Incorporating the plasmidome into antibiotic resistance surveillance in animal agriculture

Mobile genetic elements (MGE) carrying resistance genes represent a unique challenge to risk assessment and surveillance of antimicrobial resistance (AMR). Yet determining the mobility of resistance genes within animal microbiomes is essential to evaluating the potential dissemination from livestock to potential human pathogens, as well as evaluating co-selection mechanisms that may impact persistence of resistance genes with changing antibiotic use patterns. Current surveillance efforts utilize phenotypic testing and sequencing of individual isolates for tracking of AMR in livestock. In this work, we investigated the utility of using long-read sequencing of the plasmids from mixed Enterobacterales enrichments of swine fecal samples as a surveillance strategy for AMR plasmids. Enrichments were performed in either MacConkey broth without selection or with selection by addition of tetracycline or ceftriaxone, and plasmids were extracted and sequenced in order to evaluate the diversity of plasmids enriched by each method. Intact resistance plasmids were successfully assembled, as well as complex resistance transposons carrying multiple repeated elements that would interfere with assembly by short read sequencing technologies. Comparison of the assembled plasmids with representatives from public databases confirmed the quality of the assemblies and also revealed the occurrence of IncI2 plasmids carrying bla_CMY-2 in Ontario swine samples, which have not been found in previous studies.