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Diatom Communities near Acid Mine Drainage at Green Valley Lake, West Terre Haute, Indiana BRINKMANN, S.A., BRAKE, S.S, & STONE, J.R. Earth and Environmental Systems Indiana State University, Terre Haute, IN 47809 Abstract Methods Collecting Green Valley Lake is located in West Terre Haute, Indiana, near an abandoned coal mine. Acid Mine Drainage (AMD) leaching from the site of the abandoned mine has impacted this area for almost 55 years. Seasonally, the pH of effluent streams drops as low as 3 in some areas. Elevated levels of SO4, Fe, Al, and heavy metals occur in streams (and groundwater) that may contaminate Green Valley Lake. Diatoms are a golden-brown algae with a siliceous skeleton; they are an important primary producers that are abundant and typically well-preserved as fossils in most lake systems. To explore the environmental impact of AMD on this system, we sampled the lake plankton and sediments for fossil diatoms, which are known to be highly sensitive to acidity in lakes and streams. The purpose of this study is to analyze the spatial distribution of diatoms in the lake and sediments. Samples were collected from the plankton and sediment from 11 locations around the lake. Our hypothesis is that diatom diversity should increase away from areas of riverine discharge into the lake. Results from this study will help determine how AMD has influenced the structure of the diatom community in the lake and provide a baseline measurement for the modern lake system so that the long-term resilience of these communities can be analyzed in future studies. Background SITE The reclaimed Green Valley mine site, located in West Terre Haute, Indiana, contains over five million tons of waste material (Brake, Hasiotis & Dannelly, 2004). After site abandonment in 1963, levees surrounding ponds were breached by erosion, leading to uncontrolled movement of contamination from the mine area. Acid mine drainage (AMD) associated with the mine site most likely seeps into Green Valley Lake, via groundwater. AMD forms when sulfide-bearing minerals, such as pyrite, react with the atmosphere and waterways and produce iron- and aluminum- rich sulfuric acid solutions. These solutions contain elevated concentrations of total dissolved solids, iron, aluminum and sulphate. Diatom distribution is influenced by seasonal and intraseasonal changes in water temperature and chemistry, Diatoms are absent in winter due to cool water temperatures. In summer, isolated communities are present due to warmer water temperatures. When this leaching enters natural stream systems, acidity and high concentrations of dissolved constituents can adversely impact aquatic life (Brake, Hasiotis & Dannelly, 2004). RESEARCH Soluble reactive phosphorus, typically in the form of orthophosphate (PO43-), can be a nutrient for aquatic macrophytes and algae. Because phosphorus is typically a limiting nutrient, the presence of differing levels of phosphorus can alter the aquatic community, resulting in a change in the dominant phytoplankton. In lake systems with excessive phosphorus, this may cause health risks or become an aesthetic nuisance to those living near or using the waterways, because high levels of phosphorus may produce cyanobacteria blooms (Wetzel, 2001). Because diatoms are minute and virtually ubiquitous (often occurring in tap water, for example), and their frustules (shells) are resistant to decay, it is very easy to contaminate samples with either living or dead diatoms from elsewhere. Extreme cleanliness was therefore required during sampling. Procedures were followed that prevent contamination from such sources (use of reverse osmosis pure water, for example). Two types of samples were taken: planktonic and benthic. Samples were collected from a rowboat that was anchored at intervals through the lake. At each interval a scoop was sent to the lake floor, along with a measuring device. Planktonic samples were taken using a plankton net, which collects water from the top part of the water and contains anything floating within it. Site coordinates and depth were recorded for each location. The sample locations transect away from the AMD creek input, and around the lake perimeter. The samples were labeled numerically, each site consisted of a sample “A” and “B” representing planktonic and benthic, respectively. Processing It is necessary to remove any organic matter from diatom samples in order to make microscopic identification efficient. The cleaning process also allows unwanted mineral material to be removed and the concentration of diatoms to be adjusted. Since hydrogen peroxide is a powerful oxidizing agent, precautions must be taken to prevent contact with the skin and eyes. Samples are transferred into scintillation vials, where hydrogen peroxide is added and left until all organic material has been removed (this may take several days). Following the processing stage, the samples are put through a water bath, and samples were filled with RO pure water, then were left to settle out overnight between each wash or by centrifugation; this rinsing stage is repeated at least 3 times. The goal is to produce an even spread of diatoms over the cover slip. This process begins by gently heating the coverslips after the diatoms have been allowed to settle. Using pipettes, up to 0.5ml of well mixed diatom suspension is placed on each cover slip, which are then covered to keep off dust. These are left to dry on a heated table. A hotplate is then heated up to 130oC, and 1 drop of Naphrax is placed on a glass slide to drive off the toluene in the Naphrax. Results Background (Continued) Acidity, iron, and aluminum all play a role in the fate of phosphorus in aquatic systems; phosphorus is soluble at low pH, but tends to bind to aluminum and iron, particularly when it is in an oxidized state (ferric iron). Ferric iron particulate matter often dissolves in the bottom waters of most lakes as the lack of oxygen in the water transforms it into a reduced state (ferrous iron), which is soluble in water. In temperate lake systems, where iron and aluminum are present, phosphorus is often transported when it is bound to particulate matter falls through the water column and into the sediment. Under anoxic conditions, it is released and tends to be stored in the colder and denser water layers until mixing events carry it into the surface waters, where it is once again accessible to aquatic plants and algae as soluble reactive phosphorus (Wetzel, 2001). As pH in a lake or stream decreases, aluminum levels increase. In high abundances, aluminum and acid can cause chronic stress, disrupt food webs, and decrease biodiversity for aquatic organisms. Some types of organisms are able to tolerate acidic waters, while others may be acid-sensitive, this change in tolerance is represented in Green Valley lake by presence of Cyclotella meneghiniana and Stephanodiscus ("Effects of acid," 2012). By looking at a Google image of the Green Valley lake area, algal blooms are seen along the northern shore of the pond. This is a response to increased levels of nutrients, a likely cause of this is the farmland that surrounds this basin; farmland is commonly fertilized with nitrogen and phosphorus. Hypertrophication is the ecosystems response to addition of artificial or natural substances such as nitrates and phosphates, through fertilizers or sewage to an aquatic system (Schindler & Vallentyne, 2004). Enhanced growth of aquatic vegetation or phytoplankton and algal blooms disrupts normal functioning of the ecosystem (Bartram, 1999). The combined addition of Fe, Al, and acidic water to the lake system suggests that there is a strong potential for these components to disrupt the "normal" lake cycling of phosphorus. Acid mine drainage entering the lake system through groundwater may result in a change in the water chemistry, binding phosphorus temporarily and storing it in the cold dense bottom waters, until lake mixing releases it. General chemical and physical conditions which currently exist at Green Valley Lake are set forth in the section titled “Graphs”. The presence of Cyclotella and Stephanodiscus diatom genera is useful in this study to understand low nutrient (low phosphorous levels) and high nutrient (high phosphorous levels) systems in Green Valley Lake. The main purpose of this method is treating the data as a whole that way diatom community structure is highlighted and the information is condensed in a simple and interpretable way. The data matrix consisted of samples from 11 sites, one set labeled “A” representing samples from the water column, and one set labeled “B” representing benthic material. Dominant planktonic taxa included Fragilaria, Asterionella (highest abundance at site 6), Cyclotella, Stephanodiscus (relatively abundant throughout sites), and Synedra (highest abundance at site 3). Dominant benthic taxa included Fragilaria, Nitzschia (similar abundances in all samples except 5 and 8), Cyclotella, and Stephanodiscus (with their abundances peaking around sites 5 and 9). Diatom distribution in the sediment reveals a continuous increase of Cyclotella and Stephanodiscus towards the center of Green Valley lake, and a gradual decrease towards the edges of the pond, with the exception of sites nine and ten which are nearest high-nutrient input. The association of diatoms reflects the overall ecological conditions of the water body, it is not always possible to correlate the water quality of a particular area expressed by the diatom association with a chemical analysis of the water samples taken at the same time (Patrick, 1968). The measured pH of the lake water (2013) varied between 8.5-9.0. About 200 diatom frustules were counted from each planktonic and benthic sample site. Representations of Green Valley Lake surface temperatures (°C), surface electrical conductivity (µS), and bathymetry based on depths from each site (ft.). The pie charts surrounding the sample map indicate the relative abundances of planktonic samples (purple and blue charts) and benthic samples (yellow and red charts); it is important to note that it’s the samples (not the genera) that are planktonic/benthic. For simplicity, the genera Epithemia, Cocconeis, Karayevia, Nitzschia, Placoneis, Hantzschia, Eunotia, Achnanthes, Gomphonema, Psammothidium, Cymatopleura, and Navicula have been lumped together as “Benthic ” for the samples from the water column. Graphs Surface Temperatures Menoghin iana 4% Asteroinella 10% Cymbella 2% Menoghinian a 3% Lake Conductivity Nitzschia 14% 5% Fragilaria 30% 15% Synedra 2% Fragilaria 48% 18% 24% 11% 4% 15% 4% 17% 12% 2% 4% 2% 4% 31% 14% 2% 1% 26% 23% References 14% 44% 3% 12% 11% 4% 36% 4% 2% 2% 1% (n.d) Phosphorous control in water and wastewater. General Chemical, Technical Bulletin:(Wastewater), Retrieved from http://www.generalchemical.com/assets/pdf/Phosphorus_Control_in_Water_and_Wastewater.pdf 4% Turner B.L et al. (2003) Organic phosphorus in the environment. CABI publishing 7% 6% 1% 5% 25% 9% 1% 17% 13% 15% 1% 3% 2% 13% 34% 2% 17% 44% 7% 5% 13% 2% 52% 4% 2% 29% 2% 7% 22% 34% 13% 2% 2% 5% 26% 4% 6% 17% 7% 13% 14% 15% 10% 6% 18% 10% 3% 3% 14% 25% 21% Cyclotella 15% 4% 23% 24% Discostell a 1% 8% 1% 1% Benthic 22% Synedra Stephano Benthic; 28% 6% discus 12% Discostella 0% Stephanodisc us 0% Cyclotella 3% Bathymetry Likewise for the “Benthic” genera, Epithemia, Encyonema, Cymatopleura, Cocconeis, Epithemiales, Karayevia, Eunotia, Gomphonema, Placoneis, and Achnanthes are grouped together for the samples from the sediment. Diatoms react to the sum of total stresses, pollutant(s) may have a more sever effect if other environmental conditions produce stress than if this does not occur. As shown by Patrick (1968), that when the pH of the water of a diatom community, which typically lived in a mean of 7.56, was lowered to a mean pH of 5.3 in May/June when the day length and temperature were favorable for diatom growth. This lowering of pH did not cause a severe shift in the number and relative abundance of the species present. From this it was gathered that it’s not only reduction of pH that caused unfavorable effects on diatom communities, but rather this change accompanied by other environmental stresses. The ability for species of diatoms to live under conditions of concentrated organic pollutants may be because they can utilize the nutrients that are present such as amino acids and other organic forms of nitrogen; in other cases, it may be because they are able to enclose the excessive concentration of heavy metals in a vacuole or to change their chemical rate so that they do not interfere with the metabolism of the individual (Patrick, 1968). Examples of diatoms that can flourish in the presence of heavy metals include Stephanodiscus, Achnanthes, Eunotia, and Synedra; Nitzschia being an excellent indicator of increased nutrients in a body of water. Possible sources of error are associated with different sampling sites and different amounts of sample collected, primarily with the benthic samples. These species are all benthic, even though they have been found in the plankton. 1% 10% 13% 18% 42% 32% 10% 17% 6% 18% 25% Bortleson, G. C., & Lee, G. F. (1974). Phosphorous, iron, and manganese distribution in sediment cores of six wisconsin lakes. (U.S. Geological Survey, University of Texas-Dalls, Richardson) 24% 7% 12% 38% 10% 35% 6% 1% 9% 16% 11% 5% 5% 12% 11% 14% 8% 32% 13% 16% 13% Carritt, D.E. and S. Goodall. 1954. Sorption reactions and some ecological implications. Deep-sea Res. I:224-243 2% 23% Stumm, W., and J.J. Morgan. 1970. Aquatic chemistry. Wiley-Interscience. 4% 17% 4% Wetzel (2001) Limnology: Lake and River Ecosystems (3rd Edition), Academic Press, 1006 pages. EPA, (2012). Effects of acid rain - surface waters and aquatic animals . Retrieved from website: http://www.epa.gov/acidrain/effects/surface_water.html 7% Bartram, J., Wayne W. Carmichael, Ingrid Chorus, Gary Jones, and Olav M. Skulberg (1999) Chapter 1. Introduction, in: Toxic Cyanobacteria in Water: A guide to their public health consequences, monitoring and management. World Health Organization. URL: WHO document (http://www.who.int /water_sanitation_health/resourcesquality/toxicyanbact/en/) 15% 23% 2% 2% 2% 6% 2% 56% 1% 1% 30% 6% 47% 3% 4% 9% 13% 2% 14% 14% Carpenter, S.R., N.F. Caraco, and V.H. Smith. 1998. Nonpoint pollution of surface waters with phosphorus and nitrogen. Ecological Applications 8:559–568 11% 6% 5% 11% 49% Schindler, David and Vallentyne, John R. (2004) Over fertilization of the World's Freshwaters and Estuaries, University of Alberta Press, p. 1, ISBN 0888644841 40% 1% 14% 4% Brake, S. S., Hasiotis, S. T., & Dannelly, H. K. (2004). Diatoms in acid mine drainage and their role in the formation of iron-rich stromatolites. Geomicrobiology Journal, 21, 331-340. Patrick, R. (1968). Diatoms as indicators of changes in water quality. Academy of Natural Sciences, Pliladelphia, .