Monarch Butterfly Ecology, Behavior, and Vulnerabilities in North Central United States Agricultural Landscapes.

The North American monarch butterfly (Danaus plexippus) is a candidate species for listing under the Endangered Species Act. Multiple factors are associated with the decline in the eastern population, including the loss of breeding and foraging habitat and pesticide use. Establishing habitat in agricultural landscapes of the North Central region of the United States is critical to increasing reproduction during the summer. We integrated spatially explicit modeling with empirical movement ecology and pesticide toxicology studies to simulate population outcomes for different habitat establishment scenarios. Because of their mobility, we conclude that breeding monarchs in the North Central states should be resilient to pesticide use and habitat fragmentation. Consequently, we predict that adult monarch recruitment can be enhanced even if new habitat is established near pesticide-treated crop fields. Our research has improved the understanding of monarch population dynamics at the landscape scale by examining the interactions among monarch movement ecology, habitat fragmentation, and pesticide use.

Breeding season temporal and spatial trends in continental-scale migration of the monarch butterfly.

The monarch butterfly (Danaus plexippus) is a vagile species that undertakes an annual, multi-generational migration across North America. The abundance of this species at both eastern and western overwintering sites in Central Mexico and California indicates a population decline. Success of continental-scale conservation programs for a migratory species depends on providing, maintaining, and protecting habitats at appropriate temporal and spatial scales. Here, dynamics of monarch continental-scale migration and gene flow were obtained by combined stable isotope, morphological, and genetic analyses. These analyses were applied to temporal monarch samples collected from May to September during 2016-2021 at locations in Iowa, USA and spatial collections from Pennsylvania, Delaware, Iowa, Ohio, Nevada, Idaho, Hawaii, 3 Australian locations during July and August 2016, and Texas in April 2021. Evidence of seasonal multi-generational migration was obtained through δ2H analyses of spatial collections, which was corroborated by decreased wing hue (a morphological marker for non-migratory individuals). In Iowa, 10-15% of monarchs represented migrants from southern areas throughout the breeding season and 6% were migrants from the North in midsummer. Limited sequence variation detected across the mitochondrial genome impacted the capability to detect significant population genetic variation in our North American samples. However, 2 novel substitutions were identified and predicted to be fixed among Australia samples, contributing to intercontinental differentiation from counterparts in North America. Our assessment of temporal and spatial population dynamics across the North American monarch breeding range provides insight into continental-scale migration and previously undetected mitochondrial DNA variation among globally distributed populations.

Monarch butterfly-breeding habitat restoration: how movement ecology research can inform best practices for site selection.

Population dynamics, persistence, and distribution are emergent properties of animal movement behavior and the spatial configuration of resources. Monarch butterflies are a vagile species with an open-population structure. Selecting locations for monarch butterfly- breeding habitat restoration that aligns with natural movement behavior will facilitate efficient habitat utilization across the landscape, increase realized fecundity, and ultimately support increases in the overwintering population size in Mexico. Obtaining and interpreting empirical movement and space-use data through field and laboratory studies are fundamental to this effort. To gain insights into population responses at larger, spatially explicit landscape scales, the results from empirical studies can be incorporated into simulation models. Together, empirical and simulation studies can inform options for creating functional connectivity of monarch butterfly-breeding habitats. Given currently available information, we synthesize studies for the eastern monarch butterfly to illustrate how an improved understanding of movement ecology can assist in planning conservation practices.

Estimating Perceptual Range of Female Monarch Butterflies (Danaus plexippus) to Potted Vegetative Common Milkweed (Asclepias syriaca) and Blooming Nectar Resources.

Habitat loss in the summer breeding range contributes to eastern North American monarch (Lepidoptera: Nymphalidae) population decline. Habitat restoration efforts include increasing native prairie plants for adult forage and milkweed (Asclepias spp.) for oviposition and larval development. As the monarch is a vagile species, habitat establishment at a grain that matches the monarch perceptual range will facilitate efficient movement, decrease fitness costs of dispersal, and increase oviposition. We released 188 experimental monarch females 5, 25, 50, and 75 m downwind from potted milkweed and blooming forbs in 4-32 ha sod fields. Perceptual range was estimated from monarchs that flew towards and landed on the milkweed and forbs. Flight patterns of 49 non-experimental monarchs that landed on the resources were also observed. In our experimental, resource-devoid setting, wind-facilitated movement occurred most frequently. Monarchs performed direct displacement as evidenced by shallow turn angles and similarity of Euclidian and total distances traveled. We hypothesize similar monarch flight behavior when traveling over other resource-devoid areas, such as crop fields. Although the majority of experimental monarchs flew downwind, eight experimental and 49 non-experimental monarchs were observed flying upwind toward, and landing on, the potted resources from distances ranging from 3 to 125 m (mean = 30.98 m, median = 25 m, mode = 25 m). A conservative estimate of the perceptual range is 125 m, as longer distances cannot be precluded; however, the majority of observations were ≤50 m. Our findings suggest establishing habitat patches ~ 50 m apart would create functional connectivity across fragmented agricultural landscapes.

Prediction of mitochondrial genome-wide variation through sequencing of mitochondrion-enriched extracts.

Although mitochondrial DNA (mtDNA) haplotype variation is often applied for estimating population dynamics and phylogenetic relationships, economical and generalized methods for entire mtDNA genome enrichment prior to high-throughput sequencing are not readily available. This study demonstrates the utility of differential centrifugation to enrich for mitochondrion within cell extracts prior to DNA extraction, short-read sequencing, and assembly using exemplars from eight maternal lineages of the insect species, Ostrinia nubilalis. Compared to controls, enriched extracts showed a significant mean increase of 48.2- and 86.1-fold in mtDNA based on quantitative PCR, and proportion of subsequent short sequence reads that aligned to the O. nubilalis reference mitochondrial genome, respectively. Compared to the reference genome, our de novo assembled O. nubilalis mitochondrial genomes contained 82 intraspecific substitution and insertion/deletion mutations, and provided evidence for correction of mis-annotated 28 C-terminal residues within the NADH dehydrogenase subunit 4. Comparison to a more recent O. nubilalis mtDNA assembly from unenriched short-read data analogously showed 77 variant sites. Twenty-eight variant positions, and a triplet ATT codon (Ile) insertion within ATP synthase subunit 8, were unique within our assemblies. This study provides a generalizable pipeline for whole mitochondrial genome sequence acquisition adaptable to applications across a range of taxa.

Plant Preferences of Z-Pheromone Race Ostrinia nubilalis (Lepidoptera: Crambidae) Based on Leaf Tissue Consumption Rates.

European corn borer, Ostrinia nubilalis Hübner (Lepidoptera: Crambidae), has been present in the United States for over 100 yr and documented on >200 plant species, including economically valuable crops. The reported preferred host of O. nubilalis is corn, Zea mays L. (Cyperales: Poaceae), although it is considered to be a generalist agricultural pest. Life cycles of the two pheromone races, E and Z, align with the seasonality of different agricultural plants. Since the introduction of Bt corn in 1996, overall O. nubilalis presence has declined and suggests that alternative crop plants might not be suitable hosts. We investigated plant vegetation preference of third-instar Z-race O. nubilalis for leaf disks of corn and a variety of other crops using 48 h no-choice and choice tests. Z-race larvae gained more mass on V6 non-Bt field corn leaf disks in comparison to other plant species options. Additionally, a preference for non-Bt field corn leaf disks was observed in most comparisons. Higher consumption of cucumber, Cucumis sativus L. (Cucurbitales: Cucurbitaceae), leaf disks as compared to non-Bt field corn leaf disks suggested an ability to feed on excised leaf tissues of a plant species that does not induce defenses to herbivory.

Employing Very High Frequency (VHF) Radio Telemetry to Recreate Monarch Butterfly Flight Paths.

The overwintering population of eastern North American monarch butterflies (Danaus plexippus) has declined significantly. Loss of milkweed (Asclepias sp.), the monarch's obligate host plant in the Midwest United States, is considered to be a major cause of the decline. Restoring breeding habitat is an actionable step towards population recovery. Monarch butterflies are highly vagile; therefore, the spatial arrangement of milkweed in the landscape influences movement patterns, habitat utilization, and reproductive output. Empirical studies of female movement patterns within and between habitat patches in representative agricultural landscapes support recommendations for habitat restoration. To track monarch movement at distances beyond human visual range, we employed very high frequency radio telemetry with handheld antennae to collect movement bearings on a biologically relevant time scale. Attachment of 220-300 mg transmitters did not significantly affect behavior and flight capability. Thirteen radio-tagged monarchs were released in a restored prairie, and locations were estimated every minute for up to 39 min by simultaneous triangulation from four operators. Monarchs that left the prairie were tracked and relocated at distances up to 250 m. Assuming straight flights between locations, the majority of steps within the prairie were below 50 m. Steps associated with exiting the prairie exceeded 50 m with high directionality. Because butterflies do not fly in straight lines between stationary points, we also illustrate how occurrence models can use location data obtained through radio telemetry to estimate movement within a prairie and over multiple land cover types.

Survivorship of Z-Pheromone Race European Corn Borer (Lepidoptera: Crambidae) on a Range of Host Plants Varying in Defensive Chemistry.

The European corn borer, Ostrinia nubilalis (Hübner), was introduced in North America in the early 1900s and became a major pest of corn. After its introduction, it was found on > 200 other plant hosts, but corn remained its primary host. Early life history studies indicated that European corn borer had the potential of a wide host range. For nearly 80 yr before the introduction of Bt corn, the European corn borer was a major pest of corn in North America. This study investigated the growth and survivorship of the Z-pheromone race European corn borer on a range of hosts that vary in defensive chemistries and historic degree of infestation to better understand the current host plant range of Z-pheromone race of O. nubilalis. The plants tested include sweet corn, cry1F Bt field corn, non-Bt corn, cucumber, tomato, and green bean. Experiments were conducted in the growth chamber, greenhouse, and field to determine survival under different conditions. In most cases, results supported the expected outcome, with significantly higher survival on non-Bt corn hosts than the other hosts tested. Neonate larvae fed exclusively on leaves of green bean exhibited intermediate survival, whereas third-instars fed on only leaves of cucumber survived intermediately. Larvae on Bt corn and tomato had comparable low survival rates, overall suggesting that the defensive features of tomato are about as effective as Cry1F Bt corn. Non-Bt corn was found to be the most suitable host plant, overall for European corn borer among those tested.