Stream Orchid (Epipactis Gigantea) Monitoring Report 2020 Growing Season

STREAM ORCHID (EPIPACTIS GIGANTEA) MONITORING REPORT 2020 GROWING SEASON

submitted: Filoha Meadows Nature Preserve January 2021 Pitkin County, Colorado

Prepared for: Pitkin County Open Space and Trails 530 East Main Street, Suite 201 Aspen, CO 81611

Prepared by: Peak Ecological Services, LLC 301 Boulder Canyon Drive, PO Box 827 Nederland, Colorado 80466

STREAM ORCHID (EPIPACTIS GIGANTEA) MONITORING REPORT 2020 GROWING SEASON Filoha Meadows Nature Preserve Pitkin County, Colorado

revision date: March 19, 2021

Summary The stream orchid or false helleborine orchid (Epipactis gigantea) is a critically imperiled plant that occurs at Filoha Meadows Nature Preserve. This report documents the third year of a long-term monitoring study tracking annual populations trends of this orchid and investigating the effect of noxious weed control actions on the persistence of this rare plant species. In 2020, the density of the orchid population at Filoha was slightly less than in 2019, but still higher than 2018. Orchid density appears to be highly correlated with precipitation patterns the preceding winter and spring, with lower densities observed in hot dry years. We observed no adverse effects to the stream orchids from Canada thistle (Cirsium arvense) control efforts which included both herbicide and mechanical (hand-pulling) treatments. To date, the Canada thistle rust fungus, a biological control agent, has yet to become established despite three years of releases and hence its efficacy is unknown.

TABLE OF CONTENTS

1.0 INTRODUCTION ...... 4 2.0 BACKGROUND ...... 4 2.1 Geographic Range ...... 4 2.2 Life History Traits and Habitat ...... 4 2.3 Occurrences at Filoha Meadows Nature Preserve ...... 6 3.0 CLIMATE AND STREAMFLOW ...... 6 3.1 Precipitation ...... 6 3.2 Temperature ...... 8 3.3 Streamflow ...... 8 4.0 BASELINE RARE PLANT MONITORING ...... 10 4.1 Methods ...... 10 4.2 Results and Discussion ...... 12 5.0 NOXIOUS WEED CONTROL MONITORING ...... 14 5.1 Background ...... 15 5.2 Treatment Methods ...... 15 5.2.1 Biological Control ...... 15 5.2.2 Chemical Control ...... 16 5.2.3 Mechanical Control ...... 17 5.3 Monitoring Methods ...... 17 5.4 Results and Analysis ...... 18 6.0 CONCLUSIONS AND RECOMMENDATIONS ...... 22 7.0 REFERENCES ...... 23 8.0 MAPS ...... 25 9.0 TRANSECT PHOTOS (2020) ...... 28 10.0 MACROPLOT PHOTOS (2020) ...... 36 APPENDIX A. 2018, 2019 & 2020 EPIPACTIS GIGANTEA RAW DATA - MACROPLOT ...... A1-A2 APPENDIX B. 2018, 2019 & 2020 EPIPACTIS GIGANTEA RAW DATA - TRANSECTS ...... B1-B8 APPENDIX C. 2018, 2019 & 2020 CIRSIUM ARVENSE RAW DATA – TRANSECTS ...... C1-C6 APPENDIX D. STATISTICAL TEST RESULTS ...... D1-D25 APPENDIX E. PESTICIDE APPLICATION RECORD FOR MILESTONE ...... E1-E3

LIST OF TABLES Table 1. Precipitation for the 2018, 2019, and 2020 Water Years compared to Baseline ...... 7 Table 2. Epipactis density in 2018, 2019 and 2020...... 12 Table 3. Canada Thistle Treatment Methods ...... 17

Stream Orchid Monitoring Report – 2020 Growing Season - Filoha Meadows Nature Preserve Page 1 LIST OF FIGURES Figure 1. Precipitation for the 2018, 2019 and 2020 Water Years as Compared to Baseline...... 7 Figure 2. Temperature for the 2018, 2019, and 2020 Water Years as Compared to Baseline...... 8 Figure 3. Streamflow for the Crystal River. 2018 Growing Season...... 9 Figure 4. Streamflow for the Crystal River. 2019 Growing Season...... 9 Figure 5. Streamflow for the Crystal River. 2020 Growing Season...... 10 Figure 6. Macroplot (left) and Transect (right) Layout Designs...... 11 Figure 7. Epipactis density for all data. Significant differences were found between 2018-2019, 2018-2020, but not between 2019 and 2020...... 12 Figure 8. Epipactis density for macroplot only. Significant differences were found between all years of the study...... 13 Figure 9. Epipactis density for all transect data. Significant differences were found between 2018-2019, 2018- 2020, but not between 2019-2020...... 13 Figure 10. Epipactis density between 2018 and 2020 for macroplot and individual transects...... 14 Figure 11. Density of Canada Thistle over all treatment types. 2018-2020...... 18 Figure 12. Density of thistle stems for Herbicide Treatment. Significant differences were found between all years of the study...... 19 Figure 13. Density of thistle stems for hand-pull treatment. Significant differences were found between all years of the study...... 19 Figure 14. Density of Canada Thistle Stems for the Rust Treatment. Significant Differences were found between 2018-2020 and 2019-2020, but not between 2018 and 2019...... 20 Figure 15. Density of Canada thistle stems for Control (no treatment). Significant differences were found between 2018-2020 and 2019-2020, but not between 2018-2019...... 20 Figure 16. Density of Epipactis gigantea stems for Canada thistle Herbicide Treatment. Significant differences were found between all years of the study. 2018 represents the pre-treatment baseline...... 21 Figure 17. Density of Epipactis gigantea stems for Canada thistle Pull Treatment. Significant Increases were found between 2018 and 2019 and 2020, but not between 2019 and 2020. 2018 represents the pre-treatment baseline...... 21

LIST OF PHOTOS Photo 1. View of hot springs meadow with Epipactis locations flagged (Jul-02-2018)...... 4 Photo 2. Epipactis gigantea. Photo credit: Lisa Tasker...... 5 Photo 3. Canada thistle (Jul-11-2019) ...... 14 Photo 4. Canada thistle in area of prime rare plant habitat (Sep-11-2018) ...... 15 Photo 5. Karen Rosen of the Palisade Insectary applying rust fungus spores (teliospore) to Canada thistle plants (Sep-11-2018)...... 15 Photo 6. Hand painting of Milestone herbicide on Canada thistle plants (Sep-12-2018) ...... 16

Stream Orchid Monitoring Report – 2020 Growing Season - Filoha Meadows Nature Preserve Page 2 LIST OF MAPS Map 1. Occurrences of Epipactis gigantea ...... 26 Map 2. Monitoring Transects ...... 27

LIST OF TRANSECT PHOTOS Photo 1. Cirsium Transect T1. (Jul-09-2020) ...... 29 Photo 2. Cirsium Transect T2. (Jul-09-2020) ...... 29 Photo 3. Cirsium Transect T3. (Jul-09-2020) ...... 30 Photo 4. Cirsium Transect T4. (Jul-09-2020) ...... 30 Photo 5. Cirsium Transect T5. (Jul-09-2020) ...... 31 Photo 6. Cirsium Transect T6. (Jul-09-2020) ...... 31 Photo 7. Transect T7a Start. 17 deg. heading (Jul-08-2020) ...... 32 Photo 8. Transect T7b End. (Jul-08-2020) ...... 32 Photo 9. Transect T8a Start. 127 deg. heading (Jul-08-2020) ...... 32 Photo 10. Transect T8b End. (Jul-08-2020) ...... 32 Photo 11. Transect T9a Start. 222 deg. heading (Jul-08-2020)...... 33 Photo 12. Transect T9b End. (Jul-08-2020) ...... 33 Photo 13. Transect T10a Start. 16 deg. heading (Jul-08-2020)...... 33 Photo 14. Transect 10b End. (Jul-08-2020) ...... 33 Photo 15. Transect T11a Start. 4 deg. heading (Jul-08-2020)...... 34 Photo 16. Transect T11b End. (Jul-08-2020) ...... 34 Photo 17. Transect T12a Start. 336 deg. heading (Jul-08-2020)...... 34 Photo 18. Transect T12b End. (Jul-08-2020) ...... 34 Photo 19. Transect T13a Start. 349 deg. heading (Jul-08-2020)...... 35 Photo 20. Transect T13b End. (Jul-08-2020) ...... 35 Photo 21. Transect T14a Start. 13 deg. heading (Jul-08-2020)...... 35 Photo 22. Transect T14b End. (Jul-08-2020) ...... 35 Photo 1. Macroplot quadrat 1. (Jul-08-2020) ...... 37 Photo 2. Macroplot quadrat 2. (Jul-08-2020) ...... 37 Photo 3. Macroplot quadrat 3. (Jul-08-2020) ...... 38 Photo 4. Macroplot quadrat 4. (Jul-08-2020) ...... 38 Photo 5. Macroplot quadrat 5. (Jul-08-2020) ...... 39 Photo 6. Macroplot quadrat 6. (Jul-08-2020) ...... 39 Photo 7. Macroplot quadrat 7. (Jul-08-2020) ...... 40 Photo 8. Macroplot quadrat 8. (Jul-08-2020) ...... 40 Photo 9. Macroplot quadrat 9. (Jul-08-2020) ...... 41 Photo 10. Macroplot quadrat 10. (Jul-08-2020) ...... 41

Stream Orchid Monitoring Report – 2020 Growing Season - Filoha Meadows Nature Preserve Page 3

1.0 INTRODUCTION The stream orchid or false helleborine orchid (Epipactis gigantea) is a critically imperiled to imperiled plant in Colorado that occurs at Filoha Meadows Nature Preserve (Filoha), an open space managed by Pitkin County Open Space and Trails. See Photo 1. This report describes the third year of a long-term monitoring study designed to track annual population trends and investigate the effect of noxious weed control actions on the health and long-term persistence of this rare plant species.

2.0 BACKGROUND

2.1 GEOGRAPHIC RANGE Epipactis gigantea is a wide ranging species occurring south from British Columbia through the western United States as far inland as Texas, with at least one collection from Mexico (NatureServe 2018). Disjunct populations occur in Utah, Colorado, Wyoming, Montana, South Dakota, Oklahoma, Texas and Central Mexico. Although it is generally thought to be native to North America, some recent taxonomic work indicates that its global range may include parts of Asia (FNA 2003). Epipactis gigantea occupies a variety of habitats, all occurring near water. The USDA Plants website classifies its wetland status as “Obligate”, meaning the species requires wet substrate to survive. Suitable habitats include seeps, marshes near hot springs, hanging gardens, and perennial streams. In Colorado, Epipactis gigantea is known from approximately 30 locations in nine western counties, including Archuleta, PHOTO 1. VIEW OF HOT SPRINGS MEADOW WITH EPIPACTIS LOCATIONS FLAGGED (JUL-02-2018). Chaffee, Delta, Las Animas, Mesa, Montrose, Montezuma, Moffat, and Pitkin (Ackerfield 2015). The two highest elevation occurrences in Colorado are located on private land at Valley View Hot Springs in Saguache County (8,400-8,950 ft) and at Poncha Hot Springs in Chaffee County at 7,940 ft (Rocchio et al. 2006). The Filoha Meadows population at 6,800 ft in elevation, is also one of the highest elevation occurrences of this plant.

2.2 LIFE HISTORY TRAITS AND HABITAT

Epipactis gigantea is a perennial monocot and a member of the orchid family. It is a showy perennial with a large reddish flower that is not easily confused with other species. See Photo 2. The flower’s lower lip vibrates easily when moved, resulting in another commonly used name “chatterbox orchid”. The basal portion of the flower has a pouch-like appearance, which is why E. gigantea is sometimes referred to as “false ladies slipper”. This species’ large size has given it the name “giant helleborine.” Finally, the most commonly accepted name, “stream orchid,” reflects its preferred habitat. E. gigantea is pollinated by syrphid flies at Filoha (Wilson 2009). The plant attaches a pollinium to the back of the pollinator, that is then transferred to the next flower (Culver and Lemly 2013). Like all orchids, E. gigantea has tiny dust-like seeds and an association with mycorrhizae, which are critically important during orchid germination as an orchid seed has virtually no energy reserve and obtains its

Stream Orchid Monitoring Report – 2020 Growing Season - Filoha Meadows Nature Preserve Page 4 carbon from the fungal symbiont. The mycorrhizae species associates within the soil remain imperative to the plant throughout its life, providing mineral absorption.

Disturbance may be necessary for successful establishment of Epipactis gigantea. Rhizomatous species are often able to tolerate scouring and emerge from buried sediments, suggesting that E. gigantea may have a competitive advantage in environments where such disturbances are prevalent. However, data suggest that disturbance is not required for species persistence (Rocchio et al. 2006). At Filoha, the Crystal River does not often flood the upland hot springs area. However, elk and bighorn sheep congregate to calve in the spring. The hooves of these large animals may actually break up the sodden turf, which doesn’t become firmly frozen due to hot springs temperatures of up to 130°F. This disturbance may be beneficial to the orchids (Wilson, 2009).

Epipactis gigantea occurs in isolated populations scattered over a broad geographic range. Occurrences of E. gigantea in California are very different from each other, suggesting that minimal genetic information is transferred between occurrences, and that occurrences may represent genetically distinct populations. Because of this genetic variability and its self-compatibility, E. gigantea is capable of colonizing a variety of geographically isolated habitats. Genetic variability allows a species to adapt to variable habitat conditions, while self-compatibility increases the potential for successful sexual reproduction even when few individuals are present. This is consistent with some of the species’ life history characteristics, such as long-range dispersal (via wind and water) and its habitat requirements (Rocchio 2006). Pollination biology experiments at Filoha resulted in self-pollination success roughly equal to the wild controls, showing self-pollination as a reproductive method to be a normal state (Wilson, 2009). There are no data available on genetic variability or characteristics of E. PHOTO 2. EPIPACTIS GIGANTEA. PHOTO CREDIT: LISA TASKER. gigantea at Filoha; however, given that most occurrences in Colorado are isolated from each other, it is likely that genetic variability between these occurrences is considerable (Rocchio et al. 2006). The species could be vulnerable if there is low genetic diversity represented within the population at Filoha.

A potential threat to the population of Epipactis gigantea at Filoha may be its reliance on only one type of pollinator. The hot springs fed meadow is a rare habitat, in which many insect species are able to thrive because of the elevated ground temperatures year-round. Syrphid flies were the only insect documented carrying and transferring E. gigantea pollinium. The pollinator species were identified to be two species of syrphid flies, Eupeodes volucris and Sphaerophoria philanthus (Wilson, 2009). The stream orchid relies upon clonal and self- compatibility as significant reproductive methods, but the out-crossing provided by pollinators gives some needed genetic diversity within the population. The long-term effect of this mixing of genes is thought to enable a greater capacity to withstand changing climate conditions (Wilson, 2009).

With its wind-dispersed seeds and ability to colonize early successional habitat, Epipactis gigantea exhibits characteristics of a pioneer species. As indicated by its disjunct, fragmented distribution, occasional long-range dispersal events do evidently occur. However, successful dispersal events appear to be relatively uncommon. Given the characteristically large separation distances between subpopulations, the likelihood of additional migrants from the same or related founder subpopulation is very slight (Thornhill 1996).

Stream Orchid Monitoring Report – 2020 Growing Season - Filoha Meadows Nature Preserve Page 5 Throughout its wide range, Epipactis gigantea occurs infrequently but can be locally abundant. Stream orchid is considered to be globally "apparently secure" (G4) given the large number of existing populations, however in Colorado it is ranked as a S1/S2 species, critically to critically imperiled based on the relatively small number of populations (NatureServe 2018). The U.S. Forest Service also lists it as a “sensitive species” in the region.

The main threat to Epipactis gigantea is loss of suitable habitat that is caused by water development projects or any activity that lowers the water table or disturbs its wetland habitat (Rocchio et al. 2006). In Idaho and Wyoming, populations have been extirpated or reduced in size due to impacts from the recreational use of hot springs. In British Columbia, Canada, of the 34 known (including historical) subpopulations, nine have been extirpated or are likely extirpated, translating to a 26% decline over the past century (COSEWIC 2015). On the other hand, as a long-lived perennial, Epipactis gigantea occurrences are likely to be stable as long as there are no alterations to hydrology or habitat. Within Canada, numerous subpopulations are known to have persisted since at least the late 1800s or early 1900s when first recorded, attesting to their vigor and longevity (COSEWIC 2015).

At Filoha, the mineral laden, hot spring waters flow year-round, sustaining a good portion of the wetland plant communities. It is also used by the adjacent Johnson residence for heating and hot tubs. The former owner retained rights to hot springs located on Filoha to heat their property. There are still wells and piping that provide hot water to the property and it was a condition of purchase that these will remain on the property to be used by the former owner. The amount of hot spring water utilized and the influence on the groundwater table and the rare plants that it supports is current unknown, however it does remain a potential threat.

2.3 OCCURRENCES AT FILOHA MEADOWS NATURE PRESERVE Epipactis gigantea at Filoha Meadows Nature Preserve was first identified by Lisa Tasker (E.M.Ecological 2007). In 2018, we conducted a comprehensive mapping of this plant via pedestrian surveys. The field mapping resulted in the identification of a total of 29 occurrences totaling 2.0 acres. These occurrences occur as far north as Penny Hot Springs to approximately 400 ft south of the “Tall Tales” cabin. The habitat includes the hot spring wet meadow and moist terraces and floodplains of the Crystal River. Further details on the mapping as well as a list of common plant species associates may be found in the Stream orchid Monitoring Report – 2018 Growing Season, Filoha Meadows Nature Preserve (PeakEco 2019b) . See Map 1.

3.0 CLIMATE AND STREAMFLOW Seasonal and annual variations in climate such as precipitation and temperature may affect the abundance and density of plant species. In addition, streamflow influences the local alluvial aquifers which may further affect the abundance of wetland dependent species such as Epipactis gigantea. Therefore, as part of this study we have gathered and analyzed the local precipitation, temperature and streamflow data to be considered when looking at changes in plant populations over time. The precipitation, temperature and streamflow data were taken from the closest available climate stations to Filoha Meadows Nature Preserve.

Please note, due to data availability, three different stations were utilized. Precipitation and temperature data are freely available through the National Oceanic and Atmospheric Administration (NOAA) National Center for Environmental Information (NCEI) formally known as the National Climatic Data Center. See https://www.ncdc.noaa.gov/ for more details. Streamflow data was taken from the USGS National Water Information System https://waterdata.usgs.gov/co/.

3.1 PRECIPITATION Precipitation data was gathered from the Carbondale 0.5 W (US1COGF0047) station located approximately 12 miles north and about 600 feet lower in elevation than Filoha. The data show that the 2020 and 2018 water years were drier than the baseline and that the 2019 water year was much wetter than baseline. See Table 1 and Figure 1. More specifically, the total precipitation for the 2020 water year was 10.37 inches, was 18.15 inches in 2019, and 12.14 inches in 2018. The average for all years of record (2008-2020), was 14.43 inches.

Stream Orchid Monitoring Report – 2020 Growing Season - Filoha Meadows Nature Preserve Page 6 When broken down into seasons, the data show that the spring (April, May, and June) of 2020 had about 1.5 inches of less rain than baseline, 2019 had over two inches of more precipitation than baseline, and 2018 had only slightly lower precipitation (about ½ of an inch) lower than the baseline. The same pattern is evident when looking at total precipitation by month (Figure 1). Note the high precipitation levels in May, June, and July of 2019 as compared to 2018 and 2020.

The term "water year" is a TABLE 1. PRECIPITATION FOR THE 2018, 2019, AND 2020 WATER term commonly used in YEARS COMPARED TO BASELINE hydrology to describe a time BASELINE period of 12 months for which 2018 WATER 2019 WATER 2020 WATER (2008-2020 precipitation totals are YEAR YEAR YEAR measured and is defined as WATER YEARS) the 12-month period Fall (Oct – Dec) 2.87 4.51 2.34 3.26 beginning October 1, for any Winter (Jan-Mar) 2.79 5.10 3.26 3.24 given year through Spring (Apr-Jun) 3.43 5.95 2.17 3.70 September 30, of the following year. The water year Summer (Jul -Sep) 3.05 2.59 2.60 4.22 is designated by the calendar Annual Total 12.14 18.15 10.37 14.43 year in which it ends which Water Year. The 12-month period beginning October 1, for any given year through includes 9 of the 12 months. September 30, of the following year. The water year is designated by the calendar year Thus, the year ending in which it ends. September 30, 1999 is called Current Location: Elevation: 6171 ft. Lat: 39.3978° N Lon: -107.2227° W the "1999" water year. Station: CARBONDALE 0.5 W, CO (US1COGF0047). Note data prior to 2007 is not available at this station.

Precipitation 2018-2020 Station: CARBONDALE 0.5 W, CO US US1COGF0047 Elev: 6,171 ft. Lat: 39.3978° N Lon: -107.2227° W 3.5

2.5

1.5

0.5

Monthly Precipitatoin Monthly Precipitatoin in Inches 0 Oct Nov Dec Jan Feb Mar Apr May Jun Jul Aug Sep 2018 Water Year 2.06 0.38 0.43 0.72 1.2 0.87 2.24 0.4 0.79 1.21 1.4 0.44 2019 Water Year 2.58 0.9 1.03 0.94 1.2 2.96 1.61 2.25 2.09 1.84 0.37 0.38 2020 Water Year 0.95 0.66 0.73 1.16 1.17 0.93 0.6 1.09 0.48 1.23 0.22 1.15 Baseline (2008-2020) 1.27 0.79 1.20 1.07 1.07 1.11 1.36 1.60 0.73 1.77 1.20 1.25

2018 Water Year 2019 Water Year 2020 Water Year Baseline (2008-2020)

FIGURE 1. PRECIPITATION FOR THE 2018, 2019 AND 2020 WATER YEARS AS COMPARED TO BASELINE.

Stream Orchid Monitoring Report – 2020 Growing Season - Filoha Meadows Nature Preserve Page 7 3.2 TEMPERATURE

Temperature data was gathered from the Crown Colorado (USR0000CCRO) station located approximately 11 miles northeast and about 1,500 feet higher in elevation than Filoha Meadows. The data show that May 2020 had a higher average monthly temperature than baseline, similar to 2018. Then June and July of 2020 were closer to baseline values, but lower than in 2018. The data also show that the 2019 spring and early summer (May through July) had lower average temperatures than baseline years, while 2018 had much higher temperatures than baseline. These temperature differences are likely correlated with the precipitation patterns, with cooler average temperatures associated with relatively high precipitation levels (mostly as snow) as observed in 2019 and a warmer and drier season in 2018. See Figure 2.

Temperature 2018-2020 Station: The Crown Colorado, CO. USR0000CCRO Elev: 8,298 ft. Lat: 39.3528° N Lon:-107.0931 ° W F ° 80 70 60 50 40 30 20

Average Average MonthlyTemperature 10 0 Oct Nov Dec Jan Feb Mar Apr May Jun Jul Aug Sep 2018 Water Year 44.1 40.7 29.2 28.9 27.4 35 42.5 55.8 64.3 70.7 65.7 61.4 2019 Water Year 42.5 28.4 22.9 24.7 24.8 33 43.3 44.6 57.6 65.7 66.5 60.7 2020 Water Year 38.4 34.4 24.7 24.6 24.9 36.5 41.8 54.4 60.4 67.0 70.0 58.0 Baseline (2008-2020) 44.9 34.6 24.5 25.0 27.5 35.7 40.7 50.4 61.8 66.5 64.8 58.4

2018 Water Year 2019 Water Year 2020 Water Year Baseline (2008-2020)

FIGURE 2. TEMPERATURE FOR THE 2018, 2019, AND 2020 WATER YEARS AS COMPARED TO BASELINE.

3.3 STREAMFLOW Like precipitation, the streamflow for the 2018, 2019, and 2020 showed a similar pattern of above normal stream flows in 2019 and below normal in 2018 and 2020. However, 2020 river flows were still slightly higher than in 2018. Streamflow data was taken from the USGS Crystal River Above Avalanche Creek station (USGS 09081600) located in the northern portion of Filoha Meadows Nature Preserve, across the river from Penny Hot Springs. See Figures 3, 4, and 5 where blue represents the 2018, 2019, or 2020 water year and orange represents the median daily statistic of 64 years of record.

Stream Orchid Monitoring Report – 2020 Growing Season - Filoha Meadows Nature Preserve Page 8 FIGURE 3. STREAMFLOW FOR THE CRYSTAL RIVER. 2018 GROWING SEASON.

FIGURE 4. STREAMFLOW FOR THE CRYSTAL RIVER. 2019 GROWING SEASON.

Stream Orchid Monitoring Report – 2020 Growing Season - Filoha Meadows Nature Preserve Page 9 FIGURE 5. STREAMFLOW FOR THE CRYSTAL RIVER. 2020 GROWING SEASON.

4.0 BASELINE RARE PLANT MONITORING The following monitoring study was designed to provide quantitative data on the long-term trends in Epipactis gigantea density at Filoha and determine whether densities of this plant population are increasing, decreasing, or remaining stable over time. To date, three years of data have been collected: 2018, 2019 and 2020 (PeakEco 2019b, 2020). See Map 2 for a location of the sampling units and Figure 6 illustrates the macroplot and transect layouts. Appendices A-C present the raw data and Appendix D presents the statistical test results.

4.1 METHODS

Restricted Random Macroplot. A restricted random sampling approach was used to estimate the stem density at Filoha (Elzinga et al. 1998) in one large macroplot. In this monitoring method, we subjectively located a 50 meter (m) by 5m macroplot in an area of a known “hotspot” of Epipactis plants. Then the macroplot was divided into 10 equal-sized segments. Within each of these segments, a single sampling unit (5m by 2m) was randomly located and permanently marked. The total number of Epipactis gigantea ramets (both flowering and non- flowering) was recorded for each 2m wide sub-plot.

Stream Orchid Monitoring Report – 2020 Growing Season - Filoha Meadows Nature Preserve Page 10 Systematic Sampling Transects. In addition to the macroplot method described above, we used a smaller scale systematic sampling design in other locations in order to assess stem density within the smaller size occurrences throughout Filoha. In this method we used a 12m long transect, randomly located within a sub-population, and systematically placed 60 centimeter (cm) by 60 cm frames on either side of the transect. Six frames were placed to the right of the transect at the 2m, 4m, 6m, 8m, 10m, and 12m marks and six frames were placed to the left of the transect at the 1m, 3m 5m, 7m, 9m and 11m marks. Each frame was placed in front of the designated meter mark. The number of flowering and non-flowering ramets of Epipactis gigantea was recorded for each frame. In addition, the compass direction was taken at the start of the transect and photos were taken at each end. Each end was permanently marked with orange fiberglass stakes and metal 8-inch nails. Finally, a list of the vascular plant species observed within the 2m wide by 14m long belt transect was recorded. A total of eight of these 12m belt transects (T7 through T14) were sampled at Filoha. See Figure 6 and Map 2.

For both sampling designs, we determined some basic descriptive statistics for the density of ramets - both the mean (or arithmetic average) and median (the middle value) numbers of ramets per square meter (sq.m.). For highly skewed and non-normally disturbed data such as collected in this study, it was determined that the median values represented the better “average” value than the arithmetic average or mean. Statistical analyses were completed using the MedCalc computer software. Comparisons among all three years utilized the Friedman test, which is a non-parametric test similar to a repeated measures analysis of variance (ANOVA). The ANOVA test can only be used on normally distributed data, which is not represented by the data set. Furthermore, a post hoc analysis FIGURE 6. MACROPLOT (LEFT) AND TRANSECT (RIGHT) LAYOUT DESIGNS. with Wilcoxon signed-rank tests was conducted to test for differences between 2018-2019, 2018-2020, and 2019-2020. The Wilcoxon rank-sum test for paired samples is a non-parametric alternative for the paired samples t-test. All average values presented below are the median value.

Stream Orchid Monitoring Report – 2020 Growing Season - Filoha Meadows Nature Preserve Page 11 4.2 RESULTS AND DISCUSSION

Overall, when all data is combined, there is a highly significant difference in density of Epipactis gigantea TABLE 2. EPIPACTIS DENSITY IN 2018, 2019 AND ramets among the three years. There were 38.9 2020. ramets per sq.m. in 2018, 66.7 ramets per sq. m. in 2019, and 63.7 ramets per sq.m. in 2020 (Table 2). Epipactis Density - Median ramets / sq.m. When samples are compared year-to-year, Year All Macroplot Transects significant differences were found in ramet density 2018 38.9 37.6 40.3 between 2018 and 2019 and between 2018 and 2019 66.7 78.8 61.1 2020, but not between 2019 and 2020 (Figure 7, p<0.0001). 2020 63.7 67.9 59.7

When just the macro plot is analyzed, there is an average of 37.6 ramets/sq.m. in 2018, 78.8 ramets/sq.m. in 2019, and 67.9 ramets/sq.m. in 2020 (Figure 8, p<0.0001). The data are significant across all years as well as between each set of years, 2018-2019, 2018-2020, and 2019-2020.

300

250

200

150

100

Median Density ( ramets / sq.m. ) sq.m. / ramets Density( Median 50

0 2018_Density 2019_Density 2020_Density All Data

FIGURE 7. EPIPACTIS DENSITY FOR ALL DATA. SIGNIFICANT DIFFERENCES WERE FOUND BETWEEN 2018-2019, 2018-2020, BUT NOT BETWEEN 2019 AND 2020.

Finally, when just the transects T7 through T14 data are combined, the median density increases from 40.3 ramets/sq.m. in 2018 to 61.1 ramets/sq.m. in 2019, and then slightly lower to 59.7 ramets/sq.m. in 2020. The data are significant among all three years (Figure 9, p<0.0001) as well as between 2018 and 2019, 2018 and 2020, but not significant between 2019 and 2020.

Stream Orchid Monitoring Report – 2020 Growing Season - Filoha Meadows Nature Preserve Page 12 140

120

100

40 Median Density ( ramets / sq.m. ) sq.m. / ramets Density( Median 20

0 2018_Density 2019_Density 2020_Density Macroplot Only

FIGURE 8. EPIPACTIS DENSITY FOR MACROPLOT ONLY. SIGNIFICANT DIFFERENCES WERE FOUND BETWEEN ALL YEARS OF THE STUDY.

300 In addition to the group comparisons identified above, we also looked at

250 individual transect Epipactis density between 2018, 2019, and 2020. As illustrated in Figure 10, every 200 transect showed significant increases in orchid density from 150 2018 and 2019, but some transects showed slightly lower densities in

100 2020 as compared to 2019, but none

Median Density Median (ramets / sq.m.) of these differences were statistically significant. Please note, the 50 statistical year-to-year comparisons of each transect in an a priori 0 pairwise strategy is not statistically 2018_Density 2019_Density 2020_Density Transects legitimate due to an increase in the Type I error rate (Grant 2020). FIGURE 9. EPIPACTIS DENSITY FOR ALL TRANSECT DATA. SIGNIFICANT DIFFERENCES Nevertheless, the analysis is WERE FOUND BETWEEN 2018-2019, 2018-2020, BUT NOT BETWEEN 2019-2020. presented all with the other statistical test results.

Overall, the stream orchid density at Filoha is variable among years and appears to correlated with temperatures and precipitation in late spring and early summer (May, June, and July). The lowest densities of Epipactis were seen in 2018 which had very high mean temperatures and low precipitation in these months, while the densities were higher in 2019 and 2020, in which temperatures were lower and precipitation was higher. May, June, and July of 2019 had the lowest temperatures and highest precipitation as well as the highest densities of Epipactis. In addition to stem density, as presented in this report, Ms. Denise Wilson has been studying stem height, number of leaves, and flower and fruit production of Epipactis at Filoha through a grant from the Colorado Native Plant Society. Initial results of the study, which spanned 2019 and 2020 growing seasons, indicate that Epipactis plants had higher heights and more flowers and capsule production in 2019 than in 2020.

Stream Orchid Monitoring Report – 2020 Growing Season - Filoha Meadows Nature Preserve Page 13 180.0 160.0 140.0 120.0 100.0 80.0 60.0 40.0 20.0 Median Denisty (ramets / sq.m.) 0.0 M1 T7 T8 T9 T10 T11 T12 T13 T14 2018 37.6 90.3 6.9 19.4 91.7 20.8 81.9 66.7 93.1 2019 78.8 94.4 19.4 50.0 170.8 26.4 97.2 106.9 151.4 2020 67.9 111.1 22.2 54.2 148.6 26.4 91.7 108.3 165.3

2018 2019 2020

FIGURE 10. EPIPACTIS DENSITY BETWEEN 2018 AND 2020 FOR MACROPLOT AND INDIVIDUAL TRANSECTS.

5.0 NOXIOUS WEED CONTROL MONITORING The following noxious weed control monitoring study was designed to provide quantitative data on the most effective way of controlling Canada thistle (Cirsium arvense), a state listed noxious weed, without detriment to Epipactis gigantea. Canada thistle is present in significant quantities in the hot springs meadow and likely competes with the stream orchid for sunlight, nutrients, and water. Canada thistle also has allelopathic effects to neighboring plants, meaning that it produces chemicals that inhibit the growth and survival of other plants (Stachon and Zimdahl 1980). This section of the report describes the third year of a noxious weed control program designed to investigate the use of biocontrols, mechanical control (hand-pulling), and careful use of selective herbicides to control Canada thistle. Based on data obtained do date, it appears that the herbicide and hand-pulling treatments were effective at controlling the thistle and did not affect the density of the stream orchid.

PHOTO 3. CANADA THISTLE (JUL-11-2019)

Stream Orchid Monitoring Report – 2020 Growing Season - Filoha Meadows Nature Preserve Page 14 5.1 BACKGROUND

Canada thistle is one of the most serious and widespread invasive weeds worldwide and in North America. See Photo 3. It thrives in several different ecological settings including farmland, ranchland, meadows, and riparian areas. It is a deep-rooted perennial that spreads by seeds and aggressive creeping, horizontal roots called rhizomes. Canada thistle flowers in late spring and throughout the summer. It produces about 1,000 to 1,500 seeds per plant that are carried by the wind, and the seeds can survive in the soil for up to 20 years. Additionally, Canada thistle reproduces vegetatively through its root system, and quickly forms dense stands. Each fragmented piece of root is capable of forming new plants (CDA 2015). PHOTO 4. CANADA THISTLE IN AREA OF PRIME RARE PLANT HABITAT (SEP-11-2018) As depicted in Photo 4, significant stands of Canada thistle have become established in the hot spring meadow among and adjacent to populations of the stream orchid.

5.2 TREATMENT METHODS

5.2.1 BIOLOGICAL CONTROL

As detailed in the Experimental Weed Control 2018 Monitoring Report (PeakEco 2019a), attempts at controlling Canada thistle within and adjacent to stream orchid populations have included the use of biological control, mechanical control, and chemical control. Karen Rosen of the Colorado Department of Agriculture (CDA) Palisade Insectary has been attempting to establish the biological control agent, Puccinia punctiformis, which is a host-specific rust fungus on the Canada thistle at Filoha. Unlike many classical biological control agents that merely limit or control the spread of an infestation, the Canada thistle rust fungus has the potential to significantly decrease existing infestations. This rust fungus has evolved to be very effective at causing thistle mortality. According to the CDA, experimental trials showed a worst case of a 45% reduction of thistles over five years, while the best case was 100% after 18 months. The control efficacy is largely dependent on the current density of thistle and surrounding vegetation (CDA 2019). The CDA’s program has been so successful they are currently in the process of expanding in other western states (Bean 2017). To date, the rust fungus has yet to become established at Filoha. PHOTO 5. KAREN ROSEN OF THE PALISADE INSECTARY APPLYING RUST FUNGUS SPORES (TELIOSPORE) TO CANADA THISTLE PLANTS (SEP-11-2018) Stream Orchid Monitoring Report – 2020 Growing Season - Filoha Meadows Nature Preserve Page 15 Release Efforts. In 2018, Ms. Rosen utilized two different spore types representing different stages of the life cycle of P. punctiformis. The first, aeciospores, or the “spring spores” were applied on May 21, 2018 to a 12m x 6m plot (T1). The second kind of spore, teliospores, or “fall spores” were applied on September 11, 2018 to two 12m x 6m plots (T4 and T6). In 2019, no aeciospores were applied. Instead, all three treatment sites (T1, T4, and T6) were inoculated with teliospores on September 6, 2019. Each treatment received 75 grams of teliospores. See Photo 5. No additional releases occurred in 2020.

To date no rust fungus has been observed at any of the treatment sites either in 2018, 2019, or in 2020. According to Ms. Rosen, the fall inoculation sites in all years did not have many good thistle rosettes. The best rosettes for inoculation are short, healthy and abundant. Instead, spores were sprinkled along the transect on tall spindly thistle stems that are not ideal for rust fungus establishment. Ms. Rosen believes that the tall grasses in the area are likely inhibiting the important fall rosettes needed for inoculation. As discussed below in Section 5.4 Results, the Canada thistle population at Filoha appears to be in decline over the three years of the study and hence no future inoculations are planned.

5.2.2 CHEMICAL CONTROL Several different herbicides are recommend for control of Canada thistle by the Colorado Department of Agriculture including aminopyralid (Milestone), Clopyralid/Triclopyr (Prescott; Redeem; others), and Aminocyclopyrachlor/Chlorsulfuoron (Perspective) (CDA 2015). Many of these herbicides show 80% to 90% mortality rates. However, the use of herbicides may take several treatments because of root nutrient stores in the Canada thistle. In addition, herbicides, even the selective ones listed above, have the potential to affect non- target plants. Currently, the effect of these herbicides on orchids, including Epipactis gigantea, is unknown. Treatments of the herbicide Milestone have been applied for two consecutive growing seasons. In 2018, the herbicide was hand-painted on the plants and in the 2019, a back-pack sprayer was used to spot-spray the Canada thistle, which resulted in some inadvertent overspray on the senescing stream orchid leaves. No herbicide was applied in 2020.

Herbicide Application Details. Chemical control using the selective Milestone herbicide was conducted on Transects 8 and 9 by Ms. Rea Orthner. Ms. Orthner is a certified pesticide applicator with the Colorado Department of Agriculture (License No. 15431). Milestone is effective at low use rates and has a favorable ecotoxicology profile. It was registered under EPA’ s Reduced Risk Initiative. Milestone is approved for use on rangeland, permanent grass pastures, Conservation Reserve Program (CRP) acres, non-cropland areas (such as roadsides and utility rights-of-way), non-irrigation ditch banks, seasonally dry wetlands, and natural areas. It provides excellent post-emergent and residual control of over 70 broadleaf weeds and woody plants, including thistles with proven tolerance to native grasses and many shrubs and forbs (DOW 2013). As previously stated, the effect of Milestone on orchids is unknown, but it reasonable to assume it may negatively affect such plants.

In order to test the effectiveness of Milestone on treating Canada thistle, it was applied to two 12m x 6m areas around Transects 7 and 8 that contain thistles and the rare Epipactis gigantea. In 2018, we used a very conservative approach by “hand-painting” on the herbicide to Canada thistle plants within each treatment area. See Photo 6. While this method is time-consuming, it does not result in any overspray of herbicide. In 2019, after observing no impacts to orchid stem density (see Section 5.3 below), the Milestone herbicide was spot sprayed on the Canada thistle. This method is much more time-efficient but can result in a small amount of unintentional overspray on the Epipactis plants as well as other species. In both years, the herbicide was applied in mid-September, when the orchids were dying back so as to lessen adverse effects on them. Fall is also PHOTO 6. HAND PAINTING OF MILESTONE an effective time to control Canada thistle as the herbicide is HERBICIDE ON CANADA THISTLE PLANTS (SEP- 12-2018)

Stream Orchid Monitoring Report – 2020 Growing Season - Filoha Meadows Nature Preserve Page 16 translocated to the roots and underground stems during this period. A copy of the Pesticide Application Records for 2018 and 2019 are located in Appendix E and kept on file at the offices of Peak Ecological Services LLC. No herbicide was applied in 2020.

5.2.3 MECHANICAL CONTROL Mechanical control of noxious weeds can include tilling, hoeing, hand pulling, blading, grubbing, and mowing. Of all these methods, it was determined that hand-pulling of Canada thistle would be the least impactful to the neighboring rare orchid plants. However, due to Canada thistle’s extensive root system, hand-pulling and tilling are generally thought to create root fragments and stimulate the growth of new plants. However, repeated pulling of small infestations can be effective if completed frequently enough to deplete nutrient reserves in the root stocks. To date, two seasons of targeted hand-pulling have occurred. While hand-pulling may be effective in small, targeted locations at Filoha, it may be too time-consuming and labor intensive to try to implement this over a large scale.

Mechanical Control Details. In mid-September of 2018 and 2019, all Canada thistle within two 12m x 6m plots (T9 and T10) were hand pulled. All seed heads were bagged and disposed of off-site. Any remaining plant matter was hand-broadcast in the surrounding area outside of the plot. No mechanical control was performed in 2020.

A summary of the treatment methods per plot is found below in Table 3.

TABLE 3. CANADA THISTLE TREATMENT METHODS

Treatment Biological Control None Year Puccinia punctiformis Hand-Pulling Herbicide (Control) Aeciospore Teliospore T7, T8 2018 T1 T4, T6 T9, T10 T2, T3, T5 (hand-paint) T7, T8 2019 ---- T1, T4, T6 T9, T10 T2, T3, T5 (spot spray) 2020 ------

Aeciospores applied on May 21, 2018 but not in 2019; Teliospores applied September 11, 2018 and September 6, 2019. The hand-pulling/herbicide treatments occurred September 12, 2018 and again September 16, 2019. No treatments were made in 2020. 5.3 MONITORING METHODS Monitoring of the weed control treatments followed the same methods for Systematic Sampling Methods (Transects) as described in Section 4.1 above. In this method we used a 12m long transect, and systematically placed 60 cm by 60 cm frames (or quadrats) on the right side of the transect at the 2m, 4m, 6m, 8m, 10m, and 12m marks. Each frame was placed adjacent to the designated meter mark. In each frame the number of C. arvense stems was counted, being careful to look through the thatch to find all thistle plants. In addition, the start of the transect was photographed and the compass direction was taken. Each end was permanently marked with a metal T-post (T1 to T6) or orange fiberglass stakes and metal 8-inch nails (T7-T10). Finally, a list of the vascular plant species observed within the 6m wide by 12m long belt transect was recorded. Please note, the quadrat layout for the herbicide application and hand-pulling treatments (T7, T8, T9, and T10) was expanded to also include quadrats placed on the left side of the transect at the 1m, 3m, 5m, 7m, 9m, and 11m marks. Whereas, for the control and biological treatments quadrats were only placed on the right side of the transect. All rare plant monitoring transects as described in Section 4 above, had quadrats placed on both sides of the transect.

Like the Epipactis gigantea study described above, we determined some basic descriptive statistics for the density of stems-using the median to best represent the average value. Statistical analyses were completed

Stream Orchid Monitoring Report – 2020 Growing Season - Filoha Meadows Nature Preserve Page 17 using the MedCalc computer software. Comparisons among all three years utilized the Friedman test, which is a non-parametric test similar to a repeated measures analysis of variance (ANOVA). The ANOVA test can only be used on normally distributed data, which is not represented by the data set. Furthermore, a post hoc analysis with Wilcoxon signed-rank tests was conducted to test for differences between 2018-2019, 2018-2020, and 2019-2020. The Wilcoxon rank-sum test for paired samples is a non-parametric alternative for the paired samples t-test. All average values presented below are the median value.

5.4 RESULTS AND ANALYSIS The results of the study for the three treatment types on the Canada thistle as well as the control (no treatment) are illustrated below in Figures 11 through 15. Figure 11 shows the near elimination of Canada thistle in the two transects that were initially hand-wiped with herbicide and then spot-sprayed a year later. From 2018 to 2019, both the herbicide treatment and the hand-pull treatment significantly decreased thistle density over the control and rust application. However, in 2020, all transects showed significant reductions in thistle density including where no treatment was made (control) as well as rust application, which was never established.

Density of Canada Thistle 2018-2020 30.0

25.0

20.0

15.0

2018 10.0 2019 5.0 2020

0.0 Mean Thistle Mean Thistle Densite (#stems/sq. m) Herbicide Pull Rust Control 2018 13.3 8.1 21.9 25.5 2019 5.7 3.9 18.4 15.7 2020 0.2 1.0 2.9 1.2

Treatment Type

FIGURE 11. DENSITY OF CANADA THISTLE OVER ALL TREATMENT TYPES. 2018-2020.

A more detailed analysis of the herbicide and pull treatments (Figures 12 and 13), shows significant reductions in thistle density among all years of the study, including between 2018 and 2019. However, for the rust and control treatments, there was no significant difference between 2018 and 2019, but there was a significant reduction between 2019 and 2020, and 2018 and 2020 (Figures 14 and 15). The reason for the low Canada thistle stem densities in 2020, both in treated and untreated plots, is currently not known. Will Canada thistle rebound in hotter and drier conditions as observed in 2018 or will the population continue to have reduced densities in the future?

Stream Orchid Monitoring Report – 2020 Growing Season - Filoha Meadows Nature Preserve Page 18 35

10 Median ThistleMedian Density (stems / sq.m.) 5

0 2018_Cirarv_sqm 2019_Cirarv_sqm 2020_Cirarv_sqm Herbicide Treatment

FIGURE 12. DENSITY OF THISTLE STEMS FOR HERBICIDE TREATMENT. SIGNIFICANT DIFFERENCES WERE FOUND BETWEEN ALL YEARS OF THE STUDY.

10 Thistle Density (stems / sq. m.)

edian edian 5 M

0 2018_Cirarv_sqm 2019_Cirarv_sqm 2020_Cirarv_sqm Pull Treatment

FIGURE 13. DENSITY OF THISTLE STEMS FOR HAND-PULL TREATMENT. SIGNIFICANT DIFFERENCES WERE FOUND BETWEEN ALL YEARS OF THE STUDY. .

Stream Orchid Monitoring Report – 2020 Growing Season - Filoha Meadows Nature Preserve Page 19 45

10 Thistle Stem Density Median (stems / sq.m.) 5

0 2018_Cirarv_sqm 2019_Cirarv_sqm 2020_Cirarv_sqm Rust Treatment

FIGURE 14. DENSITY OF CANADA THISTLE STEMS FOR THE RUST TREATMENT. SIGNIFICANT DIFFERENCES WERE FOUND BETWEEN 2018-2020 AND 2019-2020, BUT NOT BETWEEN 2018 AND 2019.

20 Median ThistleMedian Density (stems / sq.m.) 10

0 2018_Cirarv_sqm 2019_Cirarv_sqm 2020_Cirarv_sqm Control (No Treatment)

FIGURE 15. DENSITY OF CANADA THISTLE STEMS FOR CONTROL (NO TREATMENT). SIGNIFICANT DIFFERENCES WERE FOUND BETWEEN 2018-2020 AND 2019-2020, BUT NOT BETWEEN 2018-2019.

A final component of this study is to investigate whether herbicide or hand-pulling control of Canada thistle had any adverse effects on Epipactis gigantea. As shown below in Figures 16 and 17, the Epipactis density in the transects with the herbicide application or hand-pulling treatments did not decline. In fact, a significant increase over 2018 levels were observed in both treatment types over the two years post treatment (2019 and 2020).

Stream Orchid Monitoring Report – 2020 Growing Season - Filoha Meadows Nature Preserve Page 20 300

250

200

150

100

Median EpipactisMedian Density (ramets / sq.m.) 50

0 2018_Density 2019_Density 2020_Density Herbicide Treatment

FIGURE 16. DENSITY OF EPIPACTIS GIGANTEA STEMS FOR CANADA THISTLE HERBICIDE TREATMENT. SIGNIFICANT DIFFERENCES WERE FOUND BETWEEN ALL YEARS OF THE STUDY. 2018 REPRESENTS THE PRE-TREATMENT BASELINE.

250

200

150

100

50 Median EpipactisMedian Density (ramets / sq.m.)

0 2018_Density 2019_Density 2020_Density Pull Treatment FIGURE 17. DENSITY OF EPIPACTIS GIGANTEA STEMS FOR CANADA THISTLE PULL TREATMENT. SIGNIFICANT INCREASES WERE FOUND BETWEEN 2018 AND 2019 AND 2020, BUT NOT BETWEEN 2019 AND 2020. 2018 REPRESENTS THE PRE- TREATMENT BASELINE.

Stream Orchid Monitoring Report – 2020 Growing Season - Filoha Meadows Nature Preserve Page 21 6.0 CONCLUSIONS AND RECOMMENDATIONS

The data presented in this report show a large-scale increase in stream orchid density between 2018 and both the 2019 and 2020 growing seasons, which we attribute to the very hot dry conditions of 2018 as compared to 2019 and 2020. We also show that the herbicide and mechanical (hand-pulling) treatments of Canada thistle were effective in significant reductions of this noxious weed, without detriment or Epipactis stem density during the two years of post-treatment monitoring. Finally, despite two years of inoculations of the Canada thistle rust fungus biocontrol, no rust has become established on the Filoha site. It is likely that the Canada thistle at Filoha may not be robust or dense enough to sustain a population of the rust biocontrol.

Continued monitoring of the Epipactis population at Filoha is warranted considering the continued legal withdrawal of the hot springs water from adjacent landowner, and potential for additional noxious weed treatments at this site. However, we recommend termination of the experimental weed control portion of this study. Furthermore, if Pitkin County Open Space and Trails desires to treat noxious weeds in the hot spring meadow, we recommend the following be employed to reduce the likelihood of adverse effects to this rare orchid or its associated pollinators:

• Use only hand-held or back-pack sprayers to reduce potential from compaction; • Avoid broadcast spraying; • Ensure that all project staff and contractors can correctly identify the rare plant in their various stages of growth; • Time the treatment to occur when the rare plant is dormant (not in a flowering, reproductive, or seedling phase) and not being actively visited by pollinators. Herbicide treatment should occur in mid to late-September when the Canada thistle is growing, but the rare orchids have already senesced. • Minimize potential for drift by adjusting droplet size accordingly and using a shield, like cardboard, when spraying directly adjacent to rare plants; and • Careful consideration should be made if a different herbicide should be chosen than investigated in this study.

For further information on treating noxious weeds on sites with rare plants, please see Recommended Best Management Practices for Managing Noxious Weeds on Sites with Rare Plants (Mui and Panjabi 2016). This document provides natural resource professionals, land managers, and land use decision makers with guidelines and a set of questions to consider surrounding decisions related to noxious weed management in the vicinity of rare plants.

Stream Orchid Monitoring Report – 2020 Growing Season - Filoha Meadows Nature Preserve Page 22 7.0 REFERENCES

Ackerfield, J. 2015. Flora of Colorado Page 818 pp. Brit Press Bean, D. 2017. Improving the production, distribution and post-release monitoring of Puccinia punctiformis, a naturalized rust fungus for the biological control of Canada thistle, in the West. . Grant Propoal to the USDA Forest Service BCIP Program. From: Dan Bean, Colorado Department of Agriculture, Biological Pest Control, Palisade Insectary, Palisade, CO. CDA. 2015. Canada Thistle Fact Sheet. Colorado Department of Agriculture - Conservation Service. Broomfield, CO. www.colorado.gov/ag/weeds. CDA. 2019. Canada Thistle Biocontrol. Colorado Department of Agriculture Website: https://www.colorado.gov/pacific/agconservation/canada-thistle-biocontrol. COSEWIC. 2015. Giant helleborine (Epipactis gigantea): COSEWIC assessment and status report. Page 73. Committee on the Status of Endangered Wildlife in Canada. . Culver, D. R., and J. M. Lemly. 2013. Field Guide to Colorado's Wetland Plants. Identification, Ecology and Conservation. Colorado Natural Heritage Program, Warner College of Natural Resources, Colorado State University. DOW. 2013. Invasive Plant Management with Milestone and Other Herbicides. A practical and technical guide for natural area managers. Techline Invasive Plant News. . E.M.Ecological. 2007. Filoha Meadows Open Space. Vegetation Descriptions and Stewardship. Prepared by E.M. Ecological, LLC. Natural Resource and Restoration Consulting. Prepared for Pitkin County Open Space and Trails. Elzinga, C. L., D. W. Salzer, and J. W. Willoughby. 1998. Measuring and Monitoring Plant Populations. U.S. Department of the Interior. Bureau of Land Managment. FNA. 2003. Treatment of Epipactis gigantea Douglas ex Hooker. Flora of North America. Vol. 26 Page 585. Grant, M. 2020. Personal Communication. Professor Emeritus, Univrsity of Colorado Boulder. 1228 Cornell Dr. Longmont, CO 80503. [email protected]. Mui, C., and S. Panjabi. 2016. Recommended Best Management Practices for Managing Noxious Weeds on Sites with Rare Plants. Available onlne from www.cnhp.colostate.edu. NatureServe. 2018. NatureServe Explorer: An online encyclopedia of life [web application]. Version 7.1. NatureServe Web Service, Arlington, Virginia. Available online at http://www.natureserve.org/explorer. PeakEco. 2019a. Experimental Noxious Weed Control Monitoring Report. 2018 Growing Season. Filoha Meadows Nature Preserve. Prepared for Pitkin County Open Space and Trails by Peak Ecological Services, LLC, of Nederland, Colorado. PeakEco. 2019b. Stream Orchid (Epipactis gigantea) Monitoring Report. 2018 Growing Season. Filoha Meadows Nature Preserve. Prepared for Pitkin County Open Space and Trails by Peak Ecological Services, LLC, of Nederland, Colorado. PeakEco. 2020. Stream Orchid (Epipactis gigantea) Monitoring Report. 2019 Growing Season. Filoha Meadows Nature Preserve. revsision date: Aug. 4, 2020. Prepared for Pitkin County Open Space and Trails by Peak Ecological Services, LLC, of Nederland, Colorado. Rocchio, J. 2006. Rocky Mountain Lower Montane Riparian Woodland and Shrubland Ecological System Ecological Integrity Assessment. Colorado Natural Heritage Program Warner College of Natural Resources Colorado State University 254 General Services Building Fort Collins, Colorado 80523. 80 pp. Rocchio, J., M. March, and D. G. Anderson. 2006. Epipactis gigantea Dougl. ex Hook. (stream orchid): a technical conservation assessment. [Online]. USDA Forest Service, Rocky Mountain Region. Available: http: www.fs.fed.us/r2/projects/scp/assessments/epipactisgigantea.pdf.

Stream Orchid Monitoring Report – 2020 Growing Season - Filoha Meadows Nature Preserve Page 23 Stachon, W. J., and R. L. Zimdahl. 1980. Allelopathic Activity of Canada Thistle (Cirsium arvense) in Colorado. Weed Science 28:83-86. Thornhill, A. D. 1996. Ph.D. Dissertation: Population and species level patterns of genetic diversity in a widespread temperate orchid species, Epipactis gigantea Dougl. ex. Hook (Orchidaceae): Evolutionary and conservation implications. . Rice University, Houston, TX 77005. Wilson, D. 2009. Pollination Biology of the stream orchid, Epipactis gigantea, (Dougl. Ex Hook) at three Colorado Elevations. Masters Project. University of Colorado Denver.

Stream Orchid Monitoring Report – 2020 Growing Season - Filoha Meadows Nature Preserve Page 24 8.0 MAPS

Stream Orchid Monitoring Report – 2020 Growing Season - Filoha Meadows Nature Preserve Page 25 307600 307700 307800 307900 308000 308100 308200 308300 308400 308500 308600 4345000 4345000

3 4344900 4344900

29 1 28

4344800 2 4344800 4

5 4344700 4344700

6 4344600 4344600

7 8 4344500 4344500 9 10 11

12 13 26 4344400 4344400

14 17 4344300 4344300 15

18 16 4344200 4344200

20 4344100 4344100 22 23 4344000 4344000

24 25 4343900 4343900

BASE: Pitkin County 6-inch aerials (2014) Map 1. Occurrences of Epipactis gigantea GRID: UTM NAD83 Z13 Filoha Meadows Nature Preserve Open Space Piktin County, Colorado Legend Epipactis gigantea and ID No. Date: May 2020 Scale: 1:6,000 Filoha Meadows Open Space Boundary 1 inch = 500 feet .

Notes:Data collected with Trimble Geo7x GPS unit and differenctially corrected.

Stream Orchid Monitoring Report - 2020 Growing Season - Filoha Meadows Nature Preserve Page 26 307696 307796 307896 307996 308096 308196 4344788

13a ! 4344688 4344688

12a 4344588

4344588 ! 4344488 4344488 Transect No. Plant Name T1-T6 Canada thistle only 14a T7-T10 Canada thistle & Stream orchid ! 11a T11-T14 Stream orchid only ! 4344388 4344388 Treatment Methods Transect No. Rust Fungus T1, T4, T6 Control T2, T3, T5 Herbicide T7, T8 10a ! Hand-Pulling T9, T10 4344288 4344288

8a ! 9a ! MacroPlot 4a

7a ! 4344188 4344188 ! 2a ! 3a ! 5a ! 1a ! 4344088 4344088 6a !

BASE: Pitkin County 6-inch aerials (2014) GRID: UTM NAD83 Z13 Map 2.Transect Location Map Filoha Meadows Nature Preserve Open Space Legend Piktin County, Colorado Epipactis gigantea Date: May 2020 Open Space Boundary 1:3,600 5m x 50m Macroplot 1 inch = 300 feet . 12-meter Transects ! Transect number & start Notes:Data Points collected with Tribble Geo7x unit and differenctially corrected. Stream Orchid Monitoring Report - 2020 Growing Season - Filoha Meadows Nature Preserve Page 27 9.0 TRANSECT PHOTOS (2020)

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PHOTO 1. CIRSIUM TRANSECT T1. (JUL-09-2020)

PHOTO 2. CIRSIUM TRANSECT T2. (JUL-09-2020)

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PHOTO 3. CIRSIUM TRANSECT T3. (JUL-09-2020)

PHOTO 4. CIRSIUM TRANSECT T4. (JUL-09-2020)

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PHOTO 5. CIRSIUM TRANSECT T5. (JUL-09-2020)

PHOTO 6. CIRSIUM TRANSECT T6. (JUL-09-2020)

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PHOTO 7. TRANSECT T7A START. 17 DEG. HEADING (JUL-08- PHOTO 8. TRANSECT T7B END. (JUL-08-2020) 2020)

PHOTO 9. TRANSECT T8A START. 127 DEG. HEADING (JUL-08- PHOTO 10. TRANSECT T8B END. (JUL-08-2020) 2020)

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PHOTO 11. TRANSECT T9A START. 222 DEG. HEADING (JUL-08- PHOTO 12. TRANSECT T9B END. (JUL-08-2020) 2020)

PHOTO 13. TRANSECT T10A START. 16 DEG. HEADING (JUL-08- PHOTO 14. TRANSECT 10B END. (JUL-08-2020) 2020)

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PHOTO 15. TRANSECT T11A START. 4 DEG. HEADING (JUL-08- PHOTO 16. TRANSECT T11B END. (JUL-08-2020) 2020)

PHOTO 17. TRANSECT T12A START. 336 DEG. HEADING (JUL- PHOTO 18. TRANSECT T12B END. (JUL-08-2020) 08-2020)

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PHOTO 19. TRANSECT T13A START. 349 DEG. HEADING (JUL- PHOTO 20. TRANSECT T13B END. (JUL-08-2020) 08-2020)

PHOTO 21. TRANSECT T14A START. 13 DEG. HEADING (JUL-08- PHOTO 22. TRANSECT T14B END. (JUL-08-2020) 2020)

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10.0 MACROPLOT PHOTOS (2020)

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PHOTO 1. MACROPLOT QUADRAT 1. (JUL-08-2020)

PHOTO 2. MACROPLOT QUADRAT 2. (JUL-08-2020)

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PHOTO 3. MACROPLOT QUADRAT 3. (JUL-08-2020)

PHOTO 4. MACROPLOT QUADRAT 4. (JUL-08-2020)

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PHOTO 5. MACROPLOT QUADRAT 5. (JUL-08-2020)

PHOTO 6. MACROPLOT QUADRAT 6. (JUL-08-2020)

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PHOTO 7. MACROPLOT QUADRAT 7. (JUL-08-2020)

PHOTO 8. MACROPLOT QUADRAT 8. (JUL-08-2020)

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PHOTO 9. MACROPLOT QUADRAT 9. (JUL-08-2020)

PHOTO 10. MACROPLOT QUADRAT 10. (JUL-08-2020)

Stream Orchid Monitoring Report – 2020 Growing Season - Filoha Meadows Nature Preserve Page 41

APPENDIX A. 2018, 2019 & 2020 EPIPACTIS GIGANTEA RAW DATA - MACROPLOT

Stream Orchid Monitoring Report - 2020 Growing Season - Filoha Meadows Nature Preserve APPENDIX A. 2018, 2019 2020 EPIPACTIS GIGANTEA RAW DATA - MACROPLOT

Size of Quadrat # Stems in # Total Stems / Plot/Transect Quadrat Year (sq.m.) Flr # Stems Veg quadrat total/sqm M1 01N 2018 5 37 38 75 15.0 M1 01S 2018 5 33 24 57 11.4 M1 02N 2018 5 95 147 242 48.4 M1 02S 2018 5 78 105 183 36.6 M1 03N 2018 5 54 102 156 31.2 M1 03S 2018 5 51 98 149 29.8 M1 04N 2018 5 97 63 160 32.0 M1 04S 2018 5 93 46 139 27.8 M1 05N 2018 5 174 136 310 62.0 M1 05S 2018 5 127 164 291 58.2 M1 06N 2018 5 121 83 204 40.8 M1 06S 2018 5 105 83 188 37.6 M1 07N 2018 5 202 145 347 69.4 M1 07S 2018 5 165 70 235 47.0 M1 08N 2018 5 100 88 188 37.6 M1 08S 2018 5 115 56 171 34.2 M1 09N 2018 5 77 116 193 38.6 M1 09S 2018 5 62 83 145 29.0 M1 10N 2018 5 176 112 288 57.6 M1 10S 2018 5 167 62 229 45.8 M1 01N 2019 5 91 137 228 45.6 M1 01S 2019 5 80 79 159 31.8 M1 02N 2019 5 193 274 467 93.4

M1 02S 2019 5 186 178 364 72.8 M1 03N 2019 5 117 168 285 57.0 M1 03S 2019 5 130 161 291 58.2 M1 04N 2019 5 264 102 366 73.2 M1 04S 2019 5 264 137 401 80.2 M1 05N 2019 5 357 244 601 120.2 M1 05S 2019 5 391 236 627 125.4 M1 06N 2019 5 293 170 463 92.6 M1 06S 2019 5 331 202 533 106.6 M1 07N 2019 5 294 163 457 91.4 M1 07S 2019 5 252 106 358 71.6 M1 08N 2019 5 230 173 403 80.6 M1 08S 2019 5 204 100 304 60.8 M1 09N 2019 5 170 217 387 77.4 M1 09S 2019 5 114 148 262 52.4 M1 10N 2019 5 310 149 459 91.8 M1 10S 2019 5 266 139 405 81.0 M1 01N 2020 5 63 119 182 36.4

Stream Orchid Monitoring Report - 2020 Growing Season - Filoha Meadows Nature Preserve A1 APPENDIX A. 2018, 2019 2020 EPIPACTIS GIGANTEA RAW DATA - MACROPLOT

Size of Quadrat # Stems in # Total Stems / Plot/Transect Quadrat Year (sq.m.) Flr # Stems Veg quadrat total/sqm M1 01S 2020 5 33 76 109 21.8 M1 02N 2020 5 119 315 434 86.8 M1 02S 2020 5 62 172 234 46.8 M1 03N 2020 5 93 164 257 51.4 M1 03S 2020 5 55 149 204 40.8 M1 04N 2020 5 119 248 367 73.4 M1 04S 2020 5 96 180 276 55.2 M1 05N 2020 5 140 412 552 110.4 M1 05S 2020 5 110 334 444 88.8 M1 06N 2020 5 133 242 375 75.0 M1 06S 2020 5 107 332 439 87.8 M1 07N 2020 5 126 269 395 79.0 M1 07S 2020 5 145 225 370 74.0 M1 08N 2020 5 100 242 342 68.4 M1 08S 2020 5 56 160 216 43.2 M1 09N 2020 5 91 246 337 67.4 M1 09S 2020 5 58 181 239 47.8 M1 10N 2020 5 124 293 417 83.4 M1 10S 2020 5 86 245 331 66.2

Stream Orchid Monitoring Report - 2020 Growing Season - Filoha Meadows Nature Preserve A2 APPENDIX B. 2018, 2019 & 2020 EPIPACTIS GIGANTEA RAW DATA - TRANSECTS

Stream Orchid Monitoring Report - 2020 Growing Season - Filoha Meadows Nature Preserve APPENDIX B. 2018, 2019, 2020 EPIPACTIS GIGANTEA RAW DATA - TRANSECTS

Size of Quadrat # Stems in # Total Stems / Plot/Transect Quadrat Year (sq.m.) Flr # Stems Veg quadrat total/sqm T07 1 2018 0.36 6 4 10 27.8 T07 2 2018 0.36 4 28 32 88.9 T07 3 2018 0.36 8 34 42 116.7 T07 4 2018 0.36 29 24 53 147.2 T07 5 2018 0.36 9 30 39 108.3 T07 6 2018 0.36 11 60 71 197.2 T07 7 2018 0.36 9 27 36 100.0 T07 8 2018 0.36 8 25 33 91.7 T07 9 2018 0.36 0 10 10 27.8 T07 10 2018 0.36 2 5 7 19.4 T07 11 2018 0.36 3 6 9 25.0 T07 12 2018 0.36 2 7 9 25.0 T07 1 2019 0.36 10 9 19 52.8 T07 2 2019 0.36 17 29 46 127.8 T07 3 2019 0.36 18 34 52 144.4 T07 4 2019 0.36 40 35 75 208.3 T07 5 2019 0.36 22 22 44 122.2 T07 6 2019 0.36 28 51 79 219.4 T07 7 2019 0.36 21 35 56 155.6 T07 8 2019 0.36 10 13 23 63.9 T07 9 2019 0.36 13 11 24 66.7 T07 10 2019 0.36 5 8 13 36.1 T07 11 2019 0.36 8 14 22 61.1 T07 12 2019 0.36 6 11 17 47.2 T07 1 2020 0.36 0 19 19 52.8 T07 2 2020 0.36 0 54 54 150.0 T07 3 2020 0.36 3 57 60 166.7 T07 4 2020 0.36 7 76 83 230.6 T07 5 2020 0.36 0 67 67 186.1 T07 6 2020 0.36 7 84 91 252.8 T07 7 2020 0.36 1 59 60 166.7 T07 8 2020 0.36 1 19 20 55.6 T07 9 2020 0.36 0 14 14 38.9 T07 10 2020 0.36 0 14 14 38.9 T07 11 2020 0.36 3 23 26 72.2 T07 12 2020 0.36 0 20 20 55.6 T08 1 2018 0.36 1 0 1 2.8 T08 2 2018 0.36 3 2 5 13.9 T08 3 2018 0.36 4 3 7 19.4 T08 4 2018 0.36 1 1 2 5.6

Stream Orchid Monitoring Report - 2020 Growing Season - Filoha Meadows Nature Preserve B1 APPENDIX B. 2018, 2019, 2020 EPIPACTIS GIGANTEA RAW DATA - TRANSECTS

Size of Quadrat # Stems in # Total Stems / Plot/Transect Quadrat Year (sq.m.) Flr # Stems Veg quadrat total/sqm T08 5 2018 0.36 6 1 7 19.4 T08 6 2018 0.36 8 2 10 27.8 T08 7 2018 0.36 2 2 4 11.1 T08 8 2018 0.36 1 0 1 2.8 T08 9 2018 0.36 2 0 2 5.6 T08 10 2018 0.36 2 1 3 8.3 T08 11 2018 0.36 0 0 0 0.0 T08 12 2018 0.36 0 0 0 0.0 T08 1 2019 0.36 0 2 2 5.6 T08 2 2019 0.36 4 5 9 25.0 T08 3 2019 0.36 7 11 18 50.0 T08 4 2019 0.36 6 2 8 22.2 T08 5 2019 0.36 7 3 10 27.8 T08 6 2019 0.36 12 4 16 44.4 T08 7 2019 0.36 4 1 5 13.9 T08 8 2019 0.36 3 0 3 8.3 T08 9 2019 0.36 2 4 6 16.7 T08 10 2019 0.36 5 3 8 22.2 T08 11 2019 0.36 1 0 1 2.8 T08 12 2019 0.36 1 0 1 2.8 T08 1 2020 0.36 0 0 0 0.0 T08 2 2020 0.36 2 8 10 27.8 T08 3 2020 0.36 2 15 17 47.2 T08 4 2020 0.36 2 7 9 25.0 T08 5 2020 0.36 3 12 15 41.7 T08 6 2020 0.36 3 18 21 58.3 T08 7 2020 0.36 3 5 8 22.2 T08 8 2020 0.36 1 2 3 8.3 T08 9 2020 0.36 0 6 6 16.7 T08 10 2020 0.36 1 7 8 22.2 T08 11 2020 0.36 0 2 2 5.6 T08 12 2020 0.36 0 3 3 8.3 T09 1 2018 0.36 6 4 10 27.8 T09 2 2018 0.36 4 1 5 13.9 T09 3 2018 0.36 2 1 3 8.3 T09 4 2018 0.36 0 7 7 19.4 T09 5 2018 0.36 1 3 4 11.1 T09 6 2018 0.36 0 2 2 5.6 T09 7 2018 0.36 7 4 11 30.6 T09 8 2018 0.36 2 12 14 38.9

Stream Orchid Monitoring Report - 2020 Growing Season - Filoha Meadows Nature Preserve B2 APPENDIX B. 2018, 2019, 2020 EPIPACTIS GIGANTEA RAW DATA - TRANSECTS

Size of Quadrat # Stems in # Total Stems / Plot/Transect Quadrat Year (sq.m.) Flr # Stems Veg quadrat total/sqm T09 9 2018 0.36 6 3 9 25.0 T09 10 2018 0.36 4 3 7 19.4 T09 11 2018 0.36 11 4 15 41.7 T09 12 2018 0.36 2 0 2 5.6 T09 1 2019 0.36 12 7 19 52.8 T09 2 2019 0.36 21 10 31 86.1 T09 3 2019 0.36 3 5 8 22.2 T09 4 2019 0.36 9 8 17 47.2 T09 5 2019 0.36 7 7 14 38.9 T09 6 2019 0.36 5 1 6 16.7 T09 7 2019 0.36 9 11 20 55.6 T09 8 2019 0.36 14 12 26 72.2 T09 9 2019 0.36 17 17 34 94.4 T09 10 2019 0.36 13 7 20 55.6 T09 11 2019 0.36 6 8 14 38.9 T09 12 2019 0.36 0 0 0 0.0 T09 1 2020 0.36 2 17 19 52.8 T09 2 2020 0.36 5 25 30 83.3 T09 3 2020 0.36 0 11 11 30.6 T09 4 2020 0.36 0 18 18 50.0 T09 5 2020 0.36 2 15 17 47.2 T09 6 2020 0.36 0 5 5 13.9 T09 7 2020 0.36 1 23 24 66.7 T09 8 2020 0.36 5 15 20 55.6 T09 9 2020 0.36 5 34 39 108.3 T09 10 2020 0.36 7 20 27 75.0 T09 11 2020 0.36 4 17 21 58.3 T09 12 2020 0.36 1 0 1 2.8 T10 1 2018 0.36 0 14 14 38.9 T10 2 2018 0.36 3 16 19 52.8 T10 3 2018 0.36 12 23 35 97.2 T10 4 2018 0.36 6 9 15 41.7 T10 5 2018 0.36 6 10 16 44.4 T10 6 2018 0.36 9 19 28 77.8 T10 7 2018 0.36 6 25 31 86.1 T10 8 2018 0.36 8 33 41 113.9 T10 9 2018 0.36 10 32 42 116.7 T10 10 2018 0.36 14 47 61 169.4 T10 11 2018 0.36 4 45 49 136.1 T10 12 2018 0.36 5 40 45 125.0

Stream Orchid Monitoring Report - 2020 Growing Season - Filoha Meadows Nature Preserve B3 APPENDIX B. 2018, 2019, 2020 EPIPACTIS GIGANTEA RAW DATA - TRANSECTS

Size of Quadrat # Stems in # Total Stems / Plot/Transect Quadrat Year (sq.m.) Flr # Stems Veg quadrat total/sqm T10 1 2019 0.36 1 36 37 102.8 T10 2 2019 0.36 12 43 55 152.8 T10 3 2019 0.36 20 50 70 194.4 T10 4 2019 0.36 4 15 19 52.8 T10 5 2019 0.36 13 7 20 55.6 T10 6 2019 0.36 13 15 28 77.8 T10 7 2019 0.36 29 37 66 183.3 T10 8 2019 0.36 13 48 61 169.4 T10 9 2019 0.36 23 45 68 188.9 T10 10 2019 0.36 17 53 70 194.4 T10 11 2019 0.36 17 45 62 172.2 T10 12 2019 0.36 5 77 82 227.8 T10 1 2020 0.36 0 37 37 102.8 T10 2 2020 0.36 4 49 53 147.2 T10 3 2020 0.36 6 50 56 155.6 T10 4 2020 0.36 6 12 18 50.0 T10 5 2020 0.36 6 23 29 80.6 T10 6 2020 0.36 6 29 35 97.2 T10 7 2020 0.36 6 48 54 150.0 T10 8 2020 0.36 4 45 49 136.1 T10 9 2020 0.36 8 58 66 183.3 T10 10 2020 0.36 1 82 83 230.6 T10 11 2020 0.36 4 69 73 202.8 T10 12 2020 0.36 2 68 70 194.4 T11 1 2018 0.36 4 1 5 13.9 T11 2 2018 0.36 0 0 0 0.0 T11 3 2018 0.36 1 4 5 13.9 T11 4 2018 0.36 3 3 6 16.7 T11 5 2018 0.36 3 8 11 30.6 T11 6 2018 0.36 5 0 5 13.9 T11 7 2018 0.36 5 12 17 47.2 T11 8 2018 0.36 8 1 9 25.0 T11 9 2018 0.36 8 12 20 55.6 T11 10 2018 0.36 2 0 2 5.6 T11 11 2018 0.36 7 4 11 30.6 T11 12 2018 0.36 11 6 17 47.2 T11 1 2019 0.36 4 6 10 27.8 T11 2 2019 0.36 0 0 0 0.0 T11 3 2019 0.36 3 2 5 13.9 T11 4 2019 0.36 2 2 4 11.1

Stream Orchid Monitoring Report - 2020 Growing Season - Filoha Meadows Nature Preserve B4 APPENDIX B. 2018, 2019, 2020 EPIPACTIS GIGANTEA RAW DATA - TRANSECTS

Size of Quadrat # Stems in # Total Stems / Plot/Transect Quadrat Year (sq.m.) Flr # Stems Veg quadrat total/sqm T11 5 2019 0.36 1 10 11 30.6 T11 6 2019 0.36 4 5 9 25.0 T11 7 2019 0.36 1 23 24 66.7 T11 8 2019 0.36 3 5 8 22.2 T11 9 2019 0.36 0 16 16 44.4 T11 10 2019 0.36 2 1 3 8.3 T11 11 2019 0.36 4 10 14 38.9 T11 12 2019 0.36 8 14 22 61.1 T11 1 2020 0.36 5 5 10 27.8 T11 2 2020 0.36 0 0 0 0.0 T11 3 2020 0.36 2 5 7 19.4 T11 4 2020 0.36 6 3 9 25.0 T11 5 2020 0.36 4 4 8 22.2 T11 6 2020 0.36 5 4 9 25.0 T11 7 2020 0.36 3 7 10 27.8 T11 8 2020 0.36 6 4 10 27.8 T11 9 2020 0.36 0 11 11 30.6 T11 10 2020 0.36 0 1 1 2.8 T11 11 2020 0.36 2 10 12 33.3 T11 12 2020 0.36 4 18 22 61.1 T12 1 2018 0.36 9 3 12 33.3 T12 2 2018 0.36 6 0 6 16.7 T12 3 2018 0.36 0 0 0 0.0 T12 4 2018 0.36 1 0 1 2.8 T12 5 2018 0.36 2 0 2 5.6 T12 6 2018 0.36 21 19 40 111.1 T12 7 2018 0.36 16 26 42 116.7 T12 8 2018 0.36 40 22 62 172.2 T12 9 2018 0.36 31 16 47 130.6 T12 10 2018 0.36 25 17 42 116.7 T12 11 2018 0.36 32 30 62 172.2 T12 12 2018 0.36 12 7 19 52.8 T12 1 2019 0.36 8 6 14 38.9 T12 2 2019 0.36 7 2 9 25.0 T12 3 2019 0.36 0 0 0 0.0 T12 4 2019 0.36 1 1 2 5.6 T12 5 2019 0.36 3 0 3 8.3 T12 6 2019 0.36 37 13 50 138.9 T12 7 2019 0.36 40 28 68 188.9 T12 8 2019 0.36 38 33 71 197.2

Stream Orchid Monitoring Report - 2020 Growing Season - Filoha Meadows Nature Preserve B5 APPENDIX B. 2018, 2019, 2020 EPIPACTIS GIGANTEA RAW DATA - TRANSECTS

Size of Quadrat # Stems in # Total Stems / Plot/Transect Quadrat Year (sq.m.) Flr # Stems Veg quadrat total/sqm T12 9 2019 0.36 38 35 73 202.8 T12 10 2019 0.36 19 28 47 130.6 T12 11 2019 0.36 37 36 73 202.8 T12 12 2019 0.36 9 14 23 63.9 T12 1 2020 0.36 10 8 18 50.0 T12 2 2020 0.36 10 4 14 38.9 T12 3 2020 0.36 0 0 0 0.0 T12 4 2020 0.36 1 2 3 8.3 T12 5 2020 0.36 3 2 5 13.9 T12 6 2020 0.36 16 47 63 175.0 T12 7 2020 0.36 14 35 49 136.1 T12 8 2020 0.36 20 28 48 133.3 T12 9 2020 0.36 10 54 64 177.8 T12 10 2020 0.36 22 35 57 158.3 T12 11 2020 0.36 22 50 72 200.0 T12 12 2020 0.36 6 10 16 44.4 T13 1 2018 0.36 6 10 16 44.4 T13 2 2018 0.36 7 36 43 119.4 T13 3 2018 0.36 9 32 41 113.9 T13 4 2018 0.36 2 1 3 8.3 T13 5 2018 0.36 1 4 5 13.9 T13 6 2018 0.36 0 1 1 2.8 T13 7 2018 0.36 7 8 15 41.7 T13 8 2018 0.36 22 11 33 91.7 T13 9 2018 0.36 22 34 56 155.6 T13 10 2018 0.36 29 17 46 127.8 T13 11 2018 0.36 22 10 32 88.9 T13 12 2018 0.36 0 0 0 0.0 T13 1 2019 0.36 11 40 51 141.7 T13 2 2019 0.36 13 36 49 136.1 T13 3 2019 0.36 21 31 52 144.4 T13 4 2019 0.36 3 1 4 11.1 T13 5 2019 0.36 3 7 10 27.8 T13 6 2019 0.36 0 2 2 5.6 T13 7 2019 0.36 8 9 17 47.2 T13 8 2019 0.36 16 22 38 105.6 T13 9 2019 0.36 32 41 73 202.8 T13 10 2019 0.36 22 24 46 127.8 T13 11 2019 0.36 26 13 39 108.3 T13 12 2019 0.36 3 1 4 11.1

Stream Orchid Monitoring Report - 2020 Growing Season - Filoha Meadows Nature Preserve B6 APPENDIX B. 2018, 2019, 2020 EPIPACTIS GIGANTEA RAW DATA - TRANSECTS

Size of Quadrat # Stems in # Total Stems / Plot/Transect Quadrat Year (sq.m.) Flr # Stems Veg quadrat total/sqm T13 1 2020 0.36 3 53 56 155.6 T13 2 2020 0.36 2 53 55 152.8 T13 3 2020 0.36 10 51 61 169.4 T13 4 2020 0.36 11 5 16 44.4 T13 5 2020 0.36 2 11 13 36.1 T13 6 2020 0.36 0 1 1 2.8 T13 7 2020 0.36 7 15 22 61.1 T13 8 2020 0.36 13 28 41 113.9 T13 9 2020 0.36 9 63 72 200.0 T13 10 2020 0.36 14 40 54 150.0 T13 11 2020 0.36 11 26 37 102.8 T13 12 2020 0.36 4 5 9 25.0 T14 1 2018 0.36 6 18 24 66.7 T14 2 2018 0.36 8 19 27 75.0 T14 3 2018 0.36 5 18 23 63.9 T14 4 2018 0.36 8 18 26 72.2 T14 5 2018 0.36 12 28 40 111.1 T14 6 2018 0.36 17 25 42 116.7 T14 7 2018 0.36 11 40 51 141.7 T14 8 2018 0.36 13 52 65 180.6 T14 9 2018 0.36 18 51 69 191.7 T14 10 2018 0.36 10 37 47 130.6 T14 11 2018 0.36 5 14 19 52.8 T14 12 2018 0.36 9 5 14 38.9 T14 1 2019 0.36 8 28 36 100.0 T14 2 2019 0.36 23 29 52 144.4 T14 3 2019 0.36 15 26 41 113.9 T14 4 2019 0.36 11 27 38 105.6 T14 5 2019 0.36 9 52 61 169.4 T14 6 2019 0.36 11 46 57 158.3 T14 7 2019 0.36 10 67 77 213.9 T14 8 2019 0.36 10 97 107 297.2 T14 9 2019 0.36 17 75 92 255.6 T14 10 2019 0.36 6 66 72 200.0 T14 11 2019 0.36 9 25 34 94.4 T14 12 2019 0.36 5 9 14 38.9 T14 1 2020 0.36 11 30 41 113.9 T14 2 2020 0.36 7 55 62 172.2 T14 3 2020 0.36 9 37 46 127.8 T14 4 2020 0.36 0 29 29 80.6

Stream Orchid Monitoring Report - 2020 Growing Season - Filoha Meadows Nature Preserve B7 APPENDIX B. 2018, 2019, 2020 EPIPACTIS GIGANTEA RAW DATA - TRANSECTS

Size of Quadrat # Stems in # Total Stems / Plot/Transect Quadrat Year (sq.m.) Flr # Stems Veg quadrat total/sqm T14 5 2020 0.36 11 51 62 172.2 T14 6 2020 0.36 7 50 57 158.3 T14 7 2020 0.36 10 65 75 208.3 T14 8 2020 0.36 5 89 94 261.1 T14 9 2020 0.36 13 56 69 191.7 T14 10 2020 0.36 7 58 65 180.6 T14 11 2020 0.36 1 23 24 66.7 T14 12 2020 0.36 0 15 15 41.7

Stream Orchid Monitoring Report - 2020 Growing Season - Filoha Meadows Nature Preserve B8 APPENDIX C. 2018, 2019 & 2020 CIRSIUM ARVENSE RAW DATA – TRANSECTS

Stream Orchid Monitoring Report - 2020 Growing Season - Filoha Meadows Nature Preserve APPENDIX C. CIRSIUM ARVENSE 2018, 2019, 2020 RAW DATA Transect QuadratSize of Quadrat (sq.m.)Year Treatment No. Cirarv Stems No Stems/sq.m. T01 2 0.360 2018 Rust 12 33.33 T01 4 0.360 2018 Rust 4 11.11 T01 6 0.360 2018 Rust 13 36.11 T01 8 0.360 2018 Rust 13 36.11 T01 10 0.360 2018 Rust 16 44.44 T01 12 0.360 2018 Rust 6 16.67 T02 2 0.360 2018 Control 3 8.33 T02 4 0.360 2018 Control 4 11.11 T02 6 0.360 2018 Control 5 13.89 T02 8 0.360 2018 Control 12 33.33 T02 10 0.360 2018 Control 9 25.00 T02 12 0.360 2018 Control 18 50.00 T03 2 0.360 2018 Control 3 8.33 T03 4 0.360 2018 Control 2 5.56 T03 6 0.360 2018 Control 8 22.22 T03 8 0.360 2018 Control 8 22.22 T03 10 0.360 2018 Control 4 11.11 T03 12 0.360 2018 Control 10 27.78 T04 2 0.360 2018 Rust 7 19.44 T04 4 0.360 2018 Rust 10 27.78 T04 6 0.360 2018 Rust 5 13.89 T04 8 0.360 2018 Rust 4 11.11 T04 10 0.360 2018 Rust 11 30.56 T04 12 0.360 2018 Rust 12 33.33 T05 2 0.360 2018 Control 7 19.44 T05 4 0.360 2018 Control 9 25.00 T05 6 0.360 2018 Control 9 25.00 T05 8 0.360 2018 Control 12 33.33 T05 10 0.360 2018 Control 21 58.33 T05 12 0.360 2018 Control 21 58.33 T06 2 0.360 2018 Rust 4 11.11 T06 4 0.360 2018 Rust 0 0.00 T06 6 0.360 2018 Rust 8 22.22 T06 8 0.360 2018 Rust 4 11.11 T06 10 0.360 2018 Rust 5 13.89 T06 12 0.360 2018 Rust 8 22.22 T07 1 0.360 2018 Herbicide 3 8.33 T07 2 0.360 2018 Herbicide 5 13.89 T07 3 0.360 2018 Herbicide 1 2.78 T07 4 0.360 2018 Herbicide 5 13.89 T07 5 0.360 2018 Herbicide 4 11.11 T07 6 0.360 2018 Herbicide 4 11.11 T07 7 0.360 2018 Herbicide 1 2.78 T07 8 0.360 2018 Herbicide 8 22.22 T07 9 0.360 2018 Herbicide 2 5.56 T07 10 0.360 2018 Herbicide 4 11.11

Stream Orchid Monitoring Report - 2020 Growing Season - Filoha Meadows Nature Preserve C1 APPENDIX C. CIRSIUM ARVENSE 2018, 2019, 2020 RAW DATA Transect QuadratSize of Quadrat (sq.m.)Year Treatment No. Cirarv Stems No Stems/sq.m. T07 11 0.360 2018 Herbicide 9 25.00 T07 12 0.360 2018 Herbicide 9 25.00 T08 1 0.360 2018 Herbicide 10 27.78 T08 2 0.360 2018 Herbicide 4 11.11 T08 3 0.360 2018 Herbicide 5 13.89 T08 4 0.360 2018 Herbicide 6 16.67 T08 5 0.360 2018 Herbicide 7 19.44 T08 6 0.360 2018 Herbicide 3 8.33 T08 7 0.360 2018 Herbicide 5 13.89 T08 8 0.360 2018 Herbicide 4 11.11 T08 9 0.360 2018 Herbicide 7 19.44 T08 10 0.360 2018 Herbicide 3 8.33 T08 11 0.360 2018 Herbicide 4 11.11 T08 12 0.360 2018 Herbicide 2 5.56 T09 1 0.360 2018 Pull 4 11.11 T09 2 0.360 2018 Pull 5 13.89 T09 3 0.360 2018 Pull 2 5.56 T09 4 0.360 2018 Pull 1 2.78 T09 5 0.360 2018 Pull 2 5.56 T09 6 0.360 2018 Pull 6 16.67 T09 7 0.360 2018 Pull 2 5.56 T09 8 0.360 2018 Pull 3 8.33 T09 9 0.360 2018 Pull 3 8.33 T09 10 0.360 2018 Pull 4 11.11 T09 11 0.360 2018 Pull 5 13.89 T09 12 0.360 2018 Pull 8 22.22 T10 1 0.360 2018 Pull 4 11.11 T10 2 0.360 2018 Pull 4 11.11 T10 3 0.360 2018 Pull 3 8.33 T10 4 0.360 2018 Pull 4 11.11 T10 5 0.360 2018 Pull 1 2.78 T10 6 0.360 2018 Pull 2 5.56 T10 7 0.360 2018 Pull 1 2.78 T10 8 0.360 2018 Pull 3 8.33 T10 9 0.360 2018 Pull 2 5.56 T10 10 0.360 2018 Pull 0 0.00 T10 11 0.360 2018 Pull 1 2.78 T10 12 0.360 2018 Pull 0 0.00 T01 2 0.360 2019 Rust 4 11.11 T01 4 0.360 2019 Rust 7 19.44 T01 6 0.360 2019 Rust 3 8.33 T01 8 0.360 2019 Rust 4 11.11 T01 10 0.360 2019 Rust 8 22.22 T01 12 0.360 2019 Rust 2 5.56 T02 2 0.360 2019 Control 1 2.78 T02 4 0.360 2019 Control 4 11.11

Stream Orchid Monitoring Report - 2020 Growing Season - Filoha Meadows Nature Preserve C2 APPENDIX C. CIRSIUM ARVENSE 2018, 2019, 2020 RAW DATA Transect QuadratSize of Quadrat (sq.m.)Year Treatment No. Cirarv Stems No Stems/sq.m. T02 6 0.360 2019 Control 7 19.44 T02 8 0.360 2019 Control 10 27.78 T02 10 0.360 2019 Control 3 8.33 T02 12 0.360 2019 Control 7 19.44 T03 2 0.360 2019 Control 4 11.11 T03 4 0.360 2019 Control 7 19.44 T03 6 0.360 2019 Control 7 19.44 T03 8 0.360 2019 Control 10 27.78 T03 10 0.360 2019 Control 8 22.22 T03 12 0.360 2019 Control 5 13.89 T04 2 0.360 2019 Rust 8 22.22 T04 4 0.360 2019 Rust 9 25.00 T04 6 0.360 2019 Rust 7 19.44 T04 8 0.360 2019 Rust 12 33.33 T04 10 0.360 2019 Rust 16 44.44 T04 12 0.360 2019 Rust 15 41.67 T05 2 0.360 2019 Control 13 36.11 T05 4 0.360 2019 Control 1 2.78 T05 6 0.360 2019 Control 3 8.33 T05 8 0.360 2019 Control 1 2.78 T05 10 0.360 2019 Control 5 13.89 T05 12 0.360 2019 Control 6 16.67 T06 2 0.360 2019 Rust 4 11.11 T06 4 0.360 2019 Rust 2 5.56 T06 6 0.360 2019 Rust 2 5.56 T06 8 0.360 2019 Rust 4 11.11 T06 10 0.360 2019 Rust 7 19.44 T06 12 0.360 2019 Rust 5 13.89 T07 1 0.360 2019 Herbicide 0 0.00 T07 2 0.360 2019 Herbicide 0 0.00 T07 3 0.360 2019 Herbicide 2 5.56 T07 4 0.360 2019 Herbicide 0 0.00 T07 5 0.360 2019 Herbicide 2 5.56 T07 6 0.360 2019 Herbicide 2 5.56 T07 7 0.360 2019 Herbicide 0 0.00 T07 8 0.360 2019 Herbicide 1 2.78 T07 9 0.360 2019 Herbicide 0 0.00 T07 10 0.360 2019 Herbicide 0 0.00 T07 11 0.360 2019 Herbicide 3 8.33 T07 12 0.360 2019 Herbicide 0 0.00 T08 1 0.360 2019 Herbicide 11 30.56 T08 2 0.360 2019 Herbicide 3 8.33 T08 3 0.360 2019 Herbicide 1 2.78 T08 4 0.360 2019 Herbicide 3 8.33 T08 5 0.360 2019 Herbicide 1 2.78 T08 6 0.360 2019 Herbicide 0 0.00

Stream Orchid Monitoring Report - 2020 Growing Season - Filoha Meadows Nature Preserve C3 APPENDIX C. CIRSIUM ARVENSE 2018, 2019, 2020 RAW DATA Transect QuadratSize of Quadrat (sq.m.)Year Treatment No. Cirarv Stems No Stems/sq.m. T08 7 0.360 2019 Herbicide 2 5.56 T08 8 0.360 2019 Herbicide 2 5.56 T08 9 0.360 2019 Herbicide 2 5.56 T08 10 0.360 2019 Herbicide 1 2.78 T08 11 0.360 2019 Herbicide 1 2.78 T08 12 0.360 2019 Herbicide 12 33.33 T09 1 0.360 2019 Pull 1 2.78 T09 2 0.360 2019 Pull 1 2.78 T09 3 0.360 2019 Pull 1 2.78 T09 4 0.360 2019 Pull 0 0.00 T09 5 0.360 2019 Pull 5 13.89 T09 6 0.360 2019 Pull 0 0.00 T09 7 0.360 2019 Pull 0 0.00 T09 8 0.360 2019 Pull 0 0.00 T09 9 0.360 2019 Pull 4 11.11 T09 10 0.360 2019 Pull 1 2.78 T09 11 0.360 2019 Pull 5 13.89 T09 12 0.360 2019 Pull 2 5.56 T10 1 0.360 2019 Pull 0 0.00 T10 2 0.360 2019 Pull 4 11.11 T10 3 0.360 2019 Pull 2 5.56 T10 4 0.360 2019 Pull 2 5.56 T10 5 0.360 2019 Pull 1 2.78 T10 6 0.360 2019 Pull 1 2.78 T10 7 0.360 2019 Pull 0 0.00 T10 8 0.360 2019 Pull 1 2.78 T10 9 0.360 2019 Pull 1 2.78 T10 10 0.360 2019 Pull 0 0.00 T10 11 0.360 2019 Pull 2 5.56 T10 12 0.360 2019 Pull 0 0.00 T01 2 0.360 2020 Rust 0 0.00 T01 4 0.360 2020 Rust 0 0.00 T01 6 0.360 2020 Rust 0 0.00 T01 8 0.360 2020 Rust 0 0.00 T01 10 0.360 2020 Rust 0 0.00 T01 12 0.360 2020 Rust 0 0.00 T02 2 0.360 2020 Control 0 0.00 T02 4 0.360 2020 Control 1 2.78 T02 6 0.360 2020 Control 1 2.78 T02 8 0.360 2020 Control 0 0.00 T02 10 0.360 2020 Control 1 2.78 T02 12 0.360 2020 Control 1 2.78 T03 2 0.360 2020 Control 0 0.00 T03 4 0.360 2020 Control 0 0.00 T03 6 0.360 2020 Control 0 0.00 T03 8 0.360 2020 Control 0 0.00

Stream Orchid Monitoring Report - 2020 Growing Season - Filoha Meadows Nature Preserve C4 APPENDIX C. CIRSIUM ARVENSE 2018, 2019, 2020 RAW DATA Transect QuadratSize of Quadrat (sq.m.)Year Treatment No. Cirarv Stems No Stems/sq.m. T03 10 0.360 2020 Control 1 2.78 T03 12 0.360 2020 Control 1 2.78 T04 2 0.360 2020 Rust 3 8.33 T04 4 0.360 2020 Rust 4 11.11 T04 6 0.360 2020 Rust 0 0.00 T04 8 0.360 2020 Rust 3 8.33 T04 10 0.360 2020 Rust 5 13.89 T04 12 0.360 2020 Rust 4 11.11 T05 2 0.360 2020 Control 0 0.00 T05 4 0.360 2020 Control 0 0.00 T05 6 0.360 2020 Control 0 0.00 T05 8 0.360 2020 Control 0 0.00 T05 10 0.360 2020 Control 1 2.78 T05 12 0.360 2020 Control 1 2.78 T06 2 0.360 2020 Rust 0 0.00 T06 4 0.360 2020 Rust 0 0.00 T06 6 0.360 2020 Rust 0 0.00 T06 8 0.360 2020 Rust 0 0.00 T06 10 0.360 2020 Rust 0 0.00 T06 12 0.360 2020 Rust 0 0.00 T07 1 0.360 2020 Herbicide 0 0.00 T07 2 0.360 2020 Herbicide 0 0.00 T07 3 0.360 2020 Herbicide 0 0.00 T07 4 0.360 2020 Herbicide 0 0.00 T07 5 0.360 2020 Herbicide 0 0.00 T07 6 0.360 2020 Herbicide 0 0.00 T07 7 0.360 2020 Herbicide 0 0.00 T07 8 0.360 2020 Herbicide 0 0.00 T07 9 0.360 2020 Herbicide 0 0.00 T07 10 0.360 2020 Herbicide 0 0.00 T07 11 0.360 2020 Herbicide 0 0.00 T07 12 0.360 2020 Herbicide 0 0.00 T08 1 0.360 2020 Herbicide 0 0.00 T08 2 0.360 2020 Herbicide 0 0.00 T08 3 0.360 2020 Herbicide 0 0.00 T08 4 0.360 2020 Herbicide 0 0.00 T08 5 0.360 2020 Herbicide 0 0.00 T08 6 0.360 2020 Herbicide 0 0.00 T08 7 0.360 2020 Herbicide 1 2.78 T08 8 0.360 2020 Herbicide 1 2.78 T08 9 0.360 2020 Herbicide 0 0.00 T08 10 0.360 2020 Herbicide 0 0.00 T08 11 0.360 2020 Herbicide 0 0.00 T08 12 0.360 2020 Herbicide 0 0.00 T09 1 0.360 2020 Pull 2 5.56 T09 2 0.360 2020 Pull 1 2.78

Stream Orchid Monitoring Report - 2020 Growing Season - Filoha Meadows Nature Preserve C5 APPENDIX C. CIRSIUM ARVENSE 2018, 2019, 2020 RAW DATA Transect QuadratSize of Quadrat (sq.m.)Year Treatment No. Cirarv Stems No Stems/sq.m. T09 3 0.360 2020 Pull 0 0.00 T09 4 0.360 2020 Pull 0 0.00 T09 5 0.360 2020 Pull 0 0.00 T09 6 0.360 2020 Pull 0 0.00 T09 7 0.360 2020 Pull 0 0.00 T09 8 0.360 2020 Pull 0 0.00 T09 9 0.360 2020 Pull 0 0.00 T09 10 0.360 2020 Pull 0 0.00 T09 11 0.360 2020 Pull 0 0.00 T09 12 0.360 2020 Pull 0 0.00 T10 1 0.360 2020 Pull 1 2.78 T10 2 0.360 2020 Pull 1 2.78 T10 3 0.360 2020 Pull 1 2.78 T10 4 0.360 2020 Pull 0 0.00 T10 5 0.360 2020 Pull 1 2.78 T10 6 0.360 2020 Pull 0 0.00 T10 7 0.360 2020 Pull 0 0.00 T10 8 0.360 2020 Pull 0 0.00 T10 9 0.360 2020 Pull 0 0.00 T10 10 0.360 2020 Pull 0 0.00 T10 11 0.360 2020 Pull 1 2.78 T10 12 0.360 2020 Pull 1 2.78

Stream Orchid Monitoring Report - 2020 Growing Season - Filoha Meadows Nature Preserve C6 APPENDIX D. STATISTICAL TEST RESULTS

Stream Orchid Monitoring Report - 2020 Growing Season - Filoha Meadows Nature Preserve APPENDIX D