Chemical Substances Change And Molecular Mechanism Of The Wheat Blossom Midge, Sitodiplosis Mosellana (Diptera: Cecidomyiidae) During Diapause

The wheat blossom midge (WBM), Sitodiplosis mosellana (Gehin) is one of most damaged pests distributing in the main wheat-growing regions of China and other areas of world. Many times of outbrokes and plagues caused great losses to wheat production have been recorded in history., Outbreaks and plagues of WBM have been controlled in broader area in China during recently past decade years, but WBM displayed a trend to extend their habitats at certain wheat-growing areas such as Hebei, Beijing, Shandong and has a potential threat to local wheat yield. Diapause, as a result of evolution and adapting selection of most insects fitting to natural environment changes, is one of the most important ecological and physiological strategies. It is also a special survival form which makes insects to be sustainable under bad environmental conditions for reproduction and development of their population. The wheat blossom midge, Sitodiplosis mosellana (Gehin) is one of typical diapause insects with high diapause ratio, powerful diapause potential, extremely prolonged diapause time and re-diapause. It is reported that diapause polymorphism of WBM is one of key factors affecting population fluctuation with occasional, regional and uneven dumpling characteristics in the filed.In this paper, sorbitol content, activities of trehalases, sorbitol dehydrogenase (SDH) and 3 kinds of protective enzymes, SOD, POD, CAT were determined and DNA polymorphism was studied with 5 repeat sequence primers during various diapause stages of WBM. The main results are as follows.1. Contents of sorbitol in S. mosellana were undetected in larvae collected from wheat heads or from soil before mid-October of the same year. It could be detected with a lower level in larvae from on 25th October of the same year. Sorbitol contents were increased rapidly as diapause time becom longer and temperature gradually decreased afterward. This change suggested that large amounts of sorbitol were accumulated in order to cope with cold in winter. High contents of sorbitol were regarded as index of WBM in over-wintering stage. No significant difference of sorbitol contents was found between non-cocooned larvae and cocooned larvae with the same diapause stage, but the non-cocooned larvae had a lightly high sorbitol content than the cocooned larvae. Sorbitol contents of WBM larvae with 2-year diapause period were also detectable and more than that of larvae with 1-year diapause period at the same phase.2. Activities of trehalase and SDH in WBM larvae were measured during its various diapause stages. Results showed that activity of trehalase in grown larvae collected from wheat heads was higher than that of larvae from soil. It decreased quickly during initial period of larvae entering soil. Uplifted in mid-September. Dropped again in late October of the same year and increased steeply in late February of the next year in turn. There was a close relationship between high activity of trehalase and energy transition for WBM larvae in different diapause stage. Activity of SDH in WBM larvae was not detectable from wheat mature to November. But it increased sharply from January to April. High SDH activiy was regarded as a index for WBM that was in breaking diapause stage. Activities of both trehalase and SDH in Non-cocooned larvae were higher than that of cocooned larva in the same diapause stage. An identical change trend was seen on activity of trehalase and SDH. No significant difference of activity in either trehalase or SDH was found between one-year and two-year diapause larvae.3. Activities of SOD, CAT and POD in WBM larvae were measured and result showed that there exited a remarkable difference of SOD activities in WBM larvae with the different diapause stage. SOD activities were reduced gradually after grown larvae fell into soil from wheat ears. There were not significant differences of SOD activities between no-cocooned larvae and cocooned larvae with the same diapause stage. A similar change trend of SOD activities was seen in both no-cocooned larvae and cocooned larvae. Change gradient of SOD activities of WBM larvae with one-year diapause was higher than that of WBM larvae with two-year diapause.Activities of POD and CAT in mature larvae living on the wheat head were higher than that of diapause larvae in the soil. They dropped quickly after larvae entered into soil from wheat heads. There was not a significant difference of SOD, POD and CAT activity between no-cocooned larvae and cocooned larvae at the same diapause stage. Activities of SOD, POD and CAT in WBM larvae with one-year diapause were slightly higher than that in WBM larvae with two-year diapause, there was a obvious differencc between lavae with one-year diapause and with two-year diapause.4 Chitinase activities and chitin contents were measured in larvae of the wheat midge during mature and diapause stage. The result indicated that there exited remarkable difference for chitinase activities of S. mosellana at the different diapause stage. The chitinase activitives in the cocooned larvae were higher than that of no-cocooned larvae. Chitinase activities of WBM larvae with one-year diapause were slightly higher than that in WBM larvae with two-year diapause. There existed significant differences for chitinase activities of the cocooned larvae of S. mosellana between in the one-year diapause period and in the two-year diapause period, but not existed in the no-cocooned larvae.Chitin contents of no-cocooned larvae in the different diapause stage changed slightly, not existed remarkable differences among them. But the circumstances of the cocooned larvae were opposite to no-cocooned larvae, changed steeply. Chitin contents of cocooned larvae were higher than that of no-cocooned larvae. No obvious difference of were found for chitin contents between in the one-year diapause period and in the two-year diapause period.5 The DNA polymorphism of WBM at different diapause stage was analyzed with 5 repeat sequence primers by the PCR technique. Results showed that (1) five primers generated 104 DNA bands with molecular weights from 202 to 1163bp; (2) Average similarity index of WBM at different diapause stage was in 0.573-0.936; (3) Genetic variation of WBM at different diapause stage was mainly produced from intergroups; (4) different diapause populations of the WBM may be divided into three main categories on the basis of genetic distances. The paper is the first report of the quantified genetic diversity among different diapause population of the wheat blossom midges.6. RNA was extracted from single larva of WBM firstly. An effective method of RNA extracting from the single WBM was proposed. 28S and 18S bands were all clear and not tail phenomena. The two bands of the wheat-ear larvae were lighter and wider than that of cocooned larvae, which suggested slower metabolic activities and lower quantities of RNA transcribe in the diapause larvae. Clone of diapause hormone gene of WBM was experimently carried out from genomic DNA and cDNA, but failed to get any result. There maybe has a lower homology of the diapause hormone gene between WBM and other insects. So it was difficult to design primers according to current DH gene sequence from other insects so as to amplify DH gene of WBM.