| The western flower thrips, Frankliniella occidentalis is a worldwide insect pest of horticultural crops. F.occidentalis has dispersed to many areas in China and the damage is becoming more and more serious since it was first reported in Beijing in2003. During spread of F.occidentalis, they often encounter heat stress in summer and cold stress in winter. Unfortunately, thermal tolerance and corresponding regulating mechanism in this pest have never been studied. In present study, tolerance and reproduction to cold and heat stress in F.occidentalis and its corresponding physiological mechanism were studied by ecology and molecular method. The main results are as follows:The present study examined the effects of different low temperatures, exposure duration on survival rate and fitness of F.occidentalis. Results indicated that survival rate of both adults and larvae significantly declined with decrease of temperature and extension of exposure duration. During the duration from1to6h, the temperature needed to kill50%individuals (Ltemso) for larvae was higher than that for adult females, but lower than that for adult males. Therefore, the order in cold hardness of different life stages was presented as:adult female> larvae> adult males. When larvae and adults of F. occidentalis were transferred from the normal rearing conditions at26±1℃, to cold temperatures at-13℃(for larvae) or-13.5℃(for adults) for2h, survivorship rate was only25%for larvae and27%for the adults. These two temperatures could be defined as the discriminating temperatures for the two insect stages, respectively. Survival rates of the thrips that were pretreated at0℃or5℃for2h before exposure to their discriminating temperatures increased significantly, and maximum increase in survivorships to the cold temperatures was achieved by pretreating larvae or adults at0℃for2h, which resulted in a survivorship rate of46%,54%,49%for larvae, adult females and males, respectively. When cold exposures occurred at the larval stage, there were no differences in development time, longevity and reproduction duration of enclosed adults relative to the non-treated control. However the number of eggs laid of the adults derived from cold-treated larvae was significantly less than that of the non-treated thrips. When adult thrips were exposed to the cold temperatures, the longevity, fecundity and reproduction duration all decreased significantly compared to the non-cold treated control.This study investigated the effects of high temperatures (33,35,37,39,41℃), exposure durations (0.5,1,2,4h) and life stages on survival and reproduction of the western flower thrips. Results suggested that the survival rate of adults (female and male) and larvae declined significantly with the increase in the exposure time at all high temperature examined. Therefore, the order in heat resistance of different life stages was presented as:adult female> larvae> adult males The lethal temperature needed to kill the50%of the adults and larvae decreased rapidly with the increase in the exposure time from0.5to4h. Surprisingly, preheat treatment at31(adults) or33℃for2h (larvae) significantly improved the survival rate of both stages after exposure to lethal high temperature. After2h exposure, longevity and reproduction of adults lowed remarkably with the increasing temperature. Mating experiment after exposure to high temperature demonstrated longevity and reproduction in all treatments (U(?)+S♀, S(?)+U♀, S(?)+S♀) were lower than those in control (U(?)+U♀). Longevity in U(?)+S♀treatment was not different from that in S(?)+U♀treatment, but was higher than that in S(?)+S♀treatment; while the reproduction in U(?)+S♀was significantly lower than that in S(?)+U♀, and relative to S(?)+S♀, suggesting that high temperature have adverse influence on adult female and male, but the effects on female was more serious than male.Five heat shock protein genes were cloned from F. occidentalis by RT-PCR and RACE, and were named Fohsp90, Fohsc701, Fohsc702, Fohsp60and Fohop, respectively. The complete cDNA of Fohsp90, Fohsc70, Fohsp60and Fohop are2508,2195,3065,2489,2276bp, respectively, and their respective opening reading frames are2169,1920,1983,1728,1659bp, which encode722,639,660,575and552amino acids with predicted molecular weights of83.26,69.81,72.73,60.87and62.25KDa, respectively. Sequence analysis suggested that the five Hsps shared high similarity with those of other species and contained conserved motifs found in their respective family of other species. Analysis of genomic DNA demonstrated that the five hsps contained3,4,7,5,10introns, respectively. In addition, the number and positions of introns of five hsps were highly different from those of other species. Finally, phylogenetic analysis suggested that the five Hsps were homologues of their respective family. Expression patterns of five heat shock protein genes (hsp90, hsc701, hsc702, hsp60, hop) in response to development and different temperatures were examined by Real-time PCR. Results showed that five hsps were expressed at larvae, pupae and adults stage. Among them, hsp90, hsc70and hsp60were significantly up-regulated with the process of development, while the expression level of hsc702and hop were not different among three stages. The five hsps were not very sensitive to cold, but were significantly induced up-regulation by heat, and the expression level of five hsps in larvae were higher than that in adult. In addition, pre-heat treatment lead to significant up-regulation of four genes compared to direct exposure to lethal high temperature (control).The combined effects of elevated temperature and sub-lethal dose of avermectins on biological parameters, antioxidant enzymes and heat shock protein mRNA level were studied. Results showed that temperature and avermectins concentrations significantly influenced the biological parameters. The survival, longevity and reproduction decreased with increased temperature and pesticide concentration. Elevated temperature and avermectins significantly decreased activity of SOD. Elevated temperature had no effect on activity of CAT, but its activitywas obviously improved by the combination of temperature and avermectins. Activities of POD and GST significantly increased after exposure to combination of temperature and avermectins. Expression patterns of Hsps mRNA showed that only three heat shock proteins (hsp90, hsc702and hop) was induced up-regulation by the elevated temperature, and shifted folds is very small. However, five hsps level was significantly up-regulated after exposure to conditions of the combination of elevated temperature and avermectins, and the highest level of hsp90, hsc701, hsc702and hop was reached at33℃, but the highest level of hsp60was observed at21℃.Taken together, relative strong tolerance to extreme temperature and rapid hardening response of the western flower thrips may be important factors that determine its rapid dispersal in china, and the underlying mechanism, especially under heat stress conditions, may be related to up-regulation of the molecular chaperon Hsps. |
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