| Due to the special geographical environment and climate conditions of the Hulunbuir region,forage resources are scarce in winter.The intake of nutrients by Hulunbuir sheep is severely insufficient in winter,resulting in hindered growth and development and low production efficiency.In this experiment,an animal model of energy imbalance induced by severe feed restriction during the non-pregnant period of Hulunbuir ewes was used.The effects of supplementary feeding of Leymus chinensis hay on rumen fermentation,microbial community,and liver transcription and metabolic spectrum of Hulunbuir ewes were studied.The main research results are as follows:1.The effect of supplementary feeding of Leymus chinensis hay on the growth performance,organ development and blood biochemical indicators of Hulunbuir ewesThis chapter aims to study the effect of supplementary feeding of Leymus chinensis hay on the growth performance,organ development and blood biochemical indicators of Hulunbuir ewes in winter.Forty healthy Hulunbuir ewes were selected for the experiment and randomly divided into a feed restriction group(Feed restriction,FR,n=20)and a supplementary feeding group(Ad libitum,AL,n=20).The AL group was fed according to the average daily feed intake(1.5kg)during the pre-feeding period,while the FR group’s feed intake was 40% of the supplementary feeding group(0.6kg).On the 1,7,14,21,28,42,56,70,84 and 87 days after the start of the formal experiment,their weight changes were recorded before morning feeding and blood was collected from the jugular vein to determine blood biochemical indicators such as glucose and lipid metabolism and protein synthesis in plasma.On the 87 day of the trial period,40 animals were slaughtered and their carcass weight and organ weight were recorded.The results showed that compared with the FR group,supplementary feeding significantly reduced the weight change rate of Hulunbuir ewes and increased live weight,carcass weight,liver weight,lung weight and kidney weight(P<0.05),but had no significant effect on slaughter rate and eye muscle area(P>0.05).Supplementary feeding significantly increased perirenal fat,mesenteric fat,omental fat and tail fat weight and index(P<0.05),and changed the content of total cholesterol,triglycerides,low-density lipoprotein and high-density lipoprotein in blood(P<0.05).These results indicate that insufficient intake of nutrients by sheep in winter can lead to rapid depletion of body fat reserves and cause lipid metabolism disorders.Supplementary feeding with Leymus chinensis hay can reduce weight loss and maintain stable body weight while reducing high-density lipoprotein cholesterol levels in blood due to lipid mobilization.2.The effect of supplementary feeding of Leymus chinensis hay on rumen fermentation and microbial community of Hulunbuir ewesThis chapter aims to explore the effect of supplementary feeding of Leymus chinensis hay on rumen fermentation and the composition and function of the microbial community of Hulunbuir ewes in winter.The experimental design is the same as Experiment 1.After the experiment,samples were collected by slaughtering and collecting rumen fluid and rumen tissue.Part of the rumen fluid was used to determine rumen fermentation parameters,and part was used to extract microbial DNA to study changes in microbial community composition and function through high-throughput sequencing.Rumen tissue was used to observe the morphology of the rumen epithelium,measure the height and width of the rumen papillae,and extract RNA samples to determine the relative m RNA expression levels of tight junction proteins and VFAs absorption-related genes in the rumen by RT-PCR fluorescence quantification.The results showed that compared with the FR group,supplementary feeding significantly increased the concentration of butyrate,isobutyrate,valerate and isovalerate,increased the butyrate ratio,but decreased the proportion of acetate(P<0.05),with no significant effect on rumen fluid p H,acetate and propionate content,total volatile fatty acids,propionate ratio and acetate-propionate ratio(P>0.05).Histological observation showed that supplementary feeding significantly increased the length and width of rumen papillae.Compared with the FR group,supplementary feeding significantly up-regulated the relative m RNA expression levels of tight junction proteins Claudin-1,Claudin-4 and ZO-1(P<0.05).At the same time,it also increased the relative m RNA expression levels of VFA absorption-related genes BDH1,HMGCL,MCT1,NHE1,NHE2 and PAT1(P<0.05).PCo A and NMDS analysis results showed that supplementary feeding with Leymus chinensis hay can change the composition of rumen microorganisms,but there is no difference in α diversity.A total of 23 phyla and 256 genera were detected.The genera with the highest abundance at the genus level were Saccharofermentans,Anaerovorax and Sphaerochaeta,all of which had significantly increased relative abundance in the AL group.LEf Se analysis results showed that g＿Saccharofermentans and s＿Prevotella＿ruminicola played important roles in the AL group,while f＿Christensenellaceae,g＿Prevotella and s＿Bacteroidales＿bacterium＿RM8played important roles in the FR group.Supplementary feeding significantly increased vitamin B6 metabolism,various types of n-glycan biosynthesis,AMPK signaling pathway,PI3K-Akt signaling pathway,arachidonic acid metabolism and protein digestion absorption etc.,indicating that supplementary feeding caused overall functional changes in rumen microorganisms,strengthening carbohydrate metabolism and protein metabolism to alleviate negative effects brought by insufficient nutrient intake in winter.3.The effect of supplementary feeding of Leymus chinensis hay on liver metabolism of Hulunbuir ewesThis chapter aims to explore the effect of supplementary feeding of Leymus chinensis hay on liver metabolism of Hulunbuir ewes in winter.The experimental design is the same as Experiment 1.After the experiment,samples were collected by slaughtering and collecting liver samples to explore the effect and mechanism of supplementary feeding of hay on liver metabolism in winter based on metabolomics and transcriptomics analysis.A total of 173 differential metabolites(DMs)were screened out.Compared with the FR group,there were92 up-regulated and 81 down-regulated DMs in the AL group,mainly belonging to amino acids and their metabolites,benzene and its derivatives,fatty acyls,organic acids and their derivatives,heterocyclic compounds,nucleotides and their metabolites etc.The KEGG metabolic pathways enriched by DMs include nucleotide metabolism,purine metabolism,ABC transporters,biosynthesis of cofactors,pyrimidine metabolism,cysteine and methionine metabolism and glutathione metabolism etc.A total of 5754 differentially expressed genes(DEGs)were screened out.Compared with the FR group,there were 3347 up-regulated and2407 down-regulated DEGs in the AL group.The KEGG signaling pathways enriched by DEGs include insulin signaling pathway,sphingolipid signaling pathway,endocytosis,NOD-like receptor signaling pathway,tight junctions,PI3K-Akt signaling pathway,MAPK signaling pathway and Hippo signaling pathway etc.The results showed that supplementary feeding of hay in winter significantly changed the transcriptional spectrum and metabolic spectrum of sheep liver by regulating cell proliferation and division through IGF-1,PI3K-Akt and MAPK signaling pathways to improve liver amino acid metabolism and lipid metabolism to maintain metabolic stability in the body to cope with nutrient deficiency.In summary,supplementary feeding of Leymus chinensis hay in winter can reduce weight loss of Hulunbuir sheep,change rumen fermentation pattern and microbial community composition to increase VFAs production,transport and metabolism ability as well as rumen papillae development by rumen epithelium to improve liver amino acid metabolism and lipid metabolism to maintain metabolic stability of body to cope with nutrient deficiency providing important support for healthy and efficient breeding of Hulunbuir sheep. |
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