Dataset Information

Molecular dissection of the evolution of carbapenem-resistant multilocus sequence type 258 Klebsiella pneumoniae.

ABSTRACT: Infections caused by drug-resistant bacteria are a major problem worldwide. Carbapenem-resistant Klebsiella pneumoniae, most notably isolates classified as multilocus sequence type (ST) 258, have emerged as an important cause of hospital deaths. ST258 isolates are predominantly multidrug resistant, and therefore infections caused by them are difficult to treat. It is not known why the ST258 lineage is the most prevalent cause of multidrug-resistant K. pneumoniae infections in the United States and other countries. Here we tested the hypothesis that carbapenem-resistant ST258 K. pneumoniae is a single genetic clone that has disseminated worldwide. We sequenced to closure the genomes of two ST258 clinical isolates and used these genomes as references for comparative genome sequencing of 83 additional clinical isolates recovered from patients at diverse geographic locations worldwide. Phylogenetic analysis of the SNPs in the core genome of these isolates revealed that ST258 K. pneumoniae organisms are two distinct genetic clades. This unexpected finding disproves the single-clone hypothesis. Notably, genetic differentiation between the two clades results from an ? 215-kb region of divergence that includes genes involved in capsule polysaccharide biosynthesis. The region of divergence appears to be a hotspot for DNA recombination events, and we suggest that this region has contributed to the success of ST258 K. pneumoniae. Our findings will accelerate research on novel diagnostic, therapeutic, and vaccine strategies designed to prevent and/or treat infections caused by multidrug resistant K. pneumoniae.

SUBMITTER: Deleo FR

PROVIDER: S-EPMC3977278 | biostudies-literature | 2014 Apr

REPOSITORIES: biostudies-literature

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Molecular dissection of the evolution of carbapenem-resistant multilocus sequence type 258 Klebsiella pneumoniae.

Deleo Frank R FR Chen Liang L Porcella Stephen F SF Martens Craig A CA Kobayashi Scott D SD Porter Adeline R AR Chavda Kalyan D KD Jacobs Michael R MR Mathema Barun B Olsen Randall J RJ Bonomo Robert A RA Musser James M JM Kreiswirth Barry N BN

Proceedings of the National Academy of Sciences of the United States of America 20140317 13

Infections caused by drug-resistant bacteria are a major problem worldwide. Carbapenem-resistant Klebsiella pneumoniae, most notably isolates classified as multilocus sequence type (ST) 258, have emerged as an important cause of hospital deaths. ST258 isolates are predominantly multidrug resistant, and therefore infections caused by them are difficult to treat. It is not known why the ST258 lineage is the most prevalent cause of multidrug-resistant K. pneumoniae infections in the United States a ...[more]

PMID: 24639510

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Project description:Klebsiella pneumoniae is rapidly acquiring resistance to all known antibiotics, including carbapenems. Multilocus sequence type ST258 (sequence type 258), carrying a gene encoding the K. pneumoniae carbapenemase (blaKPC) on a transmissible plasmid, is the most prevalent carbapenem-resistant Enterobacteriaceae (CRE) in the United States and has disseminated worldwide. Previously, whole-genome sequencing identified core genome single nucleotide variants that divide ST258 into two distinct clades, ST258a and ST258b. Furthermore, a subset of ST258b strains have a 347-base deletion within the enterobactin (Ent) exporter gene entS Despite the predicted inability of these strains to secrete the siderophore Ent, this clade is prevalent among clinical isolates, indicating that a full-length entS gene is not necessary for infection. To compare the transcriptional responses of ST258 subtypes to iron limitation, we performed transcriptome sequencing (RNA-Seq) in minimal medium alone or supplemented with iron or human serum and measured gene expression patterns. Iron limitation induced differential expression of distinct iron acquisition pathways when comparing ST258a and ST258b strains, including the upregulation of the hemin transport operon in entS partial deletion isolates. To measure how K. pneumoniae strains vary in iron chelation and siderophore production, we performed in vitro chrome azurol S (CAS) and Arnow assays as well as mass spectrometry. We determined that both ST258a and ST258b strains grow under iron-depleted conditions, can utilize hemin for growth, and secrete Ent, despite the partial entS deletion in a subset of ST258b strains. All carbapenem-resistant (CR) K. pneumoniae strains tested were susceptible to growth inhibition by the Ent-sequestering innate immune protein lipocalin 2.IMPORTANCE Carbapenem-resistant Enterobacteriaceae, including K. pneumoniae, are a major health care concern worldwide because they cause a wide range of infection and are resistant to all or nearly all antibiotics. To cause infection, these bacteria must acquire iron, and a major mechanism of acquiring iron is by secreting a molecule called enterobactin that strips iron from host proteins. However, a subset of carbapenem-resistant K. pneumoniae strains that lack a portion of the entS gene that is required for enterobactin secretion was recently discovered. To understand how these mutant strains obtain iron, we studied their transcriptional responses, bacterial growth, and enterobactin secretion under iron-limited conditions. We found that strains both with mutated and intact entS genes grow under iron-limiting conditions, secrete enterobactin, and utilize an alternate iron source, hemin, for growth. Our data indicate that carbapenem-resistant K. pneumoniae can use varied methods for iron uptake during infection.

Project description:Carbapenem-resistant Enterobacteriaceae, especially carbapenemase-producing Klebsiella pneumoniae, is an urgent problem in health care facilities worldwide. K. pneumoniae isolates classified as sequence type 11 (ST11) are largely responsible for the spread of carbapenem-resistant K. pneumoniae (CRKP) in China. Our previous phylogenetic reconstruction suggested that CRKP ST11 capsular locus 64 (KL64) was derived from an ST11-KL47 ancestor through recombination. However, the molecular origin of KL64 remains largely unknown, and our understanding of the recombination is incomplete. Here, we screened a global sample of 22,600 K. pneumoniae genomes and searched for KL64-harboring STs, which were found to be ST1764, ST3685, ST1764-1LV, ST30, ST505, ST147, and ST11, wherein ST1764, ST3685, ST1764-1LV, and ST30 belonged to a clonal complex, CC1764. We compared the genetic structures of representative strains from ST11-KL47, ST11-KL64, CC1764-KL64, ST505-KL64, and ST147-KL64 and further performed phylogenetic analysis and single-nucleotide polymorphism analysis among 248 isolates from all these STs. The results suggested a recombination event has occurred in a homologous ~154-kb region covering KL and the lipopolysaccharide biosynthesis locus (OL) between a recipient ST11-KL47-OL101 and a donor CC1764 (except ST30), giving rise to ST11-KL64-O2v1 strains (recombination I). Furthermore, we also found an infrequent ST11-KL64-O2v1 subclone which was not produced by recombination I but was hybridized from ST11-KL47-OL101 and ST147-KL64-O2v1 strains through recombination of a homologous ~485-kb region covering KL and OL (recombination II). Our findings provide important insights into the role of recombination in the evolution of clinical strains and the diversity of capsule and lipopolysaccharide of widely distributed KPC-associated ST11 K. pneumoniae in China. IMPORTANCE Chromosomal recombination events are considered to contribute to the genetic diversification and ultimate success of many bacterial pathogens. A previous study unravelled the molecular evolution history of ST258 strains, which have been largely responsible for the spread of KPC in the United States. Here, we used comparative genomic analyses to discover two recombination events in ST11 CRKP strains, which is a predominant KPC-associated CRKP clone in China. Two new ST11-CRKP subclones with altered capsule and lipopolysaccharide, which are two primary determinants of antigenicity and antigenic diversity among K. pneumoniae, were produced through these two recombination events, respectively. Horizontal transfer of the KL and OL appears to be a crucial element driving the molecular evolution of K. pneumoniae strains. These findings not only extend our understanding of the molecular evolutionary history of ST11 but also are an important step toward the development of preventive, diagnostic, and therapeutic strategies for CRKP.

Dataset Information

Molecular dissection of the evolution of carbapenem-resistant multilocus sequence type 258 Klebsiella pneumoniae.

Publications

Molecular dissection of the evolution of carbapenem-resistant multilocus sequence type 258 Klebsiella pneumoniae.

Similar Datasets

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