The emergence of mcr plasmid-mediated colistin-resistant extended-spectrum β-lactamase (ESBL)-producing Enterobacterales among companion dogs and cats poses a risk of the animals acting as reservoirs for cross-species transmission. However, current knowledge of mcr-harboring ESBL-producing Enterobacterales in companion dogs and cats is still limited; thus, the genetic and phenotypic characteristics of the bacterial isolates and plasmids, in companion dogs and cats, remain to be elucidated. Here, we identified mcr gene-harboring ESBL-producing Escherichia coli isolates during whole-genome sequencing of ESBL-producing E. coli isolates from a dog and a cat in Osaka, Japan. Colistin-resistant MY732 isolate from a dog carried two plasmids: mcr-1.1-harboring IncI2 plasmid and blaCTX-M-14-harboring IncFIB plasmid. Conjugation assays revealed that both plasmids can be co-transferred even though the IncFIB plasmid lacked a conjugal transfer gene cassette. The other isolate MY504 from a cat harbored two bla genes and mcr-9 on the identical IncHI2 plasmid. This isolate was not resistant to colistin, which is likely to be due to deletion of the regulatory two-component QseBC system associated with the mcr-9 expression. To the best of our knowledge, this is the first report of a colistin-resistant ESBL-producing E. coli isolate harboring mcr-1 from a companion dog in Japan. Given that the mcr gene-harboring IncI2 and IncHI2 plasmids in this study shared high homology with plasmids from human or animal-derived Enterobacterales, companion dogs and cats may act as important reservoirs for cross-species transmission of the mcr gene in the community, in Japan. Copyright © 2023 Elsevier B.V. All rights reserved.