Effects Of Feed Intake Levels On Performances,Rumen Fermentation,Microflora And Metabolome In Cows

The quantity of feed intake can reflect the amount of nutrients obtained by animals.Feed intake is the main factor to ensure the performance of animal and influences the feed utilization ratio.It is also an important prerequisite for improving productivity.Although there have been reports on feed intake and residual feed intake under limited feeding conditions,there is less conclusive evidence that the feed intake levels influence the performance of early-lactating dairy cows,rumen fermentation parameters,bacterial microbial flora and metabolites with ad libitum feeding.Of the 40 early-lactating cows,18 dairy cows were divided into three groups:low(L),middle(M)and high(H)feed intake groups(6 cows in each group),which not only assessed the effects on the performance,blood metabolism and feed intake-related hormones in the early-lactating dairy cows,but utilizing 16S rDNA amplicon sequencing and metabolome profiling to evaluate the effects of feed intake levels on apparent digestibility,rumen fermentation parameters,bacterial community and ruminal metabolites.This thesis includes two sections.Section one:Eighteen cows were allocated one of the three feed intake groups:low feed intake(L)group(mean feed intake=44.15 kg/d),medium feed take(H)group(mean feed intake=48.55 kg/d)and high feed intake(H)group(mean feed intake=53.67 kg/d)with 6 cows in each group.The trial lasted for 70 days.The results showed as follows:1)The milk yield,4%standard milk and the content of urea nitrogen from feed intake groups were significantly different(P<0.05).Compared with L group,the milk yield from H group and M group was increased by 4.79 kg and 9.83 kg,respectively(P<0.05),4%FCM yield from H group and M group was increased by 5.94 kg,9.8 kg(P<0.05),respectively and the content of urea nitrogen in the H group was decreased by 10.21%(P<0.05).2)Serum index showed the conent of albumin and triglyceride were increased significantly with the feeding intake level(P<0.05);The conent of free fatty acids,total cholesterol and insulin in the M group and the H group were significantly higher compared with the L group(P<0.05);The content of insulin-like growth factor and neuropeptide were significantly higher in the H group relative to the M group and L group(P<0.05);The content of cholecystokinin was significantly higher in the L group compared with M group and H group(P<0.05).The results suggested that dairy cows with high feed intake not only had high milk yield and the rapid metabolism of nutrients and energy,but also the biochemical indicators and hormones related to feed intake regulation and energy metabolism were more significant,relative to cows with low feed intake,which was provided scientific basis for feed intake regulation.Section two:On the day before the end of the experiment,the rumen fluid samples of 18 cows were collected at 3 h after morning feeding.For one thing,samples were used for the determination of volatile fatty acid(VFA),ammonia nitrogen(NH3-N)and microbial protein(MCP);For another,the 16S rRNA gene V3-V4 region of 18 ruminal fluid samples were sequenced by MiSeq platform of Illumina company,and the metabolites were detected and analyzed by UHPLC-MS(Ultra high performance liquid chromatography-mass spectrometry).On the last day of the experiment,fresh feces of 18 cows were collected and their apparent digestibility coefficients were calculated by ash insoluble in hydrochloric acid(AIA)method.Significant differences were identified among these groups for acid detergent fiber and cellulose digestibility coefficient.The acid detergent fiber and cellulose digestibility coefficients were significantly increased in the L group compared with M group and H group(P<0.05),whereas there was no statistical difference between M group and H group(P>0.05).Notably,significantly lower pH was observed in the L group,relative to M group,H group(P<0.05),but there was no pronounced change between group and H group.Additionally,there were significantly higher propionate and butyrate concentration in the H group compared with L group(P<0.05),while there was no obvious difference among other groups.According to the rumen fluid bacterial flora sequencing results,Bacteroidetes,Firmicutes and Proteobateria were the predominant phylums,with the sum relative abundance above 92%.At the genera level,Prevotella was the main dominant generas,with the sum of relative abundance more than 54%.There was no obvious change in diversity and richness of ruminal bacterial community among feed intake groups(P>0.05).The relative abundance of Firmicutes was significantly higher in the H group compared with M group and L group(P<0.05),whereas there was no significant difference between M group and L group(P>0.05).The ratio of Bacteroidetes/Firmicutes was significantly higher in the H group relative to M group and L group(P<0.05),but there was no significant difference between M group and L group(P>0.05).The relative abundance of Lachnospiraceae_unclassified,Veillonellaceae unclassified and Saccharofermentants was signficantly higher in the H group compared with L group and H group(P<0.05),while there was no remarkable difference among other groups(P>0.05).The relative abundance of Erysipelotrichaceae_unclassified was significantly higher in the H group relative to M group and L group,whereas there was no significant difference between M group and L group(P>0.05).Compared with L group,the relative abundance of Butyrivibrio was significantly higher in the H group,but there was no obvious change among other groups(P>0.05).According to the results of UHPLC-MS metabolite detection system,a total of 9 256 characteristic peaks were detected from 18 ruminal fluid samples.Through comparison and identification with metabolic database,a total of 232 metabolites were quantified and identified.The VIP>1 and T-test(q<0.05)were used as the basis for the identification of differential metabolites in the PLSDA analysis model,and 40 differential metabolites were obtained.Based on KEGG database,five key pathways were found including linoleic acid metabolism pathway,Alpha-Linolenic acid metabolism,Arginine and Proline metabolism,Glutathione metabolism and Valine,Leucine and Isoleucine biosynthesis,which were closely related to energy metabolism and amino acid metabolism.The key compounds in the these pathways are Linoleic acid,Stearic acid,Glutamate,Alpha-linolenic acid,L-methione,L-valine.The relative contents of Linoleic acid,Glutamate,Alpha-linolenic acid,L-methione and L-valine were significantly lower in the H group compared with M group and L group(q<0.05).Besides,the relative content of Glutamate was significantly lower in the M group compared with L group(q<0.05).Whereas,the relative content of Stearic acid was significantly higher in the H group,relative to the L group(q<0.05).In addition,the bacterial community also was closely associated with the metabolome.The our results showed that Butyrivibrio can metabolize linoleic acid and L-methionine,and involve in the metabolic pathway of Linoleic acid and Alpha-Linolenic acid.The correlation analysis revealed that Erysipelotrichaceae_unclassified was negatively associated with 27 metabolites(r>0.45,P<0.05).Saccharofermentants was negatively related with 16 metabolites(r>0.45,P<0.05).Conclusion:The study was assessed the effects of different intake on apparent digestibility,ruminal fermentation,microbial community and metabolites.There was obvious difference between bacterial community and metabolites of dairy cows with different feed intake levels.Moreover,there was a close relationship between ruminal bacterial communities and dairy cows with different feed intake levels.The key metabolic pathways and different metabolic compounds obtained were identified among these groups,which provided a scientific basis for further exploring the regulation mechanism of feed intake in dairy cows.