Function Of Peptidoglycan Recognition Proteins From Glyphodes Pyloalis In Response To Parasitic Wasp And Pesticide Stress

The insecticide resistance caused by chemical pesticides is becoming more and more serious,and it is urgent to develop new control methods.The use of natural enemies and biological pesticides or the development of new drug targets will contribute to the green control of Glyphodes pyloalis Walker.Aulacocentrum confusum is the dominant parasitic wasp at larval stage.Both chlorfenapyr and phoxim can be used to control lepidopteran pests in mulberry orchard.Bacillus thuringiensis and Empedobacter brevis are widely used in biological control as biological pesticides.Peptidoglycan recognition proteins are a family of pattern recognition receptors that can recognize peptidoglycan.They activate the immune response signaling pathway by recognizing pathogen-associated molecular patterns,and then initiate the immune response to maintain insect physiological homeostasis.In this study,the peptidoglycan recognition protein in the immune system of the G.pyloalis was used as the main experimental target to identify parasitic wasps,chemical pesticides or biological pesticides processing G.pyloalis peptidoglycan recognition protein expression patterns,analyze the immune response involved in peptidoglycan recognition proteins in the larvae of G.pyloalis when parasitized by A.confusum,investigate the response mode of peptidoglycan recognition protein of G.pyloalis to chlorfenapyr and phoxim treatment,explore the role of peptidoglycan recognition proteins in the destruction of midgut by Empedobacter brevi.The main conclusions are as follows:1.Hemolymph melanization was enhanced by peptidoglycan recognition protein induced by parasitic wasps.（1）Three short peptidoglycan recognition proteins were identified and named Gp PGRP-S1,Gp PGRP-S2 and Gp PGRP-S3,respectively,from the transcriptome of G.pyloalis parasitized by A.confusum.Bioinformatic analysis indicated that Gp PGRP-S1and Gp PGRP-S2 belonged to the sensory group,Gp PGRP-S3 has amidase activity.（2）The expression levels of Gp PGRP-S1 and Gp PGRP-S2 were the highest in female adults,and the expression levels of Gp PGRP-S3 were the highest in male adults.The expression of Gp PGRP-S1 in hemolymph was significantly higher than that in non-parasitized group after being parasitized for 3 and 5 days.The expression levels of Gp PGRP-S2 and Gp PGRP-S3 in hemolymph after 3 d and 5 d of parasitism were significantly higher than those in non-parasitic group.（3）The recombinant Gp PGRP-S2 protein and the parasitized G.pyloalis hemolymph significantly enhanced the phenoloxidase activity in healthy hemolymph in vitro,respectively and the degree of enzyme activity was significantly higher when the two were co-incubated than when they were treated alone.2.By up-regulating the expression of peptidoglycan recognition protein in midgut,fat body and other tissues,it responded to the treatment of chlorfenapyr and phoxim,and correlated with detoxification enzymes.（1）The expression levels of the three Gp PGRPs were all highly expressed in the midgut after 48 h treatment with chlorfenapyr.The expression levels of Gp PGRP-S2 and Gp PGRP-S3 were up-regulated in midgut and hemolymph after 48h of phoxim treatment.（2）The survival rate under the condition of interference with Gp PGRP-S2 was significantly lower than that of the group that under the condition without interference with Gp PGRP-S2.（3）The expression of detoxification enzyme gene Gp CXE1was significantly up-regulated without RNA interference and after interference with Gp PGRP-S3,while it was significantly down-regulated after interference with Gp PGRP-S2.In the treatment of phoxim,the expression of Gp CXE1 was significantly up-regulated without RNA interference and interference with Gp PGRP-S2,while it was significantly down-regulated after interference with Gp PGRP-S3.The results indicating that synergistic response of peptidoglycan recognition proteins and detoxification enzymes induced by chemical pesticides.3.G.pyloalis peptidoglycan recognition protein in midgut defense against bacterial infection.（1）The LC₅₀ value of E.brevis suspension with 10 billion conidia was determined to be 4.72×10⁷ spores/m L for the 4th instar larvae.（2）The expression levels of three Gp PGRPs were significantly increased in the 4^th instar larvae treated with E.brevis for 48 h,and the midgut and perimetron membrane（PM）were seriously damaged.（3）The damage degree of midgut tissue and PM of G.pyloalis larvae after 48 h infection with E.brevis with Gp PGRP-S2 was significantly higher than that without Gp PGRP-S2 infection.The study on the immune function of peptidoglycan recognition protein is helpful to understand the immune responses of G.pyloalis deeply,and then adopt more targeted methods for biological control of G.pyloalis,which provides theoretical basis for integrated pest control.