Effects Of Mannan Oligosaccharide In Milk Replacer On Serum And Urine Metabolome And Gut Immunity Of Hu Lamb

Supplementation of mannan oligosaccharides(MOS)into diets fed to young ruminants can reduce the diarrhea rate and promote growth,while few reports about MOS on lambs.In addition,it was hard to explain the specific metabolic mechanism of MOS on animals,because most of the research focused on nutrition and health in young ruminants was limited in traditional methods.So,the purposes of this study were to investigate the effects of MOS in milk replacer on serum and urine metabolome and intestinal immune mechanism of Hu lambs by metabolomics,serum biochemistry,and molecular biology techniques,with a view to provide theoretical basis for MOS nutrition research and product development in young ruminants.The main results of this study were as following.(1)The study focused on the effects of MOS in milk replacer on growth performance,rate of diarrhea,serum and urine metabolome of Hu lambs.Fifteen pairs of healthy newborn siblings Hu lambs(3.51±0.55kg)were employed in this experiment.The newborn lambs were breastfed with the ewe.At 4 days of age,all lambs were separated from the ewe and began to receive milk replacer training with feeding bottle.According to the 7-day-old weight of lambs,each pair of lambs were randomly divided into two groups by the principle of homogeneity,which were the control group(4.09 ± 0.66kg)and the MOS group(MOS,4.07 ± 0.61kg).There were fifteen lambs in each group,and each lamb was a repeat.The lambs in control group were fed with normal milk replacer,and the lambs in MOS group were fed with milk replacer that contained 0.2% MOS.Milk replacer were fed at 2% of the lamb's weight.Weighed the lambs body weigh at the age of 7,14,21 and 28 days morning before feeding,and calculated the average daily weight gain of lamb in different stages and the whole period.Diarrhea of lambs was observed and the diarrhea rate was recorded.At 28 days of age,the serum and urine sample of lambs in control group and MOS group were collected for the determination of metabolites by LC-MS.The results showed that supplementation of MOS in milk replacer can enhance the average daily weight gain of lamb in whole period(P <0.05),and reduce the rate of lamb diarrhea(P <0.05),and MOS in milk replacer can dramatically improve the concentration of 20 metabolites in serum samples including lysophosphatidylcholine,lysophosphatidylethanolamine,glyceric acid,4-Hydroxyproline,betaine,and N-acetyl-L-alanine,etc.(P <0.05).Simultaneously,the concentrations of 11 serum metabolites such as indole lactic acid,allantoin and urea,etc.were significantly reduced(P <0.05).These metabolites in serum were mainly involved in lamb's glycerolipid metabolism,glyoxylic acid and dicarboxylic acid metabolism,arginine and proline metabolism,glycine,serine and threonine metabolism,glycerophospholipid metabolism and purine metabolism pathway.In urine,MOS increased the concentration of xanthine significantly,and decreased the concentration of L-isoleucine and monoethyl malonic acid significantly(P <0.05).Xanthine,Lisoleucine and monoethyl malonic acid were related to degradation of valine,leucine and isoleucine,biosynthesis of valine,leucine and isoleucine,purine metabolism and aminoacyl-tRNA biosynthesis.Moreover,there was no significant correlation among growth performance,diarrhea rate and serum and urine metabolites of Hu lambs(P >0.05).In summary,the supplementation of MOS in milk replacer affected the metabolites in serum and urine of Hu lamb,and regulated the protein and energy metabolism pathways in lamb body,which were related to the immunity,anti-oxidation,anti-stress functions of animals.(2)The research was carried out to study the effects of MOS in milk replacer on biochemical indices and the relationship between serum biochemical indexes and differential metabolites.At 28 days of age,the serum sample of lambs were collected.The automatic veterinary biochemical analyzer was used to detect the serum biochemical indexes of lambs.The results displayed that MOS increased the content of total cholesterol,lactate dehydrogenase,alkaline phosphatase,direct bilirubin and albumin in serum significantly(P <0.05),and reduced levels of alanine aminotransferase and urea significantly(P <0.05).However,MOS did not make significant differences on other biochemical indexes(P >0.05).The concentration of allantoin,indole-lactic acid,n-acetyl-glutamic acid,o-hexanylcarnitine and urea in serum were positively correlated with aspartate aminotransferase,lactate dehydrogenase and l-hydroxybutyrate dehydrogenase(P <0.05),furthermore,the concentration of L-isoleucine was also positively correlated with creatinine,uric acid and urea in serum(P <0.05).In conclusion,the supplementation of MOS in milk replacer affected the serum biochemical indexes of Hu lambs,and the changes of serum biochemical indexes and the correlation analysis of metabolites could also illustrate the benefits from MOS well.(3)The objective of this study was to investigate the gut short-chain fatty acid(SCFA)of lambs in two groups,and analyzed the relationship between gut SCFA and serum and urine metabolites.Eight lambs in each group were selected randomly for slaughter,and jejunal and colonic content were collected for the investigation of concentration of SCFA.The results showed that concentration of SCFA in jejunal was not affected by MOS(P >0.05),but supplementation of MOS in milk replacer could increase the concentration of SCFA in colon significantly(P <0.05).In summary,the supplementation of MOS in milk replacer can promote the fermentation of microorganisms in the colon to produce SCFA.(4)The study was set up to analyze the effect of MOS on the expression of TLRs mRNA on intestinal mucosal and correlation with TLRs and metabolites.After slaughtering the lambs,jejunal and colonic mucosa were collected for the investigation of gene expression of TLRs.The result showed that supplementation of MOS in milk replacer also reduced the mRNA expression levels of TLR1,TLR2,TLR4 and TLR6 in jejunum and colonic mucosa(P <0.05),but there was no significant correlation between TLRs mRNA expression and gastrointestinal SCFA,serum and urine metabolites.In conclusion,MOS can regulate the intestinal immune function and maintain the intestinal epithelial cells homeostasis.In summary,the supplementation of MOS into the milk replacer can be good at the metabolism of Hu lamb,improve the growth performance,and enhance the intestinal tract health.