Kinetic organization of metaphase I bivalents in spermatogenesis of Lepidoptera and Trichoptera species with small chromosome numbers

Received 26 June 1996 Heredity 79 (1997) 135—143 Kinetic organization of metaphase I bivalents in spermatogenesis of Lepidoptera and Trichoptera species with small chromosome numbers KLAUS WERNER WOLF-I-, KAREL NOVAK & FRANTIEK MAREC* tlnstitut für Anatomie der Medizinischen Universität, Ratzeburger A/lee 160, D-23538 Lübeck, Germany and /nstitute of Entomology, Czech Academy of Sciences, Braniovská 31, CZ-37005 Ceské Budejovice, Czech Republic The fine structure of bivalents in metaphase I spermatocytes of two Lepidoptera species, Orgyia thyellina Butler (n = 11) and 0. antiqua (L.) (n = 14) (Lymantriidae), and a Trichoptera species, Limnephilus decipiens (Kolenat) (n = 10) (Limnephilidae) were studied using a series of ultrathin sections and transmission electron microscopy. The bulk of species in both orders possess a haploici chromosome number of about 30. Thus, the experimental species have relatively small chromosome numbers. This study showed that metaphase I bivalents in both Lepidoptera species are polykinetic; attachment of kinetochore microtubules is found scattered throughout the entire poleward chromosomal surface. The microtubules were inserted in material of medium electron density. A pair of distinct kinetochore plates, consisting of material of about the same electron density as the chromatin, was detected at each poleward chromosomal surface in metaphase I bivalents of the caddis-fly, L. decipiens. The observations suggest that DNA elements responsible for the organization of the kinetochores are dispersed throughout the chromosomes in the two Lepidoptera species, whereas they are narrowly clustered in the Trichoptera. Thus, karyotype evolution in the closely related Lepidoptera and the Trichoptera involved widely differing mechanisms. Keywords: karyotype evolution, kinetochores, meiosis, microtubules. from a common ancestor (Kobayashi & Ando, 1988; Kristensen, 1991; Neboiss, 1991; Nielsen & Common, 1991). Several synapomorphies also apply ochores were found at each poleward surface of the chromosomes. The kinetochores consisted of plates of medium electron density that were in contact with microtubules (MTs). Ephestia kuehniella (Pyralidae) and B. mon (Bombycidae) have a haploid chromosome number of 30 and 28, respectively (Traut & Mosbacher, 1968; Traut, 1976; Holm & Rasmussen, 1980). The caddis-fly, A. furcata (Limnephilidae), meiosis is of the achiasmatic type (Suomalainen, 1966; Traut & Mosbacher, 1968). Fine structure the haploid chromosome number is around 30 (fig. 5 in Wolf et al., 1992). Similarity between representa- Introduction The insect orders Lepidoptera and Trichoptera are considered sister groups on the basis of about 20 synapomorphies, common characteristics derived has not been precisely karyotyped, but published micrographs of a chromosome spread suggest that to properties of the karyotype. In both orders, females are the heterogametic sex and female tives of both insect groups with about 30 chromosomes is not restricted to chromosome structure in meiosis. In metaphase spermatogonia, kinetochore plates cover around 40 per cent of the chromosomal length (Gassner & Klemetson, 1974; Wolf et al., studies involving two Lepidoptera species, the Medi- terranean flour moth Ephestia kuehniella (Wolf & Traut, 1991) and the silkworm Bombyx mon (Holm & Rasmussen, 1980), and a Trichoptera species, Anabolia furcata (Wolf et a!., 1992) revealed that the architecture of bivalents in metaphase I spermatocytes is very similar. In all cases, two distinct kinet- 1992). Information on the kinetic organization of chromosomes in Lepidoptera and Trichoptera species with exceptionally small chromosome Correspondence. E-mail: 1997 The Genetical Society of Great Britain. 135 136 K.WWOLFETAL. numbers is scarce. In any case, electron microscopy is required because Lepidoptera chromosomes are unusually small (Goodpasture, 1975). In a previous fine structure study, the presence of polykinetic metaphase I bivalents has been mentioned in Orgyia thyellina Butler (Lymantriidae), a moth with a haploid set of 11 chromosomes (Wolf et al., 1987). In other words, MT contacts were found scattered atmosphere for dissection to Lübeck. In this species, which undergoes imaginal diapause in native conditions (Novak & Sehnal, 1963), the larvae approaching pupation appeared to be optimal for preparation of meiotic chromosomes in metaphase I (cf. Le Lannic, 1975). throughout the entire poleward surface of the chromosomes. The emphasis of the study was, however, on the comparison of eupyrene and Preparation for e/ectron microscopy apyrene meiosis and chromosome structure was not gonads were transferred to Ringer solution (Glaser, 1917: cited by Lockwood, 1961) containing 2.5 per cent glutaraldehyde. The composition of the Ringer documented in detail. In brief, Lepidoptera show double spermatogenesis. Eupyrene meiosis gives rise to spermatids which transform into fertilizing sperm, whereas apyrene meiosis leads to a non-fertilizing sperm variety (Meves, 1903; Wolf, 1994). In the present article, which is only concerned with eupyrene meiosis, we dwell on the kinetic organization of metaphase bivalents in primary spermatocytes of 0. thyellina. In addition, we present information about bivalent structure in eupyrene spermatogenesis of another species of the same genus, 0. antiqua (L.), which possesses a haploid Set of 14 chromosomes (Cretschmar, 1928). Also in this species, spindle fine structure has been previously described (Wolf, 1990), but information on the kinetic organization of the bivalents is missing. Because only eupyrene spermatocytes are involved, we dispense with this attri- bute in the text when Lepidoptera are addressed. The findings in the Lepidoptera species are compared with those in a Trichoptera species, preparation procedure has been described previously (Wolf, 1994). In brief, the dissected The solution was: 9 g NaC1, 0.42 g KC1, 0.25 g CaCI2 and 0.2 g NaHCO3 per litre doubly distilled water. After 5 mm, three volumes of 8 per cent tannic acid (Merck) in phosphate buffer (0.067 M, pH 6.8) were added. The gonads were postfixed in phosphatebuffered 0s04 (1 per cent), dehydrated in ethanol and embedded in Epon 812. Ultrathin sections, about 70 nm thick, were mounted on Pioloformcoated grids, doubly stained with uranyl acetate and lead citrate and analysed with a Philips EM 400 transmission electron microscope operated at 80 kV. In 0. thyellina, one longitudinally sectioned and one cross-sectioned metaphase I plate have been recorded using series of consecutive sections. In 0. antiqua and L. decipiens, it has been one longitudinally sectioned metaphase I spindle in each. In all cases, primary spermatocytes were studied, and the findings were confirmed by inspection of about six Limnephilus decipiens (Kolenat) (Limnephilidae), with a haploid set of 10 chromosomes (Klingstedt, 1931; for a compilation of chromosome num (...truncated)