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Journal of Natural History

ISSN: 0022-2933 (Print) 1464-5262 (Online) Journal homepage: http://www.tandfonline.com/loi/tnah20

A long-term survey of spring monarch butterfliesin north-central Florida

Lincoln P. Brower, Ernest H. Williams, Kelly Sims Dunford, James C. Dunford,Amy L. Knight, Jaret Daniels, James A. Cohen, Tonya Van Hook, EmilySaarinen, Matthew J. Standridge, Samantha W. Epstein, Myron P. Zalucki &Stephen B. Malcolm

To cite this article: Lincoln P. Brower, Ernest H. Williams, Kelly Sims Dunford, James C. Dunford,Amy L. Knight, Jaret Daniels, James A. Cohen, Tonya Van Hook, Emily Saarinen, Matthew J.Standridge, Samantha W. Epstein, Myron P. Zalucki & Stephen B. Malcolm (2018) A long-termsurvey of spring monarch butterflies in north-central Florida, Journal of Natural History, 52:31-32,2025-2046

To link to this article: https://doi.org/10.1080/00222933.2018.1510057

Published online: 10 Sep 2018.

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A long-term survey of spring monarch butterflies innorth-central FloridaLincoln P. Browera#, Ernest H. Williams b, Kelly Sims Dunforda, James C. Dunfordc,Amy L. Knightd, Jaret Danielsc, James A. Cohenc, Tonya Van Hookc, Emily Saarinene,Matthew J. Standridgec, Samantha W. Epsteinc, Myron P. Zalucki f

and Stephen B. Malcolmg

aDepartment of Biology, Sweet Briar College, Sweet Briar, VA, USA; bDepartment of Biology, HamiltonCollege, Clinton, NY, USA; cMcGuire Center for Lepidoptera and Biodiversity, Florida Museum of NaturalHistory, University of Florida, Gainesville, FL, USA; dFlorida Natural Areas Inventory, Florida State University,Tallahassee, FL, USA; eDivision of Natural Sciences, New College of Florida, Sarasota, FL, USA; fSchool ofBiological Sciences, The University of Queensland, Brisbane, Australia; gDepartment of Biological Sciences,Western Michigan University, Kalamazoo, MI, USA

Long-term springtime counts of immature and adult monarch but-terflies and their Asclepias humistrata host plants in north-centralFlorida reveal a close relationship between the milkweed’s phenol-ogy and the butterfly’s spring remigration from Mexico. Remigrantadults arrive after most frosts occur and as the milkweeds are flour-ishing but before the plants begin to senesce. The peak of adultarrival is during the first few days in April; the eggs that are laidduring this peak develop through April, leading to a second peak inadult abundance in early May. These newly emerged adults continuethe migration northward. In addition to assessing the phenology ofannual events, our long-term survey, with regular monitoring from1994–2017, has enabled us to evaluate long-term trends. Both adultsand immatures have declined in abundance from 1985 to 2017; since2005, both have declined by around 80%. This decline has occurredconcurrently with the decline in the number of monarchs at theirMexican overwintering sites.

ARTICLE HISTORYReceived 16 October 2017Accepted 1 August 2018

KEYWORDSPopulation decline; springremigration; phenology;milkweeds; monitoring

This paper presents the results of a long-term survey of early spring breeding bymonarch butterflies (Danaus plexippus L., Lepidoptera, Nymphalidae) on Asclepias humis-trata Walt. (Apocynaceae, Asclepiadoideae) in north-central Florida. Early observationsfrom 1981–1993 were followed by standardized surveys from 1994–2017. Consistentlong-term monitoring of changes in monarch spring breeding are rare for this well-studied species, even though recolonization of the upper USA and Canada depends onspring breeding in southern states. Previous work on spring migration found a delayedreturn and smaller geographic coverage of the initial wave of spring remigration(Howard and Davis 2015), with both results likely a result of declining overall abundance.Here we add to an understanding of spring migration by addressing questions about the

CONTACT Ernest H. Williams ewilliam@hamilton.edu#Deceased

JOURNAL OF NATURAL HISTORY2018, VOL. 52, NOS. 31–32, 2025–2046https://doi.org/10.1080/00222933.2018.1510057

© 2018 Informa UK Limited, trading as Taylor & Francis Group

Published online 10 Sep 2018

changing abundance of monarchs in north Florida and the phenological matching ofmilkweed host plants with monarch arrival and reproduction.

Considerable research has supported the hypothesis that the early spring monarchscollected from Texas to north Florida are individuals returning from overwintering sitesin Mexico (Lynch and Martin 1987, 1993; Malcolm et al. 1987, 1993; Martin and Lynch1988; Cockrell et al. 1993; Riley 1993; Howard and Davis 2004). Monarchs are widelyreported to reach the southern tier of states, including north Florida, during March (datafor 2000–2017, Journey North 2017). However, the geographic origin of the springmonarchs in north-central Florida continues to be debated because, in order to arrivein this area in late March and early April from their Mexican overwintering sites, themonarchs would have to either fly from Mexico across the Gulf of Mexico, which has notbeen documented and seems unlikely for several reasons (Brower 1985, 1995), or, afterreaching the Gulf coast in Texas, fly east along the coast and then south into north-central Florida. Alternatively, they could be monarchs that overwintered in coloniesalong the Gulf Coast and migrated inland in late March and early April.

While some monarchs may have flown in from coastal overwintering sites, thedistribution and occurrence of these sites is historically haphazard and their overwinter-ing numbers small (Brower 1995). Recent evidence supports the minimal influence ofcoastal sites; Journey North data on overwintering butterflies in nine coastal states overeight years, 2002–2009, summarized by Howard et al. (2010), reported 254 individualsbut no overwintering clusters, with 80% of the observations in Texas and Florida.Furthermore, our extensive cardenolide fingerprinting data (Malcolm et al. 1993)together with over a century of natural history observations reviewed in Brower (1995)strongly support the hypothesis that the majority of these April Cross Creek monarchsbred in the north the previous summer on A. syriaca, overwintered in Mexico, and flewto Cross Creek in the subsequent early spring.

Here we present the results of a long-term spring monitoring programme in whichwe recorded the phenology of a large population of Asclepias humistrata plants and thearrival time, abundance, and reproduction of monarch butterflies on this milkweed innorth-central Florida. Early observations from 1983–1994 were followed by regular andconsistent monitoring from 1994 to 2017. We give evidence that the abundance of thespring remigrant population has diminished through time, as have both the eastern andwestern North American overwintering populations (Brower et al. 2012; Xerces Society2016; Pleasants et al. 2017; Schultz et al. 2017; Malcolm 2018), and that the arrival ofmonarchs is timed to coincide with the availability of their host plant A. humistrata.

Methods

Host plant and study site

Asclepias humistrata, the sandhill or pinewoods milkweed, is the common milkweed inour study area; it is largely a south-eastern US coastal plains species occurring fromsouth-central Florida at approximately 28°N latitude northwards to North Carolina atapproximately 37°N latitude and westwards to eastern Louisiana at approximately 90°Wlongitude. It is a decumbent, herbaceous perennial plant that blooms from April to July,and its habitat includes well-drained soils on sand dunes, dry oak woods, and pine-scrub

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habitats (Woodson 1954). In north-central Florida A. humistrata is frequently abundant inovergrazed pastures and along disturbed roadsides. Based on its known US distribution(Woodson 1954; Kartesz 2015), we estimated the species range, all in the south-easternUSA, as 254,000 km2 (= 98,070 square miles). In contrast, Asclepias syriaca L., theprincipal summer foodplant of monarchs in eastern North America and Canada, has amuch more extensive distribution (3,375,000 km2 = 1,303,095 square miles) (Woodson1954; Crolla and Lafontaine 1996; Kartesz 2015). Thus, the distribution of A. humistrata isonly 7.5% that of A. syriaca.

The study area consisted principally of a cattle pasture (ca. 9 ha) at Cross Creek inAlachua Co., FL (N 29°31.75′, W 82° 11.86′), with more than 1000 A. humistrata plants(Figures 1, 2 and 3). According to the owner, Zane Hogan, the pasture had not beentreated with herbicide for several years prior to and throughout the duration of ourstudy. This allowed the survival of milkweed as well as other species undesirable forcattle, including prickly pear cactus, Opuntia humifusa Raf. (Cactaceae), tread-softly,Cnidoscolus stimulosus (Michx.) Engelm. & Gray (Euphorbiaceae), and yellow thistle,(Cirsium horridulum Michx. (Asteraceae). Grazing cattle avoided eating or damagingthe milkweeds, which contain large amounts of latex and high concentrations of toxiccardiac glycosides (Martin et al. 1992). The pasture was surrounded on the north, east,and west by ‘improved’ pasture that contained few milkweeds or other nectar sourcesand to the south by County Road 346 and a commercially planted slash pine forest,Pinus elliotti Engelm. (Pinaceae). Most land in this region is either pasture or commercialslash pine plantation. There were several large longleaf pines, Pinus palustris Mill.Pinaceae, scattered through the pasture, and a ca. 20 m wide strip of live oaks,Quercus virginiana Mill. (Fagaceae), separated the study site from the neighbouring

Figure 1. Satellite view of Hogan’s pasture (X) and the roadsides in Cross Ck, Alachua Co., FL, wherethe density, health, stage and number of Asclepias humistrata plants and stems and the springbreeding populations of monarch butterflies were assessed.

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nearly milkweed-free pasture to the east. With a population of A. humistrata that wasboth dense and isolated, this site provided prime breeding habitat for monarchs.

Surveys

Springtime monitoring of monarch adults and immatures and milkweeds began in 1981and ran typically from mid-March to mid-May. Observations from 1981 to 1985 werealong roadsides, but because of destructive roadside management (Hamilton 1987), weshifted our monitoring in 1985 to a pasture less than 1 km away, and all surveys from1986 to 2017 took place here (Hogan’s pasture; Figure 1). The data from the earlier yearsallow some comparisons to be made with the primary survey data, which were collectedfrom 1994 on.

The frequency of counts (every 3–4 days) and methods became standardized in 1994,so our analysis focuses primarily on data from 1994–2017. On each sampling date, atransect was established within the pasture with a randomized beginning location andpointed in a randomized compass direction; all plants within 1.5 m of the transect wereclosely examined for immatures until 50 plants had been monitored. If the transectreached the edge of the pasture before 50 plants had been surveyed, an additionalrandomized transect was added. All leaves, flowers, and buds on all stems of each plantwere examined, with records kept of the phenological stage of each plant, number ofstems, and numbers of monarch eggs and of each larval instar. Analyses for immaturesused the 20 years of data in which a minimum of 500 surveys were conducted (all years

Figure 2. View across Hogan’s pasture showing numerous flowering Asclepias humistrata plants. 28April 2010. Photo by K. Sims Dunford.

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Figure 3. Stages of development of Asclepius humistrata. (a) Indeterminate (without reproductiveorgans, 24 April 2010); (b) early bud (immature flower buds visible outside terminal leaves, 3 April2012); (c) bud (mature flower buds, 26 March 2009); (d) flowering (8 May 2010); (e) pod (fully matureseed pods, 19 May 2010); (f) post-pod (senesced pods that have released their seeds, 26 May 2010);(g) frost damaged plant (9 April 2009); (h) fifth instar Danaus plexippus on a flowering plant (24 April1996). Photos by L.P. Brower, K. Sims Dunford, and M. Standridge.

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from 1994 except 1997, 2000, 2001 and 2003; range 536–2987 surveys per year); at least50 plants were surveyed on each date, but the same plants were frequently surveyed ondifferent dates. Different participants were instructed on how to conduct the plant andbutterfly surveys across the years of this study. In addition a full census of the milkweedsin Hogan’s pasture was conducted each May from 2012 to 2015.

An adult survey was conducted on each survey date subsequent to the immature andplant counts. Adults were netted for one hour by one or two persons, with an attemptmade to capture each individual observed. Upon capture, each monarch adult wasplaced in a glassine envelope and held in a cooler. At the end of the collection period,each unmarked individual was sexed, tagged, and ranked for wing condition from 1(very fresh, virtually no scales missing) to 5 (worn, many scales missing). All butterflieswere re-chilled after data recording to minimize escape responses; when released, theywarmed to flight threshold within five minutes and resumed normal activity (see Kempand Zalucki 1999).

The above procedures for surveying milkweeds, immatures, and adults were followedfrom 1994 to 2017, with the following exceptions. Surveys of immatures and adults wereconducted simultaneously rather than sequentially and for up to 2 h in 1994−1999.Adults were not captured in 1997 and 2001 and only a few were in 2004. No surveyswere conducted in 2000 and 2003. Because surveys before 1994 used similar but varyingprocedures, we used records from those earlier years to provide comparative informa-tion with the data recorded more systematically from 1994 onwards. Oviposition ratesfor 1994–2017 were analysed as eggs/stem, but because oviposition rates in earlier yearswere recorded as eggs/plant, we added an analysis of eggs/plant for the full dataset.

The related queen butterfly (Danaus gilippus berenice Cramer) breeds extensively insouth-central Florida on A. humistrata (Brower 1961), is common in the nearby westcoastal areas (Moranz and Brower 1998), and lays eggs that are difficult if not impossibleto distinguish from monarch eggs. From 1995 through 2017, we recorded 19 queenadults in the pasture. Some of the eggs in our study could have been laid by queens, buttheir relative infrequency (865 monarch adults captured compared to 19 queens; 2.1%)over the same time span indicates that potential confusion was negligible. Additionally,queen larvae are easily distinguishable from monarch larvae, and none were recorded inour study area.

Statistical analysis was by SPSS (IBM Corp 2016). To evaluate long-term trends inabundance of eggs and adults, we ran linear regressions of yearly averages for the four-week period 22 March–18 April of adults captured per hour and of eggs/stem (alsoevaluated as eggs/plant), with year as independent variable. We used the Pearsoncorrelation coefficient to analyse the correlation between eggs/stem and adults/h.

Results

Milkweed phenology

Asclepias humistrata first emerges at Cross Creek around the end of February and thebeginning of March, and the development of the plants can be traced through thestages of early bud, bud, flower and pod (Figures 3 and 4). Peak flowering occurstypically from 12 to 18 April, after which pods develop and the plants then begin drying

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and senescing by late May. Some plants remain in an indeterminate or non-floweringstate throughout the season and have been excluded from the analysis of floweringstates shown in Figure 4. The timing of plant development varied from one year to thenext. Compared to the typical flowering date of 12–18 April, peak flowering in 2002occurred during the week of 5–11 April; in 2008 peak flowering occurred a week earlier,29 March–4 April; and in 2010 peak flowering occurred a week later, 19–25 April(Figure 5).

During the first week of March, 95% of the plants that will flower are in an earlybud stage, just emerging from the ground. Early budding gradually declined throughmid-May, giving way to mature buds and then full flowering plants through late May.Maturation and pod production begins in early April and continues into May. Theplants are effectively available for monarch oviposition and as larval foodplants from1 March to 23 May and for nectaring from the third week in March through to theend of April.

The size and health status of the milkweed population was assessed each May for2012–2015 (Table 1). These complete censuses establish the size and health of themilkweed population at the study site. Over the four years of these surveys, the numberof stems per plant ranged from 1 to 32, with single-stemmed plants representing from

Figure 4. Development of Asclepias humistrata at Cross Creek (data for 1997–2017; no counts for2000 and 2003). Plants without any flower development are excluded. Maximum flowering occurredthe week of 12–18 April. Based on 11,707 observations.

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61% of the total in 2014 to 79% in 2013. The maximum number of stems per plant was32 in 2015; this may have been a very old plant (Betz 1989).

Adult monarch phenology and abundance

Adult monarchs first appear at Cross Creek in mid-March, and their numbers increaserapidly towards the end of that month (Figure 6). The earliest arrival recorded for theentire period was 14 March in both 2012 and 2014. After a decline during the middle ofApril, a second increase in abundance develops in early May (Figure 6). The timing of thesecond peak, which occurs five weeks after the first, suggests that it develops from theeggs laid by adults during the first peak of appearance. Monarch development isreported to require 352 C degree-days with a 11.5°C base (Zalucki 1982). Using

Figure 5. Variation in flowering phenology of Asclepias humistrata at Cross Creek across three years;2002 was a typical year, 2008 was an early year, and 2010 was a late year.

Table 1. Number and health of the milkweed A. humistrata in Hogan’s pasture at Cross Creek, FL.Censuses from May for four seasons, 2012–2015.Dates Healthy Senesced Dead Total

18–21 May 2012 1769 (84.4%) 277 (13.2%) 49 (2.3%) 209520–22 May 2013 2399 (94.7%) 125 (4.9%) 9 (0.4%) 253326–31 May 2014 1329 (51.6%) 1090 (42.3%) 157 (6.1%) 257613–15 May 2015 1122 (35.2%) 1900 (59.6%) 166 (5.2%) 3188

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temperature records from nearby Gainesville, FL, we determined the degree-daysbetween the peaks of adult abundance (5 April and 10 May as shown in Figure 5). Forthe four years 2007–2010, degree-days ranged from 292 in 2007 to 365 in 2010, totalsclose enough to 352 to support the conclusion that the two peaks are one generationapart. In addition, Knight et al. (1999) reported a mean residence time of only 6.37 daysfor adults arriving at Cross Creek in 1995, indicating that the butterflies do not remainlong in the area. Results from the 1980s support the same patterns; for example, from1981 through 1986, the first adults and first eggs were observed at the end of March orfirst of April, and adults in May were fresher than those in April.

Additional evidence that the second peak in abundance represents new emergentadults comes from an examination of wing wear (Figure 7). Wing wear is a proxy for age;wings showed an intermediate level of wear up to 28 March, followed by greater wingwear from 29 March to 12 April, which was during the peak of oviposition. Wings wereless worn after 19 April as fresh butterflies were added to the population. By early May,most butterflies showed little wear. These data indicate that the butterflies in early Aprilwere remigrants.

In addition to the timing of average monarch return matching the pattern of milk-weed growth, there is limited evidence that this timing is synchronized within years. The

Figure 6. Appearance of adults at Cross Creek calculated as the number captured per hour,averaged for each week from 1994 through 2017 (excluding 1997, 2000–2001, 2003). The earliestarrival was 14 March (week of 8–14 March). Not all weeks were surveyed in each year. Means areshown with 95% CI. Based on observations of a total of 865 adults.

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milkweed flowering patterns in Figure 5 show that 2008 was an early year and 2010 wasa late year. Similar timing occurred in monarch numbers; the first capture in 2008 was on19 March, with a peak in numbers on 2 April; in contrast, the first capture in 2010 wasnot until 7 April with a peak in numbers on 9 April. Similar timing in the springappearance of milkweeds and monarchs likely reflects common responses to environ-mental variables.

Changes in the abundance of adult monarchs from 1994 to 2017 can be compared(Figure 8) by examining the adults/h averaged for the four central weeks, 22 March–18April, which are the weeks of initial peak abundance. The overall rate of observation ofadults has decreased significantly since 1994 (slope = −0.324, F1,15 = 12.213, p = 0.003,R2 = 0.449). Because 1994 and 1995 were extraordinary years, with nearly three times asmany monarchs as have been seen since then, we also analysed the change excludingthose two years, and the result showed no significant change from 1996 to 2017.However, from 2005 to 2017 a highly significant decrease occurred (88% by linearregression; slope = −0.313, F1,10 = 20.487, p = 0.001, R2 = 0.672). This analysis is basedon all available data for each set of years. Although the recent decline appears to parallelthe decline in abundance at the Mexican over-wintering colonies (Rendon-Salinas et al.2017), we did not find a significant correlation between spring abundance at Cross Creek

Figure 7. Wing wear of monarchs for each week through the spring flight period, from a total of 302adults combined for 2005–2017 using a scale from 1 (very fresh with no scale loss) to 5 (worn, manyscales missing). Two measures are shown for each week: the median determined from all butterfliesand the proportion of adults with wing wear 1s and 2s. Individuals in early April were mostly fresh.

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and the previous winter’s overwintering abundance in Mexico (n.s. for both eggs andadults).

Reproduction

The first eggs at Cross Creek appeared in the same week that adults first appeared, 8–14March, and egg abundance then rose rapidly to a peak during 29 March–4 April (Figure 9).From late April into May oviposition continued, though at a lower rate, indicating thatfemales continue to visit the site well into May. The higher variance before and during thepeak (Figure 9) is likely a result of year-to-year variability in arrival dates; later in April andMay, after peak arrival, the variance in oviposition rates is much less.

Eggs/stem data for the middle four weeks of greatest oviposition (22 March–18 April)and averaged each year for 1994–2017 reveal the change in oviposition rates over time(Figure 10). Egg density at Cross Creek has declined since 1994 (F1,18 = 11.501, p = 0.003,R2 = 0.390); restricting the analysis to 2005–2017 also gives a significant decline(F1,11 = 11.723, p = 0.006, R2 = 0.516). Excluding the unusual abundance in 1995 fromthe regression, the decline is approximately 80% for both comparisons (1994–2017,2005–2017), with no change in management of Hogan’s pasture. This decline is sup-ported by our earlier surveys. Because the data up to 1993 were recorded as eggs/plantrather than eggs/stem, we can compare the data from 1984 using that unit. Surveysfrom five years of data before 1994 gave an average 1.12 eggs/plant (range 0.74–1.93)compared to averages of 0.82 eggs/plant for 1994–1995, 0.41 eggs/plant for 1996–2006,and 0.12 eggs/plant for 2007–2017. The trend in egg density, analysed in different ways,

Figure 8. Adults captured per hour averaged for the four central weeks, 22 March–18 April, from1994–2017. Data were not available for 1997, 1999–2001, and 2003–2004.

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is strongly downward and reveals a significant decline in the abundance of monarchs atCross Creek.

The egg densities we calculated ranged from less than 0.1 to 0.7 eggs/stem, rates thatare similar to published values: an average maximum of 0.2 eggs/stem on A. syriaca atnon-agricultural sites (Pleasants and Oberhauser 2013); 0.098 to 1.097 eggs/stem inmidwestern fields (Pleasants et al. 2017); and 0.1 to 1.8 eggs/stem for March and Aprilin the south-central and north-central USA (Stenoien et al. 2015).

Immature phenology and survivorship

Eggs and first instar larvae first appear in the week of 8–14 March, which is a time whenmany fresh milkweeds are in bud; immatures peak in abundance around the end ofMarch/beginning of April as the milkweeds approach their peak of flowering; and theyare less abundant in late April, with a slow decline through mid-May (Figure 11). Secondand third instar larvae peak in early to mid-April at the height of flowering. Fourth andfifth instars have a broader and less well-defined peak, which ranges from early to lateApril as increasing numbers of plants enter their podded stage. The median appearancefor each stage is as follows: eggs, 29 March–4 April; 1st–3rd instars, 5–11 April; and 4thand 5th instars, 12–18 April. Overall, immatures are present for two months, from mid-March to mid-May, based on totals of 3446 eggs and 1295 larvae (1994–2017). Thus the

Figure 9. Egg abundance through the spring, recorded as mean eggs/stem for each week from 1994through 2017 (50 plants per sample, minimum of 500 plants examined each year; excludes 1997,2001, and 2004). Means are shown with 95% CI calculated from 3545 eggs found during 44,753surveys of stems.

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Figure 10. The yearly decline in oviposition rate at Cross Creek from 1994 to 2017, recorded asaverage eggs/stem (500 plant minimum each year) for the four weeks from 22 March to 18 April.(F1,18 = 11.501, p = 0.003, R2 = 0.390). Excluding the extraordinary high value in 1995, the decline bylinear regression is about 78%. Based on a total of 3545 eggs.

Figure 11. Phenology of immatures at Cross Creek, totalled for 1994–2017. Larvae are divided into1st through 3rd and 4th and 5th instar. Based on totals of 3444 eggs and 1295 larvae.

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returning monarchs exploit the milkweeds for most of the months of March and Aprilwhen the A. humistrata plants are in prime condition.

By totalling the number of eggs and larvae found across all surveys in all years, we haveestimated the survival rate of each stage of development (Table 2). From a total of 5622eggs and larvae counted from 1984–2017, first instars represented just 16.0% of the totalnumber of eggs, while 5th instars represented only 3.9% of eggs. These numbers are thepercentage of eggs that survived to 1st and 5th instars, but to determine actual survivalrates, we adjusted these percentages by the time (in degree-days) spent in each stage(Zalucki 1982). The highest rate of mortality occurred in the egg stage (77.5%), and overallsurvivorship from egg to 5th instar was only 2.9%. The first 5th instars were seen in mid-April. Records from 1987 noted that just 63 of 98 eggs hatched (64%); this finding impliesthat much of the mortality from egg to first instar occurs before the 1st instars haveemerged. Consistent with the findings of Oberhauser (2012), we recorded very fewchrysalides over the 32 years (n = 6) primarily because mature larvae move off themilkweeds to pupate. Additional chrysalis mortality from predators and parasites is likely(Zalucki and Kitching 1982; Oberhauser et al. 2017) but is difficult to quantify.

Frosts

Although temperatures in north central Florida do not fall low enough to kill the eggs orlarvae (Nail et al. 2015), frost damage to the milkweed plants results in leaves that arewilted and drying (Figure 3(g)). Frost, therefore, reduces the availability of ovipositionsites and suitable food for early instar larvae. Smaller, younger plants were damaged byfrost more than larger, older plants. Plants with less than 50% frost damage were able toregrow and produce flowers and pods, whereas plants with more than 70% frostdamage could regrow basal parts only but not reproduce. Plants with 100% frostdamage were unable to regrow. When regrowth is possible, it occurs within 10 days.The most extensive damage we observed, when more than 70% of the plants wereaffected, was found on 11 March 2011, 20 March 2004, 27 March 2006, and 8 April 2009,indicating hard frosts (low temperatures) on those dates. Following the freeze on 8 April2009, oviposition dropped significantly (5 April, 0.28 eggs/plant; 9 April, 0.02 eggs/plant).Thus, late freezes can reduce oviposition.

Our records point to intense freezes at Cross Creek being common in February,infrequent in March, and rare in April, a pattern to which the spring remigration ofmonarchs appears to have adapted. For comparison, we found that temperature recordsof nearby Gainesville for the years 1984–2011 showed the last freeze occurring after 1March for 13 of those 28 years but after 15 March for only three of the 28 years (FAWN

Table 2. Survivorship of monarch larvae by instar, totalled for 1984–2017. Survivorship rates havebeen adjusted for the degree days required for each stage (Zalucki 1982).Stage Per cent total no. eggs Adjusted per cent survivorship Survivorship per cent from previous stage

1st 16.0 22.5 22.52nd 11.3 18.1 80.43rd 9.2 17.3 95.64th 3.8 4.7 27.25th 3.9 2.9 61.7

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2015). Intense freezes also alter the spring milkweed phenology; an extended freeze inFebruary 2010 led to the delayed flowering sequence for that year (Figure 5).

Overall abundance and phenology

Both egg density and the rate of adult observations have declined significantly at CrossCreek, whether one analyses data from 1994 (decrease of 91% adults/h and, excluding1995, decrease of 78% eggs/stem) or results from only the more recent years, 2005–2017(88% adults/h and 78% eggs/stem). A highly significant correlation between the densityof eggs and the rate of adult observation documents the connection between these twocalculated values (Pearson correlation coefficient for 1994–2017 = 0.841, p < 0001;coefficient for 2005–2017 = 0.742, p = 0.006).

From 1994 to 2017, the rates of milkweed development (rate of flowering), adultarrival, and oviposition, did not change perceptibly based on the weeks in which each ofthese measures peaked in each year (all three measures regressed against year, n.s.). Theabsence of detectable phenological shifts since 1994 may result from our surveysshowing peak values week by week rather than day by day. The only evidence wehave of a phenological shift is that the peak in egg density occurred on 10 April in both1983 (Malcolm et al. 1987) and 1987, which was at least one week later than our surveyresults for 1996–2017, when the peak was 29 March–4 April. Other comparisons of earlyand late years show no obvious differences. The correlation of the appearance of adultsand eggs with milkweed development is indicated by a comparison of two years: in2008 (the early year in Figure 5), adults and eggs both peaked around 2 April, whereas in2010 (the late year in Figure 5), adults peaked on 7 April and eggs on 10 April. The dataare insufficient to examine this relationship more closely.

Discussion

Phenology of migration

Factors such as host plant abundance and mating correlate with the breaking ofdiapause in monarchs overwintering in Mexico (Goehring and Oberhauser 2004) andtheir dispersal, whereas low temperatures promote migration in a northward direction(Guerra and Reppert 2013). Our long-term study of monarchs during the spring in north-central Florida has documented a consistent pattern of remigrating monarchs arriving inMarch, with oviposition on A. humistrata and production of the new spring generationthat likely continues the migration northward late in April. Arriving during the flush ofnew growth of A. humistrata, remigrant adults appear timed to match the phenology ofthe milkweeds and to minimize the likelihood of encountering frost damage. If mon-archs arrive and oviposit too early in March, the milkweeds can freeze, desiccate, andleave the larvae starving. If they arrive too late, i.e. late April, the milkweeds will havebegun senescing, which likely means providing poor nutritional value. Thus it appearsthat there is only about a three-week window when the spring remigrant monarchs cansuccessfully establish the new spring generation and then continue the migrationnorthward. Adults during the first peak of abundance are typically worn, consistentwith evidence that they have remigrated from Mexico (perhaps with a few from south

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Florida), whereas those in the second peak, four to five weeks later, are fresher, showingthat the second peak is a new spring generation rather than late remigrants. Theseresults are consistent with our hypothesis that the eastern North American monarchpopulation has evolved with the timing of emerging spring milkweeds during theannual migratory cycle.

Remigrants oviposit and nectar on the growing milkweeds but do not remain at ourstudy site in north Florida for long. Knight et al. (1999) reported a mean residence timeof spring monarchs at our study site of only 4–6 days, with females in residence forapproximately one day longer than males. It is likely that, in the presence of freshmilkweeds, females remain longer to oviposit while males continue to migrate. Ourresults support this sex difference; from 1994 to 2017, more females were captured thanwere males (488–367). Most individuals when captured were either nectaring on themilkweed flowers or searching for oviposition sites. Thus, the female-biased sex ratio werecorded may result from their remaining at the site longer, a female-biased capturerate, and possibly the presence of more females (see Zalucki and Susuki 1987 foradditional factors). Although the number of females exceeds the number of males,both sexes arrive from Mexico.

Malcolm et al. (1987) reported a third spring generation at Cross Creek in 1983,following the arrival of the first generation and their ovipositing to produce the secondgeneration, but we found no evidence for this in other years. The new spring generation,which appears about five weeks after the arrivals from Mexico, is comprised of theoffspring of the arriving remigrants. The additional generation reported by Malcolmet al. (1987) was likely produced on fresh regrowth of the milkweed following mowingalong the roadsides by the Florida Department of Transportation. Extensive milkweedregrowth after mowing resulting in extended oviposition by monarchs has been docu-mented by Fischer et al. (2015) and Alcock et al. (2016).

Although we did not detect any phenological shifts across the years of the currentstudy, climate change may be affecting milkweed development, adult arrival anddeparture, and timing of oviposition at Cross Creek. If so, continued monitoring mayshow such changes. Because climate change can lead to mismatches in timing, thephenological matching that we found between the development of milkweeds and thearrival and reproduction of migratory monarchs could be altered in the future.

The consistent timing of the spring remigration of the monarch butterfly with thephenology of A. humistrata in north Florida raises the possibility that the spring remi-gration could also be affected by human induced changes in the geographic distributionof the milkweed flora. Monarchs appear to be well adapted to particular milkweedspecies as they migrate northward (there are 74 species of Asclepias available tomonarchs in the USA and Canada; M. Fishbein pers. comm.). Two other milkweedspecies favoured during the spring remigration in southern states are Asclepias viridisWalt. and Asclepias asperula Woods (Martin and Lynch 1988; Van Hook and Zalucki 1991;Lynch and Martin 1993).

Currently, tens of thousands of seedlings of several different milkweed species arebeing distributed to volunteers to plant in gardens and along roadsides across theeastern USA and Canada, thereby altering the natural biogeographic distribution ofmilkweeds. The goal of this planting is to restore the numbers of monarchs in thedwindling eastern population (Lovett and Taylor 2017) by expanding breeding habitat.

2040 L.P. BROWER ET AL.

Additional milkweed habitat would certainly be beneficial (Pleasants 2017), but habitatexpansion should be accomplished with attention to regional and local adaptations ofthe milkweeds to be planted. Only native milkweeds should be planted; for more aboutnatives see Monarch Watch (2018). Care must be taken especially with the non-nativetropical milkweed Asclepias curassavica L., which may not die back in southern states,thus potentially increasing the infection rate by protozoan parasites and possiblydisrupting monarch migratory behaviour (Satterfield et al. 2015). Expanding coverageof A. curassavica and loss of A. humistrata in Florida is occurring due to extensivecommercialization of A. curassavica and increasing loss of sandhill habitats. Also,Calotropis gigantea, W.T. Alton, an imported Old World milkweed, is being grown inGeorgia to feed monarch larvae (Winter 2017). We are concerned that these well-intentioned efforts to increase monarch reproduction may alter milkweeds and theirassociated communities and disrupt the strong evolutionary adaptation (Agrawal 2017)of the monarch’s migration to the native US milkweed flora.

Geographic sources and decline in abundance of the cross creek monarchs

Knight (1998) addressed the question of whether the Cross Creek spring monarchsoverwinter in Mexico or whether they may have come from some other sources bycarrying out cardenolide fingerprint analyses on 143 adult monarchs captured in CrossCreek, Florida, in April 1987, 1990, 1993, and 1994. She determined that 66% had fed aslarvae on A. syriaca, 15% on A. curassavica, 2% on an unknown milkweed, and 17% hadfingerprints too faint to assign but their foodplant was likely also A. syriaca. Because A.syriaca does not naturally occur south of North Carolina (Radford et al. 1968), themonarchs with the A. syriaca fingerprint captured in Cross Creek during April musthave developed in the north during the previous summer and therefore were springremigrants rather than a new generation of monarchs produced in south Florida orelsewhere in the tropics. Knight also fingerprinted 79 monarchs from two Florida GulfCoast overwintering populations, Cape San Blas (near Apalachicola) and HoneymoonIsland (west of Tampa). Thirteen San Blas monarchs collected in January 1988, 38Honeymoon Island monarchs collected in February 1982, and 28 collected in February1983, all had the A. syriaca fingerprint, thus indicating that they eclosed in the north andmigrated south during the previous fall to overwinter along the Florida Gulf Coast.

Two hypotheses can explain the A. curassavica butterflies captured at Cross Creek.They could have been monarchs that migrated in the spring from south Florida where A.curassavica is abundant (Malcolm and Brower 1986; Knight and Brower 2009; Dunfordet al. 2016) or they could have been monarchs that reproduced locally on A. curassavicaplants sold by nurseries. We favour the former hypothesis because our Cross Creek site isrelatively remote, and commercial A. curassavica plants were not widely available at thetime we collected the fingerprinted butterflies. Another possibility is that monarchsproduced an early spring generation on A. humistrata and other milkweeds growingfarther south in Florida, but this is largely ruled out by the fact that there were no A.humistrata butterflies and only three unknowns in Knight’s 143 April Cross Creekfingerprint samples.

While correlation is not causation, there is a growing body of evidence (Brower et al.2012; Flockhart et al. 2015; Zalucki et al. 2015; Semmens et al. 2016; Stenoien et al. 2016;

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Thogmartin et al. 2017a; 2017b; Pleasants et al. 2017; but see Inamine et al. 2016; andAgrawal 2017) supporting the contention that milkweed elimination in the midwesternUSA by the increasing annual application of glyphosate herbicide is a major contributorto the decline of monarch butterflies overwintering in Mexico. From 1997 to 2014, theamount of glyphosate applied in the USA to herbicide resistant corn and soybean cropsincreased from approximately 0.5 million to 100 million kg (USGS 2017). Pleasants (2017)estimated that since 1999 this widespread usage has caused the loss of 40% of A. syriacamilkweeds (more than 850 million individual plants) that previously had grown insidethe agricultural fields in the midwestern USA; furthermore, this loss of milkweeds inagricultural areas has led to a shift in monarch abundance towards non-agriculturalareas. It seems reasonable to hypothesize that the dwindling numbers of overwinteringmonarchs in Mexico is contributing to the dwindling numbers of monarchs reproducingeach spring in north central Florida. The decrease in adults/h and eggs/stem at CrossCreek has occurred at the same time that overwintering abundance in Mexico has beendeclining, although the decline at Cross Creek has been less steep than that for theoverwintering areas.

Conclusion

With regular monitoring from 1994 onward and supplemented by surveys from1983–1993, our survey over a total of 37 years is the longest continuing locality-basedmonarch population monitoring available. The survey has shown that the monarch’stiming of departure from Mexico at the end of their overwintering season is closelylinked to the spring phenology of A. humistrata in the south-eastern USA. Arrival is lateenough to avoid spring frosts that can kill back the freshly emergent milkweed and earlyenough to assure good plant quality for larval development. This timing is likely amanifestation of the evolved relationship of the North American milkweed flora andthe monarch’s migration biology. The long-term monitoring indicates that both adultsand immatures have declined in abundance since 1994 and strongly declined since2005. This timing correlates with the expanded use of glyphosate herbicide in themidwestern states (USGS 2017; Thogmartin et al. 2017b) where the majority of fallmigrants are produced late in the summer. Because of the possibility of altering therelationship between specific milkweeds and monarch migration, we urge caution incurrent conservation efforts to plant milkweeds widely. Seeds and seedlings should bederived from and replanted in their native biogeographic areas.

Acknowledgements

We thank Dana Griffin of the Florida State Museum for telling us the location of the A. humistratapopulation and the Hogan family, who since 1985 have generously allowed us annual access since1985 to a large population of this milkweed on their cattle farm near Cross Creek, FL. We especiallyappreciate the fact that the Hogans agreed not to use herbicides on the pasture through the durationof the study. We are grateful to the following individuals who participated in the monitoring over theyears: Martha Hoggard, Justin Saarinen, Kristin Rossetti Standridge, Joshua M. Epstein, the late AkersPence, Richard RuBino, Liz Rutkin, Barbara Cockrell, Stephen Lynch, Ron Martin, Alfonso Alonso,Cristina Dockx, Kay Eoff, Katie Barbara, Sheryl Hayes, David Ritland, Laurel Fox, Bruce Ferguson, andBob Platt. We thank Elizabeth Howard of Journey North for providing data on the spring remigration

2042 L.P. BROWER ET AL.

across the southern states. Mark Fishbein provided data on the number of milkweed species availableto monarchs in the USA and Canada. We are grateful to Linda S. Fink and to two outside reviewers forreviewing the manuscript and providing thoughtful suggestions.

Disclosure statement

No potential conflict of interest was reported by the authors.

Funding

This study has been supported by the National Science Foundation under grant DEB 0949650 toSweet Briar College, L.P. Brower, Principal Investigator, and by the October Hill Foundation.

Geolocation Information

Hogan’s pasture: 29°31.779 N, 82°11.878 W.

ORCID

Ernest H. Williams http://orcid.org/0000-0002-9438-103XMyron P. Zalucki http://orcid.org/0000-0001-9603-7577

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