In-depth Study of a Nosocomial Outbreak Caused by XDR-Pseudomonas aeruginosa Using Whole Genome Sequencing Coupled with a PCR Targeting Strain-Specific SNPs.

An outbreak involving 14 patients infected by an extensively drug resistance Pseudomonas aeruginosa strain was detected in a hospital in Madrid, Spain. Our objective was to evaluate an alternative strategy to investigate in-depth the outbreak by means of molecular and genomic approaches. Pulsed-Field-Gel-Electrophoresis was applied as a first-line approach, followed by a more refined whole genome sequencing analysis. Single nucleotide polymorphisms identified by whole genome sequencing, were used to design a specific-polymerase-chain-reaction for screening unsuspected cases infected by the outbreak strain. Whole genome sequencing alerted us of the existence of a greater diversity than initially assumed, splitting the Pulsed-Field-Gel-Electrophoresis -associated outbreak isolates into 4 groups, two of which represented coincidental transmissions unrelated to the outbreak. A multiplex allele-specific-polymerase-chain-reaction targeting outbreak specific single nucleotide polymorphisms was applied on 290 isolates, which allowed us to identify 25 additional cases related to the outbreak during 2011-17. Whole genome sequencing coupled with an outbreak strain-specific-polymerase-chain-reaction enabled us to markedly redefine the initial picture of the outbreak by i) ruling out initially suspected cases, ii) defining likely independent coincidental transmission events, iii) predating the starting point of the outbreak iv) capturing new unsuspected cases, and v) revealing that the outbreak was still active.