| As a new kind of green and environmental protection feed additive,microecologics have been widely used in animal husbandry.At present,lactic acid bacteria,bacillus,yeast,photosynthetic bacteria and so on are the most widely studied microecological additive.Among them,Bacillus has a history of more than 100 years since its discovery,and a large number of scientific research data have confirmed the use effect of Bacillus.Among Bacillus,Bacillus Megatherium has become a research hotspot due to its unique characteristics such as strong resistance to stress,high temperature resistance and easy storage.Bacillus Megatherium,as a new-type microecological additive,is a gram-positive bacterium that can produce spores.It plays a very important role in improving animal production performance and product quality,increasing the output of livestock products,reducing feed costs,etc.However,there are few studies on the nitrogen metabolism mechanism of BM1259 in young ruminants.This experiment aims to explore the nitrogen metabolism mechanism of BM1259 and its effects on growth performance,rumen fermentation characteristics and microflora changes of dairy bull calves through genomics and metabolomics methods.The specific contents are as follows:Experiment 1.Using Nanopore sequencing technique,this study analyzed the genome-wide genetic map of BM1259 and mined relevant functional gene and pathway annotation information.Results showed,the full length of B.megaterium 1259 genome is 5,043,095 bp,including 38.25%GC base,5092 encoding genes,40 rRNAs,120 tRNAs and 3 circular plasmid.No CRISPR sequence and gene island were detected in BM1259 genome.No pathogenic genes were found.Moreover,12 encoding genes related to nitrate metabolism were screened out and participates in an important pathway-nitrate assimilation,which is probably related to animal nitrogen metabolism.Experiment 2.This study performed a comparative analysis of BM1259 metabolite under different conditions,such as culture time(6 h?12 h,24 h,48 h)and nitrogen concentrations(0.1 g/L,0.5 g/L,1.0 g/L,2.0 g/L).Results showed,135 differential metabolites were identified and mainly involved in purine metabolism,arginine,proline metabolism and nitrogen metabolism.Among these,nitrogen metabolism acts as a key pathway in the process of BM1259 growth and metabolism,and some metabolites such as L-glutamic acid,AMP,L-asparagine showed a significant change.Experiment 3.Through the experiment that 12 g/head ·d of BM 1259 was added to the diet,this study analyzed the effect of BM1259 on growth performance,rumen fermentation,blood biochemical index in Holstein bull calves,in order to evaluate the safety of BM1259.Results showed,1)The ADG and DMI of experimental groups were significantly higher than those of control group(P<0.01),and increased by 12.5%and 8.79%,respectively,during 4-8 weeks after the addition of 12 g/head ·d BM1259;From 0 to 8 weeks,ADG(P<0.05)and DMI(P<0.05)in experimental group were significantly higher than those in control group,and increased by 14.9%and 6.04%,respectively.2)At the end of 4th week,the apparent digestibility of CP and NDF in experimental groups was significantly higher than that in control group(P<0.05),and increased by 5.97%and 6.70%,respectively,after adding 12 g/head·d BM1259.At the end of 8th week,the apparent digestibility of CP and NDF was significantly higher than that of the control group(P<0.01),and increased by 5.88%and 10.26%,respectively.3)At the end of 8th week,rumen fluid pH(P<0.05),MCP(P<0.05)and acetate(P<0.05)in experimental groups were significantly higher than those in control group,and increased by 9.03%,19.68%and 12.74%,respectively,after adding 12 g/head·d BM1259.At the end of the 4th week and the 8th week,NH3-N concentration in experimental group was significantly lower than that in the control group,with a decrease of 21.81%and 16.40%,respectively.4)At the end of 4th week(P<0.05)and 8th week(P<0.05),the glutamate content of rumen fluid of Holstein bull calves in experimental groups was significantly higher than that in control group,increased by 13.21%and 14.32%,respectively;At the end of 4th week,the contents of glutamate in serum(P<0.05),urine(P<0.05)and feces(P<0.05)of Holstein bull calves in experimental groups were significantly lower than those in control group,and decreased by 25.76%,33.87%and 9.23%,respectively.At the end of 8th week,the contents of glutamate in serum,urine and feces of Holstein bull calves in experimental groups were significantly lower than those in control group(P<0.01),and decreased by 26.69%,27.94%and 11.11%,respectively.5)Adding 12 g/head·d of BM1259,the urine ammonia-nitrogen content of test group was extremely significantly lower than that of control group at the end of 4th and 8th week(P<0.01),decreased by 54.60%and 40.31%,respectively.6)Adding 12 g/head·d of BM1259 had no significant effect on the level of blood biochemical indexes of Holstein bull calves.Experiment 4.Using 16S rRNA sequencing technique,this study explore the effect of 12 g/head·d BM1259 on rumen microorganisms in Holstein bull calves.Results showed,12 g/head·d BM1259 could increase the abundance and diversity of rumen bacteria,thereinto significantly improved the content of Saccharofermentans and Prevotellaceae(P<0.05).At the phylum level,Bacteroidetes and Firmicutes are the dominant bacterium in rumen of Holstein bull calves.At the Gene level,Prevotellaceae is the dominant bacterium in rumen of Holstein bull calves.In conclusion,the BM1259 genome information is consistent with its digestion and decomposition function.The supplemental level(12 g/head·d)can improve the performance and richness of rumen microflora of Holstein bull calves,and optimize the structure of intestinal microflora dominated by Prevotella,which may be the main mechanism of regulating the production performance and rumen fermentation of Holstein bull calves. |
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