Effect Of Secondary Endosymbionts Rickettsia On The Resistance Of Sitobion Avenae To Water-Defict Stress

With global warming,the frequency and intensity of extreme drought events in northwestern China are showing an increasing trend.Drought can directly affect the growth and development of individual phytophagous insects,and the response mechanisms of insects to the effects of different water deficit stresses are still unclear.The English grain aphid,Sitobion avenae（Fabricius）,belongs to Aphididae（Hemiptera）,and feeds on plant phloem sap with prickly-sucking mouthparts.The endosymbiotic bacteria of aphids are divided into two main groups:primary and secondary symbiotic bacteria.Symbiotic bacteria usually form a reciprocal symbiotic and synergistic evolutionary relationship with the host aphid.Current research on aphid symbiotic bacteria has focused on helping hosts to resist parasitic and fungal infestation by parasitic wasps,and on improving aphid tolerance to high temperatures,while it is unclear whether secondary symbiotic bacteria can assist aphids to resist water deficit stress.In this study,three Rickettsia-infected clones（infected clones,symbiotic bacteria Rickettsia）were selected based on previous laboratory studies on the secondary symbiotic bacteria Rickettsia in S.avenae.Using ecological and molecular biological methods,we investigated the role and changes of the secondary symbiotic bacteria Rickettsia on S.avenae under treatments of different levels of water deficit stress,and laid the foundation for understanding the evolution of the aphid populations in areas of different levels of drought in northwestern China.The main findings are as follows:1.Determination of the effect of the secondary symbiotic bacteria Rickettsia on the life history parameters of the S.avenae when resisting different water stresses.The results showed that the 1^stand 4^thinstar developmental time and total developmental time were significantly shorter,and the adult aphid weight was significantly higher in the symbiot-infected clone line Ric4 than in the corresponding symbiot-removed clone NRic4under severe stress.The symbiot-infected clone 5-p5 had a significantly shorter 3^rdinstar developmental time,and a significantly higher adult aphid weight and ten-day aphid production compared to the corresponding symbiot-removed clone N5-p5 under severe stress.The clonal strain ZYB34 behaved differently from the other two clonal strains in that the total developmental time of nymphs and 4^thinstar developmental time were significantly higher,and the adult aphid body weight and ten-day fecundity were significantly lower in the symbiot-infected clone line ZYB34 compared to the corresponding symbiot-removed clone NZYB34 under severe stress.It can be seen that the infection of some secondary symbiotic strains of Rickettsia shortens the developmental time of the S.avenae and increases the adult aphid weight and aphid fecundity,thus facilitating the aphid to cope with the environmental conditions of water stress.Also,the endosymbiotic bacteria produced different effects on different genotypes of the host S.avenae.2.The effect of the secondary symbiotic bacteria Rickettsia on the water balance traits（absolute water content and water loss rate）of S.avenae under different water deficit stresses was examined.The results showed that the absolute water content of symbiot-infected clones Ric4 and 5-p5 decreased gradually with increasing water stress;The water loss rate of symbiot-infected clone 5-p5 was significantly lower under both moderate and severe stress compared with the removal of the symbiot clones,and the water loss rate of the removal of the symbiot clones N5-p5 was significantly higher than that of the other treatments under severe stress;The absolute water content of the removal of the symbiot clone NZYB34 of S.avenae clone ZYB34 was significantly higher under control conditions than in the moderate and severe stress treatment groups;All three single-infested of symbiot clonal lines of S.avenae showed significantly lower water loss than the control at moderate stress.Aphids under certain water stress will increase their resistance to drought by reducing the rate of water loss,and specific Rickettsia strains are able to influence the ability of aphids to resist drought stress by affecting their rate of water loss.3.Based on EPG recordings of the feeding behavior of the secondary symbiotic bacteria Rickettsia on S.avenae under different water deficit stresses,it was found that the number of stabbing（C-wave and Pd-wave）and the retraction and puncture of the oral needle were reduced in the non-phloem feeding of the symbiot-infected clone ZYB34 under severe water stress compared with the control treatment,and the mechanical puncture time of the aphid’s oral needle in the extracellular intercellular membrane or within the cell wall was blocked was also significantly reduced.The duration of water-soluble salivary secretion waves of the symbiot-infected clone ZYB34 was found to be longer with increasing water stress,and the duration of active sap-sucking in the bast was also significantly increased.Both S.avenae aphid clones Ric4 and 5-p5 reduced the number of stings（number of C waves,number of Pd waves）and duration of stings in the non-turgid parts,and increased the duration of sap sucking in the bast and feeding in the xylem during heavy water stress compared to the corresponding symbiot-removed clones4.The copy number of Rickettsia,a secondary symbiont of the 1^stinstar and adult S.avenae,was determined by absolute quantitative PCR under different water stresses in three symbiot-infected clone lines.The results showed that the copy numbers of Rickettsia in the1^stinstar nymphs and adults of the three symbiot-infected clones was significantly higher under severe stress than under moderate stress,and the copy number of secondary symbiotic bacteria of ZYB34 and Ric4 clones under severe stress was significantly higher the control treatment.This suggests that the regulation of Rickettsia copy number may play a role in the response of this insect to water deficit stress.The copy number of the secondary symbiotic Rickettsia also varied among the different Rickettsia-infected clones,probably because different genotypic hosts have different interactions with the secondary symbionts.In conclusion,the infestation of some secondary symbiotic bacteria of Rickettsia allowed the S.avenae to shorten its developmental time,increase adult aphid weight and aphid fecundity,and reduce water loss in the face of certain water stresses.Further studies revealed that the secondary symbiotic bacteria Rickettsia might increase the adaptive capacity of the host aphid to water deficit stress by enhancing the sucking time of the aphid.Meanwhile,the copy number of the symbiotic bacteria was higher in all three clonal lines at severe stress than at moderate stress.The changes in Rickettsia colony density may be related to the adaptation of S.avenae to water stress,which may provide new ideas for understanding the population evolution of the aphid in different arid regions of northwestern China.