Feeding Behavior And Transcriptional Responses Of Phenacoccus Solenopsis Tinsley On Different Varieties Of Cotton

The cotton mealybug(Phenacoccus solenopsis)is a polyphagous insect pest of which endangers cotton,vegetables,and ornamental plantsworldwide.It has a strong adaptability making a great threat to the production of agriculture and forestry in our country.Currently,research on P.solenopsis mainly focuses on the origin,distribution,host ranges,damages,morphological and biological characteristics,prevention,and control technologies.However,researches on its feeding behaviors and molecule mechanisms of P.solenopsis are very limited.In our previous research,we found that the fertility and the ability of population growth of P.solenopsis were significantly different on different cotton varieties.For example,the egg number per insect deposited on the cotton variety Zhong-41 is 2.4 times of that deposited on the cotton variety Zhong-59.Thus,this research aimed to study its feeding behaviors on three different varieties of cotton(Zhong-41,Zhong-48 and Zhong-59)by using the electrical penetration graph(EPG)technique.At the same time,in order to reveal differential gene expressions of P.solenopsis fed on different varieties of cotton,we analyzed the genes related to growth,development and metabolism using transcriptomic approaches.The main results were summarized as follows: 1 The EPG analysis of P.solenopsis fed on different varieties of cottonElectrical penetration graphs(EPG)revealed that P.solenopsis showed nine different types of waveforms including np,A,B,C,pd,E,F,G,and R when fed on these varieties of cotton,an additional resting waveform of R similar to that of leafhoppers was found.By analyzing the feeding behavior of P.solenopsis with the two-stage-combined waveform method,we found that the feeding behavior can be divided into two stages;each with three steps.The first stage,np_ABC,is a probing process,consisting of np(probing to select feeding site),np_A(probing in epidermis)and np_BC(probing to select sucking site).The second stage,pd_BCEG process,is a sucking process.When P.solenopsis fed in the phloem,three steps included pd_BC(selection of sucking site),pd_E1(food processing)and pd_E2(sucking phloem sap).However,when the insect sucked in xylogen,pd_E1 was basically not needed.No matter where the feeding taken place(xylogen or phloem),the durations of the waveforms,pd and pd_C,on Zhong-59 were significantly longer than those on Zhong-41.For the duration of the waveform pd_B,no difference was observed between Zhong-59 and Zhong-41 when the insect sucked in phloem.In contrast,the durations of the waveform pd_B on Zhong-41 were significantly longer than that on Zhong-59.The durations of the waveforms E2 and G on Zhong-41 were significantly longer than those on Zhong-59 when P.solenopsis sucked in phloem,whereas the duration on Zhong-59 was significantly longer than those on Zhong-41 when the insect sucked in xylogen.Based on these results,we concluded that Zhong-59 showed stronger insect-resistance characteristics which may be due to the presence of some insect-resistance substances.These insect-resistance substances may exist in the phloem.2 Effect of different insect-resistant varieties of cotton on gene expression of P. solenopsis using transcriptomic approachesWe sequenced a total of 47035 transcripts.Among them,314 with significantly different expressions were selected.These transcripts include 236 down-regulated and 78 up-regulated.The transcripts with a significantly different expression accounted for 0.67% of the total expressed transcripts.The sequenced transcripts were predicted to participate in 56 gene ontology(GO)secondary pathway classifications.Among these classifications,16 showed a rate of >10% secondary classifications and were predicted to be involved in molecular function(binding and catalytic-activity),cell composition(membrane,membrane components,cell,cell components,organelle,organelle components and macromolecular compositions),biological process(cellular process,single-organism process,metabolism,biological regulations,biological process regulations,positioning and orientation).In the GO functional analysis of the differentially expressed transcripts,39 GO terms showed down-regulations,and many of the transcripts were predicted to participate in the four processes including binding,development,catalytic activity and metabolism.A total of 28 GO terms showed up-regulations and many of the transcripts were predicted to participate in the four processes including binding,organelle,catalytic activity and metabolism.From sequencing analysis,we obtained 21 enriched pathways.A total of 46 enriched transcripts were down-regulated,which accounted for 48.4% of the down-regulated unenriched transcripts.These transcripts were predicted to be involved in lipid metabolism,protein metabolism and glycometabolism.On the other hand,14 enriched transcripts were up-regulated,which accounted for 70% of the up-regulated unenriched transcripts.These transcripts are likely to be involved in energy metabolism.Our analyses also revealed the down-regulated transcripts likely involved in oxidative phosphorylation,energy and metabolism.However,several transcripts,possibly involved in defense related pathways,such as epidermal cutin,ceraceous,suberin and melanin,were up-regulated.The transcripts likely involved in the Wnt pathway were also up-regulated,suggesting decreases of feeding and metabolism after P.solenopsis ingested Zhong-59 because of insect-resistant substances in the cotton plant and increases of insect self-defense.The up-regulations of the relevant genes in the Wnt pathway may inhibit the phosphorylation that is mediated by glycogen synthase kinase and suppress the expression of related genes in the phosphorylation process and the synthesis of ATP.This result may support our notion that Zhong-59 is more resistance than Zhong-41.P450 genes relating to gossypol antagonism was screened and analyzed,we found that P450 6a13 and P450 6a14 were expressed significant difference genes.By using GO terms to map GO mechanism and functional diagrams of detoxification,we found that P.solenopsis could probably detoxicate by oxidation reduction effect.