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Species-Level Identifications and Ecological Aspects of the Chironomidae in the Fountain Creek Watershed (Colorado) USA, 2007-2008

A Master’s Thesis

Presented to the Faculty of the

College of Science and Mathematics

Colorado State University-Pueblo Pueblo, Colorado

In Partial Fulfillment Of the Requirements for the Degree of

Master of Science (Biology)

Lisa K. Helland

Colorado State University-Pueblo

August 2014

DEDICATION

I would like to dedicate this thesis to Dr. James Sublette (late Emeritus Professor of Biology,

CSU-Pueblo). He was truly one of the greatest chironomid biologists. I was very fortunate to work with him on chironomid identifications at his home in Tucson, Arizona on three separate visits. These trips were an invaluable learning experience as Dr. Sublette had so much information to share concerning chironomid identification and life histories. I am very appreciative of his time spent to identify to species my 2007 and 2008 chironomid collection from the Fountain Creek Watershed, Colorado.

ACKNOWLEDGEMENTS

I would like to thank all the members of my committee.

Dr. Scott Herrmann, thank you so much for your guidance and patience. After all the years you have in teaching biology, your enthusiasm was never lacking. That is the greatest motivation there is for a student and it certainly kept me from becoming discouraged throughout this writing process. Your contributions to CSU-Pueblo in aquatic sciences-- teaching and research-- are outstanding and I feel very fortunate to have been a part of your chironomid team!

Dr. Igor Melnykov, thank you for your time and for the incredible support with statistical analysis. This aspect of my thesis would not have been possible without you.

Dr. Del Nimmo, I am so glad you became a member of my committee. I have appreciated our conversations about this thesis and your thoughtful feedback. I also want to thank you for your time when you made yourself available to come along during my night sampling at the Dorchester Park site in Colorado Springs.

Dr. Brian Vandenheuvel, thank you for being a member of my committee. You have a great way of explaining things and I appreciate your time.

Thanks to those individuals who also contributed to this research.  John Romine for his expert midge dissection and slide preparation.  Christie Kleinert for her help in midge collection at the Monument Creek sites.  Jim Carsella for advice on water chemistry questions.  Others…You know to whom I am referring 

A very special thank you to Christopher Helland who helped me with many computer issues, you truly saved your Mom on that one!

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ABSTRACT

Species-Level Identifications and Ecological Aspects of the Chironomidae in the Fountain Creek Watershed (Colorado) USA, 2007-2008

Adult male chironomids were collected from 14 sites in the Fountain Creek Watershed using ultraviolet night lighting and sweep net techniques. Species level identification resulted in 155 species being identified including 24 undescribed species from 65 genera and 7 subfamilies/tribes. The Jaccard Coefficient of Community and Sørensen Similarity Coefficient indicated that the sites in closest proximity shared the most common species, yet a healthy variation of species existed throughout the watershed. Differences in species distributions due to elevation were evident as expected. Substrate sediment samples were collected and analyzed by size ranging from small cobble to silt/clay with coarse to fine sand being the predominate sediment. Canonical Correspondence Analysis (CCA) was used to determine which environmental variable might contribute most to community variations and distributions. Sediment was determined not to be a significant variable for species presence or absence. Selenium (Se) was determined to be the most significant contributing variable. Mean total, dissolved and pore selenium concentrations for fall and spring 2007 water samples were analyzed by CCA. Spring total, dissolved and pore Se p-values were 0.0006, 0.0006, and 0.0034, respectively; p-values for fall total and dissolved Se analysis were 0.05 and 0.0044 with fall pore Se having a nonsignificant value of 0.1266. The CCA also revealed that sites LF-4 and LF-5 located in areas of seleniferous leaching of pierre shale formations were most closely associated with high selenium concentrations.

Table of Contents

Page

Dedication iii

Acknowledgements iv

Abstract v

List of Figures viii-x

List of Tables xi

Introduction 1

Hypothesis and Specific Aims 5

Materials and Methods 6

Study Sites 6

Chironomidae Collection Methods 8

Chironomidae Dissection and Identification Procedures 9

Sediment Collection and Sieving Methods 9

Water Collection Methods for Selenium 13

Results 14

Chironomid Data 14

Physical Characteristics 35

Similarity Indices 39

Sediment Characterization 42

Canonical Correspondence Analysis 43

Discussion 50

Comparison to Western Rivers and other Aquatic Environments 50

Conclusion 57

References Cited 59

Appendix A: Collecting Site Photos Upstream and Downstream: 62 Figures 1- 30

Appendix B: Similarity Indices : Figures 42 - 53 77

Appendix C: Canonical Correspondence Analysis (CANOCO) Logs 84

List of Figures

Page

Figure 1: Sampling Site Location Map 7

Figure 2: Ultraviolet Night Light and Sweep Net Photo 8

Figure 3: Adult Chironomid Dissection Slide 10

Figure 4: Sediment Sample in PETE Collection Jar 10

Figure 5: Sediment Samples on Drying Rack 11

Figure 6: Sediment Sieves for Separating Substrate 12

Figure 7: Representative Sediments in Sieves and Weighing Boat 12

Figure 8: Pore Water Sampling Device 13

Figure 9: Chironomid Composition by Subfamily/Tribe 22

Figure 10: Percent Subfamily Composition for MC-1 2007 22

Figure 11: Percent Subfamily Composition for MC-1, 2008 23

Figure 12: Percent Subfamily Composition for MC-2, 2007 23

Figure 13: Percent Subfamily Composition for MC-2, 2008 23

Figure 14: Percent Subfamily Composition for MC-3, 2007 23

Figure 15: Percent Subfamily Composition for MC-3, 2008 24

Figure 16: Percent Subfamily Composition for MC-4, 2007 24

Figure 17: Percent Subfamily Composition for MC-4, 2008 24

Figure 18: Percent Subfamily Composition for MC-5, 2007 24

Figure 19: Percent Subfamily Composition for MC-5, 2008 25

Figure 20: Percent Subfamily Composition for UF-1, 2007 25

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Figure 21: Percent Subfamily Composition for UF-1, 2008 25

Figure 22: Percent Subfamily Composition for UF-2, 2007 26

Figure 23: Percent Subfamily Composition for UF-2, 2008 26

Figure 24: Percent Subfamily Composition for UF-3, 2007 26

Figure 25: Percent Subfamily Composition for UF-3, 2008 27

Figure 26: Percent Subfamily Composition for UF-4, 2007 27

Figure 27: Percent Subfamily Composition for UF-4, 2008 27

Figure 28: Percent Subfamily Composition for LF-1, 2007 28

Figure 29: Percent Subfamily Composition for LF-1, 2008 28

Figure 30: Percent Subfamily Composition for LF-2, 2007 28

Figure 31: Percent Subfamily Composition for LF-2, 2008 29

Figure 32: Percent Subfamily Composition for LF-3, 2007 29

Figure 33: Percent Subfamily Composition for LF-3, 2008 29

Figure 34: Percent Subfamily Composition for LF-4, 2007 30

Figure 35: Percent Subfamily Composition for LF-4, 2008 30

Figure 36: Percent Subfamily Composition for LF-5, 2007 30

Figure 37: Percent Subfamily Composition for LF-5, 2008 31

Figure 38: Number of Species Collected by UV and Sweep, 2007 31

Figure 39: Number of Species Collected by UV and Sweep, 2008 32

Figure 40: Percentage of New Species Collected Per Site 34

Figure 41: Percentage of New Species Collected by UV, Sweep or Both 34

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Figure 54: Spring Total Selenium/Silt and Clay/pH—by Sites 44

Figure 55: Spring Total Selenium/Silt and Clay/pH—by Species 44

Figure 56: Spring Dissolved Selenium/Silt and Clay/pH—by Sites 45

Figure 57: Spring Dissolved Selenium/Silt and Clay/pH—by Species 45

Figure 58: Spring Pore Selenium/Silt and Clay/pH—by Sites 46

Figure 59: Spring Pore Selenium/Silt and Clay/pH—by Species 46

Figure 60: Fall Total Selenium/Silt and Clay/pH—by Sites 47

Figure 61: Fall Total Selenium/Silt and Clay/pH—by Species 47

Figure 62: Fall Dissolved Selenium/Silt and Clay/pH—by Sites 48

Figure 63: Fall Dissolved Selenium/Silt and Clay/pH—by Species 48

Figure 64: Fall Pore Selenium/Silt and Clay/pH—by Sites 49

Figure 65: Fall Pore Selenium/Silt and Clay/pH—Species 49

List of Tables

Page

Table 1: Universal Transverse Mercator (UTM) Coordinates 6

Table 2: List of Chironomid Species Collected 15

Table 3: List of New Chironomid Species 21

Table 4: Variation of Chironomid Species by Collection Method For Site LF-3 on 3/9/2007 32

Table 5: Variation of Chironomid Species by Collection Method 33 For Site MC-5 on 9/6/2007

Table 6: Variation of Chironomid Species by Collection Method 33 For Site LF-4 on 9/1/2007

Table 7: Mean Concentration for Water Quality Parameters 37

Table 8: Elevation Differences Between Sites and Apparent Species 38 Diversity

Table 9: Matrix of Jaccard’s Similarity Coefficient vs Sørensen’s Similarity 41 Coefficient

Table 10: Mean Percentage of Fraction Sizes for Sediment Samples 42

Table 11: Chironomid Species collected by Kleinert (2008) and 52 Powell (2008).

Introduction

The Fountain Creek watershed is located within the Arkansas River Basin, Colorado, and encompasses a drainage area of 930 square miles. This watershed is bordered by Pikes

Peak to the west, the Palmer-Divide to the north, and Chico Basin to the east (USACE 2009).

Fountain Creek and Monument Creek are the two main drainages within this watershed

(Bruce, 2002). Fountain Creek, a perennial stream, originates in Woodland Park, Colorado and flows south to its confluence with the Arkansas River at Pueblo. Monument Creek, the main tributary of Fountain Creek drains 119 square miles, while the upper Fountain drains

237 square miles (USACE 2009). This study area has elevations ranging from 14,109 feet at the base of Pikes Peak to 4642 feet at Pueblo (Edelmann, 2002). Monument and Fountain

Creeks flow through eight communities within the watershed.

The streambed in the upper Fountain consists of larger boulders and cobble as well as sand and gravel, and the substrate in Monument Creek contains deep coarse sand derived from sandstone (von Guerard, 1989). The lower Fountain Creek is primarily a sandy bottom stream consisting of varying degrees of cobble and gravel and becomes braided as it flows southward into a wide alluvial valley dominated by Pierre shale deposits (von Guerard, 1989).

The Pierre shale deposits that are found predominantly in the southern branch of the lower Fountain Creek contain selenium that is continually leached into the water column in varying degrees as the shale is weathered and eroded. Selenium exists in four oxidation states from +6 to -2 and this speciation determines the bioavailability and toxicity to aquatic organisms (Martens, 2003). Further, water chemistry conditions such as pH can be a good predictor of existing selenium species (Martens, 2003). Turner (2009) found that total and dissolved water fractions taken at the same sites as this work and evaluated with ICP-MS

1 indicated the trend of increasing selenium values as Fountain Creek enters the Pierre shale formations, as well as increasing pH values southward to the confluence with the Arkansas

River.

Although selenium is a naturally occurring micronutrient necessary for metabolic function in aquatic organisms (EPA, 2004), it can be toxic to aquatic organisms at higher levels possibly resulting in bioaccumulation—the sequestering of elements into the tissues of aquatic organisms. Turner (2009) found that bioaccumulation of selenium occurred in bryophyte tissues at varying concentrations in the Fountain Creek watershed. Also, McGarvy

(2011), using the same Monument/Fountain Creek sites as this thesis, measured selenium bioaccumulation in whole body homogenates as well as ovarian, skin, liver, and muscle tissues in 6 fish species. Increased selenium concentrations were found downstream from sites LF-2 to LF-5 with the highest concentrations for both whole body and tissue samples to be those from LF-4 and LF-5. Further, (GEI Consultants, Inc, 2007) determined that mean selenium concentrations in fish tissues in the lower Fountain Creek increased from their upstream site at Fountain, CO to the Arkansas River confluence. The current EPA surface water chronic standard is 4.6 ppb (EPA, 2001).

Not only are aquatic plants and fish good bioindicators of their physical environment but also, chironomids--macroinvertebrates belonging to the order Diptera. Voshell (2002) stated that chironomids can comprise at least half of the species in an aquatic community.

Chironomids exist on every continent and species can be found in wide ranges of temperature, pH, salinity, oxygen levels, current speeds, depth, productivity, altitude and latitude

(Coffman, 1996). As such, Coffman (1996) further commented that ecologists strive to determine the ecological health of both lotic and lentic systems dependent upon certain

2 chironomid species that are found within these habitat ranges. Chironomids are holometabolous exhibiting 4 stages of life history-egg, larva, pupa and adult. Since

3 life-stages of these insects are spent in the sediments, their use in evaluating aquatic environments has great potential. Saether stated:

As a result of generally long lifecycles, the consequences both of continuing and of occasional disturbances are integrated in the distribution and occurrence of the benthos. Thus the benthos can give information impossible to obtain by merely chemical factors (Saether, 1979).

As such, the presence or absence of certain chironomid species at varying sites along with habitat evaluation can provide a means to assess the integrity or environmental condition of a riverine ecosystem. Bruce (2002) stated:

“habitat assessment promotes an understanding of the relations among physical factors that might be limiting to biological communities and provides baseline information necessary to identify future changes in habitat conditions.”

Since the substrate in the Fountain Creek ranges from small cobble and gravel to sand, a variety of habitat preferences or niches are available to chironomids. Thus, characterization of a stream’s substrate at varying sites can provide information on habitat availability. (Bruce,

2002) determined that “substrate particle size accounted for most of the variation in invertebrate community structure among sites in the Fountain Creek basin”.

Similarity Indices are commonly used to measure the likeness of species composition between two sites. Bruce (2002) used the Jaccard Coefficient of Community to determine

Chironomid community structure variations in spring and fall between sites in the Fountain

Creek Watershed. Bruce (2002) determined that similarity in community was greater for

3 tributary sites than for sites located on the main stem. von Guerard (1989) used the Similarity

Index to evaluate midge taxon similarity between five sites in Monument and Fountain Creek.

He determined that two of the ten compared sites shared the greatest similarity.

(Ruse, 2006) collected pupal exuviae exoskeletons from 51 lakes in the United Kingdom, which were identified to species, to determine the ranges of total phosphorus and pH preferred by chironomid species. Using forward stepwise regression, Ruse (2006) concluded that conductivity, catchment area, pH, total phosphorus/mean depth, nitrate, volume, latitude, and alkalinity was the order in which one could explain habitat preferences for varying species.

In this research, identifications were completed to the species-level to best assess the factors of the physical environment that may be limiting to these organisms thereby influencing their distributions. In addition to my research, species-level studies have been completed on segments of the main-stem of the Arkansas River: the upper Arkansas by Ruse, et. al. (2000) and the lower Arkansas by Kleinert (2008) and Powell (2008). Bruce (2008) stated, “to date, no systematic survey of the Colorado aquatic insect fauna has focused on plains streams” From further literature review, it is apparent that there have been no species- level studies of chironomids in an entire watershed of a Colorado sandy-bottomed stream.

This research represents a systematic survey of the adult chironomid fauna identified to species in the Fountain Creek watershed, a front- range plains stream. Species-level studies of the entire Fountain Creek watershed would fill a major void in our understanding of chironomid communities in western sandy-bottomed streams; thus, leading to my research questions concerning chironomid diversity and the environmental variables that may influence their distributions.

Hypothesis and Specific Aims

Research Questions:

 What is the chironomid species diversity in the Fountain Creek Watershed?

 What environmental variables are influencing the distributions of chironomid species

in the Fountain Creek Watershed?

Hypothesis:

 I hypothesized that some chironomid species distributions are limited by elevation,

substrate, and selected water quality parameters.

With this hypothesis in mind, I generated the following specific aims.

Specific Aims:

 Collection of adult chironomids using two different collection methods (ultraviolet

and sweep net) at fourteen sites within the Fountain Creek Watershed.

 Collection of three surficial sediment samples at each of the fourteen sites within the

Fountain Creek Watershed to assess the varying physical substrates.

 Evaluate chironomid distributions using water quality data for total, dissolved, and

pore selenium from water samples collected at each site and analyzed by ICP-MS.

 Evaluate chironomid distributions using the following abiotic water parameters at

each site: dissolved oxygen, temperature, pH, and specific conductivity.

Materials and Methods

Study Sites

Sampling sites in the Fountain Creek Watershed for this research included 4 sites in the upper Fountain Creek reach (UF 1-4), 5 sites in the Monument Creek reach (MC 1-5), and 5 sites in the lower Fountain reach (LF 1-5) (Figure 1). These sites are consistent with the sites used by Turner (2009). Site photos can be found in Appendix A. Specific site coordinates, descriptions and elevations are described in Table 1.

Table 1. Universal Transverse Mercator (UTM) coordinates, elevation and physical description for each site. Site Lat (N) Long (W) Elevation (M) Physical Description of Site Location

UF1 38.92691 -105.004 2334.77 UFC at Green Mountain Falls, CO

UF2 38.85948 -104.920 1940.66 UFC at Manitou Springs, CO

UF3 38.84629 -104.866 1859.89 UFC at 26th St., Colorado Springs, CO

UF4 38.83015 -104.842 1818.44 UFC at 8th St., Colorado Springs, CO

MC1 39.08271 -104.876 2097.22 MC at Mt. Herman Rd. near Monument, CO

MC2 39.02415 -104.844 2031.80 MC at N. Entrance, U.S. A. F. A., CO

MC3 38.95424 -104.834 1942.80 MC at S. Entrance, U.S.A.F.A., CO

MC4 38.93322 -104.817 1942.20 MC at Woodmen Rd., Colorado Springs, CO

MC5 38.84282 -104.828 1840.69 MC at Colorado College, Colo. Spgs., CO

LF1 38.81613 -104.822 1798.02 LFC at Nevada St., Colo. Spgs., CO

LF2 38.60245 -104.670 1634.95 LFC, S. Fountain, CO

LF3 38.42975 -104.598 1532.84 LFC at Pinon Bridge

LF4 38.27893 -104.602 1432.86 LFC at Hwy. 50, Pueblo, CO

LF5 LFC upstream of confluence with Arkansas 38.25572 -104.591 1414.88 River, Pueblo, CO

Figure 1. Location of sampling sites in the Fountain Creek Watershed (Turner, 2009).

Chironomidae Collection Methods

Adult chironomids were collected in the summer of 2007 (September 1-7) and 2008

(July 10-August 21) from 14 sites within the Fountain Creek Watershed. Figures 1– 28 in

Appendix A show the downstream and upstream views of all 14 sites. Sweep net and long- wave ultraviolet (UV) lamps were utilized in the collection process (Figure 2). Aerial swarm sweep net collections were conducted in the early evening using a 0.1 mm mesh net, 28 cm in diameter by 60 cm in length. Netted specimens were placed into a labeled jar containing

75% ethanol. Night light (UV) collections were conducted for one full hour after sunset by placing the ultraviolet lamp between two rectangular aluminum loaf pans filled with 75% ethanol (Figure 2). Insects collected were carefully poured from the collection pans into a labeled jar. The collected insects were later sorted specifically for male chironomid species and stored in jars of 100% undenatured ethanol and appropriately labeled with date, location and collection method.

(a) (b) Figure 2. Ultraviolet (UV) night light setup (A) and sweep net methods (B) used for aerial collections. Photo by L. Helland.

Chironomidae Dissection and Identification Procedures

Micro-dissections were performed on adult male specimens for identification purposes using a modification of slide mounting procedures as described by Wiederholm

(1989). Each male specimen was placed into a drop of euparal on a glass slide. Each wing was removed and placed on the upper left portion of the slide. One prothoracic, mesothoracic and metathoracic leg including the trochanter was removed from the thorax and placed in order directly below the wings. The antennae were also removed and aligned to the right of the mesothoracic leg. The head, thorax and abdomen were placed in a 12% KOH macerating solution for 24-48 hours dependent on specimen size to “clear” the internal anatomy. Once cleared, the KOH was neutralized with water. Another drop of euparal was added to the right side of the slide. The thorax was removed and placed to the right of the wing, and the head capsule was placed to the right of the antennae. Lastly, the abdomen was placed dorsal side up with the male anatomy toward the inside of the slide to prevent possible damage to the male sex organs from coverslipping. The euparal was thinned to allow for efficient drying for a period of two weeks. Once dry, the slide specimens were coverslipped using 12-18mm diameter coverslips depending on specimen size as shown in Figure 3. Completed slides were left to dry for four weeks before the identification process could begin. Identification of chironomid males to species level was completed by Dr. James E. Sublette (late Emeritus

Professor of Biology, CSU-Pueblo).

Figure 3. A completed adult chironomid dissection slide. Wings and legs are located on the left. The thorax, antennae, head and abdomen are on the right. Photo by C. Kleinert.

Sediment Collection and Sieving Methods

Three sediment samples were collected in March 2012, along a bank to bank transect at each of the 14 sites. The samples were collected with polyethylene terephthalate (PETE) bottles measuring 122mm in height by 64mm in diameter, with a total volume of 480 ml.

(Figure 4). The open mouth of the bottle was forced completely into the sediment filling the entire container.

Figure 4. Sediment sample in clear plastic (PETE) collection jar.

These samples were placed in pans for 48 hours at 75° C to dry (Figure 5) and later placed into labeled plastic freezer bags according to site and sample number. Each sample was sieved through 8 sieve size fractions, each sieve standardized according to the grain size classification system as described by Wentworth (1922). Stacked sieves ranged from small cobble (65-257mm), very-coarse gravel (32-64mm), coarse gravel (16-32mm), medium gravel (8-16mm), fine gravel (4-8mm), very-fine gravel (2-4mm), coarse to fine sand (0.07-

3mm), and silt and clay (4μm-62.5μm). Each fraction was placed into a lightweight plastic dish and weighed to 0.01 gram using a Denver Instrument XL-3100 balance (Figures 6 and

7). The weight of each sieve fraction was determined and fractions averaged. The weights were converted to mean percentiles of the whole.

Figure 5. Sediment samples in pans on drying rack.

(a) (b)

Figure 6. Sieves used to separate substrate sizes. (a) Sieve fraction vertical stack and (b) view from top down.

(a) (b)

Figure 7. (a) Representative sediments in sieves and (b) in weighing boat. Photos by L. Helland.

Water Collection Methods for Selenium

Dissolved, total and pore water fractions were measured by J. Turner (2009) in spring and fall 2007. Total water fractions were taken with a high density polyethylene syringe, dissolved fractions were also taken with a high density polyethylene syringe and then passed through a 32mm Acrodisc syringe filter with a 0.45 micrometer pore size. Pore water was sampled using an originally designed pore water sampling tool which consisted of a polyethylene syringe hot glued to polyethylene tubing connected to a wooden dowel (Figure

8). The sample end was inserted 10 cm into the sediment and sediment water was extracted up into the syringe.

Figure 8. Pore water-sampling device with 60 ml syringe at the top and rigid polyethylene tubing at the bottom. Photo by L. Helland.

Results

Chironomid Data

This systematic study of 714 chironomid specimens, 309 collected in 2007 and 405 collected in 2008 resulted in 154 total species (Table 2) from 65 genera and 7 subfamilies/tribes--Chironomini, Diamesinae, Orthocladiinae, Podonominae, Prodiamesinae,

Tanypodinae, and Tanytarsini (Figure 9). Orthocladiinae and Chironomini were the dominant subfamilies/tribes at 45% and 29% followed by Tanypodinae at 12%, Tanytarsini at 8%, Prodiamesinae at 3%, Podonominae at 2% and Diamesinae at 1%. Further, 24

“new/undescribed” species were included in this total representing 19 genera and 4 subfamilies – Chironomini, Orthocladiinae, Tanypodinae, and Tanytarsini (Table 3).

Table 2. List of species collected from fourteen sites in the Fountain Creek Watershed. A1 indicates species presence at a particular site, 0 indicates absence.

1 2 3 4 5

1 2 3 4

1 2 3 4 5

- - - - -

- - - -

- - - - -

LF LF LF LF LF

UF UF UF UF Code Order Subfamily/Tribe Genus Species MC MC MC MC MC CH_A Diptera Chironomini Chironomus atrella 0 0 0 1 0 0 0 0 0 0 1 0 0 1 CH_D Diptera Chironomini Chironomus decorus 1 1 1 1 1 1 0 1 1 1 1 1 1 1 CH_DL Diptera Chironomini Chironomus dilutus 0 0 0 0 0 0 0 0 0 0 1 0 0 0 CH_F Diptera Chironomini Chironomus fulvus 0 1 0 0 0 0 0 0 0 0 0 0 0 0 CH_M Diptera Chironomini Chironomus maturus 0 0 0 0 0 1 0 0 0 0 0 0 0 0 CH_S Diptera Chironomini Chironomus staegeri 1 0 1 0 0 0 0 0 0 0 0 0 0 1 CH_U Diptera Chironomini Chironomus utahensis 0 1 0 0 0 0 0 0 0 0 0 0 1 0 CH_W Diptera Chironomini Chironomus whitseli 0 0 0 0 0 0 0 0 1 0 0 0 0 0 CO_CO5 Diptera Chironomini Chironomus n.sp.CO-5 0 0 0 0 0 0 0 0 0 1 0 0 0 0 CH_CO10 Diptera Chironomini Chironomus n.spCO-10 0 0 1 0 0 0 0 1 0 0 0 0 0 0 CH_ns Diptera Chironomini Chironomus n.sp. 1 0 0 0 0 0 0 1 1 0 0 0 0 0 CL_E Diptera Chironomini Cladopelma edwardsi 0 0 0 0 0 0 1 0 0 0 0 0 0 1 CL_V Diptera Chironomini Cladopelma viridula 0 1 1 0 0 0 0 0 0 1 0 0 0 0 CY_D Diptera Chironomini Cryptochironomus digitatus 0 1 1 0 0 0 0 0 0 0 0 0 0 0 CY_F Diptera Chironomini Cryptochironomus fulvus 0 1 1 0 0 0 0 0 0 0 0 0 0 1 CY_P Diptera Chironomini Cryptochironomus parafulvus 1 0 0 0 0 0 0 0 0 0 0 0 0 0 CY_n9 Diptera Chironomini Cryptochironomus n.sp.9 0 0 0 0 0 0 0 0 0 0 0 0 0 1 CT_E Diptera Chironomini Cryptotendipes emorsus 0 0 0 0 0 0 0 0 0 0 0 1 0 0 DT_C Diptera Chironomini Dicrotendipes crypticus 0 0 0 0 0 0 0 0 0 0 0 0 0 1 DT_F Diptera Chironomini Dicrotendipes fumidus 0 0 1 0 0 1 0 1 1 1 0 0 0 1 DT_M Diptera Chironomini Dicrotendipes modestus 1 0 0 0 0 0 0 0 0 0 0 0 0 0 DT_N Diptera Chironomini Dicrotendipes nervosus 0 0 0 1 0 0 0 0 0 0 0 0 0 0 EN_N Diptera Chironomini Endochironomus nigricans 1 1 1 0 0 0 0 0 1 0 0 0 0 0 G_CO1 Diptera Chironomini Gillotia n.sp. CO-1 0 0 0 0 0 0 0 0 0 0 0 0 0 1 GL_B Diptera Chironomini Glyptotendipes barbipes 0 0 0 0 0 0 0 0 0 0 1 0 0 0 GL_L Diptera Chironomini Glyptotendipes lobiferus 1 0 0 0 0 0 0 0 0 0 1 0 0 0

H_V Diptera Chironomini Harnischia viridulus 0 0 0 0 0 0 0 0 0 0 0 0 0 1 PC_T Diptera Chironomini Parachironomus tenuicaudatus 0 0 0 0 1 0 0 0 0 1 1 0 0 0 PP_D Diptera Chironomini Paracladopelma doris 0 1 0 0 0 0 0 0 0 0 0 0 0 0 PC_ns Diptera Chironomini Paracladopelma n.sp. 0 0 1 1 0 0 0 0 0 0 1 0 0 0 PC_n1 Diptera Chironomini Paracladopelma n.sp. CO-1 0 0 0 0 0 0 0 0 0 0 0 0 0 1 PT_S Diptera Chironomini Paratendipes subaequalis 0 1 0 0 0 0 0 0 0 0 0 0 0 0 PH_P Diptera Chironomini Phaenopsectra profusa 0 1 1 0 0 0 1 1 1 0 1 1 0 0 PP_D Diptera Chironomini Polypedilum digitifer 0 0 1 0 0 0 0 0 0 0 0 0 1 0 PP_E Diptera Chironomini Polypedilum excelsius 0 0 0 0 0 1 0 0 0 0 0 0 0 0 PP_F Diptera Chironomini Polypedilum flavum 0 0 1 0 0 0 0 0 0 0 0 0 0 0 PP_I Diptera Chironomini Polypedilum illinoense 0 0 0 0 0 0 0 0 0 0 0 1 0 1 PP_LT Diptera Chironomini Polypedilum laetum 0 0 1 0 0 1 1 1 1 0 0 0 0 1 PP_O Diptera Chironomini Polypedilum obtusum 0 1 0 0 0 0 0 0 0 0 0 0 0 0 PP_P Diptera Chironomini Polypedilum prolixipartum 0 0 0 0 0 0 0 1 0 0 0 0 0 0 PP_SC Diptera Chironomini Polypedilum scalaenum 1 1 1 1 1 0 0 0 0 1 1 1 1 1 PP_SI Diptera Chironomini Polypedilum simulans 0 0 0 1 0 0 0 0 0 0 0 0 0 0 PP_SU Diptera Chironomini Polypedilum sulaceps 0 1 0 1 1 0 0 0 0 0 0 0 0 0 PP_n14 Diptera Chironomini Polypedilum n.sp. 14 0 0 0 0 0 0 0 0 0 0 0 0 1 0 PS_R Diptera Chironomini Pseudochironomus richardsoni 0 1 0 0 0 0 0 0 0 0 0 0 0 0 RB_C Diptera Chironomini Robackia claviger 0 0 0 0 0 0 0 0 0 0 0 0 1 0 SA_T Diptera Chironomini Saetheria tylus 0 1 1 1 1 0 0 1 1 1 1 1 1 1 SA_n1 Diptera Chironomini Saetheria n.sp. 1 0 0 1 1 1 0 0 0 1 1 1 0 0 0 ST_A Diptera Chironomini Stichtochironomus annulicrus 0 0 0 0 1 0 0 0 0 0 0 0 0 0 ST_V Diptera Chironomini Stichtochironomus varius 1 0 0 0 0 0 0 0 0 0 0 0 0 0 XC_B Diptera Chironomini Xestochironomus brunneus 0 0 0 0 0 0 0 0 0 0 1 1 0 1 DM_H Diptera Diamesinae Diamesa heteropus 0 0 0 0 0 0 0 0 0 0 0 1 0 0 AM_F Diptera Orthocladiinae Apometriocnemus fontanillis 0 0 0 0 0 1 0 0 0 0 0 0 0 0 BR_F Diptera Orthocladiinae Brillia flavifrons 0 1 0 0 1 1 1 1 0 1 0 0 0 0 BR_R Diptera Orthocladiinae Brillia retifinis 0 0 0 0 0 0 0 1 0 0 0 0 0 0 CC_O Diptera Orthocladiinae Cardiocladius obscurus 0 0 0 0 0 0 0 0 0 1 0 0 0 1 16

CT_A Diptera Orthocladiinae Chaetocladius astis 0 0 0 0 0 1 0 0 0 0 0 0 0 0 CO_A Diptera Orthocladiinae Corynoneura arctica 0 0 0 0 0 0 0 0 0 0 0 0 0 1 CO_T Diptera Orthocladiinae Corynoneura taris 0 0 0 0 0 0 0 0 1 0 0 1 0 0 CR_A Diptera Orthocladiinae Cricotopus annulator 0 0 0 0 0 0 0 0 0 0 0 0 0 1 CR_BC Diptera Orthocladiinae Cricotopus bicinctus 1 1 0 1 0 0 0 0 0 1 1 1 1 1 CR_BL Diptera Orthocladiinae Cricotopus blinni 1 1 0 0 1 0 0 0 0 1 1 1 0 1 CR_H Diptera Orthocladiinae Cricotopus herrmanni 1 1 1 1 0 0 0 1 1 1 0 0 0 0 CR_I Diptera Orthocladiinae Cricotopus infuscatus 1 1 1 1 1 1 0 0 1 1 1 1 1 1 CR_S Diptera Orthocladiinae Cricotopus sylvestris 0 1 1 0 0 0 0 0 0 1 0 1 0 0 CR_T Diptera Orthocladiinae Cricotopus trifascia 0 1 1 1 1 0 0 1 1 1 1 1 1 1 CR_TF Diptera Orthocladiinae Cricotopus trifasciatus 0 0 0 0 1 0 0 1 1 1 1 0 0 0 CR_V Diptera Orthocladiinae Cricotopus varipes 1 0 0 0 0 0 0 0 0 0 0 0 0 0 DC_C Diptera Orthocladiinae Diplocladius cultriger 0 1 0 0 0 0 0 0 0 0 0 0 0 0 ED_D Diptera Orthocladiinae Eudactylocladius dubitatus 0 0 0 1 0 0 0 0 0 0 0 0 0 0 ED_F Diptera Orthocladiinae Eudactylocladius fuscimanus 1 0 0 0 0 0 0 0 0 0 0 0 0 0 EK_CL Diptera Orthocladiinae Eukiefferiella claripennis 1 0 1 1 0 1 1 1 1 1 0 1 0 0 EK_CO Diptera Orthocladiinae Eukiefferiella coerulescens 1 1 1 1 1 0 0 0 1 1 1 1 0 1 EK_G Diptera Orthocladiinae Eukiefferiella gracei 1 0 0 0 0 0 0 0 0 0 0 0 0 0 EK_4 Diptera Orthocladiinae Eukiefferiella n.sp. 4 0 0 0 0 0 1 0 0 0 0 0 0 0 0 EK_9 Diptera Orthocladiinae Eukiefferiella n.sp. 9 1 0 1 0 0 0 0 0 0 0 0 0 0 0 EU_R Diptera Orthocladiinae Euorthocladius rivicola 1 1 0 0 1 0 0 0 0 0 0 1 0 0 HA_CO1 Diptera Orthocladiinae Halocladius n.sp. CO-1 0 0 0 0 0 0 0 0 0 1 0 0 0 0 HE_H Diptera Orthocladiinae Heleniella hirta 1 0 0 0 0 0 0 0 0 0 0 0 0 0 LI_A Diptera Orthocladiinae Limnophyes angelicae 1 0 0 0 0 0 0 0 0 0 0 0 0 0 LI_AS Diptera Orthocladiinae Limnophyes asquamatus 0 0 0 0 0 0 0 0 1 0 1 1 0 0 LI_F Diptera Orthocladiinae Limnophyes fumosus 0 0 0 1 1 0 0 0 0 0 0 0 0 0 LI_H Diptera Orthocladiinae Limnophyes hastulatus 0 0 1 0 0 0 0 0 1 0 0 0 0 0 LI_HU Diptera Orthocladiinae Limnophyes hudsoni 0 0 0 0 0 0 0 1 0 0 0 0 0 0 LI_M Diptera Orthocladiinae Limnophyes margaretae 0 1 1 1 1 0 0 1 0 0 1 0 1 0 LI_MI Diptera Orthocladiinae Limnophyes minimus 0 0 0 0 0 0 0 0 1 1 0 0 0 0 17

LI_N Diptera Orthocladiinae Limnophyes natalensis 1 0 1 0 0 1 1 1 0 0 1 0 0 0 LI_R Diptera Orthocladiinae Limnophyes recisus 0 0 0 1 0 1 1 1 0 0 0 0 0 0 ME_ns Diptera Orthocladiinae Metriocnemus n.sp. 0 0 0 0 0 1 0 0 0 0 0 0 0 0 NA_I Diptera Orthocladiinae Nanocladius incomptus 1 0 0 0 0 0 0 0 0 0 0 0 0 0 NA_S Diptera Orthocladiinae Nanocladius spliniplenus 0 0 1 0 0 0 0 0 0 0 0 0 0 0 OR_A Diptera Orthocladiinae Orthocladius appersoni 1 0 0 0 0 0 0 0 0 0 0 0 0 0 OR_D Diptera Orthocladiinae Orthocladius dorenus 0 0 0 0 0 0 0 0 0 1 0 0 0 0 OR_F Diptera Orthocladiinae Orthocladius frigidus 1 1 0 0 0 0 0 0 0 0 0 0 0 0 OR_O Diptera Orthocladiinae Orthocladius obumbratus 1 1 0 1 0 0 0 0 0 0 0 0 0 0 PC_ns Diptera Orthocladiinae Parachaetocladius n.sp. 0 0 1 0 0 0 0 0 0 0 0 0 0 0 PA_L Diptera Orthocladiinae Parocladius alpicola 0 0 0 0 0 0 0 0 1 0 0 0 0 0 PK_S Diptera Orthocladiinae Parakiefferiella subterrima 0 0 0 0 0 0 0 0 1 0 0 1 1 0 PM_L Diptera Orthocladiinae Parametriocnemus lundbecki 0 0 0 0 1 0 0 0 0 0 0 1 0 0 PM_ns Diptera Orthocladiinae Parametriocnemus n.sp. 0 0 0 0 0 0 0 1 0 0 0 0 0 0 PP_E Diptera Orthocladiinae Paraphaenocladius exagitans 0 0 0 1 0 0 0 1 0 0 0 0 0 0 PP_IM Diptera Orthocladiinae Paraphaenocladius impensus 0 0 0 0 0 0 0 0 0 1 1 0 0 0 PP_IN Diptera Orthocladiinae Paraphaenocladius innasus 0 0 1 0 0 0 0 0 0 1 0 0 0 0 PT_R Diptera Orthocladiinae Paratrichocladius rufiventris 0 0 0 1 0 0 0 0 1 0 0 0 0 0 PH_P Diptera Orthocladiinae Phaenopsectra profusa 1 0 0 0 0 0 0 0 0 0 0 0 0 0 PS_n8 Diptera Orthocladiinae Psectrocladius n.sp. 8 1 0 0 1 0 0 0 0 0 0 0 0 0 0 PD_F Diptera Orthocladiinae Pseudosmittia forcipatus 0 0 0 1 0 0 0 0 0 0 0 0 1 0 RH_1 Diptera Orthocladiinae Rheocricotopus n.sp. 1 0 0 1 0 0 1 0 1 0 0 0 0 0 0 SA_ns Diptera Orthocladiinae Saetheriella n.sp. 1 0 1 0 0 0 0 0 0 0 0 0 0 0 0 SM_A Diptera Orthocladiinae Smittia atterrima 0 1 0 0 0 0 0 0 1 0 0 1 0 0 SM_P Diptera Orthocladiinae Smittia polaris 1 0 0 0 0 0 0 0 0 0 0 0 0 0 TH_BA Diptera Orthocladiinae Thienemanniella barberi 0 0 0 0 0 0 0 0 0 0 0 1 0 0 TH_BO Diptera Orthocladiinae Thienemanniella boltoni 0 0 0 0 0 0 0 0 0 0 0 0 0 1 TH_E Diptera Orthocladiinae Thienemanniella elana 0 0 0 1 0 0 0 0 0 0 0 1 0 0 TH_X Diptera Orthocladiinae Thienemanniella xena 1 1 0 0 1 0 0 1 0 0 0 1 0 0 TH_7 Diptera Orthocladiinae Thienemanniella n.sp. 7 0 0 1 0 0 0 0 0 0 0 0 0 0 0 18

TH_8 Diptera Orthocladiinae Thienemanniella n.sp. 8 0 0 0 0 0 0 1 0 0 0 0 0 0 0 TV_P Diptera Orthocladiinae Tvetenia paucunca 0 0 0 0 1 1 1 1 1 0 0 1 0 0 TV_V Diptera Orthocladiinae Tvetenia vitracies 0 0 0 0 0 1 0 0 0 0 0 0 0 0 PA_K Diptera Podonominae Parochlus kiefferi 0 0 0 0 0 1 0 0 0 0 0 0 0 0 OD_F Diptera Prodiamesinae Odontomesa ferringtoni 0 0 0 0 0 0 0 0 0 0 0 1 0 0 PR_O Diptera Prodiamesinae Prodiamesa olivacea 0 0 0 0 0 0 1 1 0 0 0 0 0 0 AB_I Diptera Tanypodinae Ablabesmyia illinoensis 0 0 0 0 1 0 0 0 0 0 0 0 0 0 AB_M Diptera Tanypodinae Ablabesmyia mallochi 0 0 1 0 0 0 0 0 0 0 0 1 0 1 AB_MO Diptera Tanypodinae Ablabesmyia monilis 0 0 0 0 0 0 0 0 0 0 1 0 0 0 AB_P Diptera Tanypodinae Ablabesmyia pulchripennis 0 0 0 1 0 0 0 0 0 0 0 0 0 0 AP_F Diptera Tanypodinae Apsectratanypus florens 0 0 0 0 0 0 1 0 0 0 0 0 0 0 CO_C Diptera Tanypodinae Coelotanypus concinnus 0 0 0 0 0 0 0 0 0 0 0 0 0 1 CO_P Diptera Tanypodinae Conchapelopia pallens 0 1 0 1 1 0 0 0 0 1 1 0 0 0 CP_P Diptera Tanypodinae Conchapelopia telema 0 0 0 1 0 0 0 0 0 0 0 0 0 0 CP_ns Diptera Tanypodinae Conchapelopia n.sp. 0 0 0 1 0 0 0 0 0 0 0 0 0 0 LA_L Diptera Tanypodinae Larsia lyra 1 0 0 0 0 0 0 0 0 0 0 0 0 0 PR_B Diptera Tanypodinae Procladius bellus 1 0 1 0 0 0 0 1 0 1 0 0 1 1 PR_C Diptera Tanypodinae Procladius culciformis 0 0 0 0 0 0 0 0 1 0 0 0 0 0 PR_F Diptera Tanypodinae Procladius freemani 0 0 1 1 1 0 0 1 0 1 0 0 0 0 PR_S Diptera Tanypodinae Procladius sublettei 0 0 1 0 0 0 0 1 0 0 0 0 1 0 TA_N Diptera Tanypodinae Tanypus neopunctipennis 0 0 0 0 0 0 0 0 0 1 0 0 0 0 TA_P Diptera Tanypodinae Tanypus punctipennis 0 0 0 0 0 0 0 0 0 0 0 0 0 1 TA_S Diptera Tanypodinae Tanypus stellatus 0 0 1 0 0 0 0 0 0 0 0 0 0 0 TE_O Diptera Tanypodinae Telopelopia okoboji 0 0 0 0 0 0 0 0 0 0 0 0 1 1 CL_C Diptera Tanytarsini Cladotanytarsus crusculus 0 0 0 0 0 0 0 0 0 0 1 0 0 1 CL_F Diptera Tanytarsini Cladotanytarsus fusiformis 1 0 0 0 0 0 0 0 0 0 0 0 0 0 CL_V Diptera Tanytarsini Cladotanytarsus viridiventris 1 0 0 0 0 0 0 0 0 0 0 0 0 0 CL_7 Diptera Tanytarsini Cladotanytarsus n.sp. 7 1 0 0 0 0 0 0 0 0 0 0 0 0 0 MI_L Diptera Tanytarsini Micropsectra logani 0 0 0 0 1 0 0 0 0 0 0 1 0 0 MI_N Diptera Tanytarsini Micropsectra nigripila 0 1 1 0 1 0 0 1 0 0 0 1 1 0 19

MI_P Diptera Tanytarsini Micropsectra polita 1 1 1 0 0 1 1 1 0 0 0 0 0 0 MI_R Diptera Tanytarsini Micropsectra recurvata 1 0 0 0 0 0 0 0 0 0 0 0 0 0 PT_3 Diptera Tanytarsini Paratanytarsus n.sp. OH-3 1 0 0 0 0 0 0 0 0 0 0 0 0 0 RT_3 Diptera Tanytarsini Rheotanytarsus n.sp. 3 1 1 0 0 0 0 0 0 0 0 0 0 0 0 TT_A Diptera Tanytarsini Tanytarsus acifer 0 0 1 0 0 0 0 0 0 0 0 0 0 0 TT_H Diptera Tanytarsini Tanytarsus hastatus 0 0 0 0 0 0 0 0 0 0 1 0 0 0 TT_PA Diptera Tanytarsini Tanytarsus pallidicornis 0 0 0 0 0 0 0 0 0 0 0 0 1 0 TT_PE Diptera Tanytarsini Tanytarsus pelsuei 0 0 0 1 0 0 0 0 0 0 0 0 0 0

Table 3. List of New Chironomid Species from 2007/2008 sweep net and UV light Collections in the Fountain Creek Watershed. CODE SUBFAMILY/TRIBE GENUS LOCATION CH_CO5 Chironomini Chironomus n. sp. CO-5 LF-1 CH_CO10 Chironomini Chironomus n. sp. CO-10 UF-3 [nr.blaylock] CH_ns Chironomini Chironomus n. sp. UF-3,4, MC-1 [nr.quin] CY_n9 Chironomini Cryptochironomus n. sp. LF-5 9 G_CO1 Chironomini Gillotia n. sp. CO-1 LF-5 PC_ns Chironomini Paracladopelma n. sp. MC-3,4, LF-2 [nr.doris] PC_n1 Chironomini Paracladopelma n. sp. LF-5 CO-1 PP_n14 Chironomini Polypedilum n. sp. 14 LF-4 [=herrmanni.ms] SA_n1 Chironomini Saetheria n. sp. 1 MC-2,3,4,5, UF-4, [marki.ms.name] LF-1,2 EK_n9 Orthocladiinae Eukiefferiella n. sp. 9 MC-1,3 [cuesta.ms.name] EK_4 Orthocladiinae Eukiefferiella n. sp. 4 NM UF-1 & CO HA_CO1 Orthocladiinae Halocladius n. sp. CO-1 LF-1 ME_ns Orthocladiinae Metriocnemus n. sp. UF-1 PC_ns Orthocladiinae Parachaetocladius n. sp. MC3 PM_ns Orthocladiinae Parametriocnemus n. sp. UF-3 PS_n8 Orthocladiinae Psectrocladius n. sp. 8 MC-1,4 RH_1 Orthocladiinae Rheocricotopus n. sp. 1 MC-3, UF-3 CFW-3 [hatchi.ms.name] SA_ns Orthocladiinae Saetheriella n. sp. MC2 (marki.ms) TH_8 Orthocladiinae Thienemanniella n. sp. 8 UF-2 [=herrmanni] TH_7 Orthocladiinae Thienemanniella n. sp.7 MC-3 [luna.ms] CP_ns Tanypodinae Conchapelopia n. sp. MC-4 CL_7 Tanytarsini Cladotanytarsus n. sp. 7 MC-1,2 PT_ns Tanytarsini Paratanytarsus n. sp. MC-1 RT_3 Tanytarsini Rheotanytarsus n. sp. 3 MC-1,2 [fontanalis.ms]

Tanypodinae Tanytarsini 12% 8% Prodiamesinae Chironomini 3% Diamesinae Podonominae Chironomini Orthocladiinae 2% 29% Podonominae

Orthocladiinae Prodiamesinae Diamesinae 45% Tanypodinae 1% Tanytarsini

Figure 9. Percent composition of chironomids by subfamily/tribe from all 14 sites for 2007 and 2008.

These subfamilies/tribes were separated by year (2007 and 2008) as shown in Figures

10-37. Sampling two consecutive years ensured the collection continuity of as many subfamily/species representations as possible for a particular site. As illustrated, the subfamily/tribe composition fluctuated each year with some subfamilies/tribes being present one year but not the other. Representatives from Chironomini and Orthocladiinae were present in collections for both years at all sites.

From collections at sites MC-1 and MC-2 (Figures 10-13), the subfamily

Tanypodinae was present only in 2008.

Chironomini Tanytarsini Chironomini 17% 31% Orthocladiinae Tanypodinae Tanypodinae Tanytarsini Orthocladiinae 0% 52%

Figure 10. Percent subfamily/tribe composition of chironomids from site MC-1 2007.

Tanytarsini Chironomini 12% 29% Tanypodinae Chironomini 12% Orthocladiinae Tanypodinae

Orthocladiinae Tanytarsini 47% Figure 11. Percent subfamily/tribe composition of chironomids for site MC-1 2008.

Tanytarsini Chironomini 12% Chironomini 32% Orthocladiinae Tanypodinae Orthocladiinae Tanypodinae Tanytarsini 56% 0%

Figure 12. Percent subfamily/tribe composition of chironomids for site MC-2 2007.

Tanytarsini 8% Chironomini Tanypodinae 8% Orthocladiinae Orthocladiinae Chironomini Tanypodinae 25% 59% Tanytarsini

Figure 13. Percent subfamily/tribe composition of chironomids for site MC-2 2008.

At sites MC-3, 4 and 5 (Figures 14-19) the tribe Tanytarsini was present only in the

2007 collections.

Tanytarsini 11% Chironomini Chironomini Tanypodinae 32% 14% Orthocladiinae Tanypodinae Tanytarsini Orthocladiinae 43%

Figure 14. Percent subfamily/tribe compositon of chironomids for site MC-3 2007.

Tanypodinae 17% Chironomini Chironomini Orthocladiinae 50% Tanypodinae Orthocladiinae 33% Tanytarsini Tanytarsini 0%

Figure 15. Percent subfamily/tribe composition of chironomids for site MC-3-2008.

Tanytarsini 6% Tanypodinae Chironomini 22% Chironomini 39% Orthocladiinae Tanypodinae Tanytarsini Orthocladiinae 33%

Figure 16. Percent subfamily/tribe composition of chironomids for site MC-4 2007.

Tanypodinae 5% Chironomini Chironomini 32% Orthocladiinae Tanypodinae Orthocladiinae Tanytarsini 63% 0% Tanytarsini

Figure 17. Percent subfamily/tribe composition of chironomids for site MC-4 2008.

Tanytarsini Chironomini 6% 29% Tanypodinae Chironomini 12% Orthocladiinae Tanypodinae Tanytarsini Orthocladiinae 53%

Figure 18. Percent subfamily/tribe composition of chironomids for site MC-5 2007.

Tanypodinae 13% Chironomini Chironomini 27% Orthocladiinae Tanypodinae Tanytarsini Tanytarsini Orthocladiinae 0% 60%

Figure 19. Percent subfamily/tribe composition of chironomids for site MC-5 2008.

At site UF-1 (Figures 20 and 21), representatives from the subfamily Podonominae were collected only in 2007 while representatives from tribe Tanytarsini were collected only in 2008.

Podonominae 8% Chironomini 17% Chironomini Orthocladiinae Podonominae Tanytarsini 0% Tanytarsini Orthocladiinae 75%

Figure 20. Percent subfamily/tribe composition of chironomids for site UF-1 2007.

Tanytarsini 11% Chironomini Chironomini 33% Orthocladiinae Orthocladiinae Podonominae 56% Podonominae Tanytarsini 0%

Figure 21. Percent subfamily/tribe composition of chironomids for site UF-1 2008.

During 2007, collections from site UF-2 (Figure 22) produced only representatives from Chironomini and Orthocladiinae while 2008 samples (Figure 23) were much richer producing representatives from Chironomini, Orthocladiinae, Prodiamesinae, Tanypodinae, and Tanytarsini.

Chironomini 25% Chironomini

Orthocladiinae Tanypodinae Orthocladiinae 75% 0% Prodiamesinae Prodiamesinae 0% Tanypodinae Tanytarsini Tanytarsini 0%

Figure 22. Percent subfamily/tribe composition of chironomids for site UF-2 2007.

Tanytarsini 11% Chironomini Tanypodinae Chironomini 11% 34% Orthocladiinae Prodiamesinae Prodiamesinae 11% Tanypodinae Orthocladiinae Tanytarsini 33%

Figure 23. Percent subfamily/tribe composition of chironomids for site UF-2 2008.

Site UF-3 2008 collections (Figure 25) showed a richer subfamily/tribe representation than 2007 (Figure 24), which includes Prodiamesinae, Tanypodinae, and Tanytarsini.

Tanypodinae 8% Chironomini Chironomini 23% Orthocladiinae Prodiamesinae Prodiamesinae 0% Tanypodinae Tanytarsini Tanytarsini Orthocladiinae 0% 69%

Figure 24. Percent subfamily/tribe composition of chironomids for site UF-3 2007.

Tanytarsini 11% Chironomini Tanypodinae 33% Chironomini 11% Orthocladiinae Prodiamesinae Prodiamesinae 6% Orthocladiinae Tanypodinae 39% Tanytarsini

Figure 25. Percent subfamily/tribe composition of chironomids for site UF-3 2008.

The subfamily Tanypodinae was present in 2008 only for site UF-4 (Figures 26 and

27).

Chironomini 33% Chironomini Orthocladiinae 67% Orthocladiinae Tanypodinae Tanypodinae 0%

Figure 26. Percent subfamily/tribe composition of chironomids for site UF-4 2007.

Tanypodinae 6% Chironomini 41% Chironomini Orthocladiinae Orthocladiinae 53% Tanypodinae

Figure 27. Percent subfamily/tribe composition of chironomids for site UF-4 2008.

Collections from sites LF-1 (Figures 28 and 29) had only representatives from

Chironomini, Orthocladiinae, and Tanypodinae for both 2007 and 2008.

Tanypodinae 17%

Chironomini Chironomini 39% Orthocladiinae Orthocladiinae 44% Tanypodinae

Figure 28. Percent subfamily/tribe composition of chironomids for site LF-1 2007.

Tanypodinae 11% Chironomini 28% Chironomini Orthocladiinae Orthocladiinae Tanypodinae 61%

Figure 29. Percent subfamily/tribe composition of chironomids for site LF-1 2008.

Tanytarsini were collected from site LF-2 only in 2007 (Figure 30) while

Tanypodinae were collected only in 2008 (Figure 31).

Tanytarsini 11% Chironomini Chironomini 61% Orthocladiinae Orthocladiinae 28% Tanypodinae Tanytarsini Tanypodinae 0%

Figure 30. Percent subfamily/tribe composition of chironomids for site LF-2 2007.

Tanypodinae 17% Chironomini Chironomini 33% Orthocladiinae Tanypodinae Tanytarsini Tanytarsini Orthocladiinae 0% 50% Figure 31. Percent subfamily/tribe compostion of chironomids for site LF-2 2008.

Site LF-3 2007 (Figure 32) depicts the richest collection with representatives from 6 subfamilies/tribes and the only site in which to find the subfamily Diamesinae. Neither subfamily Prodiamesinae nor Diamesinae were collected again in 2008 (Figure 33).

Tanytarsini 9% Chironomini Tanypodinae Chironomini 5% 23% Diamesinae Orthocladiinae Prodiamesinae Diamesinae 4% 4% Prodiamesinae Tanypodinae Orthocladiinae Tanytarsini 55%

Figure 32. Percent subfamily/tribe composition of chironomids for site LF-3 2007.

Tanytarsini 7% Chironomini Chironomini 27% Tanypodinae Diamesinae 6% Orthocladiinae Prodiamesinae 0% Prodiamesinae Tanypodinae Diamesinae Orthocladiinae 0% Tanytarsini 60%

Figure 33. Percent subfamily/tribe composition of chironomids for site LF-3 2008.

The subfamily Tanypodinae and tribe Tanytarsini were found only at Site LF-4 for

2008 (Figure 35) and were not collected in 2007 (Figure 34).

Chironomini 43% Chironomini Orthocladiinae Orthocladiinae Tanytarsini Tanypodinae 57% 0% Tanytarsini Tanypodinae 0%

Figure 34. Percent subfamily/tribe composition of chironomids for site LF-4 2007.

Tanytarsini 13% Chironomini Chironomini 40% Tanypodinae Orthocladiinae 20% Tanypodinae Tanytarsini Orthocladiinae 27%

Figure 35. Percent subfamily/tribe composition of chironomids for site LF-4 2008.

Tanypodinae were collected from site LF-5 2008 (Figure 37) but not in 2007 (Figure

36). The tribe Chironomini consisted of largest percentage of chironomids from this location at 67%.

Chironomini Orthocladiinae 67% Chironomini 33% Orthocladiinae Tanypodinae Tanypodinae 0%

Figure 36. Percent subfamily/tribe composition of chironomids for site LF-5 2007.

Tanypodinae 19% Chironomini 52% Chironomini Orthocladiinae Orthocladiinae Tanypodinae 29%

Figure 37. Percent subfamily/tribe composition of chironomids for site LF-5 2008.

Figures 38 and 39 represent the number of species collected, slide prepared and identified at each site in 2007 and 2008 for both sweep net and ultraviolet light collection methods. These numbers include species duplications for some sites as certain species were collected in both methods. In 2007 (Figure 38), 124 species were collected using the sweep net method and

139 species were collected using the ultraviolet night light technique (UV), each method producing similar collection numbers. In 2008 (Figure 39), 86 species were collected using the sweep net and twice as many, 177 species, were collected using ultraviolet night lighting

(UV).

10 # Species 2007 Sweep Net # Species 2007 UV Light

Number Number Species of 5

Sites Figure 38. Number of species collected by both sweep and ultraviolet (UV) collection methods for 2007.

10 # Species 2008 Sweep Net

5 # Species 2008 UV Light Number Number Species of

Sites

Figure 39. Numbers of species collected by both sweep net and ultraviolet (UV) collection methods for 2008.

Using two different collection methods resulted in 21entirely different species of chironomids collected at site LF-3 in 2007 (Table 4). If only the net or UV method had been used exclusively, many species would not have been collected and recorded. No duplications occurred for either method at these sites even though collections were made in the same place, time and date. Site MC-5 in 2007 (Table 5) also had no duplicate species for either method resulting in 17 total species collected, as well as site LF-4 in 2007 with 7 total species collected (Table 6).

Table 4. Variation of chironomid species reported for site LF-3 on 9-3-2007 for two collection methods. LF-3 Collected 9-3-2007 Ultraviolet Night Lighting Collection Sweep Net Collection Ablabesmyia mallochi Diamesa heteropus Cricotopus bicinctus Odontomesa ferringtoni Cricotopus blinni Eukiefferiella coerulescens Cricotopus infuscatus Euorthocladius rivicola Corynoneura taris Tvetnia paucunca Cryptotendipes emerosus Micropsectra logani Eukiefferiella claripennis Micropsectra nigripila Parakiefferiella subaterrima Parametriocnemus lundbecki Phaenopsectra profusa Saetheria tylus Smittia aterrima Thienemanniella xena Xestochironomus brunneus

Table 5. Variation of chironomid species reported for site MC-5 on 9-6-2007 for two collection methods. MC-5 Collected 9-6-2007 Ultraviolet Night Lighting Collection Sweep Net Collection Chironomus decorus Saetheria tylus Polypedilum scalaenum Corynoneura arctica Saetheria n. sp. 1 Cricotopus blinni Stichtochironomus annulicrus Eukiefferiella coerulescens Cricotopus infuscatus Euorthocladius rivicola Cricotopus trifasciatus Limnophyes margaretae Ablabesmyia illinoensis Thienemanniella xena Conchapelopia pallens Micropsectra logani Micropsectra nigripila

Table 6. Variation of chironomid species reported for site LF-4 on 9-1-2007 for two collection methods. LF-4 Collected 9-1-2007 Ultraviolet Night Lighting Sweep Net Collection Chironomus decorus Polypedilum n. sp. 14 Collection Cricotopus infuscatus Saetheria tylus Limnophyes margaretae Pseudosmittia forcipata Parakiefferiella subaterrima

New species were found at all sites except LF-3 (Figure 40).

LF-4 3% MC-1 LF-5 LF-2 8% MC-1 MC-2 5% 16% MC-3 MC-4 MC-5 LF-1 8% MC-2 UF-1 11% UF-2 UF-4 UF-3 5% UF-4 LF-1 UF-3 MC-3 LF-2 8% 16% UF-2 LF-3 3% LF-4 MC-4 LF-3 UF-1 8% 0% LF-5 6% MC-5 3%

Figure 40. Percentage of new species collected per site.

Most (54%) of the new species were collected using the ultraviolet (UV) night light method.

Sweep 17%

Both 29% Sweep UV Both UV 54%

Figure 41. Percentage of new species collected by either UV or sweep net or both.

The most abundant genera collected included 11 Polypedilum species and 11 Chironomus species. For these genera in particular, Polypedilum scalaenum was found at all sites except in the upper Fountain (UF1-4). Chironomus decorus was collected at all sites except UF-2 for both 2007 and 2008 collections. The next most abundant genera included 9 Cricotopus

34 species and 9 Limnophyes species. Cricotopus species were found at all sites except UF-2 and Limnophyes species were found at all sites except LF-5.

Physical Characteristics

The data for water quality parameters of dissolved oxygen, temperature, pH, conductivity, hardness, and selenium are shown in Table 7. The data show temperature, pH, and conductivity increased downstream. As temperature increased, the concentrations of DO decreased. Hardness also increased downstream, an indication of the presence of more dissolved calcium and magnesium solutes. Dissolved, total and pore water fractions sampled at sites LF-4 and LF-5 for both spring and fall all contained selenium in greater concentrations than Colorado’s 4.6 ppb chronic level criterion for selenium. The LF-4 spring

2007 water samples had mean concentrations measuring 9.69 ppb for dissolved selenium,

9.46 ppb for total selenium, and 16.14 ppb for pore selenium; LF-4 fall 2007 concentrations were even greater at 18.59 ppb dissolved selenium, 16.04 ppb total selenium, and 20.27 ppb pore selenium. LF-5 spring 2007 concentrations were 7.91 ppb dissolved selenium, 7.72 ppb total selenium, and 11.66 ppb pore selenium; and fall 2007 concentrations were also greater at site LF-5 with 14.05 ppb, 12.25 ppb, and 8.62 ppb, respectively. These values for sites LF-

4 and LF-5 are greater than all other sites and higher than Colorado State recommended levels. At site MC-5, selenium spikes occurred for all spring and fall samples of dissolved, total, and pore fractions; mean spring selenium concentrations at MC-5 were 1.86, 1.87 and

3.69 ppb and fall concentrations 5.28, 4.68, and 2.44 ppb, respectively. Table 9 exhibits how elevation influenced species diversity by comparing sites UF-1, LF-1, and LF-4, which vary in elevation. Site UF-1 is 536.75 M higher than LF-1 and 901.91 M higher than LF-4 while

LF-1 is 365.16 M higher than LF-4. At sites UF-1, LF-1, and LF-4 19, 27, and 30 species occurred respectively, with 3 species in common. Selenium concentrations increase downstream at each of these three sites and these data are also indicated in Table 8.

Table 7. Mean concentrations for water quality parameters measured during the deployment of bryophytes in Fountain Creek, spring and fall, 2007 Spring averages Fall averages Site DO Temp pH EC Total hardness DSe TSe PSe Site DO Temp pH EC Total hardness DSe TSe PSe MC-1 10.9 4.2 7.4 129.3 39.9 0.21 0.22 0.71 MC-1 9.0 5.5 7.9 283.1 91.3 0.18 0.21 0.38 MC-2 11.0 3.9 7.5 190.0 67.1 0.28 0.29 0.29 MC-2 9.6 5.3 8.0 403.0 129.0 0.33 0.35 0.23 MC-3 11.3 4.1 7.2 194.7 75.1 0.34 0.36 0.37 MC-3 9.6 4.5 8.1 361.9 144.8 0.6 0.56 1.02 MC-4 11.5 4.1 7.5 242.6 89.3 0.44 0.44 0.76 MC-4 8.9 5.0 8.0 405.2 176.9 0.86 0.83 0.23 MC-5 11.7 4.8 7.4 301.5 136.0 1.86 1.87 3.69 MC-5 9.9 6.4 8.1 695.7 276.3 5.28 4.68 2.44 UF-1 11.0 4.6 7.2 197.8 99.6 0.19 0.21 0.05 UF-1 9.1 5.8 8.2 271.9 102.7 0.06 BDL 0.24 UF-2 12.0 4.9 7.2 218.9 101.6 0.14 0.16 0.12 UF-2 9.7 5.7 8.1 184.4 66.3 0.01 0.04 0.26 UF-3 12.2 5.7 7.6 255.2 121.6 0.67 0.66 0.05 UF-3 8.4 6.0 8.1 212.1 76.3 0.04 0.07 0.58 UF-4 12.3 6.6 7.6 340.2 146.6 1.32 1.34 0.19 UF-4 7.5 7.1 8.1 256.2 91.6 0.34 0.36 0.56 LF-1 12.1 7.7 7.5 286.9 147.3 2.05 2.0 6.31 LF-1 7.8 9.9 8.1 515.7 188.7 2.97 2.67 8.42 LF-2 10.8 9.6 7.5 691.6 203.2 2.78 2.85 3.12 LF-2 7.8 15.2 8.1 847.3 265.6 3.77 3.34 3.19 LF-3 10.2 10.6 7.6 854.2 251.4 3.29 3.12 3.26 LF-3 8.2 15.1 8.0 919.7 275.3 3.8 3.3 3.16 LF-4 9.4 11.3 7.7 923.1 296.3 9.69 9.46 16.14 LF-4 8.5 15.2 8.1 1092.3 350.9 18.59 16.04 20.27 LF-5 9.3 11.8 7.8 1024.0 297.8 7.91 7.72 11.66 LF-5 7.5 15.8 8.2 1098.0 357.7 14.05 12.25 8.62 DO dissolved oxygen is expressed as mg/L; Temp temperature as °C; Total hardness as mg/L; EC specific conductivity as μS/cm. Selenium concentrations as ppb.

Table 8. Elevation differences between three sites and occurring species diversity at each site. Selenium concentrations in ppb for 2007 are cited in the following order: dissolved, total, and pore. Species common to all sites are bolded. UF-1 LF-1 LF-4 2,334.77 M 1,798.02 M 1,432.86 M Se ppb [0.19], [0.21], [0.05](spring) Se ppb [2.05], [2.00], [6.31](spring) Se ppb [9.69], [9.46], [16.14](spring) [0.06], [BDL], [0.24](fall) [2.97], [2.67], [8.42](fall) [18.59], [16.04], [20.27](fall) Chironomini/(Total,Chironomus dissolved, poredecorus in ppb) Chironomi(Total,ni/Chironomus dissolved, decoruspore in ppb) Chironomini/Chironomus atrella

Chironomini/Chironomus maturus Chironomini/Chironomus n. sp. CO 5 Chironomini/Chironomus decorus

Chironomini/Dicrotendipes fumidus Chironomini/Cladopelma viridulus Chironomini/Chironomus staegeri

Chironomini/Polypedilum excelsius Chironomini/Dicrotendipes fumidus Chironomini/Cladopelma edswardii

Chironomini/Polydedilum laetum Chironomini/Parachironomus tenuicaudatus Chironomini/ Cryptochironomus n. sp. 9

Orthocladiinae/Apometriocnemus fontanilis Chironomini/Polypedilum scalaenum Chironomini/Cryptochironomus fulvus

Orthocladiinae/Brillia flavifrons Chironomini/Saetheria n. sp. 1 Chironomini/ Dicrotendipes crypticus

Orthocladiinae/Chaetocladius astis Chironomini/Saetheria tylus Chironomini/Dicrotendipes fumidus

Orthocladiinae/Cricotopus infuscatus Orthocladiinae/Brillia flavifrons Chironomini/Gillota n. sp. CO 1

Orthocladiinae/Eukiefferiella n. sp. 4 Orthocladiinae/Cardocladius obscurus Chironomini/Harnischia viridulus

Orthocladiinae/Eukiefferiella claripennis Orthocladiinae/Cricotopus bicinctus Chironomini/Paracladopelma n. sp. CO 1

Orthocladiinae/Limnophyes natalensis Orthocladiinae/blinni Chironomini/Polypedilum illinoense

Orthocladiinae/Limnophyes recisus Orthocladiinae/Cricotopus herrmanni Chironomini/Polypedilum lactum

Orthocladiinae/Metriocnemus n. sp. Orthocladiinae/Cricotopus infuscatus Chironomini/Polypedilum scalaenum

Orthocladiinae/Rheocricotopus n. sp. 1 Orthocladiinae/Cricotopus sylvestris Chironomini/Saetheria tylus

Orthocladiinae/Tvetenia paucunca Orthocladiinae/Cricotopus trifascia Chironomini/Xestochironomus brunneus

Orthocladiinae/Tvetenia vitraces Orthocladiinae/Eukiefferiella claripennis Orthocladiinae/Cardiocladius obscura

Podonominae/Parochlus kiefferi Orthocladiinae/Eukiefferiella coerulescens Orthocladiinae/Cricotopus annulator

Tanytarsini/Micropsectra politus Orthocladiine/Halocladius n. sp. CO 1 Orthocladiinae/Cricotopus bicinctus

Orthocladiinae/Limnophyes minimus Orthocladiinae/Cricotopus blinni

Orthocladiinae/Orthocladius dorensus Orthocladiinae/Cricotopus infuscatus

Orthocladiinae/Paraphaenocladius impensus Orthocladiinae/Cricotopus trifascia

Orthocladiinae/Paraphaenocladius innasus Orthocladiinae/Eukieffierella coerulescens

Tanypodinae/Conchopelopia pallens Orthocladiinae/Thienemanniella boltoni

Tanypodinae/Procladius bellus Prodiamesinae/Coelotanypus concinnus

Tanypodinae/Procladius freemani Prodiamesinae/Tanypus stellatus

Tanypodinae/Tanypus neopunctipennis Tanypodinae/Procladius bellus

Tanypodinae/Tanypus punctipennis

Tanypodinae/Telopelopia okoboji

Tanytarsini/Cladotanytarsus cruscula

Similarity Indices

The Jaccard Coefficient of Community calculation, J= a/(a+b+c)*, is a method used to determine taxa similarity between two sites. Results are shown in Table 9 (and Figures 42

– 44 in Appendix B) for site to site comparisons within each individual reach.

*J= Jaccard’s Coefficient of Community a= the number of taxa common to both samples. b= the number of taxa present in sample b but not in sample a. c= the number of taxa present in sample a but not in sample b.

The general trend for the three Jaccard graphs (Figure 42-44 in Appendix B) is that sites in the closest proximity share a more common pool of species. MC-1/2, 3/4 and 4/5 have greater similarity values than other sites at 0.23, 0.28, and 0.28 respectively. The pair

MC-2/5 has a value 0.03 greater than site MC-2/3 although it is two sites downstream from

MC-2. In the upper Fountain, UF-1/3, 1/2, and 2/3 have high similarity values of 0.45, 0.41, and 0.51, respectively. The similarity values in the lower Fountain share this proximity trend as well, although the Jaccard values were very similar for many site combinations.

The Sorensen Similarity Coefficient, the same as the Index of Similarity of Odum

(1971), S=2a/(2a+b+c)**, was a second method used to determine chironomid species similarity between sites. Results are shown in Table 9 (Figures 45 – 47 in Appendix B) for comparisons within each individual reach.

**S = Sorensen’s Index c = the number of taxa common to both sites. b = the number of taxa in sample at site a. a = the number of taxa in sample at site b.

Site comparisons using the Sørensen’s Index also indicated that those in closest proximity generally exhibit the most species similarity. MC-1/2 has 0.45 similarity, 0.30 for MC-3/4 and 0.44 for MC-4/5. As with the Jaccard Coefficient, the Sørensen Index also showed that

MC-2/5 has a greater similarity than MC-2/3 by 0.04 units. In the upper Fountain all UF sites in closest proximity, UF 1/2, 2/3 and 3/4 have greater similarity values. In the upper Fountain all UF sites in closest proximity, UF-1/2, 2/3 and 3/4 had high similarity values. The lower

Fountain reach had high similarity at site pairs LF-1/2 (0.48) and LF-2/3 (0.40). Site pair LF-

3/4 at 0.34 had a higher similarity value by 0.01 over site pair LF-4/5 at 0.33.

The Jaccard and Sørensen Coefficients were also used to determine species similarities between all 14 sites within the entire watershed as depicted in Table 9, which lists the commonality values for both indices for all sites and (Figures 48 – 53 in Appendix B).

Most of the Jaccard values for MC vs. UF (Figures 48 and 49 in Appendix B) were small whereas the values for the Sørensen’s Index were larger but the trends for each graph were very similar. Sites MC-3/UF-3 for the Jaccard had a 0.33 similarity value and

Sørensen’s Index value was 0.50. For both coefficients, a value of 1 indicated the most commonality between two sites. The Jaccard value for sites MC-3/UF-4 was 0.25 and the

Sørensen value was 0.41. Both similarity indices for the MC vs LF comparisons (Table 9 and

Figures 50 and 51 in Appendix B) were in agreement that sites MC-5/LF-1, MC-5/LF-2,

MC-5/LF-3, LF-1/LF-2, and MC-3/UF-3 had the most similarity; Jaccard values were 0.45,

0.32, 0.32,0.32, and 0.33 respectively, whereas Sørensen values were 0.50, 0.48, 0.49, 0.48, and 0.50. The Jaccard index showed that site UF-2/LF-4 had the least species commonality at

0.00 while the Sørensen index value for the same site pair was 0.00 (Table 9 and Figures 52 and 53 in Appendix B). Values of 0.00 indicated there were no species in common.

Table 9. Matrix of Jaccard’s Similarity Coefficient and Sørensen’s Similarity Coefficient for binary comparisons of paired sites in the Fountain Creek Watershed. Values approaching 1 indicate the most shared species; values closest to 0 indicate the fewest shared species.

Jaccard’s Similarity Coefficient

SITE MC-1 MC-2 MC-3 MC-4 MC-5 UF-1 UF-2 UF-3 UF-4 LF-1 LF-2 LF-3 LF-4 LF-5 MC-1 0.23 0.18 0.15 0.12 0.09 0.06 0.14 0.12 0.15 0.14 0.15 0.09 0.13 MC-2 0.45 0.28 0.22 0.31 0.08 0.07 0.19 0.18 0.26 0.22 0.29 0.20 0.16 MC-3 0.31 0.43 0.18 0.19 0.16 0.11 0.33 0.25 0.29 0.21 0.19 0.21 0.19 MC-4 0.31 0.36 0.30 0.28 0.09 0.05 0.16 0.19 0.26 0.27 0.18 0.20 0.15 MC-5 0.22 0.47 0.32 0.44 0.10 0.06 0.21 0.20 0.45 0.32 0.32 0.20 0.15 UF-1 0.17 0.15 0.28 0.16 0.19 0.29 0.26 0.16 0.12 0.07 0.09 0.06 0.10 UF-2 0.12 0.13 0.20 0.09 0.11 0.45 0.28 0.12 0.05 0.06 0.08 0 0.02 UF-3 0.24 0.31 0.50 0.27 0.35 0.41 0.51 0.26 0.17 0.15 0.16 0.18 0.09 UF-4 0.22 0.30 0.41 0.32 0.42 0.27 0.22 0.41 0.26 0.21 0.29 0.13 0.12 LF-1 0.26 0.41 0.45 0.41 0.50 0.21 0.10 0.36 0.42 0.32 0.21 0.18 0.23 LF-2 0.24 0.36 0.35 0.42 0.48 0.13 0.11 0.25 0.35 0.48 0.25 0.19 0.24 LF-3 0.26 0.44 0.32 0.30 0.49 0.17 0.15 0.32 0.44 0.35 0.40 0.21 0.22 LF-4 0.17 0.34 0.35 0.33 0.33 0.11 0 0.30 0.23 0.30 0.32 0.34 0.20

LF-5 0.23 0.27 0.31 0.26 0.25 0.19 0.05 0.17 0.21 0.37 0.39 0.36 0.33

Sørensen’s Similarity Coefficient

Sediment Characterization

Medium gravel, fine gravel, very fine gravel, and coarse to fine sand were the dominant components of the sediment samples collected at all 14 sites as shown in Table 11. Small cobble was not collected at any site. Very coarse gravel was collected at 5 sites only--MC-3,

UF-2, 4 and LF-1, 3. Coarse gravel was collected at all sites except UF-4 and LF-5. Silt/Clay were collected at all sites except LF-4.

Table 10. Mean percentage of fraction sizes for sediment at each site.

Coarse Site/ Very Very Small Coarse Medium Fine to Fraction Coarse Fine Silt/Clay Cobble Gravel Gravel Gravel Fine Size Gravel Gravel Sand MC1 0 0 0.14 1.46 12.55 28.67 56.39 0.49

MC2 0 0 2.87 10.62 17.14 23.39 45.72 0.25

MC3 0 2.94 16.49 16.78 14.40 20.58 28.74 0.07

MC4 0 0 2.17 4.40 13.31 29.39 50.63 0.09

MC5 0 0 0.36 5.03 14.57 28.44 51.53 0.06

UF1 0 0 1.12 7.01 12.40 16.85 62.39 0.23

UF2 0 14.30 7.42 8.51 12.67 23.63 33.22 0.25

UF3 0 0 7.27 13.29 16.31 15.34 47.58 0.21

UF4 0 10.24 0 0.99 4.00 9.75 74.13 0.89

LF1 0 2.25 3.43 10.96 21.18 27.42 34.68 0.08

LF2 0 0 3.40 5.19 10.51 24.91 55.45 0.53

LF3 0 5.14 6.68 10.53 14.83 18.30 43.79 0.74

LF4 0 0 1.31 6.99 18.43 29.76 43.50 0

LF5 0 0 0 2.43 9.29 24.11 64.14 0.04

Canonical Correspondence Analysis

Canonical Correspondence Analysis (CCA) determines which environmental variables may contribute most to community variations and distributions (Figures 54- 65). CCA was used to generate ordination diagrams using the environmental variables of mean total, dissolved, and pore selenium concentrations from the spring and fall 2007 water samples. Silt/clay was the second variable chosen. All substrate variables were not significant, so silt/clay was randomly chosen as one of the environmental variables to use in combination with selenium. pH was the third variable chosen. All values were log transformed to reduce the significance of extreme outliers. All log values are listed in the Appendix B.

Significant p-values were found for all selenium samples except fall pore selenium. The spring total, dissolved and pore selenium canonical analyses p-values were 0.0006, 0.0006, and 0.0034 respectively. The p-values for fall total, dissolved and pore selenium canonical analyses are 0.05, 0.0044, and 0.1266. The p-values for pH in spring samples were 0.7389,

0.7367, and 0.7469. Fall pH samples were significant at 0.01 for all samples, total, dissolved and pore. P-values for silt and clay were 0.4283, 0.4267 and 0.4017 in the spring and 0.5465,

0.5083 and 0.5467 in the fall. Ordination diagrams were also generated to determine which sites were most associated with high selenium concentrations. Chironomid species presence or absence at each site from the 2007 and 2008 collections represent the community aspect of the CCA analysis and are identified on the diagrams as abbreviations that are listed on Table

2 in the results section. Overlapping species on the ordination diagrams were deleted and recorded as suggested in the CANOCO manual (ter Braak, 1996) as large numbers of species make the diagram indecipherable.

Figure 54. Association of chironomids to spring total selenium/silt and clay/pH in reference to sites. Analysis suggests that sites 13 and 14, representing LF-4 and LF-5, are the most significant sites on this diagram for total selenium concentrations. The vector tagged “Log-Tot. Se” is the abbreviation of the log of the total selenium fraction of water samples.

Figure 55. Spring Total Selenium/silt and clay/pH-Species. Chironomid species data: CCA ordination diagram showing species as abbreviated codes and environmental variables (arrows). Chironomid species codes are located in the results section. The environmental variables are total selenium, pH, silt and clay. Variables were reduced to three to eliminate confounding and keep the diagram readable. Species whose distribution may be the most favorable toward selenium lie on the outermost edges. PP_I = Polypedilum illinoense , CL_C = Cladotanytarsus cruscula, TT_PA = Tanytarsus pallidicomis. PP_I was found only at LF-3 and 5, CL_C was found only at LF-2 and 5 and TT_PA was found only at LF-4. LF 4 and 5 are significant sites for high selenium concentrations as shown from the site diagram Figure 54. The vector tagged “Log-Tot. Se” is the abbreviation of the log of the total selenium fraction of water samples.

Figure 56. Association of chironomids to spring dissolved selenium/silt and clay/pH in reference to sites. Analysis suggests that sites 13 and 14, representing LF-4 and LF-5, are the most significant sites on this diagram for total selenium concentrations. The vector tagged “Log- Diss. Se” is the abbreviation for the log of the dissolved selenium fraction of water samples.

Figure 57. Spring Dissolved Selenium/silt and clay/pH-Species. Chironomid species data: CCA ordination diagram showing species as abbreviated codes and environmental variables (arrows). Chironomid species codes are located in the results section. The environmental variables are total selenium, pH, silt and clay. Variables were reduced to three to eliminate confounding and keep the diagram readable. Species whose distribution may be the most favorable toward selenium lie on the outermost edges. PP_I = Polypedilum illinoense, CL_C = Cladotanytarsus cruscula, TT_PA = Tanytarsus pallidicomis. PP_I was found only at LF-3 and 5, CL_C was found only at LF-2 and 5 and TT_PA was found only at LF-4. LF-4 and 5 are significant sites for high selenium concentrations as shown from the site diagram Figure 56. The vector tagged “Log-Diss. Se” is the abbreviation for the log of the dissolved selenium fratction of water samples.

Figure 58. Association of chironomids to spring pore selenium/silt and clay/pH in reference to sites. Analysis suggests that sites 13 and 14, representing LF-4 and LF-5, are the most significant sites on this diagram for total selenium concentrations. The vector tagged “Log-Pore Se” is the abbreviation for the log of the pore selenium fraction of water samples.

Figure 59. Spring Pore Selenium/silt and clay/pH-Species. Chironomid species data: CCA ordination diagram showing species as abbreviated codes and environmental variables (arrows). Chironomid species codes are located in the results section. The environmental variables are total selenium, pH, silt and clay. Variables were reduced to three to eliminate confounding and keep the diagram readable. Species whose distribution may be the most favorable toward selenium lie on the outermost edges. PP_I = Polypedilum illinoense, CL_C = Cladotanytarsus cruscula and TT_PA = Tanytarsus pallidicomis. PP_I was found only at LF-3 and 5, CL_C was found only at LF-2 and 5 and TT_PA was found only at LF-4. LF-4 and 5 are significant sites for high selenium concentrations as shown from the site diagram Figure 58. The vector tagged “Log-Pore_Se” is the abbreviation for the log of the pore selenium fraction of water samples.

Figure 60. Association of chironomids to fall total selenium/silt and clay/pH in reference to sites. Analysis suggests that sites 13 and 14, representing LF-4 and LF-5, are the most significant sites on this diagram for total selenium concentrations. The vector tagged “Log-Tot. Sel” is the abbreviation for the log of the total selenium fraction of water samples.

Figure 61. Fall Total Selenium/silt and clay/pH-Species. Chironomid species data: CCA ordination diagram showing species as abbreviated codes and environmental variables (arrows). Chironomid species codes are located in the results section. The environmental variables are total selenium, pH, silt and clay. Variables were reduced to three to eliminate confounding and keep the diagram readable. Species whose distribution is the most favorable toward selenium lie on the outermost edges. TT_PA = Tanytarsus pallidicomis and TE_O = Telopelopia okoboji. TT_PA was found only at LF-4 and TT_O was found only at LF-4 and 5, significant sites for selenium concentrations as shown from the site diagram Figure 60. The vector tagged “Log-Tot. Se” is the abbreviation for the log of the total selenium fraction of water samples.

Figure 62. Association of chironomids to fall dissolved selenium/silt and clay/pH in reference to sites. Analysis suggests that sites 13 and 14, representing LF-4 and LF-5, are the most significant sites on this diagram for total selenium concentrations. The vector tagged “Log-Diss. Se” is the abbreviation for the log of the dissolved selenium fraction of water samples.

Figure 63. Fall Dissolved Selenium/silt and clay/pH-Species. Chironomid species data: CCA ordination diagram showing species as abbreviated codes and environmental variables (arrows). Chironomid species codes are located in the results section. The environmental variables are total selenium, pH, silt and clay. Variables were reduced to three to eliminate confounding and keep the diagram readable. Species whose distribution is the most favorable toward selenium lie on the outermost edges. TT_PA = Tanytarsus pallidicomis and TE_O = Telopelopia okoboji. TT_PA was found only at LF-4 and TT_O was found only at LF-4 and 5, significant sites for selenium concentrations as shown from the site diagram Figure 62. The vector tagged “Log-Diss. Se” is the abbreviation for the log of the dissolved selenium fraction of water samples.

Figure 64. Association of chironomids to fall pore selenium/silt and clay/pH in reference to sites. Analysis suggests that sites 13 and 14, representing LF-4 and LF-5, are the most significant sites on this diagram for total selenium concentrations. The vector tagged “Log-Pore Se” is the abbreviation for the log of the pore selenium fraction of water samples.

Figure 65. Fall Pore Selenium/silt and clay/pH-Species. Chironomid species data: CCA ordination diagram showing species as abbreviated codes and environmental variables (arrows). Chironomid species codes are located in the results section. The environmental variables are total selenium, pH, silt and clay. Variables were reduced to three to eliminate confounding and keep the diagram readable. Species whose distribution is the most favorable toward selenium lie on the outermost edges. TT_PA = Tanytarsus pallidicomis and TE_O = Telopelopia. okoboji. TT_PA was found only at LF-4 and TT_O was found only at LF-4 and 5, significant sites for selenium concentrations as shown from the site diagram Figure 64. The vector tagged “Log-Pore Se” is the abbreviation for the log of the pore selenium fraction of water samples.

Discussion

In review of the literature, I found very few studies of Chironomidae in an entire watershed where identifications were completed to the species level.

Comparison to Western Rivers and other Aquatic Environments

Sublette et al. (1998) collected and identified 38 chironomid species, 23 genera, and 4 subfamilies in the Grand Canyon area of the Colorado River between Glen Canyon Dam and

Lake Mead, Arizona, of which only two Chironomus were listed as possible new species; this research covered a distance of 472 km and 585 m of elevation change. In comparison, the

Fountain Creek study covered a distance of approximately 124 km; Monument Creek comprises 32 km, the upper Fountain 25 km, and the lower Fountain 67 km with 919.89 m in elevation change. Only eighteen species from the Grand Canyon study were identical to those found in the Fountain Creek. From this Fountain Creek study, I collected and identified 4 times (155) as many species as Sublette’s Grand Canyon study.

From 2004 to 2006, Kleinert (2008) collected 75 chironomid species, including 8 new species all identified to the species level at five sites in the pre-Legacy Reach of the Arkansas

River below Pueblo Reservoir yet upstream from the confluence with Fountain Creek, a length of 16 km with no significant elevation changes. Chironomid species included those from the families Chironominae, Diamesinae, Orthocladiinae, Tanypodinae, and Tanytarsini;

40 of the 75 species collected were identical to the species found in the Fountain Creek; 79 more chironomid species were collected in the Fountain Creek study than from Kleinert’s study (2008).

In 2006 using the same five sites as Kleinert (2008) in the Arkansas River below the

Pueblo Reservoir (post-Legacy), Powell (2008) collected 52 species of chironomids from families Chironominae, Diamesinae, Orthocladiinae, Tanypodinae, and Tanytarsini including

4 new species; 44 of the 52 species collected were identical to the species found in the

Fountain Creek. Powell (2008) also used a sixth site for her research, located at the Hobson

Ranch above Pueblo Reservoir, a pristine area of river not impacted by either impoundment or urbanization. From this site, 40 species of chironomids from 6 different families were identified. Twenty-four of the 40 species collected were also found in the Fountain Creek watershed. In entirety, 102 more chironomid species were collected in the Fountain Creek research than in the Powell study (2008).

Chironomid species collected from Kleinert’s (2008) 2004 – 2006 chironomid collections and Powell’s (2008) 2006 collections that are identical to the species that were collected in this Fountain Creek Watershed research are listed in Table 11. All species that are highlighted in bold are species that were found in either Kleinert’s (2008) collection or

Powell’s (2008) collection but not both. Species collected from the Fountain Creek sites that are the same species as those from the Kleinert (2008) and Powell (2008) Arkansas River studies are not found only in the lower Fountain segment closest to the Arkansas confluence but they are spread throughout the Fountain Creek watershed. These included 32 similar species in the Monument Creek segment, 34 in the lower Fountain segment, and 19 in the upper Fountain segment.

Table 11. Chironomid species from the Kleinert (2008) and Powell (2008) collections that are identical to those found in the Fountain Creek Watershed 2007 and 2008 collections. Bold font indicates species that are different between Kleinert and Powell.

Kleinert 2004 - 2006 Powell 2006 Family Genus Species Family Genus Species Chiromomini Chironomus decorus Chiromomini Chironomus decorus Chironomini Chironomus n. sp. 5 Chironomini Chironomus maturus Chironomini Chironomus utahensis Chironomini Chironomus utahensis Chironomini Cladopelma edswardi Chironomini Cladopelma edswardi Chironomini Cladopelma viridulus Chironomini Cladopelma viridulus Chironomini Cryptochironomus fulvus Chironomini Cryptochironomus fulvus Chironomini Dicrotendipes fumidus Chironomini Dicrotendipes fumidus Chironomini Glyptotendipes barbipes Chironomini Glyptotendipes barbipes Chironomini Glyptochironomus lobiferus Chironomini Glyptochironomus lobiferus Chironomini Parachironomus tenuicaudatus Chironomini Parachironomus tenuicaudatus Chironomini Phaenopsectra profusa Chironomini Phaenopsectra profusa Chironomini Polypedilum digitifer Chironomini Polypedilum digitifer Chironomini Polypedilum obtusum Chironomini Polypedilum obtusum Chironomini Polypedilum scalaenum Chironomini Polypedilum scalaenum Chironomini Polypedilum laetum Chironomini Polypedilum sulaceps Chironomini Stichtochironomus annulicrus Chironomini Stichtochironomus annulicrus Chironomini Stichtochironomus varius Chironomini Stichtochironomus varius Chironomini Xestochironomus brunneus Chironomini Xestochironomus brunneus Diamesinae Diamesa heteropus Diamesinae Diamesa heteropus Orthocladiinae Cricotopus annulator Orthocladiinae Cricotopus annulator Orthocladiinae Cricotopus bicinctus Orthocladiinae Cricotopus bicinctus Orthocladiinae Cricotopus blinni Orthocladiinae Cricotopus blinni Orthocladiinae Cricotopus infuscatus Orthocladiinae Cricotopus infuscatus Orthocladiinae Cricotopus trifascia Orthocladiinae Cricotopus trifasciatus Orthocladiinae Cricotopus herrmanni Orthocladiinae Eukiefferiella claripennis Orthocladiinae Eukiefferiella claripennis Orthocladiinae Eukiefferiella coerulescens Orthocladiinae Eukiefferiella coerulescens Orthocladiinae Eukiefferiella n. sp. 9 Orthocladiinae Eukiefferiella n. sp. 9 Orthocladiinae Orthocladius obumbratus Orthocladiinae Orthocladius obumbratus Orthocladiinae Parakiefferiella subaterrima Orthocladiinae Parakiefferiella subaterrima Orthocladiinae Parametriocnemus lundbeckii Orthocladiinae Parametriocnemus lundbeckii Orthocladiinae Smittia aterrima Orthocladiinae Smittia aterrima Orthocladiinae Tvetnia vitraces Orthocladiinae Tvetnia vitraces Prodiamesinae Odontomesa ferringtoni Tanypodinae Ablabesmyia mallochi Tanypodinae Ablabesmyia mallochi Tanypodinae Procladius bellus Tanypodinae Procladius bellus Tanypodinae Procladius culciformis Tanypodinae Procladius culciformis Tanypodinae Procladius freemani Tanypodinae Procladius freemani Tanypodiinae Procladius subletti Tanypodinae Tanypus punctipennis Tanypodinae Tanypus punctipennis Tanytarsini Micropsectra logani Tanytarsini Micropsectra logani Tanytarsini Micropsectra nigripila Tanytarsini Micropsectra nigripila Tanytarsini Paratanytarsus n. sp. Tanytarsini Micropsectra polita Tanytarsini Tanytarsus acifer

As the Fountain Creek empties into the Arkansas River, this close similarity of chironomid species composition may be due to a combination of larval drift and flight compensation. The upper Fountain has significant altitudinal, thermal, flow and substrate variances in comparison to the Arkansas River and this may be a contributing factor as to why there are fewer similar species between them.

Further, Bruce (2002) collected macroinvertebrates at 10 sites with only 4 sites located in the Fountain Creek and Monument Creek from 1998-2001; 40% of the collected taxa were chironomids and these were identified at the generic level only; 22 genera were identified and 16 of the 22 chironomids collected were genera also found in our 2007 and

2008 collections. There were no new species listed in the Bruce (2002) study.

Ruse et al. (2000) collected 198 adult chironomid species from 22 sites along the East

Fork and main stem (from Leadville to Pueblo) of the Arkansas River; 46 adult species of the

198 collected in Ruses’ work were identical to those collected in this study; 16 were new species of which 2 were identical to this work. Using pupal exuviae, Ruse (2010) identified,

313 chironomid species from lakes in Great Britain; 11 of these species found in this British ecosystem were also found in our lotic samples. Using canonical correspondence analysis,

Ruse identified chironomid species that preferred certain ranges of conductivity, pH and total phosphorus.

I used two different collection techniques, ultraviolet (UV) night lighting and sweep netting, to collect the greatest possible number of different species. Night lighting attracts only adults of selected chironomid species and sweep netting collects adult males that are forming swarms for the purpose of mating as well as chironomids that have landed on riparian vegetation. Some chironomid species are strongly attracted to a UV source and

53 others are not. Figures 38 and 39 show the total number of species collected for each technique for each year 2007 and 2008. In 2007, the two techniques were similarly successful but in 2008, the UV technique resulted in far more insects. Yet, the use of two collection techniques was successful as completely different chironomid species were collected from site LF-3 on September 7, 2007, at the same time (Table 4) as well as MC-5 on September 6,

2007, (Table 5) and LF-4 on September 1, 2007, (Table 6). New species were collected at all sites except LF-3 and the majority of new species (54%) were collected with ultraviolet light traps as Figure 41 indicates, while 17% were collected with sweep nets and 29% were collected using both techniques. Also, collecting two consecutive years, 2007 and 2008, ensured the representation of different chironomid subfamilies as depicted in the percent subfamily composition graphs in Figures 10 – 37.

The elevation of Monument and Upper Fountain Creeks decrease as the stream flows to its confluence with the Arkansas River and as such, the mean water temperature increases.

Water temperatures ranged from 4.6° C at UF-1 to 11.8° C during spring sampling and 5.8°

C to 15.8° C at LF-5 during fall sampling.

Further, orthoclads are primarily cold-water midges whereas Chironominae are primarily warm-water midges (Stevens, 1998). In this study I collected only 10% more chironomid species from subfamily Orthocladiinae than from subfamily Chironominae; this difference is not large and may be due to longitudinal and elevational effects and varying water temperatures within the Fountain Creek Watershed providing ideal habitat for many representatives from both of these chironomid families. Although not shown as significant by

CANOCO, as shown in Table 8, many different species were found between sites with

54 greater elevational differences indicating that elevation does influence chironomid species distributions.

Similarity indices were used in this work to determine species similarities between sites- not only comparisons within individual branches but also among the three branches of the Fountain Creek Watershed. Two indices, the Jaccard Coefficient of Community used by

Bruce (2002) and Sorensen’s Index used by Ruse (1989), were used to ensure the most accuracy of comparison. Although the values between the indices were quite different, the trends were very much similar. For the most part, many species similarities lie within the sites that are closest in proximity. Interestingly, both indices indicate that sites MC-3 and UF-

3, located in two different branches of the watershed, have the closest similarity of all.

Overall, there seems to be a good variation of species among all branches.

A spike in selenium occurred in all three water fractions (total, dissolved and pore) at site MC-5. In Figure 1, Pierre shale formations become predominant in Monument Creek at this location and many areas at this site are scoured to bedrock. The spike in selenium at MC-

5 was most likely from continual weathering and erosion of the Pierre shale. The increase in selenium at sites LF-4 and LF-5 was also most likely related to this geological source of selenium as CANOCO analysis indicated that selenium concentrations were significant at these sites. Turner (2009) concluded from his bryophyte study of selenium bioaccumulation that concentrations begin to increase at site MC-5 as well as at UF-4 downstream to the

Lower Fountain Creek’s confluence with the Arkansas River. In this research, selenium is significant in all samples except fall pore selenium. Concentrations may determine which chironomid species may tolerate a seleniferous environment from those that do not. For example, Polypedilum illinoense and Cladotanytarsus cruscula are two species that occur in

55 every CANOCO diagram on the outermost edge (most tolerant area) of the arrow indicating the selenium variable for total, dissolved and pore spring samples. The same occurs for all the fall samples indicating that species Tanytarsus pallidicomis and Telopelopia okoboji may be the most selenium tolerant. Carsella (2014) found that selenium, when coupled with iron, was concentrated in bryophyte tissue at higher levels as the pH became more acidic. The spring water samples are more acidic than the fall samples. In my work, pH is significant at

0.01 for all the fall water samples but not significant for spring samples.

Medium, fine, very fine, and coarse to fine sand were the main sediment sample components at all sites. This is what would be expected, as the Fountain Creek is primarily a sandy-bottomed stream that becomes more pronounced as it widens to its confluence with the

Arkansas River.

Conclusion

The goal of this research was to determine the chironomid species diversity of the

Fountain Creek Watershed and to analyze the possible environmental variables of elevation, substrate and selected water quality parameters that may have a limiting effect on the distribution of these chironomid species. Overall, 154 total chironomid species were collected and identified in this Fountain Creek Watershed study that included 24 new species found at every site except LF-3. The abundance of chironomid species found in this watershed indicates that Fountain Creek provides niches adequate for the survival of these various species. Compared to the chironomid collections of other studies, this study collected a greater number and a wider range of midge species.

Bruce (2002) determined that substrate was an influencing factor for invertebrate variations between his collection sites in the Fountain Creek Watershed. In this study,

CANOCO analysis did not determine substrate size as a significant factor for species habitat preferences despite the fact that the substrate is the environment in which chironomids dwell.

CANOCO analysis identified selenium as the most significant variable in all samples except fall pore selenium. Diffuse sources of selenium occur primarily at LF-4 and LF-5, corresponding to sites 13 and 14 on the CANOCO diagrams. As shown in Table 1, pierre shale formations comprise a large part of the riverbed in the lower Fountain Creek. As a result of the pierre shale weathering process, selenium leaching into the water may be the main contributing source of selenium in these areas. Certain species found in these areas may be indicative of chironomids more tolerant of higher concentrations of selenium than others.

Future research could be done using chironomids to determine the selenium/iron/pH

57 bioaccumulation connection as J. Carsella and others (unpublished, 2014) have been studying the bryophyte, Hygrohypnum ochraceum.

Data collected from this research provides a strong baseline for future chironomid studies to evaluate environmental impact within the Fountain Creek Watershed.

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Appendix A

Collecting Site Photos Upstream and Downstream

Figure 1. Site UF-1. Upstream (facing west). Photo by L. Helland

Figure 2. Site UF-1. Downstream (facing east). Photo by L. Helland.

Figure 3. Site UF-2. Upstream (facing west). At Soda Springs Park, Manitou Springs. Photo by L.Helland.

Figure 4. Site UF-2. Downstream (facing east). Photo by L. Helland

Figure 5. Site UF-3. Upstream (facing west). Photo by L. Helland.

Figure 6. Site UF-3. Downstream (facing east). Photo by L. Helland.

Figure 7. Site UF-4. Upstream (facing north). Photo by L. Helland.

Figure 8. Site UF-4. Downstream (facing south). Photo by L. Helland.

Figure 9. Site MC-1. Upstream (facing north). Photo by L. Helland.

Figure 10. Site MC-1. Downstream (facing south). Photo by L. Helland.

Figure 11. Site MC-2. Upstream (facing north). Photo by L. Helland

Figure 12. Site MC-2. Downstream (facing south). Photo by L. Helland.

Figure 13. Site MC-3. Upstream (facing north). Photo by L. Helland.

Figure 14. Site MC-3. Downstream (facing south). Photo by L. Helland.

Figure 15. Site MC-4. Upstream (facing north). Photo by L. Helland.

Figure 16. Site MC-4. Downstream (facing south). Photo by L. Helland.

Figure 17. Site MC-5. Upstream (facing north). Photo by L. Helland.

Figure 18. Site MC-5. Downstream (facing south). Photo by L. Helland.

Figure 19. Site LF-1. Upstream (facing south). Photo by L. Helland.

Figure 20. Site LF-1. Downstream (facing north). Photo by L. Helland.

Figure 21. Site LF-2. Upstream (facing north). Photo by S. Herrmann.

Figure 22. Site LF-2. Downstream (facing south). Photo by L. Helland.

Figure 23. Site LF-3. Upstream (facing north). Photo by L. Helland.

Figure 24. Site LF-3. Downstream (facing south). Photo by L. Helland.

Figure 25. Site LF-4. Upstream (facing north). Photo by S. Herrmann.

Figure 26. Site LF-4. Downstream (facing south). Photo by S. Herrmann.

Figure 27. Site LF-5. Upstream (facing north). Photo by L. Helland.

Figure 28. Site LF-5. Downstream (facing south). Photo by S. Herrmann.

Appendix B

Similarity Indices

1 0.9 0.8 0.7 0.6 0.5 0.4 0.3 0.2 0.28 0.31 0.28 0.23 0.1 0.18 0.22 0.18 0.19 0.15 0.12 0 MC1/2 MC1/3 MC1/4 MC1/5 MC2/3 MC2/4 MC2/5 MC3/4 MC3/5 MC4/5 Sites

Figure 42. Jaccard's Coefficient of Community for sites MC 1-5, column graphs comparing chironomid species similarities between sites for 2007 and 2008 combined collections. Species similarities are greater between comparable communities as the value approaches 1. Values are unrounded.

1 0.9 0.8 0.7 0.6 0.5 0.4 0.3 0.29 0.28 0.2 0.26 0.26 0.1 0.16 0.12 0 UF1/2 UF1/3 UF1/4 UF2/3 UF2/4 UF3/4 Sites

Figure 43. Jaccard's Coeffiecient of Community for sites UF 1-4, column graphs comparing chironomid species similarities between sites for 2007 and 2008 combined collections. Species similarities are greater between comparable communities as the value approaches 1. Values are unrounded.

1 0.9 0.8 0.7 0.6 0.5 0.4 0.3 0.2 0.32 0.23 0.25 0.24 0.1 0.21 0.18 0.19 0.21 0.22 0.2 0 LF1/2 LF1/3 LF1/4 LF1/5 LF2/3 LF2/4 LF2/5 LF3/4 LF3/5 LF4/5 Sites

Figure 44. Jaccard's Coefficient of Community for sites LF 1-5, column graphs comparing chironomid species similarities between sites for 2007 and 2008 combined collections. Species similarities are greater between comparable communities as the value approaches 1. Values are unrounded.

0.5 0.45 0.4 0.35 0.3 0.25 0.47 0.45 0.43 0.44 0.2 0.36 0.15 0.31 0.31 0.3 0.32 0.1 0.22 0.05 0 MC1/2 MC1/3 MC1/4 MC1/5 MC2/3 MC2/4 MC2/5 MC3/4 MC3/5 MC4/5 Sites

Figure 45. Sørensen’s Index (Odum, 1971) for sites MC 1-5, column graphs comparing chironomid species similarities between sites for 2007 and 2008 combined collections. Species similarities are greater between comparable communities as the value approaches 1. Values are unrounded.

0.6

0.5

0.4

0.3 0.51 0.45 0.2 0.41 0.41

0.27 0.1 0.22

0 UF1/2 UF1/3 UF1/4 UF2/3 UF2/4 UF3/4 Sites

Figure 46. Sørensen’s Index for sites UF 1-4, column graphs comparing chironomid species similarities between sites for 2007 and 2008 combined collections. Species similarities are greater between comparable communities as the value approaches 1. Values are unrounded.

0.6

0.5

0.4

0.3

0.48 0.2 0.4 0.39 0.37 0.36 0.35 0.34 0.33 0.3 0.32 0.1

0 LF1/2 LF1/3 LF1/4 LF1/5 LF2/3 LF2/4 LF2/5 LF3/4 LF3/5 LF4/5 Sites

Figure 47. Sørensen’s Index for sites LF 1-5, column graphs comparing chironomid species similarities between sites for 2007 and 2008 combined collections. Species similarities are greater between comparable communities as the value approaches 1. Values are unrounded.

0.8

0.6

0.4 0.33 0.25 0.21 0.19 0.18 0.19 0.2 0.2 0.14 0.16 0.16 0.12 0.11 0.09 0.08 0.09 0.1 0.06 0.07 0.05 0.06 0

Figure 48. Jaccard's Coefficient of Community MC vs UF.

1 0.9 0.8 0.7 0.6 0.5 0.4 0.3 0.5 0.2 0.41 0.42 0.32 0.35 0.31 0.3 0.28 0.27 0.24 0.22 0.2 0.1 0.17 0.15 0.16 0.19 0.12 0.13 0.09 0.11 0

Figure 49. Sørensen’s Index MC vs UF.

1 0.9 0.8 0.7 0.6 0.5 0.4 0.3 0.2 0.45 0.320.32 0.26 0.29 0.29 0.260.27 0.22 0.2 0.21 0.21 0.2 0.2 0.1 0.15 0.15 0.16 0.19 0.19 0.18 0.15 0.15 0.14 0.090.13

MC1/LF2 MC1/LF3 MC1/LF4 MC1/LF5 MC2/LF1 MC2/LF2 MC2/LF3 MC2/LF4 MC2/LF5 MC3/LF1 MC3/LF2 MC3/LF3 MC3/LF4 MC3/LF5 MC4/LF1 MC4/LF2 MC4/LF3 MC4/LF4 MC4/LF5 MC5/LF1 MC5/LF2 MC5/LF3 MC5/LF4 MC5/LF5 MC1/LF1 Figure 50. Jaccard's Coefficient of Community MC vs LF.

1 0.9 0.8 0.7 0.6 0.5 0.4 0.72 0.3 0.5 0.480.49 0.44 0.45 0.410.42 0.2 0.36 0.35 0.35 0.34 0.32 0.31 0.3 0.33 0.33 0.260.240.26 0.23 0.27 0.26 0.25 0.1 0.17

1 0.9 0.8 0.7 0.6 0.5 0.4 0.3 0.2 0.26 0.29 0.1 0.21 0.17 0.15 0.16 0.18 0.12 0.07 0.09 0.06 0.1 0.05 0.06 0.08 0 0.02 0.09 0.13 0.12 0

Figure 52. Jaccard's Coefficient of Community UF vs LF.

1 0.9 0.8 0.7 0.6 0.5 0.4 0.3 0.2 0.42 0.44 0.36 0.35 0.32 0.3 0.1 0.21 0.25 0.23 0.21 0.17 0.19 0.15 0.17 0.13 0.11 0.1 0.11 0 0.05 0

Figure 53. Sørensen’s Index UF vs LF.

Appendix C

Canonical Correspondence Analysis (CANOCO) Logs

Spring Total Selenium

[Fri Dec 13 16:47:44 2013] Log file created [Fri Dec 13 16:49:13 2013] Running CANOCO: [Fri Dec 13 16:49:13 2013] CON file [C:\canoco\Lisa_Helland\November_15\spring_pH_Se_Silt.con] saved Program CANOCO Version 4.54 October 2005 - written by Cajo J.F. Ter Braak (C) 1988-2005 Biometris - quantitative methods in the life and earth sciences Plant Research International, Wageningen University and Research Centre Box 100, 6700 AC Wageningen, the Netherlands CANOCO performs (partial) (detrended) (canonical) correspondence analysis, principal components analysis and redundancy analysis. CANOCO is an extension of Cornell Ecology program DECORANA (Hill,1979) For explanation of the input/output see the manual or Ter Braak, C.J.F. (1995) Ordination. Chapter 5 in: Data Analysis in Community and Landscape Ecology (Jongman, R.H.G., Ter Braak, C.J.F. and Van Tongeren, O.F.R., Eds) Cambridge University Press, Cambridge, UK, 91-173 pp.

*** Type of analysis *** Model Gradient analysis indirect direct hybrid linear 1=PCA 2= RDA 3 unimodal 4= CA 5= CCA 6 ,, 7=DCA 8=DCCA 9 10=non-standard analysis Type analysis number Answer = 5

*** Data files *** Species data : C:\canoco\Lisa_Helland\November_15\species.dta Covariable data : Environmental data : C:\canoco\Lisa_Helland\November_15\spring_factors.dta Initialization file: Forward selection of envi. variables = 1 Scaling of ordination scores = 2 Diagnostics = 3 File : C:\canoco\Lisa_Helland\November_15\species.dta Title : WCanoImp produced data file Format : (I5,1X,8(I6,F3.0)) No. of couplets of species number and abundance per line : 8 No samples omitted Number of samples 14 Number of species 156 Number of occurrences 3 File : C:\canoco\Lisa_Helland\November_15\spring_factors.dta Title : WCanoImp produced data file Format : (I5,1X,3(I6,F16.9)) No. of environmental variables : 20 No interaction terms defined No transformation of species data No species-weights specified No sample-weights specified No downweighting of rare species No. of active samples: 14

No. of passive samples: 0 No. of active species: 155 Total inertia in species data= Sum of all eigenvalues of CA = 4.50600

**** Start of forward selection of variables **** *** Unrestricted permutation *** Seeds: 23239 945 N Name Extra fit 9 Silt/Cla 0.3459 12 pH Avg. 0.4104 19 Total_Se 0.4699 Environmental variable 19 tested Number of permutations= 5000

*** Permutation under reduced model *** P-value 0.0006 (variable 19; F-ratio= 1.40; number of permutations= 5000) Environmental variable 19 added to model Variance explained by the variables selected: 0.47 " " " all variables : 1.11 N Name Extra fit 12 pH Avg. 0.2942 9 Silt/Cla 0.3450 Environmental variable 9 tested Number of permutations= 5000

*** Permutation under reduced model *** P-value 0.4283 (variable 9; F-ratio= 1.03; number of permutations= 5000) Environmental variable 9 added to model Variance explained by the variables selected: 0.81 " " " all variables : 1.11 N Name Extra fit 12 pH Avg. 0.2973 Environmental variable 12 tested Number of permutations= 5000 *** Permutation under reduced model *** P-value 0.7389 (variable 12; F-ratio= 0.88; number of permutations= 5000) Environmental variable 12 added to model Variance explained by the variables selected: 1.11 " " " all variables : 1.11 No more variables to improve fit

*** End of selection *** N name (weighted) mean stand. dev. inflation factor 1 SPEC AX1 0.0000 1.0252 2 SPEC AX2 0.0000 1.0075 3 SPEC AX3 0.0000 1.0701 4 SPEC AX4 0.0000 1.0000 5 ENVI AX1 0.0000 1.0000 6 ENVI AX2 0.0000 1.0000 7 ENVI AX3 0.0000 1.0000 8 ENVI AX4 0.0000 0.0000 9 Silt/Cla 0.2879 0.2669 1.0222 12 pH Avg. 7.4737 0.1723 2.6890

19 Total_Se -0.0689 1.2552 2.6732

**** Summary **** Axes 1 2 3 4 Total inertia Eigenvalues : 0.478 0.341 0.293 0.528 4.506 Species-environment correlations : 0.975 0.993 0.934 0.000 Cumulative percentage variance of species data : 10.6 18.2 24.7 36.4 of species-environment relation: 43.0 73.6 100.0 0.0 Sum of all eigenvalues 4.506 Sum of all canonical eigenvalues 1.112 The first three eigenvalues reported above are canonical, the fourth is not since only three independent constraints can be formed from the environmental variables. [Fri Dec 13 16:49:15 2013] CANOCO call succeeded

Spring Dissolved Selenium

*** Type of analysis *** Model Gradient analysis indirect direct hybrid linear 1=PCA 2= RDA 3 unimodal 4= CA 5= CCA 6 ,, 7=DCA 8=DCCA 9 10=non-standard analysis Type analysis number Answer = 5

File : C:\canoco\Lisa_Helland\November_15\spring_factors.dta Title : WCanoImp produced data file Format : (I5,1X,3(I6,F16.9)) No. of environmental variables : 20 No interaction terms defined No transformation of species data No species-weights specified No sample-weights specified No downweighting of rare species

No. of active samples: 14 No. of passive samples: 0 No. of active species: 155 Total inertia in species data= Sum of all eigenvalues of CA = 4.50600

**** Start of forward selection of variables **** *** Unrestricted permutation *** Seeds: 23239 945 N Name Extra fit 9 Silt/Cla 0.3459 12 pH Avg. 0.4104 18 Dissolve 0.4698 Environmental variable 18 tested Number of permutations= 5000

*** Permutation under reduced model *** P-value 0.0006 (variable 18; F-ratio= 1.40; number of permutations= 5000) Environmental variable 18 added to model Variance explained by the variables selected: 0.47 " " " all variables : 1.11 N Name Extra fit 12 pH Avg. 0.2948 9 Silt/Cla 0.3452Environmental variable 9 tested Number of permutations= 5000

*** Permutation under reduced model *** P-value 0.4267 (variable 9; F-ratio= 1.03; number of permutations= 5000) Environmental variable 9 added to model Variance explained by the variables selected: 0.81 " " " all variables : 1.11 N Name Extra fit 12 pH Avg. 0.2980 Environmental variable 12 tested Number of permutations= 5000 *** Permutation under reduced model *** P-value 0.7367 (variable 12; F-ratio= 0.88; number of permutations= 5000) Environmental variable 12 added to model Variance explained by the variables selected: 1.11 " " " all variables : 1.11 No more variables to improve fit

*** End of selection *** N name (weighted) mean stand. dev. inflation factor 1 SPEC AX1 0.0000 1.0256 2 SPEC AX2 0.0000 1.0074 3 SPEC AX3 0.0000 1.0693 4 SPEC AX4 0.0000 1.0000 5 ENVI AX1 0.0000 1.0000 6 ENVI AX2 0.0000 1.0000 7 ENVI AX3 0.0000 1.0000 8 ENVI AX4 0.0000 0.0000 9 Silt/Cla 0.2879 0.2669 1.0214 12 pH Avg. 7.4737 0.1723 2.7474

18 Dissolve -0.0868 1.2849 2.7311

**** Summary **** Axes 1 2 3 4 Total inertia

Eigenvalues : 0.477 0.342 0.294 0.529 4.506 Species-environment correlations : 0.975 0.993 0.935 0.000 Cumulative percentage variance of species data : 10.6 18.2 24.7 36.4 of species-environment relation: 42.9 73.6 100.0 0.0 Sum of all eigenvalues 4.506 Sum of all canonical eigenvalues 1.113 The first three eigenvalues reported above are canonical, the fourth is not since only three independent constraints can be formed from the environmental variables.

Spring Pore Selenium

*** Type of analysis *** Model Gradient analysis indirect direct hybrid linear 1=PCA 2= RDA 3 unimodal 4= CA 5= CCA 6 ,, 7=DCA 8=DCCA 9 10=non-standard analysis Type analysis number Answer = 5

*** Data files *** Species data : C:\canoco\Lisa_Helland\November_15\species.dta Covariable data : Environmental data : C:\canoco\Lisa_Helland\November_15\spring_factors.dta Initialization file: Forward selection of envi. variables = 1 Scaling of ordination scores = 2 Diagnostics = 3 File : C:\canoco\Lisa_Helland\November_15\species.dta Title : WCanoImp produced data file Format : (I5,1X,8(I6,F3.0)) No. of couplets of species number and abundance per line : 8 No samples omitted Number of samples 14 Number of species 156 Number of occurrences 387 File : C:\canoco\Lisa_Helland\November_15\spring_factors.dta Title : WCanoImp produced data file Format : (I5,1X,3(I6,F16.9)) No. of environmental variables : 20 No interaction terms defined No transformation of species data

No species-weights specified No sample-weights specified No downweighting of rare species

No. of active samples: 14 No. of passive samples: 0 No. of active species: 155 Total inertia in species data= Sum of all eigenvalues of CA = 4.50600

**** Start of forward selection of variables **** *** Unrestricted permutation *** Seeds: 23239 945 N Name Extra fit 9 Silt/Cla 0.3459 12 pH Avg. 0.4104 20 Pore_Se_ 0.4523 Environmental variable 20 tested Number of permutations= 5000

*** Permutation under reduced model *** P-value 0.0034 (variable 20; F-ratio= 1.34; number of permutations= 5000) Environmental variable 20 added to model Variance explained by the variables selected: 0.45 " " " all variables : 1.10 N Name Extra fit 12 pH Avg. 0.2875 9 Silt/Cla 0.3496 Environmental variable 9 tested Number of permutations= 5000

*** Permutation under reduced model *** P-value 0.4017 (variable 9; F-ratio= 1.04; number of permutations= 5000) Environmental variable 9 added to model Variance explained by the variables selected: 0.80 " " " all variables : 1.10 N Name Extra fit 12 pH Avg. 0.2960 Environmental variable 12 tested Number of permutations= 5000

*** Permutation under reduced model *** P-value 0.7469 (variable 12; F-ratio= 0.87; number of permutations= 5000) Environmental variable 12 added to model Variance explained by the variables selected: 1.10 " " " all variables : 1.10 No more variables to improve fit

*** End of selection *** N name (weighted) mean stand. dev. inflation factor 1 SPEC AX1 0.0000 1.0235 2 SPEC AX2 0.0000 1.0236 3 SPEC AX3 0.0000 1.0302 4 SPEC AX4 0.0000 1.0000 5 ENVI AX1 0.0000 1.0000 6 ENVI AX2 0.0000 1.0000 7 ENVI AX3 0.0000 1.0000 8 ENVI AX4 0.0000 0.0000

9 Silt/Cla 0.2879 0.2669 1.0800 12 pH Avg. 7.4737 0.1723 1.4573 20 Pore_Se_ -0.1042 1.7441 1.4965

**** Summary **** Axes 1 2 3 4 Total inertia Eigenvalues : 0.488 0.360 0.250 0.534 4.506 Species-environment correlations : 0.977 0.977 0.971 0.000 Cumulative percentage variance of species data : 10.8 18.8 24.4 36.2 of species-environment relation: 44.5 77.2 100.0 0.0 Sum of all eigenvalues 4.506 Sum of all canonical eigenvalues 1.098 The first three eigenvalues reported above are canonical, the fourth is not since only three independent constraints can be formed from the environmental variables.

Fall Total Selenium

*** Type of analysis *** Model Gradient analysis indirect direct hybrid linear 1=PCA 2= RDA 3 unimodal 4= CA 5= CCA 6 ,, 7=DCA 8=DCCA 9 10=non-standard analysis Type analysis number Answer = 5

*** Data files *** Species data : C:\canoco\Lisa_Helland\October_25\species.dta Covariable data : Environmental data : C:\canoco\Lisa_Helland\October_25\fall_factors.dta Initialization file: Forward selection of envi. variables = 1 Scaling of ordination scores = 2 Diagnostics = 3 File : C:\canoco\Lisa_Helland\October_25\species.dta Title : WCanoImp produced data file Format : (I5,1X,8(I6,F3.0)) No. of couplets of species number and abundance per line : 8 No samples omitted Number of samples 14 Number of species 156 Number of occurrences 387 File : C:\canoco\Lisa_Helland\October_25\fall_factors.dta Title : WCanoImp produced data file Format : (I5,1X,4(I6,F9.2)) No. of environmental variables : 15 No interaction terms defined

No transformation of species data No species-weights specified No sample-weights specified

No downweighting of rare species No. of active samples: 14 No. of passive samples: 0 No. of active species: 155 Total inertia in species data= Sum of all eigenvalues of CA = 4.50600

****** Check on influence in covariable/environment data ****** The following sample(s) have extreme values Sample Environmental Covariable + Environment space variable Influence influence influence 1 13 5.8x

****** End of check ****** **** Start of forward selection of variables **** *** Unrestricted permutation *** Seeds: 23239 945 N Name Extra fit 9 Silt/Cla 0.3459 10 Se_total 0.4364 13 pH Avg. 0.4898 Environmental variable 13 tested Number of permutations= 5000 P-value 0.0100 (variable 13; F-ratio= 1.46; number of permutations= 5000) Environmental variable 13 added to model Variance explained by the variables selected: 0.49 " " " all variables : 1.24 N Name Extra fit 9 Silt/Cla 0.3244 10 Se_total 0.4248 Environmental variable 10 tested Number of permutations= 5000 P-value 0.0540 (variable 10; F-ratio= 1.30; number of permutations= 5000) Environmental variable 10 added to model Variance explained by the variables selected: 0.91 " " " all variables : 1.24 N Name Extra fit 9 Silt/Cla 0.3253 Environmental variable 9 tested Number of permutations= 5000 P-value 0.5465 (variable 9; F-ratio= 1.00; number of permutations= 5000) Environmental variable 9 added to model Variance explained by the variables selected: 1.24 " " " all variables : 1.24 No more variables to improve fit

*** End of selection *** N name (weighted) mean stand. dev. inflation factor 1 SPEC AX1 0.0000 1.0244 2 SPEC AX2 0.0000 1.0384 3 SPEC AX3 0.0000 1.0089 4 SPEC AX4 0.0000 1.0000 5 ENVI AX1 0.0000 1.0000 6 ENVI AX2 0.0000 1.0000

7 ENVI AX3 0.0000 1.0000 8 ENVI AX4 0.0000 0.0000 9 Silt/Cla 0.2879 0.2669 1.2090 10 Se_total 2.8419 4.3419 1.3353 13 pH Avg. 8.0709 0.0791 1.3787

**** Summary **** Axes 1 2 3 4 Total inertia Eigenvalues : 0.490 0.426 0.324 0.503 4.506 Species-environment correlations : 0.976 0.963 0.991 0.000 Cumulative percentage variance of species data : 10.9 20.3 27.5 38.7 of species-environment relation: 39.5 73.9 100.0 0.0 Sum of all eigenvalues 4.506 Sum of all canonical eigenvalues 1.240

Fall Dissolved Selenium

*** Type of analysis *** Model Gradient analysis indirect direct hybrid linear 1=PCA 2= RDA 3 unimodal 4= CA 5= CCA 6 ,, 7=DCA 8=DCCA 9 10=non-standard analysis Type analysis number Answer = 5

*** Data files *** Species data : C:\canoco\Lisa_Helland\November_15\species.dta Covariable data : Environmental data : C:\canoco\Lisa_Helland\November_15\fall_factors.dta Initialization file: Forward selection of envi. variables = 1 Scaling of ordination scores = 2 Diagnostics = 3 File : C:\canoco\Lisa_Helland\November_15\species.dta Title : WCanoImp produced data file Format : (I5,1X,8(I6,F3.0)) No. of couplets of species number and abundance per line : 8 No samples omitted Number of samples 14 Number of species 156 Number of occurrences 387

File : C:\canoco\Lisa_Helland\November_15\fall_factors.dta Title : WCanoImp produced data file Format : (I5,1X,3(I6,F16.9)) No. of environmental variables : 20 No interaction terms defined No transformation of species data No species-weights specified No sample-weights specified No downweighting of rare species

No. of active samples: 14 No. of passive samples: 0 No. of active species: 155 Total inertia in species data= Sum of all eigenvalues of CA = 4.50600

****** End of check ****** **** Start of forward selection of variables **** *** Unrestricted permutation *** Seeds: 23239 945 N Name Extra fit 9 Silt/Cla 0.3459 18 Dissolve 0.4837 12 pH Avg. 0.4898 Environmental variable 12 tested Number of permutations= 5000 P-value 0.0100 (variable 12; F-ratio= 1.46; number of permutations= 5000) Environmental variable 12 added to model Variance explained by the variables selected: 0.49 " " " all variables : 1.30 N Name Extra fit 9 Silt/Cla 0.3244 18 Dissolve 0.4798 Environmental variable 18 tested Number of permutations= 5000 P-value 0.0044 (variable 18; F-ratio= 1.49; number of permutations= 5000) Environmental variable 18 added to model Variance explained by the variables selected: 0.97 " " " all variables : 1.30

N Name Extra fit 9 Silt/Cla 0.3260 Environmental variable 9 tested Number of permutations= 5000 P-value 0.5083 (variable 9; F-ratio= 1.02; number of permutations= 5000) Environmental variable 9 added to model Variance explained by the variables selected: 1.30 " " " all variables : 1.30 No more variables to improve fit

*** End of selection *** *** BEWARE *** Residual for axis 1 bigger than tolerance, which is 0.0000010 N name (weighted) mean stand. dev. inflation factor 1 SPEC AX1 0.0000 1.0175 2 SPEC AX2 0.0000 1.0324 3 SPEC AX3 0.0000 1.0105 4 SPEC AX4 0.0000 1.0000

5 ENVI AX1 0.0000 1.0000 6 ENVI AX2 0.0000 1.0000 7 ENVI AX3 0.0000 1.0000 8 ENVI AX4 0.0000 0.0000 9 Silt/Cla 0.2879 0.2669 1.1713 12 pH Avg. 8.0709 0.0791 1.2270 18 Dissolve -0.2155 1.9383 1.0884

**** Summary **** Axes 1 2 3 4 Total inertia Eigenvalues : 0.493 0.478 0.324 0.491 4.506 Species-environment correlations : 0.983 0.969 0.990 0.000 Cumulative percentage variance of species data : 10.9 21.6 28.8 39.7 of species-environment relation: 38.1 75.0 100.0 0.0 Sum of all eigenvalues 4.506 Sum of all canonical eigenvalues 1.296

Fall Pore Selenium

*** Type of analysis *** Model Gradient analysis indirect direct hybrid linear 1=PCA 2= RDA 3 unimodal 4= CA 5= CCA 6 ,, 7=DCA 8=DCCA 9 10=non-standard analysis Type analysis number Answer = 5

No interaction terms defined No transformation of species data No species-weights specified

No sample-weights specified No downweighting of rare species No. of active samples: 14 No. of passive samples: 0 No. of active species: 155 Total inertia in species data= Sum of all eigenvalues of CA = 4.50600

****** End of check ****** **** Start of forward selection of variables **** *** Unrestricted permutation *** Seeds: 23239 945 N Name Extra fit 9 Silt/Cla 0.3459 20 Pore_Se_ 0.4276 12 pH Avg. 0.4898 Environmental variable 12 tested Number of permutations= 5000 P-value 0.0100 (variable 12; F-ratio= 1.46; number of permutations= 5000) Environmental variable 12 added to model Variance explained by the variables selected: 0.49 " " " all variables : 1.22 N Name Extra fit 9 Silt/Cla 0.3244 20 Pore_Se_ 0.4033 Environmental variable 20 tested Number of permutations= 5000 P-value 0.1266 (variable 20; F-ratio= 1.23; number of permutations= 5000) Environmental variable 20 added to model Variance explained by the variables selected: 0.89 " " " all variables : 1.22 N Name Extra fit 9 Silt/Cla 0.3254 Environmental variable 9 tested P-value 0.5467 (variable 9; F-ratio= 0.99; number of permutations= 5000) Environmental variable 9 added to model Variance explained by the variables selected: 1.22 " " " all variables : 1.22 No more variables to improve fit

*** End of selection *** N name (weighted) mean stand. dev. inflation factor

1 SPEC AX1 0.0000 1.0210 2 SPEC AX2 0.0000 1.0534 3 SPEC AX3 0.0000 1.0153

4 SPEC AX4 0.0000 1.0000 5 ENVI AX1 0.0000 1.0000 6 ENVI AX2 0.0000 1.0000 7 ENVI AX3 0.0000 1.0000 8 ENVI AX4 0.0000 0.0000 9 Silt/Cla 0.2879 0.2669 1.1652 12 pH Avg. 8.0709 0.0791 1.3863 20 Pore_Se_ 0.1501 1.4061 1.2395

**** Summary **** Axes 1 2 3 4 Total inertia Eigenvalues : 0.491 0.404 0.323 0.492 4.506 pecies-environment correlations : 0.979 0.949 0.985 0.000 Cumulative percentage variance of species data : 10.9 19.9 27.0 38.0 of species-environment relation: 40.3 73.5 100.0 0.0 Sum of all eigenvalues 4.506 Sum of all canonical eigenvalues 1.218