Construction And Application Of The Catalogs Of Microbial Genomes,genes And Antimicrobial Resistance Genes From Pig Gut Microbiome

Pork is the main resource of meat for human consumption.Gut microbiota plays a very important role in maintaining host physiologic,metabolic,neurological and immunological functions,as well as pig production.However,the understanding to pig gut microbiome is still limited,mainly due to the lack of a comprehensive catalog of microbial genes and genomes,which seriously limits the mining and use of metagenomic sequencing data.Meanwhile,antimicrobials have been widely used in pig production for treating diseases,improving health and promoting growth.However,the overuse of antimicrobials has promoted the emergence of antimicrobial resistance(AMR),leading to the change of the composition and diversity of gut microbiota,thereby adversely affected host health.The World Health Organization has indicated the AMR pollution as one of the major threats to global public health.Therefore,it is critical to strengthen global AMR surveillance.Understanding the diversity and distribution of antimicrobial resistance genes(ARGs)in bacteria and their resistance characteristics is fundamental to evaluate the risk of AMR to public health.However,there has still been a lack of large-scale quantitative data of ARGs in domestic pigs and wild boars,and the relationship between ARGs and bacterial species in gut microbiome has not yet been clarified.In this study,we constructed an expanded gene catalog named Pig Integrated Gene Catalog(PIGC)which contained 17,237,052 complete genes,by using 500 metagenomic sequencing data of the swine gut microbiome from a wide range of sample sources spanning various ages,sexes,breeds,gut locations,geographical areas and feeding patterns,and integrating the gene catalog of 287 pig gut metagenomes reported previously.Among these genes,28% were unknown genes.PIGC expanded the comprehension and completeness of microbial gene catalog for the pig gut microbiome.Using metagenomic binning analysis,we reconstructed 6,339 metagenome-assembled genomes(MAGs)of gut microbiota,representing 2,673 species-level genome bins(SGBs),among which 86%(2,309)of SGBs were unknown bacterial species.Using the present gene catalog and MAGs,we further compared the composition and function of gut metagenome between wild boars and commercial Duroc pigs,and identified several bacterial strains showing different abundances between two breeds.Based on the data used for constructing PIGC,we further constructed a catalog of ARGs,containing 1,299 open reading frames(ORFs)recognized as AMR protein-coding genes.These ORFs represented 350 unique types of ARGs,and could further be divided into 69 drug resistance classes based on the antimicrobials to which they confer resistance.And then,451 samples from wild boars and domestic pigs from eight various farms were used for further investigations.First,we systematically analyzed the diversity and distribution of fecal resistome in pigs under different feeding patterns(natural living,semi-free ranging and standard farms).Tetracycline resistance was most abundant in pig feces,while aminoglycoside resistance had the highest richness.Pigs raised in commercial farms had a significantly higher AMR level than Tibetan pigs under semi-free ranging and wild boars.In domestic pigs,we identified 30 and 41 resistance classes showing significantly different abundances in fecal samples between 25 and 240 days of age,and between cecum lumen and feces,respectively.However,there was no significant difference in the abundance and diversity of ARGs between cecum lumen and feces in wild boars.We also analyzed the relationship between ARGs and the bacteria carrying these ARGs.Among 128 ARGs that harbored in 24 bacterial species that were almost observed in all samples,only nine ARGs were core ARGs,suggesting that most of the ARGs in these bacterial species might be acquired.Finally,we selected three subsets of ARGs as the indicators for evaluating the pollution level of ARGs in samples with high accuracy(r=0.73?0.89).The PIGC and MAGs constructed in this study provide important resources for the studies about swine gut microbiome.And the result of ARGs in various farms under different feeding patterns provides a reference for optimizing the use of antimicrobials and evaluating the risk of the pollution by ARGs in pig farms.