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Replication of the Ordered, Nonredundant Library of Pseudomonas aeruginosa strain PA14 Transposon Insertion Mutants
Published on: May 4, 2018
A multispecies outbreak of carbapenem-resistant bacteria harboring the blaKPC gene in a non-classical transposon
Aniela Wozniak1,2, Cristian Figueroa1, Francisco Moya-Flores3,4
1Laboratory of Microbiology, Department of Clinical Laboratories, Centro Médico San Joaquín, Escuela de Medicina, Pontificia Universidad Católica de Chile, 3rd floor, Vicuña Mackenna, 4686, Santiago, Chile.
Background:
Klebsiella pneumoniae is the most frequent KPC-producing bacteria. The blaKPC gene is frequently embedded in Tn4401 transposon, and less frequently in non-Tn4401 elements (NTEKPC) variants I-III. The first case of KPC in the UC-CHRISTUS Clinical Hospital was detected in Pseudomonas aeruginosa. Soon after this event, KPC was detected in 2 additional Pseudomonas aeruginosa, 3 Escherichia coli, 3 Enterobacter cloacae, 3 Klebsiella pneumoniae, and 1 Citrobacter freundii, isolated from 6 different patients. We aimed to elucidate the possible mechanisms of genetic transfer and dissemination of the blaKPC gene among isolates of this multispecies outbreak. A molecular epidemiology analysis of the above mentioned clinical isolates (n = 13) through Multi-Locus Sequence Typing, plasmid analysis, Pulsed-Field Gel-Electrophoresis, and Whole-genome sequencing (WGS) was performed.
Results:
High-risk sequence types were found: K. pneumoniae ST11, P. aeruginosa ST654, and E. cloacae ST114. All enterobacterial isolates were not clonal except for 3 E. coli isolated from the same patient. WGS analysis in 6 enterobacterial isolates showed that 4 of them had blaKPC embedded in a novel variant of NTEKPC designated NTEKPC-IIe. Upstream of blaKPC gene there was a 570 pb truncated blaTEM-1 gene followed by an insertion sequence that was 84% similar to ISEc63, a 4473 bp element of the Tn3 family. Downstream the blaKPC gene there was a truncated ISKpn6 gene, and the inverted repeat right sequence of Tn4401. The ISec63-like element together with the blaKPC 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. P. aeruginosa isolates carried blaKPC gene embedded in a typical Tn4401b transposon in a different plasmid, suggesting that there was no plasmid transfer between Enterobacteriaceae and P. aeruginosa as initially hypothesized.
Conclusions:
Most enterobacterial isolates had blaKPC embedded in the same NTEKPC-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.
Insights
A novel genetic element, NTEKPC-IIe, facilitated the spread of the blaKPC gene among different bacterial species during a hospital outbreak. This horizontal gene transfer challenges traditional infection control methods.
Area of Science:
- Microbiology
- Genetics
- Infectious Diseases
Background:
- Klebsiella pneumoniae frequently harbors the blaKPC gene within the Tn4401 transposon or less common non-Tn4401 elements (NTEKPC).
- The first KPC detection in the UC-CHRISTUS Clinical Hospital occurred in Pseudomonas aeruginosa, followed by cases in other species including Escherichia coli, Enterobacter cloacae, K. pneumoniae, and Citrobacter freundii.
Purpose of the Study:
- To investigate the genetic transfer and dissemination mechanisms of the blaKPC gene during a multispecies KPC outbreak.
- To characterize the molecular epidemiology of clinical isolates involved in the outbreak.
Main Methods:
- Multi-Locus Sequence Typing (MLST)
- Plasmid analysis
- Pulsed-Field Gel-Electrophoresis (PFGE)
- Whole-genome sequencing (WGS)
Main Results:
- High-risk sequence types were identified: K. pneumoniae ST11, P. aeruginosa ST654, and E. cloacae ST114.
- WGS revealed a novel NTEKPC variant, NTEKPC-IIe, carrying the blaKPC gene in four enterobacterial isolates.
- The NTEKPC-IIe element, containing blaKPC and Tn4401 remnants, was inserted into a conjugative plasmid, while P. aeruginosa isolates harbored blaKPC within a typical Tn4401b transposon on a different plasmid.
Conclusions:
- The multispecies KPC outbreak was primarily driven by horizontal gene transfer of the blaKPC gene via the NTEKPC-IIe element, not clonal spread.
- The dissemination of blaKPC through a novel mobile genetic element presents significant challenges for infection control strategies.
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