Inbreeding depression in monarch butterflies

Monarch butterflies and their unique system of multigenerational migration have long fascinated the public, and concerns for the fate of this charismatic insect have grown due to the consistent declines in overwintering colony size over the last 20 years. Risks to this migratory insect have been considered in terms of climate change, habitat and thus population fragmentation, and decreased host plant availability. However, another obvious danger, that of decreased heterozygosity resulting from decreasing population size, has yet to be explored. Here we report experimental evidence for immediate inbreeding depression in individuals from the migratory population. Inbred matings produced less viable eggs and inbred offspring had higher developmental mortality and shorter lifespans. We discuss these results in the context of monarch migration extinction risk and suggest that additional genetic monitoring should be undertaken to protect this iconic animal.     

Fire creates host plant patches for monarch butterflies

Monarch butterflies (Danaus plexippus) depend on the presence of host plants (Asclepias spp.) within their breeding range for reproduction. In the southern Great Plains, Asclepias viridis is a perennial that flowers in May and June, and starts to senesce by August. It is locally abundant and readily used by monarchs as a host plant. We evaluated the effects of summer prescribed fire on A. viridis and the use of A. viridis by monarch butterflies. Summer prescribed fire generated a newly emergent population of A. viridis that was absent in other areas. Pre-migrant monarch butterflies laid eggs on A. viridis in summer burned plots in late August and September, allowing adequate time for a new generation of adult monarchs to emerge and migrate south to their overwintering grounds. Thus, summer prescribed fire may provide host plant patches and/or corridors for pre-migrant monarchs during a time when host plant availability may be limited in other areas.     

Sun compass integration of skylight cues in migratory monarch butterflies

Migrating monarch butterflies (Danaus plexippus) use a time-compensated sun compass to navigate from eastern North America to their overwintering grounds in central Mexico. Here we describe the neuronal layout of those aspects of the butterfly's central complex likely to establish part of the internal sun compass and find them highly homologous to those of the desert locust. Intracellular recordings from neurons in the monarch sun compass network reveal responses tuned to specific E-vector angles of polarized light, as well as azimuth-dependent responses to unpolarized light, independent of spectral composition. The neural responses to these two stimuli in individual neurons are mediated through different regions of the compound eye. Moreover, these dual responses are integrated to create a consistent representation of skylight cues in the sun compass throughout the day. The results advance our understanding of how ambiguous sensory signals are processed by the brain to elicit a robust behavioral response.     

Lack of genetic differentiation between monarch butterflies with divergent migration destinations

Monarch butterflies are best known for their spectacular annual migration from eastern North America to Mexico. Monarchs also occur in the North American states west of the Rocky Mountains, from where they fly shorter distances to the California Coast. Whether eastern and western North American monarchs form one genetic population or are genetically differentiated remains hotly debated, and resolution of this debate is essential to understand monarch migration patterns and to protect this iconic insect species. We studied the genetic structure of North American migratory monarch populations, as well as nonmigratory populations in Hawaii and New Zealand. Our results show that eastern and western migratory monarchs form one admixed population and that monarchs from Hawaii and New Zealand have genetically diverged from North American butterflies. These findings suggest that eastern and western monarch butterflies maintain their divergent migrations despite genetic mixing. The finding that eastern and western monarchs form one genetic population also suggests that the conservation of overwintering sites in Mexico is crucial for the protection of monarchs in both eastern and western North America.     

Juvenile hormone stimulation of sperm activator production in male monarch butterflies

A sperm activating substance is demonstrable in the male Monarch butterfly ejaculatory duct, but only during reproductively active stages. Experiments using neck-ligatures and allatectomies, with or without injections of juvenile hormone isomers, show that the post-eclosion production of sperm activator is stimulated by juvenile hormone.     

Defining behavioral and molecular differences between summer and migratory monarch butterflies

Background  

In the fall, Eastern North American monarch butterflies (Danaus plexippus) undergo a magnificent long-range migration. In contrast to spring and summer butterflies, fall migrants are juvenile hormone deficient, which leads to reproductive arrest and increased longevity. Migrants also use a time-compensated sun compass to help them navigate in the south/southwesterly direction en route for Mexico. Central issues in this area are defining the relationship between juvenile hormone status and oriented flight, critical features that differentiate summer monarchs from fall migrants, and identifying molecular correlates of behavioral state.     

Energy reserves and metabolic expenditures of monarch butterflies overwintering in southern California

Abstract. 1. Energetic expenditure and predicted requirements for overwintering metabolism were determined for monarch butterflies ( Danaus plexippus L.) in southern California.
2. Fat content of butterflies declined steadily from a maximum of 71% lean dry weight in late November to a minimum of 36% lean dry weight in late January. The energy expenditure measured by fat depletion over a 61 day period from 24 November to 25 January was 26.05 joules per day.
3. Butterflies were generally the same temperature as the air at any time they were inactive, whether they were part of a large cluster or roosting solitarily.
4. Oxygen consumption of butterflies increased in a curvilinear fashion with increasing air temperature. Thus, the predicted metabolic requirements for an inactive butterfly during their quiescent period from late November to late January was 30.32 joules per day.
5. In contrast to the steady depletion of fat reserves during their quiescent period in December and January, low and stable fat levels of butterflies in late February coincide with high levels of flight activity, mating and emigration of females from the colony.
6. Progressive tightening of the energy balance due to increasing nocturnal temperatures from northern to southern California coastal areas may limit the southern distribution and duration of overwintering aggregations.     

Sex differences in immune defenses and response to parasitism in monarch butterflies

Host susceptibility and patterns of infection are predicted to differ between males and females due to sex-based tradeoffs between the demands of reproduction and costly immune defenses. In this study, we examined immune defenses and the response to experimental infection by a protozoan parasite, Ophryocystis elektroscirrha, in male and female monarch butterflies, Danaus plexippus. We quantified two measures of immunity in late instar larvae: the concentration of circulating hemocytes and mid-gut phenoloxidase activity, and also quantified final parasite loads, body size, longevity, and wing melanism of adult butterflies. Results showed that females had greater average hemocyte counts than males in the absence of infection; males, but not females, showed an increased concentration of hemocytes in the presence of infection. However, higher hemocyte concentrations in larvae were not significantly correlated with lower adult parasite loads, and mid-gut phenoloxidase activity was not significantly associated with hemocyte counts or parasite treatments. Among unparasitized females, greater hemocyte concentrations were costly in terms of reduced body size, but for parasite-treated females, hemocyte concentrations and body size were positively associated. Across all monarchs, unparasitized butterflies showed greater wing melanism (darker forewings) than parasitized monarchs. Overall, this study provides support for differential costs of immune defenses in male and female monarch butterflies, and a negative association between parasite infection and monarch wing melanism.     

The redder the better: wing color predicts flight performance in monarch butterflies

The distinctive orange and black wings of monarchs (Danaus plexippus) have long been known to advertise their bitter taste and toxicity to potential predators. Recent work also showed that both the orange and black coloration of this species can vary in response to individual-level and environmental factors. Here we examine the relationship between wing color and flight performance in captive-reared monarchs using a tethered flight mill apparatus to quantify butterfly flight speed, duration and distance. In three different experiments (totaling 121 individuals) we used image analysis to measure body size and four wing traits among newly-emerged butterflies prior to flight trials: wing area, aspect ratio (length/width), melanism, and orange hue. Results showed that monarchs with darker orange (approaching red) wings flew longer distances than those with lighter orange wings in analyses that controlled for sex and other morphometric traits. This finding is consistent with past work showing that among wild monarchs, those sampled during the fall migration are darker in hue (redder) than non-migratory monarchs. Together, these results suggest that pigment deposition onto wing scales during metamorphosis could be linked with traits that influence flight, such as thorax muscle size, energy storage or metabolism. Our results reinforce an association between wing color and flight performance in insects that is suggested by past studies of wing melansim and seasonal polyphenism, and provide an important starting point for work focused on mechanistic links between insect movement and color.     

Serial founder effects and genetic differentiation during worldwide range expansion of monarch butterflies

Range expansions can result in founder effects, increasing genetic differentiation between expanding populations and reducing genetic diversity along the expansion front. However, few studies have addressed these effects in long-distance migratory species, for which high dispersal ability might counter the effects of genetic drift. Monarchs (Danaus plexippus) are best known for undertaking a long-distance annual migration in North America, but have also dispersed around the world to form populations that do not migrate or travel only short distances. Here, we used microsatellite markers to assess genetic differentiation among 18 monarch populations and to determine worldwide colonization routes. Our results indicate that North American monarch populations connected by land show limited differentiation, probably because of the monarch''s ability to migrate long distances. Conversely, we found high genetic differentiation between populations separated by large bodies of water. Moreover, we show evidence for serial founder effects across the Pacific, suggesting stepwise dispersal from a North American origin. These findings demonstrate that genetic drift played a major role in shaping allele frequencies and created genetic differentiation among newly formed populations. Thus, range expansion can give rise to genetic differentiation and declines in genetic diversity, even in highly mobile species.     

The relationship between mating and oogenesis in monarch butterflies (Lepidoptera: Danainae)

We studied the relationship between the timing of mating and oogenesis in monarch butterflies (Danaus plexippus) to determine (1) the potential for male nutrient input into eggs and (2) whether mating stimulates egg development. Most females mated soon after they started maturing eggs. One and 2 days after mating, females contained the same number of mature oocytes as virgin females of the same age, while 3 days after mating they contained more mature oocytes than did virgins. These results confirm the potential for male-derived nutrients to augment oocyte production, but indicate that mating is not required for oocyte maturation to occur.     

Patterns of parasitism in monarch butterflies during the breeding season in eastern North America

1. Migratory behaviour can result in reduced prevalence of pathogens in host populations. Two hypotheses have been proposed to explain this relationship: (i) ‘migratory escape’, where migrants benefit from escaping pathogen accumulation in contaminated environments; and (ii) ‘migratory culling’, where the selective removal of infected individuals occurs during migration. 2. In the host–parasite system between the monarch butterfly (Danaus plexippus Linn.) and its obligate protozoan parasite Ophryocystis elektroscirrha (OE), there is evidence to support both hypotheses, particularly during the monarchs' autumn migration. However, these processes can operate simultaneously and could vary throughout the monarchs' annual migratory cycle. Assessing the relative strength for each hypothesis has not previously been done. 3. To evaluate both hypotheses, parasite infection prevalence was examined in monarchs sampled in eastern North America during April–September, and stable isotopes (δ²H, δ¹³C) were used to estimate natal origin and infer migration distance. There was stronger support for the migratory escape hypothesis, wherein infection prevalence increased over the breeding season and was higher at southern latitudes, where the breeding season tends to be longer compared with northern latitudes. Little support was found for the migratory culling hypothesis, as infection prevalence was similar whether monarchs travelled shorter or longer distances. 4. These results suggest that migration allows individuals to escape parasites not only during the autumn, as shown in previous work, but during the monarchs' spring and summer movements when they recolonise the breeding range. These results imply a potential fitness advantage to monarchs that migrate further north to exploit parasite‐free habitats.     

Biotic predictors with phenological information improve range estimates for migrating monarch butterflies in Mexico

Although long-standing theory suggests that biotic variables are only relevant at local scales for explaining the patterns of species' distributions, recent studies have demonstrated improvements to species distribution models (SDMs) by incorporating predictor variables informed by biotic interactions. However, some key methodological questions remain, such as which kinds of interactions are permitted to include in these models, how to incorporate the effects of multiple interacting species, and how to account for interactions that may have a temporal dependence. We addressed these questions in an effort to model the distribution of the monarch butterfly Danaus plexippus during its fall migration (September–November) through Mexico, a region with new monitoring data and uncertain range limits even for this well-studied insect. We estimated species richness of selected nectar plants (Asclepias spp.) and roosting trees (various highland species) for use as biotic variables in our models. To account for flowering phenology, we additionally estimated nectar plant richness of flowering species per month. We evaluated three types of models: climatic variables only (abiotic), plant richness estimates only (biotic) and combined (abiotic and biotic). We selected models with AICc and additionally determined if they performed better than random on spatially withheld data. We found that the combined models accounting for phenology performed best for all three months, and better than random for discriminatory ability but not omission rate. These combined models also produced the most ecologically realistic spatial patterns, but the modeled response for nectar plant richness matched ecological predictions for November only. These results represent the first model-based monarch distributional estimates for the Mexican migration route and should provide foundations for future conservation work. More generally, the study demonstrates the potential benefits of using SDM-derived richness estimates and phenological information for biotic factors affecting species distributions.     

Beyond preference and performance: host plant selection by monarch butterflies,Danaus plexippus

The connection between adult preferences and offspring performance is a long‐standing issue in understanding the evolutionary and ecological forces that dictate host associations and specialization in herbivorous insects. Indeed, decisions made by females about where to lay their eggs have direct consequences for fitness and are influenced by interacting factors including offspring performance, defence and competition. Nonetheless, in addition to these attributes of the offspring, a female's choices may be affected by her own prior experience. Here we examined oviposition preference, larval performance and the role of learning in the monarch butterfly, Danaus plexippus, which encounters diverse milkweed host species across its broad range and over the course of migration. Monarch females consistently preferred to oviposit on Asclepias incarnata subspecies pulchra. This plant, however, was associated with poor caterpillar growth, low sequestration of toxins and the highest plant defences (latex and trichomes). We examined flexibility in this apparently maladaptive preference by testing the impact of previous experience and competition on preference. Experience laying on an alternative plant species enhanced preference for that species in contrast to A. i. pulchra. In addition, presence of a (competing) conspecific caterpillar on A. i. pulchra had a strongly deterrent effect and reversed host plant preferences. Thus, monarch butterflies exhibit preferences contrary to what would be expected based on offspring development and sequestered defences, but their preferences are altered by learning and competition, which may allow butterflies to shift preferences as they encounter diverse milkweeds across the landscape. Learning and perception of threats (i.e. competition or predation) may be critical for most herbivorous insects, which universally experience heterogeneity among their potential host plants.     

Oviposition preference and larval performance of North American monarch butterflies on four Asclepias species

Monarch butterflies, Danaus plexippus L. (Lepidoptera: Nymphalidae), occur world‐wide and are specialist herbivores of plants in the milkweed family (Asclepiadaceae). In North America, two monarch populations breed east and west of the continental divide in areas populated by different host plant species. To examine the population variation in monarch responses to different Asclepias species, we measured oviposition preference and larval performance among captive progeny reared from adult butterflies collected in eastern and western North America. Host plant use was evaluated using two milkweed species widely distributed in eastern North America (A. incarnata and A. syriaca), and two species common to western North America (A. fascicularis and A. speciosa). We predicted that exposure to different host plant species in their respective breeding ranges could select for divergent host use traits, so that monarchs should preferentially lay more eggs on, and larvae should perform better on, milkweed species common to their native habitats. Results showed that across all adult female butterflies, oviposition preferences were highest for A. incarnata and lowest for A. fascicularis, but mean preferences did not differ significantly between eastern and western monarch populations. Larvae from both populations experienced the highest survival and growth rates on A. incarnata and A. fascicularis, and we again found no significant interactions between monarch source population and milkweed species. Moreover, the average rank order of larval performance did not correspond directly to mean female oviposition preferences, suggesting that additional factors beyond larval performance influence monarch oviposition behavior. Finally, significant family level variation was observed for both preference and performance responses within populations, suggesting an underlying genetic variation or maternal effects governing these traits.     

The fall migration flyways of monarch butterflies in eastern North America revealed by citizen scientists

The migration of monarch butterflies (Danaus plexippus) from Canada and the United States to overwintering sites in Mexico is one of the world’s most amazing biological phenomena, although recent threats make it imperative that the resources needed by migrating monarchs be conserved. The most important first step in preserving migration resources—determining the migration flyways—is also the most challenging because of the large-scale nature of the migration. Prior attempts to determine the flyways using mark-recapture techniques with wing tags gave some clues, but this important information has never been fully obtainable until now. In 2005 the citizen-science program, Journey North, initiated a project that asked participants to record sightings of overnight roosts of monarchs during their fall migration, and this project now provides an ideal way to illustrate the flyways used by monarchs on their way to Mexico, with the assumption that roost locations indicate migration routes. We used 3 years of this data to elucidate the flyways on a continent-wide scale, that revealed two distinct flyways, but only one appears to lead directly to Mexico. This main, ‘central’ flyway begins in the American Midwest states and southern Ontario, then continues south-southwest through the states of Kansas, Missouri, Oklahoma and Arkansas, and finally passes through Texas and northern Mexico. These data also highlighted a separate, smaller flyway along the eastern and coastal states, but there was a noticeable lack of roost sightings in this flyway at lower latitudes. Since there are few recoveries of marked monarchs in Mexico originating from coastal areas, we compared the timing of roost formation in this ‘eastern’ flyway with the main, central flyway. Roosts in the eastern flyway lagged behind the central roosts in timing, suggesting that monarchs traveling in this flyway have a reduced chance of making it to the Mexico wintering site. Combined, our evidence indicates that locations in the central flyway should be considered priority areas for conserving migration resources.     

Phenology of weight, moisture and energy reserves of Australian monarch butterflies, Danaus plexippus

Abstract. 1. In the Sydney area of New South Wales, dry weights of reproductive monarch butterflies averaged 156 mg and were higher during winter than in other seasons. Dry weights of non-reproductive monarchs ranged from 216 to 324 mg and declined by 15–25% during over wintering.
2. Fat in reproductive butterflies ranged from 0.009 to 0.017g/0.1g dry weight and was lower during winter than in other seasons. Fat content of non-reproductive monarchs was higher (0.019–0.037 g/0.1g) and declined by 24–51% during over wintering.
3. Lean dry weights of reproductive monarchs were lower than those of non-reproductive individuals. Lean dry weights of non-reproductive butterflies increased rapidly at the beginning of over wintering and remained high throughout the winter. Analysis of protein content indicated the higher lean weight of non-reproductive monarchs was due to greater protein levels.
4. Moisture content of monarchs did not vary with season or reproductive status and appeared to be correlated with ambient humidity.
5. Non-reproductive monarch butterflies in New South Wales adjust bio chemically during over wintering. Energy reserve dynamics of these butterflies are comparable to those that occur in non-reproductive monarchs in North America.     

An experimental investigation of patch use by female monarch butterflies,Danaus plexippus (L.)

Monarch butterflies, which breed throughout the year in southeastern Queensland, Australia, were studied in four dense milkweed patches during the winter months (June-August) 1983. The percentage of marked females recaptured was measured in each of four 15-day sampling periods. In patches where males were experimentally removed, female recapture rate decreased compared with patches which had the same density reduction but no change in sex ratio. There was a significant correlation between female recapture rate and the proportion of males in samples, but female recapture rate was not correlated with population density, the number of males, the number of females or the proportion of young butterflies in the samples. We propose that females assessed patch quality by the sex ratios, and left a patch sooner if encounter rate with males was low. Since males provide a nutrient ‘reward’ during copulation, they may be a limited resource for females during winter. An alternative interpretation, that females left a patch sooner when female density was higher, was not supported as strongly by the data.