| Matsucoccus matsumurae (Kuwana) is the type species of the genus Matsucoccus Cockerell (Hemiptera:Coccoidea:Matsucoccidae). In1903, this pine bast scale was first discovered by Kuwana in Pinus thunbergii Parlatore in Tokyo, Japan and was published as a new species. Since invaded into China during1940s-1950s, this pine bast scale have became one of the most destructive exotic pests in pine forest and had caused a sizable economic loss. Though chemical control was carried continuously in the past half century, this pest still damages the pine forest seriously in a great scale in recent years. The main reason is that the pest is with a complex metamorphism in its life cycle and self protection from waxes secreted from the wax pores in cuticle. Therefore, it is very important to study the variety of the wax glands and wax secretions in whole life cycle for exploring new measure to control this pest. In our project, the insect survey and specimen collection were carried out as the insect development stages in the sample plot of pine forest at Jinhua, Zhejiang Province in China where this pine bast scale infests seriously. The characteristics of the wax secretions, wax pores and wax glands in different developmental stages were investigated with light microscopy, scanning electron microscopy (SEM) and transmission electron microscopy (TEM). The results are as follows.1. The development, metamorphism and wax secretion of M. matsumurae in different stagesThe results showed that the male insect of M. matsumurae is holometabolism and in order to complete the life history it must pass through6stages, the1st instar crawler,1st instar settled down nymph, the2nd instar nymph with pearl-shaped body, the3rd instar nymph that secrete wax filaments to form a wax cocoon so as to pupate, the pupa stages, and adult stages; the female insect is hemimetabolism and passes only4stages, from the1st crawler,1st instar settled down nymph, the2nd instar nymph with a pearl-shaped body, to adult stage. In adult stage, the adult males copulate with adult females, after that the adult males died while the adult females secrete wax filaments to form ovisac and lay eggs.The observation found that male scale insects had a different wax secreting with the females. The1st instar crawlers were without wax secretion on body surface, and after settled down the1st nymphs began to secrete wax. The2nd instar nymphs secreted only a little wax over body surface but the spiracles secreted long and white wax filaments. The male scale showed the critical wax-secreting action in the3rd instar nymph stage. It secreted the single, long, and white filaments on the cuticle to construct a wax cocoon. In the cocoon, the nymph pupated and was protected. The adult male was without visible wax over body surface but secreted a bunch of white and long wax filaments from the tergum of the VII abdominal segment. The female scale insect had a most hearty wax-secreting action in its adult stage. The adult female secreted abundant wax filaments from the cuticle of thorax and abdomen, and abundant at the abdomen rear, to form ovisac in which the eggs were protected.2. The observation of structure and cytological characteristics of wax glands in different development stagesThe observation found the fine structure of the wax secretions and wax pores of M. matsumurae in different stages by using scanning electron microscopy (SEM). On body surface of the1st instar nymph, almost without wax pore was observed except two lines of spiracles that played the part of wax-secreting. The spiracles were circular that comprised of an center concave area with a pore for gas exchanging, and a outer ring area with a series of pores for wax secreting. Over body surface of the2nd instar nymph, although a thin layer of wax was observed the wax pore was not visible. The spiracles became bigger and which secreted some long wax filaments from the outer ring pores. This is the first discovery about the spiracles of M. matsumurae with wax-secreting function.The3rd instar male nymph was the most important wax-secreting period in life cycle of the insect. Much more wax pores were observed over the body surface in the thorax and abdomen, in which, there were monolocular pores, biolocular pores that was with a great numbers, trilocular pores and quadrilocular pores. The wax filament was solid structure with a swollen end.On the abdominal segment VII of the male adult, a group of about18tubular ducts were observed. Each tubular duct was with five small pores that secreted the filaments. This is a typical feature of the species of the family Matsucoccidae.The adult female secreted wax filaments from wax pores on the body surface. A large number of wax pores with8-12loculars were observed on the surface of abdomen and particularly abundant at abdomen end. Such multilocular pores secreted wax filaments to form ovisacs. In addition, many dorsal scars were observed. Each dorsal scar was very like a big pit on the surface. It was said that the adult female release sex pheromone through dorsal scars. However we observed some wax around margin of dorsal scars.In prior studies, the wax filaments secreted by the species of Neococcomorpha were with a hollow structure that probably provided with a better carrying capacity in as possible as small quantity of materials used. In contrast, the most wax filaments secreted by M. matsumurae were with a solid structure which probably was a character of an ancient insect. If so, it is in accord with that the family Matsucoccidae was classified into palaeococcomorpha in Coccoidea.The complex cells of several kinds of wax glands of M. matsumurae were observed by using paraffin section microscopy and transmission electron microscopy (TEM). It was found under the open of spiracle there was a complex structure with a wax-secreting function that composed of a wax reservoir in cylinder shape located at the center and several wax-secreting cells that were closely around. Dense granules of wax synthesized were visible inside the wax-secreting cells. In the2nd instar nymph stage, some cells arranged orderly in epidermis layer were observed. They were very probable the wax-secreting cells, but they didn’t form a clear complex wax gland structure The wax gland structure of the3rd instar male nymph had two center cells with very big cubage, which composed of a wax reservoir and several small lateral cells. The wax gland of tubular duct located at the abdominal segment VII of the adult male was a complex structure comprising with a group of the wax-secreting cells in cluster at bottom, a wax reservoir placed at middle, and a tubular duct at the top leading to the pore.The wax glands of multilocular pores in the adult female were very developed. Such a wax gland composed of a center cell with bottle-shaped and several lateral cells that were bigger. By magnifying view, the synthesized wax substances were observed that filled up the wax reservoir. Some ducts connected with the center cell and led to the pores.Many rough endoplasmic reticulums (RER) and bigger nucleus were observed in the wax-secreting cells.Although the wax pores of the adult females of M. matsumurae were observed under SEM by Yang et al. our study observed wax pores on the insect body in all instars but some new wax pore were observed. In addition, we observed the cytological characteristics of wax glands by TEM. This provided a direct evidence for the process of wax synthetising, storing and secreting in the wax glands. It is the first time in the study of M. matsumurae.3. The changes of fatty acid synthase (FAS) and fatty acid elongase (FAE) inside the scale insect bodyThe fatty acid synthase (FAS) and fatty acid elongase (FAE) are associated with the wax synthesis. According to the wax-secreting characters in different stages of M. matsumurae, we determined the contents of FAS and FAE in the insect body by using ELISA methods. The result showed that the content changes of FAS and FAE have a close relativity with wax-secreting action. In the stage of the3rd instar male nymph, the content of FAS in the insect body reached the highest value with2.117±0.150nmol/L, that followed by the stage of the2nd instar nymph with1.642±0.107nmol/L, and the lowest content was detected in pupal stage with0.995±0.046nmol/L. In comparison, FAS content also changed in the body of female pine bast scales. In the stage of the2nd instar nymph, FAS content was1.642±0.107nmol/L, while the highest period appeared in the stage of adult female with2.286±0.158nmol/L before the insect starting to secrete wax and with2.203+0.104nmol/L at the medium-term of wax-secreting action; after deposited eggs, FAS content dropped quickly to1.143±0.225nmol/L. FAE content changes in body of M. matsumurae were similar to FAS, but it was relatively high in the2nd instar nymph stage. This proved that FAS and FAE were two key enzymes involved in the wax synthesis process. This result provided biochemical evidence for wax synthesis. |
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