Recent Advances in Insect Embryology in Japan
Edited by H. Ando and K. Miya.
ISEBU Co. Ltd., Tsukuba 1985

191

External Features of S ialis mitsuhashii
Embryo through Development(Megaloptera, Sialidae)

Hiroshi ANDO
Kozo MIYAKAWA
andSatoru SHlMIZU

Synopsis
Changes in external features of Sialis mitsuhashii embryo during the development observed with lightand scanning electron microscopes were described. The successive formation of mouth-parts is shown inphotographs and the external features of developing embryos in S. mitsuhashii and Protohermes grandis(Corydalidae) were compared and the primitive features of them were discussed.

Introduction
Megaloptera has been considered as the most primitive order of holometabolous insects
because the fossil records of them were known as early as at the Permian period. It is likelythat the common ancestor of Megaloptera which diverged at the Upper Carboniferous periodgave rise Mecoptera, Rhaphidioptera and Neuroptera. The modern Megaloptera (or superfamily Sialoidea) consists of two families, Sialidae and Corydalidae. Observation on the lifehistory of the Megaloptera exhibits that this insect group maintains the most primitive
features of endopterygote insects: they are; 1. the larvae are aquatic; 2. the imagoes areincapable of dispersal far from the larval habitat; 3. the eggs are deposited on leaves orrocks above the water as a batch consisted of many hundred eggs; 4. the pupae are exarate
and decticous, and have a good resemblance to adults.
The embryological features of the Megaloptera were hitherto studied for three species referable to two families; Sialidae, Sialis /utaria L. - general embryology with special reference to histological feature (Strindberg, 1915), - experimental embryology with state-192

Ando, H., K. Miyakawa and S. Shimizu

ments of normal embryogenesis (DuBois, 1938), S. mitsuhashii Okamoto -early embryogenesis (Suzuki, Shimizu and Ando, 1981), -oogenesis (Matsuzaki and Ando, 1977);
Corydalidae, Protohermes grandis Thunberg -changes of external features of the embryos
during development (Miyakawa, 1979), - homology of abdominal appendages (Miyakawa,1980). Nevertheless, the amount of embryo logical informations for this order is poor ascompared to other insect orders.
This paper aims to provide the changes of external features in the embryos of Sialis
mitsuhashii Okamoto during the development, as observed with both the light and scanningelectron microscopes, and to give a comparison with those in the embryos of Protohermes
grandis Thunberg.

Material and Methods
Adults and eggs of Sialis mitsuhashii Okamoto were collected at the Goshiki Lake on thefoot of Mt. Bandai, Fukushima Prefecture of Japan in mid-June, 1982. The external formsof early embryos were described in previous paper (Suzuki et ai., 1981). For scanningelectron microscopy, after removal of chorion and embryonic envelopes, the fixed embryos
were dehydrated with an ethyla1cohol isoamyl acetate series, dried by critical point drying
method, and coated with gold. Observation was done under the scanning electron microscope, JSM T-200 of JEOL.

Observations
Two-day old.
(Fig. 1) The protocephalon and pro to corm differentiate in the embryo
covered ventrally with the amnion. The primitive groove becomes more conspicuous andthen it is closed. The protocorm increases in length and decreases in width, exhibiting a signof the metamerization, viz. four gnathal, three thoracic, and two abdominal segments witha large caudal lobe, and the caudal end attains to the egg dorsal side over the posterior poleof the egg. The yolk cleavage is clearly observed around the germ band.
Three-day old.
(Fig. 2) The embryo elongates further due to the longitudinal growth of
the abdomen. The abdomen turns dorsally at the posterior pole of the egg and its tip
reaches the level at about one-third of the egg length from the anterior pole. Metamerizationof the embryo becomes distinct except at the posterior portion of the abdomen. In the
protocephalon there are the primordia of the labrum, antennae and the stomodaeal pit. The
rudiment of the labrum is at first one-lobed. In the gnathal region rudiments of mandible,
maxillae and labia are also formed as paired swellings. No clear boundary is detected between the narrow intercalary segment and mandibular one. Thoracic segments become
somewhat broader than the gnathal ones and bear medioposteriorly pointed small leg buds.
The abdominal part is narrow and long which consists of eleven segments each of whichhaving nearly the same length, but the width is as much the half as that of the thoracic ones.
No abdominal swellings are observed at this stage. The size of the cells composed of theembryo becomes different in this stage.

External features of Sialis embryo

193

1

2

-+---t-abs.10

Fig. 1.

Fig. 2.

Embryo (2 days old) A. Ventral view, B. Lateral view, C. Dorsal view.
IOOt-tm
Embryo (3 days old) A. Ventral view, B. Dorsal view. Scale: 100t-tm

Scale:

abs. 1, 6, 10 1st, 6th, 10th abdominal segments, am amnion, am antennal rudiment, cl
caudal lobe, cpg closed primitive groove, ins intercalary segment, Ibir labial rudiment,
Ibrr labral rudiment, mds mandibular segment, pcl protocephalic lobe, se serosa, stp
stomodaeal pit, thlr.3 rudiment of 3rd thoracic leg, ths.l 1st thoracic segment, yb yolk
block.

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Ando, H., K. Miyakawa and S. Shimizu

3
Ibrr
anr

Ibir

thlr.3
pip

4

B

p

Fig. 3.
Fig. 4.

Embryo (4 days old) A. Ventral view, B. Nearly lateral view, C. Dorsal view.
Scale: lOO,um
Embryo (5 days old) A. Ventral view, B. Dorsal view. Scale: lOO,umabs. 6, 11 6th, 11th abdominal segments, anr antennal rudiment, lbir labial rudiment, lbrr
labral rudiment, mxr maxillary rudiment, neg neural groove, pIp pleuropodium, thlr.3
rudiment of 3rd thoracic segment.

External features of Sialis embryo

195

Four-day old.
(Fig. 3) The entire length of the embryo reaches at its maximum in the
pre-revolution period, occupying about seven-eighths along the long perimeter of the egg.
The rudiments of appendages are developing. The labral rudiment now changes into bilobed. The antennal and mandibular ones become prominent. The maxillary ones have twodeveloping points, the distal and medio-basal, respectively. The paired labial ones are observed between the distal points of maxillae. The rudiments of thoracic legs are threesegmented and grow medioposteriorly. In the first abdominal segment, the pleuropodia
appear as a pair of tiny lobes. Segmentation of the abdomen becomes complete and ten
segments are counted. The neural ridge is not so clear. The invagination of the stomodaeum
becomes pronounced. The cells composed of the surface of rudimental appendages become
smaller in size and smooth in appearance.
Five-day old.
(Fig. 4) The embryo begins to increase in width and decreases in length.
Forward movement of the labial segment begins.
The antennal rudiments become three-segmented. The maxillary ones which had two
points in the previous stage now have three developing points. These are rudiments of,from the base, the lacinia, galea and the maxillary palpus. Apices on the paired la~ial
rudiments become to be located anteroposteriorly on the midline between the maxillaryrudiments, due to narrowness of the space surrounded by the latter. The thoracic leg
rudiments become three-segmented and their distal ends reach the following segment. The
neural ridge becomes deep except in the posterior portion of the abdomen.
Six-day old.
(Fig. 5) The embryo widens especially in the thoracic and first seven
abdominal segments laid on the ventral side of the egg, and the eighth abdominal one islocated at the posterior pole, and the ninth and tenth segments remain on the dorsal side ofthe egg. Morphogenetic movement proceeds in the gnathal region.
The bases of the labial rudiments move forward along the midline and reach the level ofmaxillary ones. Along with this morphogenetic movement the labial segment loses its
territory from the body sides.
Spiracular pits appear in the meso- and metathoracic segments, and three pairs of swellings are observed outside the ganglionic area of each abdominal segment. The one is locatedmedially to the spirac1e, whereas the other two just posterior to the former and on theposterior border of a segment. In the first abdominal segment, the median one of theposterior swellings is the pleuropodium and the anterior and lateral ones appear later. In thesecond abdominal segment these three swellings appear simultaneously. In the following
abdominal segments, a pair of a swelling appears at the site just medially to the tracheal pit,
which later develops into the abdominal filament or gill. This swelling is serially homologous
with two lateral (anterior and posterior) swellings of the first two abdominal segments, andquite resembles that found in corydalid Protohermes grandis embryo (Miyakawa, 1979).
Seven to eight-day old.
(Fig. 6a, b) Early revolution stage. The embryo increases inwidth to cover more than half of the ventral side of the egg, and decreases in length with aventralward turn of the abdominal end. The cephalic lobes grow and their distal ends
extend posteriorly to the posterior end of the gnathal region, thus forming the basic shape ofthe head as a union of the cephalo-gnathal regions. The last two (ninth and tenth) abdominal segments at first turn to the ventral side of the egg with the completion of dorsal c1ose-Ando, H., K. Miyakawa and S. Shimizu

196

5

c
stp

mxr

Ibir

thlr.3

pip
sabs

6a
lor

Ibir

abgr.5

Fig. 5.

Embryo (6 days old) A. Ventral view, B. Nearly lateral view, C. Dorsal view.
Scale: IOOpm

Fig. 6a.

Embryo (7 days old) A. Dorsal view, B. Lateral view. Scale: IOOpm
abgr.5 5th abdominal gill rudiment, Ibir labial rudiment, lor rudiment of lateral ocelli,
mxr maxillary rudiment, pIp pleuropodium, sabs swelling of abdominal segment, stp
stomodaeal pit, thlr.3 rudiment of 3rd thoracic leg.

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External features of Sialis embryo

6b8

Fig.6b.

Embryo (8 days old) A. Ventral view, B. Lateral view.

Fig. 7.

Embryo (9 days old) Ventral view. Scale: 100pm

Scale: 100J.lm

Fig. 8.

Embryo (10 days old) A. Ventral view, B. Nearly lateral view. Scale: 100pm
abdr.3 3rd abdominal gill rudiment, abp pore of abdominal end, lbi labium, lbir labial
rudiment, lor rudiment of lateral ocellus, mxr maxillary rudiment, mxp maxillarypalp.

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ure in this portion. As development proceeds, the eighth abdominal segment also turns
ventrally at the posterior end of the egg. In this area the yolk mass still remains and thedorsal closure is postponed at this period.
The labral rudiment remains of paired condition. All rudiments of appendages in sevendays embryos increases in size; e. g. the maxillary ones extend to the pro thoracic segment,
the rudiments of pro thoracic legs to the first abdominal segment, those of mesothoracic legs
to the third abdominal, and those of metathoracic legs to the fifth abdominal segment.
Among three abdominal swellings appeared in the previous day, the posterolateral ones
Gust posteromedial to the tracheal pit) are seen in the first seven abdominal segments asconspicuous lobes; They are the rudiments of abdominal ftlaments. The pleuropodia andcorresponding swellings observed from the second to seventh abdominal segments at theposition just lateral to the ganglionic region do not develop further, and the anterior
swellings disappear or have been incorporated into the growing posterolateral lobes.
The rudiments of lateral ocelli appear as seven tiny protrusions on the distal portion of
the cephalic lobes at the level of mandibular and maxillary segments. These are the first signof seven ocelli distributed in two rows, three in the dorsal side and four in the ventral (to thelongitudinal body axis). The pit of abdominal end becomes observable.

Nine-day old.
(Fig. 7) Late revolution stage. The embryo expands laterally to envelop
the yolk on its dorsum. This development is accompanied with a rapid consumption of
considerable amount of yolk and the longitudinal expansion of the abdominal part. The
abdomen increases five-thirds times in length during this development, whereas increase inlength of the head and thorax is not so significant. The abdomen turns ventrally as its
seventh segment localizing at the posterior pole of the egg, the tenth segment arriving at thelevel of the first abdominal one, and the tip of the caudal projection reaches the me sothoracic one. Thus, the revolution of the embryo is completed. The antennal rudiments
become three-segmented, and the mandibular rudiments become bifurcated. The developing
thoracic legs become four-segmented, which consist of the coxa, femur, tibia and tarsus.
On the midline of the gnathal region a forceps-shaped structure (in ventral view) whichlies longitudinally and bilaterally just posterior to the labral rudiment and between theantennal ones. The joint of this forceps is located anteriorly and ventral edges of the structure bear nine to ten dentations. This structure is originated from the anterior part of thelabial rudiments.
Ten-day old.
(Fig. 8) Completion of the dorsal closure. The posture of the embryo isapproximately the same as in the previous stage. The whole embryo undergoes further
extension and development of the appendages. The mouth parts unite more compactly andtheir apical portions become sharply pointed and sclerotization proceeds. Each thoracic leg
elongates further and bears setae at the coxa and the femur. The tarsal claws are sclerotized.

Eleven-day old.
(Fig. 9) The embryo becomes full grown and hatches out from theventral surface of the egg-shell.

External features of Sialis embryo

199

9

abg.l

Fig. 9.

Embryo (11 days old) A. Ventral view, B. Lateral view. Scale: 100pm
abg.l 1st abdominal gill, lbi labium.

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Ando, H., K. Miyakawa and S. Shimizu

Discussion
In many holometabolous orders, the germ disc is formed by aggregation of the blastoderm cells along the ventral midline of the egg, and the embryonic rudiment remains over
the yolk surface without sinking into the yolk. The same is true for the megalopteranembryos except that they have an exceedingly broad germ disc or ventral plate (Suzuki etal., 1981).
One of the conspicuous features of the megalopteran embryos is seen in the stage at
about beginning of the tracheal invagination in Sialis mitsuhashii (Fig. 5) and Protohermes
grandis (Miyakawa, 1979, Fig. 5). Both embryos, at this stage, exhibit two or three swellings at either side of abdominal segments, the most lateral one of which gives rise to thetracheal gill or abdominal filament, whereas other swellings are vestigial and degenerate later.
Similar swellings were found in the abdominal segments of the embryos at the corresponding
stages of neuropteran Ascaiaphus ramburi (Kamiya and Ando, 1984) and mecopteran
Panorpa and Bittacus (Suzuki, 1982; Ando and Miyakawa, unpublished) among the holometabolous orders. On the other hand, similar lobes were recently found in embryos at thecorresponding stage of lower insect orders: Thysanura (Pedetontus, Machida, 1981),
Odonata (Euphaea, Norling, 1982), Dermaptera (Anisoiabis, Fuse and Ando, 1983).
These embryonic abdominal swellings or lobes are found widely in the primitive forms of
the lower insect orders. From these evidences, authors believe that the megalopteranembryos exhibit a primitive feature among those of holometabolous orders.
Turning to comparison of the embryos between Sialidae and Corydalidae within the order
Megaloptera, some differences are observed. At the stage when the protocephalon andpro to corm are differentiated, the embryonic area relative to the total egg surface and that ofprotocephalon relative to the total embryo are larger in Sialidae than in Corydalidae. In S.
mitsuhashii and S. lutaria (DuBois, 1938) embryos, the cephalic lobes extend posterolaterally and reach the level of the mandibular segment, whereas in P. grandis embryo, thecephalic lobes extend anterolaterally and do not reach the mandibular segment. This difference of initial state of the cephalic lobes is maintained also in the later developmental stages.
The other differences are found in the labium and anal appendages of both species. These
structure become definitive at the late embryogenesis when the revolution and the dorsal
closure of the embryo take place. The outer part of labium becomes conspicuous in S.
mitsuhashii as a pair of comb-like structures (or dentate ligulae) which covers on the
midline of mouth-parts throughout, whereas it does not develop in P. grandis. The anal
lobes develop in sialid embryos as a non-paired extension, i. e., the terminal filament,
whereas those in corydalid embryos develop into a pair of pygopods with sclerotized distal
claws. These differences of the labium and the anal appendages between Sialidae andCorydalidae, however, are of postembryonic characters adaptive to the environment in theirlarval lives.

Acknowledgements
The authors wish to express their hearty thanks to Prof. H. Inoue, Akita University, andProf. M. Matsuzaki, Fukushima University, for their help on collecting materials for thepresent study.

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201

References

DuBois, A. M. 1938 La determination de revauche embryonnaire chez Sialis lutaria L.
(Megaloptera). Rev. Suisse ZoO;l 45: 1-92.
Fuse, Y. and H. Ando 1983 Embryoni€' development of the earwig, Anisolabis maritima(Dermaptera) by external observation. New Entomol. 32: 9-16. (In Japanese withEnglish synopsis.)
Machida, R. 1981 External features of embryonic development of a jumping bristle tail,
Pedetontus unimaculatus Machida (Insecta, Thysanura, Machilidae). J. Morphol. 168:339-355.
Matsuzaki, M. and H. Ando 1977 Ovarian structures of the adult alderfly, Sialis mitsuhashii Okamoto (Megaloptera: Sialidae).
Int. J. Insect Morphol. Embryol.
6:17-29.
Miyakawa, K. 1979 Embryology of the dobsonfly, Protohermes grandis Thunberg (Megaloptera: Corydalidae). I'. Changes in external form of the embryo during development.
Kontyu 47: 365-375.
- - - - - . 1980 Embryogenesis of the pleuropodia and the abdominal filaments (tracheal gills) in Protohermes grandis Thunberg (Megaloptera: Corydalidae). XVI Int.
Congr. Entomol. Abstracts: 50
Norling, U. 1982 Structure and ontogeny of the lateral abdominal gills and the caudal gills
in Euphaeidae (Odonata: Zygoptera) larvae. Zool. Jb. Anat., 107: 343-389.
Strindberg, H. 1915 HauptzUge der Entwicklungsgeschichte von Sialis lutaria L. Zool.
Anz., 46: 167-185.
Suzuki, N. 1982 The comparative embryology of the Eumecoptera (Insecta, Mecoptera).
Doctoral Thesis, Inst. BioI. Sci., Univ. Tsukuba, 1-151 with 228 figures.
_ _ _ _ , S. Shimizu and H. Ando 1981 Early embryology of the alderfly, Sialis mitsuhashii Okamoto. Int. J. Insect Morph 01. Embryol. 10: 409-418.

Authors' address:

Prof. H. Ando & Mr. S. Shimizu,
Sugadaira Montane Research Center,
University of Tsukuba, Sanada, Nagano386-22, Japan
Dr. K. Miyakawa,
1024, Imafuku, Kawagoe 356, Japan