Ovaries of Tubificinae (Clitellata, Naididae) resemble ovary cords found in Hirudinea (Clitellata)

The ultrastructure of the ovaries and oogenesis was studied in three species of three genera of Tubificinae. The paired ovaries are small, conically shaped structures, connected to the intersegmental septum between segments X and XI by their narrow end. The ovaries are composed of syncytial cysts of germ cells interconnected by stable cytoplasmic bridges (ring canals) and surrounded by follicular cells. The architecture of the germ-line cysts is exactly the same as in all clitellate annelids studied to date, i.e. each cell in a cyst has only one ring canal connecting it to the central, anuclear cytoplasmic mass, the cytophore. The ovaries found in all of the species studied seem to be meroistic, i.e. the ultimate fate of germ cells within a cyst is different, and the majority of cells withdraw from meiosis and become nurse cells; the rest continue meiosis, gather macromolecules, cell organelles and storage material, and become oocytes. The ovaries are polarized; their narrow end contains mitotically dividing oogonia and germ cells entering the meiosis prophase; whereas within the middle and basal parts, nurse cells, a prominent cytophore and growing oocytes occur. During late previtellogenesis/early vitellogenesis, the oocytes detach from the cytophore and float in the coelom; they are usually enveloped by the peritoneal epithelium and associated with blood vessels. Generally, the organization of ovaries in all of the Tubificinae species studied resembles the polarized ovary cords found within the ovisacs of some Euhirudinea. The organization of ovaries and the course of oogenesis between the genera studied and other clitellate annelids are compared. Finally, it is suggested that germ-line cysts formation and the meroistic mode of oogenesis may be a primary character for all Clitellata

Ovary cord micromorphology in the blood-sucking haemadipsid leech Haemadipsa japonica (Hirudinida: Arhynchobdellida: Hirudiniformes)

This is the first report that describes histological and ultrastructural details of ovary organization in haemadipsid leeches. In Haemadipsa japonica, the female reproductive system is organized similar to that of other haemadipsids. Each of the paired and oval ovaries of H. japonica is comprised of the ovary wall (ovisac), which encloses two elongated, thread-like ovarian units termed ovary cords. Ovary cords are comprised of germ-line cells and associated somatic cells. Each cord is polarized and contains germ-line cells in the consecutive developmental stages that are sequentially located along the long cord axis. There were three zones in each cord: the club-shaped apical part, the thread-like middle part, and the basal-most end, which contains degenerating germ cells. Outside of the reproductive period, the middle part of the cord in leeches is smooth, and no growing oocytes are visible; alternatively, in mature specimens, several growing oocytes protrude from the cord, and several huge vitellogenic oocytes that are completely detached from the cord occur within the ovisac. Ovary cord organization and functioning in H. japonica are very similar to the ‘Hirudo’ type cords that were found in several hirudiniform leeches. This conclusion supports the view that all hirudiniform leeches have conservative ovary cord organization and a similar pattern of oogenesis. Germ-line cyst composition, architecture, and functioning were also found to be evolutionarily conservative characteristics when compared with all previously examined Clitellata. In the germ-line cysts found in H. japonica each cell is connected to the central and anuclear cytoplasmic mass (cytophore) via one intercellular bridge, and, as oogenesis progresses, the fate of interconnected cell diversifies: some of them (oocytes) grow and complete oogenesis, but the majority become nurse cells and finally degenerate. Thus, oogenesis in H. japonica, similar to other clitellates, can be considered meroistic

Schematic illustration of the spatial organization of the <i>T</i>. <i>tubifex</i> ovary.

<p>The ovary is conically shaped with its narrow end attached to the intersegmental septum (is) via a thin ligament (li) and is polarized along its long axis. Germ cells at subsequent stages of oogenesis occur in three zones (I, II, III): zone I contains oogonia (oo), undifferentiated germ cells (cystocytes) (gc) occur in zone II and germ cells that are morphologically differentiated into growing oocytes (o) and smaller nurse cells (nc) occur in zone III. Oocytes grow on only one side of the ovary and gradually protrude into the body cavity. The entire ovary is composed of only one, huge germ-line cyst. Each germ cell in a cyst (ovary) is connected to the central anuclear core (cytophore) via one stable intercellular bridge (ring canal – ellipse). The cytophore (cy) is poorly developed in zones I and II, whereas it is prominent and branched in zone III. In addition to the germ cells, somatic cells (sc) occur – they are localized inside the ovary and on its surface and form a thin ovary envelope.</p

The summary of germ-line cyst organization in different groups of animals in conjunction with the cytoskeleton.

<p>No available data (-).</p><p>The summary of germ-line cyst organization in different groups of animals in conjunction with the cytoskeleton.</p

The Ovary of <i>Tubifex tubifex - Table 2 </i> (Clitellata, Naididae, Tubificinae) Is Composed of One, Huge Germ-Line Cyst that Is Enriched with Cytoskeletal Components

<p>The Ovary of <i>Tubifex tubifex - Table 2 </i> (Clitellata, Naididae, Tubificinae) Is Composed of One, Huge Germ-Line Cyst that Is Enriched with Cytoskeletal Components</p

General organization of an ovarian cyst in <i>T</i>. <i>tubifex</i>.

<p>(A) The ovary of <i>Tubifex tubifex</i> is conically shaped and polarized. Three zones can be distinguished (I, II and III). Each zone contains germ cells at subsequent stages of oogenesis: oogonia in zone I, undifferentiated cells in zone II and nurse cells and oocytes in zone III. In the third zone, several oocytes (o) grow successively on only one side of the ovary and protrude into the body cavity. An extensive cytophore (cy) is marked in this zone. Whole-mounted preparation, Nomarski interference contrast. (B) and (C) sections through zone III, in which two morphologically distinct germ cell categories occur – nurse cells (nc) and oocytes (o). The branched cytophore (cy) occupies the central position in the germ-line cyst. Arrowheads mark the cytophore regions that are enriched with cytoskeletal elements (see Figs <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0126173#pone.0126173.g002" target="_blank">2A</a> and <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0126173#pone.0126173.g003" target="_blank">3</a>). Arrows point to the somatic cells that accompany the germ cells inside the ovary and form the outer ovary envelope; nu – oocyte nucleus. B longitudinal section, fluorescence microscopy, Histocryl semi-thin section double stained with DAPI and PI; C cross section, light microscopy, Epon semi-thin section stained with methylene blue.</p

F-actin localization in a <i>T</i>. <i>tubifex</i> ovarian cyst.

<p>The entire ovary double stained with DAPI and rhodamine-conjugated phalloidin. Maximum-intensity projections of the Z stacks that cover the interior of the ovary. (A and B) whole ovaries, (C) detail of area between zones II and III. Long bundles of actin filaments occur (stars) in the branched cytophore (cy). F-actin is also localized in the cortical cytoplasm (arrows) of nurse cells (nc) and oocytes (o). Confocal microscopy.</p

Localization of microtubules in a <i>T</i>. <i>tubifex</i> ovarian cyst.

<p>Zone III. Live cells stained with Hoechst 33342 (A) and Tubulin Tracker Green (B); (C-H) merged. (A-D) Microtubules form a network that fills the entire cytoplasm of the cytophore (cy); note microtubules in the ring canals that connect the germ cells (gc) with the cytophore (arrowheads) and the bundles of microtubules in the cytophore branches (stars in D). (E-F) A prominent network of microtubules (arrows) occurs in the cytoplasm of the germ cells (gc) and in the vitellogenic oocytes (o); nu – nucleus. (G) Mitotic spindles (arrowheads) of dividing germ cells were observed in zone III; stars indicate metaphase plates. (H) Long bundles of microtubules (arrowheads) were also detected in the cytoplasmic projections of the somatic cells (arrows point to the nuclei of these cells); gc – germ cells, o – oocytes. Fluorescence microscopy.</p

Organization of F-actin in an ovarian cyst of <i>T</i>. <i>tubifex</i>.

<p>(A and B) The initial cytophore with thin F-actin bundles occurs at the narrow end of the ovary (zones I and II) (stars). The cytophore with its F-actin bundles reaches every germ cell (gc). The ring canals that connect the germ cells to the cytophore have rims that are enriched with F-actin (arrows). The F-actin also occurs in the cortical cytoplasm of the germ cells (arrowheads). Fluorescence microscopy, whole-mounted preparation stained with rhodamine conjugated phalloidin. C-E Fragment of zone II double stained with DAPI (C) and rhodamine-phalloidin (D); merged view (E). The cytophore has thin bundles of F-actin (stars) that reach each germ cell (gc). The ring canals rims are enriched with F-actin (arrows) and cortical F-actin can be observed in the germ cells (arrowheads). Fluorescence microscopy.</p

Ovary micromorphology in hormogastrid earthworms with a particular emphasis on the organization of the germline cysts

There is a gap in our knowledge of microorganization and the functioning of ovaries in earthworms (Crassiclitellata) and allied taxa. Recent analyses of ovaries in microdriles and leech-like taxa revealed that they are composed of syncytial germline cysts accompanied by somatic cells. Although the pattern of cyst organization is conserved across Clitellata – each cell is connected via one intercellular bridge (ring canal) to the central and anuclear cytoplasmic mass termed the cytophore – this system shows high evolutionary plasticity. In Crassiclitellata, only the gross morphology of ovaries and their segmental localization is well known, whereas ultrastructural data are limited to lumbricids like Dendrobaena veneta. Here we present the first report about ovarian histology and ultrastructure in Hormogastridae, a small family of earthworms inhabiting the western parts of the Mediterranean sea basin. We analyzed three species from three different genera and showed that the pattern of ovary organization is the same within this taxon. Ovaries are cone-like, with a broad part connected to the septum and a narrow distal end forming an egg string. Ovaries are composed of numerous cysts uniting a small number of cells, eight in Carpetania matritensis. There is a gradient of cysts development along the long ovary axis, and three zones can be distinguished. In zone I, cysts develop in complete synchrony and unite oogonia and early meiotic cells (till diplotene). Then (zone II), the synchrony is lost, and one cell (prospective oocyte) grows faster than the rest (prospective nurse cells). In zone III, oocytes pass the growth phase and gather nutrients; at this time, their contact with the cytophore is lost. Nurse cells grow slightly, eventually die via apoptosis, and are removed by coelomocytes. The most characteristic feature of hormogastrid germ cysts is the inconspicuous cytophore in the form of thread-like thin cytoplasmic strands (reticular cytophore). We found that the ovary organization in studied hormogastrids is very similar to that described for D. veneta and propose the term "Dendrobaena" type of ovaries. We expect the same microorganization of ovaries will be found in other hormogastrids and lumbricids.National Science Centre, Poland (NCN)Depto. de Biodiversidad, Ecología y EvoluciónFac. de Ciencias BiológicasTRUEpu