Mechanisms Of Microbial And Metabolome On Fermentation Of Corn Silage Effected By Climate Zones And Inoculations

Whole-crop corn silage is used mostly and widely around the world.It is an important basic guarantee for the development of global animal husbandry,especially the dairy industry(accounting for more than 40%feed of dairy cows).Much researches focused on developing silage additives and evaluating fermentation quality and chemical characteristics of silages,aiming at improving the efficiency of silage utilization and livestock production performance.However,the complex fermentation process and related mechanism are still not clear.Silage fermentation process and quality are mainly affected by the characteristics of the raw materials and epiphytic microorganisms.Climate factor is one of the direct factors that can affect characteristics of raw material.At present,there is no report focused on climatic factors impact the characteristics of raw material and attached microorganisms of whole-crop corn in different regions.Therefore,this study combined microbiome and metabolomics methods to investigate the effects of different climatic zeros(Subtropical monsoon climate zero,Temperate continental climate zero,Plateau alpine climate zero,Temperate monsoon climate zero and Tropical monsoon climate zero)and lactic acid bacteria on the characteristics of raw whole-crop corn,fermentation quality and nutritional composition of corn silage.The present study revealed the microecological process of whole crop corn silage fermentation,and clarified the metabolomics of microbiome and mechanism on fermentation of whole crop corn silage affected by different climatic factors and lactic acid bacteria inoculations.The aim of this study was to provide scientific and technical support for modulating safe and high-quality whole crop corn silage in different regions.The contents and main results of this study mainly include:(1)The effect of climate factors on the characteristics of raw whole crop cornThe effect of different climatic zones on the characteristics of raw whole crop`corn shows that the characteristics of raw whole crop corn were vary greatly in different climate zeros.The dry matter and content of starch in the raw whole crop corn from the plateau alpine climate zone were relatively low,and the content of woter soluble carbohydrates and fiber were relatively high.The number of lactic acid bacteria were higher in Inner Mongolia and Xinjiang of the temperate continental climate zone and Heilongjiang and Shanxi of the temperate monsoon climate zone,reaching 10^6-8 cfu/g fresh weight.The altitude and precipitation during the growth period in the climatic factors had a greater impact on the characteristics of the raw whole crop corn from different climatic regions.(2)Analysis of fermentation quality and nutrient composition of whole crop corn silage from different regionsThe results showed that the p H and ammonia nitrogen content of whole-crop corn silage from Jiangsu,Inner Mongolia,Shanxi and Heilongjiang were low,and the number of lactic acid bacteria and the concentration of lactic acid were high during the fermentation process,which indicated the fermentation quality of above these four regions were better than other regions.The content of woter soluble carbohydrates in samples from most areas decreased with the extension of ensiling time,but other nutrient components changed slightly during the fermentation process.(3)Analysis of metabolite products during fermentation process of whole crop corn silage from different regionsGC-TOF-MS method was used to detect the metabolites at different fermentation time of whole crop corn silage from different regions in China.The results showed that a total of 427 substances were detected,of which 185 substances can be qualitatively analyzed.The main metabolites were amino acids,fatty acids and carbohydrates.The composition of metabolites,dynamics and classical metabolites during the fermentation of whole crop corn silage from different regions were different.The metabolites in Guizhou,Jiangsu and Xinjiang significantly changed in early and later stage of fermentation.The metabolites in other regions were different before and after fermentation,but not significant differences during fermentation process were found.The pathways enriched by differential metabolites of whole crop corn silage from different regions and different time were mainly comprised of metabolism of alanine,aspartic acid and glutamic acid,aminoacyl-t RNA biosynthesis,metabolism of glyoxylate and dicarboxylate,metabolism of glycine,serine and threonine and valine and biosynthesis of leucine and isoleucine.The p H during ensiling process decreased with the increasement of 3,4-dihydroxycinnamic acid,serine,pyruvate and chlorogenic acid relative content,but increased with the decreasement of melezitose and palatinose relative content.(4)Microbial diversity of whole crop corn silage from different regionsThe microbial diversity of the whole-crop corn silage at different fermentation times in different regions in China was detected by SMRT(single-molecule real-time)technology.The results showed that the main epiphytic microorganisms of fresh whole crop corn in different regions were quite different,however,the microorganisms were not distinguished according to the climatic zones.Among the climate factors,the average relative humidity had a prominent effect on the microbial communities of the whole crop corn in different regions during ensiling.The dynamics of bacteria of whole crop corn silage in different regions were quite different.In general,Leuconostoc,Weissella,and Lactococcus were the mainly genus in the early stage of ensiling,while heterogeneous or facultative heterogeneous fermentation lactic acid bacteria such as Lactobacillus brevis and Lactobacillus buchneri were the mainly dominate bacteria in the later stage of silage(after 30 days).In addition,the marker microorganisms of silage which contributed to the fermentation were also different in different regions.However,Lactobacillus plantarum was a common marker microorganism during whole crop corn ensiling.(5)Based on metagenomics to study the microbial functional genes of whole crop corn silage in different regions of ChinaMetagenomics has been used to detected microorganisms of whole crop corn fermented at early and late stages in different regions of China.The results showed that the main metabolic pathways involved in the whole crop corn silage were carbohydrate metabolism,amino acid metabolism,energy metabolism,coenzyme and vitamin metabolism,and nucleic acid metabolism.The genes of silage microorganisms involved in these metabolic pathways were different during the early and late stages of ensiling in different regions.A variety of antibiotic resistance genes existed in silage microorganisms,and they can tolerate antibiotics mainly through antibiotic efflux,antibiotic inactivation and antibiotic targeted protection.Meanwhile,plasmids were the main genetic elements of resistance gene flow,thus the horizontal transfer of resistance genes might be in silage.Among the climate factors,only the average temperature had a positive effect on the resistance genes.The microbial composition was direct positively effect on the resistance genes,therefore,the microbial composition was a crucial predictor of the resistance genes.(6)Effects of lactic acid bacteria inoculants on the bacterial community and metabolome dynamics of whole crop corn silageThe effects of inoculations of Lactobacillus plantarum MTD1 or Lactobacillus buchneri 40788 on microbiome and metabolome of whole crop corn silage have been studied.The results showed that the L.Plantarum or L.buchneri regulated the bacterial communities and metabolic products in different ways,and it was found that the key microorganisms which did not depend on the relative abundance were different in different groups.In addition,many metabolites with biological functions were founded,including antibacterial,antioxidant,central nervous system inhibitory and anti-inflammatory substances.In the middle of fermentation,inoculated with lactic acid bacteria up-regulated the D-alanine metabolism pathway,flavonoid and flavonoid biosynthesis pathway,polycyclic aromatic hydrocarbon degradation pathway and glycosaminoglycan degradation pathway.Correlation analysis between metabolites and microbiome showed that the correlation between silage microorganisms and metabolites was extensive.We can screen functional lactic acid bacteria as silage additives according to the correlation analysis between metabolites and microbiome in the future studies.