Regulation Of Temperature,humidity And Photoperiod On Egg Diapause Of Penthaleus Major

Insects can survive adverse environmental conditions by entering diapause,so that individuals can survive,and maintain developmental uniformity to ensure the reproduction of the population.Understanding the diapause characteristics of insect pests is crucial for pest management.Penthaleus major(Dugés)is a major pest mite on winter wheat in China.Although previous studies have been conducted to investigate its biological characteristics,population distribution and occurrence,however,the abiotic factors inducing diapause and the underlying molecular mechanisms of egg diapause in P.major remain unknown.In this study,we explored the induction of egg diapause by abiotic factors including temperature,relative humidity and photoperiod on the egg diapause of P.major.The criteria of diapause eggs and non-diapause eggs was established.Differences in diapause duration of diapause eggs induced under different conditions were compared to identify the main factors affecting the diapause of eggs.Protective enzyme activity of P.major during diapsuse induction were determined to clarify the physiological basis underlying egg diapause.By transcriptomic analysis of diapause eggs and non-diapause eggs,differential expression genes and enrichment pathways related to diapause were screened out,and their possible roles in diapause process were analyzed.The main results are as follows:1.Induction of egg diapause by temperature,humidity and photoperiod in P.majorDiapause induction was performed by manipulating temperature(14℃,19℃ and24℃),humidity(relative humidity 84% and saturated humidity)and photoperiod(10L:14D and 16L:8D).The eggs that did not hatch within 20 days and were not decay were identified as diapause eggs.The results showed that:(1)temperature,humidity and their interaction had significant effects on the induction of diapause,while the effects of photoperiod was not significant.24℃ and 84% relative humidity induced diapause of mite eggs.At 14℃ and 19℃,the relative humidity saturation could not induce egg diapause,and the egg hatching rate was above 94%,while the relative humidity 84%induced egg diapause rate was 100%.At 24℃,the egg diapause rate was 100% under relative humidity saturation and 84% relative humidity.(2)After 20 days of induction at24℃ and relative humidity saturation conditions,the eggs of P.major entered a stable diapause state.The average diapause release time was 74.37 d,and the incubation rate was 88% after diapause release.The duration of diapause at 14℃,19℃ and 24℃ was shorter when the relative humidity was 84%.They were 10.2 d,6.93 d and 25.25 d,respectively.2.Determination of egg protective enzyme activity of P.major during diapause inductionTo clarify the changes of protective enzyme activities during the induction of diapause in P.major egg,enzyme activities were explored for the non-diapause eggs and the diapause eggs under the photoperiod of 10L:14D.The activities of superoxide dismutase(SOD),catalase(CAT)and peroxidase(POD)were measured on 5,10,15 and20 days after diapause induction at 24℃ and saturated humidity.The results showed that the activities of SOD,POD and CAT in the eggs of P.major were significantly higher than those of non-diapause individuals after 5 days of diapause induction.With an extension of diapause induction time,SOD and POD activities decreased,but the activity of CAT continued to increase.After 20 days of diapause induction,SOD and POD activities decreased to the lowest values at 6.73 U/mg prot and 33.23 U/mg prot,respectively,which were significantly lower than those in non-diapause eggs.In contrast,CAT activity was highest during the entire diapause induction process,reaching a peak of6679.89 U/mg prot at 10 days of diapause induction,and decreased by day 20,suggesting that CAT was active during diapause induction of mite eggs,which may be the main defense enzyme.It is possible that with the extension of diapause induction time,the metabolic rate of diapause eggs decreased,and the reactive oxygen species produced in the body may also decrease.At this point,the lower levels of enzyme activity were sufficient to meet the requirements for antioxidant defense,reflecting the ability of diapause eggs to cope with adverse environment.3.Analysis of differences between the transcripts of diapause and non-diapause eggsWe also analyzed differences between the transcripts of diapause and non-diapause eggs.Transcriptomic analysis showed that 5212 genes were up-regulated,while 5295 genes were down-regulated in diapause eggs compared to non-diapause eggs.We annotated a total of 7196 differential genes into the GO database,which were mainly concentrated in binding function,catalytic activity,cellular processes,and metabolic processes.Through KEGG pathway database analysis,we annotated a total of 3035 differentially expressed genes,among which 1529 genes were up regulated and 1506 genes were down regulated.These genes mapped to 303 pathways,mainly involving lysosome metabolism,ribosome and fatty acid metabolism,and carbohydrate metabolism.We screened differentially expressed genes related to lipid metabolism during diapause,such as glycerol kinase and monoglyceride lipase,which were up-regulated,while diacylglycerol kinase gene and glycerol-3-phosphate dehydrogenase 1 were downregulated.The down-regulated gene expression is consistent with the low metabolic level of diapause eggs,whereas the up-regulated gene expression may contribute to enhanced stress resistance.In this study,we elucidated the effects of temperature,humidity and photoperiod on the induction of egg diapause in P.major,and identified the main factor inducing egg diapause.The changes in activities of protective enzymes were determined during diapause induction,indicating that catalase was the main protective enzyme.The relevant genes and metabolic pathways that may be involved in diapause induction were screened,which laid the foundation for the study of the diapause mechanism of P.major.