Abstract
Shotgun metagenomics is a powerful tool to identify antimicrobial resistance (AMR) genes in microbiomes but has the limitation that extrachromosomal DNA, such as plasmids, cannot be linked with the host bacterial chromosome. Here we present a comprehensive laboratory and bioinformatics pipeline HAM-ART (Hi-C Assisted Metagenomics for Antimicrobial Resistance Tracking) optimised for the generation of metagenome-assembled genomes including both chromosomal and extrachromosomal AMR genes. We demonstrate the performance of the pipeline in a study comparing 100 pig faecal microbiomes from low- and high-antimicrobial use pig farms (organic and conventional farms). We found significant differences in the distribution of AMR genes between low- and high-antimicrobial use farms including a plasmid-borne lincosamide resistance gene exclusive to high-antimicrobial use farms in three species of Lactobacilli. The bioinformatics pipeline code is available at https://github.com/lkalmar/HAM-ART.
Highlights
The emergence of resistance to antimicrobials in bacteria can occur by spontaneous mutation or by the acquisition of mobile genetic elements carrying antimicrobial resistance (AMR) genes [1]
In this paper we describe a user-friendly, affordable and comprehensive workflow that can identify, associate and track AMR genes in bacteria
We demonstrate the efficiency and reliability of the method by comparing 50 faecal microbiomes from pig farms with high-antibiotic use, and 50 faecal microbiomes from pig farms
Summary
The emergence of resistance to antimicrobials in bacteria can occur by spontaneous mutation or by the acquisition of mobile genetic elements carrying antimicrobial resistance (AMR) genes [1] (for example, plasmids via natural transformation or conjugation, or bacteriophages via transduction [2]). Salmonella-inflicted enteropathy has been shown to elicit parallel blooms of the pathogen and of resident commensal E. coli These blooms boosted horizontal gene transfer (HGT) in general, and the transfer of a conjugative colicin-plasmid p2 from an introduced Salmonella enterica serovar Typhimurium to commensal E. coli [8]. It has been shown that the use of in-feed antimicrobials leads to a bloom in AMR genes in the bacteriophage metagenome recovered from treated pigs [9], it is unclear what the sources or destinations of these genes are. These observations suggest that HGT between pathogenic and commensal bacteria is a common occurrence in humans and animals and is likely to contribute to the persistence and spread of AMR. Many previous studies on the spread of AMR from animal sources have focused on AMR of pathogens, with less emphasis on genes within indigenous microbiota that may pass to humans from animals (and vice versa) but be difficult to culture
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