Recent Advances in Insect Embryology in Japan and Poland
Edited by H. Ando and Cz. Jura
Arthropod. EmbryoL. Soc. Jpn.
(ISEBU Co. Ltd., Tsukuba) 1987

69

Oogenesis of Nicoletia phytophila (Zygentoma, Nicoletiidae).
Preliminary Studies

Teresa SZKLARZEWICZ
Synopsis
The ovaries of Nicoletia phytophila are of panoistic type. Each ovary is composed of threeovarioles. The ovariole has terminal filament, germarium and vitellarium. The vitellogenesis isaccompanied by the development of microvilli on the oocyte surface and by changes in theultrastructure of follicular cells.

Introduction
Zygentoma and Microcoryphia, which used to be combined in one order Thysanura, are Apterygota most closely related to Pterygota. As histological and ultrastructural studies have shown, both Zygentoma and Microcoryphia have panoistic ovaries (Bitsch, 1980a, b; Bitsch and Bitsch, 1982). Vitellogenesis, of heterosynthetic ormixed type with heterosynthetic kind dominating, is typical of these insects (Bitsch,
1980b; Bitsch and Bitsch, 1982). Nicoletia phytophila is a representative of Zygentoma of
the family Nicoletiidae. Oogenesis has not yet been studied in this family and seems to
be worth a closer examination, because of the unusual life style of this species (seeMaterial and Methods). Besides of it, the analysis of the oogenesis of N. phytophila
might lead to interesting phylogenetic conclusions. This is all more likely, since studiesof oogenesis are a good basis for consideration of the relationships of groups of insects
and other invertebrates (Eckelbarger, 1983).

Material and Methods
Nicoletiaphytophila is a species of Cuban origin, where it lives in heaps of termites of
the Prorhinotormes simplex. It is a parthenogenetic species, and breeds easily in thelaboratory.

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T. Szklarzewicz

2 4 ~_
Fig. 1. Phase-contrast micrograph of an ovariole. X 180.
Fig. 2. Photomicrograph of an ovariole. Methylene blue. X 240.
Fig. 3. Photomicrograph of a germarium. Methylene blue. Xl, 000.
Fig. 4. Electron micrograph ofa germarium. X 15,000.
ge, germarium; m, mitochondria; oc, young oocyte; og, oogonia; on, oogonial nucleus; pfc,
prefollicular cells; pfn, prefollicular cell nucleus; rer, rough endoplasmic reticulum; tf, terminal filament; vt, vitellarium.

Oogenesis of Nicoletia phytophila

71

The insects were placed in a container filled with wood, taken from their naturalenvironment. After ovaries dissected out, they were placed in Hoyle's medium for insects, then photographed in the Peraval Interphaco Carl Zeiss Jena light microscope innegative phase contrast. Ovaries destined for ultrastructural studies were fixed in buffered 3 % glutaraldehyde and 2 % osmium tetroxide and embedded in epoxy resin Epon
812.
Semithin sections were stained with 1 % methylene blue. Ultrathin sections were
contrasted with uranyl acetate and lead citrate and examined with a Tesla BS 500 electron microscope.

Results
The ovaries of N. phytophila are composed of three ovarioles of the panoistic type.
In each ovariole, the terminal filament, germarium and vitellarium may be distinguished (Figs. 1, 2). The germarium is fairly small, and oogonia, prefollicular cells andyoung oocytes are found in it (Fig. 3).
The oogonial cells are isodiametric (Fig. 4), have small, regularly spherical nuclei
and a small amount of cytoplasm. Part of the chromatin is in condensed form. Numerous elements of the rough endoplasmic reticulum (RER) can be found in the cytoplasm, as well as small, spherical mitochondria, lamellar bodies and free ribosomes.
The prefollicular cells are cone-shaped. Their nuclei are ovoid, the chromatin forms
irregular aggregations inside the nucleus and along the nuclear envelope. The cytoplasm of prefollicular cells contains mainly elements of the RER and free ribosomes.
Small spherical mitochondria have also been observed. Young oocytes are larger thanoogonia, their size increases distally in the ovariole. The nucleus of young oocytes isspherical, with a diameter of 1.6 f-L m. The chromatin is diffuse, with several irregularly spaced aggregations of electron-dense material. A large amount of mitochondri~ isfound in the cytoplasm of young oocytes (Fig. 5). Oocytes are accompanied by occasional prefollicular cells. The cell membranes of young oocytes and prefollicular cells
adhere closely to one another (Fig. 8).
In the vitellarium, all the oocytes are arranged linearly. The oocytes located apically are in the stage of previtellogenesis. At this stage the oocyte cytoplasm is richer inmitochondria than before and has RER. The nuclei are spherical, and the chromatin
despiralized. Single follicular cells may be found both on the surface of oocytes, andalso between them. These cells are elongated, possess a large ovoid nucleus which
occupies the greater part of the cell. The cytoplasm which surrounds the nucleus is filled mainly by mitochondria and free ribosomes. In the stage of previtellogenesis, theoolemma begins to form a few microvilli (Fig. 9).
During vitellogenesis, the ultrastructure of the oocyte and follicular cells differsfrom that described for the stages previously studied. At the beginning of vitellogenesis
the follicular epithelium surrounds the whole oocyte (Fig. 6). The follicular cells areusually cubical and adhere closely to one another. The onset of vitellogenesis is indicated by the appearance of pinocytotic vesicles in their cytoplasm (Fig. 10). Numerous
mitochondria, ribosomes and cysterns of RER may also be found in the ooplasm. The
oolemma forms microvilli, which branch out. These microvilli lie very close to each

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T. Szklarzewicz

Fig. 5. Photomicrograph of the apical part of an ovariole. Methylene blue. X 800.
Fig. 6. Photomicrograph of early vitellogenic oocyte. Methylene blue. X 560.
Fig. 7. Photomicrograph of vitellogenic oocyte. Methylene blue. X 540.
Fig. 8. Electron micrograph of interface between young oocyte and prefollicular cell. X
26,000.
fe, follicular epithelium; oc, oocyte; on, oocyte nucleus; pfc, prefollicular cells; pfn, prefollicular cell nucleus.

Oogenesis of Nicoletia phytophila

73

Fig. 9. Electron micrograph of interface between oocyte and follicular cell. X 32,000.
Fig. 10. Electron micrograph of interface between early vitellogenic oocyte and follicularcell. X 16,000.
oc, oocyte; fc, follicular cell; fn, follicular cell nucleus; m, mitochondria; mv, microvilli; pv,
pinocytotic vesicles.

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T. Szklarzewicz

other and are often intertwined with each other. In this stage the ultrastructure of theoocyte nucleus does not change. The follicular cells have more cytoplasm, and also agreater number of cell organelIes. Mitotic figures have frequently been observed in follicular cells in the beginning of vitellogenesis (not shown).
In advanced stages of vitellogenesis, large amounts of reserve substances, such as
lipid and protein droplets appear; these eventually fill almost all the cytoplasm. The
structure of the oocyte nucleus remains unaltered in this phase of oogenesis. The nuclei
offollicular cells are branched and lobated (Fig. 7).

Discussion
The studies carried out on the oogenesis of N. phytophilahave shown that females ofthis species have typically developed panoistic ovaries. An ovary of similar structure ischaracteristic of all the Zygentoma and Microcoryphia (Bitsch, 1980a, b; Bitsch andBitsch, 1982) and also some primitive pterygote insects belonging to the following
orders: Odonata, Ephemeroptera, Plecoptera, Phasmida, Orthoptera, Grylloblattodea,
Thysanoptera (reviewed by King and Buning, 1985) and in megalopterans belonging
to the family Corydalidae (Matsuzaki et al., 1985). The ovary in Nicoletia is composedof three ovarioles. As in other Zygentoma and Microcoryphia the number of ovarioles
in the ovary is small, for example 7 in Lepismachilis (Bitsch, 1980b), 5 in Thermobia(Bitsch and Bitsch, 1982). Germ line cells are present in the ovary: these are the oogonia and oocytes. Prefollicular and follicular cells are somatic cells. The process of vitellogenesis is accompanied by changes in the organization of the follicular epithelium. In
the stage of previtellogenesis, the follicular cells are few and elongated in shape. Their
nuclei are fairly large, the cytoplasm contains mainly mitochondria and elements of the
RER. At the moment of transition from previtellogenesis to vitellogenesis, the numberof folliclilar cells grows considerably. As a result, a single layer of continuous, thinepithelium appears on the oocyte surface. Cell shapes change simultaneously, and they
become cubical. In the stage of early vitellogenesis, endomitotic division of the follicular cell nuclei has been observed. A change in the shape of the nucleus is the result of
these endomitoses, it becomes branched and lobated at this stage. Similar changes in
the follicular cell nuclei have been observed in a representative of the Microcoryphia,
Lepismachilis targionii (Bitsch, 1980a).
As the pictures obtained indicate, the chromatin despiralizes in the beginning of
the period of previtellogenesis. This state is maintained until the end of vitellogenesis.
The above observations show that intensive RNA synthesis takes place in the previtellogenic and vitellogenic oocytes. The studies, which have been made to date on RNA
synthesis during the oogenesis of insects, have shown that during the duration of thisprocess rRNA and long living mRNA are accumulated in the ooplasm (reviewed by
Te1fer, 1975). Thus it would seem that in Nicoletia, as in other insects, it is these kindsof RNA which accumulate. Further studies on the structure of the oocyte nucleus, using the electron microscope and histochemical methods will enab1~ us to gain a better
understanding of this phenomenon.
The growth of the oocyte in Nicoletia is accompanied by the complication of its cell
membrane. The oolemma forms microvilli, which are relatively few in previtellogenesis,

Oogenesis of Nicoletia phytophila

75

but during the previtellogenesis-vitellogenesis transition their number increases considerably. This brings about an increase in the surface of the cell membrane of theoocyte. The formation of microvilli in the beginning of vitellogenesis is a typical phenomenon and has been frequently decribed for various insect species (Bilinski, 1976,
1979; Bilinski et al., 1985; Bitsch, 1980b; Bitsch and Bitsch, 1982; Ksil}zkiewicz-Ilijewa,
1977; Matsuzaki, 1971; Matsuzaki et al., 1979, 1985). In early vitellogenesis, pinocytotic vesicles appear in the cortical cytoplasm of the oocyte, and grow in numbers as theprocess of yolk accumulation progresses. Thus the heterosynthetic type of vitellogenesis
occurs in Nicoletia, as is the case for other Zygentoma and Microcoryphia (Bitsch,
1980b; Bitsch and Bitsch, 1982). This would indicate the close relationship of thisgroup with pterygote insects. Moreover, the relation of Zygentoma and Microcoryphia
to Pterygota seems to be closer than with the remaining apterygote insects, in which,as has already been shown, vitellogenesis is of an autosynthetic or mixed type (Bilinski,
1976, 1977, 1979).
It is reasonable to suppose that further studies of the oogenesis of Nicoletia phytophila ought to yield interesting data on the relationship of the Zygentoma and Microcoryphia with the Pterygota on the one hand, and the Apterygota on the other.

Acknowledgements
I want to express my gratitude to Dr. F. Weyda, Czechoslovak Academy of Science, Ceske Budejovice. for providing the colony of Nicoletia and to Dr. S. Bilinski,
Jagiellonian University, Krak6w, for his suggestions and critical reading of this manuscript.

References
Bilinski, S., 1976. Ultrastructural studies on the vitellogenesis of Tetrodontophora bielanensis (Waga)(Collembola). Cell Tiss. Res. 168: 399-410.
- - - - - , 1977. Oogenesis in Acerentomon gallicum Jonescu (Protura). Previtellogenic andvitellogenic stages. Cell Tiss. Res. 179: 401-412.
- - - - - . , 1979. Oogenesis in Campodea sp. (Diplura). The ultrastructure of the egg chamber
during vitellogenesis. Cell Tiss. Res. 202: 133-143.
- - - - - , W. J. Hage and J. G. Bluemink, 1985. Gap junctions between the follicle cells
and the oocyte during oogenesis in an insect, Tribolium destructor (Coleoptera). Roux's Arch.
Devl. Bioi. 194: 296-300.
Bitsch, J., 1980a. Ultrastructure de l'epithelium folliculaire et des enveloppes de l'ovocyte chez
Lepesmachilis targionii (Grassi) (Thysanura: Machilidae). Int. J. Insect Morphol. Embryol. 9: 25-

40.

- - - - - , 1980b. Ultrastructure des ovocytes et vitellogenese chez Lepismachilis targionii(Grassi) (Thysanura, Machilidae). Int.}. Insect Morphol. Embryol. 9: 297-313.
- - - - - , and C. Bitsch, 1982. Les etapes de la vitellogenese chez Thermobia domestica (Packard) (Thysanura: Lepismatidae). Int. J. Insect Morphol. Embryol. 11: 197-212.
Eckelbarger, K. J., 1983. Evolutionary radiation in polychaete ovaries and vitellogenic mechanisms: their possible role in life history patterns. Can.}. Zool. 61: 487-504.
King, R. C. and J. Biining, 1985. The origin and functioning of insect oocytes and nurse cells. In
G. A. Kerkut and L. I. Gilbert (eds ..), Comprehensive Insect Physiology, Biochemistry andPharmacology, Vol. 1, 37-82. Pergamon Press, Oxford, New York.

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KsilJzkiewicz-Ilijewa, M., 1977. Ultrastructural study of the oocytes of Gromphadorhina brauneri(Shelf. ) (Blattodea) with special reference to yolk formation. Acta Biol. Grac., Ser. Zool. 20:
237-244.
Matsuzaki, M., 1971. Electron microscopic studies on the oogenesis of dragonfly and cricket withspecial reference to the panoistic ovaries. Devl. Growth Differ. 13: 379-398.
- - - - - , H. Ando and S. N. Visscher, 1979. Fine structure of oocyte and follicular cells
during oogenesis in Galloisiana nipponensis (Caudell and King) (Grylloblattodea: Grylloblattidae). Int.}. Insect Morphol. Embryol. 8: 257-263.
- - - - - - , A. Sasaki and M. Komuro, 1985. Panoistic ovarioles of the dobsonfly Protohermes
grandis(Megaloptera, Corydalidae). In H. Ando and K. Miya (eds. ), Recent Advances in Insect Embryology in Japan, 13-23. Arthropod. Embryol. Soc. Jpn. (ISEBU Co. Ltd., Tsukuba).
Telfer, W. H., 1975. Development and physiology of the oocyte-nurse cell syncytium. Adv. Insect
Physiol. 11: 223-319.

Author's address: MSc. T. Szklarzewicz
Department of Systematic Zoology,
Institute of Zoology, Jagiellonian
University, ul. Karasia 6, 30-060
Krak6w, Poland