| The pine wood wilt caused by Bursaphelenchus xylophilus is a worldwide quarantine disease.Since it was introduced into our country in 1982,its suitable areas have been continuously expanded.To a certain degree,the diversity of host plants and vector insects has increased the difficulty about the prevention and control of pine wood nematode.As a key environmental factor,temperature can affect the life activities of the pine wood nematode.In the early stage,our laboratory has carried out some studies on the phenotypes of the adults of B.xylophilus response to the two different temperature conditions of 15? and25?.In this research,using the method of omics sequencing,to explore the molecular level changes of adults of B.xylophilus in response to temperature,try to conduct a preliminary screening analysis of genes related to the development and reproduction of B.xylophilus;for the sex determination gene mog of B.xylophilus,analyze the transcription difference under different worm states and different temperatures,and the RNA interference method was used to analyze its effect on the development and reproduction of the B.xylophilus.The main results are as follows:1.Adults of B.xylophilus domesticated at 15? and 25? have obvious differences in transcription level,the genes related to signal transduction,lipid metabolism and development and reproduction show typically distinguishing characteristics in temperature response.The the differential genes hsp72,cbp-1,unc-68,ctl-2 in the typical signal pathway of the adults of B.xylophilus,showed up-regulated expression at 15?.In the lipid metabolism pathways of the adults of B.xylophilus,the expression level of the related genes pod-2?alh-12?asm-3?fat-2 ? fat-7 ? gpdh-2 ? gpx-5 ? sodh-1 at 15? is significantly higher than the gene expression level at 25?.The genes related to reproductive development of the adults of B.xylophilus,such as the daf gene family and the HOX gene cluster showed upregulated gene expression at 15?,while the dpy gene family and mog gene family are mostly exhibited down-regulated gene expression at 15?.The gene expression level of genes fbf-2,fem-2,mab-3,rpn-10,gld-1 and sel-10 related to the sex determination of the adults of B.xylophilus domesticated at 15? was decreased than that at 25?,while her-1 and tra-3 increased.2.The analysis found that B.xylophilus have five mog genes,namely mog-1,mog-2,mog-3,mog-4,and mog-5,which play a role in spliceosomes,and the corresponding encoded MOG proteins differ significantly in physical and chemical properties,protein structure,and other aspects.The MOG-2 protein has a leucine-rich repeat sequence structure that may be closely related to its function.The transcriptome data showed that under the condition of25°C,the mog genes in the three stages of eggs,larvae and adults all showed the highest expression level in the egg stage and the lowest expression level in the larval stage.3.Synthesized specific fragments dsRNA of two genes,mog-2 and gfp,to perform RNA interference on the second instar larvae of B.xylophilus,and found that exogenous dsRNA can enter the worm body and have a certain impact: it can cause a certain proportion of nematode death,causing abnormal peristaltic behavior,and to a certain extent,it affects the feeding and development of B.xylophilus.4.After RNA interference about the mog-2 gene of B.xylophilus,the female-to-male ratio of B.xylophilus is reduced;there is no obvious influence in the development of the males of B.xylophilus,but the female development of B.xylophilus is obviously abnormal,mainly manifested as abnormal bending of the body,forward extension of ovary,the development deformity of the pylorus structure,and atrophy of the posterior vaginal uterus. |
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