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Pseudomonas aeruginosa is one of the most critical bacterial pathogens associated with chronic infections in cystic fibrosis patients. Here we show the phenotypic and genotypic characterization of five consecutive multidrug-resistant isolates of P. aeruginosa collected during a month from a CF patient with end-stage lung disease and fatal outcome. The isolates exhibited distinct colony morphologies and pigmentation and differences in their capacity to produce biofilm and virulence potential evaluated in larvae of Galleria mellonella . Whole genome-sequencing showed that isolates belonged to a novel sequence type ST3449 and serotype O6. Analysis of their resistome demonstrated the presence of genes bla OXA-396 , bla PAO , aph(3')-IIb , catB , crpP and fosA and new mutations in chromosomal genes conferring resistance to different antipseudomonal antibiotics. Genes exoS , exoT , exoY , toxA , lasI , rhlI and tse1 were among the 220 virulence genes detected. The different phenotypic and genotypic features found reveal the adaptation of clone ST3449 to the CF lung environment by a number of mutations affecting genes related with biofilm formation, quorum sensing and antimicrobial resistance. Most of these mutations are commonly found in CF isolates, which may give us important clues for future development of new drug targets to combat P. aeruginosa chronic infections.

An extensively drug-resistant (XDR) Klebsiella pneumoniae isolate MS3802 from a tracheostomy exudate was whole-genome sequenced using MiSeq and Oxford Nanopore MinION platforms in order to identify the antimicrobial resistance and virulence determinates and their genomic context. Isolate MS3802 belonged to the clone ST23 and presented a capsular serotype K1, associated with hypervirulent K. pneumoniae (hvKp) isolates. The isolate harboured a chromosomally encoded bla CTX-M-15 gene and contained a large IncHI1B hybrid virulence/resistance plasmid carrying another copy of the bla CTX-M-15 and the virulence factors iucABCD-iutA, iroBCDN, rmpA and rmpA2. The carbapenemase gene bla OXA-48 was found in a Tn 1999 -like transposon and the 16S rRNA methylase armA gen located in the vicinity of other antibiotic-resistant genes on an IncM2 plasmid. This study represents, to the best of our knowledge, the first description of a bla CTX-M-15 -, bla OXA-48 - and armA -harbouring K. pneumoniae of ST23 and capsular serotype K1 in Spain. Our report emphasizes the importance of implementing new surveillance strategies to monitor the risk of emergence and spread of such XDR and hypervirulent K. pneumoniae isolates.