| Cassava,as an industrial energy and food crop with high comprehensive utilization values,plays an important role in the cultivation of tropical and subtropical areas.However,the damage from Tetranychus urticae has become increasingly serious in cassava-producing areas,because of its small size,rapid reproduction,strong adaptability,wide host range and capability for resistance.The study on the molecular mechanism of cassava responses induced by the mites can provide basic data for the screening of new resistance genes.Therefore,in this study,multi-omics technology was applied to the study of cassava defences against T.urticae,and the real-time differential expression of genes,proteins and metabolites was analysed to narrow the scope of target genes to screen for the systematic study of the molecular expression profiles of cassava responses to mites.The research results are as follows:1.The mite population number on SC8 was the lowest in the peak period of T.urticae damage,which was 38.7±3.8 mites/leaf.The second was NZ199,which had 53.2±8.6 mites/leaf,while the highest was SC205 with 105.6±6.9mites/leaf.Under a constant indoor temperature of 28℃,different cassava varieties,SC8,NZ199 and SC205,were used to feed T.urticae.There was no significant difference in the duration of the development of the immature stage between SC8 and NZ199,which was 9.16 d and 8.90 d,respectively,but the duration of the development of the immature stage on SC8 and NZ199 was longer than that on SC205.The number of eggs laid by the mites on SC8 and NZ199 was also less than that on SC205.Based on the results of mite resistance,it was concluded that SC8 and NZ199 were moderately resistant,while SC205 was susceptible.2.The most physiological indexes of cassava leaves detected in this paper increased after5-7d of mite infestation.The results showed that the ratio of soluble protein,SOD activity,POD activity,CAT activity and free proline increased significantly higher in SC8 than in SC205,while MDA was the opposite.It was concluded that the above indexes could be used as physiological indicators for judging the resistance of cassava varieties to mites.3.After inoculation with T.urticae,the contents of JA,ABA and IPA in the cassava leaves increased but were significantly affected by different developmental stages of cassava and different mite densities.The contents of BR and ZR and the ZR/ABA ratio decreased.Thus,it was concluded that the changes in the JA,ABA,IPA,BR,ZR contents and the ratios in cassava leaves under T.urticae stress were related to the developmental stages of cassava and mite density.In addition,the metabolic pathways of cassava leaves in different developmental stages after mite infestation were analysed by GC/MS.The 72 identified metabolites included soluble sugars,amino acids,non-protein amino acids,sphingolipids and secondary metabolites.Soluble sugars and amino acids can regulate the osmotic balance of cells and prevent mite stress from destroying the metabolism of cassava.Non-protein amino acids can restrain the activity of enzymes in plants.Sphingolipids can change the structure of cell membrane.Secondary metabolites such as gallic acid and nicotine can resist plant defences against pests.Therefore,it is speculated that the changes in these substances may be related to the defence response of cassava leaves to T.urticae infection.The induction of metabolites and related metabolic pathways were affected by the developmental stages of cassava.The metabolic pathways involved in the two developmental stages of cassava included energy metabolism,carbohydrate metabolism and the citrate cycle(TCA cycle).The specific metabolic pathways in the leaves in the tuber formation stage included nucleotide metabolism,the metabolism of cofactors and vitamins,and the biosynthesis of other secondary metabolites.The specific metabolic pathway in the leaves in the seedling stage was the metabolism of lipid,terpenoids and polyketides.The above analysis of stress metabolites and complex metabolic pathways provides new insights into the mechanism of the response of cassava to mite stress.4.The results showed that the number of differentially expressed genes in the leaves of cassava in the tuber formation stage was 698 and 2140 under lowand high-density mite infestation conditions,respectively,of which 347 genes were co-expressed.The number of differentially expressed genes in cassava seedling leaves exposed to mites at low and high densities was 8554 and 9018,respectively.Among them,6519 differentially expressed genes were co-expressed.After cassava leaves were exposed to 16 mites/leaf,the metabolic pathways related to resistance were flavonoid biosynthesis,phenylpropanoid biosynthesis,glutathione metabolism,isoflavonoid biosynthesis,pentose phosphate,brassinosteroid biosynthesis,ABC transporters,diterpenoid biosynthesis,and flavone and flavonol biosynthesis.After cassava leaves were exposed to 48 mites/leaf,the metabolic pathways related to plant resistance were plant signal hormone transduction,phenylalanine metabolic,flavonoid biosynthesis,phenylpropanoid biosynthesis,plant-pathogen interaction,glutathione metabolism,isoflavonoid biosynthesis,pentose phosphate pathway,etc.Theplant signal transduction pathway was enriched only inthe tuber root formation stage.In addition,cassava induced five transcription factor families related to plant resistance to biological stress after mite stress,namely,the WRKY,MYB,ERF,NAC and b ZIP families.The number of induced transcription factors decreased with the prolongation of the cassava growth period but increased with the increase in mite population density.It is speculated that these transcription factors are involved in regulating cassava’s defence response to mites,and they are more active in the leaves in the seedling stage than in the leaves in the tuber root formation stage and more active under high-density mite stress.5.There were 191 DEPs in the leaves in the tuber formation stage,of which 149 were upregulated and 42 were downregulated;1042 were identified,with 586 upregulated and 456 downregulated,in the leaves in the seedling stage.The number of differentially expressed proteins in the leaves in the seedling stage was greater than that in the leaves in the tuber formation stage after mite stress.A total of 115 DEPs were upregulated by the two treatments.Specific upregulated DEPs in the leaves during root tuber formation were involved in indole alkaloid biosynthesis,beta-alanine metabolism,monoterpenoid biosynthesis,pantothenate and Co A biosynthesis,and the RNA surveillance pathway.The specifically upregulated DEPs involved in resistance-related metabolic pathways in the leaves of seedling were photosynthesis-antenna protein,oxidative phosphorylation,glutathione metabolism,sphingolipid metabolism,phenylpropanoid biosynthesis,indole alkaloid biosynthesis,ascorbate and aldarate metabolism,the pentose phosphate pathway,and the tropane,piperidine and pyridine alkaloid biosynthesis pathways.6.There were 23 differential proteins in the transcriptome and proteome of cassava leaves damaged by T.urticae in the tuber root formation stage,with a correlation coefficient of 0.2953,which is a weak correlation,while 534 differential proteins were associated with cassava leaves in the seedling stage,with a correlation coefficient of 0.49.The overall metabolic network and secondary metabolic network of cassava under the stress of T.urticae at different developmental stages and at different mite densities were integrated using multi-omics data.The common metabolic pathways from multi-omics data included carbohydrate metabolism,energy metabolism,tricarboxylic acid cycle,gluconeogenesis / glycolysis,other secondary metabolism,lipid metabolism and secondary biomass biosynthesis.In the fourth chapter,the differential metabolites induced by mite stress also showed that cassava participated in the above metabolic pathway.In conclusion,the biological characteristics of T.urticae mites on cassava confirmed that the resistant cassava had a certain negative impact on the development of the mites.In this paper,the physiological response,plant hormone changes and the differences of metabolites,proteins and transcriptome levels of cassava under mite stress were systematically mined.It was confirmed that the chemical defences played a very important role in the process of cassava defences against mites,including the generation of damage signals and the transmission of resistance factors,such as jasmonate,abscisic acid,cytokinin,auxin;the changes of nutrients,such as carbohydrate synthesis and energy metabolism;induce related genes to encode defence proteins,such as protease inhibitors,chitinases,pathogen-related proteins;produce plant secondary metabolites,such as polyphenols,terpenes,flavonoids and nicotine,etc.The study provided an important basis for the anti-mite mechanism of cassava. |
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