Study On The Relationship Between The Release Of Yeast-like Symbionts And Vitellogenin In The Brown Planthopper,Nilaparvata Lugens(St?l)

Brown planthopper(BPH,Nilaparvata lugens(St?l))is the primary pest in most rice growing regions in Asia.The yeast-like symbionts(YLS)mainly exist in the fat body of brown planthopper,and vertically transmit to the next generation via the transovarial pathway.YLS is of great significance to the growth and reproduction of the brown planthopper.In this study,the correlation between YLS release and Vitellogenin(Vg)was investigated.The research results are as follows:1?The number of total yeast-like symbionts was counted by micromorphology and then the dynamic changes was analysized.The real-time PCR technology was used to determine the of the relative quantity of dominant species,Noda,in the symbiotic yeast fungi.With the growth and development of the BPH,the number of total YLS were detected increased.and the release of YLS and Noda into hemolymph was detected 6 h after the emergence of BPH.After 66 h emergence of BPH,the number of YLS in the hemolymph release per brown planthopper was about 1×105,which was 38 times,17 times,4.7 times,1.8 times and 1.2 times of 6 h,18 h,30 h,42 h and 54 h,respectively.For 66 h emergence,and the relative quantity of Noda was 19,10,3.6,1.4 and 1.1 times of 6 h,18 h,30 h,42 h and 54 h,respectively.Quantitative analysis of Vg protein relative quantity and relative expression of Vg gene was analysized by Western Blot and real-time PCR.The release of Vg protein into hemolymph was detected 18 h after the emergence of BPH,and that was lagged in the YLS.Vg gene relative expression was detected after 6 h the emergence of BPH.Form 30 h to 54 h after the emergence of BPH,the number of YLS and the relative quantity of Vg were increseased.2 ? The effects of different Vg ds RNA sequence fragments and different concentration of ds RNA on the RNAi were optimally investigated by quantitative realtime PCR.Finally,2795-3249 Vg sequence ds RNA was selected as the RNA interference fragment and the concentration was 0.8 ?g/?L.After the Vg gene was silenced,the quantities of YLS and the relative quantity of Noda were decreased.The number of YLS decreased by 63%,52% and 38% at the day2,day 3 and day 4,respectively,showing extremely significant differences.Meanwhile,the relative quantity of Noda decreased by 58%,48% and 43%,(P < 0.001)respectively.Our research showed that the release of YLS from fat body to the hemolymph is related to Vg.3?RNA sequencing results showed there were 148 genes up-regulated and 127 genes down-regulated in BPH of Vg silencing.GO-enriched analysis showed that lipid transport,lipid localization and carbohydrate metabolic process in the biological process part were closely related to the silencing of Vg gene;The top 10 terms of cell components gene were down-regulated,including ribosomal part,cytoplasmic part and so on;The relative expression of genes encoded peptidase,mannase and serine hydrolase in the molecular functional part were significant.The results of KEGG pathway analysis showed that the differentially expressed genes were annotated to ribosome and oxidative phosphorylation pathways.There are 14 significantly downregulated genes were annotated to ribosome pathways,and 11 significantly downregulated genes were annotated to oxidative phosphorylation pathways.Five genes that might be related to the release of YLS were screened,which encoding polyadenylatebinding protein,arginine kinase,50 S ribosome-binding GTPase,Calcium transports endoplasmic reticulum kinase,C terminal adenylate cyclase with specific differentiation,respectively.