Cell death and wing reduction during the metamorphosis of sex-specific flightless morphs in winter geometrid moths.

Winter geometrid moths show striking sexual dimorphism by having female-specific flightless morphs. The evolutionary grades of wing reduction in winter geometrid moths vary and range from having short wings, vestigial wings, to being wingless. Although the ontogenetic processes underlying the development of the wingless or short-wing morphs in Lepidoptera has been well studied, the mechanisms underlying the development of vestigial wing morphs in winter geometrid moths during metamorphosis are poorly understood. In the winter geometrid moth Sebastosema bubonaria Warren, 1896, the males have functional wings, but the females have vestigial wings. Here, we studied the ontogenetic processes affecting wing reduction in the winter geometrid moth S. bubonaria using light microscopy and transmission electron microscopy, and compared the ontogenetic process of wing reduction in this species with that in another species of the wingless-female winter moth that we investigated previously. Our results showed that, in the vestigial-wing morphs, the loss of pupal wing epithelium was terminated in the middle of the wing degeneration process, whereas in the wingless morph, the pupal wing epithelium disappeared almost completely and the final appearance of the wings differed slightly among flightless morphs. We propose that the extent and location of cell death in the pupal wing play an important role in the various patterns of reduced wings that are observed in flightless moths.

Early cellular development induced by ecdysteroid in sex-specific wing degeneration of the wingless female winter moth.

The winter moth, Nyssiodes lefuarius, exhibits striking sexual dimorphism in wing form; males have functional wings of normal size, whereas females lack wings. We previously found that the steroid hormone 20-hydroxyecdysone (20E) triggered massive programmed cell death (PCD) only in the female pupal wing epithelium; however, when and how early sexual trait development of the pupal wings is initiated during pupal-adult metamorphosis remains obscure. To clarify the detailed morphological changes and mechanisms underlying early sexual trait development and cell death, we examined the effects of 20E on early ultrastructural and histological changes in the pupal wing epithelium of both sexes. Before the onset of adult differentiation, no morphological differences were observed in the epithelial cells of both sexes at an ultrastructural level. When 5.4 µg of 20E was injected into pupae of both sexes at 15 days after the onset of pupation, retraction of the wing epithelium from the pupal cuticle was initiated at day 2 after 20E injection in both sexes. Although overt degeneration of wing tissue was not still obvious, apoptotic body-like structures and auto-phagosomes were visible at day 3 after 20E injection in females, whereas development of scale precursor cells started on day 4 after injection in males. Our results suggest that (1) the injection of 20E induced sexually dimorphic changes in the pattern of organelle distribution in wing epithelial cells, and (2) abnormally shaped mitochondria in the cytoplasm of the female wing epithelium might be involved in the PCD that occurs during wing tissue degeneration.

Discovery of wing imaginal discs in the penultimate instar of the lacewing Mallada desjardinsi (Insecta: Neuroptera: Chrysopidae) with histological notes on postembryonic imaginal disc development.

Holometabolous insects are alternatively named "Endopterygota" because, in the larvae of many taxa, the wing primordia in the lateral regions of the meso- and metathoracic segments form more or less invaginated structures called wing imaginal discs. Holometabolous insects exhibit differential developmental timing of the wing during ontogeny. The condition in which wing growth is deferred until the end of larval life has been considered ancestral, whereas early disc formation has been recognized as the derived condition. Even though wing disc development in holometabolous insects has been studied extensively in select groups, many questions remain about the development of the wing imaginal disc in the orders Raphidioptera, Megaloptera, Neuroptera, and Mecoptera. To clarify whether the wing imaginal disc of Neuroptera is typical of the derived condition, we examined the ontogeny of the wing imaginal discs in the lacewing Mallada desjardinsi histologically. Using both light microscopy and transmission electron microscopy, we were able to recognize wing imaginal discs in the penultimate larval instar (prefinal larval instar) of this species. To date, neuropteran insects have been characterized as having late-forming wing imaginal discs. However, our findings show that the developmental pattern of the wing imaginal discs within the Neuroptera represents a more derived pattern of development in the Holometabola than was assumed previously.

Morphological and histological examination of short-wing formation in the winter moth Protalcis concinnata (Insecta: Lepidoptera, Geometridae).

Winter geometrid moths exhibit sexual dimorphism in wing length and female-specific flightlessness. Female-specific flightlessness in insects is an interesting phenomenon in terms of sexual dimorphism and reproductive biology. In the winter geometrid moth, Protalcis concinnata (Wileman), adult females have short wings and adult males have fully developed wings. Although the developmental process for wing reduction in Lepidoptera is well studied, little is known about the morphology and the developmental pattern of short-winged flightless morphs in Lepidoptera. To clarify the precise mechanisms and developmental processes that produce short-winged morphs, we performed morphological and histological investigations of adult and pupal wing development in the winter geometrid moth P. concinnata. Our findings showed that (a) wing development in both sexes is similar until larval-pupal metamorphosis, (b) the shape of the sexually dimorphic wings is determined by the position of the bordering lacuna (BL), (c) the BL is positioned farther inward in females than in males, and (d) after the short pupal diapause period, the female pupal wing epithelium degenerates to approximately two-thirds its original size due to cell death. We propose that this developmental pattern is a previously unrecognized process among flightless Lepidoptera.

Distribution of Dall's Porpoise, Phocoenoides dalli, in the North Pacific and Bering Sea, Based on T/S Oshoro Maru 2012 Summer Cruise Data.

Dall's porpoise (Phocoenoides dalli) is a small toothed cetacean, widely inhabiting the North Pacific Ocean and adjacent seas, between about 30 and 62°N; however, only limited studies of its ecology have been made in nearshore areas. A cetacean sighting survey lasting 60 days was conducted during the 2012 summer cruise of the T/S Oshoro Maru (Hokkaido University, Japan) in the North Pacific Ocean and Bering Sea. Based on this data, the distribution of Dall's porpoises and the factors controlling it in the pelagic habitat were investigated. A total of 808 individual Dall's porpoises in 166 groups were sighted during a total of 469.6 hr and 4946.6 nm observations. The cruise consisted of three legs and the average porpoise group size was significantly larger in Leg 1. The sightings were concentrated at water depths of less than 1000 m and near eastern Aleutian passes. Sighting clusters were found on the 200 m isobath of the southeastern Bering Sea continental slope. There was a peak in sightings where the sea surface temperature (SST) was relatively cold, between 5 and 7°C. Although similar track routes were taken in Leg 1 and Leg 3, the number of sightings per unit effort was larger in Leg 1. This difference may have arisen from the significant rise in SST as the season progressed. Relatively large group size found in this study might relate with prey abundance along the Aleutian Islands.

Ecdysteroid-induced programmed cell death is essential for sex-specific wing degeneration of the wingless-female winter moth.

The winter moth, Nyssiodes lefuarius, has a unique life history in that adults appear during early spring after a long pupal diapause from summer to winter. The moth exhibits striking sexual dimorphism in wing form; males have functional wings of normal size, whereas females lack wings. We previously found that cell death of the pupal epithelium of females appears to display condensed chromatin within phagocytes. To provide additional detailed data for interpreting the role of cell death, we performed light microscopy, transmission electron microscopy, and TUNEL assay. We consequently detected two modes of cell death, i.e., dying cells showed both DNA fragmentation derived from epithelial nuclei and autophagic vacuole formation. To elucidate the switching mechanism of sex-specific wing degeneration in females of N. lefuarius, we tested the effects of the steroid hormone 20-hydroxyecdysone (20E) on pupal diapause termination and wing morphogenesis in both sexes. When 20E (5.4 µg) was injected into both sexes within 2 days of pupation, wing degeneration started 4 days after 20E injection in females, whereas wing morphogenesis and scale formation started 6 days after 20E injection in males. We discuss two important findings: (1) degeneration of the pupal wing epithelium of females was not only due to apoptosis and phagocytotic activation but also to autophagy and epithelial cell shrinkage; and (2) 20E terminated the summer diapause of pupae, and triggered selective programmed cell death only of the female-pupal wing epithelium in the wingless female winter moth.

In vitro effects of juvenile hormone analog on wing disc morphogenesis under ecdysteroid treatment in the female-wingless bagworm moth Eumeta variegata (Insecta: Lepidoptera, Psychidae).

Female adults of the bagworm moth, Eumeta variegata, lack wings completely, whereas male adults of this species have functional wings. We previously found that ecdysteroid induces apoptotic events in the female wing rudiment of E. variegata in vitro, whereas the male wing discs cultured with 20-hydroxyecdysone (20E) underwent apolysis and then cell differentiation. To investigate whether juvenile hormone (JH) in involved in sex-specific cellular response to ecdysteroid during wing development between sexes of E. variegata, we tested the effects of juvenile hormone analog (JHA), methoprene, and 20E on wing disc morphogenesis between sexes in vitro. Using transmission electron microscopy (TEM), we found that both higher concentration of JHA (5 µg/ml) and 20E (1 µg/ml) addition induced cell death (apoptosis) in the male wing discs but not induced cell death in the female wing rudiments in vitro in E. variegata. These culture experiments clearly detected the differential responses of wing discs to JHA under ecdysteroid treatment between sexes. We propose two important hypotheses: (1) JH is not significantly involved in the suppression of the female wing rudiment morphogenesis under 20E treatment, (2) female wing rudiment has lost the ability for cell proliferation in response to the stimulus of 20E.

Feminization of genetic males by a symbiotic bacterium in a butterfly, Eurema hecabe (Lepidoptera: Pieridae).

Wolbachia are symbiotic bacteria found in many arthropods and filarian nematodes. They often manipulate the reproduction of host arthropods. In the present study, female-biased sex-ratio distortion in the butterfly Eurema hecabe was investigated. Breeding experiments showed that this distorted sex ratio is maternally inherited. When treated with tetracycline, adult females of the thelygenic line produced male progeny only. After PCR using Wolbachia-specific primers for the ftsZ gene a positive result was seen in the thelygenic females, but not in male progeny from tetracycline-treated females, or individuals from a Tokyo population with normal sex ratio and reproduction. Cytological observations showed that thelygenic females lack the sex chromatin body (W chromosome). The results strongly suggest that the sex-ratio distortion in E. hecabe is due to feminization of genetic males by Wolbachia.