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Whole genome sequencing of extended-spectrum β-lactamase producing Klebsiella pneumoniae isolated from a patient in Lebanon.

Tokajian S, Eisen JA, Jospin G, Farra A, Coil DA - Front Cell Infect Microbiol (2015)

Bottom Line: Illumina paired-end libraries were prepared and sequenced, which resulted in 4,220,969 high-quality reads.Sequencing analysis revealed that the isolate harbored different β-lactamase genes, including bla oxa-1, bla CTX-M-15, bla SHV-11, and bla TEM-1b.The isolate was also characterized by the concomitant presence of other resistance determinants most notably acc(6')-lb-cr and qnrb1.

View Article: PubMed Central - PubMed

Affiliation: Department of Natural Sciences, School of Arts and Sciences, Lebanese American University Byblos, Lebanon.

ABSTRACT

Objective: The emergence of extended-spectrum β-lactamase (ESBL)-producing bacteria is now a critical concern. The ESBL-producing Klebsiella pneumoniae constitutes one of the most common multidrug-resistant (MDR) groups of gram-negative bacteria involved in nosocomial infections worldwide. In this study we report on the molecular characterization through whole genome sequencing of an ESBL-producing K. pneumoniae strain, LAU-KP1, isolated from a stool sample from a patient admitted for a gastrointestinal procedure/surgery at the Lebanese Amrican University Medical Center-Rizk Hospital (LAUMCRH) in Lebanon.

Methods: Illumina paired-end libraries were prepared and sequenced, which resulted in 4,220,969 high-quality reads. All sequence processing and assembly were performed using the A5 assembly pipeline.

Results: The initial assembly produced 86 contigs, for which no scaffolding was obtained. The final collection of contigs was submitted to GenBank. The final draft genome sequence consists of a combined 5,632,663 bases with 57% G+C content. Automated annotation was performed using the RAST annotation server. Sequencing analysis revealed that the isolate harbored different β-lactamase genes, including bla oxa-1, bla CTX-M-15, bla SHV-11, and bla TEM-1b. The isolate was also characterized by the concomitant presence of other resistance determinants most notably acc(6')-lb-cr and qnrb1. The entire plasmid content was also investigated and revealed homology with four major plasmids pKPN-IT, pBS512_2, pRSF1010_SL1344, and pKPN3.

Conclusions: The potential role of K. pneumonia as a reservoir for ESBL genes and other resistance determinants is along with the presence of key factors that favor the spread of antimicrobial resistance a clear cause of concern and the problem that Carbapenem-non-susceptible ESBL isolates are posing in hospitals should be reconsidered through systematic exploration and molecular characterization.

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Phylogenetic tree of all assembled K. pneumoniae genomes available on the NCBI ftp server as of 2-11-15, based on a concatenated alignment of 37 conserved genes obtained using PhyloSift (Darling et al., 2014). The tree was constructed using default settings in FastTree (Price et al., 2010) and edited and visualized with Dendroscope v.3 (Huson and Scornavacca, 2012). Clades with zero branch lengths were collapsed to a single representative. K. pneumoniae LAU-KP1 is indicated by a square, and the reference genome used in Figure 2 is indicated by a diamond.
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Figure 1: Phylogenetic tree of all assembled K. pneumoniae genomes available on the NCBI ftp server as of 2-11-15, based on a concatenated alignment of 37 conserved genes obtained using PhyloSift (Darling et al., 2014). The tree was constructed using default settings in FastTree (Price et al., 2010) and edited and visualized with Dendroscope v.3 (Huson and Scornavacca, 2012). Clades with zero branch lengths were collapsed to a single representative. K. pneumoniae LAU-KP1 is indicated by a square, and the reference genome used in Figure 2 is indicated by a diamond.

Mentions: Furthermore, the phylogenetic analysis of LAU-KP1 showed that it falls within a clade of other known multidrug resistant K. pneumoniae strains and demonstrated that there is considerable genomic diversity among the included isolates (Figure 1). Within the closely related isolates was K. pneumonia 7699 previously cultured from a perianal swab of a patient admitted to the intensive care unit of the University of Maryland Medical Center (UMMC) in Baltimore, MD (Hazen et al., 2014). The isolate had a 167-kb IncA/C plasmid encoding the FOX-5 β-lactamase, a CARB-2 β-lactamase, other antimicrobial resistance genes, and heavy metal resistance genes. On the other hand, a considerable phylogenetic difference was detected between LAU-Kp1 and K. pneumoniae ATCC BAA-2146, which is the first U.S. isolate found encoding the gene for the broad-spectrum and carbapenem-active metallo-β-lactamase NDM-1 (Hudson et al., 2014). Additionally, we mapped our contigs to the phylogenetically closest K. pneumoniae isolate with a complete genome, Ecl8, in order to visualize the genomic organization (Figure 2).


Whole genome sequencing of extended-spectrum β-lactamase producing Klebsiella pneumoniae isolated from a patient in Lebanon.

Tokajian S, Eisen JA, Jospin G, Farra A, Coil DA - Front Cell Infect Microbiol (2015)

Phylogenetic tree of all assembled K. pneumoniae genomes available on the NCBI ftp server as of 2-11-15, based on a concatenated alignment of 37 conserved genes obtained using PhyloSift (Darling et al., 2014). The tree was constructed using default settings in FastTree (Price et al., 2010) and edited and visualized with Dendroscope v.3 (Huson and Scornavacca, 2012). Clades with zero branch lengths were collapsed to a single representative. K. pneumoniae LAU-KP1 is indicated by a square, and the reference genome used in Figure 2 is indicated by a diamond.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4389573&req=5

Figure 1: Phylogenetic tree of all assembled K. pneumoniae genomes available on the NCBI ftp server as of 2-11-15, based on a concatenated alignment of 37 conserved genes obtained using PhyloSift (Darling et al., 2014). The tree was constructed using default settings in FastTree (Price et al., 2010) and edited and visualized with Dendroscope v.3 (Huson and Scornavacca, 2012). Clades with zero branch lengths were collapsed to a single representative. K. pneumoniae LAU-KP1 is indicated by a square, and the reference genome used in Figure 2 is indicated by a diamond.
Mentions: Furthermore, the phylogenetic analysis of LAU-KP1 showed that it falls within a clade of other known multidrug resistant K. pneumoniae strains and demonstrated that there is considerable genomic diversity among the included isolates (Figure 1). Within the closely related isolates was K. pneumonia 7699 previously cultured from a perianal swab of a patient admitted to the intensive care unit of the University of Maryland Medical Center (UMMC) in Baltimore, MD (Hazen et al., 2014). The isolate had a 167-kb IncA/C plasmid encoding the FOX-5 β-lactamase, a CARB-2 β-lactamase, other antimicrobial resistance genes, and heavy metal resistance genes. On the other hand, a considerable phylogenetic difference was detected between LAU-Kp1 and K. pneumoniae ATCC BAA-2146, which is the first U.S. isolate found encoding the gene for the broad-spectrum and carbapenem-active metallo-β-lactamase NDM-1 (Hudson et al., 2014). Additionally, we mapped our contigs to the phylogenetically closest K. pneumoniae isolate with a complete genome, Ecl8, in order to visualize the genomic organization (Figure 2).

Bottom Line: Illumina paired-end libraries were prepared and sequenced, which resulted in 4,220,969 high-quality reads.Sequencing analysis revealed that the isolate harbored different β-lactamase genes, including bla oxa-1, bla CTX-M-15, bla SHV-11, and bla TEM-1b.The isolate was also characterized by the concomitant presence of other resistance determinants most notably acc(6')-lb-cr and qnrb1.

View Article: PubMed Central - PubMed

Affiliation: Department of Natural Sciences, School of Arts and Sciences, Lebanese American University Byblos, Lebanon.

ABSTRACT

Objective: The emergence of extended-spectrum β-lactamase (ESBL)-producing bacteria is now a critical concern. The ESBL-producing Klebsiella pneumoniae constitutes one of the most common multidrug-resistant (MDR) groups of gram-negative bacteria involved in nosocomial infections worldwide. In this study we report on the molecular characterization through whole genome sequencing of an ESBL-producing K. pneumoniae strain, LAU-KP1, isolated from a stool sample from a patient admitted for a gastrointestinal procedure/surgery at the Lebanese Amrican University Medical Center-Rizk Hospital (LAUMCRH) in Lebanon.

Methods: Illumina paired-end libraries were prepared and sequenced, which resulted in 4,220,969 high-quality reads. All sequence processing and assembly were performed using the A5 assembly pipeline.

Results: The initial assembly produced 86 contigs, for which no scaffolding was obtained. The final collection of contigs was submitted to GenBank. The final draft genome sequence consists of a combined 5,632,663 bases with 57% G+C content. Automated annotation was performed using the RAST annotation server. Sequencing analysis revealed that the isolate harbored different β-lactamase genes, including bla oxa-1, bla CTX-M-15, bla SHV-11, and bla TEM-1b. The isolate was also characterized by the concomitant presence of other resistance determinants most notably acc(6')-lb-cr and qnrb1. The entire plasmid content was also investigated and revealed homology with four major plasmids pKPN-IT, pBS512_2, pRSF1010_SL1344, and pKPN3.

Conclusions: The potential role of K. pneumonia as a reservoir for ESBL genes and other resistance determinants is along with the presence of key factors that favor the spread of antimicrobial resistance a clear cause of concern and the problem that Carbapenem-non-susceptible ESBL isolates are posing in hospitals should be reconsidered through systematic exploration and molecular characterization.

Show MeSH
Related in: MedlinePlus