Effects Of Fructooligosaccharides And Astragalus Polysaccharides On Abundance And Diversity Of Antibiotic Resistance Genes In Intestinal Microbes Of Layers

Antibiotic resistance genes(ARGs)is widely distributed in environmental media such as soil,water and air,which is defined as a new environmental pollutant in recent years,and seriously threatens global ecological safety.The traditional livestock and poultry breeding mode highly relies on antibiotics,which makes the intestinal microbes of livestock and poultry select ARGs with high abundance and high diversity.The ARGs enter the natural environment through breeding manure or manure diffusion,which promotes the accumulation of ARGs in the environment.In2020,the Chinese government has banned the use of antibiotics in feed,but in recent years,it was found that "livestock and poultry intestinal organisms still contain high abundance and high diversity of ARGs without antibiotics",which posed a new challenge,and its prevention and control measures are still unclear.At the same time,there are few studies on the effects of more and more feed antibiotic substitutes on the abundance and diversity of intestinal microbes ARGs in livestock and poultry.In this study,the feed nutrition prevention and control strategy of livestock and poultry intestinal microbial drug resistance,using metagenome sequencing and fluorescent quantitative PCR technology,discussed the effect of feed additives Fructooligosaccharide(FOS)and astragalus polysaccharides(APS)on the abundance and diversity of intestinal microbial ARGS in no anti-farming mode.1081-day-old Jingfen No.1 layers were randomly divided into three groups.3 repetitive cages per group,12 feather eggs per cage.The control group was fed a no-antibiotics basal diet.Treatment group 1 and treatment group 2 were fed a no-antibiotics basal diet with 0.5% FOS and 0.1% APS,respectively.Collect fresh fecal samples from14-day-old and 56-day-old layers.The abundance and diversity of ARGs,MGEs and the composition of intestinal microbes were determined by metagenomics sequencing.The host bacteria of ARGs were found by network analysis method.The minimum inhibitory concentration of host bacteria Escherichia coli and probiotics Lactobacillus against common antibiotics was determined by micro broth dilution method.At the same time,the relative abundance of 21 main ARGs in Escherichia coli and Lactobacillus was determined by fluorescence quantitative q PCR.The main results are as follows:(1)The ARGs with the highest relative abundance in the intestinal microbes of layers are aminoglycoside resistance genes,and the MGEs with the highest relative abundance are transposases;there were 23 core ARGs,which mainly belonged toβ-lactam resistance genes and tetracycline resistance genes,and 61 core MGEs,which mainly belonged to plasmid,transposase and Tn916.(2)Adding FOS and APS to the diet reduced the total relative abundance of ARGs and MGEs of intestinal microbes in 14-day-old and 56-day-old layers,and FOS significantly reduced the total relative abundance of ARGs and MGEs of intestinal microbes in 56-day-old layers.(3)The genus with the highest relative abundance of intestinal microbes in layers is Escherichia,followed by Lactobacillus;the species with the highest relative abundance is Escherichia coli,followed by Lactobacillus crispatus and Escherichia unclassified.(4)Adding FOS and APS to the diet increased the relative abundance of Lactobacillus in the intestinal of 56-day-old layers,and significantly reduced the relative abundance of Escherichia,FOS has a better treatment effect than APS;At the species level,adding FOS to the diet significantly increased the relative abundance of Lactobacillus crispatus in 56-day-old layers and significantly reduced the relative abundance of Escherichia coli and Escherichia unclassified.(5)The total relative abundance of ARGs in the intestinal of layers was significantly positively correlated with the relative abundance of Escherichia,but significantly negatively correlated with the relative abundance of Lactobacillus.(6)The main host bacteria of ARGs were Escherichia coli,Escherichia unclassified and Klebsiella pneumoniae.Among them,Escherichia coli was the most abundant species in the intestinal of layers,and was the main host bacteria of ARGs.(7)Compared with Lactobacillus,Escherichia coli isolated from fecal samples of layers has a wider resistance phenotype to clinically used antibiotics,and carries a higher abundance of ARGs,of which the total relative abundance of ARGs of Escherichia coli isolated from layers’ feces at 14-day-old and 56-day-old is 15.1times and 12.3 times of that of Lactobacillus,respectively.In summary,FOS and APS were added to the diet of no-antibiotics breeding mode to regulate the intestinal microbes composition of layers,improve the relative abundance of Lactobacillus carrying low-abundance ARGs,reduce the relative abundance of Escherichia carrying high-abundance ARGs,and then reduce the relative abundance of intestinal microbes ARGs of layers.The study was used to reduce animal and poultry microbial args pollution from the source,reducing environmental resistance level,and mitigated theoretical guidance.