Gravisensation and modulation of gravitactic responses by other sensory cues in the monarch butterfly (Danaus plexippus).

Using the monarch butterfly (Danaus plexippus), we studied how animals can use cues from multiple sensory modalities for deriving directional information from their environment to display oriented movement. Our work focused on determining how monarchs use gravity as a cue for oriented movement and determined how cues from other sensory modalities, cues that by themselves also produce oriented movement (visual and magnetic directional cues), might modulate gravisensation. In two tests of gravisensation (movement in a vertical tube; righting behavior), we found that monarchs display negative gravitaxis only (movement opposite to the direction of gravity). Negative gravitaxis can be modulated by either visual (light) or magnetic field cues (inclination angle) that provide directional information. The modulation of gravity-mediated responses, however, depends on the relationship between cues when presented during trials, such as when cues are in accord or in conflict. For example, when light cues that elicit positive phototaxis conflicted with negative gravitaxis (light from below the monarch), monarch gravisensation was unaffected by directional light cues. We also found that the antennae play a role in gravity-mediated movement (righting), as, with antennae removed, monarch movement behavior was no longer the same as when the antennae were intact. Our results demonstrate that monarchs can use and integrate multiple, multimodal cues for oriented movement, but that the use of such cues can be hierarchical (that is, one cue dominant for movement), and the hierarchy of cues, and the responses towards them when found together, depends on the physical relationships between cues during movement.

The cost of movement: assessing energy expenditure in a long-distant ectothermic migrant under climate change.

Migration is an energetically taxing phenomenon as animals move across vast, heterogeneous landscapes where the cost of transport is impacted by permissible ambient conditions. In this study, we assessed the energetic demands of long-distance migration in a multigenerational ectothermic migrant, the monarch butterfly (Danaus plexippus). We tested the hypotheses that temperature-dependent physiological processes reduce energy reserves faster during migration than previously estimated, and that increasing climatic temperatures resulting from the climate crisis will intensify baseline daily energy expenditure. First, we reared monarchs under laboratory conditions to assess energy and mass conversion from fifth instar to adult stages, as a baseline for migratory adult mass and ontogenetic shifts in metabolic rate from larvae to adult. Then, using historical tag-recapture data, we estimated the movement propensity and migratory pace of autumn migrants using computer simulations and subsequently calculated energy expenditure. Finally, we estimated the energy use of monarchs based on these tag-recapture data and used this information to estimate daily energy expenditure over a 57 year period. We found support for our two hypotheses, noting that incorporating standard metabolic rate into estimates of migratory energy expenditure shows higher energy demand and that daily energy expenditure has been gradually increasing over time since 1961. Our study shows the deleterious energetic consequences under current climate change trajectories and highlights the importance of incorporating energetic estimates for understanding migration by small, ectothermic migrants.

Lack of evidence for a fine-scale magnetic map sense for fall migratory Eastern North American monarch butterflies (Danaus plexippus).

How first-time animal migrants find specific destinations remains an intriguing ecological question. Migratory marine species use geomagnetic map cues acquired as juveniles to aide long-distance migration, but less is known for long-distance migrants in other taxa. We test the hypothesis that naïve Eastern North American fall migratory monarch butterflies (Danaus plexippus), a species that possesses a magnetic sense, locate their overwintering sites in Central Mexico using inherited geomagnetic map cues. We examined whether overwintering locations and the abundance of monarchs changed with the natural shift of Earth's magnetic field from 2004 to 2018. We found that migratory monarchs continued to overwinter at established sites in similar abundance despite significant shifts in the geomagnetic field, which is inconsistent with monarchs using fine-scale geomagnetic map cues to find overwintering sites. It is more likely that monarchs use geomagnetic cues to assess migratory direction rather than location and use other cues to locate overwintering sites.

Oriented migratory flight at night: Consequences of nighttime light pollution for monarch butterflies.

We show that light trespass-a form of nighttime light pollution (NLP)-elicits normal daytime clock-mediated migratory behavior in fall monarch butterflies during their night-cycle. In controlled indoor flight simulator studies isolating the role of NLP on the expression of oriented migratory flight using a time-compensated sun compass,a full-spectrum light source consistent with lights used outdoors at night by the public,triggered proper fall directional flight at night in monarchs. Monarchs remained quiescent when initially placed in the flight simulator in the dark, but flight was immediately triggered when our light source was turned on. This nighttime behavior was identical to that seen in outdoor free-flying fall conspecifics during the day. The light source provided directional cues equivalent to those provided by the sun and could either phase-advance or phase-delay monarchs. Our study highlights the negative consequences of NLP on diurnal animals, especially those that rely on clock-mediated behavior.

A behavioral assay to test sensory-cue-guided oriented flight in monarch butterflies under controlled conditions.

Many animals use sensory cues to guide movement. Testing animals under conditions in which cues can be isolated and manipulated is key for understanding the function of cues. Here, we present a protocol to assess the flight of migratory monarch butterflies (Danaus plexippus). We describe procedures to optimize and conduct trials, especially under indoor conditions. This protocol facilitates testing monarchs in various experimental conditions including during their subjective night when they are not normally flying. For complete details on the use and execution of this protocol, please refer to Parlin et al. (2022).1.

Dimorphic cocoons of the robin moth, Hyalophora cecropia, reflect the existence of two distinct architectural syndromes.

The architectural design of animal structures forms part of an individual's extended phenotype that can be subjected to strong selection pressures. We examined cocoon architectural dimorphism in robin moths (Hyalophora cecropia), which construct multilayered silk-woven cocoons that possess either a 'baggy' or 'compact' morphology. These dimorphic cocoons reflect extended phenotypes that can enable survival during a critical developmental period (pupal stage to adult emergence), with cocoons occurring either sympatrically or as monomorphic groups across different climatic regions in North America. We hypothesized that cocoon dimorphism is related to the cocoon's role as a mediating barrier for moisture. We predicted that the macro- and micro-architectural differences between the cocoon morphs would be consistent with this function. We compared the cocoon morphs in terms of their orientation when spun under natural field conditions, examined how these orientations affected cocoon water absorption under simulated rain trials, and performed material surface tests to compare the hydrophobicity of cocoons. We found that compact cocoons had traits that increased water resistance, as they had significantly greater hydrophobicity than baggy cocoons, because they absorbed less water and released water vapor faster. In contrast, the increased water absorptiveness of baggy cocoons can allow for greater moisture retention, a function related to the prevention of desiccation. Our study suggests that cocoon dimorphism in robin moths reflects distinct architectural syndromes, in which cocoons are spun to optimize either water resistance or retention. These different functions are consistent with strategies that act to respond to uncertain external environmental conditions that an individual might encounter during development.

The Monarch Butterfly as a Model for Understanding the Role of Environmental Sensory Cues in Long-Distance Migratory Phenomena.

The awe-inspiring annual migration of monarch butterflies (Danaus plexippus) is an iconic example of long-distance migratory phenomena in which environmental sensory cues help drive successful migration. In this mini-review article, I begin by describing how studies on monarch migration can provide us with generalizable information on how sensory cues can mediate key aspects of animal movement. I describe how environmental sensory cues can trigger the development and progression of the monarch migration, as well as inform sensory-based movement mechanisms in order to travel to and reach their goal destination, despite monarchs being on their maiden voyage. I also describe how sensory cues can trigger season-appropriate changes in migratory direction during the annual cycle. I conclude this mini-review article by discussing how contemporary environmental challenges threaten the persistence of the monarch migration. Environmental challenges such as climate change and shifting land use can significantly alter the sensory environments that monarchs migrate through, as well as degrade or eliminate the sources of sensory cues that are necessary for successful migration.

A laboratory-based study examining the properties of silk fabric to evaluate its potential as a protective barrier for personal protective equipment and as a functional material for face coverings during the COVID-19 pandemic.

The worldwide shortage of single-use N95 respirators and surgical masks due to the COVID-19 pandemic has forced many health care personnel to use their existing equipment for as long as possible. In many cases, workers cover respirators with available masks in an attempt to extend their effectiveness against the virus. Due to low mask supplies, many people instead are using face coverings improvised from common fabrics. Our goal was to determine what fabrics would be most effective in both practices. Under laboratory conditions, we examined the hydrophobicity of fabrics (cotton, polyester, silk), as measured by their resistance to the penetration of small and aerosolized water droplets, an important transmission avenue for the virus causing COVID-19. We also examined the breathability of these fabrics and their ability to maintain hydrophobicity despite undergoing repeated cleaning. Laboratory-based tests were conducted when fabrics were fashioned as an overlaying barrier for respirators and when constructed as face coverings. When used as material in these two situations, silk was more effective at impeding the penetration and absorption of droplets due to its greater hydrophobicity relative to other tested fabrics. We found that silk face coverings repelled droplets in spray tests as well as disposable single-use surgical masks, and silk face coverings have the added advantage over masks such that they can be sterilized for immediate reuse. We show that silk is a hydrophobic barrier to droplets, can be more breathable than other fabrics that trap humidity, and are re-useable via cleaning. We suggest that silk can serve as an effective material for making hydrophobic barriers that protect respirators, and silk can now be tested under clinical conditions to verify its efficacy for this function. Although respirators are still the most appropriate form of protection, silk face coverings possess properties that make them capable of repelling droplets.

Architectural evolution in cocoons spun by Hyalophora (Lepidoptera; Saturniidae) silk moth species.

Caterpillars of the silk moth genus Hyalophora (Lepidoptera; Saturniidae) construct multilayered cocoons that function as overwintering housing during the pupal to adult developmental period. While all cocoons share the primary function of protecting the developing moth, cocoons spun by different Hyalophora silk moth species vary significantly in architectural features and in the level of intraspecific cocoon-type polymorphism. We compared the cocoons of Hyalophora species found throughout North America and investigated the evolution of architectural variation. We first characterized and compared the architectural features of cocoons at all three cocoon sections (outer envelope, inner envelope, and the intermediate section that separates them), and found that variation in the outer envelope underlies the differences in architecture between cocoons. Phylogenetic analysis indicates ancestral polymorphic architecture (both "baggy" and "compact" morphs), with diversification within Hyalophora for both monomorphic "compact" cocoons, and increased intermediate space and silk in "baggy" lineages. The evolution of these traits suggests a potential functional role for the different cocoon architectural forms.

A re-evaluation of silk measurement by the cecropia caterpillar (Hyalophora cecropia) during cocoon construction reveals use of a silk odometer that is temporally regulated.

The late 5th instar caterpillar of the cecropia silk moth (Hyalophora cecropia) spins a silken cocoon with a distinct, multilayered architecture. The cocoon construction program, first described by the seminal work of Van der Kloot and Williams, consists of a highly ordered sequence of events. We perform behavioral experiments to re-evaluate the original cecropia work, which hypothesized that the length of silk that passes through the spinneret controls the orderly execution of each of the discrete events of cocoon spinning. We confirm and extend by three-dimensional scanning and quantitative measurements of silk weights that if cocoon construction is interrupted, upon re-spinning, the caterpillar continues the cocoon program from where it left off. We also confirm and extend by quantitative measurements of silk weights that cecropia caterpillars will not bypass any of the sections of the cocoon during the construction process, even if presented with a pre-spun section of a cocoon spun by another caterpillar. Blocking silk output inhibits caterpillars from performing normal spinning behaviors used for cocoon construction. Surprisingly, unblocking silk output 24-hr later did not restart the cocoon construction program, suggesting the involvement of a temporally-defined interval timer. We confirm with surgical reductions of the silk glands that it is the length of silk itself that matters, rather than the total amount of silk extracted by individuals. We used scanning electron microscopy to directly show that either mono- or dual-filament silk (i.e., equal silk lengths but which vary in their total amount of silk extracted) can be used to construct equivalent cocoons of normal size and that contain the relevant layers. We propose that our findings, taken together with the results of prior studies, strongly support the hypothesis that the caterpillar uses a silk "odometer" to measure the length of silk extracted during cocoon construction but does so in a temporally regulated manner. We further postulate that our examination of the anatomy of the silk spinning apparatus and ablating spinneret sensory output provides evidence that silk length measurement occurs upstream of output from the spinneret.

Dimorphic cocoons of the cecropia moth (Hyalophora cecropia): Morphological, behavioral, and biophysical differences.

The larvae of the giant silk moth (Hyalophora cecropia) spin strikingly dimorphic, multilayered cocoons that are either large and fluffy (baggy) or significantly smaller and tightly woven (compact). Although these cocoon-morphs share the same function (i.e., housing for pupal to adult development during overwintering), previous work has been unable to determine why cocoon dimorphism exists. We addressed this issue in cecropia moth cocoons collected along power line right-of-way habitats in Massachusetts. We first characterized the architectural differences between cocoon-morphs for all three cocoon sections (outer and inner envelopes, and the intermediate layer separating the two). We show that outer envelope structural and ultrastructural differences are what underlie dimorphism. Using a common spinning arena, we next show that the behavioral suites used to construct the outer envelopes of the two morphs are significantly different in behavioral time investment and patterning, as well as in the location of silk placement in the common spinning arena. Finally, we compared the cocoon-morphs in response to various environmental stressors to ask whether dimorphism is an adaptive response to such pressures. In contrast to compact cocoons, we find that baggy cocoons act as heat sinks and allow greater moisture permeability; differences in outer envelope architecture underlie these characteristics. These two biophysical properties could be advantageous for pupae in baggy cocoons, during unseasonably cold or dry conditions encountered during development prior to adult emergence. Our results suggest that cocoon dimorphism in the cecropia moth may provide a bet-hedging strategy for dealing with varying environmental conditions in Massachusetts and perhaps over its entire habitat range, during pupal to adult development.

Neurobiology of Monarch Butterfly Migration.

Studies of the migration of the eastern North American monarch butterfly (Danaus plexippus) have revealed mechanisms behind its navigation. The main orientation mechanism uses a time-compensated sun compass during both the migration south and the remigration north. Daylight cues, such as the sun itself and polarized light, are processed through both eyes and integrated through intricate circuitry in the brain's central complex, the presumed site of the sun compass. Monarch circadian clocks have a distinct molecular mechanism, and those that reside in the antennae provide time compensation. Recent evidence shows that migrants can also use a light-dependent inclination magnetic compass for orientation in the absence of directional daylight cues. The monarch genome has been sequenced, and genetic strategies using nuclease-based technologies have been developed to edit specific genes. The monarch butterfly has emerged as a model system to study the neural, molecular, and genetic basis of long-distance animal migration.

Sensory basis of lepidopteran migration: focus on the monarch butterfly.

In response to seasonal habitats, migratory lepidopterans, exemplified by the monarch butterfly, have evolved migration to deal with dynamic conditions. During migration, monarchs use orientation mechanisms, exploiting a time-compensated sun compass and a light-sensitive inclination magnetic compass to facilitate fall migration south. The sun compass is bidirectional with overwintering coldness triggering the change in orientation direction for remigration northward in the spring. The timing of the remigration and milkweed emergence in the southern US have co-evolved for propagation of the migration. Current research is uncovering the anatomical and molecular substrates that underlie migratory-relevant sensory mechanisms with the antennae being critical components. Orientation mechanisms may be detrimentally affected by environmental factors such as climate change and sensory interference from human-generated sources.

A magnetic compass aids monarch butterfly migration.

Convincing evidence that migrant monarch butterflies (Danaus plexippus) use a magnetic compass to aid their fall migration has been lacking from the spectacular navigational capabilities of this species. Here we use flight simulator studies to show that migrants indeed possess an inclination magnetic compass to help direct their flight equatorward in the fall. The use of this inclination compass is light-dependent utilizing ultraviolet-A/blue light between 380 and 420 nm. Notably, the significance of light <420 nm for inclination compass function was not considered in previous monarch studies. The antennae are important for the inclination compass because they appear to contain light-sensitive magnetosensors. For migratory monarchs, the inclination compass may serve as an important orientation mechanism when directional daylight cues are unavailable and may also augment time-compensated sun compass orientation for appropriate directionality throughout the migration.

Coldness triggers northward flight in remigrant monarch butterflies.

Each fall, eastern North American monarch butterflies (Danaus plexippus) migrate from their northern range to their overwintering grounds in central Mexico. Fall migrants are in reproductive diapause, and they use a time-compensated sun compass to navigate during the long journey south. Eye-sensed directional cues from the daylight sky (e.g., the horizontal or azimuthal position of the sun) are integrated in the sun compass in the midbrain central complex region. Sun compass output is time compensated by circadian clocks in the antennae so that fall migrants can maintain a fixed flight direction south. In the spring, the same migrants remigrate northward to the southern United States to initiate the northern leg of the migration cycle. Here we show that spring remigrants also use an antenna-dependent time-compensated sun compass to direct their northward flight. Remarkably, fall migrants prematurely exposed to overwintering-like coldness reverse their flight orientation to the north. The temperature microenvironment at the overwintering site is essential for successful completion of the migration cycle, because without cold exposure, aged migrants continue to orient south. Our discovery that coldness triggers the northward flight direction in spring remigrants solves one of the long-standing mysteries of the monarch migration.

Discordant timing between antennae disrupts sun compass orientation in migratory monarch butterflies.

To navigate during their long-distance migration, monarch butterflies (Danaus plexippus) use a time-compensated sun compass. The sun compass timing elements reside in light-entrained circadian clocks in the antennae. Here we show that either antenna is sufficient for proper time compensation. However, migrants with either antenna painted black (to block light entrainment) and the other painted clear (to permit light entrainment) display disoriented group flight. Remarkably, when the black-painted antenna is removed, re-flown migrants with a single, clear-painted antenna exhibit proper orientation behaviour. Molecular correlates of clock function reveal that period and timeless expression is highly rhythmic in brains and clear-painted antennae, while rhythmic clock gene expression is disrupted in black-painted antennae. Our work shows that clock outputs from each antenna are processed and integrated together in the monarch time-compensated sun compass circuit. This dual timing system is a novel example of the regulation of a brain-driven behaviour by paired organs.

Evaluating the life-history trade-off between dispersal capability and reproduction in wing dimorphic insects: a meta-analysis.

A life-history trade-off exists between flight capability and reproduction in many wing dimorphic insects: a long-winged morph is flight-capable at the expense of reproduction, while a short-winged morph cannot fly, is less mobile, but has greater reproductive output. Using meta-analyses, I investigated specific questions regarding this trade-off. The trade-off in females was expressed primarily as a later onset of egg production and lower fecundity in long-winged females relative to short-winged females. Although considerably less work has been done with males, the trade-off exists for males among traits primarily related to mate acquisition. The trade-off can potentially be mitigated in males, as long-winged individuals possess an advantage in traits that can offset the costs of flight capability such as a shorter development time. The strength and direction of trends differed significantly among insect orders, and there was a relationship between the strength and direction of trends with the relative flight capabilities between the morphs. I discuss how the trade-off might be both under- and overestimated in the literature, especially in light of work that has examined two relevant aspects of wing dimorphic species: (1) the effect of flight-muscle histolysis on reproductive investment; and (2) the performance of actual flight by flight-capable individuals.

Flight behaviour attenuates the trade-off between flight capability and reproduction in a wing polymorphic cricket.

Flight-dimorphic insects have been used extensively to study trade-offs between energetically costly traits. Individuals may develop and maintain structures required for flight, or alternatively they may invest in reproduction. Previous experiments have not examined whether flight itself might affect investment into reproduction. As in other Gryllus species, flight-capable individuals of the wing polymorphic cricket, Gryllus texensis, incur an apparent reproductive penalty for being able to fly, expressed as smaller ovaries in females and lower courtship propensity in males, than their flight-incapable counterparts. We find that a short bout of flight eliminates the trade-off. Two days after the flight, the ovaries of flight-capable females were comparable with those of short-winged females. Similarly, flight markedly increased the probability of courtship behaviour. Our results suggest that the impact of the flight-reproduction trade-off described in earlier studies may have been overestimated.