This study aimed to investigate the identification of blaCARBA-positive multidrug-resistant Mixta calida isolates from human hosts and to elucidate their genomic determinants in a species-wide context.

Two carbapenemase-producing M. calida isolates were received by the Galway Reference Laboratory Service in Ireland between June and July 2024. One isolate originated from a sputum sample, while the other was recovered from a routine screening rectal swab. Initial identification was performed using MALDI-ToF mass spectrometry, with genomic confirmation via 16S rRNA sequencing, digital DNA-DNA hybridization, and Average Nucleotide Identity analysis. Antimicrobial susceptibility testing was conducted using a MicroScan panel, following EUCAST and CLSI guidelines. Whole-genome sequencing, plasmid replicon typing, and antibiotic-resistance-gene and virulence-factor profiling were employed. Comparative analysis included all additional canonical M. calida genomes from NCBI database.

Both Irish isolates were taxonomically placed as M. calida and exhibited multidrug resistance against penicillins, cephalosporins, monobactams and ertapenem. The acquired genes blaKPC-3, blaOXA-9, and blaTEM-122 were detected on plasmid-borne contigs, indicating horizontal acquisition. Seven plasmid replicon types were shared between the two isolates. Both plasmid replicons and acquired antimicrobial-resistance-genes (ARGs) were seldomly identified across the species. Phylogenetic inference based on core genome analysis identified a monophyletic cluster, suggesting a single introductory event.

This study documents a dual occurrence of blaCARBA-positive M. calida in human colonisation and infection. The findings highlight the potential for horizontal-gene-transfer to drive the emergence of multidrug-resistant profiles in the species, underscoring the need for enhanced surveillance, diagnostic precision, and targeted infection control strategies to mitigate public health risks.

This study reports blaESBL and blaCARBA-positive multi-drug resistant Mixta calida isolates from distinct human hosts. Genomic analysis revealed the co-occurrence of plasmid-borne resistance genes blaKPC-3, blaOXA-9, and blaTEM-122. Species-wide phylogenetic analysis grouped the two isolates into a monophyletic cluster, suggesting a single introductory event.