| Duck enteritis virus(DEV)is a member of the alpha-herpesvirus subfamily of the Herpesviridae family.It mainly causes vascular damage,body cavity hemorrhage,intestinal hemorrhage and parenchymal organ regression in ducks,geese and other Anseriformes.The infectious disease characterized by sexual lesions is duck viral enteritis.The disease has a high morbidity and mortality rate,and poses a huge threat and harm to the healthy development of the waterfowl breeding industry.At present,domestic and foreign scholars have carried out in-depth and systematic research on the pathogenic characteristics,epidemic characteristics,immune prevention and control,and detection technology of DEV,and have achieved fruitful results,but its pathogenic mechanism has not yet been fully understood.In the early stage,our laboratory used high-throughput sequencing technology to carry out transcriptome analysis of DEV-infected duck spleen and intestinal tissues,and analyzed the differential genes and related signals of DEV infection on duck spleen,liver,intestine and other tissues from the genetic level.Changes in conduction pathways,but no research and analysis have been carried out at the level of metabolitesTo explore the metabonomic changes in the body tissues of DEV-infected ducks,this study selected 24 clinically healthy green-shell ducks(body weight of 1.5kg ± 20g)and randomly divided them into 2 groups,namely the experimental group(18)and the control group(6),the experimental group ducks were injected intramuscularly with 0.2 m L of DEV virus solution(TCID50: 3.16×10-9/0.1 m L),and the control group was injected with 0.2 m L sterile saline;66 h,90 h,and 114 h after injection Capture and kill at the same time(fasting for 12 hours before killing),collect duck spleen,duodenum and ileum tissue samples,use LC-MS technology for duck body tissue metabolomics analysis,the results are as follows,After DEV infection,it was determined by clinical symptom observation,anatomical pathological examination and PCR nucleic acid detection.The results showed that the experimental group ducks clinically showed symptoms such as rough coat,depression,loss of appetite,and yellow-green feces pulling.Dissection showed vascular damage,body cavity hemorrhage,intestinal hemorrhage and cloaca mucosal hemorrhage.Specific primers are used for PCR amplification,and the corresponding specific DNA fragments(133bp)can be amplified from the liver samples of DEV ducks at 66 h,90 h and 114 h,the detection rate was100%(18/18).The ducks in the control group had no abnormal clinical manifestations and dissected lesions,and the corresponding DNA fragments were not amplified by PCR.These results indicate that this experiment successfully established a duck DEV infection model.2.Metabonomics Analysis of Duck Spleen Infected by DEVObservation shows that the chromatographic peak response intensity and retention time of DEV-infected duck spleen quality control samples basically overlap,and the PCA model graph samples are closely clustered,suggesting that the samples can be used for LC-MS analysis.According to the VIP value of the first principal component combined with the t-test FC value,the metabolites in the HMDB or KEGG database at levels 1 and 2 were analyzed.The results showed that in the positive/negative mode,DEV infected duck liver tissue was infected at 66 h,90 h,and 90 h.At 114 h,the metabolic pathways involved in different metabolites are basically the same;the metabolic pathways related to inflammation and immune response are tryptophan metabolism,tyrosine metabolism,cysteine-methionine metabolism,phenylalanine metabolism,serine-threonine Acid metabolism and histidine metabolism and other amino acid metabolism and unsaturated fatty acid metabolism and other fat metabolism.Among them,the more concentrated metabolic pathways are phenylalanine metabolism,tyrosine metabolism and tryptophan metabolism,with larger enrichment factors The metabolic pathway is vascular smooth muscle contraction and tryptophan metabolism.3.Metabonomics Analysis of Duodenum of Duck Infected by DEVAnalysis shows that in the positive and negative modes,the metabolic pathways involved in the differential metabolites of DEV infected duck duodenal tissue at 66 h,90 h,1.Duck artificially infected with DEV and 114 h are basically the same;amino acid metabolic pathways related to inflammation and immune response Consistent with the spleen of infected ducks,the metabolic pathways with a large number of differential metabolites are amino acid biosynthesis,cysteine and methionine metabolism,tryptophan metabolism,unsaturated fatty acid biosynthesis and purine metabolism,with more enrichment factors Metabolic pathways are Ig A-related intestinal immune network pathways and lysosome pathways.4.The effect of DEV infection on duck ileum metabolomeAnalysis shows that the metabolic pathways involved in the differential metabolites of the ileum tissue of DEV infected ducks and the metabolic pathways related to inflammation and immune response are the same as those in the spleen and duodenum of infected ducks,but the number of differential metabolites obtained is less;After conditional screening,DEV-infected duck ileum tissue has 16 different metabolites at 66 h infection,52 at 90 h infection,and 40 at 114 h infection.Among them,the metabolic pathway with more differential metabolite enrichment is pyrimidine metabolism.,Tyrosine metabolism,phenylalanine metabolism,tyrosine metabolism and tryptophan biosynthesis.The metabolic pathways with more enrichment factors are m TOR signaling pathway,iron death pathway,tryptophan metabolism pathway and caffeine metabolism pathway.5.Analysis of Metabonomics Difference between Duck Spleen and Intestinal Tissue of DEV InfectedA comparative analysis showed there was a certain degree of difference in the number of different metabolites between the spleen and intestinal tissues of ducks after DEV infection.Among them,the spleen tissue had the most significant metabolites,followed by the duodenum tissue,and the ileum tissue was the least;In the infection time,the number of differential metabolites obtained at 90 h of DEV infection was the largest,followed by 114 h of infection,and the least at 66 h of infection;the same differential metabolites included prostaglandin,arachidonic acid,and arachidonic ethanolamine,etc.Metabolites are mainly concentrated in amino acid metabolic pathways,but the metabolic pathways enriched in intestinal tissues with appear lg A-related intestinal immune network pathways,purine metabolic pathways,pyrimidine metabolic pathways,and lysozyme pathways.In summary,the differential metabolites caused by DEV infection in ducks are mainly concentrated in the metabolic pathways of tryptophan metabolism,amino acid biosynthesis,cysteine-methionine metabolism,unsaturated fatty acid biosynthesis and purine metabolism,among which tryptophan The most obvious enrichment of metabolic pathways can be used as the focus of follow-up research;in different tissues of DEV-infected ducks,the same different metabolites are arachidonic acid,prostaglandin and indole,etc.,which can be used to distinguish DEV-infected ducks from normal healthy ducks Potential markers. |
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