and lectures in their own classrooms. The presentations and activities were led by me and Michael Greene, the station biologist for the Turtle Cove Environmental Research Station. Specifically, participants learned the definition of a wetland and its purpose related to the health of the Pontchartrain Basin. They also learned how to identify several plant and animal species along with the differences in salinity of the region and the differences in the ecosystems of a marsh versus a swamp. The final activity was a pontoon boat exploration of Lake Pontchartrain and Pass Manchac to identify the species discussed. Michael Greene led this field trip and provided additional information about ways to restore the wetlands.

RESULTS The workshops were a success. All involved found the activities exciting and instructional. It was the first time in a canoe or boat for several of the participants, which made for quite memorable experiences. Evaluations included a written attitudinal survey of the participants, which consisted three questions using a Likert Scale rating system of Poor (1) – Fair (2) – Good (3) – Great (4). The average scores were as follows: 3.7 for the overall workshop experience, 3.6 for expertise of faculty and staff, and 3.7 for the wide variety of learning experiences offered.

Participants also gave the following responses as “life lessons learned” throughout the day: “Learning how important the wetlands are and their functions” Amazement when learning the projected rate of wetland loss “It is important for citizens to get involved in saving wetlands” “Didn’t know how beautiful the wetlands forests were” “Came afraid to get into a canoe or boat and ‘be in water’ – left an empowered citizen less afraid of water and boating and knowing much more about the importance of the wetlands” The combination of lessons learned and high Likert scores reflect the positive impact of the material and experiences provided during the workshops.

TECHNOLOGY TRANSFER This project aimed to increase public awareness of the deterioration of southeast Louisiana’s wetlands; the contribution of human activity to this deterioration; the economic, cultural, and social ramifications of the ecosystem’s demise; and the current research performed by local universities on habitat restoration and sustainability. We assume that the knowledge teachers acquired during this project will be continually transferred to students in the classroom. Each teacher may potentially impact thousands of students, thus helping to create a future citizenry that is well educated about the importance of wetlands.

In the truest sense, this is technology transfer that will reach thousands of our future citizens by educating teachers, who will educate students. The community was also directly impacted through the 20 community participants, so they can inform other stakeholders in the region, especially those stakeholders who may not have ties to the educational system.

PBRP Annual Report | 2009 65 Lake Pontchartrain Basin Research Program: Education Outreach

Although this project did not communicate directly with current policymakers, it did educate teachers who educate future policymakers, current students. We can also assume that the teachers and community members who participated in the workshops will be more likely to advocate for wetland restoration, thereby holding governmental agencies and policymakers more accountable for their actions and potentially influencing future policy decisions and elections.

66 PBRP Annual Report | 2009 De-energizing Hurricanes with Cypress/Tupelo Buffers

De-energizing Hurricanes with Cypress/Tupelo Buffers: A Plan to Restore the Repressed Swamps of the Lake Pontchartrain Basin by Using Point and Nonpoint Freshwater Sources

Gary P. Shaffer and William Bernard Wood Department of Biological Sciences; Southeastern Louisiana University

ABSTRACT This study is comprised of four objectives: 1) following the production and compositional response of the herbaceous and woody vegetation of the Maurepas swamp to the hurricanes, 2) determining the relationship between basal area of canopy trees and frequencies of windthrow, 3) isolating all substantial point and non-point sources of potential freshwater input in the basin and then building a geographic information system (GIS) of these sources and the spatial extent of currently repressed swamp that could be restored by using these sources, and 4) modeling how the area of swamp or marsh reduces the energy of storms of varying strength. Using point and nonpoint sources of fresh water to restore swamp will serve the dual benefits of improved water quality, through denitrification and assimilation, and improved wetland health. Our monitoring efforts in the Maurepas swamp have clearly identified a relationship between basal area of cypress and tupelo canopy tree and windthrow of midstory maple and ash. We also document a clear trajectory from swamp to marsh and open water over the 9-year study and show that the 2005 and 2008 hurricanes improved marsh production by detrimentally affecting tree production. Thanks to such events in our data set, we have been able to isolate specific patterns in herbaceous vegetative cover associated with tropical storms. Modeling the storm surge reduction capacity of baldcypress- water tupelo swamps will be conducted throughout the remainder of our project.

INTRODUCTION Baldcypress-water tupelo (Taxodium distichum and Nyssa aquatica, respectively) swamps historically comprised 90% of the wetlands in the Lake Pontchartrain Basin (Saucier 1963). This coverage has been radically reduced, and the swamps that remain have been classified as mostly non-sustainable (Chambers et al. 2005, Shaffer and Day 2007, Shaffer et al. 2007). Several projects have been proposed to help restore these swamps, but Hurricanes Katrina and Rita in 2005 and Gustav and Ike in 2008 have altered the way we should think about wetland restoration in general. In terms of flood- and wind-damage reduction, swamps may be far superior to other wetland habitat types. If so, we need to re-think coastal restoration strategies, tailoring them to storm-protection alternatives.

Only live oak (Quercus virginica) and palms are more resistant to windthrow (i.e., blow down) than baldcypress and water tupelo (Williams et al. 1999). Cypress-tupelo swamps faired far better than other forest types in Hurricanes Camille (Touliatos and Roth 1971), Andrew (Doyle et al. 1995), and Hugo (Gresham et al. 1991, Putz and Sharitz 1991). In addition,

PBRP Annual Report | 2009 67 De-energizing Hurricanes with Cypress/Tupelo Buffers fresh, intermediate, and brackish marshes suffered vastly greater loss in Hurricanes Katrina, Rita, Gustav, and Ike than did cypress-tupelo swamps (Barris 2006, 2008). Katrina caused windthrows of up to 80% of the bottomland hardwood forests of the Pearl River Basin (320 million trees down) while contiguous swamps remained largely intact (Chambers et al. 2007).

Initial observations in the Maurepas swamp indicate minimal windthrow of overstory trees, but light to moderate crown damage. In contrast, midstory windthrow was severe in certain areas and appears to be a function of overstory density. This study aims to quantify the relationship between windthrow and overstory tree density, which could be useful in determining optimal densities of artificial plantings. We also monitor post-hurricane tree and herbaceous ground cover production and compare it with that of normal- and drought-year conditions using Primer.

Although baldcypress-water tupelo swamps are extremely resistant to windthrow and deep flooding, they are less resistant to saltwater intrusion, thus requiring a reliable source of fresh water for system flushing following droughts and tropical storm events. We have built a geographic information system (GIS) that contains point and non-point freshwater sources on the northshore from Baton Rouge to the Pearl River. These freshwater sources are currently being prioritized using proximity to repressed swamp and potential area of benefit. The analysis focuses on wastewater treatment facilities and potential Mississippi River diversion sites, as these sources show the highest potential benefits (Shaffer and Day 2007). At present, most of these sources are input to the basin to maximize drainage efficiency.

PROJECT OBJECTIVES The original objectives of this long-term research project included the following: 1. Compare post-hurricane production and compositional response of the herbaceous and woody vegetation of the Maurepas swamp to that of “normal” and drought years 2. Determine the relationship between basal area of canopy trees and frequencies of windthrow of midstory trees 3. Isolate all substantial point and non-point sources of potential freshwater input to the degraded wetlands of the northern Pontchartrain Basin and then build a geographic information system (GIS) of these sources and the spatial extent of currently repressed swamp that could be restored by using these sources 4. Model how the area of swamp or marsh reduces the energy of storms of varying strength

Objectives 1 and 4 are ongoing, but the research activities and analyses for objectives 2 and 3 are complete and were discussed in the previous annual report. This report will briefly summarize the results to date and explain the activities and analyses that are ongoing.

METHODOLOGY This study is one of the largest and most comprehensive ecological studies ever conducted with data collection and analyses spanning almost ten years. The first phase of our data

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collection involved assessing the effects of crown damage and windthrow through measurements of wood and litter production. We also have a nine-year record of herbaceous composition and production to compare pre- and post-hurricane measurements. In addition, we have related basal area of overstory trees to windthrow of midstory trees by assessing our 52 permanent stations (625m2), which vary from sparse to dense canopy cover.

Most of our GIS data were obtained from LADEQ, focusing on small to large wastewater treatment facilities. These data were augmented with data from proposals to divert Mississippi River water at Violet, Bonnet Carré, La Branche, Romeville, Hope Canal, and Blind River. The 2007 annual report explains, in detail, how we configured the raster and vector data for the GIS. Now that the GIS is built, this year we will estimate the extent of repressed swamp that could be converted back into healthy swamp under various management scenarios.

We are still sorting and weighing 2008 leaf litter, which does not fall from the trees fully until February 2009. The diameter data on over 2,000 trees for 2007 and 2008 is in the computer, but a SAS program must be written and ran to convert that data into wood production. The leaf litter and wood production data will then be entered into SYSTAT, so they may be analyzed together. In addition to this new data and analyses performed since the 2007 report, herbaceous cover value (species composition) and clip plot (production) data for 2007 and 2008 have been entered, and we are currently conducting multivariate ordination on the cover values. Finally, we will run statistical analyses on the production data to compare it with the tree production data. We aim to have all the data analyzed by June 2009.

Our modeling efforts to quantify the wind- and flood-damage reduction capacities of cypress-tupelo swamps have begun using the parameters derived for mangrove forests subjected to tsunamis (Massel et al. 1999, Yoshihiro et al. 1997). Research on tsunamis and mangroves is applicable for estimates of storm-surge and wave-energy reduction, but obviously not wind reduction. During October, 2007, we measured stem density, basal area, and species cover of several plots contiguous with levees in St. Bernard Parish to obtain baseline data on known surge reduction. We will compare the 2008 and 2009 data with these baselines to better quantify the reduction capacities.

In general, land reduces the energy of hurricanes in three ways: detachment of the wind from the water, offering friction, and most importantly for forests that remain intact, damping surge by deflecting it in many directions, which decreases onshore movement while simultaneously de-energizing the storm. Land, in general, will detach the wind from the water. However, forested wetlands offer orders of magnitude more friction and dampening power to decrease wind velocity and storm surge, a characteristic true of those that are resistant to windthrow such as baldcypress-water tupelo swamps. To model energy reduction, we need to estimate the volume of the vegetation, its extent, and its Manning and/or Collins coefficient (Massel et al. 1999, Yoshihiro et al. 1997). Basically, the momentum equation simplifies to a balance between the slope of the water surface and the drag and dampening force offered by the land. It also depends on the spectral characteristics unique to each storm, so we will simulate a number of hurricane conditions using models housed at the Hurricane Center of LSU.

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RESULTS TO DATE Environmental Variables: The analysis of well-water salinity data revealed that salinity

followed a U-shaped pattern from 2000-2006 (quadratic contrast F1,259 = 406.16, p < 0.00001; fig. 1) with the highest salinities occurring during the severe drought of 1998-2000 followed by 2006, another drought year (2007 and 2008 also were drought years). Overall, salinity was highest at Degraded sites and lowest at Throughput sites (linear contrast F1,259 = 168.92, p < 0.00001). Soil salinity also was found to decrease with increasing distance from Pass Manchac, as well as with increasing distance from the margin of Lake Maurepas into the interior swamp. As soon as 2008 litterfall collection is complete (anticipated March 2009), we will begin the next large round of data analysis and synthesis.

Figure 1

Tree Mortality: The Maurepas swamp is in a steady state of rapid decline, perhaps best shown by the mortality of canopy and midstory trees. During the last seven years, nearly 20% of the original 1,860 trees in our study plots have suffered mortality, and recruitment of baldcypress and water tupelo saplings is essentially absent. In 2000, almost all of the mortality occurred at the Degraded sites, but the highest rates are now experienced in the Relict sites, largely because there are very few trees left to die at Degraded sites. The few remaining trees at Degraded site are nearly all baldcypress, which is more tolerant to saltwater intrusion. Mortality is highest for midstory species (fig. 2), nearly all of which are swamp red maple and green ash.

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Figure 2

Herbaceous Vegetation: For annual production, herbaceous production was highest at the Degraded sites (mean = 697.33 ± s.e. 71.84 g m-2 y-1) followed by Relict sites (mean = 381.60 -2 -1 -2 -1 ± s.e. 26.11 g m y ), and lowest for Throughput sites (64.31 ± s.e. 13.64 g m y ; F2,256 = 3.26, p = 0.040; fig. 3). Higher production in recent years is partially attributable to a shift to more salt tolerant herbaceous species and to decreased competition with tree species with continued high mortality rates, especially at the Degraded sites. We will see if this trend continues for our 2007 and 2008 data.

Most of the herbaceous biomass production could be attributed to 15 dominant ground- cover species, representing 97% of the total herbaceous cover throughout the study. We have isolated a strong post-hurricane signal (2003, 2006, 2009) of decreased perennial coverage and increased cover of annuals, especially smartweed (Polygonum punctatum).

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Figure 3

Primary Production of Trees: Total tree primary production differed between habitat types (F2,126 = 14.126, p < 0.00001; fig. 4) and years (F6,126 = 9.997, p = 0.00001; fig. 3). Bulk density and salinity were significant covariables in the model (F1,256 = 12.940, p = =0.00039, F1,256 = 2.058, p = 0.020, respectively), indicating that tree primary production was higher at sites with higher bulk densities and lower salinities. The highest rates of total tree primary production were found at the Throughput sites (mean = 737.03 ± s.e. 37.18 g⋅m-2⋅yr-1), followed by Relict sites (mean = 322.48 ± s.e. 13.88 g⋅m-2⋅yr-1), followed by Degraded sites (mean = 144.36 ± s.e. 37.18 g⋅m-2⋅yr-1). In general, leaf litter and wood production followed

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similar patterns through time (fig. 3) with the lowest production overall occurring in 2003. Interestingly, a general increase in production was experienced by the herbaceous community in 2003, but the forest community continued to decline until 2004.

Figure 4

For total annual (leaf plus wood) tree production, an interaction existed between habitat type and the three categories of species (F = 17.34, p < 0.00001; fig. 4). The interaction occurred for Relict sites where baldcypress had similar growth rates as water tupelo and ‘Other’. Although baldcypress was the least abundant species of the three categories at almost all sites (Hoeppner 2002), it had nearly twice the average growth rate (mean = 171.83 ± s.e. 8.33 g m- 2 year-1) as water tupelo (96.46± s.e. 5.70 g m-2 year-1) or ‘Other’ (mean = 91.10 ± s.e. 5.53 g m-2 year-1).

Total Primary Production: In only seven years, the Maurepas swamp has switched from an ecosystem dominated by tree production to one dominated by herbaceous production (fig. 5). From 2000-2003, overall production was similar, with a decrease in tree production compensated for by an increase in ground cover production. Overall production from 2004- 2006 was significantly greater than that of 2000-2003, yet tree production continued to fluctuate around 400 g m-2 year-1. The 2007 and 2008 data will help us better define the rates.

Degraded sites became completely dominated by herbaceous vegetation from 2000-2006, Relict sites late in the study transitioned toward Degraded sites early in the study, and Throughtput sites transitioned toward Relict sites following the severe drought (i.e., 2002), and late in the study. Analysis and synthesis of 2007-2008 data will clarify these trends.

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Figure 5

Potential Surge Reduction by Cypress Swamps: Marshes can slow down surge by storing water in the early stage of the storm, and large trees can obstruct the surge and wave propagations, thereby reducing dynamic loads on the infrastructures. Field data after Hurricanes Katrina and Rita showed that hurricane surge and wave energy could be reduced by non-structural measures such as coastal restoration and wetland creation. We are currently analyzing whether the field data after Hurricane Gustav follows these trends.

Most storm surge estimates are based on 2D models. Generally, a 2D model uses Manning’s friction coefficient to represent wetlands in the model. For large vegetation or trees (such as baldcypress-water tupelo swamps), 2D representations under-predict the surge propagation by not including the physical presence of trees in the models. In this research, we are investigating model resolutions and node elevations to better represent trees in the model and to create a flow around trees. Initial results suggest that raising nodes can mimic trees that are not totally inundated and can obstruct flows around them, thereby reducing flow velocity. At this time, we are working to improve model resolution and other available techniques to better estimate surge reduction by cypress swamps and to validate numerical results with field data.

DISCUSSION In summary, the Maurepas swamp is characterized by nutrient-poor waters, soils of extremely low strength, nearly permanent flooding in most areas, and saltwater intrusions that generally occur during the late summer and fall seasons. The Maurepas swamp is nitrogen limited, and nutrient stress is potentially as important as salt or flood stress. Furthermore, recruitment of baldcypress and water tupelo saplings throughout the swamp is very low, certainly not sufficient to sustain the aerial extent of current forest. Most of the Maurepas swamp appears to be converting to marsh and open water, primarily due to the

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lack of riverine input. Salt stress is killing trees proximal to the lake while stagnant water and nutrient deprivation seem the largest stressors at interior sites.

Although baldcypress-water tupelo swamps are extremely resistant to windthrow and deep flooding, they are less resistant to saltwater intrusion and thus require a reliable source of freshwater for system flushing following tropical storm events and during droughts. We are in the process of building a geographic information system that contains all substantial point and non-point freshwater sources, including municipal wastewater treatment facilities and potential Mississippi River diversion sites. At present, most of these sources are input to the basin to maximize drainage efficiency. Fresh water is routed into ditches and canals that carry it directly to the lakes, bypassing wetland contact. This routing creates a “lose-lose” situation as the potential for eutrophication is maximized and the wetlands remain nutrient starved (Shaffer and Day 2007). In contrast, re-routing the water to maximize sheet flow would improve water quality, increase wetland net primary production, and decrease saltwater intrusion. Furthermore, implementing the proposed Mississippi River re- introductions at Violet, Bonnet Carré, La Branche, and the Maurepas swamp (Coast 2050 1998) will greatly enhance restoration of historic salinity regimes. In addition to these benefits, increasing swamp acreage will decrease storm damage, may lead to net sediment accretion, will increase carbon sequestration (Trettin and Jorgensen 2003), will enhance biodiversity, and will improve several of the “multiple lines of defense” proposed by Lopez (2006). Ultimately, closing the Mississippi River Gulf Outlet, which is underway, will help approximate historic salinity regimes and reduce storm damage (Shaffer et al. In Review).

One concern that managers and the general public have with restoring repressed swamps is the amount of time required for swamp-like characteristics to emerge and manifest. Fortunately, given favorable hydrologic and nutrient conditions, baldcypress and water tupelo seedlings can reach greater than 10-meter heights within one decade. For example, a pilot planting of baldcypress seedlings at the Caernarvon diversion (Krauss et al. 2000) has yielded >10 m tall trees in a decade, and all of these resisted windthrow during the hurricanes of 2005 and 2008. In conclusion, if we are to reverse the trajectory of decline of coastal Louisiana swamps, we must find, and wisely use, point and non-point sources of fresh water currently being wasted.

Despite the degraded condition of the majority of the baldcypress-water tupelo swamps of the upper Lake Pontchartrain Basin, healthy areas of swamp still exist. Without exception, each of these swamps receives some form of reliable, high quality, nutrient-rich fresh water. These forests are either receiving non-point sources of fresh water from urban areas (e.g., forests of Hope Canal and Alligator Island), high quality river water (e.g., forests of Pearl River), or secondarily-treated sewage effluent (e.g., forests of Joyce Wildlife Management Area and Bayou Chinchuba). Efforts that include the proper combination of river diversions, of wetlands receiving treated sewage effluent, and of rerouted non-point source fresh water should enable restoration of Lake Pontchartrain Basin’s swamps (Shaffer and Day 2007).

TECHNOLOGY TRANSFER This study’s four objectives are vital to the restoration and sustainability of the Pontchartrain Basin. As such, policymakers and stakeholders interested in our findings include those

PBRP Annual Report | 2009 75 De-energizing Hurricanes with Cypress/Tupelo Buffers focused on reducing storm damage and those who make decisions concerning wetlands restoration in the basin. By presenting evidence at meetings sponsored by state and federal agencies, including those sponsored by local environmental groups, we will show that wetlands are not created equal with respect to resisting and reducing storm damage. We will provide a mechanism for prioritizing wetlands restoration projects in the basin by producing a GIS that displays potential point and non-point freshwater sources that could be used for nourishing and restoring swamps, improving water quality, and reducing storm damage. Other researchers could use our protocol to optimize the marriage of storm-damage reduction and wetlands restoration throughout coastal Louisiana.

More formal dissemination of our findings will occur through a final report (hard bound, CD); a master’s thesis; annual presentations at regional, national, and international conferences, and manuscripts published in international journals.

REFERENCES Barras, J.A.. 2006. Land area changes in coastal Louisiana after the 2005 hurricanes: A series of three maps. U.S. Geological Survey Open-File Report 6-1274.

Chambers, J.L., W.H. Conner, J.W. Day, S.P. Faulkner, E.S. Gardiner, M.S. Hughs, R.F. Keim, S.L. King, K.W. McKleod, C.A. Miller, J.A. Nyman, and G.P. Shaffer. 2005. Conservation, protection, and utilization of Louisiana’s coastal wetland forests. Final Report to the Governor of Louisiana.

Chambers, J.Q., J.I. Fisher, H. Zeng, E.L. Chapman, D.B. Baker, and G.C. Hurtt. 2007. Hurricane Katrina’s carbon footprint on U.S. Gulf Coast forests. Science 318:1107.

Doyle, T.W., B.D. Keeland, L.E. Gorham, and D.J. Johnson. 1995. Structural impact of Hurricane Andrew on forested wetlands of the Atchafalaya Basin in south Louisiana. Journal of Coastal Research 21:354-364.

Gresham, C.A., T.M. Williams, and D.J. Lipscomb. 1991. Hurricane Hugo wind damage to Southeastern U.S. Coastal forest tree species. Biotropica 23:420-426.

Krauss K.W., J.L. Chambers, J.A. Allen, D.M. Soileau Jr, and A.S. DeBosier. 2000. Growth and nutrition of baldcypress families planted under varying salinity regimes in Louisiana, USA. Journal of Coastal Research 16:153-163.

Lopez, J.A. 2006. The multiple lines of defense strategy to sustain coastal Louisiana. White Paper, Lake Pontchartrain Basin Foundation, New Orleans, LA, USA.

Louisiana Coastal Wetlands Conservation and Restoration Task Force and the Wetlands Conservation and Restoration Authority. 1998. Coast 2050: toward a sustainable coastal Louisiana. Louisiana Department of Natural Resources. Baton Rouge, LA, USA.

Massel, S.R., K. Furukawa, and R.M. Brinkman. 1999. Surface wave propagation in mangrove forests. Fluid Dynamics Research 24:219.

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Putz, F.E., and R.R. Sharitz. 1991. Hurricane damage to old-growth forest in Conaree Swamp National Monument, South Carolina, USA. Canadian Journal of Forest Research 21:1765-1770.

Saucier, R.T. 1963. Recent geomorphic history of the Lake Pontchartrain Basin. Louisiana State University Press, Baton Rouge, LA, USA.

Shaffer, G.P. and J.W. Day Jr. 2007. Use of freshwater resources to restore Baldcypress- Water Tupelo swamps in the Upper Lake Pontchartrain Basin. White Paper, Louisiana Department of Wildlife and Fisheries, Baton Rouge, LA, USA.

Shaffer, G.P., W.B. Wood, S.S. Hoeppner, T.E. Perkins, J.A. Zoller, and D. Kandalepas. 2009 (In Press). Degradation of Baldcypress-Water Tupelo swamp to marsh and open water in southeastern Louisiana, USA: an irreversible trajectory? Journal of Coastal Research.

Shaffer, G.P., J.W. Day, G.P. Kemp, S. Mack, M.A. Poirrier, K.A. Westpahl, D. FitzGerlad, and P.S. Penland. (In Review). The MRGO project: the worst man-made environmental, economic, and storm disaster in North American history. Journal of Coastal Research.

Trettin, C.C. and M.F. Jorgensen. 2003. Carbon cycling in wetland forest soils. Pages 311- 331 in J.M. Kimble et al., editors. The potential of U.S. forest soils to sequester carbon and mitigate the greenhouse effect. CRC Press, Boca Raton, FL, USA.

Williams, K., Z.S. Pinzon, R.P. Stumpf, and E.A. Raabe. 1999. Sea-level rise and coastal forests on the Gulf of Mexico. USGS-99-441.

Yoshihiro, M., E. Wolanski, B. King, A. Sase, D. Ohtsuka, and M. Mayoi. 1997. Drag force due to vegetation in mangrove swamps. Mangroves and Salt Marshes 1:193-199.

PBRP Annual Report | 2009 77 Mitigating the Spread of Zebra Mussels into Wetlands

Mitigating the Spread of Zebra Mussels into Wetlands from Mississippi River Diversions

William F. Font Department of Biological Sciences; Southeastern Louisiana University

ABSTRACT Mississippi River diversions are essential for navigation, flood control, and wetland restoration. However, the movement of water from the river to wetlands in southeastern Louisiana may also introduce the exotic zebra mussel, which is considered to be an aquatic nuisance species. This study is monitoring the dispersal of zebra mussels into wetlands and will determine if sustainable, reproductive populations have established. As a means of mitigating the spread of zebra mussels, a search is being conducted for parasitic castrators infecting native mussels that could infect zebra mussels and serve as biological control agents.

BACKGROUND & PRIMARY OBJECTIVES Of all alien species that have been introduced into the United States, one of the most invasive and pernicious is the zebra mussel, Dreissena polymorpha. This bivalve was first introduced into Lake St. Clair in 1988. Subsequently, it spread throughout the entire Great Lakes ecosystem and then into rivers of the northeastern United States. Zebra mussels have invaded the Mississippi River and are established as far south as New Orleans, Louisiana. The economic impact of zebra mussels, measured in millions of dollars, is coupled with its negative biological impact wherein the presence of this alien species has produced harmful effects on native species of aquatic animals.

The zebra mussel was first introduced into the Great Lakes with ship ballast water, not as adults, but rather as larvae. Briefly, the mussel has a free swimming ciliated larva called a veliger that serves as a natural dispersal stage. Ballast water was initially acquired by ships in European lakes where the zebra mussel occurs naturally. When these ships took on cargo in the Great Lakes, they released their ballast water containing veliger larvae of zebra mussels. The veliger larvae dispersed, settled, and attached to hard substrate in the Great Lakes. These mussels, attached by their adhesive byssal threads, grew and matured. They quickly established breeding populations throughout the Great Lakes and adjacent river systems.

The rapid and widespread dispersal of zebra mussels was initiated by their swimming larvae, but further spread was augmented by an additional mode of dispersal, i.e. anthropogenic activities. Because adult zebra mussels can attach to man-made objects such as boats and boat trailers, they can also be dispersed as adults by human movement associated with boat traffic and other activities.

Initial predictions stated that zebra mussels would be limited in their southward dispersion in the United States because this species is native to cold water lakes in Europe and would not

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be able to tolerate the warmer summer temperatures of southern wetlands. However, the zebra mussel was discovered in the Mississippi River in New Orleans in the early 1990s. This rapid southward spread can be explained because the water of the Mississippi River, moving south from the upper Midwest, is colder than the water of adjacent Louisiana wetlands. Nonetheless, the proximity of zebra mussels to Louisiana wetlands raises the specter of the potential of dispersal into wetlands.

Diversion of Mississippi River water into Louisiana swamps and marshes has occurred for many decades for flood control, associated with the opening of the Bonnet Carré and Atchafalaya Floodways. Recently, new diversions of Mississippi River water into wetlands of southeastern Louisiana have been constructed to rebuild marshes with freshwater and river sediments, enabling greater hurricane protection. These diversions are responsible for the introduction of zebra mussel veliger larvae into local wetlands. Therefore, this research seeks to determine whether these larvae will survive, become sexually mature adults, and establish reproductively sustainable populations in these new habitats.

In addition to conducting a survey of zebra mussels in aquatic habitats in southeastern Louisiana, this study will attempt determine if there are biological agents that might limit the spread of these exotic mussels, if they have already established breeding populations, or agents that might establish sustainable populations in the future. This search for biological control agents will center on parasites. Specifically, trematode parasites use certain mollusks, namely snails and bivalves, as first intermediate hosts in their life cycles. Theses trematodes are parasitic castrators and render their mollusk hosts sterile, making them ideal candidates for biological control agents of zebra mussels. Because species of trematodes display a rather strict specificity in their choice of first intermediate hosts, candidates for control agents would likely be mollusks that are phylogenetically closely related to zebra mussels. The sister species of the alien zebra mussel is Conrad’s false mussel, Mytilopsis leucophaeta, which occurs naturally in oligohaline marshes of south Louisiana. Thus, Conrad’s false mussel, and to a lesser extent other clams and mussels, will be intensively studied for the presence of trematode parasites that have the potential to infect and castrate zebra mussels.

RESULTS In addition to the results reported in my previous report of September 14, 2007, significant progress has been made in the past year with regard to conducting a survey for the occurrence of zebra mussels and discovering parasites of false mussels that have the potential to infect zebra mussels.

A comprehensive survey of the Lake Pontchartrain basin for the presence of Conrad’s false mussels and zebra mussels is nearing completion. False mussels occurred commonly in the western half of Lake Pontchartrain and bayous and other tributaries flowing into the lake, e.g. Bayou St. John, Bonnet Carré Floodway, Bayou Traverse, Bayou Labranche, and Pass Manchac. False mussels were rare or absent in eastern Lake Pontchartrain, probably due to higher salinities in aquatic habitats in closer proximity to the Gulf of Mexico.

False mussels were parasitized by three species trematodes that have the potential to infect zebra mussels. Taxonomic identification of these trematodes is still in progress, but one

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species has been identified as a new species of Lasiotocus. A second infection in false mussels has been determined to be a new species in the family Zoogonidae. A third trematode infecting false mussels is assigned to the family Bucephalidae. Examination of the gonads of infected mussels showed that all three species of trematodes are capable of castrating their molluskan hosts. Gravid adults of the new species of Lasiotocus were discovered in naked gobies, Gobiosoma bosc, thereby making it possible to obtain eggs in order to attempt experimental infections of zebra mussels in the laboratory. A search is underway to discover the fish host harboring gravid adults of the zoogonid trematode so that similar experimental infections can be attempted with this species of parasite. It is unlikely that the bucephalid life cycle can be maintained in the laboratory, making experimental infections of zebra mussels with this parasite problematic, although the parasite may still serve as a control agent in nature. I discovered a correlation between the presence or absence of false mussels and the presence or absence of Lasiotocus n. sp. and the zoogonid. In eastern Lake Pontchartrain, where there are no false mussels, gobies are uninfected with these two parasites. Although this implies that no biological control agents for zebra mussels exist in that part of the basin, this is not considered a problem for the following reason. Because the zebra mussel is even less tolerant of high salinities than is the false mussel, it is not likely that zebra mussels will ever invade eastern Lake Pontchartrain. Thus biological control agents are not necessary in this geographic area.

In my previous report, I discussed the potential of Lasiotocus minutus infecting rainwater killifish, Lucania parva to serve as a biological control agent for zebra mussels. Unfortunately, after much effort, I have discovered the bivalve host of this parasite is not the false mussel, but a distantly related clam, making infections of zebra mussels unlikely.

With the survey of Lake Pontchartrain nearing completion, my new research focus involves examining the Mississippi River diversions for false mussels and zebra mussels. Specifically, this research is being conducted in Bonnet Carré Floodway, Davis Canal Diversion, Caenarvon Diversion, and the navigation locks associated with the Intracoastal Waterway located in Harvey Canal and Port Allen. After several iterations, a monitoring device for detecting colonization by false mussels and zebra mussels has been perfected. This sentinel device consists of one-eighth inch nylon mesh bags filled with oyster shells. Mussels attach readily to oyster shells, and the mesh bags can be monitored seasonally to determine if mussels have colonized and to determine mussel population parameters including growth, maturation, reproduction, and acquisition of parasites. In addition, some of these bags are modified to simultaneously collect naked gobies in order to determine whether these fish are parasitized by potential biological control agents.

By far the most significant discovery during the past year was the finding of zebra mussels in the Bonnet Carré Spillway. Because of high water in the Mississippi River, the spillway was opened in spring 2008, allowing river water to flow through the spillway into Lake Pontchartrain. Mesh bags containing oyster shells were colonized by both zebra mussels from the Mississippi River and false mussels recruited from within the spillway. Survival and growth of both species of mussels was documented through late spring and summer. By late summer, with an increase in water temperature, some mortality of zebra mussels was noted, but a portion of the population survived into the fall when water temperatures declined.

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FUTURE PLANS Because of the establishment of zebra mussels in the Bonnet Carré Spillway, that study site will become the primary focus of this research. Continued monitoring of zebra mussels will assess several important biological parameters. Survival, growth, and maturation will be recorded. Additional new mesh bags of oyster shells will be placed in the spillway in early spring of 2009. Two possible scenarios will be anticipated depending on whether the spillway is reopened in 2009. First, if the spillway is not opened, recruitment of zebra mussels on the new oyster shells will be conclusive evidence that the zebra mussels have established a breeding population in the spillway. Second, if the spillway is reopened, colonization of oyster shells by zebra mussels may have two causes: either recruitment from a breeding population in the spillway or new recruitment from the Mississippi River. It will then be necessary to directly observe recruitment of established zebra mussels by bringing them into the laboratory and microscopically examining their gonads for mature eggs and sperm. Also in the lab, both zebra mussels and false mussels will be removed from oyster shells, maintained in the lab, and examined for trematode parasites.

Second only to research in the Bonnet Carré Spillway, other sources of Mississippi River water into Louisiana wetlands will be monitored for zebra mussels. In spring 2008, mesh bags of oyster shells serving as sentinels for zebra mussel recruitment were placed in other river diversions located in Caernarvon, south of New Orleans, and the Port Allen locks and Intracoastal waterway on the west side of the Mississippi River near Baton Rouge. In fall 2008, sentinel oyster shells were set in Lake Cataouche, which receives river water from the Davis Canal Diversion near Ama. Additional sentinel bags of oyster shells were placed in Harvey Canal and in the Intracoastal Waterway near Lafitte. Each of these sites will be visited regularly on a rotating basis to assess zebra mussel colonization throughout 2009.

At the annual meeting of the Louisiana Academy of Sciences in February 2009, researchers from Nicholls State University reported their discovery of a population of zebra mussels in Bayou Lafourche that was able to survive the warm summer water temperatures of a Louisiana bayou. I intend to complement their study by collecting zebra mussels in Bayou Lafourche and examining them for parasitic infections.

TECHNOLOGY TRANSFER This study’s goal is to better understand the role that Mississippi River diversions play in the actual or potential spread of zebra mussels into adjacent wetlands in southeastern Louisiana. The role of parasites in mitigating the spread of zebra mussels is also being assessed. Because trematodes castrate their molluskan hosts, they may serve as biological control agents that can limit the dispersal and reproductive success of zebra mussels. Native mussels, especially Conrad’s false mussel, will continue to be examined for trematodes that might infect zebra mussels if this alien species is introduced into Louisiana wetlands.

The success of restoration programs for Louisiana wetlands depends greatly on the rebuilding of marshes associated with the diversion of freshwater and sediments from the Mississippi River. Due to the need to maintain these diversions, we must be aware of any negative consequences associated with their presence. Without question, exotic zebra mussels will be introduced into Louisiana wetlands through these Mississippi River

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diversions in the form of veliger larvae, the dispersal stage of this mollusk. What remains unknown, though, is whether zebra mussels can establish, grow, mature, and reproduce in wetlands or whether they will be limited by high salinities and high summer temperatures. Also unknown is whether biological control agents, especially parasites, are present in Louisiana that may mitigate the spread of zebra mussels.

Hypothetically, zebra mussels may spread into wetlands although they may not initially be well adapted to survive the high water temperatures occurring in summer. However, because this species is able to evolve adaptations to new environmental conditions over time, it is further hypothesized that the potential for the establishment of sustainable populations of zebra mussels will increase in the future. A third hypothesis is that native mussels have parasites that have the potential to infect the closely related zebra mussel, and thus serve as agents that may negatively impact the reproductive success of this alien species. Given my research results to date, I recommend continually monitoring the Louisiana wetlands for the presence of zebra mussels. In addition, both native mussels and this alien mussel species should be assessed for the presence of parasites that can limit dispersal and retard population growth rates.

Federal and state wildlife agencies need to know the status of zebra mussels in Louisiana wetlands. The United States Army Corp of Engineers is charged with the creation and maintenance of Mississippi River diversions such as Bonnet Carré Floodway, Caenarvon, and Davis Canal Diversions as well as with navigation aids such as the Intracoastal Waterway and locks located at Port Allen and Harvey Canal. Jean Lafitte National Park borders Lake Cataouche, which may be colonized by zebra mussels introduced through the Davis Canal Diversion. In addition, commercial fishermen who regularly are confronted with native false mussels must be aware of the potential colonization of exotic zebra mussels and should be taught how to distinguish between these two species. Each of these state and federal agencies is impacted by the potential spread of zebra mussels. Because Mississippi River Diversions are vital to wetlands restoration, their presence will continue and the number of diversions is likely to increase, thereby increasing the potential for zebra mussels to spread. Yet continual monitoring will ensure these agencies can prepare for the negative impacts of this aquatic nuisance species and may formulate plans to control its spread.

82 PBRP Annual Report | 2009 Development of an Index of Biological Integrity for the Wetlands

Development of an Index of Biological Integrity for the Lake Pontchartrain Basin Wetlands

Janice Bossart1 and Colin Jackson2 1 Department of Biological Sciences; Southeastern Louisiana University 2 Department of Biology; University of Mississippi

ABSTRACT This collaborative study is a comparative, joint analysis of the benthic invertebrate diversity, microbial activities, and water chemistry at a strategic array of sites in the Western Lake Pontchartrain Basin (WLPB). Study sites have been selected to span and anchor both ends of a wetlands disturbance gradient. Their survey will be used to identify consistent, reliable biological indicators (metrics) and to determine how these correlate with water quality. Survey data will ultimately allow us to establish a detailed narrative of what constitutes non- degraded versus degraded swamp habitat in the WLPB. The overriding objective of our study is to use these comparative data to develop the first ever biologically-based management tool (an Index of Biological Integrity) for use by conservation managers for the rapid bioassessment of Southeastern Louisiana’s cypress swamps.

PRIMARY OBJECTIVE & METHODS This study is comprised of the following methods to attain the primary objective of creating an Index of Biological Integrity (IBI) to assess the health of Louisiana’s cypress swamps: Collect benthic macroinvertebrate, sediment microbial, and water chemistry samples on a regular recurring schedule for one year at forested wetland sites that span a habitat condition gradient. Characterize invertebrate community diversity, microbial activity levels, and water chemistry profiles at each sampling site. Quantify the relationships among community diversity, microbial activities, and measurements of water chemistry, i.e. dissolved oxygen, pH, salinity, and temperature. Identify which biological attributes (metrics) show quantitative and predictable responses to measured changes in water chemistry. Establish the reference set of baseline conditions (biological and physical) that discriminate among intact, intermediate, or degraded swamp habitats. Establish scoring criteria for each biological metric to develop an IBI.

RESULTS In this second year of the project, activities emphasized the 1) ongoing and regular sampling of our seven primary sites, 2) supervision and training of field and laboratory personnel and student workers, and 3) successful transport of sediment samples to Colin Jackson at University of Mississippi.

PBRP Annual Report | 2009 83 Development of an Index of Biological Integrity for the Wetlands

Specifically, six primary sites were identified for regular sampling: three at Joyce Wildlife Management Area (JWMA) and three in the Lake Maurepas Wetlands System (LMWS). The JWMA sites—Kleibert’s Ditch, Mainline Boat Trail, and Swamp Boardwalk—represent relatively intact cypress-tupelo swamp but are also characterized by slight differences in salinity. The LMWS sites represent intact (Alligator Island), partially degraded (Blind River), and severely degraded (Ruddock) swamp habitat. Habitat condition classes at LMSW are based on plant community analyses conducted independently by Gary Shaffer, a plant ecologist at Southeastern.

Two additional sites not outlined in the original proposed research plan were added at Three-Mile Marsh. This natural marsh lies just north of JWMA and receives affluent from the Hammond sewage treatment plant. Community samples are being collected along two 200m boardwalks that extend perpendicular from the affluent point-source into the marsh. Survey of these sites will provide both valuable comparative data with the derived marsh habitat that occurs in severely degraded areas of Lake Maurepas and vital clues about the use of nutrient-rich waste water to restore forested wetlands.

The on-going, regular sampling at the JWMA sites was initiated in May 2007 and ended in July 2008, for a total of 14 sampling bouts. Sampling at the LMWS sites was delayed because of difficulties in gaining access to sites (lack of trained drivers for boats and other logistical concerns). Although regular sampling at Blind River and Alligator Island began only a few months later (October 2007), it was not initiated at Ruddock until April 2008. Because the LMWS sites can only be accessed by boat, weather events have also impacted the frequency and quality of samples (on multiple occasions these “wetlands” have been dry). Sampling at the LMWS and Three-Mile Marsh sites is ongoing and will continue until 14 high quality samples have been collected from each.

To date, more than 6232 benthic invertebrate specimens have been collected across all sites (fig. 1). Comparison across sites is limited given the unequal number of sampling bouts that have occurred. However, many more specimens have been collected from Three-Mile Marsh-1 than at JWMA (all three sites combined) despite greater sampling effort at the latter site (13 and 42 total samples, respectively). Similarly, the specimen number collected from Three-Mile Marsh-2 and the three LMWS sites is comparable despite twice as many samples being collected at each of the three LMWS sites. High specimen numbers at the Three-Mile Marsh sites could reflect positive correlation between increased nutrient levels and number of invertebrates collected.

Taxonomic identification of the invertebrate samples collected is well underway. So far 41 families are represented in the collections. Nearly half of the specimens are insects (44%), but Crustecea, Mollusca, Annelida, and Nematoda are also well represented. Sampling is ongoing at half of the sites, so relative differences may change. But to date, family diversity is highest at Three-Mile Marsh-1 and lowest at Ruddock, our degraded site (fig. 2). The three intact JWMA sites have similar levels of family diversity. Unexpectedly, family diversity at Alligator Island, the intact LMWS site, was much lower that of the intact JWMA sites; whereas, diversity at Blind River, the intermediate LMWS site, is identical to that at the intact JWMA sites.

84 PBRP Annual Report | 2009 Development of an Index of Biological Integrity for the Wetlands

2000 d

ecte 1500 ll

1000 mens co i

spec 500 #

0 -1 -2 A d r k h h M an ve c rs rs W sl Ri do a a e r I d d M M c to in u ile ile oy a l R M M J lig B 3- 3- Al

Figure 1. Total number of benthic invertebrates collected at each study site

20 Families # 15

3-Mile Marsh-1 10 Swamp Boardwalk Mainline Kleiberts

ummulative 5 Alligator Island

C Blind River Ruddock 0 0 200 400 600 800 1000 1200 1400 1600 # Individuals Collected Figure 2. Rarefaction curve of cumulative family diversity at each site

PBRP Annual Report | 2009 85 Development of an Index of Biological Integrity for the Wetlands

Sediment samples collected at each of the sites have been assayed for activities of the hydrolytic microbial enzymes: β-glucosidase (B-gluc), cellobiohydrolase (CBH), N-acetyl-β- glucosaminidase (NAG), β-xylosidase (B-xylos), and acid phosphatase (Phos) using p- nitrophenyl linked artificial substrates; and for the activity of the oxidative enzymes phenol oxidase (Phen Ox) and lignin peroxidase (Perox) using L-DOPA. These seven microbial enzymes are involved in both organic matter decomposition (B-gluc, CBH, B-xylos, Phenol Ox, Perox) and nutrient cycling (Phos, NAG).

Activity of the four hydrolytic enzymes related to organic matter decomposition was detectable at all sites on all dates sampled. In contrast, activity of the oxidative enzymes was often absent, especially the activity of phenol oxidase, which was rarely detectable and generally inconsistent when it was detected (e.g., it may have been detected on one date at a site, but not on other dates). These results suggest that the hydrolytic enzymes will be useful for habitat condition monitoring purposes but the oxidative enzymes will not. Notably, because activity of both enzymes involved in nutrient cycling were consistently detected, these enzymes could prove valuable also for determining nutrient-related differences at sites throughout this wetlands system.

As might be expected, if drivers of enzyme activity are affecting microbial activity in general, enzyme activity levels were often highly correlated at each site. This link was particularly true for the hydrolytic enzymes, β-glucosidase and cellobiohydrolase, which showed an average correlation of 0.81 across all sites and dates (tab. 1). But this link is not unexpected given that both these enzymes are involved in cellulose degradation. Although still positive, the relationship between acid phosphatase and the other hydrolytic enzymes was generally weaker, suggesting that additional factors are driving the activity of this enzyme compared to the organic matter degrading enzymes. Despite the overall pattern of correlation between β- glucosidase and cellobiohydrolase, activity levels of these two enzymes were decoupled at Mainline-JWMA and Three-Mile Marsh-1. In general, lignin peroxidase activity was uncorrelated with the activity of any other enzymes except at Alligator Island-LMWS, where it was strongly correlated with cellobiohydrolase (0.71), and at Blind River-LMWS, where it was strongly correlated with both cellobiohydrolase (0.72) and β-xylosidase (0.91).

Table 1. Correlations of microbial enzyme activities across all sites and dates

B-gluc CBH NAG B-xylos Phos Phen-Ox B-gluc 1.0 CBH 0.807 1.0 NAG 0.682 0.787 1.0 B-Xylos 0.633 0.697 0.588 1.0 Phos 0.513 0.486 0.627 0.601 1.0 Phen-Ox 0.076 0.155 9.133 0.111 0.114 1.0 Perox 0.166 0.339 0.157 0.226 -.0116 0.420

The relationship between enzyme activities and the habitat condition class of a site is not easily interpreted. For example, although activity levels for the hydrolytic enzymes were

86 PBRP Annual Report | 2009 Development of an Index of Biological Integrity for the Wetlands

generally lowest at the three marsh sites (i.e., Ruddock and Three-Mile Marsh-1 and 2), there were notable exceptions. Acid phosphatase activity at Three-Mile Marsh-1 was double that at Three-Mile Marsh-2 and Ruddock. On average, hydrolytic enzyme activity levels were highest at Swamp Boardwalk-JWMA in all cases except for cellobiohydrolase (fig. 3), and were more similar to those at Alligator Island, the intact LMWS site, than to the other two JWMA sites. With respect to β-glucosidase, this difference was particularly apparent, measuring 27.77 and 25.98 µmol/hr/gOM for Swamp Boardwalk and Alligator Island, respectively versus measuring 16.94 and 14.5 µmol/hr/gOM for the other two JWMA sites. In general, enzyme activity levels tended to be variable across sites and across collecting dates (e.g. β-glucosidase; fig. 3).

Figure 3. Activity of the hydrolytic microbial enzyme, β-glucosidase, in sediment samples

FUTURE PLANS Sampling of the Three-Mile Marsh and LMWS sites will continue until 14 quality samples have been collected. We anticipate that sampling at the LMWS will be completed by June 2009, but not until late fall 2009 at Three-Mile Marsh. The vast majority of specimens collected thus far have been identified to family. A major project goal for the upcoming year is to identify all insects to at least genus and preferably to species, an endeavor that has proven more time consuming than originally anticipated. Non-insect specimens, such as Annelida, will be identified to lower taxonomic categories where possible. Microbial activity analyzes will continue until sampling is terminated. Analysis and interpretation of both the microbial enzyme activity and invertebrate community data is well underway and will continue until project end and beyond. Development of our Index of Biological Integrity (IBI) requires that we first identify a suite of measurable, biological attributes (metrics) that

PBRP Annual Report | 2009 87 Development of an Index of Biological Integrity for the Wetlands

are indicative of swamp health. We have already begun to identify which enzymes and taxa will be useful for monitoring the health of these forested wetlands. Coleoptera (beetles), for example, are apparently linked to high quality swamps; whereas, Diptera (flies) are more indicative of degraded swamp habitat. We expect that by early summer 2010 we will have the empirical knowledge in hand to formally develop the IBI specific to Louisiana’s cypress- tupelo swamps.

TECHNOLOGY TRANSFER The overall goal of our project is to provide a scientifically based, management tool for use by conservation stakeholders to assess the condition/health of Louisiana’s cypress-tupelo swamps in the Western Lake Pontchartrain Basin (WLPB). This management tool (an Index of Biological Integrity (IBI)) will be developed based on biological criteria (microbes and benthic invertebrates) and as such will be able to capture the complexity inherent in ecological systems that is not provided by traditional, point-in-time chemical assessments.

Forested wetlands are a predominant habitat in the WLPB. Reversing their loss and degradation has been identified by state and national stakeholders as a chief conservation priority as ecosystem sustainability is vitally linked to ecosystem health. Current efforts to evaluate and track the condition of these key ecosystems are stymied by the absence of ecologically based assessment tools. Development of an IBI specific to cypress-tupelo swamps will give stakeholders a “yardstick” by which to measure swamp condition and a powerful tool for compiling the critical ecological data needed by managers and decision- makers. Development of an IBI specific to the WLPB additionally links directly to the imminent restoration activities planned for the basin. This biologically-based management tool will let stakeholders easily track, for example, changes in the health of Maurepas Swamp resulting from the Mississippi River freshwater diversion project.

Our target groups, microbes and benthic invertebrates, are both superlative and proven indicators of, and directly linked to, ecosystem health. Their survey will provide information on swamp condition over both the short- (days-months; microbes) and mid- (months-years; invertebrates) term. This index will allow decision-makers to more easily assess real-time and ongoing changes in these ecosystems that might occur in response to natural events, such as hurricanes, or anthropogenic impacts, such as freshwater diversions or increased north shore development and pollution. Corrective measures could then be implemented earlier rather than later, thus reducing the breadth and impact of the disturbance. We imagine that results of our study will be most immediately and directly valuable to state and federal employees who are working to manage and restore the wetland forests of Louisiana and elsewhere.

88 PBRP Annual Report | 2009 Determining the Potential for Algal Bloom in Lake Maurepas

Determining the Potential for Algal Bloom in Lake Maurepas: Effects of Changing Nutrient Load from Freshwater Diversion and Changes in Human Population

Philip Voegel1, James Pinckney2, and Sophia Passey3 1 Department of Chemistry and Physics; Southeastern Louisiana University 2 Department of Biological Sciences; University of South Carolina 3 Department of Biology, University of Texas, Arlington

ABSTRACT This project included two phases of research to determine the potential for algal bloom in Lake Maurepas. During the first year of the project, the concentrations of phosphate and silicate, along with pH and water temperature, were determined at nine locations in Lake Maurepas and then compared to published historical data. During the second year of the project, the opening of the Bonnet Carré Spillway provided a unique opportunity to directly study the effects of freshwater diversion on Lake Pontchartrain. Concentrations of salinity, nitrate, phosphate, and silicate were determined and then compared to changes observed in chlorophyll concentrations at six sites near the spillway before, during, and after its opening.

PRIMARY OBJECTIVES During the first year of the project, we focused on Lake Maurepas. First, we monitored water-quality parameters including nutrient and chlorophyll concentrations at nine locations in Lake Maurepas. Then we compared those parameters to historical data for the same areas. This comparison enabled us to determine which water-quality parameters are most likely to lead to increases in algal growth in Lake Maurepas.

The second year of the project focused on the Bonnet Carré Spillway and Lake Pontchartrain. For this study, we monitored water-quality parameters including nutrients and chlorophyll concentrations at six locations near the Bonnet Carré Spillway both while the spillway was actively diverting water into Lake Pontchartrain and after the spillway was closed. Then we determined which water-quality parameters were most likely to lead to increases in algal growth in Lake Pontchartrain, and we determined the amount of time required for the water-quality parameters to return to normal levels following the closing of the spillway.

RESULTS: MAUREPAS STUDY Overview: In the study of water quality in Lake Maurepas, temperature, pH, phosphate, and silicate were successfully recorded during sampling trips from June 8, 2007 to March 28, 2008. Nitrate levels were initially examined using an ion-selective electrode method; however, nitrate levels in Lake Maurepas were below the detection limit for this method. To correct this limitation, the cadmium reduction spectroscopic method for nitrate detection

PBRP Annual Report | 2009 89 Determining the Potential for Algal Bloom in Lake Maurepas

was attempted. Unfortunately, this method never proved consistent or reproducible in the hands of undergraduate researchers. Filtered algae samples were submitted to Jay Pinckney at the University of South Carolina for analysis by HPLC. The concentrations of algal pigments determined using this method were generally lower than expected and generally below the method’s detection limit. Similarly, the limited concentration of phytoplankton cells made identification by light microscopy by a second collaborator, Sophia Passey at the University of Texas at Arlington, impractical. Compared to historical data from studies occurring prior to Hurricane Katrina, statistically relevant increases in phosphate and silicate are observed in this study. No statistical differences in pH were observed.

Sampling: In April 2007, three samples were taken from with 500m of each other at each of nine locations in Lake Maurepas. Because no significant differences were observed in nutrient levels for these samples, only one sample was taken from each of the nine sites during future sampling trips. Sampling sites included the four primary sources of freshwater entering Lake Maurepas (Amite, Blind, Tickfaw Rivers and Reserve Canal), the primary outflows from the lake (Pass Manchac and North Pass), and three sites in the middle of the lake. Table 1 shows the recorded GPS coordinates for each site, which are mapped in figure 1. At each location, a 2.2L horizontal alpha water sampler was used to obtain water from just below the lake’s surface. Water temperature was immediately measured. The samples were transferred to plastic bottles, placed in a storage cooler, and transported to a laboratory at Southeastern Louisiana University where they were refrigerated at 4°C until analyzed.

Table 1. Sampling locations Inflow Sites Amite 30°17.94±0.02N x River 90°33.35±0.01W Blind 30°12.46±0.01N x River 90°35.20±0.04W Reserve 30°10.35±0.02N x Canal 90°32.94±0.01W Tickfaw 30°20.47±0.04N x River 90°28.51±0.03W Mid-Lake Sites 1 30°13.09±0.02N x 90°31.91±0.01W 2 30°16.23±0.02N x 90°31.69±0.02W 3 30°16.38±0.02N x 90°28.25±0.03W Figure 1. Lake Maurepas sampling sites Outflow Sites North 30°18.64±0.07N x Pass 90°25.25±0.02W Pass 30°24.84±0.01N x Manchac 90°24.84±0.01W

90 PBRP Annual Report | 2009 Determining the Potential for Algal Bloom in Lake Maurepas

pH Analysis: pH levels of water samples were determined with an Orion pH meter after filtration. The analysis of pH was completed 11 times from June 8, 2007, through January 29, 2008 (fig. 2). The overall pH range was from a minimum of 6.25±0.11 on January 29, 2008, at the Tickfaw River site to a maximum of 8.28±0.01 on November 22, 2007, at the Mid-Lake 2 site.

8.5 8.5 8 8 7.5 7.5 7 7 6.5 6.5 pH pH 6 6 5.5 5.5 5 5 4.5 4.5 4 4 6/8/07 7/15/07 8/21/07 9/27/07 11/3/07 12/10/07 1/16/08 6/8/07 7/16/07 8/23/07 9/30/07 11/7/07 12/15/07 1/22/08

Reserve Tickfaw Amite Blind Manchac North Pass Midlake 1 Midlake 2 Midlake 3 Figure 2. pH at inflow sites (left) and outflow and mid-lake sites (right)

Phosphate Analysis: Phosphate levels were determined usin a standard spectroscopic method. Phosphate analysis was completed 12 times from June 8, 2007, through March 28, 3- 2008 (fig. 3). Phosphate levels ranged from a minimum of 0.037±0.009ppm PO4 at Pass 3- Manchac on June 22, 2007, to a maximum of 0.421±0.009ppm PO4 at Reserve Canal on June 8, 2007.

0.450 0.450 0.400 0.400 0.350 0.350 0.300 0.300 0.250 0.250 ] ppm ] ppm ] 3- 3-

4 0.200 4 0.200 0.150 0.150 [PO [PO 0.100 0.100 0.050 0.050 0.000 0.000 6/8/07 7/28/07 9/16/07 11/5/07 12/25/07 2/13/08 6/8/07 7/28/07 9/16/07 11/5/07 12/25/07 2/13/08

Reserve Tickfaw Amite Blind Manchac North Pass Midlake 1 Midlake 2 Midlake Figure 3. Phosphate at inflow sites (left) and outflow and midlake sites (right)

Silicate Analysis: Silicate levels were determined using a standard spectroscopic method. Silicate analysis was completed eight times from September 10, 2007, through March 28, 2- 2008 (fig. 4). Silicate levels ranged from a minimum of 3.358±0.007ppm SiO3 at Mid-Lake 2 2- on November 2, 2007, to a maximum of 12.12±0.05ppm SiO3 at Reserve Canal on January 14, 2008.

PBRP Annual Report | 2009 91 Determining the Potential for Algal Bloom in Lake Maurepas

14 14 12 12

10 10 8 8 ] ppm ] ppm ] 2- 2- 3 6 3 6

[SiO 4 [SiO 4

2 2 0 0 9/10/07 10/30/07 12/19/07 2/7/08 3/2 9/10/07 10/30/07 12/19/07 2/7/08 3/2

Reserve Tickfaw Amite Blind Manchac North Pass Midlake 1 Midlake 2 Midlake Figure 4. Silicate at inflow sites (left) and outflow and mid-lake sites (right)

RESULTS: BONNET CARRÉ SPILLWAY STUDY Overview: In the study of water quality in Lake Pontchartrain from just before the April 11, 2008, opening of the Bonnet Carré Spillway until well after its closing on May 8, 2008, temperature, pH, phosphate, silicate, nitrate, ammonia, conductivity and chlorophyll a were successfully recorded. Sampling trips occurred April 11-August 14, 2008. Because of the previous difficulties when using the cadmium reduction method for nitrate analysis and because of the lack of sensitivity for nitrate ion-selective electrodes, nitrate was determined by simple background-corrected spectroscopic analysis at 220nm. Limited interference by organic matter in Lake Pontchartrain was observed, and this simple method proved quite successful in the hands of undergraduate researchers.

Sampling: Due to high water in the Mississippi River, the U.S. Army Corp of Engineers announced plans to open the Bonnet Carré Spillway. This unique opportunity to study of a large scale freshwater diversion into the Lake Pontchartrain Basin was deemed as more important than continuing to collect baseline information on nutrient levels in Lake Maurepas. Thus, in early April 2008, six sites were selected in Lake Pontchartrain near the Bonnet Carré Spillway for study. Four sampling sites were located at approximately one and two miles directly out from the spillway’s guide levees with two additional sites located approximately one mile to the east and west of the guide levees. Table 2 shows the recorded GPS coordinates for each site, which are mapped in figure 5. At each location, a 2.2L horizontal alpha water sampler was used to obtain water from just below the lake’s surface. Water temperature was immediately measured. The samples were transferred to plastic bottles, placed in a storage cooler, and transported to a laboratory at Southeastern Louisiana University where they were refrigerated at 4°C until analyzed.

92 PBRP Annual Report | 2009 Determining the Potential for Algal Bloom in Lake Maurepas

Table 2. Sampling locations Site 1 30°5.12±0.01N x 90°23.18±0.01W 2 30°4.34±0.01N x 90°21.84±0.01W 3 30°4.00±0.01N x 90°20.05±0.01W 4 30°5.27±0.01N x 90°21.23±0.01W 5 30°5.99±0.01N x 90°22.67±0.01W 6 30°6.41±0.01N x 90°24.50±0.01W

Figure 5. Lake Maurepas sampling sites

Analysis of Water Quality Parameters: The level of pH was determined using an Orion pH meter. Conductivity and temperature were determined using a Vernier LabQuest datalogger with calibrated conductivity and temperature probes. Temperatures ranged from 16.2°C at site 6 on April 15th to 32.0°C at site 6 on June 2nd. Temperature was not recorded on August 14 th due to equipment failure. Conductivities ranged from 223μS/cm at site 1 on April 15th to 6848μS/cm at site 3 just after the opening of the spillway on April 11th. The pH ranged from 7.20±0.09 at site 1 on April 17th to 8.92±0.01 at site 4 on June 2nd. The concentration of chlorophyll a was determined by filtration with glass fiber filter papers and extraction of chlorophyll into 90% aqueous acetone followed by spectroscopic analysis. Just before the opening of the spillway on April 11th, chlorophyll a levels were below the detection limit for the UV-visible spectroscopic method of analysis employed at 5 of the 6 sites. The chlorophyll a levels were 1.9±1.7 ppb at site 4, but ranged from 1 ppb at site 4 to 15ppb at site 1 (one of the sites closest to the spillway). The average values for each date in the study area are shown in figure 6.

Chemical Analyses of Phosphate, Silicate, Nitrate, and Ammonia: Standard spectroscopic methods were employed for the analysis of each analyte. Nitrate used a simple direct UV detection method at 220nm following background subtraction to account for potential interference from dissolved organic matter. Phosphate levels ranged from 0.04±0.01ppm at site 1 on May 27th to 0.86±0.02 ppm at site 6 on June 9th. Silicate

PBRP Annual Report | 2009 93 Determining the Potential for Algal Bloom in Lake Maurepas

concentrations ranged from 0.27±0.11 ppm at site 4 on April 11th, just before the opening of the spillway, to 12.43±0.06 at site 5 on April 17th. Nitrate levels were below the detection limit at most sites on April 11th (just after the spillway’s opening), on June 2nd, and on June 9th (approximately a month after the spillway’s closing). The highest rate recorded was 2.115±0.005 ppm at site 2 on April 17th. Ammonia concentrations were below the detection limit at all 6 sites on April 11th (just after the opening of the spillway), on April 15th, and on April 17th. But ammonia concentrations increased to a maximum of 29.4±0.2 ppbN at site 6 on June 9th. The complete data are summarized as the daily averages for all sites in the study area in figure 6.

DISCUSSION: MAUREPAS STUDY The levels of pH, phosphate, and silicate observed in this study were compared to historical data available from a number of resources. First, we compared them to data collected by the USGS from April 15, 1975 through January 19, 1981.1 The data obtained from USGS for various sites near the center of the lake were combined into a single average for comparison to the combined data from the three mid-lake sites in our study (see table 3 for a summary). The combined average pH for the lake, as a whole, in this study is not statistically different

from that observed in the USGS data (texp=1.20, tcrit=1.97); however, when specifically comparing the average mid-lake pH to that determined by USGS, the current values are

statistically higher (texp=5.40, tcrit=1.97). The range of pH values observed in the current study is also, overall, higher than that observed by USGS from 1975-1981, particularly with respect to the mid-lake samples.

Table 3. Comparison of pH from this study to historical data Site n Average Range Our study Lake Maurepas 99 7.19±0.06 6.25 – 8.28 Inlet Sites 44 6.99±0.08 6.25 – 7.57 Outlet/Mid-Lake 55 7.35±0.06 6.88 – 8.28 Mid-Lake 33 7.42±0.09 7.16 – 8.28 USGS Mid-Lake 137 7.15±0.05 6.10 – 7.90

Phosphate levels determined in our study were compared to data collected from April 15, 1975 through January 19, 1981 by the USGS1 near the center of the lake and were compared to data collected at Pass Manchac, the Reserve Canal, and the Blind River from April 4, 2002 through May 13, 2003 by Day, et al.2 The phosphate data is summarized in table 4. The average phosphate level throughout Lake Maurepas over the course of this study is

comparable to that observed by the USGS near the center of the lake (texp=0.06, tcrit=1.97). Similarly, when comparing mid-lake samples from our study (0.127±0.012ppmP) to those

obtained by USGS (0.143±0.029ppmP), there was no statistical difference (texp=0.53,

1 Full address for data from USGS (April 15, 1975 – January 19, 2981): http://nwis.waterdata.usgs.gov/ nwis/qwdata?site_no=301500090300000&agency_cd=USGS&format=brief_list 2 Full address for data from Day et al. (April 4, 2002 – May 13, 2003): http://epa.gov/region6/water/ ecopro/em/cwppra/maurepas/wq_hydrologic_modeling_final_report.pdf

94 PBRP Annual Report | 2009 Determining the Potential for Algal Bloom in Lake Maurepas

tcrit=1.97). Comparisons to more recent data collected by Day et al., prior to Hurricane Katrina, however, show that a significant increase in phosphate levels has occurred in Lake Maurepas since Hurricane Katrina.

7000 35 6000 30 C) o

S/cm) 5000 25 μ 4000 20

3000 15 2000 10

1000 ( Temperature 5 Conductivity ( Conductivity 0 0 4/10/08 5/5/08 5/30/08 6/24/08 7/19/08 8/13/08 4/10/08 4/25/08 5/10/08 5/25/08 6/9/08

10 45 40 9

] ppb 35 8 a 30 25 7 pH 20 6 15 10 5 5 [Chloropophyll 4 0 4/10/08 5/5/08 5/30/08 6/24/08 7/19/08 8/13/08 4/10/08 5/5/08 5/30/08 6/24/08 7/19/08 8/13/08

0.6 1.8 1.6 0.5 1.4 0.4 1.2 1 ] ppm ] ppm

0.3 - 3- 3

4 0.8 0.2 0.6 [NO [PO 0.4 0.1 0.2 0 0 4/10/08 5/5/08 5/30/08 6/24/08 7/19/08 8/13/08 4/10/08 5/5/08 5/30/08 6/24/08 7/19/08 8/13/08

12 25

10 20 8 15 ] ppm 6 ] ppb N ] ppb 2- + 3 4 10 4 [SiO [NH 2 5

0 0 4/10/08 5/5/08 5/30/08 6/24/08 7/19/08 8/13/08 4/10/08 5/5/08 5/30/08 6/24/08 7/19/08 8/13/08

Figure 6. Summary of Bonnet Carré Spillway monitoring results

PBRP Annual Report | 2009 95 Determining the Potential for Algal Bloom in Lake Maurepas

The concentrations of phosphate observed by Day from 2002-2003 were 0.07±0.03ppmP, 0.06±0.03ppmP, and 0.06±0.03ppmP at Pass Manchac, Reserve Canal, and the Blind River, respectively. At corresponding sites in our study, phosphate concentrations were 0.12±0.04ppmP, 0.19±0.05ppmP, and 0.18±0.04ppmP, showing an increase in concentration of 2-3 times more than the Day et al. study. These increases in concentration were all

statistically significant at the 95% confidence level with texp being 5.52, 4.97, and 5.10 for comparison of Pass Manchac, Reserve Canal, and the Blind River, respectively. In all cases tcrit was 2.07.

Table 4. Comparison of phosphate concentrations to historical data

3- Site n [PO4 ] (ppmP) Range (ppmP)

Lake Maurepas 108 0.14±0.01 0.04-0.42 Inlet Sites 48 0.16±0.02 0.07-0.42 Outlet Sites 24 0.12±0.02 0.04-0.26 Pass Manchac 12 0.12±0.04 0.04-0.26

Our study Reserve Canal 12 0.19±0.05 0.13-0.42 Blind River 12 0.18±0.04 0.09-0.26 Mid-Lake 36 0.13±0.01 0.05-0.22

Mid-Lake 137 0.14±0.03 0.01-1.70 USGS

Pass Manchac 12 0.07±0.03 0.00-0.15 Reserve Canal 12 0.06±0.03 0.00-0.13

Day et al. Blind River 12 0.06±0.03 0.00-0.16

The concentration of silicate was also higher at corresponding locations in our study than in the study by Day et al. In our study, the average concentration of silicate was 2- 2- 2- 7.4±0.9ppmSiO3 , 8.9±1.3ppmSiO3 , and 8.8±1.2ppmSiO3 at Pass Manchac, Reserve Canal, and the Blind River, respectively. In the study by Day et al., the concentrations were 2- 2- 2- 4.3±1.5ppmSiO3 , 4.8±1.6ppmSiO3 , and 5.3±1.4ppmSiO3 . In general, the concentration of silicate fluctuated significantly from one sampling date to the next in both our study and in the study by Day et al., resulting in a wide range of concentrations at each site over time. Despite this wide range, the higher average silicate concentrations observed in our study are statistically different from those observed by Day et al. with texp values of 3.48, 4.07, and 3.87 for data associated with Pass Manchac, Reserve Canal, and the Blind River, respectively. The critical t-value is 2.10 in all three cases.

96 PBRP Annual Report | 2009 Determining the Potential for Algal Bloom in Lake Maurepas

Table 5. Comparison of silicate concentrations to historical data

2- 2- Site n [SiO3 ] (ppm) Range [SiO3 ] (ppm) Lake Maurepas 72 7.9±0.3 3.4 – 12.1 Inlet Sites 32 8.5±0.5 6.0 – 12.1 Outlet Sites 40 7.5±0.3 3.4 – 8.6 Mid Lake Sites 24 7.6±0.5 3.3 – 8.6

Our Study Pass Manchac 8 7.4±0.9 5.1 – 8.3 Reserve Canal 8 8.9±1.3 7.4 – 12.1 Blind River 8 8.8±1.2 6.9 – 11.4 Pass Manchac 12 4.3±1.5 0.7 – 7.9 Reserve Canal 12 4.8±1.6 0.3 – 8.8

Day et al. Blind River 12 5.3±1.6 1.2 – 8.7

DISCUSSION: BONNET CARRÉ SPILLWAY STUDY To relieve high waters on the Mississippi River, the Bonnet Carré Spillway was opened from April 11, 2008 until May 8, 2008. During this time an average of 160,000cfs of freshwater was diverted into Lake Pontchartrain. Because nitrate and silicate are important to the growth of phytoplankton and other algae and are expected to cause increased algal growth, we tracked their concentrations. The concentration of nitrate and silicate immediately rose to much higher than normal levels while the concentration of phosphate in the study area, near the spillway’s outflow, declined (fig. 6). While phosphate levels did show some decrease in concentration during the time when increasing chlorophyll a levels were observed, this decrease was much less significant than the initial decrease in phosphate concentration following the opening of the spillway; thus, the decrease in phosphate concentration does not appear to control algal growth. While increased algal growth was visible in Lake Pontchartrain, excessive algal growth did not occur significantly until after the closing of the spillway.

The worst areas were on the northern shore of Lake Pontchartrain. Readily visible algal growth did not occur in the study area near the spillway. However, chemical analysis of water samples taken from the spillway area showed significant increases in chlorophyll a levels in the study area beginning after the closing of the spillway (fig. 6). Figure 7 combines concentrations of chlorophyll a, nitrate, and silicate to demonstrate the relationship between algal growth and nutrient concentration. It is clear from figure 7 that the dramatic increases in nitrate and silicate concentrations lead to the increased algal growth, as evidenced by increasing chlorophyll a levels following the closing of the spillway. It is also clear that as chlorophyll a levels begin to dramatically increase, the nitrate and silicate are being rapidly assimilated through algal growth. When algal growth exceeds the ability of nitrate and silicate to support that rate of growth, chlorophyll a levels begin to decrease precipitously, indicating a decrease in algal population. While increasing silicate levels may have aided in the observed

PBRP Annual Report | 2009 97 Determining the Potential for Algal Bloom in Lake Maurepas algal growth, its average concentration reached a minimum of 1.8ppm on May 27th and then begins to rise, without being fully depleted, by the rapidly growing algae population. On the other hand, nitrate levels decreased to a level below the detection limit on June 2nd, after being fully depleted by the rapidly growing algae, making it the limiting nutrient. After this point, the concentrations of nitrate, silicate, phosphate, ammonia, and chlorophyll a began to return to more normal levels. By the end of the summer, the concentrations of these substances reached levels that are similar to those observed just prior to the opening of the spillway as has the water’s conductivity.

45 12 40 10 35 30 8 ] (ppm) 2- 25 3 6 20 ] & [SiO [Chl a] (ppb) a] [Chl 15 4 - 3 10 2 [NO 5 0 0 4/10/08 5/5/08 5/30/08 6/24/08 7/19/08 8/13/08

[Chl a] [NO3-] [SiO32-]

Figure 7. Comparison of nutrient and chlorophyll a levels

FUTURE DIRECTIONS Plans for slowing coastal deterioration still include regular freshwater diversions into Manchac Swamp through the Hope Canal on the southeastern shore of Lake Maurepas. For this reason continued monitoring of water quality in Lake Maurepas will be useful to evaluate if changes in water quality occur once the diversion is in operation. The results from the study of Lake Pontchartrain make it a reasonable hypothesis that nitrate will be the nutrient that will be of greatest importance in future studies of Lake Maurepas and the Hope Canal Diversion. Preliminary studies completed in this laboratory where water samples from Lake Maurepas were amended with additional phosphate and silicate support this hypothesis in that additional algal growth was not observed in the laboratory. Continued monitoring near in Lake Pontchartrain is also recommended to better understand changes in its water chemistry during and following future openings of the Bonnet Carré Spillway. This monitoring should be expanded geographically to cover a wider portion of Lake Pontchartrain given the observance of high levels of algal growth on the northern shore.

Finally, to have more consistent data for analyzing changes in the water chemistry of Lakes Maurepas and Pontchartrain, it is recommended that greater automation be employed in future studies. At minimum, a discrete analyzer system should be employed for analysis of water samples taken manually by boat from the lakes. This system would enable faster chemical analyses and would produce less chemical waste, thereby “greening” the process. The faster analysis time would allow more samples to be taken and analyzed. Scheduling of significant blocks of time on a regular basis for undergraduate students to collect samples and, during certain parts of the year, significant periods of poor weather, however, will continue to limit the consistent collection of samples. A second option for improving data

98 PBRP Annual Report | 2009 Determining the Potential for Algal Bloom in Lake Maurepas

collection for these studies would be the use of automated analysis using YSI or Hach multiparameter sondes for constant monitoring. To provide significant coverage of the lakes, a relatively large number of sondes would be needed.

TECHNOLOGY TRANSFER The goal of this project was to evaluate the effect of freshwater diversions on the water quality in Lakes Maurepas and Pontchartrain. The concentration of nutrients in Lake Maurepas was monitored and compared to historical data. This additional data now can serve as a baseline for comparing nutrient levels in Lake Maurepas following the Hope Canal Diversion to determine any detrimental effects.

The underlying question of our research, and a question that should underlie future research, was whether fresh water diverted into Manchac Swamp from the Mississippi River through the Hope Canal Diversion was likely to lead to excessive nutrient enrichment in Lake Maurepas that causes excessive algal growth? This study demonstrated that levels of silicate and phosphate are higher in Lake Maurepas than in the past; however, the low levels of chlorophyll a show that these increases have not led to excessive algal bloom. Coupled with the study of freshwater diversion through the Bonnet Carré Spillway, we believe the limited flow of freshwater from the Mississippi River into Manchac Swamp through the Hope Canal is unlikely to lead to excessive algal growth.

We recommend that slightly higher diversion flow rates through the Hope Canal Diversion be considered. Algal growth was clearly observed both in this and other studies in Lake Pontchartrain following the opening of the Bonnet Carré Spillway. However, the Bonnet Carré opening represents a much larger diversion with flow rates averaging 160,000cfs and led to only limited algal bloom. The Bonnet Carré opening had flow rates that are more than 100 times greater than the planned Hope Canal Diversion. We also recommend that nutrient monitoring continue in Lakes Maurepas and Pontchartrain before and after the projected opening of the Hope Canal Diversion and that the monitoring program be much more automated than what was undertaken in this study.

This study included work in Livingston, St. Charles, St. John the Baptist, and Tangipahoa parishes, and water quality issues examined in this study can clearly affect other parishes in the Lake Pontchartrain Basin such as Jefferson, Orleans, and St. Tammany. Other agencies that may be interested in nutrient and chlorophyll levels as determined in this study include the US Army Corp of Engineers, the USGS, the US EPA, the Louisiana Department of Environmental Quality, and the Louisiana Department of Wildlife and Fisheries. Stakeholders interested in results of this study may include recreational and commercial fisherman and crabbers.

The results of this study demonstrate that nitrate is the most important nutrient likely to be of interest in understanding the effects of planned freshwater diversions through projects such as the Hope Canal. These results demonstrate that higher levels of flow may be possible without leading to excessive algal growth and further studies should examine this possibility. We recommend continuing and expanding the water quality monitoring in Lake Maurepas both prior to and after the Hope Canal Diversion begins.

PBRP Annual Report | 2009 99 Information Transfer and Outreach for the Pontchartrain Basin Research Program

Information Transfer and Outreach Program for the Lake Pontchartrain Basin Research Program

Robert Moreau1, Lacy Landrum2, and Tiffany McFalls3 1 Turtle Cove Environmental Research Station; Southeastern Louisiana University 2 Lake Pontchartrain Basin Research Program; Southeastern Louisiana University 3 Department of Biological Sciences; Southeastern Louisiana University

ABSTRACT During the last year, the outreach activities for the Lake Pontchartrain Basin Research Program (PBRP) have continued steadily. The goal of PBRP is to translate the knowledge gained in research activities to both a) decision-makers in the regulating agencies and organizations and b) stakeholders in the basin such as citizens and news media. Ultimately, our goal is to ensure that the program’s research is applied appropriately to the current environmental and ecological issues and, in particular, to the effective management of the Pontchartrain basin minimizing the human impact on the physical environment. This goal ensures reducing environmental and economic damage to the basin, thereby ensuring the health of the basin’s stakeholders and communities.

INTRODUCTION The purpose of the Information Transfer and Outreach program is to ensure that the knowledge gained in the PBRP research activities is disseminated widely to decision-makers and technical professionals in the regulating agencies and organizations as well as to other important stakeholders and the citizens and news media in the Lake Pontchartrain basin. The goal is to ensure that the PBRP research is applied appropriately to the current environmental and ecological issues, and in particular, to the effective management of the Pontchartrain basin. Ultimately, effective information transfer and outreach educates the public and potentially motivates them to advocate for restoring the wetlands and the overall health of the Pontchartrain region to reduce future impact from hurricanes.

PRIMARY OBJECTIVES From 2007 to 2008, the Information Transfer and Outreach program accomplished the following objectives: 1. Implement the Information Transfer and Outreach plan 2. Create and disseminate Research Highlights as PBRP projects completed their findings 3. Create a Basin Update discussing the mitigation potential on the northshore and the use of sewage effluent to improve the basin’s wetlands 4. Compile a list of the program’s research accomplishments since its inception

100 PBRP Annual Report | 2009 Information Transfer and Outreach for the Pontchartrain Basin Research Program

RESULTS The PBRP outreach has continued steadily for the last year. The former assistant director, Thais Perkins, left PBRP in June 2008, and her position remained open for the remainder of the year. Fortunately, Lacy Landrum is now serving as assistant director and will be able to complete and extend many of the outreach activities.

The first two objectives of the Information Transfer and Outreach program align with each other and have been accomplished concurrently. The first objective addresses implementing the outreach plan for PBRP, and the main goal of the Information Transfer and Outreach plan is to disseminate the research findings of PBRP investigators so that the information is timely and relevant to the needs of those living and working in the Pontchartrain basin. Efforts were made to email current Research Highlights to the staff of state policymakers; however, a more substantial database of contacts, including state and federal policymakers along with advocacy organizations, needs to be established. This database needs to be updated and then used effectively for disseminating PBRP’s Research Highlights and Basin Updates.

The second objective involves creating and disseminating Research Updates. These one-page information handouts summarize the research findings of each PBRP project with hopes that the information facilitates further communication among policymakers and stakeholders. The following Research Updates are currently being written: Genetic Variation between Lake Maurepas and Mississippi River Basin Fishes [K. Piller] A Whole-System Approach for Restoring the Wetlands [G. Shaffer] Amphibian and Reptile Monitoring in the Pontchartrain-Maurepas Region [B. Crother] Salinity as a Stressor of the Freshwater Turtle in the Lake Pontchartrain Basin [R. Valverde]

Streaming video of Samuel Hyde’s documentary was also added to the PBRP website. Hyde’s documentary traces the socio-economic and political forces that directly caused the crisis facing the western and northern Pontchartrain areas. By linking human behavior to environmental degradation, Hyde predicts how the crisis will worsen if policymakers fail to curtail human sprawl and advocate wetland restoration.

Creating a Basin Update that addresses the mitigation potential for restoring the wetlands has been a more complicated objective. This Update depends on research findings of the mitigation project, which began late due to the 2005 storms. In lieu of using this information for the Update, we decided to produce an Update about restoring the wetlands as part of the Pontchartrain basin’s “lines of defense,” an argument that the wetlands lessen the effects of hurricane winds and storm surge, meaning they serve as our first line of defense.

PBRP Annual Report | 2009 101 Information Transfer and Outreach for the Pontchartrain Basin Research Program

FUTURE PLANS Future plans for the Information Transfer and Outreach program include redesigning the PBRP website to make it compliant with the accessibility guidelines outlined by the World Wide Web Consortium. As each PBRP project concludes, the research findings will be summarized in Research Updates and disseminated to the contacts in our database of policymakers and advocacy groups. Additionally, the program plans to create webcasts that feature the PIs discussing their research. These webcasts could be downloaded and used in high school or college classrooms, or they could be disseminated to policymakers, so they are informed of the current projects that document and impact the health of the Lake Pontchartrain Basin.

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