| Plutella xylostella (diamondback moth, DBM) is a destructive cosmopolitan pest of cruciferous crops. As reported, the fitness of the insecticide-resistant (R) and-susceptible (S) DBM was decreased sharply under high temperature. Effects of heat stress on expression of caspase and hsp70genes and level of resistance to insecticide in the R and S DBM were studied by biochemistry, molecular biology and resistant bioassays. The results showed that resistance level of DBM was decreased significantly under heat stress, and this decrease was heritable.Caspase is the key factor in apoptosis. The caspase gene, named Px-caspase-1, was isolated and sequenced from the DBM. The full-length of Px-caspase-1cDNA was1172bp, including an903bp open reading frame (ORF) that encoded300amino acids. The dedueed protein had a computed molecular mass of33.6kDa. The Px-caspase-1belonged to effector caspase based on its high identity of amino acid sequences to the effector caspase genes from other insect species.The expression of Px-caspase-1and the enzyme activity of caspase-9and caspase-3under heat stress (32.1、36、40、42、45、47℃) were studied. The results showed that the basic levels of Px-caspase-1expression and caspase-9and caspase-3activities could be detected at25℃. However, the gene expression and the two enzyme activities increased significantly when the adults of DBM were pretreated with high temperature. With the increase of pretreatment temperature and time period, the Px-caspase-1mRNA expression and activities of caspase-9and caspase-3reached the peak value earlier under heat stress than those at25℃. In addition, high temperature could result in the increase of mortality of adult DBM. The results indicated that apoptosis was a normal physiological procession in DBM at normal temperature, and heat stress could significantly induce apoptosis in DBM and cause death of DBM.The expression of Px-caspase-1between R and S adults DBM under heat stress (32.1、40、42℃) were studied. Significantly increase of Px-caspase-1expression was found in female and male adults of R and S DBM when the adults were pretreated with high temperature, and the extent of up-regulation of Px-caspase-Ⅰ expression in R DBM was significantly higher than in S DBM.The heat shock protein70family were invoved in apoptosis under heat stress. The heat shock protein gene, named Px-hsp70, was isolated and sequenced from DBM. The full-length of Px-hsp70cDNA was2128bp, including an1899bp open reading frame (ORF) that encoded632amino acids. The deduced protein had a computed molecular mass of69.3kDa. The Px-hsp70belonged to hsp70gene family based on its high identity of amino acid sequences to the hsp70genes from other insect species.The basic expression levels of Px-hsp70in female and male larvae were lower than those in female and male adults in R and S DBM, in general. There were no significantly difference of Px-hsp70expression in female and male larvae between25℃and high temperature treatments (37and42℃). However, significantly increase of Px-hsp70expression was found in female and male adults of R and S DBM when the adults were pretreated with high temperature, and the extent of up-regulation of Px-hsp70expression in S DBM was significantly higher than in R DBM.The results indicated that the fitness of the R and S DBM was inhibited significantly by high temperature, and the R DBM was inhibited much than S DBM.The level of resistance to insecticide of DBM was influenced markedly by high temperature. Based on the field resistance monitoring, the level of resistance to insecticide decreased sharply in July. The level of resistance to avermectin in the progenies of DBM declined sharply when the insects were reared at33.5℃for one generation. The great decline of avermectin resistance in the progenies of R DBM,when the DBM were reared at high temperature for one generation, might be associated with its significantly lower fitness. |
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