To address the proposed objectives, STNRD-WMP will work with project collaborators from Loyola University of Chicago (LUC) and the University of Wisconsin-Oshkosh (UWO). Over the last six years, STNRD-WMP has partnered with LUC on work related to invasive species, monitoring of vegetation using HRMI, and initial manoomin restoration work within this system. Past work has spurred current partnerships and resulted in the development of a peer-reviewed article detailing the results of research using HRMI to monitor wetland plant responses to ecological restoration (Lishawa et al. 2017). Partners from LUC bring necessary skillsets related to vegetation management, monitoring using HRMI, and analysis of remote sensing data that fill a gap in Sault Tribe’s current capacity. For this work, we will initiate a new partnership with UWO. Partners at UWO are well-versed in research related to environmental tolerances of aquatic vegetation and specifically those of manoomin (e.g., Pillsbury and McGuire 2009). Partners from UWO bring expertise that will be integral to the success of this project. Construction of Seed Germinating Units (SGUs) The basic experimental unit for Objective 1 will be Seed Germinating Units (SGUs). Part of our work for this proposal will be to develop a replicable design to test seed germination while optimizing: 1) ease of construction and deployment, 2) low cost of construction, and 3) durability. Viable manoomin seeds (quantity based on treatment) will be placed inside flattened mesh cloth bags (50cm X 50cm, 0.5 cm mesh). The mesh bags will be divided into 6 compartments to ensure that the seed is distributed evenly, which is important for density studies. The flattened mesh bags will be fitted to a square PVC pipe frame (50 cm x 50 cm frame, constructed of 0.5 in diameter PVC). When deployed, the PVC frame tubes will be filled with sand and secured with long stakes so that they rest flat against the wetland sediment. This will expose germinating seeds to the chemical conditions of the sediment surface where seeds normally germinate. Roots and shoots can develop normally, and shoots will emerge by growing through the mesh. The SGUs will prevent waterfowl or carp from eating the seeds. This also allows seeds (the most sensitive stage of manoomin development) to be easily retrieved. Deployment locations will be marked using GPS and a pole above water or underwater using metal tags and metal detectors. The response of manoomin to its environment will be measured by: % seed germination, % seedlings at floating leaf stage, % plants at emergent leaf state, and (after harvesting SGUs) root mass (dry weight), shoot mass (dry weight), and root:shoot ratio. To determine the optimal density of manoomin to plant in the St. Marys River, Michigan Determining the optimal density for seeding manoomin is important because seeding efforts can be expensive and planting manoomin at low densities can result in failure due to consumption by wildlife or competition from other macrophytes. Conversely, planting at too high a density can be wasteful. We will construct 30 SGUs utilizing three different densities (n = 10 replicates/treatment) at 80 seeds/m2, 50 seeds/m2 and 25 seeds/m2. All SGUs will be deployed soon after ice out at a uniform depth (60 cm - 80cm). Next to each SGU a procedural control (PC) will also be deployed. The PC will consist only of a PVC frame that will be seeded with the same density of seeds, but seeds will not be protected by the mesh. This will help determine the likely threat of wildlife consumption of seeds. To determine the optimal depth for planting manoomin in the St. Marys River, Michigan Determining the optimal depth for planting manoomin is important for managers since St. Marys River water levels can fluctuate greatly within one season and from year to year. Knowing how manoomin responds specifically to these conditions will inform managers where they should concentrate their efforts. We will construct 30 SGUs and deploy them at three different depths (n= 10 replicates/treatment); 30 cm, 60 cm and 80 cm each with 50 seeds/m2. All SGUs will be deployed soon after ice out. Next to each SGU a procedural control (PC) will also be deployed. Statistical analysis: Data collected will be assessed first by running a series of ANOVAs (analysis of variance) with depth and density as independent factors and response variables such as % germination, % floating leaf plants, % emergent leaf plants, seed/plant production, root mass, shoot mass, and root:shoot ratios. A follow-up two-way ANOVA will then be conducted on the same range of factors but will also include the procedural controls. This will test if (and at what stage) wildlife has a negative impact on planted manoomin. To evaluate chemical and biological tolerances of manoomin in the St. Marys River, Michigan We will further evaluate the environmental tolerances of manoomin in the St Marys by collecting a broad suite of chemical and biological data from the recently planted stands (i.e., Munuscong Bay focal site), at SGU deployment plots, and within at least two well-established manoomin stands in the region. We will characterize the chemical condition of the water at each plot by collecting the following data: water depth, dissolved-oxygen, pH, temperature, turbidity, oxidation reduction potential (ORP), ammonium (NH4), nitrate (NO3), phosphate (PO 4), sulfate (SO4), and chloride (Cl). We will use field installed data loggers (In-situ Rugged Troll 100) to record continuous water depth and water temperature data throughout the growing season at each site. We will use a LUC-owned multi-parameter instrument (YSI Pro-Plus multiparameter probe) to record dissolved oxygen, temperature, turbidity, pH, and ORP. We will collect water samples at each sampling plot and determine ammonium, nitrate, phosphate, sulfate, and chloride content at the University of Michigan Biological Station chemistry lab. In addition, we will collect comprehensive plant community data of all plant species present within each field plot using standard percent cover methods. We will statistically test the correlation between manoomin cover and stem density data with the chemical and biological data collected in the study. With these results, we will attempt to parameterize the chemical and biological tolerances of manoomin in the St. Marys River. Using the resulting data, we will achieve the following four goals: 1. Determine the optimal density of manoomin seed to plant in the St. Marys River, Michigan 2. Determine the optimal depth for planting manoomin in the St. Marys River, Michigan 3. Determine the effects wildlife has on the planting of manoomin across a range of planting densities and depths 4. Assess a novel and inexpensive approach to determine the above goals (1-3) by constructing Seed Germinating Units (SGUs) and explore the standardization of using SGUs to enhance the overall power of manoomin research.