POTENTIAL FOR PONDWEED CONTROL IN LAKE
TAHOE USING BOTTOM BARRIERS
Allison Gamble1†
Thomas Barr2
Brant Allen1
Katie Webb1
John Reuter3 (PI)
Marion Wittmann1† (Co-PI)
Sudeep Chandra4 (Co-PI)
Geoff Schladow5
June 30, 2013
Prepared by:
1
University of California, Davis, Tahoe Environmental
Research Center and John Muir Institute for the
Environment, 291 Country Club Dr., Suite 320 Incline
Village, NV 89451
2
University of California, Davis, Department of Plant
Sciences, One Shields Avenue, Davis, CA 95616
3
University of California, Davis, Tahoe Environmental
Research Center and Department of Environmental Science
and Policy, One Shields Avenue, Davis, CA 95616
4
Department of Natural Resources and Environmental Science,
University of Nevada-Reno, 1000 Valley Road/ MS 186,
Reno, Nevada 89512
5
University of California, Davis, Tahoe Environmental
Research Center and Department of Civil & Environmental
Engineering, One Shields Avenue, Davis, CA 95616
a
Current address is Minnesota Department of Natural
Resources Ecological and Water Division, 50499 Samantha
Lake State Park Road Waterville, MN 56096
b
Current address is Department of Biological Sciences,
University of Notre Dame, Notre Dame, IN 46556
Prepared for:
Tiffany van Huysen, Tahoe Science Program, Pacific
Southwest Research Station, Tahoe Environmental Science
Center, 291 Country Club Dr., Incline Village, NV 89451
Table of Contents
Executive Summary. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . iv
List of Figures. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . vii
List of Tables. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . viii
Acknowledgements. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . ix
1. Introduction. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1
1.1 Background. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1
1.2 Nuisance aquatic macrophytes in Lake Tahoe: A brief overview. . . . . . . . . . 2
1.3 Use of benthic barriers to control aquatic species. . . . . . . . . . . . . . . . . . . . . . 2
1.4 Role of turions in curly-leaf pondweed life cycle. . . . . . . . . . . . . . . . . . . . . . 6
1.5 Scope of work. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .7
2. Methods and Results. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 9
2.1 Objective 1: Profile five sites around the South Shore of Lake Tahoe with
existing curlyleaf pondweed populations. . . . . . . . . . . . . . . 9
2.1.1 Methods. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 9
2.1.1.1 Site location. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 9
2.1.1.2 Surveying the five sites. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 9
2.1.2
2.1.2.1
2.1.2.2
2.1.2.3
2.1.2.4
2.1.2.5
Results and Discussion . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 11
Species composition and diversity, plant density and plant biomass. . . .11
Density and biomass of curly-leaf pondweed. . . . . . . . . . . . . . . . . . . . . .21
Turion density and biomass. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 24
Temperature, water chemistry, sediment and tissue composition. . . . . . 26
Sediment particle size analysis. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .28
2.2 Objective 2: Select three of the profiled sites to deploy three types bottom
barriers over adult curly-leaf pondweed populations and
turions to determine the impact of anoxia. . . . . . . . . . . . . . 32
2.2.1 Methods. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 32
2.2.1.1 Field experimentation and surveys in Lake Tahoe. . . . . . . . . . . . . . . . . . 32
2.2.1.2 Laboratory testing of turion germination. . . . . . . . . . . . . . . . . . . . . . . . . .34
2.2.1.3 Further testing of turion germination following anoxia. . . . . . . . . . . . . . .34
2.2.2
2.2.2.1
2.2.2.2
2.2.2.3
Results and Discussion. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .35
Field Surveys. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 35
Laboratory germination of Lake Tahoe turions. . . . . . . . . . . . . . . . . . . .39
Laboratory germination of turions – bench and microcosm-scale
ii
experiments with turions from the SF Bay-Delta. . . . . . . . . . . . . . . . . . .40
3. Conclusions and Recommendations . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .44
4. References. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .45
Appendix A. Benthic Barriers for Control for Aquatic Weed Propagules
Except from T. Barr Ph.D. dissertation. . . . . . . . . . . . . . . . . . . . . .50
iii
Executive Summary
Potamogenton crispus or curly-leaf pondweed is a relatively recent example of a
nearshore aquatic invasive species in Lake Tahoe. Because of its unique life history
traits, including turion production, this species has competitive advantages over native
aquatic plants. Indeed, curly-leaf pondweed has been called one of the most widely
distributed, nuisance-forming taxon in North America (Crowell 2003, Johnson 2007).
Due to its recent introduction, and restricted range in the southern portion of Lake
Tahoe, curly-leaf pondweed is considered to be in an early invasion stage and thus could
be a potential candidate for eradication or control if a feasible methodology can be
found. The goal of this research was to study turions (vegetative buds) produced by
curly-leaf pondweed to determine their relationship to plant biomass and assess their
susceptibility to non-chemical treatment using gas permeable and impermeable benthic
bottom barriers.
Plant and Turion Density and Biomass
Plant biomass and turion surveys were conducted once during the summer at five sites
along the south shore, on one occasion during the summer, when these vegetative buds
(turions) were abundant. The five sites were not intended to be representative of the
entire south shore nor was the survey done to assess changes in the spatial extent of the
entire P. crispus population. Three of the sites were located in the open-water while two
were associated with marina.
Nine species of aquatic macrophytes were identified during the August survey. Total
plant density, measured as number of stems m-2 ranged from approximately 400-1,330
while total plant biomass ranged from 31-92 g dry wt. m-2. The ratio of total plant
biomass to stem density was fairly consistent in the range of 0.07 to 0.11. Macrophyte
biomass in the present study was within the range of values reported in the literature,
although not as large as that seen in highly infested lakes. Community structure was
evaluated using Simpson’s Index of Diversity (1-D). This Index ranged from 0.42-0.78
in Lake Tahoe with the greatest value seen inside the Tahoe Keys. In general, the
Simpson’s Index of Diversity for macrophytes ranges 0.75->0.90.
Stem density for curly-leaf pondweed was 124-456 stems m-2 while its biomass ranged
from 8.8 to 52.4 g dry wt. m-2, and again, not dissimilar to literature values for other
lakes. Turion density and biomass were not insignificant when compared with other
findings. Turion biomass ranged from 2 to 24 g dry wt. m2. Similar to observations for
turion density, biomass showed differences between marina and outside marina sites.
Despite Lake’s Tahoe ultra-oligotrophic status, it contains significant aquatic plant
community biomass, albeit a limited whole-lake distribution. This was also true for
curly-leaf pondweed and the reproductive turions in the present study. In other words,
macrophyte/curly-leaf pondweed/turion are low in Lake Tahoe when taken as a wholelake average (in contrast to many small lakes that have complete infestation). However,
in those areas of Lake Tahoe where macrophytes exist, their density and biomass can be
relatively high. While specific tests for “physiological-well being” for P. cripsus were
iv
not performed, it appeared as though curly-leaf pondweed in Lake Tahoe is not on the
decline.
Control of Turion Germination
We utilized benthic bottom barriers to potentially control curly-leaf pondweed biomass
by inhibiting turion sprouting. This test was based on recent literature where scientists
found that the sprouting percentage of turions was 68-72 percent lower under highly
anoxic treatment conditions relative to the control. In Lake Tahoe, turions collected from
the Tahoe Keys were placed in mesh bags and placed under jute, polyethylene or rubber
barriers that were deployed in situ. The results of these field experiments at Lake Tahoe
showed that turion sprouting was higher after 8-weeks under the barriers than the
controls. This was an unexpected result thought to be due to the effect of cold-water
temperature on pre-mature sprouting. A second in situ set of trials using barriers and
bags of turions was mobilized in the fall, however, extreme weather and wave conditions
destroyed the experimental set up in the lake.
It appeared that benthic barriers were successful in reducing the existing biomass of
curly leaf pondweed populations, with jute being the most cost-effective and easily
deployed bottom material. Examination of the test sites one-half year and one year after
application of the barriers indicated that pondweed populations did not appear to be
primarily using turions for recolonization of the plots, but were instead spreading via
rhizomes.
Given the uncertainty associated with the turion sprouting experiments in Lake Tahoe as
well as some inconsistency in the literature, we went beyond the scope of work to
further investigate this question using turions collected from the San Francisco BayDelta by performing bench-scale tests and mesocosm experiments. This set-up was
considered applicable to the management question at hand since turions from this
location were available and the tests were designed to evaluate the use of three types of
bottom barriers on turion sprouting. Bench-scale treatments (without sediment) showed
that with the rubber barrier treatment, 48 percent, or less than one-half, of the turions
sprouted relative to 100 percent sprouting in the controls (p ˂ 0.0001). Turions in the
jute treatment had 100 percent sprouting with these was 97 percent sprouting in the
polyethylene treatment.
Mesocosm scale treatments (with sediment) also showed the rubber barrier treatment
had a strong and highly significant effect on sprouting turions of Potamogeton crispus (p
˂ 0.0001). The rubber barriers showed 30 percent sprouting compared to the 98 percent
germination in the controls. The jute benthic bottom barrier treatments had a 72 percent
sprouting rate. Polyethylene showed a 70 percent sprouting. Comparison of treatment
means (p-value of 0.05 or lower) showed rubber benthic bottom treatments where
significantly different from polyethylene, jute and the control. Additionally, means for
jute and polyethylene were significantly different from both the control and the rubber
benthic bottom barrier treatments. Jute and polyethylene benthic bottom barrier
treatments were not significantly different from each other (p = 0.99).
v
Management Implication
In reference to aquatic macrophyte control applied to infested waterbodies in general,
Barr (2013) concluded that benthic bottom barriers alone cannot eradicate 100 percent of
the turions on their own, but that non-porous benthic bottom barriers could possibly be
used in conjunction with other integrated methods for eradication of Potamogeton
crispus turions. Such combined treatments may be able to selectively take advantage of
the anoxic conditions that will enhance efficacy. This study does not recommend the use
herbicide or other toxics for treatment of curly-leaf pondweed in the open waters of
Lake Tahoe. Rather, we provide comments on ‘combined treatments’ to inform future
discussions on this matter should the need arise.
Conclusion
As hypothesized in the literature (Wu et al. 2009), while a standing condition of anoxia
in a water body may be an important mechanism inhibiting sprouting of turions and
growth of curly-leaf pondweed, this appears to be fundamentally different than
establishing a temporary condition of anoxia for the purpose of inhibiting turion
sprouting. Since turions are produced each year, bottom barriers would have to be
installed annually, and then with only a 20-30 percent reduction in sprouting (i.e.
incomplete control/management). We did not find evidence to suggest that once the
barriers were removed that in situ turion germination would not occur in the treated
plots, provided the appropriate environmental conditions were present. Bottom barriers
have been used to control plant growth and biomass in Emerald Bay; however, since
these barriers affect photosynthesis by blocking light, a distinction between material that
are porous or non-porous to dissolved oxygen is not an issue. In contrast the use of nonporous material is essential if the objective is to inhibit turion sprouting.
Recommendation
Based on our experiments we see no reason to recommend the large-scale application of
non-porous bottom barriers for managing existing curly-leaf pondweed populations in
Lake Tahoe that employs control of turion sprouting as a primary mechanism.
vi
List of Figures
Fig. 1-1
Fig. 1-2
Fig. 1-3
Fig. 1-4
Fig. 1-5
Fig. 1-6
Fig. 2-1
Fig. 2-2
Fig. 2-3
Fig. 2-4
Fig. 2-5
Fig. 2-6
Fig. 2-7
Fig. 2-8
Fig. 2-9
Fig. 2-10
Fig. 2-11
Fig. 2-12
Fig. 2-13
Fig. 2-14
Fig. 2-15
Fig. 2-16
Fig. 2-17
Photographs of turions. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1
Distribution of Eurasian watermilfoil (Myriophyllum spicatum) and curly-leaf
pondweed (Potamogeton crispus) in Lake Tahoe, 1995 – 2006. . . . . . . . . . . . . 4
Distribution of curly-leaf pondweed along the south shore of Lake Tahoe,
Including data from 2009. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5
Examining dense growth of curly-leaf pondweed. . . . . . . . . . . . . . . . . . . . . . . . 5
Role of turions in curly-leaf pondweed life cycle. . . . . . . . . . . . . . . . . . . . . . . . .6
Seasonal aspects of curly-leaf pondweed life cycle. . . . . . . . . . . . . . . . . . . . . . . 7
Location of five sampling sites. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .10
Total plant community density and biomass. . . . . . . . . . . . . . . . . . . . . . . . . . . .18
Simpson’s Index of Diversity. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 19
Density and biomass for individual species at each site. . . . . . . . . . . . . . . . . . .21
Relative distribution - density and biomass for individual species. . . . . . . . . . .22
Photograph of curly-leaf pondweed growing in Lake Tahoe. . . . . . . . . . . . . . . 23
Density and biomass for curly-leaf pondweed and turions. . . . . . . . . . . . . . . . .24
Annual cycles of temperature at four biomass collection sites. . . . . . . . . . . . . .26
Nutrient chemistry in macrophyte beds; mid-column and near bottom. . . . . . . 27
Curly-leaf pondweed tissue composition (C, N, P). . . . . . . . . . . . . . . . . . . . . . .30
Sediment characterization of clay, silt and sand. . . . . . . . . . . . . . . . . . . . . . . . .31
Photograph of bottom barrier deployment by divers. . . . . . . . . . . . . . . . . . . . . 33
Macrophyte growth in field treatment plots 8 months after barrier removal. . . 36
Dissolved oxygen concentrations under selected bottom barriers. . . . . . . . . . . 38
Results of Lake Tahoe turion sprouting experiments. . . . . . . . . . . . . . . . . . . . .42
Germination rate for bench-scale tests using SF Bay-Delta turions. . . . . . . . . .43
Germination rate for mesocosm tests using SF Bay-Delta turions. . . . . . . . . . .43
vii
List of Tables
Table 2-1
Species-specific density and biomass values for plants and turions. . . . . . . . . . . . 13
viii
Acknowledgements
This project was funded by Research Joint Venture USDA No. 11JV11272170091
between the USFS Pacific Southwest Research Station and UC Davis under master
California Cooperative Ecosystems Study Unit agreement No. 08JV11272150052. A
major portion of the funding originated with the Department of Interior Bureau of Land
Management through the Southern Nevada Public Land Management Act (SNPLMA).
We thank Jonathan Long and Tiffany van Huysen with PSW for their patience and
continual help during this study.
We especially acknowledge the contribution of Ben Bradford at the USDA’s ARS at
Davis, CA for his invaluable help collecting samples and hard work throughout this
project. Cooperation with locating field sites and on-lake logistics was kindly provided
at Lakeside Marina and Tahoe Keys Marina. We appreciate our partnership with
members of the Lake Tahoe Aquatic Invasive Species Working Group and other
stakeholders for their continued support and efforts to manage AIS in Lake Tahoe.
George Malyj’s wizardry in program management was once again evident.
Report Preparers:
Funding & Administration:
ix
10
1. Introduction
1.1 Background
Aquatic invasive species introductions to Lake Tahoe have increased in recent decades.
Invasive aquatic plants are dispersing and impact the nearshore/shallow-water condition
in this ultra-oligotrophic lake. The rooted aquatic macrophyte, curly-leaf pondweed
(Potamogeton crispus) was initially seen in the southern portion of the lake and has
expanded to other areas along the littoral zone. Because curly-leaf pondweed is
relatively recently established and still has a restricted range, management may still be
considered a possible, albeit, costly option.
The scientific literature suggests that depriving the early life-stage of pondweed of
oxygen can significantly inhibit both laboratory and in situ germination (literature
presentation to follow). Based on these findings it was hypothesized that non-permeable
bottom barriers, that sufficiently reduce oxygen, may be able to lower the sprouting rate
of pondweed turions. Turions are vegetative reproductive structures (buds) produced by
pondweed plants that can sprout in the sediment to grow new plants (Figure 1-1).
With this backdrop, researchers from the University of California, Davis (TERC) and
the University of Nevada (Aquatic Ecosystems Laboratory) received a Round 11 science
research grant from the federal Sierra Nevada Public Lands Management Act
(SNPLMA) as part of its Lake Tahoe program. The project was to investigate the
potential for pondweed control in Lake Tahoe using bottom barriers to reduce oxygen
levels and inhibit turions from sprouting. This project was funded under the
‘Watersheds’ theme and the ‘Understanding the impacts of aquatic invasive species’
subtheme.
Figure 1-1. Photograph of turions on plant (left), attached to a removed set of leaves, detached
from leaves (center) and in the process of sprouting (right). Turions are produced in early
summer, fall off and enter the sediment, then sprout in the fall. These turions are about 1 cm in
size (taken from Anderson 2010, 2012).
1.2 Nuisance aquatic macrophytes in Lake Tahoe: A brief overview
Aquatic invasive species have been identified in recent years as a major threat to Lake
Tahoe, and there have been a number of rapid responses by basin management agencies
to monitor and control various non-native invertebrate, macrophyte and warm-water fish
populations. In particular, three recent invaders, Asian clam (Corbicula fluminea)
(identified in 2002), Eurasian watermilfoil (Myriophyllum spicatum) (identified 1995,
Anderson and Spencer 1996) and curly-leaf pondweed (P. crispus) (identified 2003) are
moving outward from their area of origin, and are establishing or threatening other
portions of Lake Tahoe. In the Tahoe Keys, aquatic macrophytes are present in
excessive abundance and provide habitat for warm-water invasive fishes (Kamerath et
al. 2008). At the same time these submerged plants pump phosphorus into the water
column potentially leading to increased algae production (Walter 2000), and are a
navigation and aesthetic nuisance to homeowners, who incur an annual harvesting cost
of $260,000 for this species (Dotson 2007). Both M. spicatum and P. crispus have
spread across a large portion of the south shore of Lake Tahoe since their initial
discovery (Figure 1-2, Figure 1-3). Curly-leaf pondweed has been called the most
widely dispersed, nuisance forming invasive aquatic macrophytes in North American
lakes where it can inhibit recreation, increase water column phosphorus concentrations
and impair water quality (Bolduan et al. 1994, Woolf and Madsen 2003).
As part of the Lake Tahoe Aquatic Invasive Species Working Group (LTAISWG) and
the Lake Tahoe Aquatic Invasive Species Coordinating Committee (LTAISCC),
resource agencies and scientists are collaborating to abate both the spread and extent of
thee invasive aquatic plants (e.g. Anderson 2012, Shaw et al,. 2012). These activities
primarily focus on the use of benthic barriers and localized harvesting by hand when
appropriate (Figure 1-4).
1.3 Use of benthic barriers to control aquatic species
The use of benthic barriers to inhibit vital physiological or metabolic requirements such
as photosynthesis or respiration is well known as a non-chemical management strategy
for aquatic nuisance species (e.g. Engel 1984, Ussery et al. 1997, Gunnison and Barko
1992, Eichler et al. 1995, Wittmann et al. 2012). In Lake Tahoe, the use of polyethylene
fabric bottom barriers to control Eurasian watermilfoil has been implemented since 2007
with localized population reductions observed after treatment (Shaw et al. 2012). In
2009, UCD and UNR researchers, in cooperation with Tahoe basin agencies
experimented with ethylene propylene diene monomer (EPDM) rubber benthic barriers
on Asian clam populations and achieved complete mortality in 28 days – during the
warmer summer months - due to the reduction of dissolved oxygen concentrations under
the barriers (Wittmann et al. 2010). Under colder water temperatures it required 2-3
months for mortality (Wittmann et al. 2011). In 2010, one-acre of EPDM benthic barrier
was applied to Asian clam populations in Lake Tahoe as part of a 120-day experiment
using this method (Wittmann et al. 2012).
2
Life history and ecophysiological characteristics of submersed aquatic macrophytes
make these species another ideal candidate for treatment with benthic barriers. Aquatic
plants require light for growth, and the use of benthic barriers block light, inhibiting
photosynthesis until shoot mortality occurs (USACE 2005). Traditional benthic barriers
(i.e. gas permeable) have been used on curly-leaf pondweed with variable results.
Mayhew and Runkel (1962) and Mayer (1978) reported that covering curly-leaf
pondweed populations with benthic barriers (polyethylene and Aquascreen™ (fiber
glass coated sheeting)) was effective at reducing biomass, but did not report on
subsequent recolonization. Madsen and Crowell (2002) suggest that bottom barriers are
effective to prevent the growth of rooted curly-leaf pondweed, however, long-term
management requires the elimination of vegetative buds called turions to interrupt the
life cycle of curly-leaf pondweed.
Bottom barriers have been successfully used in Emerald Bay, Lake Tahoe for the
treatment of submerged aquatic macrophytes (e.g. Shaw et al, 2012, TRCD 2012).
3
Figure 1-2. The distribution of Eurasian watermilfoil (Myriophyllum spicatum) and curly-leaf
pondweed (Potamogeton crispus) in Lake Tahoe, 1995–2006. Red circles indicate the presence
of M. spicatum, yellow triangles indicate the presence of curly-leaf pondweed, which was
discovered in 2003. Map created and provided by Lars Anderson, USDA-ARS. 4
Figure 1-3. Distribution of curly-leaf pondweed along the south shore of Lake Tahoe. View of
photo is from south to north. Slide created by and data is from Lars J. Anderson (ret.), USDA
Aquatic Weeds lab, Davis, CA (Anderson 2010). Includes data from 2009.
Figure 1-4. Ted Thayer of the TRPA examining dense growth of curly-leaf pondweed.
5
1.4 The role of turions in the curly-leaf pondweed life cycle
Curly-leaf pondweed sprouts from dormant shoot segments called turions (see Figure 11). In the American northeast, turions fall from the stems (Figure 1-4) and are observed
to germinate in late summer or fall, and the plants overwinter as small plants and resume
growth in spring when water temperatures rise (Figure 1-5). Individual stems may
spread locally by rhizome growth and curly-leaf pondweed biomass often reaches its
maximum in early summer while flowering and fruiting occur from April to May. This
seasonal pattern allows curly-leaf pondweed to avoid competition from other species,
giving it ecological advantages as an invasive species in freshwater systems (Tobiessen
and Snow 1984). Turion production is curly-leaf pondweed main source of vegetative
reproduction (Rogers and Breen 1980) and sexual reproduction occurs through seed
production (Waisel 1971). However, seed germination is extremely low (<0.1%) and not
commonly thought of as an important source of recruitment (Rogers and Breen 1980).
Additionally, turions also act as a storage unit for carbohydrate (Madsen and Crowell
2002), which can enable dormancy until environmental conditions are more favorable.
Though curly-leaf pondweed is known to be susceptible to contact herbicide treatment,
plant biomass re-growth often occurs due to turion formation as the major form of reinfestation during the following growing season (Netherland et al. 2000). Thus, curlyleaf pondweed turions are not only the main means of population propagation,
reproduction and dispersal, but also can expand the window of opportunity for
successful recruitment for this species through carbohydrate storage, herbicide evasion
and season-based competition.
Figure 1-5. Schematic diagram depicting a typical annual growth pattern for curly-leaf
pondweed highlighting the production of turions on more mature plants late in the growth cycle.
These reproductive buds are dropped to the sediment and are able to subsequently sprout into
new plants (from Johnson 2012).
6
While turion germination can enable the sustainability of curly-leaf pondweed
populations, it can be limited by a number of physical factors. Laboratory and field
studies have found that sprouting of dormant turions is generally controlled by
temperature, light intensity and photoperiod (Rogers and Breen 1980, Sastroutomo 1980,
Kadono 1982, Tobiessen and Snow 1984, Jian et al. 2003). However, in Asian systems
where curly-leaf pondweed is native and used to restore degraded lakes, turion sprouting
has ceased as a result of low light availability and anoxic conditions (Wu et al. 2009,
Short et al. 1987, Lauridsen et al. 1993, Clarke and Wharton 2001, Ni 2001).
Understanding turion sprouting behavior is key to assessing the population dynamics of
this species and the potential for non-chemical treatment in Lake Tahoe. More broadly,
the understanding of the interaction between the plant physiology, ecology, and
management is key toward a successful control program. Figure 1-6. Curly-leaf pondweed life cycle as observed in North America (Madsen and Crowell
2002). Seasonal patterns observed in Lake Tahoe have different temporal scales than plant
populations observed in the northeastern regions of North America (L. Anderson, pers. comm.). 1.5 Scope of Work
Project tasks were:
1. Profile five sites around the South Shore of Lake Tahoe containing existing
curly-leaf pondweed populations for density (stems m-2) and biomass (g dry
weight m-2) of both plants and turions. At each site survey species-specific
density and biomass for other submerged aquatic macrophytes.
2. Select three of the five profiled sites and deploy three types of bottom barriers
over adult curly-leaf pondweed populations and turions to determine the impact
of anoxia in these plots. These include jute, polyethylene, and EDPM rubber
barriers.
7
3. Conduct a fall turion survey of the three barrier deployment sites to quantify the
Tahoe turion bank as it relates to plant biomass/density, and habitat quality. We
were unable to complete this aspect of the project due to limited time and
resources. As explained below (section 2.2) the sprouting experiment at Lake
Tahoe did not produce reliable results and therefore the project team conducted
an extensive set of addition experiments (beyond the original scope) designed to
further address Task 2.
8
2. Methods and Results
2.1 Objective 1: Profile five sites around the South Shore of Lake Tahoe with existing
curly-leaf pondweed populations.
a. Determine macrophyte community species composition, density and
biomass
b. Determine the number and biomass of curly-leaf pondweed turions
c. Measure, year-round, lake temperature in vicinity of sampling sites
d. Measure soluble N and P in macrophyte beds and plant tissue
composition
2.1.1 Methods
2.1.1.1 Site location
Five sites along the south shore of Lake Tahoe were selected based on initial
observational field surveys that demonstrated the presence of curly-leaf pondweed as
well as a proximity to local marinas. The selected sites were located inside and outside
Lakeside Marina, (38°57.540’ N, 119°57.101’ W; 38°57.590’ N, 119°57.070’ W),
outside Ski Run Marina (38°57.159’ N, 119°57.560’ W), and inside and outside the
Tahoe Keys (38°56.104’ N, 120°01.034 W; 38°56.529’ N, 120°00.529’ W) (Figure 2-1).
The site inside Tahoe Keys was located in a dead-end channel - the closest road location
was 2008 Aloha Dr, South Lake Tahoe, CA. The proposal originally called for sampling
inside Ski Run marina, but permission was denied, and so we switched to inside
Lakeside Marina, where the owner agreed to participate. This profile of five sites was
not intended to duplicate the much larger lake survey conducted by Lars Anderson
(USDA Aquatic Weeds Lab, Davis, CA) between 1995-2006 (refer to Figures 1-2 and 13).
2.1.1.2 Surveying the five sampling sites
The surveys were made prior to the barrier deployment experiment. Biomass sampling
was conducted at four of the five sites (inside Lakeside Marina, outside Lakeside
Marina, outside Ski Run Marina, and outside Tahoe Keys East) on August 17th, 2011,
and inside the Tahoe Keys on August 18th, 2011. Five replicate samples for density and
biomass were collected per site, using ¼ m2 quadrants. The ¼ m2 quadrants were
designed to pull apart to allow the halves to fit around large, tall clumps of plants. All
plants that were within the quadrant were collected and brought to the surface and
separated by species.
Stem counts per plant species were recorded, along with turion counts. All plant species
were separated, counted as number of stems and dried for determination of dry weight.
Subsamples of dried curly-leaf pondweed were transported to the UC Division of
Agriculture and Natural Resources analytical laboratory (UC Davis campus) for
measurement of plant carbon (SOP 522.01), nitrogen (SOP 522.01) and phosphorus
9
(SOP 540.02). Turions were separated from plant stems, counted and stored separately.
Water samples from inside the macrophyte beds were collected on August 23rd from
each site at both mid-water column and sediment-water interface locations. Hand-held
bottles were used by divers to collect water from the mid-water depths and hand-held
syringes were used to sample at the sediment-water interface. Water was analyzed for
nitrate (NO3--N), ammonium (NH4+-N) and soluble reactive phosphorus (SRP) using the
standard protocols of UC Davis Tahoe Environmental Research Center for low-level
nutrients (Liston et al. 2013).
Figure 2-1. Location of five sampling sites used in this study; South Lake Tahoe, California.
In October 2011, the biomass sampling sites were assessed for sediment particle size
distribution (sediment structure) using triplicate petite ponar grabs (15 cm x 15 cm) of
the upper 2-4 cm of sediment later analyzed at the UC Division of Agriculture and
Natural Resources analytical laboratory (SOP 470.03). Particle size analysis for sand,
silt and clay was done using the hydrometer method. Samples were collected at Outside
Tahoe Keys, Inside Tahoe Keys, Inside Ski Run Marina, Outside Ski Run Marina and
Outside Lakeside Marina. Note that these differ somewhat from the sites for the August
biomass sampling.
10
To assess seasonal patterns in water temperature conditions, five Tidbit temperature
loggers (HOBO UTBI-001 TidbiT Temp, accuracy ± 0.2 °C) were deployed from
October 2011 to October 2012 at the sites used for biomass collections. The Tidbits were
staked to the sediment as close to the biomass collection locations as possible, and
recorded temperature every hour. Four temperature loggers were retrieved in October
2012, however the Tidbit outside of Ski Run Marina went missing. It appeared that this
temperature logger was taken, as the stake that it was attached to was still present, but
had been pulled up and was lying on the sediment surface.
2.1.2 Results and Discussion
2.1.2.1 Species composition and diversity, plant density and plant biomass
Nine species of aquatic macrophytes were identified during the August survey. These
included curly-leaf pondweed (Potamogeton crispus), Eurasian water milfoil
(Myriophyllum spicatum) - considered an invasive species, Andean milfoil (M.
quintense), Elodea (Elodea canadensis), Richardson’s pondweed (P. richardsonii), leafy
pondweed (P. folisus), water crowfoot (Ranunculus multifidus), white water buttercup
(Ranunculus aquatilis) and an unidentified pondweed (Potamogeton sp.). Two species
of macro-algae were also identified Nitella (Nitella sp.) and Chara (Chara sp.) (Table 21 a-e).
Total plant density in August, measured as number of stems m⁻2 ranged from
approximately 400-1,330 with a standard error to mean ratio between 14-53 percent.
These latter values indicate a large degree of variation between replicate samples and are
not unexpected given the heterogeneity typical of bottom dwelling organisms. At 1,068
and 1,332, total stems m-2 the sites located in quite, semi-isolated areas (Inside Lakeside
Marina and Inside Tahoe Keys, respectively) were on the order of twice as dense as the
three sites along the lake shore outside the marinas (~400-608 stems m-2) (Figure 2-2a).
Total plant biomass ranged from a low of 31 g dry wt. m-2 inside the Lakeside Marina to
a high of 92 g dry wt. m-2 in the Tahoe Keys (Figure 2-2b). The distribution of
macrophyte biomass among sites was similar to stem density with the striking exception
of inside the Lakeside Marina. This was due to the large contribution of the macro-alga
Nitella at the site; Nitella has a much lower biomass/stem ratio than the species of
aquatic angiosperms - 4-6 times lower in the present study.
The ratio of total plant biomass to stem density was fairly consistent in the range of 0.07
to 0.11. As mentioned above, this ratio was uncharacteristically low in the Lakeside
Marina due to large contribution of the macro-alga Nitella to stem density (Figure 2-2c).
Macrophyte biomass in the present study was within the range of values reported in the
literature. Biomass values include 200-260 g dry wt. m-2 (Duarte and Kalff 1986); 1,000
g dry wt. m-2 (Hopson and Zimba 1993); 100-1011 g dry wt. m-2 (annual values over an
eight year period, Harman and Albright 2004); 150-350 g dry wt. m-2 (Bosch et al.
2010); 307-1,100 (Ginn 2011); 50-200 g dry wt. m-2 (Schloesser and Manny 1984);
0.07-496 g dry wt. m-2 (median of 120 g dry wt. m-2, Wetzel 1983); 5-250 g dry wt. m-2
11
(Dowling and Anderson 1985). Downing and Anderson summarized data from 1,200
publications and found a range of 0.002-3,700 g dry wt. m-2. While the Lake Tahoe
macrophyte community clearly supported much less biomass than many lakes, the
measured range during this study (31- 92 g dry wt. m-2) was not exceedingly low.
Community structure was also evaluated using Simpson’s Index of Diversity (1-D). This
value ranges from 0-1 with greater diversity represented by the higher values. This index
represents the probability that two individuals randomly selected from a sample will
belong to different
12
Table 2-1 a-e. Plant and turion density and biomass (m-2) for the five profiles made along the
south shore of Lake Tahoe in August 2011. Values expressed as mean±standard deviation.
Values in parentheses denote medians. Curly-leaf pondweed and Eurasian water milfoil are
considered invasive in Lake Tahoe. Turions on both plants and on the bottom sediment were
included in these profiles.
13
Table 2-1 cont.
14
Table 2-1 cont.
15
Table 2-1 Cont.
16
Table 2-1 Cont.
17
Figure 2.2. Stem density (#/m2) and biomass (g dry weight/m2) for all plants in the submerged
macrophyte community at the five sites in Lake Tahoe. Vertical bars denote standard error.
18
species. The sites with the lowest Simpson’s Index of Diversity were Outside Lakeside
Marina (0.50) and Outside Tahoe keys (0.42)(Figure 2-3). Inside Lakeside Marina (0.67)
and outside Ski Run Marina had values of 0.65-0.67, while inside Tahoe Keys had the
most diverse community (0.78). Diversity was not related to either stem density or
community biomass. The Simpson Index of Diversity for macrophytes in lakes typically
is on the order of 0.75->0.90 (e.g. Nichols et al. 2000, Mouillot et al. 2005, Harmony
Environmental 2006, Endangered Resource Services 2008,Williamson and Kelsey
2008). Nichols et al. (2000) found that values of 0.50-0.60 were uncommonly low.
Figure 2-3. Simpson’s Index of Diversity (1-D).
19
Samples collected inside Lakeside Marina primarily consisted of a mixture of curly-leaf
pondweed (124 stems m-2), Elodea (112 stems m-2), leafy pondweed (496 stems m2) and
the macro-alga Nitella (316 plants m2) (Table 2-1a, Figure 2-4). These species accounted
for 98 percent of the total stem density. Dry weight biomass was slightly more
distributed among the five species than seen for stem density with leafy pondweed (12.4
g dry wt. m-2), curly-leaf pondweed (8.8 g dry wt. m-2) and Nitella (6.8 g dry wt. m-2)
accounting for 83 percent of the total biomass (Table 2-1a, Figure 2-4, Figure 2-5).
Outside Lakeside Marina, curly-leaf pondweed accounted for the majority of stem
density (352 stems m-2 or 67 percent of the total) and biomass (36.4 g dry wt. m-2 or 79
percent) (Table 2-1b, Figure 2-4, Figure 2-5). Stem density for Nitella was 112 stems
m-2 and accounted for 21 percent of the total stem number. However, with a biomass of
3.0 g dry wt. m-2 this macro-alga comprised only percent of the total biomass.
At the site outside of the Ski Run Marina, stem density and biomass were very similar to
those seen outside of the Lakeside Marina (Table 2-1c, Figure 2-4, Figure 2-5). Curlyleaf pondweed accounted for the majority of stem density (232 stems m-2 or 56 percent
of the total) and biomass (34.4 g dry wt. m-2 or 80 percent). Richardson’s pondweed was
the next most abundant species with a density of 80 stems m-2 (19 percent) and a
biomass of 5.8 g dry wt. m-2 (12 percent). Richardson’s pondweed was only this
abundant at Outside Ski Run Marina.
At the Outside Tahoe Keys Marina (east) site, curly-leaf pondweed dominated stem
density (456 stems m-2, 75 percent) and biomass (52.2 g dry wt. m-2, 90 percent) (Table
2-1d, Figure 2-4, Figure 2-5). Other species had minor contributions. The macrophyte
community inside the Tahoe Keys, and especially stem density, was well distributed
(Figure 2-5). Curly-leaf pondweed contributed a stem density of 324 stems m-2 (24
percent) with a biomass of 43.5 g dry wt. m-2 (47 percent). Eurasian watermilfoil was
much higher in the Tahoe Keys than the other locations at 140 stems m-2 (11 percent of
total at this site) and a biomass of 9.4 g dry wt. m-2 (47 percent). Other species included
Elodea (424 stems m-2 and 22.6 g dry wt. m-2), leafy pondweed (260 stems m-2 and 12.4
g dry wt. m-2) (Table 2.1e). White water buttercup was only found at this site with a
density of 98 stems m-2 and a biomass of 4.8 g dry wt. m-2.
The relative density (56-75 percent) and biomass (79-90 percent) of curly-leaf
pondweed, compared to the other species at each site) was highest for the three site
outside of marinas. Inside the two marinas the relative stem density of curly-leaf
pondweed was 12-24 percent. As highlighted in Figures 2-4 and 2-5, (1) stem number
and dry weight for curly-leaf pondweed were much greater than the other species, (2)
curly-leaf pondweed was lowest inside the Lakeside Marina, although similar at the
other four locations, and (3) Eurasian water milfoil was only significant inside the Tahoe
Keys. Elodea was also the highest inside the Tahoe Keys. Nitella was only abundant at
the two Lakeside Marina sites.
20
Figure 2-4. Density (stems m-2) and biomass (g DW m-2) for submerged macrophyte species
found at each site in Lake Tahoe.
21
Figure 2-5. Relative distribution (as percent of total) of density and biomass for individual
species at each sampling site. Data comes from Table 2-1. Panels represent the following: a –
Inside Lakeside Marina stem density, b – Inside Lakeside Marina biomass, c – Outside Lakeside
Marina stem density, d – Outside Lakeside Marina biomass, e – Outside Ski Run Marina stem
density, f – Outside Ski Run Marina biomass, g – Outside Tahoe Keys stem density, h – Outside
Tahoe Keys biomass, i – Inside Tahoe Keys stem density and j – Inside Tahoe Keys biomass.
22
2.1.2.2 Density and biomass of curly-leaf pondweed
As curly-leaf pondweed (Figure 2-6) is the focus of this research project, density and
biomass are specifically summarized in Figure 2-7a,b. Curly-leaf pondweed biomass
during this study ranged from 8.8 g dry wt. m-2 to 52.4 g dry wt. m-2. Woolfe and
Madsen (2004) reported a maximum biomass range for curly-leaf pondweed between
80-110 g dry wt. m-2 for four lakes in southern Minnesota. Similarly, Owens et al.
(2007) found curly-leaf pondweed biomass at 43 g dry wt. m-2. A common range for in
other lakes for curly-leaf pondweed lies between <50 g dry wt. m-2 (Rogers and Breen
1980) and 530 g dry wt. m-2 (Quade et al. 1994) during the growing season.
Curly-leaf pondweed density at the Lake Tahoe sites ranged from 124-456 stems m-2.
Other lakes commonly support densities of 200-1000 stems m-2 (e.g. McComas 2011,
McComas and Stuckert 2012). An important point here is that while Lake Tahoe is
classified as ultra-oligotrophic with regard to water column (phytoplankton)
productivity, it has moderate, yet significant, significant growth of curly-leaf pondweed
in those areas of the Lake where it has established. Values are in the range of some
eutrophic systems.
Figure 2-6. Photograph of curly-leaf pondweed growth in Lake Tahoe. (original source
of photo unknown, taken from Anderson 2010).
23
Figure 2.7. Density and biomass for curly-leaf pondweed and turions at the five sampling sites in
Lake Tahoe. Vertical bars represent standard error. The ratios of # turions:# stems (panel e) and
turion biomass:curly-leaf pondweed biomass (panel f) are also shown.
2.1.2.3 Turion density and biomass
The sampling design allowed us to measure turion density (turions m-2) and turion
biomass (g dry wt. m-2) (Table 2-1). Turions were found at all five sampling sites with
density ranging from 64 turions m-2 (Inside Lakeside Marina) to 408 turions m-2
(Outside Tahoe Keys) (Figure 2-7c). Turion density was generally related to curly-leaf
pondweed stem density with a ratio of 0.5-0.9 (Figure 2-7e). The mean±standard
deviation turion density for all five sites was 214±124 turions m-2. Turion density
(number m-2) in other lakes have been reported at 2,100 (Sastroutomo et al. 1979),
1,000-1,500 (Rogers and Breen 1980), 200-1,800 (Kunii 1982), 2,000-7,000 (Kunii
24
1989), 900-2,800 (Woolfe and Madsen 2003), 40-820 (Newman and Roley 2006), 340
(Owen et al. 2007), 850 to a maximum of 4,400 (Johnson 2007), 525 (Johnson 2010)
and 300-1,000 (Johnson 2012). The mean±standard deviation of the mid-point values
from each of these studies is 1,530±1,295 turions m-2. The median for all the reported
values given above is 1,000 turions m-2. While these summary values were 5-7 times the
mean found in Lake Tahoe, the highest Lake Tahoe value (408 turions m-2; outside
Tahoe Keys was 25-40 percent of the literature values. The mean density of 214 turions
m-2 and maximum site specific density of 408 turions m-2 found in Lake Tahoe were
within the lower range report by others. However, based on the literature, Johnson
(2012) developed a classification system where a density of turions in the range of 0-50
m-2 is associated with the potential for little or no recreational impairment, 100-150
turions m-2 is associated with slight recreational impairment, 250-300 turions m-2 is
associated with moderate recreational impairment, and 400+ turions m-2 suggests severe
impairment. This observation again highlights that while curly-leaf pondweed may have
a low, lake-wide distribution, it is at recreational impairment levels where it is
established.
The ratio of turion number to curly-leaf pondweed stem density ranged 0.5 to 0.9, inside
Lakeside Marina and outside Tahoe Keys, respectively (Figure 2-7e). The
mean±standard deviation of this ratio all sites was 0.7±0.2. This is in contrast to reported
ratios of 4-10 turions per stem (Kunii 1982) and 5-7 turions per stem (Bolduan et al.
1994). Since this project required a survey on one date during the summer when turion
production was seasonally active, perhaps sampling did not occur during the specific
period of maximum turion production. Another possibility is that turions could have
been buried in the sediments or that turion production varied with water depth (e.g.
Newman and Roley 2006). A full seasonal study would be needed to investigate this
further, however, the key findings here are that curly-leaf pondweed in Lake Tahoe does
produce turions at high densities, and that this turion production is significant.
Turion biomass was also measured (Table 2-1, Figure 2-7d). Values in Lake Tahoe
ranged from 2 g dry wt. m-2 (Inside Lakeside Marina) to 24 g dry wt. m-2 (Outside Tahoe
Keys). The mean (±standard deviation) for all five sites was 11±8 g dry wt. m-2. Similar
to observations for turion density, biomass did show differences between marina versus
outside marina sites. Turion biomass is not frequently reported in the literature, rather
turion density is much more often reported. Based on literature values for the number of
turions m-2 and accompanying curly-leaf pondweed biomass and using the
mean±standard deviation for the turion:plant biomass discussed below (0.30±0.12), the
estimated turion biomass reported by others could range from approximately 10-150 g
dry wt. m-2 with a median value of approximately 20-25 g dry wt. m-2. Further, Kunii
(1989) summarized data for turion density and biomass from various studies. This
information, along with an additional study by Neil and Graham (1994) resulted in an
estimate of 0.08 g dry wt per turion. When multiplied by the mean number of turions
from the Lake Tahoe sites the resulting an estimated turion biomass of 17 g dry wt. m-2
with a range of (5-33 g dry wt. m-2).
25
The ratio of turion biomass to curly-leaf pondweed biomass was on the order of 0.20 to
0.45 (Figure 2.7f), i.e. a significant amount of biomass for this species was present as
turions at the time of sampling. These values are well within the range reported by
others, e.g. 0.23 (Rogers and Breen), 0.42 (Kunii 1982), 0.29 (Kunii 1989), 0.14 (Neil
and Graham 1994) and 0.22-0.58 (Woolfe and Madsen 2003).
2.1.2.4 Temperature, water chemistry, bottom sediment and tissue composition
Water temperature - Temperature patterns were similar across the four sites, and
followed the pattern of cold-water temperatures in the winter months and increasing
temperatures beginning in April (Figure 2-8). The consistency between sites suggests
that water temperatures at the outside Ski Run site (not measured due to lack of
instrumentation) would have followed a similar pattern.
Maximum temperature was in August for all four sites ranging between 21 and 23 °C
(Figure 2-8). Minimum temperature was in January-February at approximately 5 °C.
Water temperature patterns inside the Tahoe Keys were the most different from the other
sites (higher in spring-summer and lower in winter), due to the ability of the open-water
to better buffer temperature changes, and the lack of connectivity between the Key and
the open water. A consistent pattern of warming was seen between March to August.
Temperature dropped somewhat slowly after the August peak; after October it declined
rapidly.
-
Figure 2.8. Temperature values (°C) at four biomass collection sites over the course of one
calendar year – 10/15/11 to 9/14/12.
Water chemistry – On August 25th, water samples were taken inside the macrophyte
beds at each site. Depths of collection included mid-water column and as close to the
bottom without disturbing the sediment-water interface. Nitrate concentrations were
very low in all samples ranging from <1 µg N L-1 to 3 µg N L-1 (Figure 2-9). These were
26
similar to values reported for the nearshore (lake-wide) by Loeb and Reuter (1984).
There was no evidence of difference between the mid-water column and near-bottom
concentrations. Phosphorus concentrations in the present study were in the range of 1 µg
L-1 with the obvious exception on inside the Tahoe Keys. Here the mid-water column
and near bottom values were much higher at 9 and 14 µg L-1 (Figure 2-9). Loeb and
Reuter (1984) reported typical nearshore phosphorus levels of 2-6 µg L-1 during the
summer; concentrations inside Tahoe Keys were not included in the Loeb and Reuter
(1984) study. Tahoe Keys is semi-isolated from the open water and is has a very high
level of biological productivity, especially its large, dense macrophyte community. As
plants die and decompose, or exude extracellular products, total and soluble phosphorus
can accumulate. Also, soluble phosphorus can enter the water column under localized
conditions of anoxia or if the bottom is disturbed. Since dilution of Tahoe Keys water is
so slow compared to the open-water, the high phosphorus concentrations are not
necessarily unexpected. Ammonium was much higher that nitrate at all sites ranging
from 2-16 µg N L-1 but most commonly at 5-10 µg N L-1 (Figure 2-9). With the
exception of inside Tahoe Keys there was no apparent mid-water column and near
bottom difference. This was also observed for soluble reactive-P. There are little to no
relevant data for ammonium in the nearshore; however, the elevated values observed in
the macrophyte beds appears to reflect that this represents an environment that is high in
organic matter which decomposes to form ammonium.
Figure 2-9. Nutrient concentrations at each of the sampling sites taken inside the macrophyte
bed. The designation of “mid” denotes that sample was taken at the mid-point between the
top of the plants and the sediment, while “sed” was taken just above the sediment-water
interface. All samples hand-collected using SCUBA. Concentrations expressed as µg N L-1
and µg P L-1.
Macrophyte tissue composition – Plant carbon, nitrogen and phosphorus were measured
in curly-leaf pondweed at three locations – Outside Ski Run, Inside Tahoe Keys and
Outside Tahoe Keys. Values are expressed as percent of C, N or P per unit dry weight of
curly-leaf pondweed biomass. Mean percent C was very similar between all three sites at
levels of 39 percent, 37 percent and 35 percent for Outside Ski Run (OSR), Inside Tahoe
27
Keys (ITK) and Outside Tahoe Keys (OTK), respectively (Figure 2-10). These values
were lower, but in the vicinity of values of 45-50 percent reported in the literature (e.g.
Ho 1979, Hakanson and Boulion 2002). Median percent carbon was also similar
between sites and similar to the means. Median values were 38 percent at OSR, 37
percent at ITK and 38 percent at OTK Percent carbon for OSR showed much less
variation that the other two sites.
Curly-leaf pondweed nitrogen content ranged from 1.1 percent outside Ski Run Marina
to 1.3 percent outside of the Tahoe Keys to 1.7 percent inside the Tahoe Keys (Figure 210). Literature values are commonly in the range of 1-2 percent (e.g. Ho 1979, Boulduan
et al. 1994; Fernandez-Alaez 1999; Owens et al. 2007). As was the case for plant
carbon, and based on literature values, there is no reason to suspect that there was an
ecological difference between the sites.
Mean total plant phosphorus ranged from 0.07-0.08 percent (Figure 2-10). Owens et al.
(2007) found plant phosphorus in curly-leaf pondweed to be 0.2±0.01 percent. In their
review of curly-leaf pondweed, Bolduan et al. (1994) report a range of 0.11 to 0.8
percent with values most commonly in the 0.2-0.4 percent range. Ho (1979) published
similar values (0.2-0.4 percent) for other macrophytes.
By comparing the coefficients of variation (CV) (defined as mean÷standard deviation)
we see that percent carbon had the lowest degree of difference between the 15-16
replicate plants analyzed – CV=5, 10, 14 percent at OSR, ITK and OTK, respectively.
At these same sites, the CV for percent N was 46, 55 and 38 percent, respectively. The
CV for percent P was 25, 43 and 55 percent respectively. While it is not within the scope
of this project to investigate these differences, the observation that the CV for the plant
growth nutrients N and P were higher and similar, while there was little variation in the
carbon content may reflect low and patchy levels of nitrogen and phosphorus in the
environment.
The N:P ratio for curly-leaf pondweed in Lake Tahoe was on the order of 17 by weight.
Other studies report this ratio as 5:1-9:1 (Eilers 2005), 10:1 (Fernandez-Alaez 1999;
Owens et al. 2007) and 12:1 (Duarte 1992). The 17:1 ratio in Lake Tahoe curly-leaf
pondweed is considerably larger suggesting possible P-limitation (Eilers 2005).
However, we stress caution in that this study was not intended to evaluate nutrient
limitation in this species and sampling was limited to a very narrow time period.
2.1.2.5 Sediment particle size analysis
Sand dominated the percent weight at all the sampled sites with the glaring exception of
Inside Tahoe Keys (Figure 2-11). The contribution of sand was 90, 95, 97 and 95
percent at OTK, OLS, ISR and OSR, respectively. At these sites the contribution of silt
to total sediment weight was 2-6 percent while the clay accounted for 1-4 percent. Inside
the Tahoe the contribution of sand declined dramatically to only 48 percent while silt
and clay rose to 39 and 13 percent, respectively. This was not unexpected as the Tahoe
Keys is found in a dredged location (formerly part of a large wetland), with quiescent
waters that receive non-point source drainage.
28
Barko and Smart (1986) studied the growth of Eurasian water milfoil and Hydrilla in
laboratory experiments, using 40 different sediments from 17 geographically widespread lakes in North America. They found diminished growth of macrophytes in highdensity, inorganic sand (>75 percent sand by mass). The authors also report that this
general finding is complicated by sediment density and percent organic matter, both of
which were not measured in this study. Clearly, however, the presence and spread of
submerged macrophytes along the shore of Lake Tahoe demonstrates that these plants
can grow under the higher levels of sand observed.
29
Figure 2-10. Curly-leaf pondweed tissue composition for carbon, nitrogen and phosphorus.
Expressed as percent of total plant biomass.
30
Figure 2-11. Sediment characterization at the five study locations based on percent clay, silt and
sand as dry weight.
31
2.2 Objective 2: Deploy three types of bottom barriers in curly-leaf pondweed beds to
determine the impact of anoxia on turion sprouting.
a. Deploy and retrieve barriers
b. Collect turions after experimental deployment of barriers and assess
dormant or non-dormant status and viability, as germination in laboratory
experiments
c. Measure dissolved oxygen (DO) underneath the barriers
d. Survey the impact of the barriers on adult plant populations immediately
following barrier removal, at approximately eight months, and
approximately one year
2.2.1 Methods
2.2.1.1 Field experimentation and surveys in Lake Tahoe
Barrier deployment/removal and in-lake experimental design – The three sites used for
barrier deployment were Outside Tahoe Keys (OTK), Inside Tahoe Keys (ITK) and
Outside Ski Run Marina (OSR) (Figure 2-1). It was originally intended to have a site in
the vicinity of Lakeside Marina, however, the only suitable site was inside the
breakwater, extremely close to the swimming beach in shallow water. Since barriers
placed at this location would be easily accessible to all beach patrons, and thus easily
tampered with, it was decided to place the third set of barriers outside of the Ski Run
Marina.
Jute, polyethelyne, and EDPM rubber barriers (one of each) were deployed at each site
on August 21-23, 2011 by UC Davis TERC research divers. Barriers measured 3 m2 and
sufficient rebar was used to weight each barrier down (see Figure 2-12 for an example of
deployment). Each jute barrier consisted of two jute fabric layers placed on top of each
other. This was done because the weave in the jute was considered too large for the
purpose of isolating the bottom from dissolved oxygen in the overlying water.
Turions from curly-leaf pondweed were collected in a batch from the west end of Tahoe
Keys marina in June of 2012 (coordinates: 38.933173, -120.015990). Turions were kept
in cold water (4 °C) in the dark until placement under the in situ bottom barriers was
initiated. Consequently, all turions used in this experiment represented a single cohort.
Some turions began to sprout in the short time period between initial collection in the
Tahoe Keys and placement under the bottom barriers. This pre-treatment germination
was accounted for in the results on post-treatment germination.
Spencer and Ksander (1997) suggested that variation in propagule size may lead to
differences in survivorship, thus two size classes were used, small 50 to 100 mg and
large turions 150 to 250 mg. Labeled mesh bags of turions were placed under each
barrier, and attached to short pieces of rebar. The use of mesh bags helped insure
retrieval of turions from under each of the treatment barriers. Two bags of turions were
placed under each barrier. One contained small turions (50-100 mg, n=15) and the other,
large turions (150-250 mg, n=15). Identical mesh bags containing turions (n=15, at all
three sites) were zip-tied to a stake driven in the sandy sediment near, but outside of
32
each of the three experiment barriers– these served as controls barriers.
At the Outside Tahoe Keys site, two dissolved oxygen probes (Zebra-Tech, Ltd. D-Opto
Logger, accuracy +/- 0.1°C for water temperature and one percent of the measured value
for dissolved oxygen) were placed underneath the jute and polyethylene barriers.
Dissolved oxygen probes were not placed underneath the EDPM rubber barrier since an
extensive data already existed that documented the ability of EDPM rubber barriers to
achieve anoxic conditions (Wittmann et al. 2010, 2011).
Barriers were retrieved at our three experiment sites (three barrier types per location) in
October 2011, approximately two months after deployment. Barriers were removed
following HACCP protocol (hazard analysis and critical control points) for mitigating
impacts of potential invasive species contained in the barriers. Bagged turions were
retrieved during removal. As the barriers were removed, the barrier plots were
delineated. Visual assessment of re-growth was performed at approximately eight
months and approximately one year after the barrier deployment. The exception to this
was inside the Tahoe Keys. Visibility was extremely poor at this site (< 15 cm inches),
and we were unable to place any kind of markers at the corners of these plots.
Consequently, it was not possible re-survey the pondweed re-growth inside the Tahoe
Keys. However, when we collected the temperature logger from this site in October
2012 (one year after barrier removal), visual observations indicated no diminishment in
the pondweed population.
Figure 2-12. Research divers roll out a bottom barrier over macrophyte bed.
33
2.2.1.2 Laboratory testing of Lake Tahoe turion sprouting
Post-treatment turion viability was based on visual inspection of new sprouts following a
vernalization protocol1 (Sastroutomo 1981). New or existing sprouts indicated turions
were viable and survived the treatments. These post-treatment sprouting tests were
performed on the University of California, Davis campus in September, 2011. Two
hundred milliliters of deionized water was placed into 250 ml beakers with 10 mg of
osmocoat slow release fertilizer. Turions were removed after an 8 week barrier
treatments and then placed in a growth chamber under the following conditions:
photoperiod: 12 hour daylight: 12 hour night; photosynthetic photon flux (area) density:
1126 µmole sec-¹ m-² light intensity, and temperatures of 5°C for two weeks, followed
by 1 week at 30 °C (Sastroutomo 1981).
2.2.1.3 Further testing of turion germination following anoxia
Given the uncertainty associated with the turion germination experiments in Lake Tahoe
as well as in the literature, we went considerably beyond both the scope of work and the
budgetary allocation of this project to further investigate this question. Since turions
were not available from Lake Tahoe (due to natural seasonality), they were collected
from the San Francisco Bay-Delta where turions were abundant. Even though the turions
were not from Lake Tahoe, the results of the experimental treatment (i.e. use of three
types of bottom barriers) were considered applicable to the management question at
hand.
Thomas Barr (UC Davis Plants Sciences, Ph.D.) conducted these extra experiments.
Bench-scale tests for sprouting germination following jute, polyethylene and rubber
barrier treatment were conducted on the UC Davis campus and mesocosm experiments
were run at the California Department of Food and Agriculture “B” Street facility in
Sacramento.
The bench-scale tests were conducted in 250 ml glass beakers covered with jute,
polyethylene and rubber. There was no sediment in these beakers. Turions were
subjected to an 8-week treatment. The rubber barrier beakers were purged with nitrogen
gas and sealed to maintain anoxia. Following the 8-week treatment turions were
sprouted as described above in section 2.2.1.2.
The mesocosm experiments were conducted 200 cm x 125 cm x 25 cm fiberglass
chambers (see photograph on last page of this report. Turions were placed in 15 cm
diameter x 6 cm deep pots containing sediment and placed in the experimental
chambers. Pots were covered with the three types of barrier material. The sprouting
protocol was as above.
Details of the methodology used in these bench-scale and mesocosm experiments are
provided in Appendix A of this report.
1
Vernalization is the acquisition of a plant's ability to flower or germinate in the spring by exposure to the
prolonged cold of winter.
34
2.2.2 Results and Discussion
2.2.2.1 Field surveys
Re-growth of macrophytes following barrier removal - Plant species composition and regrowth were assessed in June 2012 (eight months after deployment) inside the
delineated plots that were previously covered by the three types of barriers. Very little
re-growth was observed (Figure 2-13). All re-growth at Outside Tahoe Keys was
Richardson’s pondweed, whereas all re-growth at Outside Ski Run was curly-leaf
pondweed. Plant re-growth in the rubber barrier plots was between 0-2 stems m-2. Plant
re-growth in the jute barrier plots varied between sites (no re-growth at Outside Tahoe
Keys and 3-5 stems m-2 at Outside Ski Run Marina). Plant re-growth was also variable
in the polyethylene barrier plots, averaging around 3-5 stems m-1 at Outside Tahoe Keys
and 1-2 stems m-1 at Outside Ski Run.
The second and final assessment of the re-growth in the barrier treatment plots occurred
in October 2012, one full year after barrier removal. The differences in re-growth
between the sites were far more pronounced, with extremely low levels of plant
coverage in the Outside Ski Run Marina plots and high plant coverage at plots at the
Outside Tahoe Keys location. Both the rubber and jute barrier plots outside the Ski Run
Marina showed less than five percent plant coverage. The polyethylene plot had
approximately 30 percent coverage of short (7-10 cm) curly-leaf pondweed. The
polyethylene plot outside the Tahoe Keys had 60 percent plant coverage, primarily
curly-leaf pondweed with10 percent was water milfoil. The jute plot outside the Tahoe
Keys Marina had the lowest plant coverage at this location, approximately 30 percent.
Again the plants were primarily curly-leaf pondweed with a small percentage of water
milfoil. All plants were short, approximately 15 cm tall. Coverage was highest at the
rubber plot, at 80 percent. The plants were dominated by curly-leaf, with 10 percent
water milfoil. The pondweed plants in the rubber barrier plot were the tallest plants in all
of the barrier plots, on the order of 60 cm. It appeared to still be peak growth season
outside the Tahoe Keys Marina, as there was no sign of senescence or sloughing in any
of the plants.
We hypothesize that the plants in the barrier plots were using rhizomes for re-growth in
place of turions, based on visual observation made in the re-growth treatment plots.
35
!"
!"
!"
Figure 2-13. Example of invasive weed growth in barrier plots eight months after the removal of
the polyethylene (A), EDPM rubber (B), and jute (C) bottom barriers (June 2012).
36
Dissolved oxygen levels under the bottom barriers - Dissolved oxygen (DO) was
measured underneath the jute and polyethylene barriers at the Outside Tahoe Keys site.
Unfortunately, after being downloaded, the DO data from underneath the polyethylene
barrier was corrupted and unusable. Observations of the polyethylene barriers suggest
that DO levels were most likely relatively high, given the extremely poor sedimentwater seal and gapping in the material that occurred in all of the deployments of that
type of barrier material. Additionally, many living crayfish were found underneath the
polyethylene barriers during barrier removal.
Results from the jute barrier were unexpected - DO levels underneath this barrier type
reached zero approximately three weeks in to the deployment (Figure 2-14, top panel).
To obtain more data from this barrier type, we conducted a second barrier deployment in
the same area, from October to November 2011. Given the poor performance of the
polyethylene barrier to seal to the bottom, with transport of lake water most likely under
this material, we placed one DO probe underneath a rubber barrier, and one underneath a
jute barrier. These DO probes were successfully retrieved and downloaded at the end of
November 2011 (Figure 2-14, bottom panel). While rubber bottom barriers have been
reported to create anoxic conditions elsewhere in Lake Tahoe (e.g. Wittmann et al.
2012) and showed a rapid decline to zero or near zero DO levels in the present study, the
jute barrier DO levels also became anoxic. Although experiments to further understand
DO dynamics under the jute mats was beyond the scope of this project, we hypothesize
that the decomposition of both ambient organic matter in the sediments and organic
matter from the jute material may have been sufficient for bacterial consumption of
oxygen. However, anoxic conditions were maintained for the remainder of the sampling
period underneath the rubber barrier, whereas the jute barrier showed periodic and
temporary spikes in DO, perhaps related to storm or wind events.
37
Figure 2-14. Average daily dissolved oxygen (DO) concentrations (± standard deviation) under
selected bottom barriers during the experimental deployment. The upper panel represents data
from jute barriers only; DO data from underneath the polyethylene barrier was corrupted, and
could not be analyzed. The lower panel represents data from jute and rubber barriers (red and
blue lines, respectively) from a later deployment of bottom barriers in the same location as in
panel A. Both panels represent data collected in 2011.
38
2.2.2.2 Laboratory sprouting of Lake Tahoe turions
Unexpectedly, a greater proportion of the turions placed under the bottom barriers were
found to sprout than in the controls, suggesting no impact of the barrier treatment on
germination (Figure 2-15). For all the controls, combined by site and size, the mean
(±SD) of germination for the controls was low at 25±15 percent. In their study of
sediment anoxia and turion sprouting, Wu et al. (2009) reported a 60-70 percent
sprouting rate for the controls. In the current study it was observed that for all three sites
combined the sprouting rate in the controls was higher for the large turions (36±8
percent) compared to the small turions (16±14 percent). This may be related to
nutritional stores in the larger turions, however, it was not in the scope of this study to
investigate this further.
For the small turions (50-100 mg) germination for each treatment was generally >1.5
times the control. This was not the case for jute and rubber from the Outside Tahoe Keys
location. Since there was no sprouting of the control turions from the Ski Run Marina
location is it not possible to calculate a relative percent sprouting; however, the absolute
sprouting rates for the small turions at this location were jute – 7 percent, polyethylene –
73 percent and rubber – 73 percent, with the latter two among the highest rates of
sprouting observed.
Treatment sprouting was less than the controls for only the Outer Tahoe Keys rubber
(30-35 percent less than controls) and for the polyethylene treatment at Inside Tahoe
Keys, also 30-35 percent lower. This was observed for the large turions (150-250 mg)
(Figure 2-15). For the remainder of the large turion experiments sprouting among all
treatments ranged from 1.0-3.2 times higher than the control with a mean (±SD) of
2.0±0.8 times higher than controls. There was no ecologically significant difference
between the response of the small versus large turions.
As noted above, the finding that the turions from under the in situ barrier treatments
sprouted more than the controls was not expected based on the study of Wu et al. 2009.
These authors found that the sprouting rate of turions was 68-72 percent lower under
highly anoxic treatment conditions than in the control. Wu et al. (2009) reported a
sprouting rate of approximately 20 percent even under highly anoxic conditions. Jian et
al. (2003) suggested that substrate oxygen condition had no influence on turion
sprouting, but Wu et al. (2009) speculated that the dissolved oxygen levels in the Jian et
al. experiments may have been too high to see an effect. Wu et al. (2009) used their
findings to suggest that ambient conditions of anoxia in waterbodies inhibit growth from
turions in pondweed populations. Under prolonged, seasonal/annual periods of anoxia,
this may well be the case.
Inside the Tahoe Key’s marina where turions were plentiful enough to be harvested, the
environmental parameters are much different compared to the open lake. Temperature
exposures during the harvesting and sorting potentially played a role in sprouting. Lake
Tahoe temperatures were 9 °C at time of harvest and air temperatures during sorting
were 32 °C. This may have led to premature sprouting for some turions that may have
39
otherwise been environment or treatment suppressed. Turions are adapted to sprout in
the cooling fall season to overwinter as young plants. Colder temperatures within Lake
Tahoe during treatments could have potentially resulted large sprouting differences
between experiments once the post-treatment sprouting protocol were performed.
It appeared that benthic barriers were successful in reducing the existing biomass of
curly leaf pondweed populations, with jute being the most cost-effective and easily
deployed bottom material. Examination of the test sites one-half year and one year after
application of the barriers indicated that pondweed populations did not appear to be
primarily using turions for recolonization of the plots, but were instead spreading via
rhizomes. This rhizome spread appeared to result in shorter, less dense pondweed
populations than were previously observed at the sites (pre-bottom barrier deployment).
We believe that the influence of temperature may have been at the core of the Lake
Tahoe sprouting findings. We attempted to set up a second in situ trial using barriers and
bags of turions to further investigate our results, however, extreme weather and wave
conditions destroyed the experimental set up in the lake.
2.2.2.3 Laboratory germination of turions – bench-scale and mesocosm experiments
with turions from the SF Bay-Delta
As with the methodology for the additional bench-scale and mesocosm experiments a
detailed results and discussion section provided in Appendix A. We summarize the
findings below.
Bench scale treatments showed that rubber barrier treatment had a highly significant
effect on sprouting turions of P. crispus with a p-value ˂ 0.0001 (Figure 2-16). Rubber
(no light, no oxygen) benthic bottom barrier treatments had a 48 percent sprouting
(standard error = 11 percent; n=16). The control (light and oxygen) and jute (reduced
light, oxygen) treatments had the same sprouting rate at 100% (SE= 0; n=16), whereas
polyethylene benthic bottom barrier treatments (no light, oxygen) had a small, nonsignificant reduction in percent sprouting of 97 percent (standard error = 2 percent;
n=16). Comparison of the treatment means using the Tukey-Kramer HSD test set for α
at 0.05, showed rubber benthic bottom treatments were significantly different from
polyethylene, jute and the control (Figure 2-16).
Mesocosm scale treatments also showed the rubber barrier treatment had a strong and
highly significant effect on sprouting turions of P. crispus (p ˂ 0.0001) (Figure 2-17).
Rubber (no light, no oxygen) benthic bottom barrier treatments had 30 percent sprouting
(standard error = 6 percent; n=16) which was less than seen in the bench-scale
experiments. The control (light and oxygen) had a 98 percent sprouting rate (standard
error = 2 percent; n=16). Jute (low light, oxygen) benthic bottom barrier treatments had
a 72 percent sprouting rate (standard error = 4 percent; n=16). Jute benthic bottom
barriers rapidly colonized with periphyton algae at this scale, reducing light after two
weeks into the test. Polyethylene (no light, oxygen) benthic bottom barrier treatments
had 70 percent sprouting (standard error = 5 percent; n=16). Means comparison using
the Tukey-Kramer HSD test set for α at 0.05 showed rubber benthic bottom treatments
40
were significantly different from polyethylene, jute and the control (Figure 2-17).
Additionally, means for jute and polyethylene were significantly different from both the
control and the rubber benthic bottom barrier treatments. Jute and polyethylene benthic
bottom barrier treatments were not significantly different from each other (p = 0.99).
We found that rubber – the only non-porous bottom barrier material – was the most
effective at inhibiting sprouting in turions of P. crispus (Figures 2-16 and 2-17). Porous
benthic bottom barriers (jute and polyethylene) had no effect on sprouting in the benchscale test without sediments, while the mesocosm experiments with sediments had
significant differences compared to the control (Figure 2-16). There may be several
reasons for the differences in the bench-scale and mesocosm experiments. First, the
bench-scale experiment controls lacked algae growth in the beakers, while the
mesocosms had dense periphyton growth occurring after two weeks into the experiment.
The sediments covered with algae may have high oxygen concentrations near the water
soil interface due to photosynthesis while the jute and polyethylene barrier treatments
acted as an algal shield, protecting the turions from algal growth and elevated oxygen.
Second, the thick coverage of algae on the porous barriers acted as its own barrier by
reducing gas exchange. Third, the addition of sediments to the mesocosms reduced
oxygen concentrations. All three effects could have potentially combined to reduce the
proportion sprouting through anoxia in both the porous and non-porous treatments at the
mesocosm scale. The effects of light appear to be minimal based on the experiments and
concurs with Wu et al. (2009).
Wu et al. (2009) used sucrose additions to the sediments to reduce oxygen levels in their
mesocosm aquariums in order to replicate field conditions. We used barriers as our
experiment was the test the efficacy of bottom covers as a management technique. These
two studies had similar results even though the methodology was different, i.e. mean
sprouting rate for P. crispus turions was approximately 20 percent in Wu et al. (2009)
and 30 percent in the current study. Wu et al. (2009) controls had lower sprouting rates
(70 percent) compared to our controls (100 percent). This difference may be due to the
time of year that turions were collected as well as ambient temperature between the
experiments. If turions are collected in autumn, they tend to be brown and smaller, these
have a lower proportion of sprouting (mean = 68 percent), while green turions collected
in the spring have the highest proportion of sprouting (mean 100 percent) (Sastroutomo,
1981). We collected only spring cohort P. crispus turions for this experiment.
41
Figure 2-15. Results of Lake Tahoe turion sprouting experiments performed in the
laboratory following treatment (coverage) with the three bottom barrier types: jute
polyethylene and rubber. Panels show the proportion sprouting in the post -treatment
trials relative to the controls. Small turions are 50-100 mg, large turions 150-250 mg.
42
Figure 2-16. Proportion sprouting (percent) of Potamogeton crispus turions after an 8-week
exposure under benthic bottom barrier treatments at the bench scale. Means are ± 1 SE (n= 16).
Data are combined from repeated experiments and size classes. Levels not connected by the
same letter are significantly different (Tukey HDS; p≤0.05).
Figure 2-17. Proportion sprouting (percent) of Potamogeton crispus turions versus benthic
bottom barrier treatment material for 8-week exposure at the mesocosm scale. Means are ± 1 SE
(n= 16). Data are combined from repeated experiments and size classes. Levels not connected by
the same letter are significantly different (Tukey HDS; p≤0.05).
43
3. Conclusions and Recommendations
`
Management Implications
In reference to aquatic macrophyte control applied to infested waterbodies in general,
Barr (2013) concluded that benthic bottom barriers alone cannot eradicate 100 percent of
the turions on their own, but that non-porous benthic bottom barriers could possibly be
used in conjunction with other methods for eradication of Potamogeton crispus turions
(e.g. heat treatment, acetic acid, etc.). Such combined treatments may be able to
selectively take advantage of the anoxic conditions that will enhance efficacy. Use of
these “other methods” must be very mindful of their impact on non-target organisms and
existing water quality regulations. This study does not recommend the use biocides for
treatment of curly-leaf pondweed in the open waters of Lake Tahoe. Rather, we provide
comments on ‘combined treatments’ to inform future discussions on this matter should
the need arise.
Conclusion
As hypothesized in the literature (Wu et al. 2009), while a standing condition of anoxia
in a water body may be an important mechanism inhibiting the sprouting of turions and
growth of curly-leaf pondweed, this appears to be fundamentally different that
establishing a temporary condition of anoxia for the purpose of inhibiting turion
sprouting. Since turions are produced each year, bottom barriers would have to be
installed annually, and then with only a 20-30 percent reduction in sprouting (i.e.
incomplete control). We did not find evidence to suggest that once the barriers were
removed that in situ turion sprouting would not occur in the treated plots, provided the
appropriate environmental conditions were present. Bottom barriers have been used to
control plant growth and biomass in Emerald Bay; however, those barriers affect
photosynthesis by blocking light, a distinction between material that are porous or nonporous to dissolved oxygen is not an issue. In contrast the use of non-porous material is
essential when the objective is the stop turion sprouting.
Recommendation
Based on our experiments we see no reason to recommend the large-scale application of
non-porous bottom barriers for managing existing curly-leaf pondweed populations in
Lake Tahoe that employ control of turion sprouting as a primary mechanism.
44
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49
Appendix A. Benthic Barriers for Control of Aquatic Weed Propagules
Chapter 1 from T.C. Barr, Ph.D. dissertation.
Barr, T.C., 2013. Integrative Control of Curly Leaf Pondweed Propagules Employing
Benthic Bottom Barriers: Physical, Chemical and Thermal Approaches. Horticulture and
Agronomy. University of California, Davis. 138 p.
The following is the abstract from the full dissertation along with the Table of Contents.
This is included in case the reader is interested in related research not part of the current
project’s scope of work. Chapter 1, as provided by TCB is below.
Abstract The effective management of submersed aquatic macrophytes depends on understanding their reproductive biology. Potamogeton crispus L. (curlyleaf pondweed, Potamogetonaceae) produces numerous asexual propagules that make traditional management difficult. It has spread to roughly half of the counties in California (USA) from alpine habitats such as Lake Tahoe to the tidally influenced Sacramento‐San Joaquin delta. Studies were conducted from May 2012 till October 2012 at the bench and mesocosm scales in Davis and Sacramento California to explore the effects of benthic barrier control measures on the propagules (turions) of Potamogeton crispus. The first study examined the effects of three benthic barrier materials (jute, polyethylene and rubber) had on turion sprouting. Jute benthic barrier material allows some light and oxygen through the fabric, while polyethylene allows oxygen, but not light. Rubber barrier material blocks light and oxygen exchange. Turion viability, as determined by sprouting, was then assessed post‐treatment. Results showed no significant differences at the bench scale for the untreated control (100% sprouting, SE=0%), jute (100% sprouting, SE=0%), or polyethylene treatments (96.9%, SE=2.1%)(n=16 for all treatments, α = 0.05). Rubber treatments resulted in 48.4% sprouting (SE= 10.6%; n=16). Results for the mesocosm experiments showed significant differences between the control and the jute and polyethylene treatments (control = 98.4% sprouting, SE= 1.6%; jute = 71.9% sprouting, SE= 4.5%; polyethylene= 70.3% sprouting, SE= 4.7%, n=16 for all treatments). Jute and polyethylene treatments were not significantly different in the mesocosm experiment. Rubber mesocosm experiments significantly reduced sprouting (29.7% sprouting, SE= 6.1%; n=16) compared to other treatments. While light had minimal impact on sprouting, anoxia appeared to be the main factor inhibiting sprouting using benthic bottom barriers. Barrier induced anoxic stress combined with herbicides may potentially offer enhanced efficacy. The second study explored enhancement of the impermeable rubber barrier material with dilute acetic acid loaded into cassava starch “pearls”. Turions were exposed for 2 weeks and then assessed for viability via post treatment sprouting protocol with and without hydrosoil at the bench‐ and mesocosm‐scale. Results for the bench‐scale showed 20.8 mmol L⁻¹ acetic acid treatments were not significantly different (p=0.4231) compared to the untreated control (Tukey HDS; p≤0.05). However, 41.6 mmol L⁻¹ acetic acid treatments were highly significantly different from the controls (p‐value < 0.0001) 50
at the bench‐scale, but did not completely inhibit sprouting (mean sprouting of 31.25% (SE= 11.97)). Complete inhibition of sprouting turions occurred for both experiments at and above acetic acid concentrations of 83.3 mmol L⁻¹ (SE= 0). Results showed that tapioca starch saturated with acetic acid and combined with impermeable benthic barriers may offer an effective chemical treatment for the control of Potamogeton crispus. The final study examined hot water exposures under the barriers to kill and inhibit sprouting in turions. Heated water circulated under an insulated benthic bottom barrier may potentially offer a simple non‐
chemical rapid method to target surface proagules on the sediment, subterranean propagules and young plants. Heated water was used to treat P. crispus turions at the bench and mesocosm scales (25°C, 40°C, 50°C, 60°C, 70°C and 80°C exposures for 30 to 300 seconds). Heated water exposures inhibited sprouting turions at 50°C and 60°C at the mescosm and the bench scales, however, did not completely inhibit sprouting for all time exposures except at the bench scale 60°C treatment for 300 seconds. For 70°C and 80°C treatment exposures, there was a slight difference at the 30‐second exposure mark, but at 60 second and beyond, all 70°C and 80°C treatments provided 100% inhibition. Cost to raise the temperature 60°C from ambient water temperature under the contained limited volume under insulated barriers suggests approximately 2 USD$ per 100 square feet for 5 minute treatments or 1400$ per acre. 51
52
Chapter 1 Benthic Bottom Barriers for Control of Aquatic Weed Propagules Abstract Aquatic weed propagules pose a serious long‐term management problem for water stakeholders. Propagules, such as turions, can evade traditional chemical treatment methods in the water column by resting in the sediment‐water interface. In many aquatic systems, such as Lake Tahoe, California (USA), herbicides are not authorized for use. In effort to target this source of long term propagule bank using non chemical methods, benthic bottom barrier materials were tested on Potamogeton crispus turions at the bench and mesocosm scales. Jute, polyethylene and rubber sheet materials covered turions for 8 weeks and then assessed for viability via post treatment sprouting protocol with and without soil at the bench and mesocosm scale. Results showed no significant differences at the bench scale for the control, jute or polyethylene (α = 0.05). Rubber treatments resulted in 48.4375% proportion sprouting (SE= 10.5743%; n=16). Results for the mesocosm experiments showed significant differences between the control and the jute and polyethylene treatments (control = 98.4375% proportion sprouting, SE= 1.5625%; jute = 71.875% proportion sprouting, SE= 4.49247%; polyethylene= 70.3125% proportion sprouting, SE= 4.6875% n=16 for all treatments). Mesocosm jute and polyethylene treatments were not significantly different. Rubber was significantly different from all mesocosm treatments with 29.6875% proportion sprouting (SE= 6.1317%; n=16). We conclude that light has a minimal impact on proportion sprouting and that anoxia is the main driver for inhibition when using benthic bottom barriers. Potential implications for barrier induced anoxic stress integrated with herbicides are discussed. 53
Introduction First application of benthic barriers Benthic bottom barriers have been widely used for aquatic weed management since the 1970’s in the United States and Europe (Born et al., 1973; Nichols, 1974; Mayer, 1978; Murphy, 1988; Eichler et al.; 1995; Caffrey et al., 2010). Benthic bottom barriers are gas porous or nonporous materials that block light and/or gas exchange, killing the plants underneath. Treatment is similar to terrestrial weed barrier fabrics used for horticulture and landscaping. Barrier materials are placed over the area of concern and then secured with “u” shaped rebar metal stakes or weights. Benthic bottom barriers may offer effective management options for limiting plant growth and establishment as a low cost rapid response tool to control establishment of new aquatic weed infestations (Gettys et al., 2009). Unlike terrestrial barrier systems, aquatic sediments produce gas bubbles which rise and collect under the barrier. Gas accumulation can upend and dislodge barriers over time due to billowing and buoyancy. Porous barrier material facilitates gas exchange and thus prevents billowing and allows benthic fauna to survive underneath the mats. Where management goals are less concerned about benthic infauna and billowing can be reduced or eliminated (e.g. with gas check valves, sealing flaps), non‐porous materials can be used. Rubber non‐porous benthic bottom barrier material was used to kill the Asian clam (C. fluminea) in Lake Tahoe by creating anoxic conditions (Wittmann et al., 2012). Non‐porous benthic bottom barriers typically have anoxic conditions underneath the mats (Wittman et al., 2012). Typical materials used for benthic bottom barriers consist of jute, polyethylene fabric, rubber and fiberglass fabric. Jute is attractive since it is a natural plant product that sinks within a few minutes, slowly decomposes and requires no removal the following season. Jute is highly porous and tends to allow more light underneath than other porous materials such as fiber glass woven mats or polyethylene (unless several layers of the fabric are used) (Caffrey et al., 2010). Polyethylene fabric is less porous and used widely in terrestrial systems as a weed light block barrier. Polyethylene fabric is used for aquatic weed management typically for regions around boat marinas and docks. It tends to be buoyant and subjected to billowing. Porous fiberglass mats have been used in the past with success as a semi barrier by pressing the weed beds down and decomposing them at the roots (Mayer 1978). Sedimentation on top of the barrier materials can contribute to regrowth unless the barriers are removed yearly and redeployed (Mayer, 1978). Several studies showed that benthic bottom barriers were ineffective for eradication due to limited area coverage and rapid post treatment regrowth (Ussery et al., 1997). Benthic bottom barriers are generally not considered economically cost effective for large areas. If water managers are not authorized to use chemical or other mechanical methods, then the economic trade off cost for benthic bottom barriers may provide a viable management option. 54
Economic cost For smaller systems with deployment by the pond or marina owners, cost can be relatively moderate, but redeployment year to year can increase cost. Cost are typically 0.1$ to 0.5$ per square foot area. For private vendor installations, rates are 0.35$ to 1.10$ per square foot. Bailey and Calhoun (2008) calculated actual cost at 2.68$ per square foot with SCUBA labor cost included. Barriers can be made up ahead of time and then divers with SCUBA can be hired to place and secure the barriers on the sediment. Such partial set up may cost 0.3$ to 0.7$ square foot. There is a high potential for barrier uplifting around boat docks, marinas and navigation channels due to strong currents. Uplifted barriers may cause boating and navigation hazards. Continued redeployment may have added cost depending on the site if barriers are uplifted frequently. Where localized management for control is warranted, benthic bottom barriers may provide a good option, however, if the management goal is eradication, then it may be far more costly and difficult. Bailey and Calhoun (2008) suggested that barriers combined hand removal may offer excellent control and potentially eradication for small new infestations in lakes. Eradication of the invasive marine macroalgae Caulerpa taxifolia was achieved over two regions, 104.6 and 2.6 acre sites using both chemical control and nonporous benthic barrier materials in California at a cost of 3.5 Million Dollars $ (Merkel & Associates, 2004). Total area of treatment however was much lower due to patchy Caulerpa taxifolia distribution (0.32 acres). While the cost was high with the Caulerpa taxifolia example, stopping the early infestations at the initial stages prevents long term and more costly economic management options (Anderson, 2007). Year to year deployment is not required if eradication is achieved, however, the infestation needs to be managed at the earliest stages to be cost effective and practical. Benthic bottom barriers have no requirement for permitting for chemical use, greatly reducing cost for small patchy new infestations. Where integrated methods have been used with non‐porous benthic bottom barriers and the chemical containment, the amount of chemicals required for small patchy regions is greatly reduced and minimized the effects on non‐target organisms (Merkel & Associates, 2006). Combining integrated methods such chemical or thermal treatment under benthic bottom barriers has not been widely considered, but appears to have good potential for eradication within new and isolated patchy sites. This approach may prove cost effective for large lake systems and marine sites where whole lake or bay chemical treatments are unfeasible or practical. Development for large‐scale benthic barriers Large‐scale development within California has been demonstrated in Lake Tahoe for the Asian clam using rubber non‐porous benthic bottom material (Wittmann et al., 2012). One hundred foot long rolls, ten feet wide, were deployed from boats onto the lake bottom. Similar control methods were employed in Lake Tahoe’s South Lake Tahoe Keys Marina for aquatic weed control over several acre test sites in 2011 using 55
polyethylene and jute. Management using these larger scale methods have several tradeoffs: the high cost of SCUBA labor may become an economic factor as the method is scaled up for deployment; as with fiberglass and other materials, sedimentation build‐up and regrowth on top of the benthic bottom barrier material will require removal or redeployment; heterogeneous lake, pond, stream, irrigation canal bathymetry can present challenges at larger scales. Reduced deployment treatment times (minutes instead of months) may potentially be possible with thermal blanket barriers with heated water (70 Celsius) to allow cost effective rapid systematic treatments over large areas (under 0.1$ per square foot for fuel cost). Design of weighted barrier materials may also provide some reduction in cost for larger scale deployment. Baffles in rubber barriers may be filled with gravel or sand on site. The weighted rubber barriers could then be positioned over the target area from boats or the shore without the need for SCUBA divers. Boat mounted fabric rolls on wenches were used for deployment to control Lagarosiphon major in Irish lakes (Caffery et al., 2010). Cost for jute runs 0.05$ square foot not including deployment cost and labor. Jute has several advantages over polyethylene: jute slowly biogrades eliminating removal cost, allows more gas permeability which reduces billowing, does minimal harm to the benthic fauna and jute is not buoyant, it will sink within a few minutes of soaking in the water (Caffery et al., 2010). Fundamental Effects and Mechanisms Porous benthic bottom barriers kill and inhibit plant growth by blocking light by stopping or reducing photosynthesis. Jute or fiberglass benthic bottom appears to kill weeds effectively in some systems by compacting existing growth which subsequently rots (Mayer 1978; Caffrey, 2010). While such materials may appear effective for some species, they do not target long‐lived propagules such as Potamogeton turions (Caffrey, 2010). In colder oligotrophic lakes such as Lake Tahoe, (California, USA) benthic bottom barriers have recently been authorized. However, it is not clear if these materials have a significant impact on aquatic weed propagules (Reuter, 2011 pers. comm). There have been some successful eradication projects with Eurasian milfoil (Myriophyllum spicatum) in a Seattle (Washington, USA) reservoir (Zisette, 2001). In Lake George, New York, rapid recolonization occurred (Eichler et al., 1995). Benthic bottom barriers were not successful in Clear Lake, California with Hydrilla (King, 1999). Deployment has been troublesome for smaller ponds for Hydrilla in Yuba County, California, USA (California Department of Food and Agriculture, 2009). Eichler et al. (1995) reported that no plants survived under the nonporous benthic bottom barriers while the porous barriers had plants survive continuously throughout the experiment. In both cases, the treated areas recolonized rapidly. Aquatic macrophytes are generally low light adapted plants that are able to grow at as little as 12 umol m‐² sec‐¹ (Van et al., 1976; Bowes et al., 1977). Aquatic weed propagules are able to survive on starch reserves without light for years in the hydrosoil (Kunii, 1989; Neatherland, 1997; Madsen and Owens, 1998). Wu et al. (2009) suggested that 150‐200 umol m‐² sec‐¹ was most favorable for Potamogeton crispus turion growth and could ameliorate the negative impacts from 56
slight anoxia, however, under strong anoxia, light had little effect. Non‐porous benthic bottom barrier materials will alter the physical and chemical habitat under the barriers by increasing ammonium and eliminating oxygen and light (Ussery et al., 1997). It has been suggested that the anoxic conditions inhibit the sprouting and growth of aquatic weed propagules of Potamogeton crispus turions rather than light (Wu et al., 2009). Strong anoxia in the hydrosoil can lead to the production of microbial phytotoxins such as sulfide, methane and acetic acid (Van Wijck et al., 1992; Terrados et al., 1999). Spencer and Ksander (1995, 1997) and Spencer et al. (2003) demonstrated that Hydrilla, American pondweed (Potamogeton nodosus) winter buds and sago pondweed (Stuckenia pectinatus) tubers were inhibited with dilute acetic acid treatments. Acetic acid appeared to more effectively depolarize the cell membranes, leading to metabolite leakage under anoxic conditions (Spencer and Ksander, 1999). Fermentation is often associated with assumed sucrose production from starch reserves in anaerobic conditions. Bouny and Saglio (1996) suggest that anaerobic tolerance is not due to changes in the rate of fermentation, rather, the ability to sustain fermentation under prolonged anoxia. In plants that are sensitive to anoxic soil conditions such as corn (Zea mays), rapid potentially damaging cytoplasmic acidification occurs (Sachs et al., 1980). Fermentation in aquatic weed propagules may explain why there is an absence of cytoplasmic acidification (Schwartz, 1969; Bouny and Saglio, 1996; Drew, 1997, Jackson and Summer, 2009). Turions of Potamogeton crispus, tubers of sago pondweed (Stuckenia pectinatus), winter buds in American pondweed (Potamogeton nodosus), yelllow pond‐lily (Nuphar advena) and Hydrilla tubers have starch‐laden reserves, similar to rice seed and potato (Yeo et al., 1984; Woolf and Madsen, 2003). In seeds, high starch content have been shown to be particularly tolerant of anoxia due to the ability to maintain higher energy metabolism rates compared to fatty seeds. However, there is wide variation in response to anaerobiosis by aquatic weed progagules (Wu et al., 2009; Summers, 2000). Tolerance of anoxic stress varies with plant species, age, cell type, and acclimation conditions (Drew, 1997). Similarly, there is considerable variation in germination rates amongst starchy seed types in rice systems (Tsuji, 1973; Alpi and Beevers, 1983). Exogenously supplied glucose was shown to stimulate ethanol production rates by threefold in the anoxia ‘intolerant’ rice genotypes, which may suggest intolerance to anoxia may be due to sugar starvation (Gibbs et al. 2000). Potamogeton crispus turions contain 30‐40% starch, while Hydrilla and sago pondweed tubers are higher in starch content (50‐70%), they also are able to withstand long periods of strong anaerobic conditions (4‐7 years) in the hydrosoil whereas Potamogeton crispus turions are able to survive in the water hydrosoil interface for roughly 1‐4 years (Kunii, 1989; Neatherland, 1997; Madsen and Owens, 1998).They may be some differences between aquatic weed propagule response and chemical treatments under non porous benthic barriers compared to the more commonly used porous materials. In rice agricultural systems, Raymond et al. (1985) suggests that seeds may be grouped into classes based on their responses to oxygen concentration. There may be a similar relationship between aquatic macrophyte propagules classes when applying benthic bottom barriers for control. 57
Benthic Bottom Barriers in Lake Tahoe Attempts were made to test jute, polyethylene and rubber benthic bottom materials for control of Potamogeton crispus turions in the field in Lake Tahoe, near the city of South Tahoe (California, USA) in 2010. We deployed 3 barrier types (3.2 meters x 3.2 meters each) at 3 locations (one replicate per location site). These treatments did not show any differences between barrier materials and the control for each site. Jute treatments had a high density of turions attached (200 per meter²) on the outside surface, acting as a substrate but turions were absent outside the barrier areas. In the summer of 2011, we set up another test site with 3 replicates of each barrier treatment and a control at the same location approximately 200 meters north of the Tahoe Key’s marina. However, in early December 2011, large wind driven waves destroyed the experimental set up. In effort to address survival of turions of Potamogeton crispus under benthic bottom barriers in more controlled conditions, bench scale and mesocosm scale experiments were conducted at the University of California, Davis California. Since the benthic bottom barrier effectiveness has been varied in the field, we asked the following questions to better elucidate this issue with turions of Potamogeton crispus. 1‐ Can turions of Potamogeton crispus be killed or inhibited using benthic bottom barriers at the bench and mesocosm scales? 2‐ Will non‐porous barrier materials be more effective at killing or inhibit sprouting of turions of Potamogeton crispus? 3‐ Is there an effect between light and oxygen that influences sprouting of turions of Potamogeton crispus under benthic bottom barriers? These questions are critical to address benthic bottom barrier approaches to aquatic weed propagule management. Methods and Results Methods Turions of Potamogeton crispus were collected from the north end of Fisherman’s Cut near Brannon Island State Park, California in June of 2012 (38.084713, ‐121.646722). Turions were then harvested and then kept in cold water (4C) in the dark until the experimental treatments started. All turions represented a single cohort at the time of harvest. Spencer and Ksander (1997) suggested that variation in propagule size may lead to differences in survivorship, thus two size classes were used at 50 to 100 milligrams and then 150‐250 milligrams. Additionally, some turions were sprouted to ensure viability using the protocols from Sastroutomo (1981) prior to testing and repeated for each set of turions used in each experiment. Turions were covered 3 types 58
of benthic bottom barrier materials; 4 millimeter thick Rubber, Polyethylene fabric and mesh jute fabric. Controls had the same conditions except for barrier material. Table 1 outlines treatments by each barrier type. Post treatment turion viability was based on visual inspection of new sprouts following a vernalization protocol (Sastroutomo, 1981). New sprouts suggest the turions are viable and survived the treatments. Table 1 summarizes the treatments applied to each replicate of Potamogeton crispus turions. The experimental design for bench and mesocosm scale were both complete randomized designs with 4 replicates for each of the 3 treatments and a control. Large and small turion size classes were initially split into groups to detect potential differences between size classes and efficacy of treatments, however, both data set means were not significantly different and normally distributed based on Chi squared test (Chi squared X²= 2.7833 with df= 7), and therefore combined. Each experiment was repeated. The repeated experiments were also combined based in the similar means between each experiment (Chi squared X²=27.866 with df= 15). To ensure homogeneity of variances, a Levene’s test was performed prior to the ANOVA test. Computations of significant differences (Tukey‐Kramer HSD, p ˂0.05) were based on analyses of variance. Means quoted in the text carry their associated standard errors (SE). Statistical analyses were performed using SAS JMP 8.0 statistical software (2009). Experiment 1: Bench scale Experiments were performed at Robbins Hall, University of California Davis, California in June, 2012. 200 milliliters of deionized water was placed into 250 milliliter flask with 10 milligrams of osmocoat slow release fertilizer. Lighting was supplied by fluorescent fixtures using a pair of new Phillips 865 bulbs 32 watt bulbs at 140 umol m‐² sec‐¹. Flasks were covered with 3 types of benthic bottom materials: jute with 60 millimeter pore mesh size, Polyethylene weed block fabric and 4 millimeter rubber sheeting. Light meter readings were measured under each barrier type (jute= 24 umol m‐² sec‐¹, polyethylene= 3 umol m‐² sec‐¹and rubber= 0 umol m‐² sec‐¹ respectively) using an Apogee light meter (model: MQ‐200 Quantum with a SQ ‐120 sensor). Photoperiod was 12 hours for the 8‐week treatment exposure. Rubber barrier flask water was purged of oxygen using nitrogen gas and then quickly sealed to replicate anoxic conditions. The control, jute and polyethylene treatments were left unsealed. Temperature was stable at 26 Celsius in the laboratory. Turions were removed after 8 weeks and then placed in a growth chamber with the following conditions: photoperiod: 12 hour daylight: 12 hour night; photosynthetic photon flux (area) density: 1126 µmole sec‐¹ m‐² light intensity and temperature: 5 degrees Celsius for two weeks, followed by 1 week at 30 degrees Celsius (Sastroutomo, 1981). Bench scale treatments showed the rubber barrier treatment had a highly significant effect on sprouting turions of Potamogeton crispus with a p‐value ˂ 0.0001 (Figure 1). Rubber (no light, no oxygen) benthic bottom barrier treatments had a 48.4375% proportion sprouting (SE= 10.5743%; n=16). The control (light and oxygen) and jute (reduced light, oxygen) treatments were the same sprouting at 100% (SE= 0; n=16), 59
whereas polyethylene benthic bottom barrier treatments (no light, oxygen) had a small non‐significant reduction in % sprouting of 96.875% (SE=2.13478%; n=16). Means comparison Tukey‐Kramer HSD test set for α at 0.05 showed rubber benthic bottom treatments where significantly different from polyethylene, jute and the control (Table 2). Experiment 2: Mesocosm mesocosms Experiments were performed at California’s Department of Food and Agriculture’s “B” street facility in Sacramento in June, 2012. Mesocosm conditions were as follows: temperatures varied from 22C to 35C and light at noon was 1210 umol m‐² sec‐¹ on June 21st, 2012. Mesocosms were made from fiberglass and were 200 cm x 25 cm x 125cm in volume (516 liters). Water depth was set at 25 cm and fresh carbon filtered tap water was set to slowly continuously drip into the mesocosms. Tap water had an alkalinity of 37 mg/l and a pH of 7.81. Temperatures in the water ranged from 24C to 29C range during the experiment. Pots size was 15 centimeters in diameter x 6 centimeters deep plastic trays with a 3 centimeter deep sediment. Replicates had 4 small turions and 4 large turions placed on the sediment in the middle of each pot. Each pot was covered with each of the 3 types of benthic bottom materials: jute with 60 millimeter pore mesh size, Polyethylene weed block fabric and 5 millimeter rubber sheeting. Light meter readings were measured under each barrier type (jute= 68 umol m‐² sec‐¹, polyethylene= 10 umol m‐² sec‐¹and rubber= 0 umol m‐² sec‐¹ respectively). Sediments were collected from Owl Harbor (Twitchell Island, Isleton, CA, USA), screened (63 millimeter mesh size) to remove larger particles and homogenized in a mixer. Sediment reduction oxidation potential was measured at 3 centimeters for all. Platinum electrodes were constructed using the methods from Reddy and DeLaune (2008) using a calomel reference electrode and an Orion meter (Model 290A). Four electrodes were placed in one of each treatment type and left in place for the duration of the experiment. Millivolt readings were recorded weekly. 10 Grams of osmocoat fertilizer (15‐15‐15) was added to each mesocosm. Mesocosm scale treatments also showed the rubber barrier treatment had a stronger highly significant effect on sprouting turions of Potamogeton crispus with a p‐value ˂ 0.0001 (Figure 2) compared to the other treatments. Rubber (no light, no oxygen) benthic bottom barrier treatments had a 29.6875% proportion sprouting (SE= 6.1317%; n=16). The control (light and oxygen) had a 98.4375% proportion sprouting (SE= 1.5625%; n=16). Jute (low light, oxygen) benthic bottom barrier treatments had a 71.875% proportion sprouting (SE= 4.49247%; n=16). Jute benthic bottom barriers rapidly colonized periphyton algae at the mesocosm scale, reducing light after 2 weeks into the test. Polyethylene (no light, oxygen) benthic bottom barrier treatments had a 70.3125% proportion sprouting (SE= 4.6875%; n=16). Means comparison Tukey‐Kramer HSD test set for α at 0.05 showed rubber benthic bottom treatments where significantly different from polyethylene, jute and the control (Table 4). Additionally, means for jute and polyethylene were significantly different from both the control and 60
the rubber benthic bottom barrier treatments (Table 3). Jute and polyethylene benthic bottom barrier treatments were not significantly different from each other with a p‐
value ˂ 0.9949(Table 3). Summarized reduction oxidation potentials of soil in the mesocosm experiment are presented in the appendix in Figure 5. Rapid reduction occurred in all soil treatments, however, the control had consistently less reduction values compared to the other treatments with barriers after the second week which also correlated with the rapid growth of periphyton on the barrier (the soil for the control) surface on all treatment surfaces (Figure 5). The control treatments had no barriers and the algae grew in and on the sediment directly. Under the barriers for the benthic bottom barriers, there was no periphyton growth, only on top of the barriers. Discussion We found that the nonporous benthic bottom barrier materials were most effective at inhibiting sprouting in turions of Potamogeton crispus (Figure 1 and 2). Porous benthic bottom barriers (jute and polyethylene) had no effect on sprouting in the bench scale test without sediments, while the mesocosm experiments with sediments had significant differences compared to the control (Figure 2). There may be several reasons for the differences in the bench and mesocosm scale experiments. First, the bench scale experiment controls lacked algae growth in the flask, while the mesocosms had dense periphyton growth occurring after two weeks into the experiment. The sediments covered with algae may have high oxygen concentrations near the water soil interface due to photosynthesis while the jute and polyethylene barrier treatments acted as an algal shield, protecting the turions from algal growth and elevated oxygen. Second, the thick coverage of algae on the porous barriers acted as its own barrier by reducing gas exchange. Thirdly, the reduction in light by the periphyton growth may have enhanced the effects of slight anoxic. Wu et al. (2009) found moderate light helped reduce the impact of slight anoxic. Fourth, the addition of sediments to mesocosms reduced the oxygen concentrations. All four effects could have potentially combined to reduce the proportion sprouting through anoxia in both the porous and non‐porous treatments at the mesocosm scale. Oxygen sensors placed at the immediate location of the turions being treated would be more informative. The effects of light appear to be minimal based on the experiments. This concurs with Wu et al., (2009) results. Wu et al., (2009) used sucrose additions to the sediments to reduce oxygen levels in mesocosm aquariums to replicate field conditions rather than non‐porous barrier materials. We report a similar result with different methodology (mean for mesocosm test: 29.69% and a mean roughly 20% sprouting under high anoxia for Wu et al. (2009)), however Wu et al., (2009) controls had relatively low proportion sprouting (70%) comparative to our control data (100%). This difference may be due to the time of year that turions were collected between the experiments. If turions are collected in autumn, they tend to be brown and smaller, these have a lower proportion of sprouting (mean = 67.5%), while green turions collected in the spring have the highest proportion of sprouting (mean 100%) (Sastroutomo, 1981). We 61
collected only spring cohort Potamogeton crispus turions for this experiment. Additionally, by using the bench scale treatments, we removed sediments, bacterial interactions, metabolites and algae as dependent variables, illustrating the non‐
significant effect of light at an intensity of 140 umol m‐² sec‐¹ which is roughly the range of Wu et al. (2009) “optimal” range of 10% full sunlight for sprouting under mild anoxia. Aquatic sediment anaerobiosis leads to the production of microbial phytotoxins such as sulfide, methane and acetic acid (Van Wijck et al., 1992; Terrados et al., 1999). Non porous benthic bottom barriers enhance anaerobic conditions in the sediments (Wittman et al., 2010). We noted that stressed Potamogeton crispus plants sealed in jars or bags tended to produce weak elongated smaller turions late in the season (autumn), and also in culture in the mesocosm and storage inside dark 4C refrigeration. Researchers found that in Potamogeton distinctus, anoxia contributed to elongated turions and this effect was enhanced additions of 2,4‐dichlorophenoxyacetic acid (2,4‐
D) and suppressed by additions of sorbitol, 2‐deoxyglucose (2‐dGlc) and abscisic acid (ABA)(Harada et al., 2005). Elongated turions also occurred with treatments of gibberellic acid (GA3) (Wang et al., 2012). Wang et al. (2012) reported inhibition (complete or partial) of turion formation in Potamogeton crispus with treatments of 6‐
benzyladenine (6‐BA), a cytokinin substances and gibberellic acid (GA3), propagule starch allocation was inhibited and cytokinin was a more effective substance in regulating propagule differentiation. Summers and Jackson (2006) found anaerobic stem growth in tubers of Stuckenia pectinata was inhibited by exogenous abscisic acid (ABA) while exogenous indole‐3‐acetic acid (IAA) promoted stem elongation under anaerobic conditions. Such effects by plant growth regulators may offer insight into how anoxia can help provide potential tools for management of aquatic weed propagules. Many aquatic herbicides target plant growth regulators and the combined effects under non‐porous benthic bottom barriers may offer an effective combination for integrated management under some conditions. The mechanism how aquatic weed propagules are able to support respiration under anoxia may have some implications for further study. For example, Summers et al. (2000) demonstrated minimal acidification in the cytoplasm (≤0.2 pH) in absence of oxygen for with Stukenia pectinata tubers. In non‐flood tolerant plants like corn and pea, pH declines of 0.5–0.6 (Fox et al., 1995) and 0.7 were observed post anoxia (Summers et al., 2000). Recovery also occurred far more rapidly with Stukenia pectinata than with pea (Pisum sativum) internodes following post treatment re‐
oxygenation (Summers et al., 2000). Jackson and Summer (2000) suggest that in sago pondweed (Stuckenia pectiantus) tubers are able to avoid toxic effects of acetaldehyde accumulation and potentially damaging acidification of the cytoplasm under anoxia by vigorous fermentation via a large Pasteur effect in the shoot. Rice (Oryza sativa) transgenic mutant lines that lack appreciable alcohol dehydrogenase (ADH) levels die under anoxia (Matsumura et al., 1998). Mutants overexpressing ADH did not appear to have enhanced survival under anoxic conditions (Agarwal et al., 2007). This suggests that ADH activity in rice is often sufficient to guarantee efficient fermentative 62
metabolism. Since Stukenia pectinata suppresses post‐anoxic acetaldehyde accumulation thereby avoiding possible self‐poisoning (Summers et al., 2000), alcohol dehydrogenase (ADH) maybe a potential target for ADH inhibiting herbicides such as triazines for aquatic macrophyte propagules high in starch using non‐porous barriers. Such treatments may only require a very small amount of herbicide and brief exposure to be effective. It is clear that benthic bottom barriers alone cannot eradicate 100% of the propagules on their own, however, due to specific stressors caused by anoxia, more effective targeted delivery and reduced chemical application could potentially be used under nonporous barriers. For example, applying solid formulations of naturally occurring phytoxins such as dilute acetic acid, herbicides such as triazines or synthetic plant growth regulators that target specific chemical metabolic pathways that are stressed under anoxia may potentially be effective. Such approaches may be effective in persistent small patchy sites in larger rivers, irrigation canals, marinas and lake systems where whole lake or river treatments are impractical. Investigating synergism occurring between non porous anoxic benthic bottom barriers and chemical treatments may offer another tool for aquatic weed propagule management. We conclude that nonporous benthic bottom barriers need to be used in conjunction with other integrated methods for eradication of Potamogeton crispus propagules. Such combined treatments may be able to selectively take advantage of the anoxic conditions that will enhance efficacy. References Agarwal S., Kapoor A., Lakshmi O.S., Grover A. (2007) Production and phenotypic analysis of rice transgenics with altered levels of pyruvate decarboxylase and alcohol dehydrogenase proteins. Plant Physiology and Biochemistry, 45, 637–646. Anderson, LW (2007). Control of invasive seaweeds. Botanica Marina, 50(5/6), 418‐437. Bailey JE, & Calhoun JK 2008. Comparison of Three Physical Management Techniques for Controlling Variable‐leaf Milfoil in Maine Lakes. Journal of Aquatic Plant Management, 46(2), 163. Born SM, Wirth TL, Brick EM, Peterson JP 1973. Restoring the recreational potential of small impoundments. The Marion Millpond experience. Department of Natural Resources, Madison, WI. Technical Bulletin No. 71. Bouny J.M., Saglio P.H. (1996) Glycolytic flux and hexokinase activities in anoxic maize root tips acclimated by hypoxic pretreatment. Plant Physiology, 111, 187–194. Bowes, G. A. S. Holaday, T. K. Van and W. T. Haller. 1977. Photosynthetic and photorespiratory carbon metabolism in aquatic plants. In Proceedings 4th Int. Congress of Photosynthesis, Reading (UK) pp. 289‐298. Caffrey JM 1993. Aquatic plant management in relation to Irish recreational fisheries development. Journal of Aquatic Plant Management 31: 162‐168. Caffrey, JM Millane, M Evers, S Moran, H & Butler, M 2010. A novel approach to aquatic weed control and habitat restoration using biodegradable jute matting. 63
Aquatic Invasions 5: 123‐129. The California Department of Food and Agriculture Hydrilla Eradication Program annual progress report 2011. Eakin HL, Barko JW 1995. Evaluation of the effect of benthic barrier placement on sediment physical and chemical conditions. U.S. Army Engineer Waterways Experiment Station, Vicksburg, MS, Technical report A‐95‐2. Eichler LW, Bombard RT, Sutherland JW, Boylen CW 1995. Recolonization of the littoral zone by macrophytes following the removal of benthic barrier material. Journal of Aquatic Plant Management 33: 51‐54. Gibbs J., Morrell S., Valdez A., Setter T.L., Greenway T. 2000 Regulation of alcoholic fermentation in coleoptiles of two rice cultivars differing in tolerance to anoxia. Journal of Experimental Botany, 51, 785–796. Gunnison D., Barko JW 1992. Factors influencing gas evolution beneath a benthic barrier. Journal of Aquatic Plant Management 30: 23‐28. King, P. 1999. Clear Lake Aquatic Weed Pilot Project. Greater Lakeport Chamber of Commerce and Lake County Department of Public Works, Water Resources Division. Kunii, H., 1982. Life cycle and growth of Potamogeton crispus L. in a shallow pond, Ojaga‐ika. Bot. Mag. (Tokyo) 95, 109–124. Madsen, J. D. and C. S. Owens. 1998. Seasonal biomass allocation in dioecious Hydrilla. J. Aquat. Plant Manage. 36:138‐145. Madsen J.D. 2000. Advantages and disadvantages of aquatic plant management techniques. U.S. Army Engineer Research and Development Center. Vicksburg, MS, Final report, ERDC/EL MP‐00‐1, 31 pp. Mayer JR 1978. Aquatic weed management by benthic semibarriers. Journal of Aquatic Plant Management 16: 31‐33. Merkel & Associates, Inc 2006. Final report on the eradication of the invasive seaweed Caulerpa taxifolia from Agua Hedionda Lagoon and Huntington Harbour, California. Southern California Caulerpa Action Team, San Diego. Murphy K.J. 1988. Aquatic weed problems and their management: a review. II. Physical control measures. Crop Protection 7: 283‐302. Netherland, M. D. (1997). Turion ecology of Hydrilla. Journal of Aquatic Plant Management, 35, 1‐10. Nichols, S. A. 1974. Mechanical and habitat manipulation for aquatic plant management; A review of techniques. Technical Bulletin No. 77, Department of Natural Resources, Madison, WI. Mayer, J. R. 1978. Aquatic weed management by benthic semi‐barriers. Journal of Aquatic Plant Management 21: 31‐33. Raymond, P., Al‐ani, A., Pradet, A. 1985. ATP production by respiration and fermentation, and energy charge during aerobiosis and anaerobiosis in twelve fatty and starchy germinating seeds. Plant Physiol 79: 879‐884. Reddy, K R, & DeLaune, R D 2008. Biogeochemistry of wetlands: science and applications. CRC. Reuter, John personal communication 2011. Acting Associate Director, Tahoe 64
Environmental Research Center Director, Lake Tahoe Interagency Monitoring Program, Scientific Director, Lake Tahoe TMDL Program. Sachs MM, Freeling M, Okimoto R.1980. The anaerobic proteins of maize. Cell20: 761–
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Figure 1: Proportion sprouting % of Potamogeton crispus turions after 8 week exposure under benthic bottom barrier treatments at the bench scale. Means are ± 1 SE (n= 16). Data is combined from repeated experiments and size classes. Levels not connected by same letter are significantly different (Tukey HDS; p≤0.05). 67
Figure 2: Proportion sprouting % of Potamogeton crispus turions versus benthic bottom barrier treatment material for 8 week exposure at the mesocosm scale. Means are ± 1 SE (n= 16). Data is combined from repeated experiments and size classes. Levels not connected by same letter are significantly different (Tukey HDS; p≤0.05). 68
Figure 3. Variance test for benthic bottom barrier treatments at the bench scale.
Figure 4. Variance test for benthic bottom barrier treatments at the mesocosm scale. 69
fi Figure 5. Mesocosm Overlay Plot for Reduction Oxidation Potential by Time (8 week) Based on Benthic Bottom Barrier Material Treatment in Mesocosm experiment. Depth of redox probes = 3cm. 70
71
Table 2. One way Analysis of Proportion sprouting % By Treatment at the bench scale
Oneway Analysis of Proportion sprouting % By Treatment (bench scale)
Summary of Fit
Rsquare 0.523889
Adj Rsquare
0.500083
Root Mean Square Error 0.215753
Mean of Response
0.863281
Observations (or Sum Wgts)
64
Analysis of Variance
Source DF
Sum of Squares Mean Square
F Ratio Prob > F
Treatment
3
3.0732422
1.02441 22.0070 <.0001*
Error
60
2.7929688
0.04655
C. Total 63
5.8662109
Means for Oneway Anova
Level
Number Mean
Control 16
1.00000
Jute
16
1.00000
Ploy
16
0.96875
Rubber 16
0.48438
Std Error
0.05394 0.89211
0.05394 0.89211
0.05394 0.86086
0.05394 0.37648
Lower 95%
1.1079
1.1079
1.0766
0.5923
Upper 95%
Std Error uses a pooled estimate of error variance
Means Comparisons
Comparisons for all pairs using Tukey-Kramer HSD
q*
Alpha
2.64252 0.05
Abs(Dif)-LSD
Control Jute
Ploy
Rubber
Control -0.20157 -0.20157 -0.17032 0.314053
Jute
-0.20157 -0.20157 -0.17032 0.314053
Ploy
-0.17032 -0.17032 -0.20157 0.282803
Rubber 0.314053
0.314053
0.282803
-0.20157
Positive values show pairs of means that are significantly different.
Level
Control A
Jute
A
Ploy
A
Rubber
B
Mean
1.0000000
1.0000000
0.9687500
0.4843750
Levels not connected by same letter are significantly different.
Level
Control
Jute
Ploy
Control
Jute
Jute
- Level
Rubber
Rubber
Rubber
Ploy
Ploy
Control
Difference
0.5156250
0.5156250
0.4843750
0.0312500
0.0312500
0.0000000
Std Err Dif
0.0762803
0.0762803
0.0762803
0.0762803
0.0762803
0.0762803
Lower CL
0.314053
0.314053
0.282803
-0.170322
-0.170322
-0.201572
.
72
Upper CL
0.7171973
0.7171973
0.6859473
0.2328223
0.2328223
0.2015723
p-Value
<.0001*
<.0001*
<.0001*
0.9766
0.9766
1.0000
Table 2 Contiuned.
Level
Control
Jute
Ploy
Rubber
Test
O'Brien[.5]
Brown-Forsythe
Levene
Count
Std Dev
16
16
16
16
0.0000000
0.0000000
0.0853913
0.4229731
F Ratio
31.7109
40.2941
44.0271
MeanAbsDif to
Mean
0.0000000
0.0000000
0.0546875
0.3613281
DFNum
3
3
3
73
DFDen
60
60
60
MeanAbsDif to
Median
0.0000000
0.0000000
0.0312500
0.3593750
Prob > F
<.0001*
<.0001*
<.0001*
Table 3. One Way Analysis of Proportion sprouting % by Treatment at the Mesocosm Scale
Oneway Anova
Summary of Fit
Rsquare
Adj Rsquare
Root Mean Square Error
Mean of Response
Observations (or Sum Wgts)
Analysis of Variance
Source
Treatment
Error
C. Total
DF
3
60
63
Means for Oneway Anova
Level
Number
Control
16
Jute
16
Ploy
16
Rubber
16
0.661925
0.645021
0.181322
0.675781
64
Sum of Squares
3.8623047
1.9726563
5.8349609
Mean
0.984375
0.718750
0.703125
0.296875
Mean Square
1.28743
0.03288
Std Error
0.04533
0.04533
0.04533
0.04533
Lower 95%
0.89370
0.62808
0.61245
0.20620
F Ratio
39.1584
Prob > F
<.0001*
Upper 95%
1.0750
0.8094
0.7938
0.3875
Std Error uses a pooled estimate of error variance
Means Comparisons
Comparisons for all pairs using Tukey-Kramer HSD
q*
Alpha
2.64252
0.05
Abs(Dif)-LSD
Control
Jute
Ploy
Rubber
Control
-0.1694
0.096221
0.111846
0.518096
Jute
0.096221
-0.1694
-0.15378
0.252471
Ploy
0.111846
-0.15378
-0.1694
0.236846
Rubber
0.518096
0.252471
0.236846
-0.1694
Positive values show pairs of means that are significantly different.
Level
Control
Jute
Ploy
Rubber
A
B
B
C
Mean
0.98437500
0.71875000
0.70312500
0.29687500
Levels not connected by same letter are significantly different.
Level
Control
Jute
Ploy
Control
Control
Jute
- Level
Rubber
Rubber
Rubber
Ploy
Jute
Ploy
Difference
0.6875000
0.4218750
0.4062500
0.2812500
0.2656250
0.0156250
Std Err Dif
0.0641069
0.0641069
0.0641069
0.0641069
0.0641069
0.0641069
74
Lower CL
0.518096
0.252471
0.236846
0.111846
0.096221
-0.153779
Upper CL
0.8569040
0.5912790
0.5756540
0.4506540
0.4350290
0.1850290
p-Value
<.0001*
<.0001*
<.0001*
0.0003*
0.0006*
0.9949
Table 3 Continued
Level
Control
Jute
Ploy
Rubber
Test
O'Brien[.5]
Brown-Forsythe
Levene
Count
Std Dev
16
16
16
16
0.0625000
0.1796988
0.1875000
0.2452677
F Ratio
5.5639
7.0188
9.1762
MeanAbsDif to
Mean
0.0292969
0.1367188
0.1523438
0.2089844
DFNum
3
3
3
75
DFDen
60
60
60
MeanAbsDif to
Median
0.0156250
0.1250000
0.1406250
0.2031250
Prob > F
0.0020*
0.0004*
<.0001*
Supportive Figures Figure 6. Mesocosm Reduction Oxidation Potential by Time (8 week) for the Control Treatment in the Sediment Based on Benthic Bottom Barrier Material Treatment in Mesocosm Experiment. Depth of Redox Probes = 3cm. 76
Figure 7. Mesocosm Reduction Oxidation Potential by Time (8 week) for the Jute
Treatment in the Sediment Based on Benthic Bottom Barrier Material Treatment in
Mesocosm Experiment. Depth of Redox Probes = 3cm.
77
Figure 8. Mesocosm Reduction Oxidation Potential by Time (8 week) for the
Polyethylene Treatment in the Sediment Based on Benthic Bottom Barrier Material
Treatment in Mesocosm Experiment. Depth of Redox Probes = 3cm.
78
Figure 9. Mesocosm Reduction Oxidation Potential by Time (8 week) for the Rubber
Treatment in the Sediment Based on Benthic Bottom Barrier Material Treatment in
Mesocosm Experiment. Depth of Redox Probes = 3cm.
79
Table 4 Oneway Analysis of control By Time for Redox (weeks after flooding).
Oneway Anova
Summary of Fit
Rsquare
Adj Rsquare
Root Mean Square Error
Mean of Response
Observations (or Sum Wgts)
0.987855
0.984256
18.87115
55.97222
36
Analysis of Variance
Source
DF Sum of Squares Mean Square F Ratio Prob > F
Time (weeks after flooding) 8
782079.72
97760.0 274.5138 <.0001*
Error
27
9615.25
356.1
C. Total
35
791694.97
Means for Oneway Anova
Level
Number
0
4
1
4
2
4
3
4
4
4
5
4
6
4
7
4
8
4
Mean
407.75
221.75
46.75
-28.75
-46.50
-45.50
-24.25
-7.75
-19.75
Std Error
9.4356
9.4356
9.4356
9.4356
9.4356
9.4356
9.4356
9.4356
9.4356
Lower 95%
388.4
202.4
27.4
-48.1
-65.9
-64.9
-43.6
-27.1
-39.1
Upper 95%
427.1
241.1
66.1
-9.4
-27.1
-26.1
-4.9
11.6
-0.3898
Std Error uses a pooled estimate of error variance
Means Comparisons
Comparisons for all pairs using Tukey-Kramer HSD
q*
Alpha
3.36470
0.05
Abs(Dif)-LSD
0
1
2
7
8
6
3
5
4
0
-44.8983
141.1017
316.1017
370.6017
382.6017
387.1017
391.6017
408.3517
409.3517
1
141.1017
-44.8983
130.1017
184.6017
196.6017
201.1017
205.6017
222.3517
223.3517
2
316.1017
130.1017
-44.8983
9.6017
21.6017
26.1017
30.6017
47.3517
48.3517
7
370.6017
184.6017
9.6017
-44.8983
-32.8983
-28.3983
-23.8983
-7.1483
-6.1483
8
382.6017
196.6017
21.6017
-32.8983
-44.8983
-40.3983
-35.8983
-19.1483
-18.1483
6
387.1017
201.1017
26.1017
-28.3983
-40.3983
-44.8983
-40.3983
-23.6483
-22.6483
Positive values show pairs of means that are significantly different.
Level
0
1
2
7
8
6
3
5
4
A
B
C
D
D
D
D
D
D
Mean
407.7500
221.7500
46.7500
-7.7500
-19.7500
-24.2500
-28.7500
-45.5000
-46.5000
Levels not connected by same letter are significantly different.
80
3
391.6017
205.6017
30.6017
-23.8983
-35.8983
-40.3983
-44.8983
-28.1483
-27.1483
5
408.3517
222.3517
47.3517
-7.1483
-19.1483
-23.6483
-28.1483
-44.8983
-43.8983
4
409.3517
223.3517
48.3517
-6.1483
-18.1483
-22.6483
-27.1483
-43.8983
-44.8983
Table 5 Oneway Analysis of Jute By Time for Redox (weeks after flooding).
Oneway Anova
Summary of Fit
Rsquare
Adj Rsquare
Root Mean Square Error
Mean of Response
Observations (or Sum Wgts)
0.993051
0.990993
20.01944
-47
36
Analysis of Variance
Source
DF Sum of Squares Mean Square F Ratio Prob > F
Time (weeks after flooding) 8
1546491.0
193311 482.3406 <.0001*
Error
27
10821.0
401
C. Total
35
1557312.0
Means for Oneway Anova
Level
Number
0
4
1
4
2
4
3
4
4
4
5
4
6
4
7
4
8
4
Mean
399.00
227.50
28.00
-148.25
-186.75
-185.75
-191.00
-178.50
-187.25
Std Error
10.010
10.010
10.010
10.010
10.010
10.010
10.010
10.010
10.010
Lower 95%
378.5
207.0
7.5
-168.8
-207.3
-206.3
-211.5
-199.0
-207.8
Upper 95%
419.5
248.0
48.5
-127.7
-166.2
-165.2
-170.5
-158.0
-166.7
Std Error uses a pooled estimate of error variance
Means Comparisons
Comparisons for all pairs using Tukey-Kramer HSD
q*
Alpha
3.36470
0.05
Abs(Dif)-LSD
0
1
2
3
7
5
0
-47.6303 123.8697 323.3697 499.6197 529.8697 537.1197
1
123.8697 -47.6303 151.8697 328.1197 358.3697 365.6197
2
323.3697 151.8697 -47.6303 128.6197 158.8697 166.1197
3
499.6197 328.1197 128.6197 -47.6303 -17.3803 -10.1303
7
529.8697 358.3697 158.8697 -17.3803 -47.6303 -40.3803
5
537.1197 365.6197 166.1197 -10.1303 -40.3803 -47.6303
4
538.1197 366.6197 167.1197 -9.1303 -39.3803 -46.6303
8
538.6197 367.1197 167.6197 -8.6303 -38.8803 -46.1303
6
542.3697 370.8697 171.3697 -4.8803 -35.1303 -42.3803
Positive values show pairs of means that are significantly different.
Level
Mean
0
A
399.0000
1
B
227.5000
2
C
28.0000
3
D
-148.2500
7
D
-178.5000
5
D
-185.7500
4
D
-186.7500
8
D
-187.2500
6
D
-191.0000
Levels not connected by same letter are significantly different.
81
4
538.1197
366.6197
167.1197
-9.1303
-39.3803
-46.6303
-47.6303
-47.1303
-43.3803
8
538.6197
367.1197
167.6197
-8.6303
-38.8803
-46.1303
-47.1303
-47.6303
-43.8803
6
542.3697
370.8697
171.3697
-4.8803
-35.1303
-42.3803
-43.3803
-43.8803
-47.6303
Table 6 Oneway Analysis of Polyethylene By Time for Redox (weeks after flooding).
Oneway Anova
Summary of Fit
Rsquare
Adj Rsquare
Root Mean Square Error
Mean of Response
Observations (or Sum Wgts)
0.996023
0.994844
15.63176
-55.1111
36
Analysis of Variance
Source
DF Sum of Squares Mean Square F Ratio Prob > F
Time (weeks after flooding) 8
1652242.1
206530 845.2167 <.0001*
Error
27
6597.5
244
C. Total
35
1658839.6
Means for Oneway Anova
Level
Number
Mean
Std Error
0
4
418.25
7.8159
1
4
212.75
7.8159
2
4
20.50
7.8159
3
4
-201.00
7.8159
4
4
-202.00
7.8159
5
4
-186.75
7.8159
6
4
-186.75
7.8159
7
4
-186.75
7.8159
8
4
-184.25
7.8159
Std Error uses a pooled estimate of error variance
Lower 95%
402.2
196.7
4.5
-217.0
-218.0
-202.8
-202.8
-202.8
-200.3
Upper 95%
434.3
228.8
36.5
-185.0
-186.0
-170.7
-170.7
-170.7
-168.2
Means Comparisons
Comparisons for all pairs using Tukey-Kramer HSD
q*
Alpha
3.36470
0.05
Abs(Dif)-LSD
0
1
2
8
7
5
0
-37.1911 168.3089 360.5589 565.3089 567.8089 567.8089
1
168.3089 -37.1911 155.0589 359.8089 362.3089 362.3089
2
360.5589 155.0589 -37.1911 167.5589 170.0589 170.0589
8
565.3089 359.8089 167.5589 -37.1911 -34.6911 -34.6911
7
567.8089 362.3089 170.0589 -34.6911 -37.1911 -37.1911
5
567.8089 362.3089 170.0589 -34.6911 -37.1911 -37.1911
6
567.8089 362.3089 170.0589 -34.6911 -37.1911 -37.1911
3
582.0589 376.5589 184.3089 -20.4411 -22.9411 -22.9411
4
583.0589 377.5589 185.3089 -19.4411 -21.9411 -21.9411
Positive values show pairs of means that are significantly different.
Level
0
1
2
8
7
5
6
3
4
A
B
C
D
D
D
D
D
D
Mean
418.2500
212.7500
20.5000
-184.2500
-186.7500
-186.7500
-186.7500
-201.0000
-202.0000
Levels not connected by same letter are significantly different.
82
6
567.8089
362.3089
170.0589
-34.6911
-37.1911
-37.1911
-37.1911
-22.9411
-21.9411
3
582.0589
376.5589
184.3089
-20.4411
-22.9411
-22.9411
-22.9411
-37.1911
-36.1911
4
583.0589
377.5589
185.3089
-19.4411
-21.9411
-21.9411
-21.9411
-36.1911
-37.1911
Table 7 Oneway Analysis of Jute By Time for Redox (weeks after flooding).
Oneway Anova
Summary of Fit
Rsquare
Adj Rsquare
Root Mean Square Error
Mean of Response
Observations (or Sum Wgts)
0.993315
0.991335
19.5479
-56.5278
36
Analysis of Variance
Source
DF Sum of Squares Mean Square F Ratio Prob > F
Time (weeks after flooding) 8
1533127.7
191641 501.5199 <.0001*
Error
27
10317.2
382
C. Total
35
1543445.0
Means for Oneway Anova
Level
Number
Mean
Std Error
0
4
395.00
9.7739
1
4
208.50
9.7739
2
4
15.75
9.7739
3
4
-185.00
9.7739
4
4
-192.75
9.7739
5
4
-196.25
9.7739
6
4
-191.50
9.7739
7
4
-185.00
9.7739
8
4
-177.50
9.7739
Std Error uses a pooled estimate of error variance
Lower 95%
374.9
188.4
-4.3
-205.1
-212.8
-216.3
-211.6
-205.1
-197.6
Upper 95%
415.1
228.6
35.8
-164.9
-172.7
-176.2
-171.4
-164.9
-157.4
Means Comparisons
Comparisons for all pairs using Tukey-Kramer HSD
q*
Alpha
3.36470
0.05
Abs(Dif)LSD
0
1
2
8
3
7
6
4
5
Level
0
1
2
8
3
7
6
4
5
0
1
2
8
-46.5084
139.9916
332.7416
525.9916
533.4916
533.4916
539.9916
541.2416
544.7416
139.9916
-46.5084
146.2416
339.4916
346.9916
346.9916
353.4916
354.7416
358.2416
332.7416
146.2416
-46.5084
146.7416
154.2416
154.2416
160.7416
161.9916
165.4916
525.9916
339.4916
146.7416
-46.5084
-39.0084
-39.0084
-32.5084
-31.2584
-27.7584
A
B
C
D
D
D
D
D
D
Mean
395.0000
208.5000
15.7500
-177.5000
-185.0000
-185.0000
-191.5000
-192.7500
-196.2500
Levels not connected by same letter are significantly different.
83
3
7
6
533.4916 533.4916
346.9916 346.9916
154.2416 154.2416
-39.0084 -39.0084
-46.5084 -46.5084
-46.5084 -46.5084
-40.0084 -40.0084
-38.7584 -38.7584
-35.2584 -35.2584
539.9916
353.4916
160.7416
-32.5084
-40.0084
-40.0084
-46.5084
-45.2584
-41.7584
Photograph: Drip irrigation overflow mesocosms at the CDFA’s “B” street facility in
Sacramento California, USA. Water was dripped slowly into the mesocosms and then
overflowed onto the ground below.
84