The peculiar extra-acrosomal structure of the Collembola (Hexapoda) spermatozoa.

The springtail Collembola are characterized by having rolled spermatozoa, with a long cylindrical extracellular structure adhering to the acrosome. This structure is produced by the secretory activity of the testes epithelial cells at almost the end of spermiogenesis. At the beginning of its formation, it is a thin extension with a helical wall and a dense axial region. Later the cylindrical structure shows an inner organization which is different in the several species examined: species of Entomobryidae contain material with a paracrystalline structure, whilst some of Symphypleona contain ovoid structures. The outer envelope of the extracellular structure consists of two overlapped layers orthogonally arranged, clearly identified by cryo-preparations. Immunoblot analysis and lectin stainings have indicated that the cylindrical structure has a glycoproteic composition. As the structure is no longer visible after the sperm transfer into the female spermatheca, it is suggested that it could contain enzymes able to activate the sperm unwinding process and possibly allowing the reacquisition of sperm motility.

Deep phylogeographic divisions and long-term persistence of forest invertebrates (Hexapoda: Collembola) in the North-Western Mediterranean basin.

The North-Western Mediterranean basin is well known for its high number of relictual endemic taxa, and has been indicated as one of the world's major biodiversity hotspots at the species level. A possible contributing factor may be long-term persistence of populations and their prolonged stability. This study was designed to investigate the phylogeographic structure of three common species of the genus Lepidocyrtus (Hexapoda: Collembola), soil-dwelling arthropods characterized by limited dispersal capabilities and generally associated with forest habitats. We observed a remarkable geographic structure, with numerous deeply divergent genetic lineages occupying islands as well as mainland sites with no apparent gene flow among most sites, even across distances of only tens of kilometres. The reconstructed time frame for the evolution of these lineages suggests divergence between 5 and 15 Ma. This indicates a remarkably ancient origin and long-term persistence of individual lineages over a fine geographic scale despite the occurrence of abrupt sea level and climatic fluctuations in the area. This further suggests that currently recognized morphological species might be a serious underestimation of the true springtail biodiversity within this region.

Different sperm number in the spermatophores of Orchesella villosa (Geoffroy) (Entomobryidae) and Allacma fusca (L.) (Sminthuridae).

Members of the basal hexapod group Collembola perform indirect sperm transfer; males deposit spermatophores on the soil and these are then picked up by females for insemination. Orchesella villosa exhibits a dissociated mating behavior, while Allacma fusca has either associated mating behavior or, more commonly, produces spermatophores without pairing. Sperm ultrastructure in the two species has been studied by TEM and SEM observations to estimate the volume of these cells. The sperm number into each spermatophore has been determined by counting the fluorescent nuclei after Hoechst staining of sperm droplets squash preparations. Sperm droplets at the apex of spermatophores contain numerous spermatozoa of typical appearance, wound up and with a long anterior appendage (peduncle). Sperm droplets of Orchesella villosa are smaller (about 10 times) than in A. fusca, however they contain substantially more sperm cells (about three times). These differences could be explained as follows: (a) sperm of O. villosa are shorter than those of A. fusca (58 and 107microm, respectively) and thus the volume of a single sperm of O. villosa is about 10 times smaller than in A. fusca; (b) considering the volume of sperm droplets and of sperm cells, the estimated number of sperm cells would be higher in A. fusca than in O. villosa; and (c) the conspicuous reduction of the sperm cells observed in A. fusca is likely linked to the aberrant meiosis present in this species, as well as in all Symphypleona studied so far, that causes a loss of 50% of their sperm.

Fine structure of the secretory and sensory organs on the cephalon and the first pereionite of Trichoniscus alexandrae Caruso (Crustacea, Isopoda).

Trichoniscus alexandrae Caruso is a blind troglobiont isopod; males possess secretory and sensory organs on the cephalon and 1st pereionite consisting of cuticular pits hosting a tuft of setae and gland openings. Such organs are absent in females. Three types of cuticular structures have been observed: (a) lamellar setae, which likely play a role in protecting the gland openings and favouring the evaporation of secretions; (b) contact chemoreceptors, each provided with six bipolar sensory cells, a scolopale cell and enveloping cells; (c) a secretory cell complex, consisting of a long cylindrical slender duct-forming cell, with the function of transporting to the cuticular surface a secretion produced by two deeper secretory cells. The duct-forming cell is characterized by the presence of numerous microtubules in its cytoplasm, and is provided with a flattened duct. It is suggested that the secretion produced by the secretory cells could serve for sex-recognition.

Sperm structure and spermiogenesis in Coletinia sp. (Nicoletiidae, Zygentoma, Insecta) with a comparative analysis of sperm structure in Zygentoma.

The spermatozoon of Coletinia sp. has a bilayered acrosome, a short nucleus (4 microm) and a relatively short sperm tail with two mitochondrial derivatives. The chromatin is uniformly dense except for several electron-lucid channels or strands which permeate the nucleus and which originate in the spermatid as invaginations of the nuclear envelope. The invaginations occur mostly or exclusively along two meridians of the spermatid that are also characterized by the presence of a longitudinal rod of medium electron density. The two rods (designated as 'mid-spermatid rods') evidently are instrumental in the formation of the electron-lucid channels. The significance of this elaborate system of intranuclear channels is not understood. The sperm tail has a 9 + 9 + 2 axoneme with each of the nine microtubular doublets accompanied by an accessory microtubule; scant intertubular material can also be distinguished. Hence, the tail axoneme resembles that of many pterygote insects. Each of the two mitochondrial derivatives contains a crystalline inclusion that has periodically spaced layers going in different directions on either side of the midline. Two synapomorphic traits appear to be shared by Ateluridae and Nicoletiidae, namely the invaginations of the nuclear membrane along two meridians of the nucleus and the shape of the crystalline inclusions of the mitochondrial derivatives. Four species from the family Lepismatidae were also examined as to their sperm ultrastructure. Three of them, Allacrotelsa kraepelini, Ctenolepisma longicaudata and Ctenolepisma sp., were found to be very similar to the two previously examined lepismatids, Thermobia domestica and Lepisma saccharina. On the other hand, spermatozoa of Tricholepisma aurea were aggregated in small groups rather than pairwise joined as seen in the other lepismatids. Sperm characters are also used to reconstruct a phylogenetic hypothesis which suggests a close relationship between Ateluridae and Nicoletiidae.

The complete mitochondrial DNA sequence of the basal hexapod Tetrodontophora bielanensis: evidence for heteroplasmy and tRNA translocations.

We present the complete 15,455-nt mitochondrial DNA sequence of the springtail Tetrodontophora bielanensis (Arthropoda, Hexapoda, Collembola). The gene content is typical of most metazoans, with 13 protein-coding genes (PCGs), 2 genes encoding for ribosomal RNA subunits, and 22 tRNA genes. The nucleotide sequence shows the well-known A+T bias typical of insect mtDNA; its A+T content is lower (72.7%) than that observed in other insect species, but still higher than that in other arthropodan taxa. The bias appears to be uniform across the whole molecule, unlike other insect taxa, which show increased A+T content in the so-called A+T-rich region. However, the bias is slightly higher in the third codon positions of the PCGs (81.4%). Anomalous initiation codons have been observed in the nad2 and the cox1 genes. In the latter, the ATTTAA hexanucleotide is suggested to be involved in the initiation signaling. All tRNAs could be folded into the typical cloverleaf secondary structure, but the tRNA for cysteine appears to be missing the DHU arm. Long tandemly repeated regions (193 nt) were found in the A+T-rich region, which in turn was shown to have the possibility of forming a complex array of secondary structures. One of these structures encompassed the junction between the repeats. The A+T-rich region was also interesting in that it showed heteroplasmy in the number of repeats. Three haplotypes were found, possessing 2, 3, and 4 identical repeats, respectively. The order of protein coding and rRNA genes in the molecule was determined and was identical to that of all insects studied so far. However, two tRNA translocations were found which were unprecedented among Arthropoda. These involved the trnQ, which was found between the rrnS and the A+T-rich region, and the trnS(ucn), which was located between trnM and trnI. A preliminary phylogenetic analysis based on the amino acid sequence of the PCGs failed to find support for the monophyly of Hexapoda.

Aberrant spermatogenesis and the peculiar mechanism of sex determination in Symphypleonan Collembola (Insecta).

Light and electron microscopy evidence have been obtained to describe the peculiar spermatogenesis in the collembolan species Sminthurus viridis and Allacma fusca (Sminthuridae). In these two species, the two sexes differ for the lack of two chromosomes (the sex chromosomes) in males (males, 2n = 10; females, 2n = 12). While oogenesis seems to proceed normally, spermatogenesis is peculiar because the two daughter cells of the first meiotic division have different chromosome numbers (six and four). The cell receiving four chromosomes degenerates, while the cell receiving six chromosomes completes meiosis and produces identical spermatozoa (n = 6). At fertilization, pronuclei with six chromosomes fuse together to form zygotes with 2n = 12. Male embryos must lose two sex chromosomes during the first zygotic mitosis, as all male cells have 2n = 10 chromosomes. The sex chromosome system of these species can be identified as X1X1X2X2:X1X20. Electron microscopy observations show that the same peculiar spermatogenesis occurs also in two others species of the same family, Caprainea marginata and Lipothrix lubbocki. The peculiar sex determination system described is similar but not identical to what is observed in other insect orders, and it may represent an evolutionary step toward parthenogenesis. It is suggested that this peculiar spermatogenesis is common to all Symphypleona.

Chromosome elimination and sex determination in springtails (Insecta, Collembola).

A post-zygotic mechanism of sex determination is described in the two symphypleonans Dicyrtomina ornata (Nicolet) and Ptenothrix italica Dallai. The process consists of the loss of two sex chromosomes from the male embryo. At the end of the first meiotic division of spermatogenesis, a second chromosome elimination occurs, allowing half the secondary spermatocytes, later transformed into spermatids, to receive a complete haploid set of chromosomes. The secondary spermatocytes, which receive an incomplete set of chromosomes, degenerate. Males of the two collembolan species, therefore, produce a reduced number (50%) of spermatozoa. Females of D. ornata have 2n = 12 and males 2n = 10 chromosomes; females of P. italica have 2n = 14 and males 2n = 12 chromosomes. In both species, oogenesis proceeds normally and chromosomes pair and form chiasmata in meiotic prophase. The adaptive significance of this post-zygotic mechanism of sex determination is discussed. The mechanism seems to be a characteristic feature of the suborder Symphypleona. The neanurid Arthropleona Anurida maritima (Guérin), which was studied for comparative analysis, has 2n = 8 chromosomes and normal spermatogenesis producing haploid nuclei with four chromosomes. J. Exp. Zool. (Mol. Dev. Evol.) 285:215-225, 1999.

Ultrastructure of the male accessory glands ofAllacma fusca(Insecta, Collembola).

The accessory glands ofAllacma fusca(L.) (Insecta, Collembola, Sminthuridae) consist of a series of secretory units that are arranged in parallel and open into the ejaculatory duct. Each unit is composed of microvillate cells stacked around a common cavity. Basal cells are involved in ion-control of fluids from the hemocoel to the cavity. The intermediate and apical cells, which have a laminar appearance and contain many microtubules, are involved in the structural integrity of the unit. Supporting cells ensheath the most apical cells. Large openings in the cuticle allow the gland secretion to flow into the ejaculatory duct lumen. These openings are protected by a porous cuticle different from that lining the epithelium of the ejaculatory duct. Conspicuous muscle fibers run along the lateroventral side of the ejaculatory duct beneath the insertion of the accessory glands. The fine structure of the accessory glands indicates that they are type I ectodermic glands as defined by Noirot & Quennedey (1974). Their function could be to control the fluidity of the material for spermatophore formation and to ensure the proper physiological conditions for spermatozoa stored in the ejaculatory duct lumen.