Bacteriology

Population Structure of Klebsiella pneumoniae Causing Bloodstream Infections at a New York City Tertiary Care Hospital: Diversification of Multidrug-Resistant Isolates

Angela Gomez-Simmonds, Michelle Greenman, Sean B. Sullivan, Joshua P. Tanner, Madeleine G. Sowash, Susan Whittier, Anne-Catrin Uhlemann
K. C. Carroll, Editor
DOI: 10.1128/JCM.03455-14

ABSTRACT

Despite the growing importance of carbapenem-resistant Klebsiella pneumoniae (CRKP), the clonal relationships between CRKP and antibiotic-susceptible isolates remain unclear. We compared the genetic diversity and clinical features of CRKP, third-generation and/or fourth-generation cephalosporin-resistant (Ceph-R) K. pneumoniae, and susceptible K. pneumoniae isolates causing bloodstream infections at a tertiary care hospital in New York City between January 2012 and July 2013. Drug susceptibilities were determined with the Vitek 2 system. Isolates underwent multilocus sequence typing and PCR sequencing of the wzi and bla KPC genes. Clinical and microbiological data were extracted from patient records and correlated with molecular data. Among 223 patients, we identified 272 isolates. Of these, 194 were susceptible, 30 Ceph-R, and 48 CRKP, belonging to 144 sequence types (STs). Susceptible (127 STs) and Ceph-R (20 STs) isolates were highly diverse. ST258 dominated CRKP strains (12 STs, with 63% ST258). There was minimal overlap in STs between resistance groups. The bla KPC-3 gene (30%) was restricted to ST258/wzi154, whereas bla KPC-2 (70%) was observed for several wzi allele types. CRKP infections occurred more frequently among solid organ transplant (31%) and dialysis (17%) patients. Mortality rates were high overall (28%) and highest among CRKP-infected patients (59%). In multivariable analyses, advanced age, comorbidities, and disease severity were significant predictors of 30-day mortality rates, whereas the K. pneumoniae susceptibility phenotype was not. Among CRKP infections, we observed a borderline significant association of increased mortality rates with ST258 and the wzi154 allele. Although the clonal spread of ST258 continues to contribute substantially to the dissemination of CRKP, non-ST258 strains appear to be evolving. Further investigations into the mechanisms promoting CRKP diversification and the effects of clonal backgrounds on outcomes are warranted.

INTRODUCTION

Infections due to carbapenem-resistant Enterobacteriaceae have emerged as an important public health problem over the past decade and are now considered an urgent antibiotic-resistant threat by the Centers for Disease Control and Prevention (CDC), the category of greatest concern (1). In the United States, carbapenem resistance among Enterobacteriaceae is primarily attributable to the production of the Klebsiella pneumoniae carbapenemase (KPC) (2, 3), which is plasmid mediated and most commonly encountered in K. pneumoniae in nosocomial settings (4, 5). Bloodstream infections (BSIs) caused by carbapenem-resistant K. pneumoniae (CRKP) are associated with particularly high mortality rates (6–8), with previous studies reporting hospital mortality rates of 40 to 70%, compared to rates of 20 to 30% among matched patients with bacteremia due to susceptible K. pneumoniae (4, 9).

A single clone, multilocus sequence type 258 (ST258), has been found to account for the majority of CRKP infections in the United States and was identified among 70% of isolates sent to the CDC for resistance testing between 1996 and 2008 (2). More recently, whole-genome sequencing revealed that ST258 includes at least two distinct genetic lineages (10), highlighting the ongoing evolution and adaptation of this strain. Its divergence was based on differences in a hypervariable region encoding a K. pneumoniae capsule component, as well as the presence of blaKPC-2 or blaKPC-3. In a separate investigation, ST258 harboring blaKPC-3 was exclusively associated with wzi allele type wzi154, which corresponds to the novel capsule type C200 (11). This KPC-3-expressing strain was found to have decreased virulence in a Galleria mellonella (waxworm) model, compared to strains carrying blaKPC-2.

Despite this close association between blaKPC variants and ST258 sublineages, several studies have demonstrated that the Tn4401 transposon known to carry the blaKPC gene can be found on a variety of plasmids that are transferrable between Enterobacteriaceae (12–14), although the nature of recipient strains has not been well defined. While most carbapenem resistance in K. pneumoniae is plasmid mediated, alternative mechanisms, such as the presence of porin mutations in isolates that harbor extended-spectrum beta-lactamases (ESBLs), have also been described (15). However, given that most hospital epidemiological investigations focus on identifying direct transmission events involving single clones, the contributions of plasmid-mediated horizontal gene transfer and other mechanisms to the CRKP epidemic remain largely unknown.

To better understand the interactions between susceptible and drug-resistant K. pneumoniae populations, we investigated the molecular epidemiology of K. pneumoniae BSIs at a tertiary care hospital in New York City. We assessed molecular variables in order to identify isolate characteristics (including sequence and capsule types) associated with the acquisition of multidrug resistance determinants. We also evaluated host factors associated with third-generation and/or fourth-generation cephalosporin-resistant (Ceph-R) and CRKP infections and the effects of molecular factors on outcomes. Understanding the isolate and host characteristics that promote the emergence and transmission of CRKP among hospitalized patients may inform the design of interventions that limit dissemination.

MATERIALS AND METHODS

Study isolates and chart review.This study was reviewed and approved by the institutional review board of Columbia University Medical Center. We retrospectively identified and retrieved all K. pneumoniae isolates grown from blood cultures that had been systematically stored by the microbiology laboratory between January 2012 and July 2013. The microbiology laboratory processes all clinical specimens obtained from the Columbia University hospital system, which includes large academic teaching hospitals for adults and children and a smaller community hospital that serves as a referral center for local skilled nursing facilities. Repeat isolates collected from a single patient within a 14-day period were not available for analysis. We decided to include repeat isolates that were >2 weeks apart in our description of clonal diversity to account for some degree of within-patient heterogeneity.

Isolates were retrospectively matched to patient medical records, and clinical information was extracted by chart review. Detailed information on baseline comorbidities and disease severity at the time of the positive bloodstream culture was used to calculate Charlson comorbidity index scores (CCISs) and Pitt bacteremia scores (PBSs), respectively. Clinical outcomes were defined as (i) deaths within 30 days of the date of bacteremia and (ii) deaths during the index hospitalization. The presumed sources of infection were determined using CDC/National Healthcare Safety Network (NHSN) surveillance definitions (16) and then grouped into the following categories: central line associated, respiratory tract, intra-abdominal, soft tissue (including wounds), urinary tract, other infection, or unknown source of infection.

In addition, microbiology records were reviewed for results of antibiotic susceptibility testing, which was performed according to routine microbiology laboratory protocols using the Vitek 2 automated system, with additional E-testing as needed (bioMérieux, Durham, NC). Susceptibility breakpoints were derived from Clinical and Laboratory Standards Institute guidelines (17). CRKP isolates were defined by nonsusceptibility to any carbapenem, and Ceph-R isolates were nonsusceptible to ceftazidime, cefotaxime, ceftriaxone, or cefepime, in accordance with infection control guidelines at our institution. Susceptible K. pneumoniae isolates did not meet the criteria for multidrug-resistant (MDR) phenotypes (Ceph-R or CRKP). For patients from whom multiple isolates were collected, only the initial isolate susceptibility profiles were considered in assessments of clinical factors and outcomes and are reported in detail.

Sequence typing.Genetic relatedness among K. pneumoniae bloodstream isolates was determined using multilocus sequence typing (MLST), as described previously (18). Sequencing of the wzi gene locus was also performed for select isolates, including all Ceph-R and CRKP isolates and susceptible isolates that shared a ST with Ceph-R or CRKP isolates (18); wzi sequencing has been shown in previous studies to be a molecular method for rapid capsule serotyping and a proxy for K-typing (11). Unique allele and sequence type numbers were requested from the Institut Pasteur for clones that had not been reported previously.

All K. pneumoniae bloodstream isolates were screened for the presence of genes conferring carbapenem resistance, including blaKPC, blaVIM, blaIMP, and blaNDM, by PCR amplification using previously established primers and PCR parameters (19). For isolates found to harbor blaKPC, amplification followed by bidirectional DNA sequencing of the blaKPC PCR product was carried out in order to differentiate among blaKPC alleles (20).

Statistical analysis.Clinical parameters were described as frequencies or as mean ± standard deviation or median values, for normally or nonnormally distributed variables, respectively, as appropriate. For patients with multiple positive blood cultures, only the first available isolate was used in the assessments of clinical variables and outcomes. Univariable analyses were used to identify clinical and isolate characteristics associated with MDR infections and outcomes, using the χ2 test or Fisher's exact test for categorical variables and analysis of variance (ANOVA) or the Kruskal-Wallis test for continuously distributed variables, as appropriate. To further analyze factors associated with 30-day mortality rates, clinical variables with P values of < 0.1 were selected for inclusion in a parsimonious multivariable model using backward stepwise selection, to which a categorical resistance phenotype variable was added a priori. Similar multivariable models were constructed to assess the relationships between molecular variables selected a priori and 30-day mortality rates. In all analyses, P values of <0.05 were considered statistically significant. Data were analyzed using SAS 9.3 (SAS Institute Inc., Cary, NC).

RESULTS

Clinical characteristics of patients.During the 18-month study period, 223 patients with K. pneumoniae bloodstream infections were identified. Multiple isolates were collected for 29 patients, resulting in a total of 272 K. pneumoniae blood isolates. Of the 223 patients, 170 had susceptible, 24 Ceph-R, and 29 carbapenem-resistant K. pneumoniae infections, based on Vitek 2 testing of their initial bloodstream isolates. Patient demographic and clinical characteristics are delineated in Tables 1 and 2. Across all three groups, patients were noted to be elderly (median age, 62 years), to have multiple comorbid conditions (30% with CCISs of ≥4), and to have high rates of previous hospitalizations (61% within the previous 6 months).

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TABLE 1

Baseline characteristics of patients with K. pneumoniae bloodstream infections

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TABLE 2

Clinical characteristics of initial bloodstream infections

In univariable analyses, differences between the three groups were seen in underlying comorbidities (Table 1) and timing and risk factors for BSIs (Table 2). Patients who developed Ceph-R or CRKP BSIs were often chronically ill (42% had CCISs of ≥4). Large proportions of these patients were solid organ transplant (SOT) recipients (26%), had end-stage renal disease (ESRD) requiring chronic dialysis (13%), or had cirrhosis (13%). Many resided in long-term-care facilities (26%), although these patients accounted for a larger proportion of Ceph-R infections (29%) than CRKP infections (24%). The sources of infection did not differ significantly between patient groups.

Solid organ transplant (SOT) recipients accounted for the largest proportion of patients with CRKP infections (31%), followed by nursing home residents (24%) and patients with end-stage renal disease (17%) or cirrhosis (14%). Among SOT recipients, CRKP infections were most common among liver transplant patients (56% of CRKP BSIs occurred among SOT recipients), although at our center liver transplantation is performed only 30 to 40% more often than heart or lung transplantation and one-half as often as kidney transplantation (21). All patients with CRKP BSIs had been exposed to antibiotics within the past 6 months, and only 28% had received carbapenems. Patients with susceptible and Ceph-R infections often had initial positive blood cultures obtained at or soon after admission; CRKP bacteremia tended to develop later in the hospital course. Finally, at the time of their bacteremia, patients with CRKP infections were more likely to be severely ill (69% had PBSs of ≥4 on the date of the positive blood culture and 90% were in an intensive care unit [ICU]) than their counterparts with susceptible or Ceph-R isolates. Recent invasive surgery, central venous catheterization, Foley catheter use, and mechanical ventilation were correspondingly common in this group and might have served as additional risk factors for MDR infections.

Population structure of K. pneumoniae bloodstream isolates.We performed MLST on all 272 isolates and detected 144 different STs (Fig. 1a). Overall, ST258 was the predominant clone and was detected for 32 isolates (12%), followed by ST20 (n = 14), ST17 (n = 12), and ST37 (n = 10). There were 13 STs comprising 4 or more isolates, and a unique ST was identified for 108 isolates.

FIG 1
FIG 1

Multilocus sequence type and wzi allele type diversity in susceptible, third-generation and/or fourth-generation cephalosporin-resistant (Ceph-R), and carbapenem-resistant K. pneumoniae (CRKP) isolates. (a) ST diversity among all isolates, with the number of isolates belonging to each ST being represented by a section of the pie graph. (b) ST variability according to susceptibility phenotype. Among the 194 susceptible isolates, 127 different STs were identified, compared to 20 different STs among 30 Ceph-R isolates and only 10 STs among 48 CRKP isolates. (c) Numbers of STs shared by the 3 resistance groups, which were limited. (d and e) wzi allele type variability among Ceph-R and CRKP (d) and ST258 (e) isolates, with the number of isolates belonging to each allele type being depicted by a section of the pie graph. There were 18 wzi allele types identified among 30 Ceph-R isolates and 12 wzi allele types seen among 47 CRKP isolates. The 32 ST258 isolates were subdivided into six wzi allele types. Among Ceph-R and CRKP isolates, there were 4 shared wzi allele types.

We observed the highest number of STs among the 194 susceptible isolates (Fig. 1b). We identified 127 different STs, and most of those were singletons (99 STs). ST20 was the most common susceptible K. pneumoniae type, with 12 isolates (19%), followed by ST37 (n = 10) and ST45 (n = 6). Moreover, only 5 STs were responsible for 4 or more isolates. Among the 30 Ceph-R isolates, we detected 20 different STs, including 14 STs with a single isolate. We did not observe a dominant Ceph-R clone. The most frequently encountered ST, ST11, was noted for only 4 isolates. In contrast, we found that CRKP isolates (n = 48) were restricted to 10 STs. The majority of CRKP isolates were accounted for by ST258 (n = 30 [63%]). ST17 included 6 isolates (13%) and ST392 included 5 isolates (10%), whereas the remaining 7 isolates were singletons.

Few STs included isolates with differing susceptibility phenotypes (Fig. 1c). Only ST17 and ST35 included susceptible, Ceph-R, and CRKP isolates. Of 12 ST17 isolates, five were susceptible, one was Ceph-R, and six were CRKP; ST35 included one isolate from each group. Susceptible and Ceph-R isolates shared six additional STs, i.e., ST15, ST20, ST34, ST152, ST299, and ST919 (Fig. 1c). The majority of ST258 isolates were CRKP, but two Ceph-R isolates were also identified as ST258. ST392 included one Ceph-R isolate and 5 CRKP isolates. ST36 was the only ST to include isolates that were susceptible (n = 2) and CRKP (n = 1).

To further investigate the variability in capsule-encoding loci, we carried out wzi allele sequencing for all Ceph-R and CRKP isolates and for susceptible isolates that shared STs with Ceph-R or CRKP isolates. The locus exhibited high levels of diversity, with 34 different wzi allele types among 108 isolates, including seven new wzi alleles (Table 3). Among 30 Ceph-R isolates, 18 wzi alleles were identified; among 47 CRKP isolates (amplification of the gene locus was unsuccessful for one isolate), 12 wzi alleles were encountered (Fig. 1d). In some cases, mixing of wzi alleles and STs was seen. Several wzi alleles were represented across isolates of various susceptibilities (Table 3).

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TABLE 3

wzi allele type frequencies and associated sequence types and antibiotic susceptibilities

Within ST258, we encountered a variety of wzi alleles (Fig. 1e); wzi154 was most frequently observed (n = 16) and included almost all ST258 isolates harboring the blaKPC-3 gene (n = 12). This observation is consistent with a new variant described as C200 by Diago-Navarro and colleagues at another large medical center in New York City (11). The blaKPC-2-positive ST258 isolates primarily carried the wzi29 allele (n = 8) or wzi83 allele (n = 6). Conversely, all ST17 isolates, irrespective of their drug susceptibilities, carried the wzi141 allele.

Resistance gene identification.The presence of the blaKPC gene was detected by PCR amplification in all 49 CRKP isolates and no Ceph-R or susceptible K. pneumoniae isolates and was presumably responsible for the carbapenem resistance phenotype in the CRKP isolates. Sequencing of the full length of blaKPC, which was completed for 47 of the CRKP isolates, identified blaKPC-2 and blaKPC-3 in 70% and 30% of the isolates, respectively. No other blaKPC allele was detected. All ST17 and ST392 isolates carried the blaKPC-2 gene. Among ST258 isolates, 61% harbored blaKPC-2 and 39% carried blaKPC-3. With the exception of 3 isolates, all blaKPC-3 genes occurred in ST258 isolates (all except one positive for the wzi154 allele). Only 3 blaKPC-positive isolates had meropenem MICs of ≤8 mg/liter. Genes encoding other known carbapenemases, including blaVIM, blaIMP, and blaNDM, were not detected in any of the bloodstream isolates. Corresponding isolate susceptibility profiles for all initial Ceph-R and CRKP isolates collected from each patient are shown in Table 4.

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TABLE 4

In vitro susceptibilities of initial Ceph-R and CRKP bloodstream isolates

Serial bloodstream isolates.During the study period, 29 patients had multiple bloodstream isolates collected, including 22 susceptible isolates belonging to 18 different STs, eight Ceph-R isolates belonging to five STs, and 19 CRKP isolates belonging to six STs, including eight belonging to ST258 (25% of all ST258 isolates). For 19 of these patients, all collected isolates belonged to the same clonal background, whereas nine patients had isolates from more than one ST (Fig. 2). Among isolates with matching STs, no variability in antibiotic susceptibility was seen.

FIG 2
FIG 2

Clonal diversity of serial K. pneumoniae bloodstream infections in select patients. There were 9 patients with multiple positive blood cultures for whom collected isolates were of more than one sequence type (ST). Each bloodstream isolate is depicted by a rectangle. Susceptibility phenotypes are differentiated by color; gray bars, susceptible isolates; pink bars, Ceph-R isolates; green bars, CRKP isolates. Isolates of different STs that have the same susceptibility phenotype are differentiated by pattern but share the same color background. Thus, only patients 1, 2, and 9 have isolates of different susceptibility phenotypes and STs; all others have multiple isolates with the same resistance pattern but differing STs.

Among the nine patients who were infected with different K. pneumoniae clones, susceptibilities varied between isolates of different STs in only three patients (Fig. 2). In one patient, a CRKP infection occurred after bacteremia with a susceptible K. pneumoniae isolate; in two instances, a Ceph-R infection was followed by a CRKP infection. All of the other patients with serial BSIs had isolates of the same susceptibility phenotype but different genotypes.

Patient outcomes.Overall, 24% of patients died within 30 days after having a blood culture positive for K. pneumoniae and 28% died during the index hospitalization. Patients with either Ceph-R or CRKP BSIs had significantly worse outcomes than patients infected with susceptible isolates, including 30-day mortality rates (35% versus 20%; P = 0.02) and hospital mortality rates (45% versus 22%; P = 0.001). While the 30-day mortality rates did not differ significantly between patients with CRKP and Ceph-R infections (41% versus 29%; P = 0.4), the hospital mortality rate was significantly higher among patients with CRKP infections (59%) than among those with susceptible infections (22%; P < 0.0001) or Ceph-R infections (29%; P = 0.03).

Advanced age, the presence of multiple chronic comorbidities, and select disease severity markers were associated with 30-day mortality rates in univariable analyses (Table 5). After backward stepwise selection, the final multivariable model included age, PBS, CCIS, and ICU admission as significant predictors of 30-day mortality rates. No significant association between resistance phenotype and 30-day mortality rates remained (P = 0.7) in the final model.

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TABLE 5

Clinical predictors of 30-day mortality rates

CRKP genotypes and outcomes.We then assessed the associations between select molecular characteristics and 30-day mortality rates in patients with CRKP BSIs. These underpowered analyses were limited by the relatively small number of patients. The 30-day mortality rate was higher among patients infected with a ST258 strain (n = 21), compared to non-ST258 strains (n = 8), but the difference was not statistically significant (52% versus 13%; P = 0.09). After adjustment for CCISs and PBSs in a multivariable model, infection with a ST258 strain versus a non-ST258 strain also showed a borderline significant association with increased 30-day mortality rates (odds ratio [OR], 8.3 [95% confidence interval [CI], 0.7 to 100.4]; P = 0.09). The 30-day mortality rates did not differ significantly between patients infected with isolates positive for blaKPC-2 (n = 16) versus blaKPC-3 (n = 11) (38% versus 55%; P = 0.5). Lastly, patients with CRKP BSIs caused by ST258 strains did not have significantly worse outcomes if the isolate carried wzi154 (n = 12) versus other wzi allele types (n = 16) (58% versus 31%; P = 0.15). However, after adjustment for PBSs and CCISs, infection with a wzi154-carrying ST258 isolate versus a non-wzi154 isolate showed a borderline significant association with increased mortality rates (OR, 16.7 [95% CI, 1.0 to 283]; P = 0.051).

DISCUSSION

In this study, we describe the molecular epidemiology of K. pneumoniae strains causing bloodstream infections at a tertiary care hospital in New York City, an area of high CRKP endemicity (22). We detected substantial differences in clonal structure and diversity across K. pneumoniae antibiotic resistance groups. While a high level of diversity among antibiotic-susceptible isolates was anticipated, the degree of variability among both susceptible and Ceph-R or CRKP isolates was remarkably high. In multivariable analyses, advanced age, comorbidities, and illness severity were highly predictive of 30-day mortality rates, whereas the susceptibility phenotype was not. Taken together, these data suggest that poor outcomes for K. pneumoniae bloodstream infections were driven by patient vulnerability and disease severity.

Despite small numbers, we observed a borderline significant association of higher mortality rates with ST258 or isolates carrying the wzi154 allele in CRKP-infected patients, accounting for comorbidities and illness severity, whereas the blaKPC type was not associated with significantly different outcomes. The remarkably high levels of diversity in the clonal backgrounds of susceptible and Ceph-R isolates, combined with the restricted overlap in clonal backgrounds with CRKP isolates, precluded analysis of the impact of clonal backgrounds irrespective of the susceptibility phenotype. This high level of clonal diversity of susceptible K. pneumoniae isolates, combined with the observed onset of bacteremia within 72 h after hospital admission, is most consistent with community-onset infections attributable to a colonizing heterogeneous clonal population. The Ceph-R isolates were similarly diverse, supporting widespread diversification of strains in nonoutbreak settings in areas in which K. pneumoniae is endemic (23).

Although ST258 accounted for the majority of CRKP infections, we detected a larger proportion of non-ST258 strains, compared to previous reports from New York and elsewhere (2, 11, 24). Given that blaKPC is found on plasmids that are transmissible among Enterobacteriaceae and can produce novel CRKP strains (24, 25), plasmid-mediated uptake of antibiotic resistance determinants by our study isolates might have accounted for some of the unique resistant STs. However, we observed limited overlap in STs between susceptible or Ceph-R and CRKP isolates, including non-ST258 CRKP isolates, and no clear progression of resistance among K. pneumoniae isolates obtained from the same patient, to suggest unconstrained plasmid transfer. However, uptake of plasmids carrying blaKPC and other β-lactamase genes might have been limited to a subset of clones, such as ST17, that were present in all three susceptibility classes. Alternatively, recipient clones might have resided as colonizers and emerged in the setting of antibiotic exposure. In light of the high meropenem MICs observed here and the universal presence of the blaKPC gene in all CRKP isolates, the diversity of CRKP clones is less likely to have been accounted for by novel porin gene mutations. Further study is needed to better understand the contributions of plasmid transfer to MDR K. pneumoniae diversification.

The heterogeneity in wzi types and blaKPC variants among ST258 isolates encountered in our study is consistent with emerging literature on variability in capsule types and associated resistance determinants in this ST (10, 11, 26). We found that blaKPC-3 was almost entirely restricted to wzi154-positive ST258 CRKP isolates, whereas blaKPC-2 was detected in isolates with several sequence and wzi alleles. A blaKPC-3-encoding wzi154 clone was recently documented in another New York City hospital (11), raising the possibility of local expansion or interhospital spreading of isolates. However, a study conducted in Israel also noted restriction of blaKPC-3 to ST258 isolates carrying a common plasmid (pKpQIL), while blaKPC-2 was present in several different plasmids associated with different K. pneumoniae STs (27). This supports the hypothesis that blaKPC-2 might be present on mobile resistance determinants that are more readily transmissible. Differences in mortality rates according to the blaKPC variant or wzi allele were not noted here, although our sample size was limited and mortality rates might have been affected by other factors.

Several patient subpopulations were identified as important target populations for further study. Although severe illness and debility are known risk factors for CRKP infections (28), solid organ transplant (SOT) recipients were disproportionately affected by CRKP BSIs in our population. A recent review by Satlin et al. reported 3 to 10% incidence of CRKP infections among SOT recipients in areas of high CRKP endemicity and suggested similar rates of infections among liver, kidney, lung, and heart transplant recipients (29). We noted particularly large burdens of CRKP bacteremia among liver, and to a lesser extent kidney, transplant recipients. Given that twice the number of patients received kidney transplants, compared to liver transplants, at our center in 2012 and 2013 (21), our data suggest that liver transplant patients are more frequently affected by CRKP and thus are at greater risk for death resulting from K. pneumoniae BSIs.

Patients with ESRD and those who resided in long-term-care facilities, including long-term acute-care hospitals (LTACHs) and nursing homes, also represented large proportions of the patients with CRKP bacteremia. Previous studies showed that residents of long-term-care facilities have relatively high rates of CRKP colonization (30) and LTACHs may serve as important transmission hubs for CRKP (31). A similar phenomenon may occur in dialysis units. Although we were not able to determine whether patients with similar strains shared long-term-care facilities or dialysis centers, nonhospital nosocomial transmission might have been responsible for some CRKP strain acquisition in our study.

Several limitations of our study need to be considered. First, this study examined patients who were admitted to a single tertiary hospital system with an active solid organ transplant center in New York City. Therefore, our results reflect the local at-risk population and do not examine regional differences in carbapenem resistance prevalence or associated molecular markers. Second, because this was a retrospective analysis, only stored clinical specimens could be examined, and we were not able to assess colonization or infection with K. pneumoniae in noncultured sources. Sites of K. pneumoniae colonization, especially the gastrointestinal tract, may serve as settings for strain diversification in high-risk patients and should be investigated in more detail. Third, although MLST and wzi allele typing are able to broadly characterize strain heterogeneity, they lack the resolution necessary to identify patterns of genetic diversification among related strains. High-resolution whole-genome sequencing studies are needed to better understand the evolution of clonal types and the development of resistance in particular strains. Last, despite the relatively large sample size, the number of CRKP infections was limited. Therefore, some of our negative findings have to be taken cautiously. Larger multicenter investigations are needed to further evaluate the effects of genotypes and resistance phenotypes on patient outcomes.

K. pneumoniae is a heterogeneous bacterial pathogen that is adept at developing antibiotic resistance, including resistance to broad-spectrum antibiotics. Although nosocomial transmission of ST258 continues to play an important role in the spread of CRKP, the diversifying clonal background of carbapenem-resistant strains suggests that plasmid-mediated transfer of resistance determinants is contributing to this epidemic. While only select strains appear to be able to take up resistance determinants, the clonal background of CRKP isolates might be associated with virulence and patient outcomes. A more detailed understanding of the genetic and host factors that enable the uptake of resistance determinants by novel strains will be critical to efforts to limit the spread of MDR K. pneumoniae.

ACKNOWLEDGMENTS

This work was supported in part by a NIH training grant in infectious diseases (grant 5T32AI100852-02 to A.G.-S.), a Columbia University Paul A. Marks scholarship (A.-C.U.), and NIAID/NIH grant 1R01AI116939 (A.-C.U.).

We thank the clinical microbiology laboratory for assistance in sample collection. We also acknowledge Valerie Passet and the Institut Pasteur MLST database for assistance in identifying and assigning new STs and wzi alleles.

We have no conflicts of interest to declare.

FOOTNOTES

    • Received 5 December 2014.
    • Returned for modification 12 January 2015.
    • Accepted 9 April 2015.
    • Accepted manuscript posted online 15 April 2015.

REFERENCES

  1. 1.↵
    Centers for Disease Control and Prevention. 2013. Antibiotic resistance threats in the United States, 2013. Centers for Disease Control and Prevention, Atlanta, GA. http://www.cdc.gov/drugresistance/threat-report-2013.
  2. 2.↵
    1. Kitchel B,
    2. Rasheed JK,
    3. Patel JB,
    4. Srinivasan A,
    5. Navon-Venezia S,
    6. Carmeli Y,
    7. Brolund A,
    8. Giske CG
    . 2009. Molecular epidemiology of KPC-producing Klebsiella pneumoniae isolates in the United States: clonal expansion of multilocus sequence type 258. Antimicrob Agents Chemother 53:3365–3370. doi:10.1128/AAC.00126-09.
    OpenUrlAbstract/FREE Full Text
  3. 3.↵
    1. Munoz-Price LS,
    2. Poirel L,
    3. Bonomo RA,
    4. Schwaber MJ,
    5. Daikos GL,
    6. Cormican M,
    7. Cornaglia G,
    8. Garau J,
    9. Gniadkowski M,
    10. Hayden MK,
    11. Kumarasamy K,
    12. Livermore DM,
    13. Maya JJ,
    14. Nordmann P,
    15. Patel JB,
    16. Paterson DL,
    17. Pitout J,
    18. Villegas MV,
    19. Wang H,
    20. Woodford N,
    21. Quinn JP
    . 2013. Clinical epidemiology of the global expansion of Klebsiella pneumoniae carbapenemases. Lancet Infect Dis 13:785–796. doi:10.1016/S1473-3099(13)70190-7.
    OpenUrlCrossRefPubMedWeb of Science
  4. 4.↵
    1. Patel G,
    2. Huprikar S,
    3. Factor SH,
    4. Jenkins SG,
    5. Calfee DP
    . 2008. Outcomes of carbapenem-resistant Klebsiella pneumoniae infection and the impact of antimicrobial and adjunctive therapies. Infect Control Hosp Epidemiol 29:1099–1106. doi:10.1086/592412.
    OpenUrlCrossRefPubMedWeb of Science
  5. 5.↵
    1. Nordmann P,
    2. Cuzon G,
    3. Naas T
    . 2009. The real threat of Klebsiella pneumoniae carbapenemase-producing bacteria. Lancet Infect Dis 9:228–236. doi:10.1016/S1473-3099(09)70054-4.
    OpenUrlCrossRefPubMedWeb of Science
  6. 6.↵
    1. Borer A,
    2. Saidel-Odes L,
    3. Riesenberg K,
    4. Eskira S,
    5. Peled N,
    6. Nativ R,
    7. Schlaeffer F,
    8. Sherf M
    . 2009. Attributable mortality rate for carbapenem-resistant Klebsiella pneumoniae bacteremia. Infect Control Hosp Epidemiol 30:972–976. doi:10.1086/605922.
    OpenUrlCrossRefPubMedWeb of Science
  7. 7.↵
    1. Zarkotou O,
    2. Pournaras S,
    3. Tselioti P,
    4. Dragoumanos V,
    5. Pitiriga V,
    6. Ranellou K,
    7. Prekates A,
    8. Themeli-Digalaki K,
    9. Tsakris A
    . 2011. Predictors of mortality in patients with bloodstream infections caused by KPC-producing Klebsiella pneumoniae and impact of appropriate antimicrobial treatment. Clin Microbiol Infect 17:1798–1803. doi:10.1111/j.1469-0691.2011.03514.x.
    OpenUrlCrossRefPubMed
  8. 8.↵
    1. Neuner EA,
    2. Yeh JY,
    3. Hall GS,
    4. Sekeres J,
    5. Endimiani A,
    6. Bonomo RA,
    7. Shrestha NK,
    8. Fraser TG,
    9. van Duin D
    . 2011. Treatment and outcomes in carbapenem-resistant Klebsiella pneumoniae bloodstream infections. Diagn Microbiol Infect Dis 69:357–362. doi:10.1016/j.diagmicrobio.2010.10.013.
    OpenUrlCrossRefPubMed
  9. 9.↵
    1. Ben-David D,
    2. Kordevani R,
    3. Keller N,
    4. Tal I,
    5. Marzel A,
    6. Gal-Mor O,
    7. Maor Y,
    8. Rahav G
    . 2012. Outcome of carbapenem resistant Klebsiella pneumoniae bloodstream infections. Clin Microbiol Infect 18:54–60. doi:10.1111/j.1469-0691.2011.03478.x.
    OpenUrlCrossRefPubMed
  10. 10.↵
    1. Deleo FR,
    2. Chen L,
    3. Porcella SF,
    4. Martens CA,
    5. Kobayashi SD,
    6. Porter AR,
    7. Chavda KD,
    8. Jacobs MR,
    9. Mathema B,
    10. Olsen RJ,
    11. Bonomo RA,
    12. Musser JM,
    13. Kreiswirth BN
    . 2014. Molecular dissection of the evolution of carbapenem-resistant multilocus sequence type 258 Klebsiella pneumoniae. Proc Natl Acad Sci U S A 111:4988–4993. doi:10.1073/pnas.1321364111.
    OpenUrlAbstract/FREE Full Text
  11. 11.↵
    1. Diago-Navarro E,
    2. Chen L,
    3. Passet V,
    4. Burack S,
    5. Ulacia-Hernando A,
    6. Kodiyanplakkal RP,
    7. Levi MH,
    8. Brisse S,
    9. Kreiswirth BN,
    10. Fries BC
    . 2014. Carbapenem-resistant Klebsiella pneumoniae exhibit variability in capsular polysaccharide and capsule associated virulence traits. J Infect Dis 210:803–813. doi:10.1093/infdis/jiu157.
    OpenUrlCrossRefPubMed
  12. 12.↵
    1. Gootz TD,
    2. Lescoe MK,
    3. Dib-Hajj F,
    4. Dougherty BA,
    5. He W,
    6. Della-Latta P,
    7. Huard RC
    . 2009. Genetic organization of transposase regions surrounding blaKPC carbapenemase genes on plasmids from Klebsiella strains isolated in a New York City hospital. Antimicrob Agents Chemother 53:1998–2004. doi:10.1128/AAC.01355-08.
    OpenUrlAbstract/FREE Full Text
  13. 13.↵
    1. Cuzon G,
    2. Naas T,
    3. Truong H,
    4. Villegas MV,
    5. Wisell KT,
    6. Carmeli Y,
    7. Gales AC,
    8. Venezia SN,
    9. Quinn JP,
    10. Nordmann P
    . 2010. Worldwide diversity of Klebsiella pneumoniae that produce β-lactamase blaKPC-2 gene. Emerg Infect Dis 16:1349–1356. doi:10.3201/eid1609.091389.
    OpenUrlCrossRefPubMedWeb of Science
  14. 14.↵
    1. Naas T,
    2. Cuzon G,
    3. Villegas MV,
    4. Lartigue MF,
    5. Quinn JP,
    6. Nordmann P
    . 2008. Genetic structures at the origin of acquisition of the β-lactamase blaKPC gene. Antimicrob Agents Chemother 52:1257–1263. doi:10.1128/AAC.01451-07.
    OpenUrlAbstract/FREE Full Text
  15. 15.↵
    1. Tsai YK,
    2. Fung CP,
    3. Lin JC,
    4. Chen JH,
    5. Chang FY,
    6. Chen TL,
    7. Siu LK
    . 2011. Klebsiella pneumoniae outer membrane porins OmpK35 and OmpK36 play roles in both antimicrobial resistance and virulence. Antimicrob Agents Chemother 55:1485–1493. doi:10.1128/AAC.01275-10.
    OpenUrlAbstract/FREE Full Text
  16. 16.↵
    Centers for Disease Control and Prevention. 2014. CDC/NHSN surveillance definitions for specific type of infections. Centers for Disease Control and Prevention, Atlanta, GA. http://www.cdc.gov/nhsn/PDFs/pscManual/17pscNosInfDef_current.pdf.
  17. 17.↵
    Clinical and Laboratory Standards Institute. 2011. Performance standards for antimicrobial susceptibility testing; 21st informational supplement. M100-S21. Clinical and Laboratory Standards Institute, Wayne, PA.
  18. 18.↵
    1. Diancourt L,
    2. Passet V,
    3. Verhoef J,
    4. Grimont PA,
    5. Brisse S
    . 2005. Multilocus sequence typing of Klebsiella pneumoniae nosocomial isolates. J Clin Microbiol 43:4178–4182. doi:10.1128/JCM.43.8.4178-4182.2005.
    OpenUrlAbstract/FREE Full Text
  19. 19.↵
    1. Poirel L,
    2. Walsh TR,
    3. Cuvillier V,
    4. Nordmann P
    . 2011. Multiplex PCR for detection of acquired carbapenemase genes. Diagn Microbiol Infect Dis 70:119–123. doi:10.1016/j.diagmicrobio.2010.12.002.
    OpenUrlCrossRefPubMed
  20. 20.↵
    1. Francis RO,
    2. Wu F,
    3. Della-Latta P,
    4. Shi J,
    5. Whittier S
    . 2012. Rapid detection of Klebsiella pneumoniae carbapenemase genes in Enterobacteriaceae directly from blood culture bottles by real-time PCR. Am J Clin Pathol 137:627–632. doi:10.1309/AJCP9SNHJG2QGLWU.
    OpenUrlCrossRefPubMed
  21. 21.↵
    Organ Procurement and Transplantation Network. 2014. Transplant center data, 2012–2013. Organ Procurement and Transplantation Network, Richmond, VA. http://optn.transplant.hrsa.gov/converge/LatestData/stateData.asp?type=center.
  22. 22.↵
    1. Landman D,
    2. Babu E,
    3. Shah N,
    4. Kelly P,
    5. Olawole O,
    6. Backer M,
    7. Bratu S,
    8. Quale J
    . 2012. Transmission of carbapenem-resistant pathogens in New York City hospitals: progress and frustration. J Antimicrob Chemother 67:1427–1431. doi:10.1093/jac/dks063.
    OpenUrlCrossRefPubMedWeb of Science
  23. 23.↵
    1. Essack SY,
    2. Hall LM,
    3. Pillay DG,
    4. McFadyen ML,
    5. Livermore DM
    . 2001. Complexity and diversity of Klebsiella pneumoniae strains with extended-spectrum β-lactamases isolated in 1994 and 1996 at a teaching hospital in Durban, South Africa. Antimicrob Agents Chemother 45:88–95. doi:10.1128/AAC.45.1.88-95.2001.
    OpenUrlAbstract/FREE Full Text
  24. 24.↵
    1. Chen L,
    2. Chavda KD,
    3. Melano RG,
    4. Jacobs MR,
    5. Koll B,
    6. Hong T,
    7. Rojtman AD,
    8. Levi MH,
    9. Bonomo RA,
    10. Kreiswirth BN
    . 2014. Comparative genomic analysis of KPC-encoding pKpQIL-like plasmids and their distribution in New Jersey and New York hospitals. Antimicrob Agents Chemother 58:2871–2877. doi:10.1128/AAC.00120-14.
    OpenUrlAbstract/FREE Full Text
  25. 25.↵
    1. Mathers AJ,
    2. Cox HL,
    3. Kitchel B,
    4. Bonatti H,
    5. Brassinga AK,
    6. Carroll J,
    7. Scheld WM,
    8. Hazen KC,
    9. Sifri CD
    . 2011. Molecular dissection of an outbreak of carbapenem-resistant Enterobacteriaceae reveals intergenus KPC carbapenemase transmission through a promiscuous plasmid. mBio 2(6):e00204–11.
    OpenUrlCrossRefPubMed
  26. 26.↵
    1. Kitchel B,
    2. Sundin DR,
    3. Patel JB
    . 2009. Regional dissemination of KPC-producing Klebsiella pneumoniae. Antimicrob Agents Chemother 53:4511–4513. doi:10.1128/AAC.00784-09.
    OpenUrlAbstract/FREE Full Text
  27. 27.↵
    1. Leavitt A,
    2. Carmeli Y,
    3. Chmelnitsky I,
    4. Goren MG,
    5. Ofek I,
    6. Navon-Venezia S
    . 2010. Molecular epidemiology, sequence types, and plasmid analyses of KPC-producing Klebsiella pneumoniae strains in Israel. Antimicrob Agents Chemother 54:3002–3006. doi:10.1128/AAC.01818-09.
    OpenUrlAbstract/FREE Full Text
  28. 28.↵
    1. Schwaber MJ,
    2. Klarfeld-Lidji S,
    3. Navon-Venezia S,
    4. Schwartz D,
    5. Leavitt A,
    6. Carmeli Y
    . 2008. Predictors of carbapenem-resistant Klebsiella pneumoniae acquisition among hospitalized adults and effect of acquisition on mortality. Antimicrob Agents Chemother 52:1028–1033. doi:10.1128/AAC.01020-07.
    OpenUrlAbstract/FREE Full Text
  29. 29.↵
    1. Satlin MJ,
    2. Jenkins SG,
    3. Walsh TJ
    . 2014. The global challenge of carbapenem-resistant Enterobacteriaceae in transplant recipients and patients with hematologic malignancies. Clin Infect Dis 58:1274–1283. doi:10.1093/cid/ciu052.
    OpenUrlCrossRefPubMed
  30. 30.↵
    1. Endimiani A,
    2. Depasquale JM,
    3. Forero S,
    4. Perez F,
    5. Hujer AM,
    6. Roberts-Pollack D,
    7. Fiorella PD,
    8. Pickens N,
    9. Kitchel B,
    10. Casiano-Colon AE,
    11. Tenover FC,
    12. Bonomo RA
    . 2009. Emergence of blaKPC-containing Klebsiella pneumoniae in a long-term acute care hospital: a new challenge to our healthcare system. J Antimicrob Chemother 64:1102–1110. doi:10.1093/jac/dkp327.
    OpenUrlCrossRefPubMedWeb of Science
  31. 31.↵
    1. Won SY,
    2. Munoz-Price LS,
    3. Lolans K,
    4. Hota B,
    5. Weinstein RA,
    6. Hayden MK
    . 2011. Emergence and rapid regional spread of Klebsiella pneumoniae carbapenemase-producing Enterobacteriaceae. Clin Infect Dis 53:532–540. doi:10.1093/cid/cir482.
    OpenUrlCrossRefPubMed
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