Axonal pathfinding in developing labial palps of the butterflies,Pieris rapae and Pieris brassicae

Summary Electron microscopy was used to study the developmental changes in the pupal labial palp of Pieris rapae L. and P. brassicae L. prior to axogenesis of the peripheral receptor organs. Two cells emigrate from the anlage of the labial palp-pit organ (LPPO). They develop growth cones and filopodia, which are directed distally. The filopodia insert into stem cells of the LPPO. The perikarya of the cells proceed into the hemolymph space of the palp such that, eventually, these cells bridge the gap between the LPPO and the apical scolopidial organ (ASO) of the labial palp. This bridge is established at about 11% of total pupal development. Only 6 h later, at 15% of pupal development, the cells are no longer visible. We suggest that these cells are luminal neurons and name them pit-organ luminal (POL) cells. These POL cells may act as primary guiding structures for the axons of the sensory cells of the LPPO, which are assumed to orientate along this structure once they reach the ASO.Supported by the Deutsche Forschungsgemeinschaft (H.A.: SFB4/G1).     

UV reflectance of inter-subspecific hybrid females obtained by crossing cabbage butterflies from Japan (Pieris rapae crucivora) with those from New Zealand (P. rapae rapae)

Cross-mating was performed between subspecies of cabbage butterfly Pieris rapae crucivora from Japan and P. rapae rapae from New Zealand, whose females are known to reflect and absorb ultraviolet (UV) light, respectively. In the female F ₁ hybrids UV reflectance was intermediate between the two parent females. Further, the UV reflectance was significantly higher in the female F ₁ hybrids derived from females of the New Zealand subspecies than in those derived from females of the Japanese subspecies.     

Pattern of spatial distribution of the common cabbage butterfly,Pieris rapae in a cabbage farm

---

Contributions from the Entomological Laboratory, Kyoto University, No. 203.     

Structure,development and death of sensory cells and neurons in the pupal labial palp of the butterflies Pieris rapae L. and Pieris brassicae L. (Insecta,Lepidoptera)

Summary The development of neurons possibly related to the outgrowth of axons from the labial palp-pit organ was studied in Pieris rapae. Serial sections of six successive stages between pupation and emergence of the imago were examined with the electron microscope. At pupation the palp contains an apical scolopidial organ (ASO) and cellular strands connected to it. The ASO consists of three type-1 scolopidia, which are characterized by the presence of a ciliary 9 × 2 + 0 pattern throughout the dendritic outer segment and a ciliary dilation beneath the cap. The scolopidia show two special features: (i) the dendritic outer segments reach beyond the cap, and (ii) an intricate junctional complex develops between the dendritic inner segments and the scolopale cells. The cellular strands comprise two types of cells: (1) bipolar cells regarded as neurons due to their cytological features, and (2) enveloping cells, which are wrapped around the bipolar cells. The strands degenerate about 10 h after pupation. The sensory cells of the ASO degenerate consecutively between 28 h and 130 h after pupation. However, their enveloping cells survive and endure in the imago, which emerges about 160 h after pupation. An ASO similarly lacking sensory cells was observed in imagines of Pieris brassicae. It is hypothesized that the ASO and the bipolar neurons of the strands play a role in pathfinding of the axons of the labial palp-pit organ.Supported by the Deutsche Forschungsgemeinschaft (SFB 4/G1)     

Purification,characterization, and synthesis of vitellin from the cabbage butterfly Pieris rapae L.

Vitellin from the cabbage butterfly Pieris rapae L. was purified and characterized by electrophoresis. Vitellin from P. rapae is a phosphorylated glycolipoprotein of 380,000 ± 10,000 molecular weight as determined by nondenaturing polyacrylamide gel electrophoresis. Two subunits with an M_r of 150,000 and 40,000 were obtained from vitellin. The native molecule is thought to be a tetramer composed of two molecules of each of these subunits. The isoelectric point, as determined by isoelectric focusing on polyacrylamide gels, is 6.10. Vitellin and vitellogenin were indistinguishable by immunological methods such as double diffusion and tandem-crossed immunoelectrophoresis. Vitellogenin from the hemolymph and vitellin from the ovary were quantified by rocket immunoelectrophoresis. Vitellogenin and vitellin were first detected in 6-day-old pupae, and their levels increased continuously during ovarian development. Vitellogenin synthesis by the fat body in 4-day-old female pupae could be induced by juvenile hormone I.     

Mate recognition in two sympatric species of butterflies,Pieris rapae andP. melete

Female butterflies should distinguish conspecific males from males of different species. The aim of the present study was to evaluate the female role in avoiding allospecific mating between 2 sympatric pierids,Pieris rapae crucivora (P. rapae hereafter) andP. melete. Since it is relatively rate to observe responses of a wild female to a courting male of different species, we substituted the wings of a female with those of a female of the other species. Such a female attracted allospecific males. FemaleP. melete took “mate-refusal posture” to courting males, regardless whether the male belonged to different species or not. FemaleP. melete discontinued the posture only when the male was conspecific. Such a behavioural difference indicates that the female discriminated its conspecific male. On the other hand, when only the mate-refusal posture was taken into account, a femaleP. rapae did not differently respond to courlship of males of bothP. melete andP. rapae. Having once approached, the maleP. melete attempted more frequently to copulate with its conspecific females than with allospecific ones, while the maleP. rapae indiscriminately attempted to copulate with both allospecific and conspecific females. The maleP. melete tended to persist in its courtship to females of bothP. rapae andP. melete who took the mate-refusal posture. The mate recognition system by male and female in these 2 species is discussed in connexion with the difference in mating behaviour pattern of the species.     

Associative learning in the ovipositional behaviour of the cabbage butterfly, Pieris rapae

ABSTRACT. When tested with familiar and novel oviposition sites, gravid female Pieris rapae (L.) tended to land on sites of the same appearance as those they had experienced 24 h earlier. This acquired preference could be reversed by subsequent e xperience. An ovipositional preference, shown by groups of insects, paralleled the landing preferences of single insects. These influences of experience were shown with intact cabbage plants of different kinds, with discs cut from the leaves of different kinds of cabbage, and with discs of blotting paper of different shades of green and blue, wetted with sinigrin solution or water. A single landing by a gravid female on paper wetted with sinigrin solution induced a landing preference for the same colour of paper in a choice test made 24 h later. Gravid females thus associated a favourable chemical stimulus of oviposition with the appearance of the acceptable site. Such learning by trial and error would facilitate host-selection from a distance. In the laboratory it induced oviposition on 'neutral' sites of the same appearance as 'host' sites.     

The hormonal control of diuresis in the cabbage white butterfly Pieris brassicae.

The diuresis which follows the pupal-adult ecdysis of Pieris brassicae is hormonally controlled. Use of the isolated Malpighian tubules as a bioassay shows the presence of substantial diuretic activity in homogenates of the brain and corpus cardiacum-corpus allatum complex. The hormone is probably produced in the brain and released from a storage site in the corpora cardiaca. The tubules of the butterfly are maximally responsive to the diuretic hormone at the time of eclosion.     

Sexual dichroism and pigment localization in the wing scales of Pieris rapae butterflies

The beads in the wing scales of pierid butterflies play a crucially important role in wing coloration as shown by spectrophotometry and scanning electron microscopy (SEM). The beads contain pterin pigments, which in Pieris rapae absorb predominantly in the ultraviolet (UV). SEM demonstrates that in the European subspecies Pieris rapae rapae, both males and females have dorsal wing scales with a high concentration of beads. In the Japanese subspecies Pieris rapae crucivora, however, only the males have dorsal wing scales studded with beads, and the dorsal scales of females lack beads. Microspectrophotometry of single scales without beads yields reflectance spectra that increase slightly and monotonically with wavelength. With beads, the reflectance is strongly reduced in the UV and enhanced at the longer wavelengths. By stacking several layers of beaded scales, pierid butterflies achieve strong colour contrasts, which are not realized in the dorsal wings of female P. r. crucivora. Consequently, P. r. crucivora exhibits a strong sexual dichroism that is absent in P. r. rapae.     

Feeding, growth, and the thermal environment of cabbage white caterpillars, Pieris rapae L

Laboratory studies of temperature effects on short-term feeding and growth rates were combined with field data on thermal environments to explore the consequences of temperature variation for growth of caterpillars of the cabbage white butterfly, Pieris rapae. Mean short-term (24-h) consumption and growth rates of fourth-instar P. rapae feeding on collard leaves increased continuously with increasing temperatures between 10 degrees and 35 degrees C, peaked at 35 degrees C, and declined rapidly with temperatures above 35 degrees C. Physical models can mimic temperatures of real fifth-instar caterpillars under collard leaves within 1 degrees -2 degrees C in sunny summer conditions in Seattle, Washington. Continuous recordings of operative temperatures of model caterpillars in a collard garden suggest that, at the timescale of the duration of the fifth instar (5-8 d in the field), P. rapae caterpillars frequently experience temperatures spanning a 25 degrees C range, they spend most of their time at temperatures well below those that maximize growth, and they encounter substantial variation in the frequency distribution of operative temperatures between time periods. Combining these data on growth rate as a function of temperature and the distribution of operative temperatures in the field, I illustrate how growth rates at higher temperatures can make disproportionate contributions to the overall mean growth rates even when higher temperatures are relatively infrequent. Fluctuating thermal conditions may generate variable patterns of selection on reaction norms for growth rate in the field.     

A comparative study of the egg-laying behaviour and larval development of Pieris rapae L. and P. brassicae L. on the same host plants

Summary Pieris rapae and P. brassicae feed on the same host plants and have synchronized seasons. P. brassicae, whose larvae are twice the size of P. rapae, lays eggs in clusters of 40–100 eggs whereas P. rapae lays single eggs. In this paper we examine how egg clustering may be advantageous for P. brassicae. The larval development of each species was studied, and found not to differ significantly. P. brassicae larvae were observed to migrate from their host plant after defoliating it. A comparison of the efficiency of host plant utilization by the two pierid species was undertaken by measuring the effect of larval feeding on the growth of their host plants (kale and brussel sprouts). The results show that egg clustering is advantageous for larval fitness in terms of host resource exploitation, and we suggest that P. brassicae is adapted for ovipositing on clumped vegetation, while P. rapae is selected for exploiting isolated plants.     

Purification and cloning of pierisin-2, an apoptosis-inducing protein from the cabbage butterfly, Pieris brassicae.

The cabbage butterfly Pieris rapae contains a strong apoptosis-inducing substance, pierisin, against human cancer cell lines, which is thought to act via ADP-ribosylation. Here we report the purification and cloning of an apoptosis-inducing substance, designated as pierisin-2, from another cabbage butterfly, Pieris brassicae. Pierisin-2 was purified from pupae by sequential chromatography and its cytotoxic and apoptosis-inducing activities to various cancer cells were similar to those of pierisin, designated as pierisin-1, from P. rapae. cDNA cloning of pierisin-2 was performed on the basis of the partial amino-acid sequence. The nucleotide sequence indicated that the cDNA encodes an 850-amino-acid protein with a calculated molecular mass of 97 986. The deduced amino-acid sequence of pierisin-2 was 91% identical with that of pierisin-1. In vitro expressed protein in the reticulocyte lysate exhibited apoptosis-inducing activities against human gastric carcinoma TMK-1 and cervical carcinoma HeLa cells, similar to the purified native pierisin-2 from the pupae. Pierisin-2 shows regional sequence similarities with certain ADP-ribosylating toxins such as the A-subunit of cholera toxin. The results from site-directed mutagenesis at Glu165, a conserved residue among ADP-ribosylating enzymes necessary for NAD binding, and from experiments with ADP-ribosylating enzyme inhibitors suggested that pierisin-2 could be considered as an ADP-ribosylating toxin like pierisin-1.     

Avoidance of maladaptive,precocious copulation in the cabbage white butterfly,Pieris rapae crucivora

In the cabbage white butterfly, Pieris rapae crucivora, copulation hinders normal expansion and hardening of the wings of newly emerged females. The resulting permanent wing deformation makes it impossible for females to fly and therefore, to find an oviposition site and nectar sources. An attempt was made to clarify whether the newly emerged female butterfly avoids copulation. Observation of wing expansion and hardening reveals that the wings are fully expanded and hard by 20–30 min after emergence. In the field, presentation of females with soft wings to males shows that males will attempt to copulate with these females. However, newly emerged females prevent successful completion of copulation by assuming the mate refusal posture, and thereby avoid a potentially maladaptive copulation. The discussion focusses on the question as to why females and not males avoid early copulation.     

Constituents of host- and non-host plants deterring oviposition by the cabbage butterfly, Pieris rapae

Oviposition by Pieris rapae L. on cabbage was deterred by homogenized cabbage tissue sprayed onto intact plants. Ether extracts of cabbage also were deterrent, but water extracts were not. Hexane extracts of other host plants deterred oviposition and water extracts had little or no effect. Polar as well as non-polar extracts of non-host plants inhibited oviposition. Polar deterrents in the non-host crucifers, Erysimum cheiranthoides and Capsella bursa-pastoris may explain the avoidance of these plants by P. rapae. Chemical deterrents, as well as stimulants, apparently play a major role in the acceptance or rejection of plants as hosts by ovipositing female butterflies.

---

Zusammenfassung Die Eiablage von Pieris rapae auf Kohl wurde gehemmt von einem Homogenisat aus Kohlblättern, das auf intakte Pflanzen gesprüht wurde. Ether-Extrakte der Kohlblätter wirkten im Gegensatz zu Wasserextrakten abschreckend. Ebenso zeigten Hexanextrakte anderer Wirtspflanzen eine hemmende Wirkung auf die Eiablage, während Wasserextrakte anderer Wirtspflanzen keine Wirkung aufwiesen. Dagegen aber verhinderten sowohl polare als auch unpolare Extrakte von Nicht-Wirtspflanzen die Eiablagen. Polare, abschreckende Substanzen in den Nicht-Wirtspflanzen Erysimum cheiranthoides und Capsella bursa-pastoris (Cruciferae) könnten Ursache dafür sein, dass P. rapae diese Pflanzen meidet. Chemische Substanzen mit sowohl abschreckender als auch stimulierender Wirkung spielen offensichtlich eine bedeutende Rolle für legebereite Schmetterlingsweibchen bei der Auswahl von Pflanzen als geeigneten Eiablageplatz.

     

On the distribution of the common cabbage butterfly,Pieris rapae crucivora,in the field

---

Contributions from the Department of Biology, Miyazaki University, No. 45.     

The learning abilities of the white cabbage butterfly,Pieris rapae,foraging for flowers

This study examines the role of learning and memory in the butterflyPieris rapae crucivora Boisduval during foraging for flowers. In an outdoor cage with 6 flower species,P. rapae showed various visiting patterns: some visited only one species, while others visited several species in a day. The foraging process for flowers ofErigeron annuus (L.) Pers. could be divided into two successive steps: (1) landing on the nectaring caputs, and (2) finding the source of nectar in the caput. Butterflies learned to proceed through the two steps more efficiently with successive attempts: they gradually decreased landings on nectarless caputs and probings on the nectarless petals of ligulate flowers respectively. As a result, handling time per unit caputs became shorter, and apparent rewards per unit time, i.e. the efficiency of collecting nectar, increased. In addition, once learned,P. rapae could remember a rewarding flower color for 3 days, which was not interfered with by learning another flower color. This indicates thatP. rapae keeps memory for a period longer than 3 days, and that they can remember at least two flower species as suitable flower resources. Furthermore, data indicated that they sometimes can apply the foraging skills obtained on other flower species to a novel one. These abilities could enable butterflies to easily switch flower species, or to enhance labile preference. It has been known thatP. rapae also shows flower constancy, which may be due to memory constraints. Therefore, they may appropriately use two foraging tactics: visit consistency and labile preference, to get enough nectar according to their circumstances.     

Quality of light effects on immature stages and adults of Pieris rapae (L.) (Lepidoptera: Pieridae)

Larvae of Pieris rapae (L.), the imported cabbage worm, were reared to the adult stage under controlled light conditions. Spectral quality affected their development. The effect of the long wave ultraviolet light was to significantly decrease developmental time. The light source used also appeared to exert an effect on pupal colouration.