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

255

Differentiation and Behaviour of Primordial Germ Cells
during the Early Embryonic Development of

Parnassius glacialis Butler, Luehdorfia japonica Leech
and Byasa (Atrophaneura) alcinous alcinous Klug
(Lepidoptera: Papilionidae)

Masahiro T ANAKA
Synopsis
Differentiation and behaviour of the primordial germ cells of Pamassius glacialis, Luehdorfia
japonica and Byasa (Atrophaneura) alcinous alcinous are described. Tlie primordial germ cells of P.
glacialis are formed through mitotic divisions and ivagination of.the cells at mid-line of the region
about one-third of ventral plate;before.completion of formation of serosa. Those of L. japonica andB. alcinous alcinous appear in a group at the regions about one-third of the germ band before theinner layer formation. When the inner layer has formed, the primordial germ cells are surrounded by the mesodermal cells and come to be undistinguishable from surrounding cells. Afterthe segmentation of ectodermal part of embryo and the metamerization of inner layer, the primordial germ cells divide into two lateral groups in the fourth to sixth abdominal segments, andthen they are carried to the lateral-inner sides of the ectodermal parts with the mesodermal cells.
During this process, the primordial germ cells grow and assume their characteristic appearance.
Their number on each side in the fourth to the sixth abdominal segment varies more or lessamong the species and individuals. One to four primordial germ cells are counted on one side ofeach segment in P. glacialis, one to three in L. japonica and two to five in B. alcinous alcinous.

Introduction
Differentiation and behaviour of the primordial germ cells are the subject of study
attracting interest of insect embryologists. A considerable number of researches on thegerm cells of the lepidopteran insects have been done. The time and the manner of pri-M. Tanaka

256

mordial germ cell formation, however, differ remarkably among the species or the author's opinion, and there remain some uncertain problems concerning the differentiation and behaviour of the primordial germ cells in the Lepidoptera. Different opinions
and uncertain problems are probably caused by the difficulty to distinguish the primordial germ cells from the somatic cells in the early stage of embryonic development.
In the present paper, the author describes the formation and the behaviour of primordial germ cells in Pamassius glacialis, Luehdorfia japonica and Byasa (Atrophaneura) alcinous alcinous.

Materials and Methods
To obtain eggs for the present study, pregnant females of Pamassius glacialis Butler,
Luehdorfia japonica Leech and Byasa (Atrophaneura) alcinous alcinous Klug were captured in
Gifu Prefecture in 1980 to 1984. Eggs of P. glacialis were laid in May, those of L.japonica in April and those of B. alcinous alcinous in June, and kept at room temperature in thelaboratory.
The eggs at various ages were fixed in Carnoy's fluid for 30-40 min or alcoholic
Bouin's fluid for 60-90 min. After fixation, the eggs removed of their chorion were dehydrated, embedded in paraffin, sectioned in 8 fl m, and stained with Delafield'shaematoxylin and eosin.

Observations
For the convenience of description, the duration of the developmental period, from
oviposition to formation of rudiments of appendages, is divided into seven stages asshown in Table 1.
Table I.

Time table of the early embryonic development of Pamassius glacialis, Luehdorfia
japonica and Byasa (Atrophaneura) alcinous alcinous at room temperature.

Stage

State of Development

Approximate Age of Eggs
P. glacialis(20-23°C)

L. japonica(12-15 O CJ

B. alcinous(25-30 o C)

Maturation and Fertilization

0- 1 hr

0- 1 hr

0- 1 hr2

Cleavage

2-16 hr

2-14 hr

2- 8 hr3Formation of Blastoderm and Appearanceof Primary Dorsal Organ

17-20 hr

15-18hr

9-12hr4

Formation of Ventral Plate and Serosa

21-24 hr

19-23 hr

13-15hr5Formation of Germ Band and Anatrepsis
Formation of Protocephalon and Protocorm

1- 1 .5 days

1 day

16-20 hr

2- 3.5 days

1.5- 2days

1 day

4- 6 days

3- 5 days

1.5-2 days

6

7

Formation of Inner Layer

and Segmentationof Ectoderm and Mesoderm

Embryo Longest in Pre-revolution Stage
Formation of Rudiments of Appendages andof Neural GroovePrimordial germ cells of Papilionidae

257

vs
l

Figs. 1-6. Formation of primordial germ cells in Parnassius glacialis.
1. Cross section of about 23 hr-old egg through a region about one-third of the egg
length from its posterior end. 2. Cross section through the lateral side of ventral
plate at about 23 hr after oviposition, showing the mitotic figures of the somatic
cells. 3, 4. Cross sections through the ventral side of ventral plate at the samestage of Fig. 2, showing the mitotic figures (Fig. 3) and the invagination (Fig. 4) ofprimordial germ cells. 5, 6. Cross sections through the ventral side of ventral
plate at about 27 hr after oviposition (Fig. 5) and at the slightly later stage thanthat shown in Fig. 5 (Fig. 6).
DS, dorsal side of ventral plate; PG, primordial germ cell; RSE, rudimentary serosa; SC,
somatic cell; SE, serosa; SVM, secondary vitelline membrane; VI, vitellophagus; VM, vitelline membrane; VS, ventral side of ventral plate; Y, yolk.

M. Tanaka

258

Figs. 7 - 11.

Primordial germ cells of embryo of Pamassius glacialis, Luehdorfia japonica andByasa alcinous alcinous.

7. Sagittal section of 45 hr-old embryo of P. glacialis, showing a cluster of primordial germ cells. 8. Cross section of 12 hr-old embryo of B. alcinous alcinous, showing primordial germ cells. 9. Sagittal section of 24 hr-old embryo of L. japonica,
showing primordial germ cells. 10. Sagittal section of 2. 5 day-old embryo of P.
glacialis, showing a furrow at the region where the cluster exists. 11. Sagittal section of 2. 5 day-old embryo of P. glacialis, showing the region in which primordial
germ cells are located .
GB, germ band ; FU, furrow on the ventral surface of germ band; PCE, protocephalon;pco, protocorm; PGC, primordial germ cell; SE, serosa; YN, yolk nucleus. Scales: 30 fL m
in Figs. 7 -10, 100 fL m in Fig. 11.

Primordial germ cells of Papilionidae

259

1. Parnassius glacialis
Like in other lepidopteran eggs, there are neither polar granules nor pole cells in
the egg of this species.
During stage 4, the equatorial region of the blastoderm becomes thickened to forma ventral plate, and then the ventral plate is covered with the extra-embryonic area,
the developing serosa (Fig. 1). At this time, the cells of ventral plate are cuboidal inshape and continue to increase in number by cell proliferation. Several mitotic figures
are found at the mid-line of the region at about one-third of the ventral plate. These
mitotic figures are of primordial germ cells and are clearly distinguishable from thoseof the somatic cells by the location and the direction of spindles. The mitotic spindlesof primordial germ cells do not show any definite directions (Fig. 3), while those of thesomatic cells are always parallel to the surface of the ventral plate (Fig. 2). This is thefirst appearance and multiplication of the primordial germ cells. A few cells, invaginating from the ventral plate into the yolk, are found in same material. They are also theprimordial germ cells. At the region where the primordial germ cells invaginate into
the yolk, a furrow is formed between the cells (Fig. 4). In other cases, two-cell layer
produced by the divisions and one or more primordial germ cells lying in close contact
with the ventral plate (Figs. 3 - 5) were observed. The feature of primordial germ cells
is quite the same as that of invaginating cells, namely those cells are polygonal andhave a distinct nucleus with one or two nucleoli and many tiny chromatin granules.
The morphological difference between primordial germ cells and somatic cells is not
yet remarkable except the shape. The nuclei of the former, however, are slightly larger
than those of the latter and are almost the same as those of the yolk cells. Thus theprimordial germ cells are clearly distinguishable from the somatic cells before the completion of serosa.
At the next step, the furrows resulted by invagination of the primordial germ cells
are completely closed, but yet the arrangement of somatic cells at this position isirregular.
By the middle stage 5, in the cup-like embryo sunk into the yolk, multiplied primordial germ cells aggregate and make a group or cell-mass at the region where theydifferentiated originally. At this time the mitotic figures of primordial germ cells arestill seen (Fig. 6). In a sagittal section, in the latter half of stage 5, the embryo shows
C-shape and has a remarkable cluster of primordial germ cells, protruding into theyolk (Fig. 9). The proliferation of primordial germ cells is lost at this stage. The number of primordial germ cells in a cluster is 25 - 35, shows a remarkable variation
among the individuals. Each nucleus of primordial germ cells is about 9 - 10 f! m indiameter, is slightly larger than that of somatic cells and has one or two distinct nucleoli stained with haematoxylin. Difference of affinity for dyes is not yet remarkable between the primordial germ cells and the somatic cells. Position and structure of thecluster remain for a while.
At the beginning of stage 6, the pot-like embryo is deeply embedded by the yolk in
its ventral side. In cross and sagittal sections the embryo is C-shaped. The primordial
germ cells are located at the middle portion of protocorm of the embryo (Fig. 11). At
this stage a shallow furrow is formed temporally, at the region where the cluster exists(Fig. 10). In advance of development, the embryo turns sideways in the posterior half

260

M. Tanaka

Figs. 12-15. Primordial germ cells of Pamassius glacialis and Luehdorfiajaponica.
12, 13. Sagittal sections of the protocorm of embryo of P. glacialis, showing agroup of primordial germ cells at the same stage in Fig. 11. (12) and at the slightly later stage than that in Fig. 12 (13). 14. Cross section of the fifth abdominal
segment of L. japonica at about five days after oviposition, showing the migrating
primordial germ cell. 15. Longitudinal section of the fifth and the sixth abdominal segments of L. japonica at the same stage in Fig. 14, showing primordial germ
cells surrounded by mesodermal cells.
ABS5, 6, fifth and sixth abdominal segments; AM, amnion; ECT, ectoderm; GB, germ
band; MES, mesoderm; NB, neuroblast; NG, neural groove; PGC, primordial germ cell;SE, serosa; YN, yolk nucleus. Scales: 30 f1 m.

Primordial germ cells of Papilionidae

261

of the egg. During this process the connection among the primordial germ cells is gradually lost and rearrangement of the cells occurs to form a cell-mass in close contact
with the inside of the embryo (Fig. 12), and in some cases the germ cells lie apart fromthe wall of the embryo in a space between the spherical yolk-masses (Fig. 13).
Early in stage 6, just before the beginning of the inner layer formation, a shallow
groove, i. e. primitive groove, makes its appearance along the mid-line of the embryo.
The primordial germ cells stay as a single cell-mass at their original position, but inrare cases they are divided into two lateral groups by the groove. As the middle part, i.
e. middle plate sinks inside and forms the inner layer, the primordial germ cells become
scattered in the limited area and are surrounded by the mesodermal cells. It becomesdifficult to discriminate the former from the latter at this stage, because the structuraldifferences between them are not yet distinct, except the size of nuclei and the amountof cytoplasm.
Up to the middle of stage 6, the ectodermal parts of embryo undergo segmentationand the mesodermal cells extend gradually to the lateral side along the inner surface ofthe ectoderm. The author failed to trace the behaviour of primordial germ cells during
the latter half of stage 6.
In stage 7, as the formation of neural groove progresses, the mesoderm is divided
into lateral groups, and primordial germ cells are also divided into two symmetricalgroups by this groove. Then the germ cells are carried to the lateral sides of theembryo together with the mesodermal cells. In this stage the primordial germ cells,
localized in the fourth to the sixth abdominal segment, become to be characterized,that is, they are of light appearance, much larger in size than those of mesodermal cells
because of rich cytoplasm and a large nucleus with one or two dark-stained nucleoli
and many tiny chromatin granules. The number of primordial germ cells on each sideand in each segment varies more or less among the individuals. For example, in one
embryo, two, three and two cells were counted on one side of the fourth to the sixth
abdominal segment respectively, and one, two and two on the other side. In otherembryo, their number was three, three and two on one side, and none, four and two on
the other.
The primordial germ cells do not increase in number since they were differentiated.

2. Luehdorjia japonica
Neither the polar granules nor the pole cells are found in the early stage of development. The first differentiation of the primordial germ cells could not be observed.
Late in stage 4, the broad equatorial zone, i. e. ventral plate, comes to be distinguished and the cells of this region divide actively to form the columnar cells during theformation of the serosa. The germ band increases in length, diminishing its width, andbecomes L-shaped in a sagittal section. It encircles three-fouths of the equator of theegg. At this stage several cells, between the germ band and the yolk, are observed at
the region about one-third of the germ band from its posterior end (Fig. 9). These are
the primordial germ cells.
At the end of stage 5, the germ band differentiates to the protocephalon and the262

M. Tanaka

protocorm, and their ends curve into the yolk. The yolk undergoes cleavage to form
large spherical masses. At this time 20 - 30 primordial germ cells between the embryoand the yolk spherical masses are observed. These cells do not distribute separately,
but assemble as a group or cell-mass. Each cell is almost spherical in shape, about 16
p. m in diameter and has a distinct nucleus with one or two nucleoli deeply stained.
During stage 6, the embryo assumes the coiled form, directs to the anterior pole ofthe egg, and lies wrapped around the yolk. The primordial germ cells lie at the region
one-fifth of the embryo from its posterior end, along the mid-line. They are distributed
somewhat separately, closely contacting with the inside of embryo. As the formation of
inner layer progresses, the primordial germ cells come to be embedded in the mass of
inner layer and to be undistinguishable from neighbour cells.
At the middle of stage 6, the embryo sinks deeply into the yolk. The segmentationof ectoderm and inner layer take place. At the next step the mesodermal cells spreadlaterally and divide into two lateral groups. The primordial germ cells with large nuclei, about 8 p. m in diameter, make their appearance on the both sides in the fourth tothe sixth or the fifth to the sixth abdominal segment (Fig. 14). In the longitudinal sections they lie at the anterior half of each segment (Fig. 15). The number of primordial
germ cells is five or six on one side of the body, namely one to three on one side ofeach segment. The divisions of primordial germ cells are not found until the stage ofgonad formation.

3. Byasa alcinous alcinous
In a newly laid egg and the early stage of development, i. e. from cleavage to thedifferentiation of embryonic and extra-embryonic areas, the primordial germ cells are
not found.
At the end of stage 4, the germ band lying on the ventral side of the egg, is Cshaped in sagittal section and encircles two-thirds of the egg periphery in cross section
through the equator of the egg. Twenty-five to 35 primordial germ cells are observed at
the region about one-third of the germ band from its posterior end, along the mid-line.
They lie in close contact with irregularly arranged, columnar cells of the germ band,making a cell-mass loose assembled as observed in L.japonica (Fig. 8).
When the differentiation of protocephalon and protocorm takes place in the middleof stage 5, the mass of primordial germ cells is located between the embryo and thespherical yolk masses at the region about one-forth of embryo from its posterior end.
Each primordial germ cell has a distinct nucleus which is as large as that of somatic
cells, and it does not represent remarkable characteristic as seen in the later stage ofdevelopment.
By the end of stage 5, the elongated embryo, directing its cephalic part to theanterior pole of the egg, occupies the periphery from the lateral side to the posteriorpole of the egg. The primordial germ cells are in close contact with the inner surface ofembryo as a single cell-mass, and some of them lie among the somatic cells at the posterior region of the embryo. This state is similar to that of L. japonica.
In the early part of stage 6, the middle part of the embryo sinks into the yolk, andfinally the inner layer is formed. Through this process the primordial germ cells arePrimordial germ cells of Papilionidae

263

embedded in the mass of inner layer and come to be undistinguishable from the othercells.
By the end of stage 6, the segmentation of ectodermal part and of mesoderm of theembryo takes place. At this time the primordial germ cells begin to spread laterallywith the mesodermal cells, and then they are completely divided into two symmetricalgroups by a neural groove. In stage 7, the typical primordial germ cells become clear
observable on both sides of the fourth to the sixth abdominal segment. The number ofthem varies among the individuals as that of other species. Table 2 shows the distribution and the nl:lmber of primordial germ cells on each side and segments at the stage ofgonad formation.

*

Table 2. Number and distribution of primordial germ cells in the fourth to the sixth abdominal segment of Byasa alcinous alcinous.
dominalSegment
SampleNumberNumber and Distribution of
Primordial Germ Cells

Total Numberof Primordial
Germ Cells

4th

5th

6th

3
5

4
4

5
4

12
13

2

5
4

5
5

4
4

14
13

3

3
2

4
5

2
4

9
11

4

4
4

5
5

5
4

14
13

5

3
3

4
5

4
4

11
12

Discussion
Many investigations concerning the period and the process of germ cell formation
in insects have been done. Results obtained, however, differ remarkably according to
the species, families and orders of insects. Though considerable number of works onthese subjects have been also reported in the Lepidoptera (Toyama, 1902, in Bombyx;
Schwangart, 1905, in Endromis; Yamauchi, 1917, in Antheraea; Johannsen, 1929, in Diacrisia; Eastham, 1930, in Pieris; Sehl, 1931, Ephestia; Lautenschlager, 1932, in Solenobia;
Saito, 1937, in Antheraea; Kawaguch and Miya, 1943, in Bombyx; Miya, 1950, 1951,
1952, 1953, 1957, 1958, 1959, 1975, in Bombyx; Presser and Rutschky, 1957, in Heliothis;Okada, 1960, in Chilo; Bassand, 1965, in Zeiraphera; Anderson and Wood, 1968, in
Epiphyas; Tanaka, 1970, in Ancylolomia; Ando and Tanaka, 1980, in Endoclita;
Kobayashi, 1983, in Neomicropteryx; Kobayashi and Ando, 1984, in Neomicropteryx; Tanaka, 1985, in Amata), the author's opinions are not always the same among different species or even in the same species.

264

M. Tanaka

Nelsen (1934) and Johannsen and Butt (1941) classified the insects into six andfive groups respectively, considering the time of primordial germ cell appearance.
Furthermore, Miya (1958) divided insects into several groups according to the process
and the position of germ cell differentiation. In his classification the lepidopterous insects belong to three groups. Namely first, primordial germ cells differentiate at or near
the posterior pole of egg, but polar granules are obscure. Next, primordial germ cells
differentiate, or probably differentiate, at the definite region of ventral side of egg, andthey appear in a group and scatter in several segments later. Last, it is the same as thesecond type, but primordial germ cells are already scattered in several segments at thetime of appearance. Almost lepidopterous insects, including Parnassius, Luehdorfia andByasa belong to the second type.
The last type is known in Antheraea (Yamauchi, 1917; Saito, 1937), Diacrisia(Johannsen, 1929), Pie'ris (Eastham, 1930) and Neomicropteryx (Kobayashi, 1983;
Kobayashi and Ando, 1984). Generally, newly differentiated primordial germ cells have
not yet striking feature, and the typical features of them make during the pFoceeding ofdevelopment. For instance, in P. glacialis, L. japonica and B. alcinous alcinous the primordial germ cells become remarkable at the time when the mesodermal cells spread laterally, along the inner surface of ectoderm, after the segmentation of ectodermal part ofembryo and of inner layer. Thus there is a time lag between the first appearance andthe emergence of striking feature of primordial germ cells.
In Bombyx mori, Kawaguchi and Miya (1943) described that primordial germ cells
originated as a group of cells from the primitive ectoderm before the mesoderm formation, and Miya (1950, 1951, 1952, 1953, 1958, 1959) made it clear that they wereformed through the process of invagination at the ventral side of germ band when theblastoderm differentiated to the germ band and extra-embryonic region. Moreover, he(1950, 1957, 1958) presumed, by the results of cauterization experiments, that the cells
destined to be differentiated to the primordial germ cells were predetermined in theblastoderm stage. In Heliothis zea, Presser and Rutschky (1957) and in Epiphyas postvittana, Anderson and Wood (1968) also observed the primordial germ cell differentiation
through the invagination process. Therefore, it seems to be the dominant opinion thatthe first multiplication of primordial germ cells is finished during the blastoderm stage,
and then they are distinguished from the somatic cells through the invagination process
shortly after germ band formation. Recently Tanaka (1985), in Amata fortunei, reported
that several mitotic figures distinguished from those of the somatic cells existed at the
mid-line near the center of newly formed germ band, and they show the first appearance of the primordial germ cells.
In P. glacialis, the morphological differences between the primordial germ cells andthe somatic cells were not yet recognized at the stage when the blastoderm began todifferentiate to the ventral plate and the rudimentary serosa. In the next stage, however, the multiplication and the invagination of primordial germ cells underwent in theregion about one-third from the posterior end of ventral plate, and then they aggregated to form a single cell-mass consisting of 25 - 35 cells. These facts, as observed in
A. fortunei and in P. glacialis, suggest that several cells localized at the definite region of
the blastoderm are destined to differentiate to the primordial germ cells, that is, theprimordial germ cells are predetermined at the blastoderm stage as pointed out by
Miya in Bombyx mori.

Primordial germ cells of Papilionidae

265

Acknowledgements
The author wishes to express his cordial thanks to Prof. Hiroshi Ando, SugadairaMontane Research Center, University of Tsukuba, and Prof. Czeslaw Jura, Jagiellonian University, for their kind advices and critical reading of the manuscript.

References
Anderson, D. T. and E. C. Wood, 1968. The morphological basis of embryonic movements inthe light brown apple moth, Epiphyas postvittana (Walk. ) (Lepidoptera: Tortricidae). Aust.].
Zool. 16: 763-793.
Ando, H. and M. Tanaka, 1980. Early embryonic development of the primitive moth, Endoclita
signifer Walker and E. excrescens Butler (Lepidoptera: Hepialidae). Int. ]. Insect MOTphol.
Embryol. 9: 67-77.
Bassand, D., 1965. Contribution a I'etude de la diapause embryonnaire et de l'embryogenese de
Zeiraphera griseana Hubner (=z. diniana Guenee) (Lepidoptera: Tortricidae). Rev. Suisse Zool.
72: 431-542.
Eastham, L. E. S., 1930. The embryology of Pieris rapae. Organogeny. Phil. Trans. Roy. Soc., SeT. B
219: I-50.
Johannsen, O. A., 1929. Some phases in the embryonic development of Diacrisia viTginica Fabr.
(Lepidoptera).]. MOTPhol. 48: 493-541.
- - - - - , and F. H. Butt, 1941. Embryology of Insects and Myriapods. McGraw-Hill, New
York, London.
Kawaguchi, E. and K. Miya, 1943. Embryonic development of the germ cells in the silkworm,
Bombyx mori L. (inJapanese).Jpn.]. Genet. 19: 133-134.
Kobayashi, Y., 1983. The Embryology of Primitive Moths, Neomicropteryx nipponensis Issiki and
Nepticula castanopsiella Kuroko (Insecta, Lepidoptera). Doctoral thesis, University of Tsukuba.
- - - - - , and H. Ando, 1984. Mesodermal organogenesis in the embryo of the primitive
moth, Neomicropteryx nipponensis Issiki (Lepidoptera, Micropterygidae).]. Morphol. 181: 29-47.
Lautenschlager, F., 1932. Die Embryonalentwicklung der weiblichen Keimdruse bei der Psychide
Solenobia triquetrella. Zool.Jb. Anat. Ontog. 56: 121-162.
Miya, K., 1950. Studies on the development of gonad of the silkworm, Bombyx mori L. I. On theregion and period of differentiation of germ cells (in Japanese with English summary). Trans.
Sapporo Nat. Hist. Soc. 19: 36-39.
- - - - - , 1951. On the numerical variation of germ cells in the embryos of four strains of
the silkworm (Studies on the development of gonad of the silkworm 11) (in Japanese). ].
Seric. Sci. jpn. 20: 221-225.
- - - - - , 1952. On the distribution of germ cells in the embryo of silkworm (Studies on thedevelopment of the gonad in silkworm Ill) (in Japanese with English summary). Jpn. J.
Genet. 27: 48-55.
- - - - - , 1953. The presumptive genital region at the blastoderm stage of the silkworm egg.
]. Fac. Agr., Iwate Univ. I: 223-227.
- - - - - , 1957. Studies on the development of the gonad in the silkworm, Bombyx mori L. 7.
Differentiation of germ cells in eggs cauterized at the cleavage stage. Jpn.]. Genet. 32: 153157.
- - - - - , 1958. Studies on the embryonic development of the gonad in the silkworm, Bombyx mori L. Part. I. Differentiation of germ cells.]. Fac. Agr., [wate Univ. 3: 436-467.
- - - - - " 1959. Studies on the embryonic development of the gonad in the silkworm, Bombyx mori L. Part 11. Formation of the gonad.J. Fac. Agr., [wate Univ. 4: 126-151.
- - - - - , 1975. Ultrastructural changes of embryonic cells during organogenesis in the silkworm, Bombyx mori. I. The gonad.]. Fac. AgT., Iwate Univ. 12: 329-363.

266

M. Tanaka

Nelsen, O. E., 1934. The segregation of the germ cells in the grasshopper, Melanoplus differentialis.
J. Morphol. 55: 545-574.
Okada, M., 1960. Embryonic development of the rice stem-borer, Chilo suppressalis. Sci. Rep. Tokyo
Kyoiku Daigaku, Sec. B 9: 243-296.
Presser, B. D. and C. W. Rutschky, 1957. The embryonic development of the corn earworm,
Heliothis zea (Boddie) (Lepidoptera, Phalaenidae). Ann. Entomol. Soc. Amer. 50: 133-164.
Saito, S., 1937. On the development of the tusser, Antheraea pernyi Guerin-Meneville, with specialreference to the comparative embryology of insects. J. Fac. Agr., Hokkaido Imp. Univ. 40: 35109.
Schwangart, F., 1905. Zur Entwicklungsgeschichte der Lepidopteren. Biol. Zhl. 25: 721-729.
Sehl, A., 1931. Furchung und.. Bildung der Keimanlage bei der Mehlmotte Ephestia kuhniella Zell.
J}ebst einer allgemeinen Ubersicht iiber den Verlauf der Embryonalentwicklung. Z. Morphol.
Okol. Tiere 20: 535-598.
Tanaka, M., 1970. Embryonic development of the rice webworm, Ancylolomia japonica Zeller. 11.
From yolk segmentation to gnathal segment formation (in Japanese with English summary).
New Entomol. 19: 65-71.
- - - - - , 1985. Early embryonic development of Amatafortunei (Lepidoptera, Amatidae). In
H. Ando and K. Miya (eds. ), Recent Advances in Insect Embryology in Japan, 139-155.
Arthropod. Embryo!. Soc. Jpn. (ISEBU Co. Ltd., Tsukuba).
Toyama, K., 1902. Contribution to the study of silkworm. I. On the embryology of the silkworm. Bull. Coll. Agr., Tokyo Imp. Univ. 5: 73-118.
Yamauchi, G., 1917. On the embryonic development of the tusser, Antheraea pernyi (in Japanese).
Sanshigakuho, 2: 356-368.

Author's address: Dr. M. Tanaka
Kan6 High School, Nany6-ch6, Kan6,
Gifu 500, Japan