| Summary: | Colistin is classified as a high-priority critical antimicrobial by the World Health Organization (WHO). A better understanding of the biological cost imposed by <i>mcr</i>-plasmids is paramount to comprehending their spread and may facilitate the decision about the ban of colistin in livestock. This study aimed to assess the prevalence of <i>mcr</i> and ESBL genes from 98 <i>Escherichia coli</i> and 142 <i>Salmonella enterica</i> isolates from food-producing animals and the impact of the <i>mcr-1</i> acquisition on bacterial fitness. Only <i>mcr-1</i> was identified by multiplex PCR (<i>mcr-1</i> to <i>mcr</i>-<i>10</i>) in 15.3% of <i>E. coli</i>. Colistin MICs ranged between 8–32 mg/L. In four isolates, <i>bla</i><sub><i>TEM-1</i></sub>, <i>bla</i><sub><i>CTX-M-1</i></sub>, and <i>bla</i><sub><i>CTX-M-15</i></sub> co-existed with <i>mcr-1</i>. The IncH12, IncHI1, IncP, IncN, and IncI plasmids were transferred by conjugation to <i>E. coli</i> J53 at frequencies of 10<sup>−7</sup> to 10<sup>−2</sup> cells/recipient. Growth kinetics assays showed that transconjugants had a significantly lower growth rate than the recipient (<i>p</i> < 0.05), and transconjugants’ average growth rate was higher in the absence than in the presence of colistin (1.66 versus 1.32 (<i>p</i> = 0.0003)). Serial transfer assay during 10 days demonstrated that plasmid retention ranged from complete loss to full retention. Overall, <i>mcr-1</i>-bearing plasmids impose a fitness cost, but the loss of plasmids is highly variable, suggesting that other factors beyond colistin pressure regulate the plasmid maintenance in a bacterial population, and colistin withdrawal will not completely lead to a decrease of <i>mcr-1</i> levels.
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