Study On Methionine Metabolism Mechanism Of Rumen Bacteria In Goats

In ruminant livestock,when production performance is improved,small intestine amino acid transport,absorption and utilization and microbial protein production are restricted,resulting in an increase in amino acid requirements,showing the restrictive effect of some amino acids.Dietary ratio affects the amino acid composition,directly affects the restriction order of amino acids,which can be the first,second and third restriction amino acids according to the degree of restriction.In corn-soybean meal diet,methionine is the primary limiting amino acid,while the presence of rumen microorganisms leads to the uncertainty of amino acids entering the small intestine.The amino acid metabolism of rumen bacteria mainly includes catabolism(dissimilation)and anabolism(assimilation).Catabolism of rumen bacteria refers to the decomposition of complex organic matters that cannot be used by animals into glucose,organic acids,etc.,for use by animal bodies.There are many ways of catabolism,mainly deamination and decarboxylation.Rumen microbial anabolism refers to the synthesis of microbial proteins from ammonia,peptides and amino acids produced by the degradation of dietary proteins as nitrogen sources to provide essential nutrients and energy sources for animal bodies.The purpose of this study was to investigate the metabolic mechanism of methionine in rumen microorganisms.Experiment 1:The aim of this study was to investigate the use of methionine(Met)by rumen microorganisms and the effect of Met on microbial nitrogen transfer by using isotope labeling.(Method)Total mixed ration consisted of concentration and corn stover(50:50)was used for fermentation substrates,The study was conducted as one-factor block experimental design,and two supplemental levels of Met were added to the fermenter to explore the influence of Met on in vitro fermentation p H,ammonia nitrogen(NH₃-N)content,volatile fatty acid(VFA)content,microbial nitrogen,microbial degradation rate and the abundance of ¹³C in microbial.The results showed that:1)supplemental of Met could significantly increase the p H value of the fermentation liquid(P<0.01);2)supplemental of ¹³C-Met could significantly increase the abundance of ¹³C in microorganisms(P<0.01);3)supplemental of ¹³C-Met had no significant effect on NH₃-N content,VFA content,nitrogen degradation rate,microbial nitrogen and microbial Met content(P>0.1).Therefore,it can be inferred that rumen microorganisms could effectively utilize Met.Met had some influence on fermentation characteristics,but did not affect the flow of microbial nitrogen.Experiment 2:The objective of this study was using a dual-flow continuous culture system to evaluate effects of difference dietary protein sources including soybean meal(SBM),cottonseed meal(CSM)or rapeseed meal(RSM)on nutrient digestibility,rumen fermentation and microbiota.This study involved a replicated 3×3 Latin square design with 3 dietary protein sources as treatment factors with 3 experimental periods.Each period lasted 8 days,which consisted of 5 days for diet adaption and 3 days for sample collection.The results showed that dietary protein sources of either CSM,RSM or SBM had no effect on nutrient disappearance(P>0.1).Inclusion of CSM or RSM significantly reduced(P<0.01)p H value and enhanced(P<0.01)NH₃-N concentration versus SBM.As compared to RSM and CSM,total volatile fatty acids(VFA)contents were greater(P=0.05)in SBM;molar proportion of acetate was lesser(P<0.01)while molar proportion of propionate was greater(P<0.01)in RSM than in SBM,consequently a reduced acetate to propionate ratio(A/P)in RSM.Protozoa enumeration tended(P=0.08)to be lesser in SBM than RSM or CSM.Principal coordinate analysis(PCo A)displayed that the bacterial community was different in RSM or CSM versus in SBM.Bacteroidetes,Firmicutes,and Proteobacteria were detected as the dominant phyla,and the relative abundance of Spirochaetae(P<0.01)and Chlorobi(P<0.05)declined in CSM and RSM groups as compared to SBM.At genus level,Prevotella?1was the dominant genus;as compared to SBM,the relative abundance was greater(P<0.01)in CSM and RSM groups;whereas,Prevotellaceae?Ga6A1 and Christensenellaceae?R7 was lesser(P<0.05)in CSM group,Eubacterium?oxidoreducens and Treponema?2 were lower(P<0.01)in both CSM and RSM groups,as other genera were not different(P>0.10)across treatments.In conclusion,although the microbial community changed with different dietary protein sources,the nutrient disappearances were not affected,suggesting that CSM and RSM could be well used by rumen bacterial like SBM does,and they will be suitable protein sources for ruminant diets.Experiment 3:The purpose of this study was to evaluate the effects of different protein sources on the amino acid composition,the utilization of methionine and methionine metabolism of microorganisms using a dual-flow continuous culture fermentation system.This study involved a repeat 3×3 Latin square design,with 3dietary protein sources as treatment factors with 3 experimental periods.Each period lasted 8 days,which consisted of 5 days for diet adaption and 3 days for sample collection.Additional 0.097%Met was added to each fermenter,on day 6,40%Met was replaced by 20%[1-¹³C]-Met and 20%methyl-²H₃-Met.Different protein sources had no significant influence on the total microbial amino acid content(P>0.1),the bacteria Ser content in the SBM and RSM group was significantly higher than that of CSM group(P=0.01),Gly in RSM group had a significantly higher content than that of SBM and CSM groups(P=0.02),and other bacteria amino acids content was not affected by protein sources(P>0.1);Different protein sources had no significant effect on APE of[1-¹³C]-Met(P>0.1),but the bacteria APE of[methyl-²H₃]-Met in the RSM group was significantly higher than that in the CSM group(P=0.00).After Met converted to SAM,about 80%of SAM converted to decarboxylated SAM,and about 20%entered the homocysteine cycle.The results of metabolomics analysis of microbial hydrolysates showed that different protein sources mainly affected the metabolic pathways such as ABC transporter,protein degradation and absorption,and there were significant differences between SBM,RSM and CSM.Experiment 4:The experiment was conducted to evaluate Met on growth performance,slaughter performance,meat quality,muscle amino acids and fatty acids composition in xiangdong black goat.10 goats(18.73±1.8kg)were randomly divided into two groups using a completely randomized block experiment design,control group fed based diet,treatment group add 1.27%Met,goats were slaughted at the ending of experiment.The results showed that Met supplementation had no significant effect on growth performance and slaughter performance(P>0.1).Dietary Met supplementation had no effect on the visceral index of goats(P>0.1).The p H,color,drip loss,water loss rate,and cooking loss of longissimus dorsi muscle and vastus lateralis muscle were not different(P>0.1),Met will significantly increase the p H value at 24 h of gluteus maximus(P=0.04),and other meat quality index is not affected by Met(P>0.1).Met did not affect the content of free amino acids in blood,but the content of Gly tend to increas(P=0.07).The most aboundant amino acid in muscle were Glu,Asp,Lys and Leu,respectively,Met did not affect total amino acid content in longissimus dorsi muscle(P>0.1),but significantly improve content of Gly(P<0.01)and Pro(P<0.01),as other amino acids were not affected(P>0.05),Met tended to increase Met content in the vastus lateralis muscle(P=0.06),but other amino acids is not affected by Met(P>0.1),Met(P=0.10)and Glu(P=0.05)content has a tendency to increase in vastus lateralis muscle as Met added,but other amino acids were not affected by Met(P>0.1).Met had no effect on taste amino acids,essential amino acids,non-essential amino acids,acidic amino acids,basic amino acids,and efficacy amino acids of all muscles(P>0.1).After Met was added to the diet,the relative content of eicosenoic acid was significantly reduced(P=0.04),and the relative content of anti-octadecenoic acid was tended to decreased(P=0.07).The relative content of other fatty acids was not affected by Met(P>0.1),and the relative content of saturated fatty acids and unsaturated fatty acids was not affected by Met(P>0.1).Met supplementation did not affect the NH₃-N concentration and VFA content in the rumen.In summary,Met supplementation had no effect on growth performance,slaughter performance and viscera index of goats.Met may have the potential function to improve meat quality.In general,Met has little effect on the amino acids composition of muscle,but Met may have the potential function of improving Met content in muscle,But Met might be detrimental to the synthesis of unsaturated fatty acids in muscle;Met had no effects on rumen fermentation in goats.