In Portugal, the epidemiological stage for the spread of carbapenemase-producing Enterobacteriaceae (CPE) increased from sporadic isolates or single hospital clones (2010–2013), to hospital outbreaks, later. Here we report data from a 6-month study performed under the European Survey on Carbapenemase-Producing Enterobacteriaceae (EuSCAPE). During the study period, 67 isolates (61 Klebsiella pneumoniae and 6 Escherichia coli) non-susceptible to carbapenems were identified in participant hospital laboratories. We detected 37 blaKPC–type (including one new variant: blaKPC–21), 1 blaGES–5, and 1 blaGES–6 plus blaKPC–3, alone or in combination with other bla genes. Bioinformatics analysis of the KPC-21-producing E. coli identified the new variant blaKPC–21 in a 12,748 bp length plasmid. The blaKPC–21 gene was harbored on a non-Tn4401 element, presenting upstream a partial ISKpn6 (ΔISKpn6/ΔtraN) with the related left IR (IRL) and downstream a truncated Tn3 transposon. PFGE and MLST analysis showed an important diversity, as isolates belonged to distinct PFGE and STs profiles. In this study, we highlighted the presence of the high-risk clone E. coli sequence-type (ST) 131 clade C/H30. This worldwide disseminated E. coli lineage was already detected in Portugal among other antibiotic resistance reservoirs. This study highlights the intra- and inter-hospital spread and possible intercontinental circulation of CPE isolates.

We studied a commensal colistin-resistant Escherichia coli isolated from a swine cecum sample collected at a slaughter, in Portugal. Antimicrobial susceptibility phenotype of E. coli LV23529 showed resistance to colistin at a minimum inhibitory concentration of 4 mg/L. Whole genome of E. coli LV23529 was sequenced using a MiSeq system and the assembled contigs were analyzed for the presence of antibiotic resistance and plasmid replicon types using bioinformatics tools. We report a novel mcr-1 gene variant (mcr-1.9), carried by an IncX4 plasmid, where one-point mutation at nucleotide T1238C leads to Val413Ala substitution. The mcr-1.9 genetic context was characterized by an IS26 element upstream of the mcr-pap2 element and by the absence of ISApl1. Bioinformatic analysis also revealed genes conferring resistance to β-lactams, sulphamethoxazole, trimethoprim, chloramphenicol and colistin, corresponding to the phenotype noticed. Moreover, we highlight the presence of mcr-1.9 plus blaCTX-M-8, a blaESBL gene rarely detected in Europe in isolates of animal origin; these two genes were located on different plasmids with 33,303 and 89,458 bp, respectively. MCR-1.9-harboring plasmid showed high identity to other X4-type mcr-1-harboring plasmids characterized worldwide, which strongly suggests that the presence of PMCR-encoding genes in food-producing animals, such as MCR-1.9, represent a potential threat to humans, as it is located in mobile genetic elements that have the potential to spread horizontally.