| China is the largest broiler market and production country in the world,antimicrobials are widely used in broiler production.Amoxicillin,chlortetracycline and florfenicol are widely used in broiler production due to their good antibacterial effect and less side effects.Therefore,to explore the effects of amoxicillin,chlortetracycline and florfenicol on gut resistome and microbiome in broilers is helpful to guide the rational use of antimicrobials in broiler production and reduce the generation and spread of antimicrobial resistance.Based on this background,this research used metagenomic approach to investigate the effects of infeed amoxicillin,chlortetracycline and florfenicol on chicken gut resistome and microbiome.Thirty 15 d old commercial broilers were divided into five groups: blank control group,amoxicillin group,chlortetracycline group,florfenicol group and mixed administration group.Each group had three replicates,two broilers per replicate.The administrated concentrations of amoxicillin group,chlortetracycline group and florfenicol group were 0.06 g/L,0.1 g/L,0.06 g/L,respectively.The order of administration in the mixed group was amoxicillin,chlortetracycline and florfenicol.Broilers were fed for 7 days before the treatment.The fecal samples were collected at different time points before and after the treatment.The collection time points were T0(before administration),T5(after the first administration),T10(after the second administration)and T15(after the third administration).Fecal metagenomic were extracted to high-throughput sequencing and the sequencing data were spliced and assembled.The antimicrobial resistance genes and species annotation were obtained after gene sequence alignment,then statistically analyzed the data.The results showed that the most abundant antimicrobial resistance genes in broiler intestine were multidrug resistance gene(40.4%),aminoglycoside resistance gene(8.4%),polymyxin resistance gene(7.4%),aminocoumarin resistance gene(7.4%)and tetracycline resistance gene(6.9%).The intestinal tract of the broilers in control group also contained high abundance of antimicrobial resistance genes.The abundance of the amoxicillin,chlortetracycline and mixed group were significantly higher than control group.Multidrug resistance genes,nitrofurantoin resistance genes,quinolones resistance genes and sulfanilamide resistance genes were significantly correlated with the abundance of total resistance genes in the antimicrobial-treated groups.The tetracycline resistance gene tet A was the most abundant ARG subtype in the florfenicol,chlortetracycline,mixed group on T15,while sulfonamide resistance gene sul2 was the predominant gene in amoxicillin group.the mcr-1 gene significantly increased on T15 compared with T0 in mixed group.Throughout our experiment,the genera harboring most of the ARGs detected were Escherichia(84%),followed by Klebsiella(5.1%),Bacteroides(3.9%),Shigella(2.9%),and Clostridium(1.7%).Escherichia was the host of most resistant genes.The ARG-carrying host significantly changed on T10.Escherichia and Bacteroides were the two most significantly changed ARG-carrying host.The results of this experiment are helpful to provide scientific basis for the rational use of antimicrobial in production and to deal with the problem of antimicrobial resistance.The four predominant taxonomic phyla in fecal microbiota of broiler chickens were Proteobacteria(69.8%),Bacteroidetes(15.3%),Firmicutes(10.9%).Escherichia(67.7%)and Bacteroides(15.3%)were the two predominant genera in the broilers intestinal.The predominant genera in broiler intestine significantly changed after the second time of antimicrobial administration,Escherichia significantly increased in all groups,while Bacteroides significantly decreased in the amoxicillin,chlortetracycline and mixed group.Klebsiella significantly decreased in the florfenicol group,while significantly increased in the amoxicillin and chlortetracycline group.The significant increase of Escherichia is the reason of the increase of Proteobacteria,while the significant decrease of Bacteroides is the reason of the decrease of Bacteroidetes.38 characteristic taxa were identified by Le FSe analysis.Proteobacteria and Bacteroidetes,Escherichia and Bacteroides are the most characteristic phylum and genus in this study,respectively.The abundance of antimicrobial resistance gene in amoxicillin,chlortetracycline and mixed group were significantly higher than that in the control group,antimicrobial administration may increase the abundance of antimicrobial resistance genes in broiler intestine.The abundance of multidrug resistance genes was the highest in the gut resistome,and was significantly related to the abundance of total antimicrobial resistance genes.Not all antimicrobial resistance genes have significant changes under the effects induced by antimicrobial.The abundance of mcr-1 significantly increased in the mixed group.This study investigated the response characteristics of chicken gut resistome and microbiome under the pressure of in-feed antimicrobials,providing theoretical basis for rational use of antimicrobial s and the reduction of antimicrobial resistance in animal husbandry. |
