Morphology of the larval and adult brains of the monarch butterfly, Danaus plexippus plexippus, L

Cell population and neuropile morphology of larval and adult brains of the monarch butterfly, Danaus plexippus plexippus, L., are compared. The larval brain is in continuous transition, the processes of adult brain development being underway from the earliest larval stages. It is characterized by a less diverse population of cells and more homogenous fiber areas than those of the adult. Neuroblasts, which divide to form the neurones of the adult brain, occur either in discrete proliferation centers or scattered among the larval ganglion cells. The larval brain contains, in addition to small homogeneous antennal centers and a distinct larval optic center, rapidly developing adult optic centers, corpora pedunculata, and protocerebral bridge. The larval brain lacks a central body. Major differences between larval and adult brains are clearly related to the increased dependence of the adult upon sensory input from the eyes and antennae.     

Sialic acids inTrichoplusia ni andDanaus plexippus egg glycoproteins

Baculovirus expression vector insect cell system (BEVIS) is useful for high level production of human therapeutic proteins. However, it has been reported that recombinant glycoproteins produced from this system are, in many cases, biologically inactive or less active than authentic counterparts, due to incomplete glycosylation potential of insect cells used so far, producing recombinant proteins with only high-or paucimannosidic oligosaccharides without sialylation. The presence of sialic acids in insects is still controversial. Egg proteins ofTrichoplusia ni andDanaus plexippus were isolated, and the presence of sialic acids was examined using reverse-phase fluorescent HPLC after derivatization of samples with 1,2-diamino-4,5-methylenedioxybenzene (DMB). The proteins of both eggs were shown to contain 5-N-acetylneuraminic acid. The results suggest that both insects may be able to produce proteins with sialylated complex-type oligosaccharide chains.     

Pyrrolizidine alkaloids from Senecio vulgaris sequestered and stored by Danaus plexippus

Danaus plexippus reared on Asclepias curassavica , sprayed with homogenized leaves of Senecio vulgaris , were found to contain mean pyrrolizidine alkaloid concentrations of 38.8±6.08 μg/male butterfly and 16.5 ± 3–63 μg/female butterfly.     

Migration of the monarch butterfly, Danaus plexippus: energy sources,

The origin and nature of the metabolic reserves of the monarch butterfly during its autumn migration through Missouri were examined. Information was obtained about the butterfly's protein, glycogen, lipid, and lipid-soluble pigment composition. The principal results showed that lipids, which made up ca. 45 per cent of the insect's dry weight, were the major reserve. Analyses of the lipid classes showed that triglycerides were the dominant class in each sex. Separate analyses of the thorax and abdomen showed that >90 per cent of the lipids were present in the massive fat body which is restricted to the abdomen. The adult female rapidly incorporated U-¹⁴C-d-glucose into abdominal glycerides. The rôle of larval reserves and nectar consumption in migration and ovarial maturation was discussed.     

Simulation of movement and egglaying inDanaus plexippus (Lepidoptera: Nymphalidae)

A model simulating the movement and oviposition of monarch butterflies over a female's life time is presented. The model's predictions compare favourably with observed data and suggest that females who lay eggs in an optimal fashion should have low directionalities in areas with high host plant density (patches and single plants) and high directionalities in areas with low host plant density. The model also provides one means of combining individual animal processes and spatial heterogeneity into population dynamics.     

Overwintering ecology of the monarch butterfly, Danaus plexippus L., in California

Abstract. 1. Population and physiological measurements were made on monarch butterflies in three overwintering colonies along the California coast during the 1975–76 season. 2. Mark, release and recapture studies indicated that two northern colonies (Muir Beach and Santa Cruz) stabilized at maxima of about 40000 and 95 000 individuals with little movement in or out of colonies, whereas the southern one at Santa Barbara appeared dynamic with a maximum of about 45 000. 3. Initially low, sexual activity builds to a massive mating ceremony in February, followed by rapid colony dispersal. The butterflies deteriorate gradually during the overwintering period, but males do so to a greater extent because of their involvement in frenzied courting flights. 4. Changes in several parameters at the Muir Beach colony were measured between 1 November and 2 February. Right forewing length did not change with time, indicating no differential mortality with respect to size. However, both sexes lost about 25% of their total dry weight, and 49–60% of their lipid contents. Minimum fat levels were about 20 mg and some butterflies were as lean on arrival as were the most depleted overwintering survivors. 5. The importance of nectaring in restoring lipid reserves is potentially great but remains uninvestigated. The hypothesis is put forward that the distance spring migrants are capable of flying is dependent largely upon their lipid reserves following the overwintering period. This and the intracolony mass-mating ceremony are seen as randomizing factors which prevent geographic differentiation and allow the maintenance of the generalized, nature of the migration, which in turn enables the monarch to exploit the vast milkweed flora across the North American continent. 6. Cardenolides also diminished during the wintering, but to a substantially lesser extent than the lipids. There was no evidence for a change in the proportion of cardenolide-positive butterflies, suggesting no differential elimination by predation of the poison-free palatable individuals. This is the first field evidence in support of the theory of automimicry. 7. The overwintering behaviour of the monarch butterfly is seen as an integrative force in optimizing foodplant exploitation, reproduction, anti-predator, and migration strategies.     

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.     

Effect of photoperiod and temperature on reproduction of the monarch butterfly,Danaus plexippus

Optimal posteclosion ovarian development in monarch butterflies occurs at about 28°C. Higher and lower temperatures appear progressively inhibitory. Reproductive gland development in males follows a similar pattern. Juvenile hormone injections stimulate oöcyte maturation in animals held at 20 and 35°C but are most effective at the higher temperature. Juvenile hormone injections are ineffective at 10 and 15°C. The stimulatory effect of increasing photophase on ovarian development was most apparent at optimal temperature. The possible significance of our findings to monarch migration is discussed.     

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.     

Response of Danaus plexippus to pollen of two new Bt corn events via laboratory bioassay

Laboratory bioassays were conducted to evaluate the response of first instar larvae of the monarch butterfly, Danaus plexippus L. (Lepidoptera: Danaidae), a non‐target species, to pollen from corn, Zea mays L. (Commelinales: Poaceae), from two new corn hybrids genetically modified to express different types of insecticidal proteins derived from the bacterium Bacillus thuringiensis Berliner (Bacillales: Bacillaceae) (Bt). One hybrid expresses both Cry1Ab and Cry2Ab2 proteins (MON 810 × MON 84006), active against lepidopteran pests, and the other expresses Cry3Bb1 protein (MON 863), targeted against coleopteran pests. First instar larvae were placed on milkweed leaves (Asclepias syriaca L.) (Gentianales: Asclepiadaceae) dusted with doses of either Bt pollen or its nonexpressing (isoline) pollen counterpart ranging from 50 to 3200 grains cm^?2 of milkweed leaves, or no pollen at all. Larvae were exposed to pollen for 4 days, then moved to pollen‐free leaves and observed for another 6 days. Survival was observed after 2, 4, and 10 days. Weight gain was estimated after 4 and 10 days, leaf consumption after 2 and 4 days, and larval development after 10 days. Exposure to pollen of the Cry1Ab/Cry2Ab2‐Bt expressing hybrid reduced larval survival approximately 7.5–23.5% at the dose ranges tested relative to a no pollen control. Larval weight gain and consumption were reduced for larvae exposed to pollen of this hybrid and a small minority of larvae (3.1%) never developed past the third instar after 10 days of observation. Exposure to pollen of the Cry3Bb1‐Bt expressing hybrid had no negative effects on larval mortality, weight gain, consumption, or development relative to the consumption of Bt‐free corn pollen. The relevance of these findings to the risk that these Bt corn hybrids pose to monarch populations is discussed.     

Influence of acclimation temperature on the shivering behavior of the butterfly Danaus plexippus (L.)

Summary Most of the monarch butterflies kept at 4–5° C for a few days shivered when released at a test temperature of 15–16° C, whereas fewer of the butterflies kept at 23–24° C did so. Cold-acclimated butterflies shivered more readily, as indicated by the length of the interval between release at the test temperature and the onset of shivering, and they shivered for longer periods of time. The effects of cold acclimation were reversible, but in only 1 out of 3 replicates was the warm acclimation clearly reversed. Cool animals shivered at room temperature, indicating that body temperature and not ambient temperature is important in releasing the behavior. It is suggested that the acclimation involves alteration in the central neurons controlling the activity of muscles involved in shivering.I thank Miss Janice Ruppert and Mr. C. J. Doughty for their valuable technical assistance. The co-operation of the administrators of New Brighton Beach State Park in permitting me to collect in the park is appreciated.     

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.     

Contact chemoreception related to host selection and oviposition behaviour in the monarch butterfly, Danaus plexippus

Abstract. Behavioural events during host selection by ovipositing monarch butterflies (Danaus plexippus (L.), Danainae, Nymphalidae) include tapping the leaf surface with fore-tarsi and touching this surface with mid-tarsi (‘drumming’) and antennae. Flavonoids identified from host plant extracts are known to stimulate oviposition. Scanning electron microscopy revealed the presence of contact-chemoreceptor sensilla on all appendages that contact the leaf surface. This electrophysiological study was conducted to identify the contact chemoreceptors that are sensitive to the known oviposition stimuli and are therefore probably involved in host recognition. Receptor cells of conspicuous sensilla grouped in clusters on fore-tarsi of females were sensitive to the behaviourally active butanol fraction of host plant (Asclepias curassavica) extract. However, these receptors generally had low sensitivity to three oviposition-stimulating flavonoids identified from this fraction, but they were also sensitive to the butanol fraction of a non-host (Brassica oleracea). Chemoreceptors in sensilla of the tarsomers 2–4 of the mid-legs also responded to the behaviourally active fraction of host plant extract and showed some sensitivity to two of the flavonoids that stimulate oviposition. Similar results were obtained from receptor cells in sensilla on the tip of the antennae. Most of these sensilla had cells responding to the butanol fraction of A. curassavica extract but only 25% of them were also sensitive to one of the behaviourally active flavonoids. These electrophysiological results, in combination with behavioural observations, suggest that host selection in monarch butterflies relies on a complex pattern of peripheral sensory information from several types of tarsal and antennal contact chemoreceptors.     

Dynamics of oviposition in Danaus plexippus (Insecta: Lepidoptera) on milkweed, Asclepias spp

Where a butterfly chooses to lay an egg will influence the subsequent survival of her offspring. In this paper we report on the effect of a number of variables which influence the choice of oviposition site in Danaus plexippus L. Experiments were conducted under both flight cage and field conditions. The field observations consisted of recording the within and between plant egg dispersions across different patch sizes. Laboratory experiments looked at egg laying preferences as affected by plant species, age and condition. Butterflies were selective in their choice of oviposition site. Eggs were laid singly on the underside of medium sized leaves towards the top of a plant. More eggs were laid per plant on single isolated plants than on plants within a patch. The number of eggs per plant increases with plant height but decreases with plant age. Females preferred young plants or plants with fresh regrowth of leaves. These characteristics could override species preferences which were, in decreasing order, Asclepias curassavica. A. fruticosa and A. physocarpa. Species preferences varied between butterflies and with female age. Neither the presence of eggs nor larvae on a plant deterred oviposition. These results are compared with previous observations of egg laying in this species.     

Sex ratios and Ophryocystis elektroscirrha infection levels of Danaus plexippus during spring migration through Oklahoma,USA

Monarch butterflies, Danaus plexippus L. (Lepidoptera: Nymphalidae), have a multiple brood migration in the spring as they move between their overwintering grounds and summer breeding grounds. In Oklahoma, USA, monarchs produce at least one generation in the spring, which develops and continues the northward migration, leaving Oklahoma without a breeding population during the hot summer months. Female monarchs leave the overwintering grounds prior to males, but it is not clear whether females re‐colonize areas along the migration route prior to, or at the same time as males. Male‐to‐female ratios are 1:1 at emergence, but studies have identified a male‐biased sex ratio in the field. Both males and females are susceptible to infection by the obligate protozoan parasite, Ophryocystis elektroscirrha McLaughlin & Myers (OE), which reduces flight abilities and life spans of infected individuals. We examine sex ratios during the spring migration through Oklahoma and whether sex ratios or OE infection estimates vary with capture technique (active or passive). Our data suggest populations are male‐biased during the 1st week of spring migration in Oklahoma, but shift to female‐biased by the 3rd week in both cool and warm springs. Therefore, males may leave southern areas prior to females or migrate longer distances per day. Active sampling (i.e., netting) did not bias sex compared to passive sampling (i.e., sticky traps). Significantly fewer OE‐carrying monarchs (with two or more spores) were captured via netting than by sticky traps which may be caused by sticky trap glue affecting tape sampling effectiveness, but there was no difference in the number of heavily infected individuals (more than 100 spores). Therefore, data from netted monarchs may underestimate OE infection rates within populations.     

The structure and function of the bursa copulatrix of the monarch butterfly (Danaus plexippus)

The bursa compulatrix of the Monarch butterfly was investigated utilizing light microscopy, histochemistry, and scanning and transmission electron microscopy in order to relate its morphology to the release of sperm from the spermatophore. The bursa has a row of large chitinous teeth on either side of the organ. The dorsal and ventral surfaces are covered with chitinous plates, the plates having bristles on one side. A single layer of cells lies under both the plates and teeth, one columnar cell under each plate, one cuboidal cell under each tooth. The toothed area has no muscle cells. However, the dorsal and ventral hemispheres of the bursa each have a crescent-shaped packet of muscle fibers that traverse the organ; there are no longitudinal fibers. Spermatophores with thick walls were found in the bursal lumen. Morphological evidence suggests that the presence of the spermatophores is sensed by the bristles and that the packets are opened by contraction of the muscles bringing the large teeth into contact with the spermatophore wall.     

Molecular basis for the insensitivity of the Monarch (Danaus plexippus) to cardiac glycosides.

The Monarch (Danaus plexippus) sequesters cardiac glycosides for its chemical defence against predators. Larvae and adults of this butterfly are insensitive towards dietary cardiac glycosides, whereas other Lepidoptera, such as Manduca sexta and Creatonotos transiens are sensitive and intoxicated by ouabain. Ouabain inhibits the Na+,K(+)-ATPase by binding to its alpha-subunit. We have amplified and cloned the DNA sequence encoding the respective ouabain binding site. Instead of the amino acid asparagine at position 122 in ouabain-sensitive insects, the Monarch has a histidine in the putative ouabain binding site, which consists of about 12 amino acids. This change may explain the ouabain insensitivity.     

Consistent differences among individual monarch butterflies (Danaus plexippus L.) in flight activity at low temperatures

Previous work suggested differences in allozyme frequencies between samples of monarch butterflies collected at different times of the day. This study examines the prediction that some individuals are active consistently earlier in the day than others. Four specific hypotheses were tested: (1) Individuals caught early on one day were more likely than those caught late to be caught early on subsequent days. This was true in some experiments. (2) Individuals caught early in the field were more likely than those caught late to fly early in outdoor flight cages. This was true when outdoor temperatures were low (between 10 and 20^oC) but not when they were above 20^oC. (3) Individuals caught early in the field were more likely to be able to fly at low body temperatures than those caught late. This was not true. (4) Individuals that flew early in outdoor flight cages were more likely to be able to fly at low body temperatures. This was not true. Overall we conclude, on the basis of field and flight cage experiments, that some individuals are active consistently earlier in the day than others. The lack of a relationship between field and temperature cabinet results indicates that this is not purely a result of differences in ability to fly at low body temperatures. The implications of these results are discussed.     

Thoracic temperature,shivering, and flight in the monarch butterfly,Danaus plexippus (L.)

Summary Monarch butterflies, Danaus plexippus (L.), display a warm-up behavior characterized by wingstrokes of small amplitude. Thoracic temperature during this shivering and during fixed flight was measured by means of a smallbead thermistor inserted into the thorax. At ambient temperatures of 15–16°C, once shivering is initiated the thoracic temperature rises at a maximum rate of 1.3°C/min, and a thoracic temperature 4.0°C greater then ambient is produced (Table 1). Fixed flight at these low ambient temperatures results in a similar rate of increase in thoracic temperature, and a similar temperature excess is produced (Fig. 3). At ambient temperatures between 22 and 35°C the thoracic temperature of an animal starting to fly rises at a faster rate, 3.6°C/min, and reaches a greater excess, 7.9°C (Fig. 4). The wingbeat frequency of animals in fixed flight increases with increasing thoracic temperature (Fig. 2). In the absence of direct solar radiation, shivering typically occurs prior to flight at low ambient temperatures (13–17°C), and the resulting increase in thoracic temperature allows monarch butterflies to fly at these cool temperatures.I thank Miss Janice Ruppert and Mr. C. J. Doughty for their valuable technical assistance. The co-operation of the administrators of New Brighton Beach State Park in permitting me to collect in the park is appreciated. Financial support for this study was provided in part by a faculty research grant from the University of California.