Assessing potential for spread and impacts of Eurasian watermilfoil Myriophyllum spicatum) in Lake Tahoe using in situ transplants, microcosms, and bioassays Katey M. Walter, Department of Environmental Studies and Policy, UC Davis, 95616 Dr. Lars W. Anderson, USDA-ARS Exotic and Invasive Weed Research Lab, UC Davis, 95616 Dr. Charles R. Goldman, Department of Environmental Studies and Policy, UC Davis, 95616   Introduction This project was conducted to determine the ecosystem effects of the invasion of Eurasian watermilfoil (Myriophyllum spicatum) at Lake Tahoe and compare these effects to the native plant, Elodea canadensis. Submersed macrophytes, such as these, have been shown to link littoral sediments with the overlying water column by releasing nutrients, such as nitrogen and phosphorus, that contribute to the process of eutrophication (Barko et al., 1991, Jackson et al., 1994, James et al, 1996, Barko and James, 1998). The majority of these studies have been conducted in mesotrophic and eutrophic systems, leaving a paucity of research in ultra-oligotrophic systems, such as Lake Tahoe. The invasion of M. spicatum in Lake Tahoe is of great concern to those that value the uniquely pristine ultra-oligotrophic environment because of its potential to impair water quality and change the sensitive nutrient cycles of the littoral ecosystem. Objectives Monitor the occurrence and spread of M. spicatum around Lake Tahoe. Estimate the potential for infestation of new areas around the lake Determine relative threates of M. spicatum and E. canadensis to lake water quality through the release of nutrients and enhancement of algae growth.   Approach and Methods Lake Surveys (Objective 1) We conducted aerial, boat, and SCUBA surveys to monitor the current distribution of M. spicatum populations around Lake Tahoe (Fig. 1). We established 35-meter transects across plant communities at four locations: Meeks Bay, Emerald Bay, Crystal Bay Marina, and Obexer’s Marina. Along each transect, 10 permanent sampling sites were surveyed four times over the plant’s growing season (July, August, September, Oct-November, 1999) to monitor changes in density of M. spicatum.   Reciprocal transplant (Objective 2) Apical meristems of M. spicatum (20-cm length) were collected from Cove East Lagoon and Meeks Bay Marina on August 21, 1999 and planted into (1-gallon) acid-washed plastic containers filled with sediment from various Lake Tahoe sites. We used the following reciprocal transplant design to combine sources of sediment, sources of plants, and growth locations:                             Five cuttings with apical tips were placed in a container, and each treatment (Site-plant source-sediment source) was replicated four times. Containers were placed on the bottom of the lake at depths of 10-12 feet. Following nine weeks of growth, containers were removed from the transplant sites on October 23, 1999. Plant survivorship and growth were determined (% survivorship and plant height).   32P cycling in hydroponic microcosms (Objective 3) Apical meristems of M. spicatum (20cm, without roots) and E. canadensis (with a few roots) were planted in containers of sediment in the Tahoe Keys East Cove Lagoon on August 24, 1999 and left to establish roots for 12 days prior to transfer to laboratory microcosm tubes. Plants were grown hydroponically in filtered lake water microcosms for 45 days. Glass root compartments held 135 ml of filtered lake water with a 10 m Ci addition of carrier free 32P-PO34-. Microcosm treatments consisted of one plant per tube: M. spicatum (32P, n=10), E. canadensis (32P, n=9), M. spicatum (no 32P, n=10), and filtered water (no plants, no 32P, n=6). Treatments were split randomly into two separate growth chambers with long (14 hr)- and short-day (10hr) photoperiods. Temperature and light levels were maintained equally in both chambers at ~18° C and 275 m mol respectively. We stirred water columns and sampled 1-ml for liquid scintillation counting (LSC) 36 days of the 45-day experimental period (September 9, 1999 to October 20, 1999) to monitor leakage of 32PO34- from plant shoots during growth and senescence. The activity in root compartments was sampled at the start of the experiment (September 9, 1999), on September 15, 1999, and at the end of the experiment (October 20, 1999). We filtered suspended particulate matter in the water columns on the final day of the experiment to determine how much 32P was associated with plankton and detritus. Outdoor sediment-plant microcosms and 14C phytoplankton bioassay (Objective 3) We established microcosms of M. spicatum (n=5) and E. canadensis (n=5) in clear, Plexiglas tubes (1.5 liters) using lake sediment from the Tahoe Keys and unfiltered lake water from near Tahoe City (see map) on September 24, 1999. Two types of controls without plants included (1) tubes with lake sediment and unfiltered lake water (n=5), and (2) tubes with unfiltered lake water alone (n=4). Initial sediment and water were sample from microcosms for nutrient and chlorophyll-a analyses. Following a five-week growth period outside, under natural photoperiod and temperature regime, sediments and water were collected from individual microcosms for nutrient and chlorophyll-a analyses. Water from microcosms was composited by treatment and filtered through HA MilliporeÒ membrane filters for use in a bioassay to test the response of natural phytoplankton populations to filtered exudates from microcosms treatments. A small amount of radioactive tracer 14C-NaHCO3 was mixed into lake water containing natural phytoplankton we added filtered water containing exudates from the sediment-plant microcosms in concentrations of 1% and 10% to bioassay flasks. An additional set of control flasks included deionized water (DI) at 1% and 10% concentration. All treatments were in triplicate, except the DI controls. The flasks were placed in a laboratory incubator under fluorescent lighting (275 m mol) at ambient lake temperature (15ºC) and day length (10 hours). The growth response of phytoplankton was measured every two days over a 6-day period according to in vivo fluorescence and 14C radioactivity accumulated in phytoplankton filtered onto HA MilliporeÒ membrane filters (Goldman, 1968). Results and Discussion Reciprocal transplant (Objective 2) Myriophyllum spicatum grew successfully at every site and in each type of sediment, except at Boatworks Marina under extreme wave conditions (Fig. 2). Survivorship was highest when plants from a particular location were grown at that location. Crayfish preferentially grazed foreign transplants. Myriophyllum spicatum grew taller at the Tahoe Keys, the greatest source of M. spicatum at Lake Tahoe, than at any of the other lake sites. At every site, M. spicatum grew taller in sediment from the Tahoe Keys than in sediment from any other source. 32P cycling in hydroponic microcosms (Objective 3) At the end of the experiment, up to 0.45% of the original amount of 32P introduced into sealed root compartments of M. spicatum was released into the water columns, while less than 0.05 % was released into the water columns of E. canadensis microcosms (Fig. 3). Photoperiod did not affect the release of 32P. 32P activities in the water of root compartments were greater in jars containing M. spicatum than E. canadensis on the mid-experiment sampling date (September 15, 1999) (t=3.613, p = .0010), suggesting greater uptake of 32P by E. canadensis plants. 32P associated with suspended particulate matter (SPM) in water columns was not significantly greater in M. spicatum microcosms than E. canadensis (ANOVA, F = 3.64111,18 , p = .0725). These results suggest that regardless of photoperiod, the invasive macrophyte, M. spicatum, releases phosphorus into the water column during growth and senescence to a greater extent than the native plant, E. canadensis, thereby contributing a nutrient source for algal growth. Outdoor sediment-plant microcosms and 14C Phytoplankton Bioassay (Objective 3) Chlorophyll-a was greater in outdoor microcosms with M. spicatum than in the initial lake water, E. canadensis microcosms, and control microcosms. Total phosphorus was greater in the water of microcosms containing M. spicatum than in microcosms with E. canadensis, sediment, and lake water. NO3 concentrations were greater in the initial unfiltered lake water and in microcosms with M. spicatum, than in microcosms with E. canadensis, sediment or lake water. There was not enough information to distinguish effects of microcosm treatments on water nutrients, SRP, NH4, and DP, or on sediment nutrients, TKN and Olsen-P. In all bioassay treatment flasks, in vivo chlorophyll measurements and 14C counts on HA Millipore membrane filters indicated that phytoplankton populations increased over the six-day incubation period. At the 10% filtered exudate dilution level, the M. spicatum treatment yielded higher in vivo fluorescence in bioassay flasks than E. canadensis and the control treatments. Similarly, E. canadensis exudates enhanced phytoplankton productivity as measured by in vivo fluorescence more than the control treatments. Phytoplankton productivity measured by 14C-uptake was greater in flasks with M. spicatum exudates than flasks with E. canadensis exudates. Exudates from both M. spicatum and E. canadensis enhanced phytoplankton growth more than the no-plant controls (sediment, lake water, and deionized water). It is possible that high TP and NO3 levels from the sediment-plant microcosms were responsible for increased phytoplankton productivity. However, future studies should consider more detailed chemical analyses of plant exudates including dissolved organic carbon and secondary compounds (Wetzel, 1975, Gross et al., 1996, Wetzel and Sondergaard, 1998). These results suggest that the invasive plant, M. spicatum may have a more negative affect on water quality at Lake Tahoe than E. canadensis.     Conclusions M. spicatum populations are expanding around Lake Tahoe and have a potential to establish in protected sites where it is not yet present. Despite the greater capacity of E. canadensis to acquire 32P in its tissues, release of 32P into the water columns appears to have been greater in microcosms with M. spicatum than microcosms with E. canadensis. Although results were marginally significant, we detected higher levels of 32P in suspended particulate matter of the hydroponic M. spicatum microcosms, suggesting some uptake of 32P by phytoplankton and bacterioplankton in the water column. Similarly, the outdoor sediment-plant microcosms demonstrated higher chlorophyll-a levels in water where M. spicatum grew than in water with E. canadensis or control treatments without plants. Furthermore, filtered exudates from M. spicatum microcosms enhanced productivity of mid-lake natural phytoplankton in 14C bioassays more than E. canadensis or filtered sediment and lake water from control microcosms. Both the in vivo chlorophyll-a and 14C-uptake responses of natural phytoplankton productivity in this bioassay support the hypothesis that M. spicatum accelerates growth of phytoplankton at Lake Tahoe more than the native plant, E. canadensis. Results of this study suggest that in late summer, the invasive macrophyte, M. spicatum, contributes to a loss of water quality in ultra-oligotrophic Lake Tahoe to a greater extent than the native plant, E. canadensis, by releasing nutrients into the water column and enhancing the productivity of phytoplankton.   References Barko, J.W., D. Gunnison, and S.R. Carpenter. 1991. Sediment interactions with submerged macrophyte growth and community dynamics. Aquatic Botany, 41:41-65. Barko, J.W., and James, W.F. 1998. Effects of submerged aquatic macrophytes on nutrient dynamics, sedimentation, and resuspension. In The Structuring Role of Submerged Macrophytes in Lakes. 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