2019 Update: Dykstra’s Interpopulation Crosses

Small remnant Echinacea populations may suffer from inbreeding depression. To assess whether gene flow (in the form of pollen) from another population could “rescue” these populations from inbreeding depression, we hand pollinated Echinacea from six different prairie remnants with pollen from a large prairie remnant (Staffanson Prairie) and from a relatively small population that we call “Northwest Landfill.” We also performed within-population crosses as a control. Amy Dykstra planted achenes (seeds) that resulted from these crosses in an experimental plot at Hegg Lake WMA.

Plants in the crossing plots were originally found as seedlings like this one

We sowed a total of 15,491 achenes in 2008. 449 of these achenes germinated and emerged as seedlings. Each summer we census the surviving plants and measure them. This summer we found 48 surviving plants. None of these plants has flowered, but we think some of them are close! The largest plant we measured had 4 leaves, the longest of which was 35 cm.

You can read more about the interpopulation crosses, as well as links to prior flog entries mentioning the experiment, on the background page for this experiment.

Start year: 2008

Location: Hegg Lake WMA

Data collected: Plant fitness measurements (plant status, number of rosettes, number of leaves, and length of longest leaf), and notes about herbivory. Contact Amy Dykstra to access this data.

Products: Dykstra, A. B. 2013. Seedling recruitment in fragmented populations of Echinacea angustifolia. Ph.D. Dissertation. University of Minnesota. PDF

2019 Update: Echinacea Ploidy

To see how ploidy varies in Echinacea species in our study site, in fall 2019, wecollected and dried tissue from E. pallida, E. angustifolia, and E. purpurea. We also collected tissue from potential hybrids and known hybrids. We brought the dried tissue back to the Chicago Botanic Garden, where we plan to analyze ploidy using a flow cytometer, a device that can be used to find relative genome size.

The flow cytometer used to assess relative genome size. Although it just looks like a box, it is truly a powerful machine.
Example of dried Echinacea tissue… We thought it might be a hybrid when we visited it in the field!

Unfortunately, preparing samples for the flow cytometer is difficult, so we are going to first optimize our Echinacea tissue preparation protocol using live tissue. To do this, we withdrew accessions of E. angustifolia, E. pallida, and E. purpurea from Millennium Seed Bank at the CBG. For E. pallida, we took seed from collections throughout its range to see if its ploidy varies with latitude. We are currently germinating this live tissue to use for ploidy analysis with Elif. We are very excited to see what we find – any finding will help expand genomic knowledge for the genus Echinacea!

Start year: 2019

Location: Hegg Lake WMA, various prairie remnants and restorations, hybrid experimental plots

Overlaps with: Echinacea hybrids (exPt6, exPt7, exPt9), Echinacea pallida flowering phenology

Data/ materials collected: Dried tissue from plants throughout the study area; samples are currently held at Chicago Botanic Garden, in a small box in the glass cupboard to the right upon entering room 159, the Population Biology Lab. Updates will be posted when genome data is available.

Background: If you’d like to learn more about this experiment, check out the background flog post!

2019 Update: Pulse-steady pollination experiment

In 2019 Team Echinacea conducted a new experiment called “Pulse-Steady,” with roots in Ashley Barto’s 2017 REU project. The experiment investigates whether flowering Echinacea plants which received a resource pulse (pollination every three days) set seed at a different rate to Echinacea which received a steady flow of resources (pollination every day.)

Shea expresses frustration with the bees who beat us to the pollen—bagging flowering plants to ensure we had pollen sources became critical at the end of flowering!

Stuart and Gretel selected 48 flowering Echinacea with single flowering heads and assigned 24 to the pulse treatment, and the other 24 to the steady treatment. The team placed pollinator exclusion bags on the heads of all plants prior to the beginning of flowering to ensure that humans were the only pollinators. The team returned to exPt 2 every day from July 16 to August 7 to count anthers and styles and hand-pollinate the 48 heads, though rain caused pollen to present at strange times or not at all on some days. The team collected pollen from other flowering plants in exPt2 as well as bagged heads around Hegg Lake. Pollen samples included a minimum of four sires to ensure that compatible S-alleles were present in the mix. Pollinators collected additional pollen from heads in the experiment after pollinating the styles, to prevent self-pollen from clogging the styles and to replenish dwindling pollen supplies. Human pollinators frequently competed with insect pollinators, as pollen was scarce at the end of the flowering season, and had to wave off bees from taking pollen from experimental heads and pollen donors in the plot.

In December, Carleton externs Jack, Eli and Emma worked on a modified ACE protocol to process the harvested pulse-steady heads. They cleaned the heads and carefully separated the achenes based on their position in the head so that we can investigate whether seed set differs at the beginning, middle and end of flowering between the treatments, as well as whether seed set differs based on style “freshness” in the pulse treatment. They also scanned the heads with achenes separated out by location in the head.

Start year: 2019

Location: exPt 2, Douglas County, MN

Overlaps with: Flowering phenology in experimental plots, common garden experiment

Data/materials collected: The team harvested 48 heads in the experiment which have been cleaned and are ready to be randomized and x-rayed at the CBG. Each head has 8 envelopes associated with it (7 envelopes of achenes and 1 of chaff.)

Maps and datasheets for the field experiment are located at ~Dropbox\teamEchinacea2019\pulseSteady

The cleaning protocol and datasheets relevant to cleaning are located at ~Dropbox\CGData\150_clean\clean2019\inb2PulseSteady

2019 update: Amy Dykstra’s local adaptation experiment

This experiment was designed to study how well adapted Echinacea populations are to their local environments. Amy collected achenes from Echinacea populations in western South Dakota, central South Dakota, and Minnesota, and then sowed seeds from all three sources into experimental plots near each collection site. You can read more about the experiment and see a map of the seed source sites on the background page for this experiment.

This summer, during our annual census of the experimental plots, we found 67 living Echinacea plants in the western South Dakota plot, including 9 flowering plants. We found 116 living plants, including one flowering plant in the Minnesota plot. This was the first flowering plant in the Minnesota plot! (We abandoned the central SD plot after it was inadvertently sprayed in 2009, killing all the Echinacea). For more details and graphs, please read this brief report.

First flowering plant in the MN local adaptation plot!

Start year: 2008

Location: Grand River National Grassland (Western South Dakota), Samuel H. Ordway Prairie (Central South Dakota), Staffanson Prairie Preserve (West Central Minnesota), and Hegg Lake WMA (West Central Minnesota).

Overlaps with: Dykstra’s interpopulation crosses

Data/ materials collected: Plant fitness measurements (plant status, number of rosettes, number of leaves, and length of longest leaf), heads from all flowering plants

Products: Dykstra, A. B. 2013. Seedling recruitment in fragmented populations of Echinacea angustifolia. Ph.D. Dissertation. University of Minnesota. PDF

Winter Internship Week 2

This past week, I continued work on multiple projects. I continued to recheck and label cleaned Echinacea heads, growing the supply of achenes that are ready to be scanned, so that we can keep the samples moving through the steps. I also spent considerable time on randomizing. I resorted the informative and uninformative achenes from some of the 2013 and 2014 collection so that they are organized in the same way as more recent samples and so protocol is consistent across the board. After resorting was complete, I did the standard randomization protocol on achenes from 2018.

In addition to working with the Echinacea collection, I continued to organized the native bee collection. One task in particular has been going through the specimens, checking the SPID numbers, and checking them off on a data sheet to confirm which specimens still exist in the collection and which ones have been removed or discarded. I have labeled each smaller box within the cases with Roman numerals and have recorded on the data sheet in which box each specimen can be found. I will continue this project in the last few days of my internship and hopefully complete it so that there is a definite record of the specimens in the native bee collection.

2019 Update: Seedling Establishment

This field season the team continued the seedling recruitment experiment begun in 2007. The original goal of the project was to determine the rates of establishment and growth of seedlings in remnant populations of Echinacea angustifolia. From 2007 to 2013, plants which had flowered in the preceding year were visited in the spring to find any resulting seedlings. Each fall since then the team visits the plants, re-finds the seedlings and measures the living offspring.

In 2019 Team Echinacea visited 69 focal maternal plants in 12 prairie remnants to determine the survival and growth of their offspring. The team searched for 128 of the original 955 seedlings and found 87 of them (10 fewer than were found in 2018.) However, the team also re-found 27 seedlings which could not be located in 2018. None of the original seedlings flowered this year.

In addition to the annual seedling search, in 2019 the team began working on a project to relate the fitness and mating scene of maternal plants to their offspring. In late October 2019, Erin and Riley staked to the locations of 451 maternal plants included in the original experiment which now have “inactive” circles with no living seedlings. They visited these maternal plants because they were not included in demo in 2018 or 2019, and their goal was to determine the status of every original maternal plant. Since then, a group of Team Echinacea alums and current members has begun working to prepare the data for analysis in 2020 with the goal of submitting a manuscript in the spring.

The first day of searching occurred at East Elk Lake Road, where 13 of the original maternal plants still have living offspring.

Sites with seedling searches
East Elk Lake Road, East Riley, East of Town Hall, KJ’s, Loeffler’s Corner, Landfill, Nessman, Northwest of Landfill, Riley, Steven’s Approach, South of Golf Course, Staffanson Prairie

Start year: 2007

Location: Douglas County, MN

Overlaps with: Demographic census in the remnants

Data/materials collected: The EchinaceaSeedlings repository holds the data for this experiment. Lea Richardson restructured the repo in December 2019 to facilitate collaboration on the new project.

Notes on the project and master datasheet scans are at ~Dropbox\remData\115_trackSeedlings\slingRefindsFall2019

Data specific to the new 2019 project, including maps and datasheets used to refind the inactive circle maternal plants, is at ~Dropbox\slingProject2019

Team members involved with the 2019 project: Lea Richardson, Erin Eichenberger, Riley Thoen, Drake Mullett, Amy Waananen, Scott Nordstrom, Will Reed, Amy Dykstra, Gretel Kiefer , Stuart Wagenius

Products: Amy Dykstra used seedling survival data from 2010 and 2011 to model population growth rates as a part of her dissertation.

You can read more about seedling establishment, as well as links to prior flog entries mentioning the experiment, on the background page for this experiment.

2019 update: Demographic census in remnants

Poor weather conditions delayed demo this year but did not dampen our spirits! In 2019 Team Echinacea added 4031 visor records to demo and 1431 GPS points to surv. The largest effort this summer occurred at Aanenson where eight team members took demo records on over 200 flowering plants. We found flowering plants at Northwest of Landfill with tags from 1995 in situ, meaning the plants were at least several years older than most of the team. We also found non-angustifolia Echinacea invading prairie remnants Dog and Yellow Orchid Hill.

The team does demo at everyone’s favorite site: East Riley! Also known as Whose Loc Is It Anyway? Where the status is “Can’t find” and the GPS points don’t matter.

This year we performed demo and surv at 32 prairie remnants and 10 additional sites with angustifolia populations. For our smaller sites we visit every mature plant (ones which have flowered before.) At the larger sites we measure a subsample of the mature population. At every site we also perform flowering demo, where we visit plants which flowered for the first time or were not included in the subsample. We record the status of each plant we visit, its neighbors and the number of heads it produced. All flowering plants are tagged and shot in surv, and in the coming months we will use demap to reconcile the 2019 demo and surv records with each other as well as those from previous years to construct our spatial dataset of reproductive fitness in the tallgrass prairies of our study area.

Total Demo
Bill Thom’s Gate, Common Garden, Dog, East of Town Hall, Golf Course, Hegg Lake, Martinson’s Approach, Nessman, North of Golf Course, REL, RHE, RHP, RHS, RHX, RKE, RKW, Randt, Railroad Crossing Douglas County, South of Golf Course, Sign, Town Hall, Tower, Transplant Plot, West of Aanenson, Woody’s, Yellow Orchid Hill

Annual Sample
Aanenson, Around Landfill, East Elk Lake Road, East Riley, KJ’s, Krusmark’s , Loeffler’s Corner, Landfill, North of Railroad Crossing, Norwest of Landfill and North of Northwest of Landfill (lumped,) On 27, Riley, Railroad Crossing, Steven’s Approach, Staffanson Prairie

In addition to the annual collection of data, this year Erin began developing a study to investigate whether flowering return interval and isolation by distance are correlated in Echinacea.

Start year: 1995

Location: Unbroken (never tilled) remnant prairies in Douglas County, MN, located along roadsides, nature preserves, railroad right-of-ways and privately-owned land.

Overlaps with: Flowering phenology in remnants, fire and flowering at SPP, reproductive fitness in remnants, EA fire & fitness

Data: Access the most recent copies of allDemoDemo.RData and allSurv.RData at ~Dropbox\demapSupplements\demapInputFiles. Demap accepts these files and the demap team will clean and reconcile them in the demap repository. ~Dropbox\geospatialDataBackup2019 houses the raw GPS jobs while the aiisummer2019 repo houses the raw demo records.

Data related to the flowering isolation study can be found in the floiso repository (contact Erin Eichenberger for access) and in the folder ~Dropbox\floweringIsolation2019

Products: Amy Dykstra’s dissertation included matrix projection modeling using demographic data

Project “demap” merges phenological, spatial and demographic data for remnant plants

You can read more about the demographic census in the remnants, as well as links to prior flog entries mentioning the experiment, on the background page for this experiment.

2019 Update: Common Garden Experiment

Since 1996, members of Team Echinacea have walked, crawled, and ~sometimes~ run next to rows of Echinacea angustifolia planted in common garden experiments. Although protocol varies depending on the common garden, every year team members record flowering phenology data, measuring data, and harvest the heads of the thousands of plants we have in common garden experiments. The Echinacea Project currently has 10 established experimental plots: exPts1-10. Due to the repetitiveness of yearly phenology, measuring, and harvesting, this project status report will include updates on all common garden experiments except for Amy Dykstra’s plot (exPt3), qgen2qgen3 (exPt8), and the West Central Area common garden (exPt10).

exPt1: Experimental plot 1 was first planted in 1996 (cleverly termed the 1996 cohort), and has been planted numerous times in subsequent years, with the most recent planting being inbreeding 2. It is the largest of the experimental plots, with over 10,000 planted positions; experiments in the plot include testing fitness differences between remnants (1996, 1997, 1999) effects of inbreeding (inb1, inb2), and quantitative genetics experiments (qgen1). There are also a number of smaller experiments in it, including fitness of Hesperostipa spartea, aphid addition/exclusion, and pollen addition/exclusion. In 2019, we visited 4392 of the original 10,622 planted and found 3486 alive. Only 70 plants were classified as “flowering” in exPt1 this year. This is a drastic change from the nearly 1000 plants that flowered in summer 2018 – perhaps it is a testament to the benefits of controlled burning (we burned in spring 2018 but not in 2019). In summer 2019, we harvested 52 total Echinacea heads in exPt1. In the fall, we added 789 staples to positions where plants were gone for three straight years – this was desperately needed because no staples were added to positions with dead plants after 2018.

exPt2: Heritability of flowering time is the name of the game in exPt2. Planted in 2006, exPt2 was planted to assess if flowering start date and duration was heritable in Echinacea. In summer 2019, we visited 2050 positions of the 3961 positions originally planted. We measured 1802 living plants, of which 654 were flowering. In the fall, we harvested ~1100 heads from exPt2. We do not have an exact number of heads harvested from exPt2 yet, as we have not had time to complete head reconciliation. Location: Hegg Lake WMA

Stuart explains Echinacea capitulum morphology to Shea in one of the experimental plots.

exPt4: Experimental plot 4 was planted to determine if Echinacea from small remnant populations could be genetically rescued via an outcross to larger, more genetically diverse populations. Caroline Ridley and other members of Team Echinacea sowed 3584 achenes at Hegg Lake WMA in 2008, and we have assessed survival and basal plant characters every year since. Survival in exPt4 is incredibly low. We only visited 21 plants in 2019 and only 7 were alive. No plants have flowered in this plot yet. Location: Hegg Lake WMA

exPt5: The only experimental plot planted at Staffanson Prairie Preserve (SPP), exPt5, was planted in an attempt to compare progeny of maternal plants from burned and unburned sections of SPP. There were 2800 plants planted originally, but high mortality made it impossible to visit the plot row-by-row. Now, we and treat the plot like demography. We use a GPS to find plants in exPt5 that have previously flowered and add more plants to the stake file if new plants in the plot flower. In 2019, we visited 10 plants in the plot, all of which were alive! There were no plants flowering in exPt5 in 2019, though. Location: Staffanson Prairie Preserve

exPt6: Experimental plot 6 was the first E. angustifolia x E. pallida hybrid plot planted by Team Echinacea. A total of 66 Echinacea hybrids were originally planted; all have E. angustifolia dams and E. pallida sires. In 2019, we visited 40 positions and found 28 living plants. No plants have flowered in this plot yet. As of January 2020, all exPt6 measure data through summer 2019 is uploaded to the SQL database. Location: near exPt8

exPt7: Planted in 2013, experimental plot 7 was the second E. pallida x E. angustifolia plot. It contains conspecific crosses of each species as well as reciprocal hybrids. There were 294 plants planted. Out of the 205 plants we visited in 2019, we found 161 plants still alive and basal; there were no flowering plants this year. For some context, survival of pure E. angustifolia crosses was lower than all other cross types. As of January 2020, all exPt7 measure data through summer 2019 is uploaded to the SQL database. Location: Hegg Lake WMA

exPt9: Experimental plot 9 is another hybrid plot, but unlike the other two hybrid plots, we do not have a perfect pedigree of the plants. That is because E. angustifolia and E. pallida maternal plants used to generate seedlings for exPt9 were open-pollinated. We need to do paternity analysis to find the true hybrid nature of these crosses (assuming there are any hybrids). There were originally 745 seedlings planted in exPt9, and in 2019 we visited 510 positions. It was one of the harder plots to measure because over half of the positions did not have a plant and we do not use staples at Hegg Lake WMA. We found 308 living plants in 2019, one of which was flowering! We know the flowering plant has an E. pallida mother, but we are still unsure of the paternity of the flowering plant. When we know, we will post an update. As of January 2020, all exPt9 measure data through summer 2019 is uploaded to the SQL database. Location: Hegg Lake WMA

Summer 2019 measures exPt9… so many “Can’t Finds!”

For more information on survival in common garden experiments, see this flog post about survival in common gardens.

Start year: Various, see individual listings above. First ever planting was 1996.

Location: Various, see above

Overlaps with: Pretty much everything we do

Data/ materials collected: Measure data for all plots. All raw measure data available in cgData repository. Processed data should eventually be available in SQL database; ask GK for status of SQL database. GPS points were shot for the exPt9 flowering plant, as well as for all surviving plants in exPt4. Find the GPS jobs containing the exPt4 and exPt9 points in ~Dropbox\geospatialDataBackup2019, saved in three formats in temporaryDarwBackups2019, convertedXML2019 and convertedASVandCSV2019. The job name is SURV_20191002_DARW and the points have names that distinguish them by the experimental plot. The stake file to find exPt5 plants is here: ~Dropbox\geospatialDataBackup2019\stakeFiles2019\exPtFiles\exPt05stakeFile.csv

Products: Many publications and independent projects.

2019 Update: Experimental plot management

This summer Jay Fordham conducted an experiment in experimental plot 8 to determine an effective method for eradicating Fraxinus pennsylvanica, or green ash. Team Echinacea is concerned with the spread of green ash in our experimental plots because it crowds out native herbs. The Echinacea in exPt8 may be at particular risk due to their young age relative to individuals in other plots. A prior attempt to manage ash in exPt8 with triclopyr (brand name Garlon) largely failed, resulting in only 3% mortality.

Jay devised three treatments of  triclopyr application to green ash. The three treatments were 1) A foliar application where he painted all leaves with triclopyr; 2) A bark application where he cut each ash 10cm from the base and applied triclopyr to the remaining above-ground stem; and 3) A cambium application where he cut each ash 10cm from the base, scraped off the exterior bark with a knife, and applied triclopyr to the remaining above-ground stem. He divided exPt8 into 35 treatment sections and randomly assigned a treatment to each section. He then randomized the treatment application order and, with the help of the team, treated 438 green ash trees from July 22nd to August 8th. Jay then assessed ash mortality on August 27th and 28th and found the cambium application to be most effective. Jay presented his findings at the 2019 Midstates Undergraduate Research Symposium in St. Louis on November 1-2. His presentation is available to view and download here.

Triclopyr treatment Mean Proportion Dead Stems
Foliar application 0.005
Cutting and bark application 0.333
Cutting and cambium application 0.498

In addition to the green ash management experiment in exPt8, the team removed Bird’s-foot trefoil from exPt1 and along the bordering road. The team also removed sweet clover from within and around exPt1, exPt2 and exPt3. The team cut back sumac from the easternmost rows of exPt1.

Jay brandishing sweet clover plants that the team pulled in exPt1

In June the team planted Asclepias viridiflora in exPt1 at regular intervals. Stuart initially assessed approximately 124 surviving plugs prior to planting. Erin and Riley, while pulling flags marking the planting locations in September, did not observe any surviving milkweed plants. The team also planted Carex gravida and Carex brevior in the path around exPt1. The team planted three of the same carex species at each location in a triangular configuration. Erin shot the planting positions with the GPS pole in the center of the three plants, or between two where two survived, or north of a single plant where one survived. In October she observed that approximately 2/3rds of the carex plantings were present.

Location: exPts 1, 2 and 8

Data/materials collected: Weeds were discarded outside the plot as they were removed.

Find information about Jay’s experiment at ~Dropbox\teamEchinacea2019\jayFordham

Find information about the planting locations of the Asclepias viridiflora at ~Dropbox\CGData\Asclepias\plantPlugs2019.csv

Find the two GPS jobs containing the carex locations in ~Dropbox\geospatialDataBackup2019, saved in three formats in temporaryDarwBackups2019, convertedXML2019 and convertedASVandCSV2019. The job names are CAREX_P1_20190801_DARW and CAREX_P1_20191003_DARW.

2019 Update: Pollen Interference

The pollen interference experiment series examines the influence of heterospecific pollen application on the shriveling of Echinacea styles. The shriveling of Echinacea styles has previously been considered a signal of compatible pollen receipt, but shriveling can also occur after the application of pollen from other, closely related species without the creation of a viable seed. The pollen interference project aims to determine which species successfully cause shrinkage of Echinacea styles and whether this interference occurs because of an interspecies match in morphology or interparental genetic compatibility. 

During the 2019 summer field season, Julie Bailard performed hand-crosses with pollen isolated from individuals of either Heliopsis helianthoides or Ratibida pinnata on styles of Echinacea angustifolia. The goal of this experiment was to determine if heterospecific pollination of Echinacea with closely-related asters resulted in style shriveling. Pollen from each pollen donor was collected in separate microfuge tubes and applied to Echinacea styles the same day, using a fresh toothpick for each donor. In total, 19 Heliopsis and 8 Ratibida sires were crossed with 16 Echinacea dames for a total of 96 interspecies crosses. In 2009, Allegra Halverson applied pollen from Heliopsis helianthoides, Coreopsis, and Carduusonto Echinacea styles and observed that application of Heliopsis pollen often preceded successful style shriveling. 

Start year: 2009

Location: exPt02

Data/ materials collected: The data are composed of binary outcomes (y or n) representing either successful style shriveling or style persistence for each pollinated style, recorded 24 and 48 hours after initial heterospecific pollen. Each style’s records are paired with the identity of its maternal plant and the species (application 2009 and 2019) and individual identity code (2019) of the heterospecific pollen donor. Datasheets are found in Julie Bailard’s teamEchinacea2019 Dropbox folder in.

Previously worked on by: Allegra Halverson (Allegra’s pollination dataflog entry)