Evaluating the Suitability of Roadway Corridors for Use by Monarch Butterflies (2020)

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29 Background The protocol includes information on road type, adjacent land use, management practices, forb species richness and percent cover, noxious weed presence and percent cover, and milk- weed species richness and abundance (Table 3) (field data sheet and protocol instructions pro- vided in Appendix C). The research team developed both a paper data sheet and an electronic data form that could be filled using a tablet or smartphone in the field. The RA was designed in Survey123 for ArcGIS (Esri), which affords several benefits for road- side management authorities. States or other entities can collect, manage, and view their own datasets using their own Esri Enterprise account. The RA is installed within each agency’s ArcGIS Online platform. It comes pre-populated with a comprehensive nectar plant list for their state (derived from the USDA plants database). Then managers may customize their assessment by selecting the noxious weeds they wish to track and set default answers regarding herbicide use and mowing practices, if desired. Within their own Survey123 website, transportation managers can view site locations, field data, and monarch habitat quality scores derived from data collected using the RAs. The electronic form of the RA provides the field user advantages such as the ability to auto- matically record the location, date, and time of the assessment. The survey also provides features such as a searchable drop-down list of plant species that enables one to type in letters from either the common name or the Latin name to select the species. There are also choices based on genera, such as “Solidago/goldenrod species” to lump plant species that are difficult to distin- guish. The assessment is flexible in that observers may also tally plant types they cannot identify and choose to estimate milkweed plant abundance in categories rather than count individual plants (e.g., depending on the abundance of the milkweed and time constraints). The researchers incorporated several factors identified as important to roadside managers, including the need to assess sites quickly and once per growing or monarch breeding season, the ability to specify weeds of local or state importance, and the ability to specify the width of the area to be surveyed with regard to mowed areas, which are described subsequently. Given the strong preference of roadside managers for a protocol that could adequately charac- terize the habitat quality of a site in a single visit per year, the research team required a proxy for the availability of nectar throughout the growing season. The team defined a term “potentially blooming nectar plants” (hereafter “nectar plants”) to describe forbs and shrubs that could provide nectar to pollinators (e.g., excluding grasses), whether or not blooming on the date of assessment. This broad categorization encompasses plants that may provide nectar, regardless of their nativity or the amount or quality of nectar they may provide. The number of nectar plant species may be important because a greater number of species may represent a greater number of bloom times and thereby provision nectar for a greater proportion of a season of monarch C H A P T E R 4 Product B: Rapid Assessment of Roadside Habitat for Monarchs

30 Evaluating the Suitability of Roadway Corridors for Use by Monarch Butterflies Habitat Component Significance Measure Categories Road Exposure to collisions, road salt, and chemicals from cars Road type 2 lane, 4 lane, > 4 lane Landscape Exposure to pesticides, proximity to existing habitat Adjacent land use type (within 100 ft) CROP1, HCR, DEV, HDE, WOOD, DIV, NDI, WET Milkweed Required host plants for monarch eggs and larvae Milkweed abundance (count plants or choose category) 0, 1–5, 6–10, 11–25, 26– 50, 51–250, >250 Species richness may increase seasonal availability Milkweed number of species Nectar Required for adult monarch foraging Potential blooming nectar plant2 % cover 0%, 1–5%, 6–10%, 11– 25%, 26–50%, 51–75%, >75% Species richness increases seasonal availability of nectar Potential blooming nectar plant number of species Native species may have higher resilience, sustainability, and provide habitat to pollinators and other native organisms Native potential blooming nectar plant number of species Weeds3 Threatens native milkweed and nectar plants; may require management that could temporarily remove habitat Weed % Cover 0%, 1–5%, 6–10%, 11– 25%, 26–50%, 51–75%, >75% Greater species richness of weeds may require more and/or multiple control effort(s) Weed number of species Herbicide Use Frequency of use Herbicide use on site None, spot treat noxious weeds, spot treat woody species, treat grass to stimulate forbs, broadleaf applied in clear zone 1x/yr; broadleaf applied in clear zone >1x/yr; broadleaf applied throughout the ROW4 Mowing Mowing, at least temporarily, reduces nectar availability and destroys eggs and larvae; the width of frequently mowed areas impacts the amount of available habitat Mowed width (ft) Frequent mowing of the full width of the ROW reduces nectar availability and survival of egg and larval monarchs Frequency of mowing full ROW width Never, every few years, 1x/yr, 2x/yr, >2x/yr, don't know 1. CROP = cropland, no barrier; HCR = Crop with woody barrier or hedgerow; DEV = Developed, lawn or paved; HDE = Developed with woody barrier or hedgerow; DIV = Diverse grassland/natural habitat; NDI = Not diverse grassland with few forbs; WOOD = Woody habitat; WET = Wetland habitat 2. Potential blooming nectar plants are forbs and shrubs that can provide nectar for monarchs or other pollinators, whether or not blooming on the survey date. 3. Weeds are defined to be of management interest by the transportation authority; may include noxious weeds and other invasive species under active surveillance or management. 4. ROW = right-of-way Table 3. Habitat components (and data collected) for roadside ROW habitats using the RA.

Product B: Rapid Assessment of Roadside Habitat for Monarchs 31 use or use by other pollinators. The researchers identified plants to species when possible and also estimated the aerial percent cover of nectar plants as a group. To make the protocol usable for people with varying skills in identifying plants to species, an option was included for tallying unidentified types of plants. To accommodate variation in the list of invasive species, weeds, or non-native species of management concern from state to state, the team created a customizable weed list. When transportation managers initially set up the protocol for their organization, they can populate a custom list of weed species they want to include in the assessment. Weeds of interest can be specified for each field assessment if important weed species vary across the jurisdiction. Observers will then report whenever those species are present on the assessment areas and estimate aerial cover for those species as a group to describe their prevalence. A survey of roadside managers indicated that the frequency and widths of mowing in the ROWs were highly variable; mowing the full ROW width was done multiple times per growing season to sometimes only once every several years. Some mow a safety strip (e.g., first 10–12 feet) monthly during the growing season, while others mow the strip only once per year (and some do not mow from May to July for wildlife and pollinators). Furthermore, some roadside managers expressed interest in using the RA to gain information about the effects of their mowing prac- tices on pollinator habitat. Mowing is needed to maintain safety strips along the road margins and also may be used to control woody and invasive species beyond the safety strip. Reduced mowing during the monarch breeding season is recommended to reduce direct mortality for monarch eggs and caterpillars and to preserve more plant blooms as nectar sources (Monarch Joint Venture 2017). Reduced mowing also can be associated with greater species richness of blooming nectar plants (Halbritter et al. 2015). However, mowing can also stimulate growth of new milkweed leaves preferred by egg-laying monarchs (Alcock et al. 2016, Baum and Mueller 2015, Fischer et al. 2015). In 2017, the research team collected data across the entire ROW. In 2018, after recognizing how difficult it is to assess vegetation in safety strips if they are frequently mowed (e.g., monthly or more), the team collected data from the unmowed area. Ultimately, to accommodate regional variation in mowing practices, managers or surveyors were allowed to choose whether to conduct their assessments in full ROWs, unmowed areas, or in mowed and unmowed areas separately. Finally, because some departments of transportation were interested in monarch breeding activity in their roadside areas, the researchers included optional data fields for recording monarch eggs, larvae, and adults. This section also includes a place to record the species and number of milkweed plants searched. Rapid Assessment Protocol RAs are completed for a 45.7-m (150-ft) length of roadway, implemented at random loca- tions or systematically (e.g., every mile or 10 miles) in a road system (see protocol in Appen- dix C). They may be combined to depict average conditions or to compare habitat quality among sites. Alternatively, sites may be selected because they are of interest to the observer, such as construction, restoration or remnant native habitat areas. Upon arrival at a location of interest, the observer walks parallel to the road, toward traffic, pacing the 150 ft of distance (Figure 13). Next, the width of the vegetated ROW (perpendicular to the road) is estimated (e.g., paced). These two distances bound the rectangular assessment area that extends from the road to the back of the ROW. The observer walks back through the ROW to the starting point, systematically zigzagging back and forth throughout the roadside habitat, while record- ing data. The observer records the number of milkweed plants by species, where stems sepa- rated by soil are counted as plants regardless of whether they are clonal or genetic individuals

32 Evaluating the Suitability of Roadway Corridors for Use by Monarch Butterflies (following Kasten et al. 2016 and CEC 2017), the species or number of nectar plants (also notes for each species if it is blooming or not), and the presence of weeds (as defined by their roadside organization). Percent aerial cover is also estimated by classes for nectar plant species collectively and for weeds of concern. The observer records the dominant adjacent land use, mowing, and herbicide application information. As an option, observers may examine milkweed plants by species for monarch eggs and larvae, recording the number of plants searched and number of eggs and larvae detected. To maintain efficiency when milkweed is abundant, observers may choose to monitor every 2nd, 3rd, or 5th milkweed plant encountered to gain a sample size of 50–100 milkweed plants searched. Integrated Monarch Monitoring Program Methods IMMP sampling employs a total of 100 quadrats placed along ten transects arrayed diago- nally from the road edge to the back of the ROW along a 400- to 500-m length of roadway (see Figure 14). Transects are 50 meters in length and quadrats are placed every 5 m (however, in 2017, researchers placed quadrats every 2 m along 25-m transects, with 25 m between each transect). Quadrats consist of a 1.0 m by 0.5 m sampling frame placed to either side of the transect line for a 2.0 m by 0.5 m or 1-m2 quadrat area. Within each quadrat, observers count milkweed plants (same definition as above) to estimate milkweed density (milkweed plants/ha). All blooming plants are identified to species and assigned to the first subplot (area within the quadrat) in which they occur (first 0.5 × 0.5 m, 1.0 × 0.5 m, or 2.0 × 0.5 m) to generate a fre- quency score (proportion of subplots occupied; not presented in this paper). Plants that are not blooming on the date of the assessment are not recorded. Figure 13. Depiction of a Rapid Assessment, showing how an observer moves 150 ft from a starting point along a roadside, systematically zigzagging throughout the ROW to characterize habitat conditions. Figure 14. Overlay of RA and IMMP for the comparison of protocols. The IMMP uses ten 50-m-long transects arrayed diagonally 400–500 m along the roadway. In comparison trials, 2–4 RAs were completed for each IMMP site, typically established at the ends and middle of each IMMP site.

Product B: Rapid Assessment of Roadside Habitat for Monarchs 33 To account for the different sizes of the survey areas for each protocol, at each of these sites, the research team completed one IMMP survey and typically three RAs spaced 200–250 m apart within the footprint of the IMMP site (Figure 14). One site in 2017 had four RAs and one site had only two; in 2018 three sites had only two RAs. Field Trials The RA was tested in Minnesota and Oklahoma. In Minnesota, the research team collected comparative data to allow the RA to be compared to the IMMP and to address specific ques- tions about the interaction between landscape and roadside habitat and use by monarchs. In Oklahoma, the team focused on conducting a statewide survey, including evaluating the contributions of single or clustered sites. Minnesota—Protocol Comparison For 2017 field trials, the research team chose 14 sites from a set of randomly selected roadside sites in Minnesota that had been surveyed for milkweed and monarchs and contained milkweed in 2015 (Kasten et al. 2016; Figure 15). In 2018, 15 new sites were selected through the IMMP, which uses generalized random tessellated stratified sampling (GRTS) to identify random 10 × 10 km blocks and random point locations within them stratified by land-use sector and pri- oritized to accommodate for variable inclusion probability (Cariveau et al. 2019b). Sites in 2018 were randomly selected using the GRTS list of point locations; 13 sites were within the 15 highest ranked blocks in Minnesota (with vegetated roadsides at least 4 m wide) plus two additional sites within the 25 highest ranked blocks, for a total of 15 sites. Sites in both years represented variation in roadway types (except freeways, which were excluded due to safety concerns). Figure 15. Twenty-nine field sampling locations in Minnesota where RAs and IMMP protocols were compared (Kasten et al. 2016).

34 Evaluating the Suitability of Roadway Corridors for Use by Monarch Butterflies Minnesota—Broadscale Surveys After refining the protocols based on the 2017 season, in 2018 the researchers sent a field crew to locations throughout Minnesota to run RAs. There were several primary objectives in the 2018 season. The primary objective was to determine how the habitat quality scores would be distributed through the RA protocol. To augment the Minnesota surveys, the team also field-tested the protocol in several states and in cooperation with DOTs in order to incorporate their reviews. The team also addressed research questions regarding the interaction between landscape and roadside habitat and use by monarchs as follows: • On sites that were adjacent to natural lands (i.e., not developed, not agriculture), and closer to habitat core areas, – Was milkweed more abundant? – Were the metrics representing nectar plants (potentially blooming plant species richness, native species richness, cover by potentially blooming plants, or blooming cover) higher? – Was cover by noxious weeds lower? – Was monarch use greater? In terms of distance to habitat core areas, did use of sites differ among close, near, and far? • For all four metrics above, were there observable differences when comparing sites adjacent to developed, agricultural, and natural land-use types? • Was density of milkweed or nectar plant indices inversely related to percent cover by noxious weeds? • Were Habitat Calculator scores correlated to scores from the Landscape Prioritization Model? The researchers created a stratified-random process for selecting field sites to address ques- tions about the relative importance of landscape context versus on-site vegetation characteristics in terms of predicting milkweed availability and breeding use by monarchs. The team drew a sample of 15 sites in each of the categories below (36 categories) for a total of 520 samples. The sample of random sites was selected from a factorial of these types of roads: • Road Types/ROW Widths – Local (2 lane; < 10 meters wide) – Moderate (2–4 lane; 10–20 meters wide) – Interstate (4+ lanes; > 20 meters wide)—subject to permissions • Adjacent Land Use – Cropland – Developed – Grassland – Wooded • Distance from Core Habitat Areas – Mainland (Adjacent to core, within 1 km of core) – Stepping Stone (intermediate distance from core area) – Island (isolated from core area by at least 10 or 15 km) Core metrics were: milkweed density, nectar plant species richness, percent cover by poten- tially blooming plants, percent of area covered by blooms, percent cover by native blooming plants; percent cover by noxious weeds, and use by monarchs. Statistical Analyses The researchers calculated milkweed plants/ha based on the number of milkweed plants counted (all species combined) and the area searched at each site and converted to hectares.

Product B: Rapid Assessment of Roadside Habitat for Monarchs 35 For the IMMP, the area searched was 100 m2 based on the 100 1-m2 quadrats. For the RA, the area searched was estimated as 45.7 m (the length of the plot) multiplied by the ROW width. The team presented monarchs/plant as the sum of all monarch eggs and larvae observed, divided by the number of milkweed plants searched. For the IMMP protocol, the number of milkweed plants searched differed from the number of milkweed plants in the density estimate, because observers could search additional milkweed plants between the quadrats to look for monarch eggs and larvae. The team focused analyses on sites with at least ten milkweed plants to ensure robustness of the density statistics (larvae/plant). The team also estimated monarchs/ha by multiplying the average number of monarchs/plant times the average number of milkweed plants/ha using the IMMP method. The research team presented the number of blooming species to represent species richness of blooming plants as an index of nectar resource availability. For the IMMP protocol, this is a list of all blooming species encountered in the quadrats. For RAs in 2017, the team listed all of the blooming plant species encountered; in 2018, all of the potentially blooming nectar plants were identified and noted whether or not plants were blooming. Here, the team presented the blooming subset in comparison to the IMMP data. The nectar plant species lists across the several RAs for each IMMP site were combined in two ways. First, the number of blooming species was determined for each RA, and then the number was averaged across the several RA for each IMMP survey location; this is called RA averaged. Second, because of known relation- ships between species richness and area, the team also depicted the number of blooming species determined when summing the species across the RAs for each IMMP site (removing duplicates), which is called RA summed. The team computed statistics using R version 3.5.1 (R Core Team 2018). For milkweed plants/ha and monarchs/plant, the team compared the mean of the 2–4 RAs to the IMMP measure for each site. To determine if protocol type had a significant effect on response vari- ables, the team ran generalized linear mixed models with year and protocol type as fixed effects and site as a random effect for each of the response variables of milkweed density, monarchs/ plant, and number of blooming species (“nlme” package; Pinheiro et al. 2018). The team reported an interaction term for year and protocol type when significant. The sample size was 113 visits to 29 sites for the plant data; because the team found no milkweed plants dur- ing 17 visits, the model for monarchs per plant contained 96 visits to 29 sites. For number of blooming species, the researchers compared the estimates by the IMMP protocol to the RA averaged and RA summed in a generalized linear mixed model with year and protocol type as fixed effects, site as a random effect, and a year by protocol type interaction effect. For clarity, researchers also compared the numbers of blooming species by the IMMP protocol to the RA averaged and RA summed for each year separately. The researchers also compared the mean of the RAs per IMMP site to the IMMP measure with a Kendall Rank Correlation for each of the same response variables. Data were plotted in Excel and ggplot2 (Wickham 2016). Field Trials—Oklahoma In 2018, from May 28 through June 18, 143 sites were visited across Oklahoma with every ecoregion represented (Figure 16). The selected sites were a subset of those from a 2016 statewide milkweed survey. At each site, either a single survey was conducted (73 sites) or three surveys were conducted approximately 500 m apart (70 sites). A subset of sites was revisited in the Cross Timbers and Central Great Plains ecoregions in July and August 2018. This subset consisted of nine sites with three surveys and 11 sites with single surveys for a total of 38 surveys. At each site the team implemented the RA protocol, which included recording information about road type, mowed width, ROW width, and adjacent land use. The team also estimated

36 Evaluating the Suitability of Roadway Corridors for Use by Monarch Butterflies overall forb cover, flowering plant cover, and noxious weed cover based on the following categories: <5%, 6–25%, 26–50%, 51–75%, 76–95%, or >96%. The number of milkweed plants was also estimated in categories, including 0, 1–10, 11–50, 51–100, 101–500, or >500 plants for each species present at a site. The team also recorded all forbs present at a site and indicated if they were flowering or not. The total time to complete a single survey, without including travel time, averaged 9 min- utes (range: 0–39 min) (Figure 17a). The total time to complete a set of three surveys (located approximately 500 m apart), including travel time between sites, averaged 45 minutes (or 15 minutes per single survey; range: 27–88 min). The average ROW width (calculated as mowed width (i.e., safety zone width) + ROW width) was smaller for two-lane roads (10.5–201 ft; safety zone: 16.5 ft) than four-lane roads (17–348 ft; safety zone: 20.7 ft) (Figure 17b). Seven species of milkweed were recorded during surveys, with green antelopehorn (Asclepias viridis) being the most common (40.93% of sites) (Figure 18). Clasping milkweed (A. amplexicaulis) and Englemann’s milkweed (A. englemanniana) were the least commonly encountered milkweed species (present in <1% of surveys). No milkweed plants were observed in 53.74% of surveys. Figure 16. Map of Oklahoma with 143 visited sites indicated by the white and black circles. Sites were visited from 28 May through 18 June 2018. All 12 ecoregions, represented by color, were sampled in the full dataset. Solid black circles indicate a subset of 20 sites revisited in July and August 2018 within the Cross Timbers and Central Great Plains ecoregions.

Product B: Rapid Assessment of Roadside Habitat for Monarchs 37 53.74 40.93 3.91 3.56 2.14 1.42 1.07 0.36 0.36 0.00 10.00 20.00 30.00 40.00 50.00 60.00 NONE ASCVIR ASCASP ASCVIF ASCTUB ASCLAT ASCSTE ASCAMP ASCENG Pe rc en t o f S ur ve ys w ith M ilk w ee d Pr es en t Milkweed Species Figure 18. Percent of each milkweed species observed during the survey. Milkweed codes are defined in Table 4. 0:00 0:07 0:14 0:21 0:28 0:36 0:43 0:50 0:57 single triple A ve ra ge L en gt h of S ur ve y (m in ) Type of Survey 0 10 20 30 40 50 60 70 80 90 2 lane 4 lane A ve ra ge R O W W id th (ft ) Type of Road a b Figure 17. a) Average amount of time (min = minutes) to complete a single survey and triple surveys (located approximately 500 m apart). b) Average ROW width (ft) for two-lane roads and four-lane roads.` Species Common name Code Asclepias viridis green antelopehorn ASCVIR Asclepias asperula spider milkweed (antelopehorns) ASCASP Asclepias viridiflora green comet milkweed ASCVIF Asclepias tuberosa butterfly milkweed ASCTUB Asclepias latifolia broadleaf milkweed ASCLAT Asclepias stenophylla slimleaf milkweed ASCSTE Asclepias amplexicaulis clasping or blunt leaved ASCAMP Asclepias engelmanniana Engelmann's milkweed ASCENG Table 4. Milkweed species encountered during surveys with common names and codes.

38 Evaluating the Suitability of Roadway Corridors for Use by Monarch Butterflies Milkweed species richness was highest in roadsides adjacent to diverse grasslands and low- est in roadsides adjacent to cropland regardless of road type (Figure 19). However, the differ- ence in species richness between the highest and lowest sites was only one additional species. When comparing the number of milkweed species observed among triple surveys, 40 sites had a different number of milkweed species among surveys, while only eight sites had the same number of milkweed species among surveys. Thus, conducting triple compared to single surveys increased the number of milkweed species observed in approximately 83% sites. Average forb species richness ranged from 7.5–14 species. The most common forb species encountered during surveys were fleabane species (Erigeron spp.) followed by green antelope- horn (A. viridis) (Table 5). Four non-native forb species were frequently recorded, including annual yellow sweetclover (Melilotus indicus), yellow salsify (Tragopogon dubius), prickly lettuce (Lactuca serriola), and curly dock (Rumex crispus). Field Surveys with Multiple Protocols In 2017, 14 sites were assessed between June 29 and August 22. All sites were located along paved roads, 11 along two-lane roads, and three along four-lane roads. Eight sites were adjacent to cropland, with two sites each by woodland, grassland, and developed land. ROW widths from the RAs varied from 3 m to 21.5 m (mean = 12.35 m, standard deviation (SD) = 3.71); widths were not recorded by the IMMP protocol in 2017. In 2018, 15 sites were surveyed between July 23 and August 29; all sampled sites were along two-lane roads; 12 were paved; three were dirt/gravel. In 2018, adjacent land uses included: crop- land (7), woodland (3), grassland (2), and wetland (3). The widths of the ROWs by RAs varied from 5 to 52 m (mean = 14.07, SD = 12.79). The average width of the ROWs in 2018 recorded by the IMMP was 9.43 m (SD = 3.70, range 3.5–19.5 m). A single RA took an average of 22 minutes in 2017 (SD = 15 minutes; range 4–88 min) and 20 minutes in 2018 (SD = 12 minutes; range 5–59 minutes). IMMP visits took 134 min- utes on average (SD = 67 minutes; range 68–345 minutes) in 2017 and 167 minutes in 2018 0 0.5 1 1.5 2 2.5 CROP DEV DIV NDI WET WOOD CROP DEV DIV NDI OTH WET WOOD 4 lanes 2 lanes Av er ag e M ilk w ee d Sp ec ie s Ri ch ne ss Road Type and Adjacent Land Use Figure 19. Average milkweed species richness by adjacent land use and road type. CROP = agricultural land, DEV = developed/industrialized land, DIV = diverse forb grassland, NDI = non-diverse forb grassland, WET = wetlands and/or water bodies, WOOD = prominently wooded/forested areas, OTH = other or not recorded.

Product B: Rapid Assessment of Roadside Habitat for Monarchs 39 (SD = 56 minutes; range 92–274 minutes). Variations in the duration of visits was likely related to the number of nectar plant species present and the number of milkweed plants counted and examined for monarch eggs and larvae. Milkweed Density Milkweed was detected at all sites in 2017 and 14 of the 15 sites (93%) in 2018 using the IMMP protocol. The vast majority of milkweed was Asclepias syriaca (common milkweed, 96%); other species were A. incarnata (swamp milkweed, 3%), A. verticillata (whorled milk- weed, 0.69%), A. sullivantii (Sullivant’s milkweed, 0.2%), and A. tuberosa (butterfly weed, 0.01%). The mean milkweed density for all species of milkweed combined using the IMMP protocol was 1,242 plants/ha (SD = 1,303) in 2017, 2,807 plants/ha (SD = 4,864) in 2018, and for both years combined, 2,052 plants/ha (SD = 3639; median = 800; range 0–18,000) (Fig- ure 20a). Averaging the RAs per site, the mean milkweed density for all species of milkweed across sites in 2017 was 1,508 plants/ha (SD = 2,082), 1,545 plants/ha (SD = 2,377) in 2018, and 1,527 plants/ha for years combined (SD = 2,199; median = 625; range 0–8966). Milkweed density did not vary with year (t27 = 0.415, p = 0.681) or survey type (t83 = –0.639; p = 0.524, df = 83). Milkweed density as estimated by the two protocols was correlated (Kendall’s rank correlation tau = 0.568, z = 4.257, df = 27, p < 0.001). See Figure 21a. Monarch Eggs and Larvae The mean number of milkweed plants searched for monarch eggs and larvae in 2017 was 40.93 (SD = 47.66) with the IMMP and 76.11 (SD = 91.15) with the RA. In 2018 the mean num- ber of milkweed plants searched for monarch eggs and larvae was 113 (SD = 134.48) with the IMMP and 36.27 (SD = 44.38) with the RA. In 2017, using the IMMP method, monarch eggs or larvae were found at six of 14 sites (43%); the RA monarch eggs or larvae were found at seven of 14 sites (50%). In 2018, using the IMMP method or the RA, monarch eggs or larvae were found in 11 of 15 sites (73%) or in 11 of 14 sites containing milkweed (79%). If considering RAs independently from one another, then in 2017, monarch eggs or larvae were found in 11 of # Survey Locations Code | forb species | common name Native? 126 Other - Erigeron spp.| fleabanes Yes 114 ASVI2| Asclepias viridis | green antelopehorn Yes 99 MEIN2| Melilotus indicus | annual yellow sweetclover No 95 ACHIL| Achillea | yarrow 92 Other - Cirsium spp. | thistles 83 MINU6| Mimosa nuttallii | Nuttall's sensitive-briar Yes 80 TRDU| Tragopogon dubius | yellow salsify No 78 DAPU3| Daucus pusillus | American wild carrot Yes 75 LASE| Lactuca serriola | prickly lettuce No 74 COCA5| Conyza canadensis | Canadian horseweed Yes 72 SOEL| Solanum elaeagnifolium | silverleaf nightshade Yes 71 Unknown 61 RUHI2| Rudbeckia hirta | blackeyed Susan Yes 58 Other - Plantago spp. | plantains 54 RUCR| Rumex crispus | curly dock No 51 SOLAN| Solanum | nightshade 50 HELIA3| Helianthus | sunflower Yes Table 5. The most frequently encountered forb species during surveys. The native status of each forb is included.

40 Evaluating the Suitability of Roadway Corridors for Use by Monarch Butterflies 42 (26%) RAs or 11 of 37 (30%) sites with milkweed. In 2018, monarch eggs or larvae were found in 19 of 42 (45%) RAs or 19 of 30 (63%) sites with milkweed. When restricting analysis to sites with at least ten milkweed plants examined by each protocol (10 sites), in 2017, monarch egg or larvae were found at 40% of the sites with the IMMP protocol and 50% with the RA protocol (summed per site). In 2018 (n = 11 sites with at least ten milk- weed plants examined by each protocol), monarch eggs or larvae were found at 82% of the sites with the IMMP protocol and 91% with the RA. In 2017, monarchs/plant with the IMMP pro- tocol was 0.010 (SD = 0.014) and 0.011 (SD = 0.025) with the RA; in 2018, the mean number a b Figures 20. a) Mean milkweed density (plants/ha) and b) mean monarch eggs and larvae per milkweed plant with two sampling methodologies, IMMP and averaged values for 2–4 RAs. Mean values are indicated by the “x”; median values by a horizontal line. Boxes indicate 25 and 75% quartiles; bars indicate the upper and lower quartiles, and outliers more than 1.5 the 75% quartile are depicted by dots. ba Figures 21. a) Comparison of milkweed density (milkweed plants/ha), log10 transformed, for sites monitored in 2017 (red) and 2018 (blue) using the IMMP and RA averaged for each site, with 95%CI indicated in gray. b) Monarch eggs and larvae per milkweed plant searched (monarchs/plant), log10 transformed, for sites monitored in 2017 (red) and 2018 (blue), same two methodologies.

Product B: Rapid Assessment of Roadside Habitat for Monarchs 41 of monarchs/plant was 0.099 (SD = 0.105) with the IMMP and 0.153 (SD = 0.173) with the RA. In 2018, the mean number of monarchs/plant was 0.099 (SD = 0.105) with the IMMP and 0.153 (SD = 0.173) with the RA (Figure 20b). For monarchs/plant, year was a significant factor (t27 = 2.373, p = 0.025) with more eggs and larvae found in 2018 than 2017, but protocol type did not have a significant effect on monarch density (t66 = 0.118; p =0.906). Monarchs/ plant measured with the two protocols were correlated (Kendall’s rank correlation tau = 0.489, z = 2.71, p = 0.007) (Figure 21b). An estimate of the average number of monarch eggs and larvae per hectare, using the overall IMMP mean was 115 monarchs/ha (2,052 plants/ha × 0.056 monarchs/plant) across both years. For 2017, the estimate was 12 monarchs/ha (1,242 × 0.01), and for 2018, 253 monarchs/ha (2,807 × 0.09). Blooming Nectar Plants The average number of blooming species per site in 2017 was 6.71 (SD = 4.50, range 1–18) with the IMMP protocol, 6.72 (SD = 2.56, range 1–12.33) with RA averaged, and 12.14 (SD = 4.45, range = 5–19) with RA summed. In 2018, the average number of blooming species per site was 10.40 (SD = 6.40, range = 1–23) with the IMMP protocol, 6.57 (SD = 2.85, range = 2–11.33) with RA averaged, and 12.00 (SD = 5.35, range = 1–20) with RA summed (Figure 22). Comparing the number of blooming species by IMMP to the RAs (taking each RA inde- pendently as in milkweed and monarch analyses), the significance of the factors in the model was as follows: year (t27 = 2.33, p = 0.027), protocol type (t82 = –0.047; p = 0.963), and protocol type by year interaction (t82 = –2.86; p = 0.005). In 2017, the number of blooming species estimated by IMMP did not differ from the RA averaged (t26 = 0.007, p = 0.995) but was lower Figure 22. Mean number of blooming species as estimated by IMMP, averaging across RAs (RA averaged) per IMMP site and summing across RA per IMMP site. Mean values are indicated by the “x”; median values by a horizontal line. Boxes indicate 25 and 75% quartiles, and bars indicate the upper and lower quartiles.

42 Evaluating the Suitability of Roadway Corridors for Use by Monarch Butterflies than the RA summed (t26 = 6.247, p < 0.001). In 2018, for the same comparison, IMMP results did not differ from RA summed (t28 = 1.532, p = 0.136), but were higher than RA averaged (t28 = −3.463, p = 0.002). In 2017, the number of blooming species by IMMP protocol was correlated with RA averaged (Kendall’s tau = 0.457, z = 2.22, p = 0.027) and RA summed (Kendall’s tau = 0.568, z = 2.684, p = 0.007) (Figure 23a). In 2018, the number of blooming species by IMMP protocol was correlated with RA averaged (Kendall’s tau = 0.617, z = 3.105, p = 0.002) and RA summed (Kendall’s tau = 0.596, z = 2.967, p = 0.003) (Figure 23b). Discussion The research team designed and tested an RA protocol for monarch habitat within roadside ROWs. While other monarch monitoring programs exist, none were developed for ready appli- cation in the roadside ROWs context. The results suggest that the RA provides a standardized, rapid, accurate way to describe habitat conditions for monarch butterflies in roadside ROWs. The research team sought input from roadside vegetation managers through a survey, semi- structured interviews, and by observing representative users applying the protocol in the field. The survey and interviews provided insights into important considerations for design of the protocol and identified constraints facing roadside vegetation managers. Many managers have pollinator programs and were interested in information on the distribution of potential mon- arch habitat as well as ways to assess the current quality of their roadsides as monarch habitat. Furthermore, many (but not all) managers indicated that they could provide staff to assess habitat conditions and that their staff could learn to identify key habitat features, including milkweed species and noxious weeds. However, the amount of time they could dedicate to habitat assessments was limited. Observations of representative users in field visits supported refinement of the protocol by uncovering common mistakes and suggesting ways of custom- izing the protocol for users with differing abilities and roadside management authorities with dif- fering needs. The researchers used this information to design a habitat assessment protocol that would meet the needs of roadside managers. Through the course of developing the RA, including field visits with state DOTs, it was learned that meeting the wide range of needs of DOTs required a flexible survey design. Departments a b Figure 23. Number of blooming plant species in 2017 (a) and 2018 (b) comparing data by RA averaged (in orange) or RA summed (in gray), compared to the number derived from the IMMP for each site.

Product B: Rapid Assessment of Roadside Habitat for Monarchs 43 differ in their capacities to identify vegetation. For instance, some field staff are knowledgeable about vegetation and would like to quantify not only the number of nectar plant species present but also how many are native. Others are only able to quantify numbers of plants that look distinct from one another without identifying them all to species. The research team developed a convenient plant lookup table from which a surveyor can pick plants by either common or Latin names and simply tally unknown types. Departments differ also in the tracking of noxious weeds, from no tracking to extensive lists of species that differ state to state and sometimes by counties or bioregions within states. Therefore, when setting up their survey, road management entities also can include a custom list of species they wish to track. Additionally, they can enter default values regarding management practices of herbicide (type, targets, and frequency of application) and mowing—or these may be left to vary by site. The RA focuses observers on a small area along the roadway (150 ft, which may easily be paced); within each area the observer counts milkweed plants and types of nectar plants and estimates cover of nectar plants and noxious weeds. The protocol may be conducted across par- ticular areas (e.g., where a pollinator planting was installed or where construction is planned) or repeated at intervals along a roadway to depict the conditions in that area (e.g., once per half mile for 10 miles). It should be noted that it is important for managers to conduct assessments at preselected random or systematic (e.g., every half mile) locations if they wish to effectively characterize larger areas without being biased by sampling in locations where habitat conditions are known to be high. Because managers indicated that only a limited number of days and people would be available for assessments, a survey was designed to be conducted once per growing season. To accom- modate the single yearly sample, the researchers created the term “potentially blooming nectar plants” to represent all of the plants that could provide nectar for monarchs and other pollina- tors, regardless of whether they were blooming on the date of the survey (and without informa- tion regarding nectar quality or quantity). This is consistent with a pollinator scorecard being designed by the Rights-of-Way as Habitat Working Group of the Energy Resource Center at the University of Illinois-Chicago (A. Cariveau, personal communication). Because it is gener- ally more difficult to identify plants if they are not blooming, it is recommend that surveys be conducted in peak blooming season within the period(s) of time when monarchs are present to facilitate identification, or at least differentiation, of species (usually mid- to late-summer). The RA protocol was well received by state DOTs for simplicity in implementation without the need for measuring tapes or other sampling gear. While estimating distances will introduce variation into the areas measured, it is likely to be small relative to the inherent variability in the distribution of milkweed and nectar plants in roadside habitats. All of the departments of trans- portation that provided input use Esri software and many employ data collection tablets in the field for other projects and in many cases already use Survey123. As an indication of the interest in this project, personnel at the Delaware DOT implemented the RA at nearly 100 locations in summer 2018 to learn about monarch habitat along their roadways. The research team evaluated the efficiency and efficacy of the protocol by comparing time requirements and parameter estimates obtained through the RA with those required and obtained through the IMMP, a similar but more rigorous protocol. The team sought to deter- mine whether a more rapid assessment, necessary given constraints on time and funding, could characterize the quality of monarch habitat sufficiently to meet the needs and objectives of road- side vegetation managers. The IMMP employs a more rigorous sampling protocol that pro- vides additional data, particularly regarding nectar plant frequency and diversity. This highly repeatable protocol is stronger for objectives of the IMMP, such as tracking changes in habitats throughout seasons and across years and for comparing monarch habitat quality and use across

44 Evaluating the Suitability of Roadway Corridors for Use by Monarch Butterflies land-use sectors. However, the results suggest that, for assessing and comparing ROWs habitat, the rapid assessment produces sufficiently accurate results for the purposes of roadside habitat restoration and management. The research team found the RA to be efficient; the two-person field crew completed assess- ments in an average of 21 minutes, including time spent looking for monarch eggs and larvae. The RA was significantly more efficient than the IMMP, even when repeating the protocol three times over the footprint of the IMMP (sum of 62 minutes as compared to an average duration of 2.5 hours). The RAs may be spread out further, enabling managers to sample from a larger landscape more readily, and in one day a crew could complete 10 to 20 assessments. Experi- enced crews, after learning how to identify the plants in the ROW, are expected to be faster than employees who are conducting assessments for the first time. However, observers typically become faster through practice, and it is expected that most transportation entities focus on habitat assessment rather than systematically looking for monarch eggs and larvae, which would make the surveys even faster. There may be concerns about whether road crews could effectively collect the data required for the RA. However, several other monitoring programs, such as the Monarch Larva Monitor- ing Project and the IMMP, have success in citizen scientists collecting similar data. The RA was effective for measuring milkweed density and monarch eggs and larvae per plant. When averaging RAs, milkweed density estimates and monarch eggs and larvae per plant were not statistically different than those derived by the IMMP protocol, and the estimates by the two methods were moderately correlated across sites (tau = 0.568, 0.489, respectively; Figures 21a and 21b). Milkweed is patchily distributed across the landscape, and the spatial distribution of quadrats sampled with the IMMP protocol was more reliable for detection of milkweed than a single RA, although milkweed detection was similar when combining the several RAs per site. Also, it is likely that the use of quadrats in the IMMP provided more accurate estimates of milk- weed density by focusing observer attention into small areas, but the estimates obtained by the RA were similar and highly correlated. Averaging parameter estimates for multiple RAs gener- ally yielded more consistent results than any single RA from sites, suggesting that combining multiple RAs to characterize areas is preferred over single samples. Numbers of species of blooming nectar plants also were highly correlated between survey protocol types (tau = 0.457–0.617, depending on the comparison; Figures 23a and 23b). A greater number of nectar plant species was detected when summing results across the RAs per site (excluding duplicates) than by a single RA, due to variation in species composition across the area. These results suggest that completing multiple RAs along a roadway will yield better results than single samples in depicting nectar plant availability. Differences were also noted between years in numbers of blooming species. Using the IMMP protocol and data summed across RAs, the average number of blooming plants in 2018 was higher than in 2017, which may have been due to different sampling locations in each year and the particular composition of the flora at the sites sampled. However, the number of nectar plants estimated by the RA averaged across sites was not similarly higher in 2018. It is possible that in these locations blooming plants were patchily distributed across sites, such that in at least one of the three RAs, the blooming species richness was very low (e.g., grass-dominated), thus decreasing the estimate of species richness when averaging as compared to the IMMP. The high milkweed density documented in the study in Minnesota (2,052 plants/ha (834 plants/ac) by the IMMP method and 1,527 plants/ha (620 plants/acre) by RA) confirms that roadside ROWs can provide significant amounts of breeding habitat for monarchs (Kasten et al. 2016). The 2017 estimate could have been inflated due to the fact that sites were selected from a set that contained milkweed in a prior study, but the 2018 average milkweed density was higher,

Product B: Rapid Assessment of Roadside Habitat for Monarchs 45 and these sites were selected through a random process. These milkweed densities are higher than other studies in the upper Midwest (508 plants/ha (Kasten et al. 2016) or 141 plants/ha, as converted from Hartzler and Buhler 2000 in Thogmartin et al. 2017b and used to estimate levels in current roadside ROWs). However, the sample size was small, and the research team did not sample all types of roads, such as those in developed areas that do not typically provide habitat or those that appeared to be less than 4 m wide when reviewed online. Overall estimates of habitat availability must take into account different roadway types and potential variation by region; data collected from more locations will greatly assist in ongoing assessments of monarch habitat availability. The levels of monarch use for reproduction suggest these roadways are serving a significant function for breeding habitat. The per plant density of monarch eggs and larvae ranged from 0.01 monarchs/plant in 2017 to 0.099 in 2018 (IMMP protocol), bracketing the 0.059 reported for roadsides by Kasten et al. (2016) and 0.043 eggs/plant reported by Nail et al. (2015) from Monarch Larva Monitoring Project data from non-roadside areas, primarily gardens. The team detected a strong difference among years in monarch abundance, which is not surprising given high inter-annual variation in monarch numbers (Thogmartin et al. 2017a). This suggests that, if monarch use is a primary focus for a roadside manager, collecting data in more than one year (and comparing across sites within the same year(s)) would be advisable. Repeat surveys within a year would also greatly improve information about monarch use. The presence of late instar larvae indicate that monarchs can develop in these habitats. Providing more milkweed dispersed across the landscape may improve monarch larval sur- vival in lower density patches of milkweed (Zalucki and Kitching 1982), and having access to milkweed across the landscape should increase the number of eggs females lay (Zalucki and Lammers 2010, Grant et al. 2018, Zalucki et al. 2016). However, monarch eggs and larvae sus- tain high levels of mortality due to predation, weather, disease, and other factors (Nail et al. 2015). Also, roadside areas may support lower densities of monarchs than adjoining agricul- tural habitats (Pitman et al. 2018), although it is not known if these patterns reflect differences in habitat quality or other factors, such as behavioral responses to linear landscape features. There- fore, more information about the survival of monarch eggs and larvae in roadside habitats compared to other habitat types will be highly informative in assessing the relative benefits of roadside habitat for producing monarchs. While the results and a handful of previous studies highlight the promise of roadsides as monarch habitats, these areas also bring a suite of threats to monarchs and other pollinators including collisions with vehicles and chemical inputs (Skorka et al. 2013, Keilsohn et al. 2018, Pitman et al. 2018, Snell-Rood et al. 2014). However, larger butterflies such as monarchs may sustain a lower rate of mortality from car collisions than smaller butterflies (Skorka et al. 2013). Furthermore, mortality from cars is lower in roadside habitats with certain characteristics, such as greater plant species richness (Ries et al. 2001, Skorka et al. 2013). The width of the ROW habitat as well as the composition of adjacent lands also may affect collision mortality rates, such that wider habitats with greater access to adjoining habitats may reduce collision mortality (Munguira and Thomas 1992, Skorka et al. 2013, but see Saarinen et al. 2005). Chemicals, including sodium and heavy metal runoff from roadways, are incorporated into roadside vegetation (Snell-Rood et al. 2014, Munoz et al. 2015). These chemicals could affect the development of monarch eggs and larvae or even affect adults through contamination of nectar resources. Further study of roadside areas to profile monarch egg and larval survival as well as chemical or traffic-induced mortality would allow better understanding of how roadside habitats perform as monarch breeding areas. Roadside management authorities are becoming aware of the impact of management poli- cies on roadside habitat, and exemplary programs with deferred mowing, re-establishment of

46 Evaluating the Suitability of Roadway Corridors for Use by Monarch Butterflies native plants, control of noxious weeds, and integrated vegetation management occur around the country. While providing protocols for the assessment of pollinator habitat in ROWs is an important first step, additional work is needed to interpret the resulting data in the context of pollinator habitat quality. The research team has developed the Roadside Monarch Habitat Calculator to score monarch habitat quality for sites assessed by the RA protocol. Additional challenges include balancing the multiple management needs for ROWs and communicating the benefits of native, uncut vegetation to shift public preferences for well-manicured turf grass along roadways. Because of the importance of the breeding season to the monarch annual cycle (Oberhauser et al. 2017), the strong connection between habitat loss in the core of the eastern population’s breeding range, low monarch numbers (Thogmartin et al. 2017a), and use of roadsides for monarch breeding (Kasten et al. 2016), roadside restoration and management is promising for monarch conservation. Furthermore, roadside areas managed for monarch habitat provide native plants that could benefit other wildlife, such as small mammals, birds, pollinators and other beneficial insects. Ongoing communication and research around the potential conserva- tion benefits of well managed roadside ROWs will be highly beneficial. Important findings from this work include the high level of interest and motivation within DOTs for providing habitat for monarchs. The research team was impressed by the number of interested departments as well as the number that are implementing exemplary practices. The survey revealed the needs for simple ways to assess habitat both in the landscape setting as well as at the level of the vegetation present. Communication tools are also highly valued by trans- portation administrators who are not experts in the area of monarch habitat development and who interface with varied audiences from high level management, to vegetation management crews, to the public.