<jats:title>Abstract</jats:title><jats:sec>
<jats:title>Background</jats:title>
<jats:p><jats:italic>Klebsiella pneumoniae</jats:italic> is the most frequent KPC-producing bacteria. The <jats:italic>bla</jats:italic><jats:sub>KPC</jats:sub> gene is frequently embedded in Tn4401 transposon, and less frequently in non-Tn4401 elements (NTE<jats:sub>KPC</jats:sub>) variants I-III. The first case of KPC in the UC-CHRISTUS Clinical Hospital was detected in <jats:italic>Pseudomonas aeruginosa</jats:italic>. Soon after this event, KPC was detected in 2 additional <jats:italic>Pseudomonas aeruginosa</jats:italic>, 3 <jats:italic>Escherichia coli</jats:italic>, 3 <jats:italic>Enterobacter cloacae</jats:italic>, 3 <jats:italic>Klebsiella pneumoniae,</jats:italic> and 1 <jats:italic>Citrobacter freundii</jats:italic>, isolated from 6 different patients. We aimed to elucidate the possible mechanisms of genetic transfer and dissemination of the <jats:italic>bla</jats:italic><jats:sub>KPC</jats:sub> gene among isolates of this multispecies outbreak. A molecular epidemiology analysis of the above mentioned clinical isolates (<jats:italic>n</jats:italic> = 13) through Multi-Locus Sequence Typing, plasmid analysis, Pulsed-Field Gel-Electrophoresis, and Whole-genome sequencing (WGS) was performed.</jats:p>
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<jats:title>Results</jats:title>
<jats:p>High-risk sequence types were found: <jats:italic>K. pneumoniae</jats:italic> ST11, <jats:italic>P. aeruginosa</jats:italic> ST654, and <jats:italic>E. cloacae</jats:italic> ST114. All enterobacterial isolates were not clonal except for 3 <jats:italic>E. coli</jats:italic> isolated from the same patient. WGS analysis in 6 enterobacterial isolates showed that 4 of them had <jats:italic>bla</jats:italic><jats:sub>KPC</jats:sub> embedded in a novel variant of NTE<jats:sub>KPC</jats:sub> designated NTE<jats:sub>KPC</jats:sub>-IIe. Upstream of <jats:italic>bla</jats:italic><jats:sub>KPC</jats:sub> gene there was a 570 pb truncated <jats:italic>bla</jats:italic><jats:sub>TEM-1</jats:sub> gene followed by an insertion sequence that was 84% similar to ISEc63, a 4473 bp element of the Tn3 family. Downstream the <jats:italic>bla</jats:italic><jats:sub>KPC</jats:sub> gene there was a truncated ISKpn6 gene, and the inverted repeat right sequence of Tn4401. The ISec63-like element together with the <jats:italic>bla</jats:italic><jats:sub>KPC</jats:sub> gene plus Tn4401 remnants were inserted in the Tra operon involved in conjugative transfer of the plasmid. This NTE was carried in a broad host-range IncN plasmid. <jats:italic>P. aeruginosa</jats:italic> isolates carried <jats:italic>bla</jats:italic><jats:sub>KPC</jats:sub> gene embedded in a typical Tn4401b transposon in a different plasmid, suggesting that there was no plasmid transfer between <jats:italic>Enterobacteriaceae</jats:italic> and <jats:italic>P. aeruginosa</jats:italic> as initially hypothesized.</jats:p>
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<jats:title>Conclusions</jats:title>
<jats:p>Most enterobacterial isolates had <jats:italic>bla</jats:italic><jats:sub>KPC</jats:sub> embedded in the same NTE<jats:sub>KPC</jats:sub>-IIe element, suggesting that this multispecies KPC outbreak was due to horizontal gene transfer rather than clonal spread. This poses a greater challenge to infection control measures often directed against containment of clonal spread.</jats:p>
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