Spatial And Temporal Distribution Of The Intracellular Symbiotic Intracellular Bacteria Wolbachia In Trichogramma Pretiosum And Trichogramma Dendrolimi

Wolbachia is the most widely distributed intracellular symbiotic bacterium in nature,infecting arthropods and nematodes.Wolbachia regulates a variety of reproductive phenotypes in the host,these include Thelytoky,Cytoplasmic Incompatibility(CI),Feminization(F),and male-killing(MK).In Trichogramma,Wolbachia leads to the failure of the first mitosis of the unfertilized eggs of Trichogramma,doubling the number of chromosomes,and inducing parthenogenesis in females.It has a good application prospect.The stability of the parthenogenetic(PI)phenotype of Trichogramma induced by Wolbachia depends on.The distribution pattern and titer of Wolbachia in host.It was found that the titer and distribution of Wolbachia in the host could be affected by factors such as strain,host species and genetic background,host development stage,antibiotics and temperature.In this study,female parthenogenetic Trichogramma pretiosum(TP)and Trichogramma dendrolimi(TD)were selected.Through fluorescence in situ hybridization and fluorescence quantitative determination of the effects of egg laying intensity and development period on Wolbachia titer and distribution,this study will provide a reference for revealing the interaction between Wolbachia and host.The results are as follows:1.To investigate the effect of oviposition intensity on Wolbachia distribution and titer in Trichogma wasp,Wolbachia distribution and titer,fecundity,male ratio of offspring,individual size of offspring and emergence rate of TD and TP females with different oviposition intensity(nee,AH and NAH)were measured.The results showed that the male ratio of AH was significantly higher than that of NAH and NE.Wolbachia titer in AH was significantly lower than NAH and NE.Wolbachia titer of AH abdomen was significantly lower than that of NAH and NE.In TP strain,there was no significant difference in Wolbachia titer between AH,NAH and NE chest.Wolbachia was detected only in the chest and head of AH.NAH Wolbachia was detected only in abdomen.The fecundity and progeny of AH were significantly lower than NAH and NE,and NAH was lower than NE.The emergence rate of AH was lower than NAH and NE,and NAH was lower than NE.In TD strain,the Wolbachia titer in AH chest was higher than that in NAH,but lower than that in NE.In NE,Wolbachia mainly distributed in the abdomen and chest,and a few in the head.The fecundity and progeny of AH were significantly lower than NAH and NE,and there was no significant difference between NAH and NE.The emergence rate of AH was lower than that of NAH and NE,and there was no significant difference between NAH and NE.TD was significantly larger than TP.The continuous oviposition behavior of TD and TP overconsumed Wolbachia in the ovary leading to the decrease of Wolbachia titer in the ovary and the decrease of Wolbachia-induced parthenogenesis intensity.2.Fluorescence in situ hybridization and fluorescence quantification were used to detect the distribution and titer of Wolbachia in TD and TP females at different developmental stages.The distribution pattern of Wolbachia at different stages of TD and TP development was as follows: Wolbachia accumulates at the back end of early embryo,and gradually migrates to the middle and front end in late embryo.In larval stage,Wolbachia mainly distributed in the front and back ends.Wolbachia migrated from the front end of pre-pupal stage to the middle end,and most of Wolbachia concentrated in the middle and back end.From pupal stage to from pupa to adult,the distribution pattern did not change significantly,and distributed in the head,chest and abdomen.The titer of Wolbachia in TD and TP immature stages(embryonic to pre-pupal stage)increased significantly with the prolongation of development time,and the proliferation rate of Wolbachia was the high in the whole development stage.Wolbachia titer in TD pre-pupal stage was significantly lower than that in pupal stage and adult stage,and there was no significant difference between pupal stage and adult stage.There was no significant difference in Wolbachia titer between TP prepupal and pupal stage and adult stage.3.In order to clarify the changes of Wolbachia in early TD and TP embryos,the fluorescence in situ hybridization was used to locate Wolbachia in early TD and TP embryos at 30 min,50 min,60 min,70 min,90 min and 120 min,and the titer and density of Wolbachia were determined.DAPI staining was performed on the nuclei of embryos to record the mitotic frequency and the number of nuclei in early embryos.The results showed that Wolbachia was mainly distributed in the rear end of TD and TP embryos,and then migrated to the middle and front end.Wolbachia density,mitosis frequency,nuclear number and embryo size increased with the development time.