Dietary zinc supplementation inhibits bacterial plasmid conjugation in vitro by regulating plasmid replication ( rep ) and transfer ( tra ) genes

Abstract

Humans use dietary supplements for several intended effects, such as supplementing malnutrition. While these compounds have been developed for host end benefits, their ancillary impact on the gut microbiota remains unclear. The human gut has been proposed as a reservoir for the prevalent lateral transfer of antimicrobial resistance and virulence genes in bacteria through plasmid conjugation. Here, we studied the effect of dietary zinc supplements on the incidence of plasmid conjugation in vitro . Supplement effects were analyzed through standardized broth conjugation assays. The avian pathogenic Escherichia coli (APEC) strain APEC-O2-211 was a donor of the multidrug resistance plasmid pAPEC-O2-211A-ColV, and the human commensal isolate E. coli HS-4 was the plasmid-free recipient. Bacterial strains were standardized and mixed 1:1 and supplemented 1:10 with water, or zinc derived from either commercial zinc supplements or zinc gluconate reagent at varying concentrations. We observed a significant reduction in donors, recipients, and transconjugant populations in conjugations supplemented with zinc, with a dose-dependent relationship. Additionally, we observed a significant reduction ( P < 0.05) in log conjugation efficiency in zinc-treated reactions. Upregulation of the mRNA for the plasmid replication initiation gene repA and the subset of transfer genes M , J , E , K , B , P , C , W , U , N , F , Q , D , I , and X was observed. Furthermore, we observed a downregulation of the conjugal propilin gene traA and the entry exclusion gene traS . This study demonstrates the effect of dietary zinc supplements on the conjugal transfer of a multidrug resistance plasmid between pathogenic and commensal bacteria during in vitro conditions. IMPORTANCE This study identifies dietary zinc supplementation as a potential novel intervention for mitigating the emergence of multidrug resistance in bacteria, thus preventing antibiotic treatment failure and death in patients and animals. Further studies are required to determine the applicability of this approach in an in vivo model.