Mapping the Evolution of Hypervirulent Klebsiella pneumoniae

ABSTRACTHighly invasive, community-acquiredKlebsiella pneumoniaeinfections have recently emerged, resulting in pyogenic liver abscesses. These infections are caused by hypervirulentK. pneumoniae(hvKP) isolates primarily of capsule serotype K1 or K2. Hypervirulent K1 isolates belong to clonal complex 23 (CC23), indicating that this clonal lineage has a specific genetic background conferring hypervirulence. Here, we apply whole-genome sequencing to a collection ofK. pneumoniaeisolates to characterize the phylogenetic background of hvKP isolates with an emphasis on CC23. Most of the hvKP isolates belonged to CC23 and grouped into a distinct monophyletic clade, revealing that CC23 is a unique clonal lineage, clearly distinct from nonhypervirulent strains. Separate phylogenetic analyses of the CC23 isolates indicated that the CC23 lineage evolved recently by clonal expansion from a single common ancestor. Limited grouping according to geographical origin was observed, suggesting that CC23 has spread globally through multiple international transmissions. Conversely, hypervirulent K2 strains clustered in genetically unrelated groups. Strikingly, homologues of a large virulence plasmid were detected in all hvKP clonal lineages, indicating a key role inK. pneumoniaehypervirulence. The plasmid encodes two siderophores, aerobactin and salmochelin, and RmpA (regulator of the mucoid phenotype); all these factors were found to be restricted to hvKP isolates. Genomic comparisons revealed additional factors specifically associated with CC23. These included a distinct variant of a genomic island encoding yersiniabactin, colibactin, and microcin E492. Furthermore, additional novel genomic regions unique to CC23 were revealed which may also be involved in the increased virulence of this important clonal lineage.IMPORTANCEDuring the last 3 decades, hypervirulentKlebsiella pneumoniae(hvKP) isolates have emerged, causing severe community-acquired infections primarily in the form of pyogenic liver abscesses. This syndrome has so far primarily been found in Southeast Asia, but increasing numbers of cases are being reported worldwide, indicating that the syndrome is turning into a globally emerging disease. We applied whole-genome sequencing to a collection ofK. pneumoniaeclinical isolates to reveal the phylogenetic background of hvKP and to identify genetic factors associated with the increased virulence. The hvKP isolates primarily belonged to clonal complex 23 (CC23), and this clonal lineage was revealed to be clearly distinct from nonhypervirulent strains. A specific virulence plasmid was found to be associated with hypervirulence, and novel genetic determinants uniquely associated with CC23 were identified. Our findings extend the understanding of the genetic background of the emergence of hvKP clones.

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Whole-Genome Sequencing of Human ClinicalKlebsiella pneumoniaeIsolates Reveals Misidentification and Misunderstandings ofKlebsiella pneumoniae,Klebsiella variicola, andKlebsiella quasipneumoniae

mSphere ◽

10.1128/mspheredirect.00290-17 ◽

2017 ◽

Vol 2 (4) ◽

Cited By ~ 46

Author(s):

S. Wesley Long ◽

Sarah E. Linson ◽

Matthew Ojeda Saavedra ◽

Concepcion Cantu ◽

James J. Davis ◽

...

Keyword(s):

Homologous Recombination ◽

Klebsiella Pneumoniae ◽

Whole Genome Sequencing ◽

Genome Sequencing ◽

Clinical Microbiology ◽

Whole Genome ◽

Beta Lactamase ◽

Opportunistic Pathogens ◽

Content Type ◽

Strain Collection

ABSTRACTKlebsiella pneumoniaeis a major threat to public health, causing significant morbidity and mortality worldwide. The emergence of highly drug-resistant strains is particularly concerning. There has been a recognition and division ofKlebsiella pneumoniaeinto three distinct phylogenetic groups:Klebsiella pneumoniae,Klebsiella variicola, andKlebsiella quasipneumoniae.K. variicolaandK. quasipneumoniaehave often been described as opportunistic pathogens that have less virulence in humans thanK. pneumoniaedoes. We recently sequenced the genomes of 1,777 extended-spectrum-beta-lactamase (ESBL)-producingK. pneumoniaeisolates recovered from human infections and discovered that 28 strains were phylogenetically related toK.variicolaandK. quasipneumoniae. Whole-genome sequencing of 95 additional non-ESBL-producingK. pneumoniaeisolates recovered from patients found 12K. quasipneumoniaestrains. Matrix-assisted laser desorption ionization–time of flight mass spectrometry (MALDI-TOF MS) analysis initially identified all patient isolates asK. pneumoniae, suggesting a potential pitfall in conventional clinical microbiology laboratory identification methods. Whole-genome sequence analysis revealed extensive sharing of core gene content and plasmid replicons among theKlebsiellaspecies. For the first time, strains of bothK. variicolaandK. quasipneumoniaewere found to carry theKlebsiella pneumoniaecarbapenemase (KPC) gene, while anotherK. variicolastrain was found to carry the New Delhi metallo-beta-lactamase 1 (NDM-1) gene.K. variicolaandK. quasipneumoniaeinfections were not less virulent thanK. pneumoniaeinfections, as assessed by in-hospital mortality and infection type. We also discovered evidence of homologous recombination in oneK. variicolastrain, as well as one strain from a novelKlebsiellaspecies, which challenge the current understanding of interrelationships between clades ofKlebsiella.IMPORTANCEKlebsiella pneumoniaeis a serious human pathogen associated with resistance to multiple antibiotics and high mortality.K. variicolaandK. quasipneumoniaeare closely related organisms that are generally considered to be less-virulent opportunistic pathogens. We used a large, comprehensive, population-based strain collection and whole-genome sequencing to investigate infections caused by these organisms in our hospital system. We discovered thatK. variicolaandK. quasipneumoniaeisolates are often misidentified asK. pneumoniaeby routine clinical microbiology diagnostics and frequently cause severe life-threatening infections similar toK. pneumoniae. The presence of KPC inK. variicolaandK. quasipneumoniaestrains as well as NDM-1 metallo-beta-lactamase in oneK. variicolastrain is particularly concerning because these genes confer resistance to many different beta-lactam antibiotics. The sharing of plasmids, as well as evidence of homologous recombination, between these three species ofKlebsiellais cause for additional concern.

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Genomic Analysis of a Pan-Resistant Isolate ofKlebsiella pneumoniae, United States 2016

mBio ◽

10.1128/mbio.00440-18 ◽

2018 ◽

Vol 9 (2) ◽

Cited By ~ 16

Author(s):

Tom J. B. de Man ◽

Joseph D. Lutgring ◽

David R. Lonsway ◽

Karen F. Anderson ◽

Julia A. Kiehlbauch ◽

...

Keyword(s):

Public Health ◽

United States ◽

Antimicrobial Resistance ◽

Klebsiella Pneumoniae ◽

Whole Genome Sequencing ◽

Genome Sequencing ◽

The United States ◽

Whole Genome ◽

Beta Lactams ◽

Content Type

ABSTRACTAntimicrobial resistance is a threat to public health globally and leads to an estimated 23,000 deaths annually in the United States alone. Here, we report the genomic characterization of an unusualKlebsiella pneumoniae, nonsusceptible to all 26 antibiotics tested, that was isolated from a U.S. patient. The isolate harbored four known beta-lactamase genes, including plasmid-mediatedblaNDM-1andblaCMY-6, as well as chromosomalblaCTX-M-15andblaSHV-28, which accounted for resistance to all beta-lactams tested. In addition, sequence analysis identified mechanisms that could explain all other reported nonsusceptibility results, including nonsusceptibility to colistin, tigecycline, and chloramphenicol. Two plasmids, IncA/C2 and IncFIB, were closely related to mobile elements described previously and isolated from Gram-negative bacteria from China, Nepal, India, the United States, and Kenya, suggesting possible origins of the isolate and plasmids. This is one of the firstK. pneumoniaeisolates in the United States to have been reported to the Centers for Disease Control and Prevention (CDC) as nonsusceptible to all drugs tested, including all beta-lactams, colistin, and tigecycline.IMPORTANCEAntimicrobial resistance is a major public health threat worldwide. Bacteria that are nonsusceptible or resistant to all antimicrobials available are of major concern to patients and the public because of lack of treatment options and potential for spread. AKlebsiella pneumoniaestrain that was nonsusceptible to all tested antibiotics was isolated from a U.S. patient. Mechanisms that could explain all observed phenotypic antimicrobial resistance phenotypes, including resistance to colistin and beta-lactams, were identified through whole-genome sequencing. The large variety of resistance determinants identified demonstrates the usefulness of whole-genome sequencing for detecting these genes in an outbreak response. Sequencing of isolates with rare and unusual phenotypes can provide information on how these extremely resistant isolates develop, including whether resistance is acquired on mobile elements or accumulated through chromosomal mutations. Moreover, this provides further insight into not only detecting these highly resistant organisms but also preventing their spread.

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MCR-1 and OXA-48 In Vivo Acquisition in KPC-Producing Escherichia coli after Colistin Treatment

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.02540-16 ◽

2017 ◽

Vol 61 (8) ◽

Cited By ~ 32

Author(s):

Racha Beyrouthy ◽

Frederic Robin ◽

Aude Lessene ◽

Igor Lacombat ◽

Laurent Dortet ◽

...

Keyword(s):

Escherichia Coli ◽

Klebsiella Pneumoniae ◽

Whole Genome Sequencing ◽

Genome Sequencing ◽

Whole Genome ◽

Content Type ◽

First Report ◽

E Coli ◽

Carbapenem Resistant

ABSTRACT The spread of mcr-1-encoding plasmids into carbapenem-resistant Enterobacteriaceae raises concerns about the emergence of untreatable bacteria. We report the acquisition of mcr-1 in a carbapenem-resistant Escherichia coli strain after a 3-week course of colistin in a patient repatriated to France from Portugal. Whole-genome sequencing revealed that the Klebsiella pneumoniae carbapenemase-producing E. coli strain acquired two plasmids, an IncL OXA-48-encoding plasmid and an IncX4 mcr-1-encoding plasmid. This is the first report of mcr-1 in carbapenemase-encoding bacteria in France.

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Evolution and Population Dynamics of Clonal Complex 152 Community-Associated Methicillin-Resistant Staphylococcus aureus

mSphere ◽

10.1128/msphere.00226-20 ◽

2020 ◽

Vol 5 (4) ◽

Author(s):

Sharmin Baig ◽

Anders Rhod Larsen ◽

Patrícia Martins Simões ◽

Frédéric Laurent ◽

Thor Bech Johannesen ◽

...

Keyword(s):

Staphylococcus Aureus ◽

Whole Genome Sequencing ◽

Genome Sequencing ◽

Clonal Complex ◽

Whole Genome ◽

Methicillin Resistant ◽

Content Type ◽

Type V ◽

Panton Valentine Leukocidin ◽

Valentine Leukocidin

ABSTRACT Since the late 1990s, changes in the epidemiology of methicillin-resistant Staphylococcus aureus (MRSA) were recognized with the emergence of community-associated MRSA (CA-MRSA). CA-MRSA belonging to clonal complex 152 (CC152), carrying the small staphylococcal cassette chromosome mec (SCCmec) type V and encoding the Panton-Valentine leukocidin (PVL), has been observed in Europe. The aim of this study was to investigate its origin, evolution, and dissemination. Whole-genome sequencing was performed on a global collection of 149 CC152 isolates spanning 20 years (93 methicillin-susceptible S. aureus [MSSA] and 56 MRSA isolates). Core genome phylogeny, Bayesian inference, in silico resistance analyses, and genomic characterization were applied. Phylogenetic analysis revealed two major distinct clades, one dominated by MSSA and the other populated only by MRSA. The MSSA isolates were predominately from sub-Saharan Africa, whereas MRSA was almost exclusively from Europe. The European MRSA isolates all harbored an SCCmec type V (5C2&5) element, whereas other SCCmec elements were sporadically detected in MRSA from the otherwise MSSA-dominated clade, including SCCmec types IV (2B), V (5C2), and XIII (9A). In total, 93% of the studied CC152 isolates were PVL positive. Bayesian coalescent inference suggests an emergence of the European CC152-MRSA in the 1990s, while the CC152 lineage dates back to the 1970s. The CA-MRSA CC152 clone mimics the European CC80 CA-MRSA lineage by its emergence from a PVL-positive MSSA ancestor from North Africa or Europe. The CC152 lineage has acquired SCCmec several times, but acquisition of SCCmec type V (5C2&5) seems associated with expansion of MRSA CC152 in Europe. IMPORTANCE Understanding the evolution of CA-MRSA is important in light of the increasing importance of this reservoir in the dissemination of MRSA. Here, we highlight the story of the CA-MRSA CC152 lineage using whole-genome sequencing on an international collection of CC152. We show that the evolution of this lineage is novel and that antibiotic usage may have the potential to select for the phage-encoded Panton-Valentine leukocidin. The diversity of the strains correlated highly to geography, with higher level of resistance observed among the European MRSA isolates. The mobility of the SCCmec element is mandatory for the emergence of novel MRSA lineages, and we show here distinct acquisitions, one of which is linked to the successful clone found throughout Europe today.

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Klebsiella pneumoniae Carbapenemase (KPC)-Producing K. pneumoniae at a Single Institution: Insights into Endemicity from Whole-Genome Sequencing

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.04292-14 ◽

2015 ◽

Vol 59 (3) ◽

pp. 1656-1663 ◽

Cited By ~ 83

Author(s):

Amy J. Mathers ◽

Nicole Stoesser ◽

Anna E. Sheppard ◽

Louise Pankhurst ◽

Adam Giess ◽

...

Keyword(s):

Klebsiella Pneumoniae ◽

Whole Genome Sequencing ◽

Genome Sequencing ◽

Clinical Isolates ◽

Whole Genome ◽

Single Institution ◽

Content Type ◽

Klebsiella Pneumoniae Carbapenemase ◽

Distinct Sequence ◽

Sequence Types

ABSTRACTThe global emergence ofKlebsiella pneumoniaecarbapenemase-producingK. pneumoniae(KPC-Kp) multilocus sequence type ST258 is widely recognized. Less is known about the molecular and epidemiological details of non-ST258K. pneumoniaein the setting of an outbreak mediated by an endemic plasmid. We describe the interplay ofblaKPCplasmids andK. pneumoniaestrains and their relationship to the location of acquisition in a U.S. health care institution. Whole-genome sequencing (WGS) analysis was applied to KPC-Kpclinical isolates collected from a single institution over 5 years following the introduction ofblaKPCin August 2007, as well as two plasmid transformants. KPC-Kpfrom 37 patients yielded 16 distinct sequence types (STs). Two novel conjugativeblaKPCplasmids (pKPC_UVA01 and pKPC_UVA02), carried by the hospital index case, accounted for the presence ofblaKPCin 21/37 (57%) subsequent cases. Thirteen (35%) isolates represented an emergent lineage, ST941, which contained pKPC_UVA01 in 5/13 (38%) and pKPC_UVA02 in 6/13 (46%) cases. Seven (19%) isolates were the epidemic KPC-Kpstrain, ST258, mostly imported from elsewhere and not carrying pKPC_UVA01 or pKPC_UVA02. Using WGS-based analysis of clinical isolates and plasmid transformants, we demonstrate the unexpected dispersal ofblaKPCto many non-ST258 lineages in a hospital through spread of at least two novelblaKPCplasmids. In contrast, ST258 KPC-Kpwas imported into the institution on numerous occasions, with otherblaKPCplasmid vectors and without sustained transmission. Instead, a newly recognized KPC-Kpstrain, ST941, became associated with both novelblaKPCplasmids and spread locally, making it a future candidate for clinical persistence and dissemination.

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Whole-Genome Sequencing To Identify Drivers of Carbapenem-Resistant Klebsiella pneumoniae Transmission within and between Regional Long-Term Acute-Care Hospitals

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.01622-19 ◽

2019 ◽

Vol 63 (11) ◽

Cited By ~ 3

Author(s):

Jennifer H. Han ◽

Zena Lapp ◽

Frederic Bushman ◽

Ebbing Lautenbach ◽

Ellie J. C. Goldstein ◽

...

Keyword(s):

Klebsiella Pneumoniae ◽

Whole Genome Sequencing ◽

Genome Sequencing ◽

Treatment Options ◽

Health Care Facilities ◽

Resistant Organism ◽

Whole Genome ◽

Content Type ◽

Transmission Rates ◽

Carbapenem Resistant

ABSTRACT Carbapenem-resistant Klebsiella pneumoniae (CRKP) is an antibiotic resistance threat of the highest priority. Given the limited treatment options for this multidrug-resistant organism (MDRO), there is an urgent need for targeted strategies to prevent transmission. Here, we applied whole-genome sequencing to a comprehensive collection of clinical isolates to reconstruct regional transmission pathways and analyzed this transmission network in the context of statewide patient transfer data and patient-level clinical data to identify drivers of regional transmission. We found that high regional CRKP burdens were due to a small number of regional introductions, with subsequent regional proliferation occurring via patient transfers among health care facilities. While CRKP was predicted to have been imported into each facility multiple times, there was substantial variation in the ratio of intrafacility transmission events per importation, indicating that amplification occurs unevenly across regional facilities. While myriad factors likely influence intrafacility transmission rates, an understudied one is the potential for clinical characteristics of colonized and infected patients to influence their propensity for transmission. Supporting the contribution of high-risk patients to elevated transmission rates, we observed that patients colonized and infected with CRKP in high-transmission facilities had higher rates of carbapenem use, malnutrition, and dialysis and were older. This report highlights the potential for regional infection prevention efforts that are grounded in genomic epidemiology to identify the patients and facilities that make the greatest contribution to regional MDRO prevalence, thereby facilitating the design of precision interventions of maximal impact.

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Predicting Phenotypic Polymyxin Resistance in Klebsiella pneumoniae through Machine Learning Analysis of Genomic Data

mSystems ◽

10.1128/msystems.00656-19 ◽

2020 ◽

Vol 5 (3) ◽

Cited By ~ 3

Author(s):

Nenad Macesic ◽

Oliver J. Bear Don’t Walk ◽

Itsik Pe’er ◽

Nicholas P. Tatonetti ◽

Anton Y. Peleg ◽

...

Keyword(s):

Machine Learning ◽

Klebsiella Pneumoniae ◽

Whole Genome Sequencing ◽

Genome Sequencing ◽

Genomic Data ◽

Whole Genome Sequencing Data ◽

Whole Genome ◽

Sequencing Data ◽

Pr Genes ◽

Content Type

ABSTRACT Polymyxins are used as treatments of last resort for Gram-negative bacterial infections. Their increased use has led to concerns about emerging polymyxin resistance (PR). Phenotypic polymyxin susceptibility testing is resource intensive and difficult to perform accurately. The complex polygenic nature of PR and our incomplete understanding of its genetic basis make it difficult to predict PR using detection of resistance determinants. We therefore applied machine learning (ML) to whole-genome sequencing data from >600 Klebsiella pneumoniae clonal group 258 (CG258) genomes to predict phenotypic PR. Using a reference-based representation of genomic data with ML outperformed a rule-based approach that detected variants in known PR genes (area under receiver-operator curve [AUROC], 0.894 versus 0.791, P = 0.006). We noted modest increases in performance by using a bacterial genome-wide association study to filter relevant genomic features and by integrating clinical data in the form of prior polymyxin exposure. Conversely, reference-free representation of genomic data as k-mers was associated with decreased performance (AUROC, 0.692 versus 0.894, P = 0.015). When ML models were interpreted to extract genomic features, six of seven known PR genes were correctly identified by models without prior programming and several genes involved in stress responses and maintenance of the cell membrane were identified as potential novel determinants of PR. These findings are a proof of concept that whole-genome sequencing data can accurately predict PR in K. pneumoniae CG258 and may be applicable to other forms of complex antimicrobial resistance. IMPORTANCE Polymyxins are last-resort antibiotics used to treat highly resistant Gram-negative bacteria. There are increasing reports of polymyxin resistance emerging, raising concerns of a postantibiotic era. Polymyxin resistance is therefore a significant public health threat, but current phenotypic methods for detection are difficult and time-consuming to perform. There have been increasing efforts to use whole-genome sequencing for detection of antibiotic resistance, but this has been difficult to apply to polymyxin resistance because of its complex polygenic nature. The significance of our research is that we successfully applied machine learning methods to predict polymyxin resistance in Klebsiella pneumoniae clonal group 258, a common health care-associated and multidrug-resistant pathogen. Our findings highlight that machine learning can be successfully applied even in complex forms of antibiotic resistance and represent a significant contribution to the literature that could be used to predict resistance in other bacteria and to other antibiotics.

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Whole-Genome Sequencing of Emerging Invasive Neisseria meningitidis Serogroup W in Sweden

Journal of Clinical Microbiology ◽

10.1128/jcm.01409-17 ◽

2018 ◽

Vol 56 (4) ◽

Cited By ~ 13

Author(s):

Lorraine Eriksson ◽

Sara Thulin Hedberg ◽

Susanne Jacobsson ◽

Hans Fredlund ◽

Paula Mölling ◽

...

Keyword(s):

Neisseria Meningitidis ◽

Whole Genome Sequencing ◽

Genome Sequencing ◽

Clonal Complex ◽

Invasive Disease ◽

South American ◽

Whole Genome ◽

The Novel ◽

Content Type ◽

Low Incidence

ABSTRACT Invasive disease caused by Neisseria meningitidis serogroup W (MenW) has historically had a low incidence in Sweden, with an average incidence of 0.03 case/100,000 population from 1995 to 2014. In recent years, a significant increase in the incidence of MenW has been noted in Sweden, to an average incidence of 0.15 case/100,000 population in 2015 to 2016. In 2017 (1 January to 30 June), 33% of invasive meningococcal disease cases (7/21 cases) were caused by MenW. In the present study, all invasive MenW isolates from Sweden collected in 1995 to June 2017 ( n = 86) were subjected to whole-genome sequencing to determine the population structure and to compare isolates from Sweden with historical and international cases. The increase of MenW in Sweden was determined to be due to isolates belonging to the South American sublineage of MenW clonal complex 11, namely, the novel U.K. 2013 lineage. This lineage was introduced in Sweden in 2013 and has since been the dominant lineage of MenW.

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ISEcp1-Mediated Transposition Leads to Fosfomycin and Broad-Spectrum Cephalosporin Resistance in Klebsiella pneumoniae

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.00150-20 ◽

2020 ◽

Vol 64 (5) ◽

Author(s):

Nicolas Kieffer ◽

Laurent Poirel ◽

Linda Mueller ◽

Stefano Mancini ◽

Patrice Nordmann

Keyword(s):

Klebsiella Pneumoniae ◽

Whole Genome Sequencing ◽

Genome Sequencing ◽

Broad Spectrum ◽

Whole Genome ◽

Genetic Event ◽

Content Type ◽

Hybrid Promoter ◽

Cephalosporin Resistance ◽

Fosfomycin Resistance

ABSTRACT A fosfomycin-resistant and carbapenemase (OXA-48)-producing Klebsiella pneumoniae isolate was recovered, and whole-genome sequencing revealed ISEcp1-blaCTX-M-14b tandemly inserted upstream of the chromosomally encoded lysR-fosA locus. Quantitative evaluation of the expression of lysR and fosA genes showed that this insertion brought a strong hybrid promoter leading to overexpression of the fosA gene, resulting in fosfomycin resistance. This work showed the concomitant acquisition of resistance to broad-spectrum cephalosporins and fosfomycin due to a single genetic event.

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Complete Genome Sequence of the Temperate Klebsiella pneumoniae Phage KPP5665-2

Genome Announcements ◽

10.1128/genomea.01118-17 ◽

2017 ◽

Vol 5 (43) ◽

Cited By ~ 1

Author(s):

Gaby Carl ◽

Claudia Jäckel ◽

Josephine Grützke ◽

Stefan Hertwig ◽

Mirjam Grobbel ◽

...

Keyword(s):

Klebsiella Pneumoniae ◽

Whole Genome Sequencing ◽

Genome Sequencing ◽

Genome Sequence ◽

Complete Genome Sequence ◽

Complete Genome ◽

Whole Genome ◽

The Novel ◽

Content Type ◽

Narrow Host Range

ABSTRACT We describe here the genome sequence of the novel temperate Klebsiella pneumoniae phage KPP5665-2 isolated from a Klebsiella pneumoniae strain recovered from milk in Germany in 2016. The phage exhibited a narrow host range and a siphoviridal morphology. KPP5665-2-related prophage sequences were detected in whole-genome sequencing (WGS) data of various Klebsiella species isolates.

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