Klebsiella pneumoniae: adaptive immune landscapes and vaccine horizons.

Klebsiella pneumoniae is among the most common antibiotic-resistant pathogens causing nosocomial infections. Additionally, it is a leading cause of neonatal sepsis and childhood mortality across the globe. Despite its clinical importance, we are only beginning to understand how the mammalian adaptive immune system responds to this pathogen. Further, many studies investigating potential K. pneumoniae vaccine candidates or alternative therapies have been launched in recent years. Here, we review the current state of knowledge on the adaptive immune response to K. pneumoniae infections and progress towards developing vaccines and other therapies to combat these infections.

Identification of a second glycoform of the clinically prevalent O1 antigen from Klebsiella pneumoniae.

Carbapenemase and extended ß-lactamase-producing Klebsiella pneumoniae isolates represent a major health threat, stimulating increasing interest in immunotherapeutic approaches for combating Klebsiella infections. Lipopolysaccharide O antigen polysaccharides offer viable targets for immunotherapeutic development, and several studies have described protection with O-specific antibodies in animal models of infection. O1 antigen is produced by almost half of clinical Klebsiella isolates. The O1 polysaccharide backbone structure is known, but monoclonal antibodies raised against the O1 antigen showed varying reactivity against different isolates that could not be explained by the known structure. Reinvestigation of the structure by NMR spectroscopy revealed the presence of the reported polysaccharide backbone (glycoform O1a), as well as a previously unknown O1b glycoform composed of the O1a backbone modified with a terminal pyruvate group. The activity of the responsible pyruvyltransferase (WbbZ) was confirmed by western immunoblotting and in vitro chemoenzymatic synthesis of the O1b terminus. Bioinformatic data indicate that almost all O1 isolates possess genes required to produce both glycoforms. We describe the presence of O1ab-biosynthesis genes in other bacterial species and report a functional O1 locus on a bacteriophage genome. Homologs of wbbZ are widespread in genetic loci for the assembly of unrelated glycostructures in bacteria and yeast. In K. pneumoniae, simultaneous production of both O1 glycoforms is enabled by the lack of specificity of the ABC transporter that exports the nascent glycan, and the data reported here provide mechanistic understanding of the capacity for evolution of antigenic diversity within an important class of biomolecules produced by many bacteria.

Capsular polysaccharide inhibits vaccine-induced O-antigen antibody binding and function across both classical and hypervirulent K2:O1 strains of Klebsiella pneumoniae.

Klebsiella pneumoniae presents as two circulating pathotypes: classical K. pneumoniae (cKp) and hypervirulent K. pneumoniae (hvKp). Classical isolates are considered urgent threats due to their antibiotic resistance profiles, while hvKp isolates have historically been antibiotic susceptible. Recently, however, increased rates of antibiotic resistance have been observed in both hvKp and cKp, further underscoring the need for preventive and effective immunotherapies. Two distinct surface polysaccharides have gained traction as vaccine candidates against K. pneumoniae: capsular polysaccharide and the O-antigen of lipopolysaccharide. While both targets have practical advantages and disadvantages, it remains unclear which of these antigens included in a vaccine would provide superior protection against matched K. pneumoniae strains. Here, we report the production of two bioconjugate vaccines, one targeting the K2 capsular serotype and the other targeting the O1 O-antigen. Using murine models, we investigated whether these vaccines induced specific antibody responses that recognize K2:O1 K. pneumoniae strains. While each vaccine was immunogenic in mice, both cKp and hvKp strains exhibited decreased O-antibody binding in the presence of capsule. Further, O1 antibodies demonstrated decreased killing in serum bactericidal assays with encapsulated strains, suggesting that the presence of K. pneumoniae capsule blocks O1-antibody binding and function. Finally, the K2 vaccine outperformed the O1 vaccine against both cKp and hvKp in two different murine infection models. These data suggest that capsule-based vaccines may be superior to O-antigen vaccines for targeting hvKp and some cKp strains, due to capsule blocking the O-antigen.

Heptavalent O-Antigen Bioconjugate Vaccine Exhibiting Differential Functional Antibody Responses Against Diverse Klebsiella pneumoniae Isolates.

Klebsiella pneumoniae is the leading cause of neonatal sepsis and is increasingly difficult to treat owing to antibiotic resistance. Vaccination represents a tractable approach to combat this resistant bacterium; however, there is currently not a licensed vaccine. Surface polysaccharides, including O-antigens of lipopolysaccharide, have long been attractive candidates for vaccine inclusion. Herein we describe the generation of a bioconjugate vaccine targeting 7 predominant O-antigen subtypes in K. pneumoniae. Each bioconjugate was immunogenic in isolation, with limited cross-reactivity among subtypes. Vaccine-induced antibodies demonstrated varying degrees of binding to a wide variety of K. pneumoniae strains. Furthermore, serum from vaccinated mice induced complement-mediated killing of many of these strains. Finally, increased capsule interfered with the ability of O-antigen antibodies to bind and mediate killing of some K. pneumoniae strains. Taken together, these data indicate that this novel heptavalent O-antigen bioconjugate vaccine formulation exhibits limited efficacy against some, but not all, K. pneumoniae isolates.

A novel chimeric vaccine containing multiple epitopes for simulating robust immune activation against Klebsiella pneumoniae.

BACKGROUND: Due to antibiotic resistance, the Klebsiella genus is linked to morbidity and death, necessitating the development of a universally protective vaccine against Klebsiella pathogens. METHODS: Core sequence analysis prioritized non-redundant host molecules and expected lipid bilayer peptides from fully sequenced Klebsiella genomes. These proteins were refined to identify epitopes, examining their immunogenicity, toxicity, solubility, and interaction with MHC alleles. Epitopes were linked to CPG ODN C274 via EAAAK, HEYGAEALERAG, and GGGS linkers to enhance immunological responses. The vaccine's tertiary structure was modelled and docked with MHC-I and MHC-II. RESULTS: Fifty-five proteins were recognized in the Vaxign collection as having remarkable features. Twenty-three proteins with potential pathogenicity were then identified. Eight options for vaccines emerged after the immunogenicity of proteins was examined. The best antigens were three proteins: MrkD, Iron-regulated lipid membrane polypeptides, and RmpA. These compounds were selected for their sensitivity. The structural protein sequences of K. pneumoniae were utilized to identify seven CTL epitopes, seven HTL epitopes, and seven LBL epitopes, respectively. The produced immunization displayed a stable contact with the receptors, based on molecular dynamic simulations lasting 250 nanoseconds. Intermolecular binding free energies also indicated the dominance of the van der Waals and electrostatic energies. CONCLUSION: In summary, the results of this study might help scientists develop a novel vaccine to prevent K. pneumoniae infections.

Oral administration of Lactobacillus plantarum expressing aCD11c modulates cellular immunity alleviating inflammatory injury due to Klebsiella pneumoniae infection.

BACKGROUND: Klebsiella pneumoniae (KP), responsible for acute lung injury (ALI) and inflammation of the gastrointestinal tract, is a zoonotic pathogen that poses a threat to livestock farming worldwide. Nevertheless, there is currently no validated vaccine to prevent KP infection. The development of mucosal vaccines against KP using Lactobacillus plantarum (L. plantarum) is an effective strategy. RESULTS: Firstly, the L. plantarum strains NC8-pSIP409-aCD11c' and NC8-pLc23-aCD11c were constructed via homologous recombination to express the aCD11c protein either inducibly or constitutively. Both NC8-pSIP409-aCD11c' and NC8-pLc23-aCD11c strains could enhance the adhesion and invasion of L. plantarum on bone marrow-derived dendritic cells (BMDCs), and stimulate the activation of BMDCs compared to the control strain NC8-pSIP409 in vitro. Following oral immunization of mice with NC8-pSIP409-aCD11c' and NC8-pLc23-aCD11c, the cellular, humoral, and mucosal immunity were significantly improved, as evidenced by the increased expression of CD4+ IL-4+ T cells in the spleen, IgG in serum, and secretory IgA (sIgA) in the intestinal lavage fluid (ILF). Furthermore, the protective effects of L. plantarum against inflammatory damage caused by KP infection were confirmed by assessing the bacterial loads in various tissues, lung wet/dry ratio (W/D), levels of inflammatory cytokines, and histological evaluation, which influenced T helper 17 (Th17) and regulatory T (Treg) cells in peripheral blood and lung. CONCLUSIONS: Both the inducible and constitutive L. plantarum strains NC8-pSIP409-aCD11c' and NC8-pLc23-aCD11c have been found to stimulate cellular and humoral immunity levels and alleviate the inflammatory response caused by KP infection. These findings have provided a basis for the development of a novel vaccine against KP.

Deep Intraclonal Analysis for the Development of Vaccines against Drug-Resistant Klebsiella pneumoniae Lineages.

Despite its medical relevance, there is no commercial vaccine that protects the population at risk from multidrug-resistant (MDR) Klebsiella pneumoniae infections. The availability of massive omic data and novel algorithms may improve antigen selection to develop effective prophylactic strategies. Up to 133 exposed proteins in the core proteomes, between 516 and 8666 genome samples, of the six most relevant MDR clonal groups (CGs) carried conserved B-cell epitopes, suggesting minimized future evasion if utilized for vaccination. Antigens showed a range of epitopicity, functional constraints, and potential side effects. Eleven antigens, including three sugar porins, were represented in all MDR-CGs, constitutively expressed, and showed limited reactivity with gut microbiota. Some of these antigens had important interactomic interactions and may elicit adhesion-neutralizing antibodies. Synergistic bivalent to pentavalent combinations that address expression conditions, interactome location, virulence activities, and clone-specific proteins may overcome the limiting protection of univalent vaccines. The combination of five central antigens accounted for 41% of all non-redundant interacting partners of the antigen dataset. Specific antigen mixtures represented in a few or just one MDR-CG further reduced the chance of microbiota interference. Rational antigen selection schemes facilitate the design of high-coverage and "magic bullet" multivalent vaccines against recalcitrant K. pneumoniae lineages.

Case Commentary: Unlocking the Potential of Bacteriophage to Prevent Recurrent Urinary Tract Infections after Kidney Transplantation.

Recurrent urinary tract infections (UTIs) are common in kidney transplant recipients, and novel prevention approaches are needed. The case presented by Le et al. (Antimicrob Agents Chemother, in press) describes a patient with recurrent UTIs due to extended-spectrum ß-lactamase-producing Klebsiella pneumoniae who was successfully treated with bacteriophage therapy. This commentary highlights the potential for bacteriophage therapy to prevent recurrent UTIs, as well as outstanding questions that require further investigation.

Klebsiella pneumoniae vaccine studies in animal models.

The treatment of Klebsiella pneumoniae is faced with challenges demanding the development of a vaccination strategy. However, no approved and globally available vaccine exists yet. This study aimed to systematically review all published data on K. pneumoniae vaccines in animal models. Without time restrictions, electronic databases were searched using appropriate keywords. The retrieved studies were screened and the data of those that matched our inclusion criteria were collected and analyzed. In total, 2027 records were retrieved; of which 35 studies were included for systematic review. The most frequently used animal model was BALB/c mice. Proteins, polysaccharides, and their combinations (conjugates) were the most common vaccine candidates used. The amount of antigen, the route used for immunization, and the challenge strategy was varying in the studies and were chosen based on several factors such as the animal model, the type of antigen, and the schedule of immunization. Almost all studies claimed that their vaccine was effective/protective, indicated by increasing survival rate, reducing organ bacterial load, and eliciting protective antibody and/or cytokine responses. Altogether, the information presented here will assist researchers to have a better look at the K. pneumoniae vaccine candidates and to take more effective steps in the future.

Identifying the drivers of multidrug-resistant Klebsiella pneumoniae at a European level.

Beta-lactam- and in particular carbapenem-resistant Enterobacteriaceae represent a major public health threat. Despite strong variation of resistance across geographical settings, there is limited understanding of the underlying drivers. To assess these drivers, we developed a transmission model of cephalosporin- and carbapenem-resistant Klebsiella pneumoniae. The model is parameterized using antibiotic consumption and demographic data from eleven European countries and fitted to the resistance rates for Klebsiella pneumoniae for these settings. The impact of potential drivers of resistance is then assessed in counterfactual analyses. Based on reported consumption data, the model could simultaneously fit the prevalence of extended-spectrum beta-lactamase-producing and carbapenem-resistant Klebsiella pneumoniae (ESBL and CRK) across eleven European countries over eleven years. The fit could explain the large between-country variability of resistance in terms of consumption patterns and fitted differences in hospital transmission rates. Based on this fit, a counterfactual analysis found that reducing nosocomial transmission and antibiotic consumption in the hospital had the strongest impact on ESBL and CRK prevalence. Antibiotic consumption in the community also affected ESBL prevalence but its relative impact was weaker than inpatient consumption. Finally, we used the model to estimate a moderate fitness cost of CRK and ESBL at the population level. This work highlights the disproportionate role of antibiotic consumption in the hospital and of nosocomial transmission for resistance in gram-negative bacteria at a European level. This indicates that infection control and antibiotic stewardship measures should play a major role in limiting resistance even at the national or regional level.

The impact of public health interventions on the future prevalence of ESBL-producing Klebsiella pneumoniae: a population based mathematical modelling study.

BACKGROUND: Future prevalence of colonization with extended-spectrum betalactamase (ESBL-) producing K. pneumoniae in humans and the potential of public health interventions against the spread of these resistant bacteria remain uncertain. METHODS: Based on antimicrobial consumption and susceptibility data recorded during > 13 years in a Swiss region, we developed a mathematical model to assess the comparative effect of different interventions on the prevalence of colonization. RESULTS: Simulated prevalence stabilized in the near future when rates of antimicrobial consumption and in-hospital transmission were assumed to remain stable (2025 prevalence: 6.8% (95CI%:5.4-8.8%) in hospitals, 3.5% (2.5-5.0%) in the community versus 6.1% (5.0-7.5%) and 3.2% (2.3-4.2%) in 2019, respectively). When overall antimicrobial consumption was set to decrease by 50%, 2025 prevalence declined by 75% in hospitals and by 64% in the community. A 50% decline in in-hospital transmission rate led to a reduction in 2025 prevalence of 31% in hospitals and no reduction in the community. The best model fit estimated that 49% (6-100%) of observed colonizations could be attributable to sources other than human-to-human transmission within the geographical setting. CONCLUSIONS: Projections suggests that overall antimicrobial consumption will be, by far, the most powerful driver of prevalence and that a large fraction of colonizations could be attributed to non-local transmissions.

Preparation and evaluation of a new combined conjugated vaccine against Klebsiella pneumonia and Pseudomonas aeruginosa.

AIMS: Lower respiratory tract infections (LRTIs) have been identified by the World Health Organization as the most deadly infectious diseases and a pervasive public health problem, causing increased hospital admissions, mortality and antibiotic use. This study aims to determine the most common and resistant bacteria that cause LRTIs and prepare an appropriate vaccine to reduce and prevent potential future infections. METHODS AND RESULTS: Our survey was conducted by collecting respiratory exudate specimens. The most predominant and resistant types were Klebsiella pneumonia and Pseudomonas aeruginosa. The lipopolysaccharides (LPS) were extracted using a modified hot phenol method to prepare the vaccine. The LPS were then activated and conjugated. The immunogenicity of the prepared singles and combined vaccines was determined through an in vivo assay using BALB/c mice. The prepared vaccine provided high protection against the lethal dose of both bacteria in mice. The combined vaccine shows a significant value in achieving high immunization. CONCLUSION: These findings demonstrate the potential of the bacterial LPS molecules to be used as effective vaccines. SIGNIFICANCE AND IMPACT OF STUDY: Developing an effective single and combined vaccine against P. aeruginosa and K. pneumonia can protect and reduce LRTI incidence.

A promising bioconjugate vaccine against hypervirulent Klebsiella pneumoniae.

Hypervirulent Klebsiella pneumoniae (hvKp) is globally disseminating as a community-acquired pathogen causing life-threatening infections in healthy individuals. The fact that a dose as little as 50 bacteria is lethal to mice illustrates the dramatic increase of virulence associated with hvKp strains compared with classical K. pneumoniae (cKp) strains, which require lethal doses greater than 107 bacteria. Until recently, these virulent strains were mostly antibiotic-susceptible. However, multidrug-resistant (MDR) hvKp strains have been emerging, spawning a new generation of hypervirulent "superbugs." The mechanisms of hypervirulence are not fully defined, but overproduction of capsular polysaccharide significantly impedes host clearance, resulting in increased pathogenicity of hvKp strains. While there are more than 80 serotypes of K. pneumoniae, the K1 and K2 serotypes cause the vast majority of hypervirulent infections. Therefore, a glycoconjugate vaccine targeting these 2 serotypes could significantly reduce hvKp infection. Conventionally, glycoconjugate vaccines are manufactured using intricate chemical methodologies to covalently attach purified polysaccharides to carrier proteins, which is widely considered to be technically challenging. Here we report on the recombinant production and analytical characterization of bioconjugate vaccines, enzymatically produced in glycoengineered Escherichia coli cells, against the 2 predominant hypervirulent K. pneumoniae serotypes, K1 and K2. The K. pneumoniae bioconjugates are immunogenic and efficacious, protecting mice against lethal infection from 2 hvKp strains, NTUH K-2044 and ATCC 43816. This preclinical study constitutes a key step toward preventing further global dissemination of hypervirulent MDR hvKp strains.

Transmission of Klebsiella pneumoniae carbapenemase (KPC)-producing Klebsiella pneumoniae: the role of infection control.

BACKGROUND: The worldwide spread of carbapenemase-producing Gram-negative bacteria (GNB) in healthcare settings is worrying. Of particular concern is the occurrence of Klebsiella pneumoniae carbapenemase (KPC)-producing Klebsiella pneumoniae (KP). In recent years, several guidelines and recommendations have focused on the control of carbapenem-resistant GNB. It remains, however, unknown to what extent individual infection control measures are effective. Our aim was to critically review the recent evidence regarding the effectiveness of measures to control KPC-KP spread in healthcare settings. METHODS: Critical review of the literature aiming to evaluate, in accordance with published recommendations, all available studies reporting infection control (IC) measures to control KPC-KP published in the past 5 years. RESULTS: Among 11 included studies, the majority consisted of outbreak reports, where application of measures was reported in the absence of control groups. Variability was observed related to the frequency of application of recommended measures for control of KPC-KP. Additional measures were recorded, such as environmental sampling and staff screening, whereas compliance with hand hygiene was measured in relatively few studies. Finally, mortality in patients harbouring KPC-KP was notable, reaching 42.9% of included patients. CONCLUSIONS: Despite successful control of KPC-KP spread during outbreaks, the impact of individual IC measures is difficult to assess, as the quality of published evidence is low and controlled intervention studies are lacking. The distribution of studies, the number of reported cases and the high mortality rates, clearly show that KPC-KP remains a major healthcare problem worldwide.

The role of antimicrobial stewardship in preventing KPC-producing Klebsiella pneumoniae.

Antimicrobial stewardship programmes are widely considered to be a core component of the response to the antimicrobial resistance threat. However, a positive impact of these interventions in terms of microbiological outcomes remains difficult to demonstrate, especially when focusing on specific resistant phenotypes. The first part of this review aims to explore the complex relationship between antibiotic exposure and resistance development in KPC-producing Klebsiella pneumoniae. In the second part we aim to summarize published examples of antimicrobial stewardship interventions intended to impact on the epidemiology of KPC-producing K. pneumoniae. For this purpose, a literature search was performed and seven studies were included in the review. Both restrictive and non-restrictive interventions were associated with an overall reduction in antibiotic consumption, and a decrease in carbapenem resistance rates was observed in five studies. The overall quality of the evidence was low, mainly due to the poor reporting of microbiological outcomes, lack of a control group and suboptimal study design. Although the link between antibiotic use and resistance development is supported by strong evidence, demonstrating the impact of antimicrobial stewardship interventions on microbiological outcomes remains difficult. Studies with adequate design and appropriate outcome measures are needed to further promote antimicrobial stewardship and elucidate which interventions are more successful for controlling the spread of KPC-producing K. pneumoniae.

Evaluation of riboflavin and ultraviolet light treatment against Klebsiella pneumoniae in whole blood-derived platelets: A pilot study.

BACKGROUND: Bacterial contamination of platelet concentrates (PCs) is the predominant cause of infectious transfusion reactions. The Pathogen Inactivation Mirasol system was implemented at the King Faisal Specialist Hospital (Saudi Arabia) to reduce the risk of transfusing contaminated PCs. This pilot study evaluated the effectiveness of Mirasol against Klebsiella pneumoniae, a pathogen associated with transfusion reactions, in whole blood-derived PCs. STUDY DESIGN AND METHODS: Whole blood (WB) units inoculated with one of six K. pneumoniae strains (five clinical isolates and ATCC-700603) at a concentration of 3-38 CFU/unit, were processed using the platelet-rich plasma (PRP) method. Each spiked PC was pooled with four unspiked units. The pooled PC was split into three Mirasol storage bags: an untreated unit (control), and two units treated with Mirasol at 26 and 32 h post-WB collection, respectively. PC samples obtained before and after Mirasol treatment were used for BacT/ALERT cultures and determination of bacteria quantification. Each experiment was repeated three independent times. RESULTS: Five strains were detected prior to PC treatment (24 h post-WB spiking), while one clinical isolate was not detected. Mirasol treatment after 26 h of WB collection resulted in complete inactivation of all K. pneumoniae strains. However, treatment 32 h post-WB collection resulted in the breakthrough of one clinical isolate in two of the three replicates with ~7.8 log10 CFU/unit detected on day 5 of PC storage. CONCLUSION: Delayed Mirasol treatment from 26 to 32 h post-WB collection, resulted in one breakthrough. These results highlight the importance of minimizing the time between WB collection and PI treatment.

Prekallikrein inhibits innate immune signaling in the lung and impairs host defense during pneumosepsis in mice.

Prekallikrein (PKK, also known as Fletcher factor and encoded by the gene KLKB1 in humans) is a component of the contact system. Activation of the contact system has been implicated in lethality in fulminant sepsis models. Pneumonia is the most frequent cause of sepsis. We sought to determine the role of PKK in host defense during pneumosepsis. To this end, mice were infected with the common human pathogen Klebsiella pneumoniae via the airways, causing an initially localized infection of the lungs with subsequent bacterial dissemination and sepsis. Mice were treated with a selective PKK-directed antisense oligonucleotide (ASO) or a scrambled control ASO for 3 weeks prior to infection. Host response readouts were determined at 12 or 36 h post-infection, including genome-wide messenger RNA profiling of lungs, or mice were followed for survival. PKK ASO treatment inhibited constitutive hepatic Klkb1 mRNA expression by >80% and almost completely abolished plasma PKK activity. Klkb1 mRNA could not be detected in lungs. Pneumonia was associated with a progressive decline in PKK expression in mice treated with control ASO. PKK ASO administration was associated with a delayed mortality, reduced bacterial burdens, and diminished distant organ injury. While PKK depletion did not influence lung pathology or neutrophil recruitment, it was associated with an upregulation of multiple innate immune signaling pathways in the lungs already prior to infection. Activation of the contact system could not be detected, either during infection in vivo or at the surface of Klebsiella in vitro. These data suggest that circulating PKK confines pro-inflammatory signaling in the lung by a mechanism that does not involve contact system activation, which in the case of respiratory tract infection may impede early protective innate immunity. © 2019 Authors. Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland.

Synergistic Action of Antimicrobial Lung Proteins against Klebsiella pneumoniae.

As key components of innate immunity, lung antimicrobial proteins play a critical role in warding off invading respiratory pathogens. Lung surfactant protein A (SP-A) exerts synergistic antimicrobial activity with the N-terminal segment of the SP-B proprotein (SP-BN) against Klebsiella pneumoniae K2 in vivo. However, the factors that govern SP-A/SP-BN antimicrobial activity are still unclear. The aim of this study was to identify the mechanisms by which SP-A and SP-BN act synergistically against K. pneumoniae, which is resistant to either protein alone. The effect of these proteins on K. pneumoniae was studied by membrane permeabilization and depolarization assays and transmission electron microscopy. Their effects on model membranes of the outer and inner bacterial membranes were analyzed by differential scanning calorimetry and membrane leakage assays. Our results indicate that the SP-A/SP-BN complex alters the ultrastructure of K. pneumoniae by binding to lipopolysaccharide molecules present in the outer membrane, forming packing defects in the membrane that may favor the translocation of both proteins to the periplasmic space. The SP-A/SP-BN complex depolarized and permeabilized the inner membrane, perhaps through the induction of toroidal pores. We conclude that the synergistic antimicrobial activity of SP-A/SP-BN is based on the capability of this complex, but not either protein alone, to alter the integrity of bacterial membranes.

Handwashing Sink Contamination and Carbapenem-resistant Klebsiella Infection in the Intensive Care Unit: A Prospective Multicenter Study.

BACKGROUND: Handwashing sinks can become contaminated by carbapenem-resistant Klebsiella (CRK), including carbapenem-resistant Klebsiella pneumoniae (CRKP) and carbapenem-resistant Klebsiella oxytoca (CRKO), but whether they are major sources of CRK infections remains unknown. METHODS: We performed a prospective multicenter study in 16 intensive care units (ICUs) (9 general and 7 neonatal) at 11 hospitals. All sinks at these locations were sampled to screen CRK. All CRK clinical isolates recovered between 2 weeks before and 3 months after sampling in ICUs with CRK-positive sinks or other participating ICUs at the same hospital were collected. Whole-genome sequencing of all isolates was performed. Isolates of the same sequence type (ST) were assigned to clones by calling single-nucleotide polymorphisms. RESULTS: Among 158 sinks sampled, 6 CRKP and 6 CRKO were recovered from 12 sinks in 7 ICUs, corresponding to a 7.6% CRK contamination rate. Twenty-eight clinical isolates were collected, and all were CRKP. The 34 CRKP isolates belonged to 7 STs, including ST789 (n = 14, all had blaNDM-5); ST11 (n = 12, 5 belonged to KL64 and 7 to KL47, all had blaKPC-2); ST709 (n = 4, all had blaNDM-5); and ST16, ST20, ST1027, and ST2407 (n = 1 each). One particular ST789 clone caused an outbreak and contaminated a sink. ST11_KL47 sink isolates were likely the source of a cluster of clinical isolates. Two ST11_KL64 isolates belonged to a common clone but were from 2 hospitals. CONCLUSIONS: Contaminated sinks were not the major source of CRK in our local settings. ST789 blaNDM-5-carrying CRKP might represent an emerging lineage causing neonatal infections.

Novel Subclone of Carbapenem-Resistant Klebsiella pneumoniae Sequence Type 11 with Enhanced Virulence and Transmissibility, China.

We aimed to clarify the epidemiologic and clinical importance of evolutionary events that occurred in carbapenem-resistant Klebsiella pneumoniae (CRKP). We collected 203 CRKP causing bloodstream infections in a tertiary hospital in China during 2013-2017. We detected a subclonal shift in the dominant clone sequence type (ST) 11 CRKP in which the previously prevalent capsular loci (KL) 47 had been replaced by KL64 since 2016. Patients infected with ST11-KL64 CRKP had a significantly higher 30-day mortality rate than other CRKP-infected patients. Enhanced virulence was further evidenced by phenotypic tests. Phylogenetic reconstruction demonstrated that ST11-KL64 is derived from an ST11-KL47-like ancestor through recombination. We identified a pLVPK-like virulence plasmid carrying rmpA and peg-344 in ST11-KL64 exclusively from 2016 onward. The pLVPK-like-positive ST11-KL64 isolates exhibited enhanced environmental survival. Retrospective screening of a national collection identified ST11-KL64 in multiple regions. Targeted surveillance of this high-risk CRKP clone is urgently needed.