Microbial Assessment of Thermal Impacts of Dworshak Reservoir Releases A. Abstract The primary purpose of this project is to develop a methodology to evaluate the impacts of cold water releases from Dworshak Reservoir using innovative microbial assessment techniques. The hypothesis is that incomplete mixing of cold water from the reservoir negatively impacts steelhead and Chinook rearing habitat in the lower Clearwater. A robust, cost-effective methodology is needed to evaluate potential impacts caused by management decisions regarding releases. The project involves collecting and analyzing microbial populations and water temperature data on the Clearwater River downstream of Dworshak Reservoir during summer release periods of July, August and September. Similarities and differences in habitat characteristics are evident at the most fundamental level; the microbial community. The information would be used in a numerical model to evaluate the effects of reservoir releases as a function of total mainstem flow and ambient water temperatures on the Clearwater upstream of its confluence with the North Fork of the Clearwater. The results could be used to formulate guidelines for releases or demonstrate the need for channel improvement structures to enhance mixing at the confluence in Orofino. These results could also be used to help understand the system-wide impacts of altering thermal regimes. Microbial populations would be sampled from surface and near surface sediments at ten crosssections along the lower Clearwater and at one location in the tailrace of Dworshak Reservoir and one location upstream of the North Fork-Clearwater confluence. At each cross-section, three samples will be taken representing i) near right-bank, ii) near left-bank, and iii) near midchannel. The samples will be taken twice a month during the months of July, August and September. Four replicate samples will be taken at one cross-section to demonstrate repeatability and variability of the method. Temperature probes will be operated at these locations during the same period so that comparisons to temperature regimes can be made. Researchers from Washington State University (Dr. Jeremy Rentz and Dr. Michael Barber) will be responsible for conducting the work with the assistance of appropriate graduate level students and technicians. Samples will be collected in the Clearwater River basin and transported to the Civil and Environmental Engineering laboratories in Pullman, Washington for analysis. Finally, cost-effective solutions for uniformly appraising the impacts of habitat and water quality projects throughout the Columbia River basin are needed to determine which projects are having the most significant impact. If successful, this microbial assessment methodology could be used to evaluate the effectiveness of all habitat restoration projects in the basin. B. Scientific Background Cold water releases from Dworshak Reservoir are necessary to partially mitigate downstream summer temperature regimes on the Lower Snake River. However, these releases can significantly alter the natural temperatures in the lower Clearwater River below the confluence with the North Fork Clearwater tributary. This problem is controlled by the ratio of unregulated discharges in the Clearwater versus Dworshak releases and is exasperated by the fact that 1 complete mixing does not occur immediately at the confluence. Aerial images of the study area show the tendency of tributary sediment plumes to persist for considerable distances downstream similar to the evidence indicating complete mixing of the Clearwater and Snake Rivers does not occur until the water is well into the Lower Granite pool. While cool temperatures are generally preferable to warm water, excessively cold water can adversely impact rearing habitat by modifying the food supply and slowing the development of fry and smolt growth rates. This has been shown to negatively impact the survival rate of migrating juvenile steelhead and Chinook salmonids in direct opposition to the management goals of the Council’s Fish and Wildlife Program. Traditional methods of habitat assessment, such as macroinvertebrate investigations and temperature profiling of the substrate material, are plagued with difficulties related to the measuring in deep pools, quantifying natural variability across seasonal variations in life-stage of indicator organisms, and overall cost. What is needed, not only for this watershed but for all habitat restoration projects, is a rapid, cost-effective, reliable method for evaluating the impact of management decisions. This innovative method proposes to examine the use of microbial population distribution and diversity as direct measures of environmental disturbance. This project could have profound implications to the operation of Dworshak Reservoir and thus the flows and temperatures in all four lower Snake River dam pools. C. Rationale and Significance The Northwest Power and Conservation Council recognizes the negative impact hydrosystem operation can have on fish populations, specifically with respect to water temperature (page 25 of 2000 Fish & Wildlife Program). Within the Clearwater Sub-basin, the focus of this proposal, understanding impacts associated with water temperature amelioration using Dworshak Reservoir waters is a primary goal. Thus, the rationale for our proposed study is the recognition that understanding thermal impacts of Dworshak Reservoir releases will contribute directly to goals set by the NPCC and the Clearwater Sub-basin Plan. Our project is significant because it will lead to specific fish impacts and to the development of an assessment tool that can be used throughout the Columbia River Basin. Clearwater River steelhead will benefit from the studies through the development of new Dworshak Reservoir management plans. These plans will also mitigate high temperatures on the lower Snake River, which benefits Chinook. Microbial community analysis developed during this project will be transferable to other reservoirs in the basin that significantly affect water temperature and to habitat restoration projects that comprise much of the work funded by the BPA. As a rapid, costeffective method, microbial community analysis could be easily implemented to evaluate impacts associated with management decisions. D. Relationship to other projects This project is directly related to a number of projects on the Snake and Clearwater Rivers related to restoration and protection of instream habitat. In particular, it relates to the considerable efforts being expended on Dworshak Reservoir because of its status as a major storage reservoir on the system. For example, the Nez Perce Tribe has been working on 2 examining the impacts of Dworshak Reservoir releases on productivity within the reservoir. The Idaho Fish and Game, in cooperation with the US Army Corps of Engineers, is examining how artificial fertilization might improve reservoir fisheries. The US Army Corps of Engineers has been studying how to minimize reservoir spill because of TDG concerns and multiple agencies have been involved in evaluating the use of Dworshak in improving downstream water temperatures on the lower Snake River. While tied directly to existing concerns, this project is envisioned as an independent effort. No immediate collaborations are necessary to successfully demonstrate the feasibility of the proposed methodology. However, over the course of the work, it is hoped that other groups can be brought in to provide input into the study. This project has potentially broad impacts throughout the watershed. First, when combined with other objectives and constraints, it could help define a new management strategy for releases from Dworshak Reservoir. It would clearly demonstrate the positive or negative impacts of modified water temperatures in the lower Clearwater. Second, the proposed microbial fingerprinting methodology could be used to help define the effectiveness of habitat restoration projects well beyond the current proposed project. If successful, it could become a cost-effective means of rapidly assessing disturbance impacts such as new logging roads. This would significantly enhance the monitoring and evaluation efforts required by the Council. E. Proposal objectives, work elements, methods, and monitoring and evaluation As previously stated, the overall goal of this project is to investigate the impacts of Dworshak Reservoir releases on the lower Clearwater River using an innovative microbial fingerprinting technique. In order to reach this goal, the specific objectives of this proposed study are to: 1. Determine the extent that Dworshak Reservoir releases in July, August and September impact habitat function in the lower Clearwater, and 2. Evaluate the effectiveness of using microbial populations as a measure of disturbance. In order to complete these objectives, the following work elements will be completed: 1. Complete reconnaissance temperature survey profile to identify locations for temperature loggers and microbial sampling stations, 2. Install and monitor temperature arrays, 3. Collect and analyze microbial populations from surface sediments, 4. Develop numerical model to evaluate affects of management decisions regarding the ratios and absolute flows between Clearwater and Dworshak Reservoir releases, 5. Statistically analyze combined relationships between water temperature and microbial populations, and 6. Demonstrate management implications on operation of reservoir. 3 Detailed descriptions of these work elements are provided below. It should be noted that since the two objectives are related, some of the work elements will be used to satisfy both objectives. For instance, Objective 1 will include work elements 1, 2, 4, 5, and 6 while Objective 2 will involve work elements 1, 3, 5 and 6. Work Element No. 1 The first step of the project will be to complete a reconnaissance temperature survey profile to identify locations for temperature loggers and microbial sampling stations. Factors such as channel geometry, islands, and tributary inflow all act to influence the mixing of water in the lower Clearwater. Visual inspection of the study reach will be required to evaluate the specific influence of these factors. In addition, temperature profiles (horizontal and vertical) will be collected using a hydrolab or similar device. Substrate material will also be examined to determine uniformity of bed material between sites. Based on this information, researchers will use a global positioning system (GPS) and maps to identify ten (10) cross-sections on the lower Clearwater having characteristics representative of the river reach. Complete profiles of channel geometry will be collected at those sites as well as additional sites near tributary inflows. Other sites may also be surveyed depending on the results of the reconnaissance trip to insure sufficient input data for the temperature model (see Work Element 4). Two additional sites, one on the North Fork of the Clearwater immediately downstream of the dam and one upstream of the confluence will be used as control sites. Work Element No. 2 Once these sites are identified, temperature probes will be installed near the bottom substrate. At each cross-section, three probes will be installed to determine near right bank, near left bank and near mid channel. A total of 30 probes will be used in the mainstem study reach. Additional probes will be used at the upstream control sites with three probes anticipated for the Clearwater cross-section upstream of Orofino and up to three probes for the cross-section downstream of the reservoir depending on the distribution of temperatures in that short reach. The probes will be installed in late June and operated continuously through the end of September. The loggers will be programmed to collect temperature data at ½ hour intervals. Data will be downloaded bi-monthly during July, August and September during the six field trips that will sample microbial populations (see Work Element 3) in order to insure that data loss due to malfunctioning equipment will be minimized. GIS maps of maximum, minimum, and average bi-weekly measured stream temperatures will be generated for visual comparison. Initially, the assumption that variations between stations are linear unless directly influenced by tributary inflows such as those from the Potlatch and Lapwei River basins will be used to present measured data. This assumption will be examined further in the modeling task. Traditional time-dependent plots of temperature and 7-day min/max averages will also be produced. Work Element No. 3 Surficial sediment samples will be collected from locations adjacent to water temperature monitoring stations at bi-monthly intervals during July, August and September. Microbial 4 community fingerprints will be determined for each sediment sample using the culture independent technique terminal-Restriction Fragment Length Polymorphism (Moyer et al., 1996). The analysis requires extraction of microbial DNA from sediments, which will be used as a template to amplify a characteristic gene from all bacteria present in the sample. Genes from different bacterial types will then be separated chromatographically to yield microbial community fingerprints. Microbial community fingerprints generated using t-RFLP can be examined several ways (Brito et al., 2006; Engebretson and Moyer, 2003). First, plots will be visually inspected to observe differences in microbial community structure. Distribution and relative abundance of microbial types will be evaluated in this manner. Microbial diversity will also be determined by summing the different microbial types present within a sediment sample. It is difficult to predict whether diversity for pristine environments will be higher or lower than polluted environments, but differences will be observed (Nocker et al., 2007). Work Element No. 4 A two-dimensional depth averaged numerical model of the lower Clearwater River will be developed based on information on channel geometry collected during the reconnaissance study, aerial photographs, and maps. FLOW2D will be used as the base model. The model will be calibrated to the temperature data collected and information regarding releases and stream flow from the USACE and the USGS. A portion of the measured data will be used to run the model in verification mode. Once calibration and validation are complete, potential operating scenarios for reservoir releases will be evaluated. The spatial distribution of temperature as a function of the ratio between mainstem flows upstream of the confluence and reservoir releases will be thoroughly investigated. Management options, including channel modifications to increase mixing will be examined. Work Element No. 5 The results from Work Element 3 will yield spatial and temporal variations in microbial communities at 30 locations in the lower Clearwater River. This task will statistically analyze the combined relationships between sediment microbial populations and water temperature using analysis of variance (ANOVA) methods. Microbial community parameters to be considered include distribution and relative abundance of microbial types and overall community diversity. The hypothesis is that a cause/effect relationship can be identified and used to predict the impact of management decisions. Work Element No. 6 Using indexes of microbial populations and the temperature results, we will examine the implications of release patterns on the Clearwater River. Comparison of the projected changes will be used with known information regarding the growth rates of steelhead as a function of temperature to demonstrate the potential impact of release schedules compared to reservoir inflow and upstream discharges in the Clearwater. The goal will be to end up with the same reservoir pool elevation on October 1. Ultimately, this will be one more parameter to factor into the operation of the reservoir. It is not the purpose of this work to suggest it is the only parameter 5 that should be considered. The downstream impacts below the Snake-Clearwater confluence will not be adequately addressed in this study. Nor does the investigation attempt to analyze the impact on Kokanee in the reservoir. Critical Assumptions An underlying critical assumption of the proposed project is that microbial communities are directly influenced by water temperatures and that population differences can be uniquely identified with sufficient certainty to permit definitive cause-effect relationships. While we know the first part of this is true, temperature do impact microbial development, variability in a natural environment has not been studied adequately enough to guarantee the latter. Monitoring and Evaluations Interpretation of results will be subject to rigorous statistical analyses. These analyses will be verified by consulting with the Statistics Department at Washington State University, a service routinely provided by the department. In situ measurements of temperature during field trips will be compared to recorded values to insure that probes are operating correctly. Four microbial community fingerprints will be determined for three sampling locations to ascertain the inherent variability of sample results. A final report will be submitted documenting the procedures, data, results and conclusions of this work. We plan on presenting the findings to the US Army Corp of Engineers office in Walla Walla, Washington at which time representatives from NOAA fisheries will also be invited. In addition, we anticipate presenting the results at a minimum of one regional and one national conference and submitting a peer-reviewed journal paper to help disseminate the findings. F. Facilities and equipment Washington State University is a major research institution in the region. As a result, it is fully equipped with routine field equipment, motor pool vehicles, and laboratory facilities. In addition, the Department of Civil and Environmental Engineering has modern computer facilities that will be used to process and analyze data and run the numerical model. Dr. Rentz occupies a new microbiology laboratory (~900 sq ft) outfitted to complete the project, and he also has access 10,000 sq ft of laboratory space controlled by the Environmental Research Center (ERC). Project funds will be used to purchase temperature probes, a thermal cycler and miscellaneous supplies related to installation of the probes. In addition, disposable laboratory supplies related to collection and processing of microbial samples will be needed. The WSU DNA core will be used for t-RFLP analysis. 6 G. Literature Cited Brito,E.M., Guyoneaud,R., Goni-Urriza,M., Ranchou-Peyruse,A., Verbaere,A., Crapez,M.A. et al. (2006) Characterization of hydrocarbonoclastic bacterial communities from mangrove sediments in Guanabara Bay, Brazil. Res. Microbiol. 157: 752-762. Engebretson,J.J. and Moyer,C.L. (2003) Fidelity of select restriction endonucleases in determining microbial diversity by terminal-restriction fragment length polymorphism. Appl. Environ. Microbiol. 69: 4823-4829. Moyer,C.L., Tiedje,J.M., Dobbs,F.C., and Karl,D.M. (1996) A computer-simulated restriction fragment length polymorphism analysis of bacterial small-subunit rRNA genes: efficacy of selected tetrameric restriction enzymes for studies of microbial diversity in nature. Appl. Environ. Microbiol. 62: 2501-2507. Nocker,A., Lepo,J.E., Martin,L.L., and Snyder,R.A. (2007) Response of Estuarine Biofilm Microbial Community Development to Changes in Dissolved Oxygen and Nutrient Concentrations. Microb Ecol. H. Key Personnel The project will be conducted by Dr. Jeremy Rentz and Dr. Michael Barber. Each researcher will contribute 348 hours to the project over the 18 month period. This is equivalent to approximately two person-months per investigator. Dr. Rentz will be responsible for designing the experimental work related to the microbial sampling and analysis. As indicated in the attached resume, Jeremy’s graduate and post-doctoral research in environmental microbiology provided him with the skills necessary to conduct this portion of the work. Dr. Rentz will also be responsible for project management and supervision of the graduate student. Dr. Barber will be responsible for field work related to temperature monitoring and numerical model development. Michael has more than 15 years experience in field data collection and modeling. He will also be involved in the combined analysis of temperature/microbial population data. One full-time Graduate Students will be funded by this project. Responsibilities shared by the students include collecting field samples and completing microbial community analysis. 7 JEREMY A. RENTZ EDUCATION Ph. D., Civil and Environmental Engineering The University of Iowa, Iowa City, IA, 2004 Degradative repression and co-metabolism, plant-microbe interactions affecting polycyclic aromatic hydrocarbon phytoremediation. Jerald L. Schnoor, Thesis advisor B. S., Chemical Engineering The University of Iowa, Iowa City, IA, 1999 PROFESSIONAL EXPERIENCE Assistant Professor Washington State University, Pullman, WA, 2006 – Present 50% Teaching & 50% Research appointment Post Doctoral Fellow Northwestern University, Evanston, IL, 2005 – 2006 Post Doctoral Research Scientist American Type Culture Collection, Manassas, VA, 2004 – 2005 EXPERTISE I conduct general environmental microbiology research that evaluates both natural and engineered systems. Natural systems are investigated using molecular methods to determine the function and structure of microbial communities. Engineered systems focus on biological degradation of polycyclic aromatic hydrocarbons (PAH) and biological removal of phosphorus and heavy metals from water supplies. PUBLICATIONS Rentz, J. A., Alvarez, P. J. J., & Schnoor, J. L. 2007. Benzo[a]pyrene degradation by Sphingomonas yanoikuyae JAR02. Environmental Pollution (In Press). Rentz, J. A., Alvarez, P. J. J., & Schnoor, J. L. 2005. Benzo[a]pyrene co-metabolism in the presence of plant root extracts and exudates: Implications for phytoremediation. Environmental Pollution 136, 477-484. Kamath, R., Rentz J. A., Schnoor, J. L., & Alvarez, P. J. J. 2004. Phytoremediation of hydrocarbon-contaminated soils: principles and applications. In Petroleum Biotechnology: Developments and Perspectives Studies in Surface Science and Catalysis. R. Vazquez-Duhalt and R. Quintero-Ramirez (eds.). Elsevier Science, Oxford, UK. pp. 447-478. Rentz, J. A., Alvarez, P. J. J., & Schnoor, J. L. 2004. Repression of Pseudomonas putida phenanthrene-degrading activity by plant root extracts and exudates. Environmental Microbiology. 6, 574-583. Rentz, J. A., Chapman, B., Alvarez, P. J. J., & Schnoor, J. L. 2003. Stimulation of hybrid poplar growth in petroleum-contaminated soils through oxygen and soil nutrient amendments. International Journal of Phytoremediation. 5, 57-72. 8 MICHAEL ERNEST BARBER State of Washington Water Research Center Department of Civil and Environmental Engineering Washington State University Pullman, Washington 99164-3002 Phone: (509) 335-6633 C Email: meb@wsu.edu Education Institution University of Texas at Austin Purdue University University of New Hampshire Degree Ph.D. MSCE BSCE Year 1991 1983 1981 Employment History From 12/01 to Present, Director - State of Washington Water Research Center & From 8/99 to Present, Associate Professor, Washington State University, Pullman, WA. From 8/94 to 8/99, Assistant Professor, Washington State University, Pullman, WA. From 8/91 to 8/94, Assistant Professor, Tulane University, New Orleans, LA. From 9/88 to 8/91, Research Assistant, University of Texas, Austin, TX. From 6/83 to 7/88, Project Engineer, KKBNA Inc., Wheat Ridge, CO. From 9/81 to 12/82, Teaching Assistant, Purdue University, West Lafayette, IN. Summers 80/81, Civil Engineer, US Army Corps CRREL, Hanover, NH. Professional Registered Professional Engineer in State of Colorado Associate Member of American Society of Civil Engineers Member of American Water Pollution Control Federation Member of American Water Resources Association Member of American Geophysical Union Selected Publications 1. G.N. Teasdale and M.E. Barber, “Aerial Assessment of Ephemeral Gully Erosion from Agricultural Regions in the Pacific Northwest” accepted ASCE Journal of Irrigation and Drainage Engineering, Winter 2005. 2. G. Fu, M.E. Barber and S. Chen, “The Impacts of Climate Change on Regional Hydrological Regimes in the Spokane River Watershed,” accepted ASCE Journal of Hydrologic Engineering - HE/2005/022945, Fall 2005. 3. Guang-Te Wang, S. Chen, M.E. Barber, and D.R.Yonge, (2004). “Modeling Flow and Pollutant Removal of a Wet Detention Pond Treating Stormwater Runoff,” Journal of Environmental Engineering, American Society of Civil Engineers, Vol.130, No. 11, pp 1315-1321. 4. M.E. Barber, S.G. King, D.R. Yonge, and W.E. Hathhorn, (2003). “The Ecology Ditch: A BMP for Stormwater Mitigation,” Journal of Hydrologic Engineering, American Society of Civil Engineers, Vol 8, No. 3, pp 111-122. 5. E. Rowland, R.H. Hotchkiss, and M.E. Barber, (2003). “Predicting Fish Passage Design Flows at Ungaged Streams in Eastern Washington,” Journal of Hydrology, Vol. 273, pp 177-187. 9 10