Carbapenemase-producing Klebsiella Pneumoniae Research Articles

Synergy Testing by E-Test and Microdilution Checkerboard for Fosfomycin Combined with Tigecycline against KPC-Producing Klebsiella pneumoniae

KPC-producing Klebsiella pneumoniae (KPC-Kp) has become a serious threat to patients worldwide, as the treatment options are limited. The combination of fosfomycin with other antibiotics has been reported but with inconsistent results. Thus, we performed synergy testing of fosfomycin combined with tigecycline by E-test, which was easy to perform and to determine results, compared with microdilution checkerboard, which is considered to be the "gold standard", to evaluate the agreement between the two methods. Thirty non-repetitive KPC-Kp isolates from different patients were included in this study. Bacterial identification and routine antibiotic susceptibility testing were performed by a VITEK 2 Compact automated system. The KPC producing isolates were identified by modified Carbapenem Inhibitory Method (mCIM) and PCR amplification of carbapenemase genes. Synergy testing of fosfomycin combined with tigecycline was performed by E-test (E-test stripes were placed at 90° angle), with microdilution chequerboard performed in parallel. Fractional inhibitory concentration index (FICI) was calculated. Statistical analyses were performed by SPSS 18.0 software. All 30 KPC-Kp were mCIM test positive and KPC-2 producing. The susceptibility rates of fosfomycin and tigecycline were 36.7% (11/30) and 63.3% (19/30), respectively. Both checkerboard and E-test results showed that most MICs of fosfomycin and tigecycline decreased in the combination group. FICI showed 13.3% - 16.7% isolates were synergistic, 30.0% - 36.7% were additive, and 50.0% - 53.3% were indifferent. No antagonism was found. There was no significant difference between the two groups (p > 0.05), and the overall agreement (with FICI difference ≤ 0.25 in each isolate) between the two methods was 76.7% (23/30). The synergy testing results determined by E-test correlated well with microdilution checkerboard. Thus E-test synergy testing has the potential to be used in routine clinical laboratory.

KPC-producing Klebsiella pneumoniae gut decolonisation following ceftazidime/avibactam-based combination therapy: A retrospective observational study.

KPC-producing Klebsiella pneumoniae (KPC-Kp) gut colonisation is a major risk factor for developing systemic infection. Ceftazidime/avibactam (CAZ/AVI) may have a role as decolonisation therapy in special situations. This was a retrospective, observational, multicentre study. The KPC-Kp gut decolonisation rate of CAZ/AVI-based therapy (Group A) was compared with other antimicrobial regimens (Group B) in patients with KPC-Kp infection. Among 12 patients in Group A, 11 (91.7%) achieved gut decolonisation. None of the 24 patients of Group B were decolonised. CAZ/AVI-based therapy could be useful in KPC-Kp gut decolonisation in high-risk patients.

Healthcare associated infections caused by plasmid-encoded blaKPC and blaNDM strains of Klebsiella pneumoniae in Ibadan, Nigeria

Introduction: Carbapenemase producing Klebsiella pneumoniae (CPKP) has recently emerged as major cause of healthcare associated infections (HAIs). These strains are now classified by the US Center for Disease Control as superbugs of urgent concern. This study investigated the occurrence of CPKP in HAIs among hospitalized patients in University College Hospital (UCH), Ibadan. Methods: Non-duplicate specimens obtained from 250 patients with clinical HAIs were cultured by standard microbiological methods over a period of 6 months. K. pneumoniae was identified using GNB 24E Microbact kit and invitro susceptibility to selected antibiotics was performed by the disk diffusion test. Carbapenemase production was tested by Modified Hodge technique. K pneumoniae carbapenemase (blaKPC) and New Delhi Metallo-β-lactamase (blaNDM) genes were detected by conventional PCR assay. Data were analyzed for the 50 (20%) patients who were culture positive for K pneumoniae infections. Results: Isolates were resistant to ampicillin (100%), cefuroxime (96%), amoxicillin (92%), gentamicin (86%), nitrofurantoin (80%), ciprofloxacin (80%), ceftazidime (78%) and ofloxacin (72%) but 76% were sensitive to meropenem. No carbapenemase enzyme was detected but 8 of 12 iso-lates resistant to meropenem carried blaKPC while 2 carried both blaKPC and blaNDM genes. The study shows that blaKPC and blaNDM K pneumoniae are involved in clinical infections in Ibadan. Conclusion: This study reports the second case of K pneumoniae NDM clinical infection in Nigeria. There is need to strengthen the infection control programme of Nigerian hospitals to prevent nationwide spread of these organisms.

Triple combination of meropenem, colistin and tigecycline was bactericidal in a dynamic model despite mere additive interactions in chequerboard assays against carbapenemase-producing Klebsiella pneumoniae isolates.

Combination schemes are commonly used for the treatment of infections due to carbapenemase-producing Klebsiella pneumoniae (CP-Kp). We therefore investigated the in vitro effectiveness of double and triple combinations of meropenem, colistin and tigecycline against CP-Kp isolates with different resistance mechanisms in a static broth microdilution model and a pharmacokinetic-pharmacodynamic model. One WT isolate and seven CP-Kp isolates with different carbapenem resistance mechanisms and increasing MICs of meropenem (4-512 mg/L), colistin (0.5-32 mg/L) and tigecycline (0.25-4 mg/L) were tested with a 3D chequerboard microdilution method. Combinations were then assessed in an in vitro pharmacokinetic-pharmacodynamic model simulating 50 and 100 mg of tigecycline q12h as a 1 h infusion, 4.5 million units of colistin q12h as a 1 h infusion and 1 g of meropenem q8h as 1 and 0.5 h infusions for 2 days. In the chequerboard assay, interactions within the triple combination were mainly additive with a median (range) fractional inhibitory index of 0.66 (0.22-1.26). In the dynamic model, meropenem alone was bactericidal against isolates with MICs up to 4 mg/L, whereas bactericidal activity was found with the double combination meropenem + colistin and the triple combination meropenem + colistin + tigecycline against CP-Kp isolates with meropenem MICs of 16 and 256 mg/L, respectively. A high dose (100 mg) of tigecycline and a prolonged infusion (1 h) of meropenem increased the efficacy of the triple combination. Despite the merely additive interactions in the chequerboard assay, the triple combination of meropenem, tigecycline and colistin was bactericidal in the dynamic model against highly resistant CP-Kp isolates. This effect was more pronounced if prolonged infusion of meropenem and high tigecycline dosing were used.

Increased Relative Abundance of Klebsiella pneumoniae Carbapenemase-producing Klebsiella pneumoniae Within the Gut Microbiota Is Associated With Risk of Bloodstream Infection in Long-term Acute Care Hospital Patients.

An association between increased relative abundance of specific bacterial taxa in the intestinal microbiota and bacteremia has been reported in some high-risk patient populations. We collected weekly rectal swab samples from patients at 1 long-term acute care hospital (LTACH) in Chicago from May 2015 to May 2016. Samples positive for Klebsiella pneumoniae carbapenemase-producing Klebsiella pneumoniae (KPC-Kp) by polymerase chain reaction and culture underwent 16S rRNA gene sequence analysis; relative abundance of the operational taxonomic unit containing KPC-Kp was determined. Receiver operator characteristic (ROC) curves were constructed using results from the sample with highest relative abundance of KPC-Kp from each patient admission, excluding samples collected after KPC-Kp bacteremia. Cox regression analysis was performed to evaluate risk factors associated with time to achieve KPC-Kp relative abundance thresholds calculated by ROC curve analysis. We collected 2319 samples from 562 admissions (506 patients); KPC-Kp colonization was detected in 255 (45.4%) admissions and KPC-Kp bacteremia in 11 (4.3%). A relative abundance cutoff of 22% predicted KPC-Kp bacteremia with sensitivity 73%, specificity 72%, and relative risk 4.2 (P = .01). In a multivariable Cox regression model adjusted for age, Charlson comorbidity index, and medical devices, carbapenem receipt was associated with achieving the 22% relative abundance threshold (P = .044). Carbapenem receipt was associated with increased hazard for high relative abundance of KPC-Kp in the gut microbiota. Increased relative abundance of KPC-Kp was associated with KPC-Kp bacteremia. Whether bacteremia arose directly from bacterial translocation or indirectly from skin contamination followed by bloodstream invasion remains to be determined.

Gut Microbiota and Clinical Features Distinguish Colonization With Klebsiella pneumoniae Carbapenemase-Producing Klebsiella pneumoniae at the Time of Admission to a Long-term Acute Care Hospital.

BackgroundIdentification of gut microbiota features associated with antibiotic-resistant bacterial colonization may reveal new infection prevention targets.MethodsWe conducted a matched, case–control study of long-term acute care hospital (LTACH) patients to identify gut microbiota and clinical features associated with colonization by Klebsiella pneumoniae carbapenemase-producing Klebsiella pneumoniae (KPC-Kp), an urgent antibiotic resistance threat. Fecal or rectal swab specimens were collected and tested for KPC-Kp; 16S rRNA gene-based sequencing was performed. Comparisons were made between cases and controls in calibration and validation subsamples using microbiota similarity indices, logistic regression, and unit-weighted predictive models.ResultsCase (n = 32) and control (n = 99) patients had distinct fecal microbiota communities, but neither microbiota diversity nor inherent clustering into community types distinguished case and control specimens. Comparison of differentially abundant operational taxonomic units (OTUs) revealed 1 OTU associated with case status in both calibration (n = 51) and validation (n = 80) subsamples that matched the canonical KPC-Kp strain ST258. Permutation analysis using the presence or absence of OTUs and hierarchical logistic regression identified 2 OTUs (belonging to genus Desulfovibrio and family Ruminococcaceae) associated with KPC-Kp colonization. Among clinical variables, the presence of a decubitus ulcer alone was independently and consistently associated with case status. Combining the presence of the OTUs Desulfovibrio and Ruminococcaceae with decubitus ulcer increased the likelihood of KPC-Kp colonization to >38% in a unit-weighted predictive model.ConclusionsWe identified microbiota and clinical features that distinguished KPC-Kp gut colonization in LTACH patients, a population particularly susceptible to KPC-Kp infection. These features may warrant further investigation as markers of risk for KPC-Kp colonization.

KPC-producing Klebsiella pneumoniae bloodstream isolates from Brazilian hospitals: What (still) remains active?

This study assessed susceptibility to polymyxin B (PMB) and alternative antimicrobials, with focus on aminoglycosides and tigecycline, according to different breakpoints in KPC-producing Klebsiella pneumoniae (KPC-Kp) bloodstream isolates from Brazilian hospitals. Bloodstream K. pneumoniae isolates non-susceptible to any of the three carbapenems (meropenem, imipenem or ertapenem) from four Brazilian tertiary-care hospitals were selected. Antimicrobial susceptibility was determined and interpreted according to distinct breakpoints. Twenty-nine PMB-resistant KPC-Kp isolates were selected for molecular typing. A total of 158 KPC-Kp were analysed. MIC50/90 values for PMB were 0.25/16mg/L; 40 isolates (25.3%) were resistant to PMB. MIC50/90 values for meropenem were 32/≥256mg/L; no isolates were susceptible to meropenem according to CLSI, but 10 isolates were intermediate using EUCAST breakpoints (1, MIC=4mg/L; 9, MIC=8mg/L). MIC50/90 values for tigecycline were 2/8mg/L; 53 (33.5%) and 94 (59.5%) isolates were susceptible according to EUCAST and FDA breakpoints, respectively. MIC50/90 values were 32/≥64mg/L for amikacin and ≥16/≥16mg/L for gentamicin; 48 (30.4%), 28 (17.7%) and 16 (10.1%) were susceptible to amikacin according to CLSI, EUCAST and USCAST, respectively, but susceptibility rates to gentamicin were <7.0%. Eighteen distinct clonal profiles were identified among 29 PMB-resistant isolates by DNA macrorestriction. Most clones belonged to CC11. Elevated rates of PMB-resistant KPC-Kp bloodstream infections were found in four Brazilian hospitals, mostly of polyclonal origin. Alternative antimicrobials with the highest in vitro activity were tigecycline and amikacin, although susceptibility rates significantly decreased using criteria with stricter breakpoints (e.g. EUCAST, USCAST).

Critical Review of Double-Carbapenem Therapy for the Treatment of Carbapenemase-Producing Klebsiella pneumoniae.

To review the clinical data on the effectiveness and safety of double carbapenem therapy (DCT) in patients infected with carbapenemase-producing Klebsiella pneumoniae (CP-Kp). A literature search was performed utilizing PubMed and EMBASE (from 1966 to May 2018); bibliographies of the retrieved articles were also searched. Articles were included if they evaluated patients with infections caused by CP-Kp and were treated with DCT. Meeting abstracts, editorials, and animal and in vitro studies were excluded. The search strategy revealed 8 case reports and 6 clinical studies (total of 171 patients) that evaluated the administration of ertapenem followed by prolonged infusions of meropenem or doripenem. Most patients were critically ill and commonly had infections in the blood, lungs, and urine. Clinical and microbiological success were reported in 70% of the patients and mortality in 24%. Adverse events, which included mostly seizures, sodium disorders, and gastrointestinal symptoms, were reported in 16 patients; none required interruption of treatment. Relevance to Patient Care and Clinical Practice: This review evaluated the clinical experience of DCT in the treatment of CP-Kp infections, based on case reports and clinical studies, for the potential role of DCT as a therapeutic option. Despite the limited studies, current data suggest that DCT may be an effective and safe strategy to treat CP-Kp. However, large randomized controlled trials are necessary to clearly define the role of DCT.

Infections by Carbapenemase Producing Klebsiella Pneumoniae Bacteria in Solid Organ Transplant Recipients

Infections by Carbapenemase Producing Klebsiella Pneumoniae Bacteria in Solid Organ Transplant Recipients

Colistin resistance in carbapenemase-producing Klebsiella pneumoniae bloodstream isolates: Evolution over 15 years and temporal association with colistin use by time series analysis

Colistin is often the only available treatment option against infections caused by carbapenemase-producing Klebsiella pneumoniae (CP-Kp). In this study, the evolution of colistin resistance among CP-Kp and its relationship with colistin use in a tertiary-care hospital in Athens, Greece, was investigated. All CP-Kp blood isolates recovered between January 2002 and June 2016 were tested for susceptibility to colistin by agar dilution and broth microdilution methods. Data on colistin use were collected from the pharmacy database. Time series of colistin use and resistance were analysed using the Box and Jenkins method. A transfer function model was built to quantify the dynamic relationship between colistin use and resistance. Overall, 313 CP-Kp isolates were identified. The percentage colistin resistance increased from 0% in 2002 to 26.9% in 2016 (R2 = 0.5, P < 0.01). A temporal association between colistin use and resistance was observed; an increase in colistin use by 1 DDD/100 patient-days led to a 0.05 increase in the incidence rate of colistin resistance. The time lag between the effect of colistin use on subsequent variations in colistin resistance was 3 months. Colistin use and prior levels of colistin resistance could explain 69% of colistin resistance; in the remaining 31%, other factors might have played a role. The results presented here demonstrate a significant temporal association between colistin use and colistin resistance. These findings have important implications in implementing strategies to contain colistin resistance.

Carbapenem Nonsusceptible Klebsiella pneumoniae in Taiwan: Dissemination and Increasing Resistance of Carbapenemase Producers During 2012\u20132015

Before 2011, the prevalence rates of carbapenemase-producing Klebsiella pneumoniae (CPKP) among carbapenem nonsusceptible K. pneumoniae (CnSKP) isolates were below 10% in Taiwan. The study presents the dissemination and increased antimicrobial resistance of CPKP from January 2012 to August 2015, as shown by Taiwanese multicenter surveillance. Isolates with minimum inhibitory concentrations (MICs) of >1 μg/mL for imipenem or meropenem were collected, screened for various carbapenemase genes by PCR, and tested for antimicrobial susceptibility. Among 1,457 CnSKP isolates, 1,250 were collected from medical centers. The CnSKP prevalence in medical centers increased by 1.7-fold during the study. Among all CnSKP isolates, 457 were CPKP. The CPKP rate among CnSKP increased by 1.5-fold and reached 36.8% in 2015. The CPKP nonsusceptibility rate to aztreonam, fluoroquinolones, and aminoglycosides increased yearly. Six CPKP isolates carried dual carbapenemase genes. Three Ambler classes were identified in 451 isolates with a single carbapenemase: classes A (315 blaKPC-2, 2 blaKPC-3, 28 blaKPC-17, 2 blaKPC-34), B (26 blaIMP-8, 2 blaNDM-1, 36 blaVIM-1), and D (40 blaOXA-48). The blaOXA-48 rate among CPKP increased by 6-fold over three years. Most KPC and OXA-48 producers were ST11. CnSKP was increasingly prevalent, owing to CPKP dissemination. Additionally, CPKP became more resistant during the study period.

Efficacy of Colistin and Its Combination With Rifampin in Vitro and in Experimental Models of Infection Caused by Carbapenemase-Producing Clinical Isolates of Klebsiella pneumoniae.

Despite the relevance of carbapenemase-producing Klebsiella pneumoniae (CP-Kp) infections there are a scarce number of studies to evaluate in vivo the efficacy of combinations therapies. The bactericidal activity of colistin, rifampin, and its combination was studied (time–kill curves) against four clonally unrelated clinical isolates of CP-Kp, producing VIM-1, VIM-1 plus DHA-1(acquired AmpC β-lactamase), OXA-48 plus CTX-M-15 (extended spectrum β-lactamase) and KPC-3, respectively, with colistin MICs of 0.5, 64, 0.5, and 32 mg/L, respectively. The efficacies of antimicrobials in monotherapy and in combination were tested in a murine peritoneal sepsis model, against all the CP-Kp. Their efficacies were tested in the pneumonia model against the OXA-48 plus CTX-M-15 producers. The development of colistin-resistance was analyzed for the colistin-susceptible strains in vitro and in vivo. In vitro, colistin plus rifampin was synergistic against all the strains at 24 h. In vivo, compared to the controls, rifampin alone reduced tissue bacterial concentrations against VIM-1 and OXA-48 plus CTX-M-15 strains; CMS plus rifampin reduced tissue bacterial concentrations of these two CP-Kp and of the KPC-3 strain. Rifampin and the combination increased the survival against the KPC-3 strain; in the pneumonia model, the combination also improved the survival. No resistant mutants appeared with the combination. In conclusion, CMS plus rifampin had a low and heterogeneous efficacy in the treatment of severe peritoneal sepsis model due to CP-Kp producing different carbapenemases, increasing survival only against the KPC-3 strain. The combination showed efficacy in the less severe pneumonia model. The combination prevented in vitro and in vivo the development of colistin resistant mutants.

In vivo evolution of resistant subpopulations of KPC-producing Klebsiella pneumoniae during ceftazidime/avibactam treatment.

KPC-producing Klebsiella pneumoniae (KPC-Kp) represent a serious problem worldwide. Herein, we describe the evolution of ceftazidime/avibactam resistance by sequencing longitudinal clinical isolates from a patient with KPC-Kp bloodstream infection undergoing ceftazidime/avibactam treatment. WGS was performed on one ceftazidime/avibactam-susceptible KPC-Kp (BOT-CA-S) and two phenotypically different ceftazidime/avibactam-resistant KPC-Kp with low (BOT-CA-R) and high (BOT-EMO) carbapenem MICs. The population diversity was assessed by the frequency of allele mutations and population analysis profiles (PAPs). Phylogenetic analysis demonstrated clonal relatedness of the KPC-Kp isolates, all belonging to the clone ST1519. The D179Y mutation in blaKPC-3 was detected in both of the ceftazidime/avibactam-resistant KPC-Kp, whereas it was absent in the ceftazidime/avibactam-susceptible isolate. The mutation emerged independently in the two ceftazidime/avibactam-resistant isolates and was associated with a significant reduction in carbapenem MICs in BOT-CA-R, but not in BOT-EMO. WGS analysis revealed that the frequency of the D179Y mutation was 96.32% and 51.05% in BOT-CA-R and BOT-EMO, respectively. PAP results demonstrated that carbapenem resistance in BOT-EMO was due to the coexistence of mixed subpopulations harbouring WT and mutated blaKPC-3. A bacterial subpopulation with high ceftazidime/avibactam resistance for BOT-EMO KPC-Kp showed low carbapenem MICs, whereas a subpopulation with high meropenem resistance had a low MIC of ceftazidime/avibactam. Our analysis indicates that mixed subpopulations of ceftazidime/avibactam-resistant KPC-Kp emerge after ceftazidime/avibactam treatment. The evolution of different subpopulations that are highly resistant to ceftazidime/avibactam likely contributes to treatment failure, thereby highlighting the need for combination treatment strategies to limit selection of ceftazidime/avibactam-resistant KPC-Kp subpopulations.

Use of Monte Carlo simulation to evaluate the efficacy of tigecycline and minocycline for the treatment of pneumonia due to carbapenemase-producing Klebsiella pneumoniae

Background: Pneumonia caused by carbapenemase-producing Klebsiella pneumoniae (CP-KP) are increasingly encountered in hospitals worldwide, causing high mortality due to lack of treatment options. The goal of this study was to assess the efficacy of tigecycline and minocycline for CP-KP hospital-acquired pneumonia (HAP) by using Monte Carlo simulation.Methods: A total of 164 non-duplicated CP-KP strains were collected from sputum or blood in patients with HAP. The MICs for antimicrobials were determined by the agar dilution method. A 10,000-patient Monte Carlo Simulation based on a PK/PD model incorporating the MICs and population pharmacokinetic parameters were conducted to calculate probability of target attainment (PTA) at each MIC value and total cumulative fraction of response (CFR).Results: The susceptibility rate of tigecycline and minocycline were 79.9% and 41.5%, respectively. At recommended doses, an optimal PTA of 90% was obtained for treating HAP caused by CP-KP with MICs of tigecycline ≤0.5 mg/L or minocycline ≤4 mg/L. The CFR of tigecycline at the recommended dose and double dose (100 mg q12h) were 71.2% and 90.2%, respectively. The CFR of minocycline at recommended dose and double dose (200 mg q12h) was 53.4% and 77.2%, respectively.Conclusions: The findings of this study suggest that the recommended dose of tigecycline was not effective in HAP caused by CP-KP, and a higher CFR indicating a better clinical efficacy can be gained by doubling the dose (100 mg q12h). minocycline (200 mg q12h) might be a potential alternative of tigecycline to against strains with MICs ≤ 8 mg/L.

Molecular Characterization of Carbapenemase Producing Klebsiella Pneumoniae Dominance of OXA-48, KPC, VIM and NDM Producers in Khartoum, Sudan

Background and Objectives: Little is known about carbapenemase producing Klebsiella pneumoniae (CPK) in Sudan. This study aimed to determine the prevalence of CPK in a major hospital in Khartoum, Sudan between may 2015 - January 2017 and to characterize the isolates and detect the types of carbapenemase (s) they produced. Materials and Methods: The study was done in the Department of Molecular Microbiology, Faculty of Medical Laboratories Science, Al-Neleen University. All the isolates were obtained from clinical samples of patients treated inside the hospitals. Strains of K. pneumoniae resistant to at least one carbapenem (imipenem or meropenem) by using disc diffusion technique according to the CLSI guidelines were included in this study. Molecular detection of carbapenemase genes was achieved using Real-Time PCR (Sacace Biotechnologie, Italie). Results: A total 96 strains of K. pneumoniae of different non duplicated isolates were obtained from different hospitals. Seventy-two percent (70/96) isolates were positive for carbapenemase genes; 59.4% (57/96) were positive for blaKPC genes, 57.3% (55/96) were positive for blaNDM genes, 37.5% (36/96) were positive for blaVIM genes and 35.4% (34/96) were positive for blaOXA-48 genes. Nineteen isolates possessed four genes (blaKPC, blaNDM, blaVIM and blaOXA-48), fourteen isolates possessed three genes{( blaNDM, blaVIM and blaOXA-48=6), (blaKPC, blaNDM, and blaOXA-48=3), (blaKPC, blaNDM and blaVIM =3), (blaKPC, blaVIM and blaOXA-48=2)}, 27 isolates possessed two genes{( blaKPC and blaNDM =21), (blaKPC, blaOXA-48=2), (blaNDM and blaVIM =3), (blaNDM and blaOXA-48=1)}, 10 isolates possessed only one gene (blaKPC =8, blaOXA-48=1 and blaVIM =1) and the remaining 26 isolates were free from these genes. Conclusion &amp; Recommendation: In Sudan, the most common type of carbapenemase gene multidrug-resistant K. pneumoniae is KPC. Co-production of KPC, VIM, NDM and OXA-48 genes are found in K. pneumoniae. To our knowledge, this study was done for the first time in Sudan. Therefore, it is necessary to determine carbapenem resistance in K. pneumoniae isolates and take essential infection control precautions to avoid spread of this resistance.

Expansion of KPC-producing Klebsiella pneumoniae with various mgrB mutations giving rise to colistin resistance: the role of ISL3 on plasmids

mcr-1 has been reported as the first plasmid-encoded gene conferring colistin resistance. In KPC-producing Klebsiella pneumoniae (KPC-KP), however, colistin resistance is rapidly emerging through other mechanisms. Resistance is frequently due to disruption of the mgrB gene by insertion sequences, e.g. ISL3. The aim of this study was to investigate the expansion of mgrB-mutated KPC-KP isolates. In addition, the localisation and targets of ISL3 sequences within the core and accessory genome of common KPC-KP lineages were identified. A total of 29 clinical K. pneumoniae isolates collected from Italian patients were randomly selected. Whole genome sequences were analysed for resistance genes, plasmids and insertion sequences. In addition, 27 colistin-resistant KPC-KP isolates from a previous study from Crete (Greece) were assessed. Clonal expansion of KPC-KP isolates with various mutations in mgrB among all lineages was observed. In two Italian MLST ST512 isolates and eight Greek ST258 isolates, an identical copy of ISL3 was inserted in mgrB nucleotide position 133. ISL3, a transposable restriction–modification system of 8154 nucleotides, was located on pKpQIL-like plasmids and may transpose into the chromosome. In four isolates, chromosomal integration of ISL3 in diverse inner membrane proteins other than mgrB was identified. Colistin resistance is most often explained by clonal expansion of isolates with mutated mgrB. pKpQIL-like plasmids, which are omnipresent in KPC-KP, carry insertion sequences such as ISL3 that have mgrB as a target hotspot for transposition. Transposition of insertion sequences from plasmids and subsequent clonal expansion may contribute to the emerging colistin resistance in KPC-KP.

In vitro interaction of ceftazidime–avibactam in combination with different antimicrobials against KPC-producing Klebsiella pneumoniae clinical isolates

ObjectivesCombination therapy has been recommended when using ceftazidime–avibactam (CAZ–AVI) for the treatment of KPC-producing Klebsiella pneumoniae (KPC-Kp), but the optimal combination is unknown. Six common antimicrobial agents (ertapenem, imipenem, meropenem, gentamicin, tigecycline, and ciprofloxacin) were evaluated for synergy with the recently approved cephalosporin–β-lactamase inhibitor combination CAZ–AVI in this study. MethodsDifferent antimicrobial combinations were tested against 13 KPC-Kp, including CAZ–AVI-susceptible (n=11) and resistant (n=2) clinical isolates. In vitro interactions of CAZ–AVI with different antimicrobials were tested using the gradient synergy test. Changes in the minimum inhibitory concentration (MIC) value were interpreted using the fractional inhibitory concentration (FIC) index and susceptible breakpoint index (SBPI). ResultsThe combination of CAZ–AVI with gentamicin or ciprofloxacin displayed no synergism against any of the KPC-Kp isolates, whereas synergistic activity was observed with imipenem and meropenem against all KPC-Kp isolates. Notably, CAZ–AVI reduced MICs for meropenem and imipenem below the resistance breakpoints against all strains. The SBPI analysis showed that CAZ–AVI in combination with imipenem achieved higher SBPI values than other CAZ–AVI-based combinations. ConclusionsThese data suggest that combinations of CAZ–AVI with imipenem may be considered a useful therapeutic option for the treatment of KPC-Kp infections.

Population Pharmacokinetics of High-Dose Continuous-Infusion Meropenem and Considerations for Use in the Treatment of Infections Due to KPC-Producing Klebsiella pneumoniae.

We assessed the population pharmacokinetics of high-dose continuous-infusion (HDCI) meropenem in a cohort of patients with Klebsiella pneumoniae carbapenemase (KPC)-producing Klebsiella pneumoniae (KPC-Kp) infections. Monte Carlo simulations were used to define the permissible HDCI meropenem regimens that could be safely considered for the treatment of KPC-Kp infections due to meropenem-resistant strains. Permissible doses were arbitrarily defined as those associated with a ≤10% to 15% likelihood of meropenem steady-state concentrations (Css) of >100 mg/liter. Probabilities of target attainment (PTA) of four incremental pharmacodynamic determinants for meropenem efficacy (100% T>1×MIC, 100% T>2×MIC, 100% T>3×MIC, and 100% T>4×MIC, where "T>MIC" represents the time during which the plasma concentration of this time-dependent antibacterial agent is maintained above the MIC for the pathogen) in relation to different classes of renal function were calculated. The cumulative fractions of response (CFR) for the permissible HDCI meropenem regimens were calculated against the MIC distribution of the KPC-Kp clinical isolates that were collected routinely at our University Hospital between 2013 and 2016 (n = 169). Ninety-seven meropenem Css were included in the analysis. The final model included creatinine clearance (CrCL) as a covariate and explained 94% of the population variability. Monte Carlo simulations based on licensed dosages of up to 6 g/day predicted an acceptable PTA (>80%) of 100% T>1×MIC against KPC-Kp with a meropenem MIC of ≤32 mg/liter in patients with a CrCL level of <130 ml/min. Dosages of 8 g/day were needed for achieving the same target in patients with CrCL at levels of 130 to 200 ml/min. In dealing with pathogens with a meropenem MIC of 64 mg/liter, HDCI regimens using meropenem at higher than licensed levels should be considered. In these cases, real-time therapeutic drug monitoring could be a useful adjunct for optimized care. The predicted CFR were >75% in all of the classes of renal function.

Covert dissemination of carbapenemase-producing Klebsiella pneumoniae (KPC) in a successfully controlled outbreak: long- and short-read whole-genome sequencing demonstrate multiple genetic modes of transmission.

BackgroundCarbapenemase-producing Enterobacteriaceae (CPE), including KPC-producing Klebsiella pneumoniae (KPC-Kpn), are an increasing threat to patient safety.ObjectivesTo use WGS to investigate the extent and complexity of carbapenemase gene dissemination in a controlled KPC outbreak.Materials and methodsEnterobacteriaceae with reduced ertapenem susceptibility recovered from rectal screening swabs/clinical samples, during a 3 month KPC outbreak (2013–14), were investigated for carbapenemase production, antimicrobial susceptibility, variable-number-tandem-repeat profile and WGS [short-read (Illumina), long-read (MinION)]. Short-read sequences were used for MLST and plasmid/Tn4401 fingerprinting, and long-read sequence assemblies for plasmid identification. Phylogenetic analysis used IQTree followed by ClonalFrameML, and outbreak transmission dynamics were inferred using SCOTTI.ResultsTwenty patients harboured KPC-positive isolates (6 infected, 14 colonized), and 23 distinct KPC-producing Enterobacteriaceae were identified. Four distinct KPC plasmids were characterized but of 20 KPC-Kpn (from six STs), 17 isolates shared a single pKpQIL-D2 KPC plasmid. All isolates had an identical transposon (Tn4401a), except one KPC-Kpn (ST661) with a single nucleotide variant. A sporadic case of KPC-Kpn (ST491) with Tn4401a-carrying pKpQIL-D2 plasmid was identified 10 months before the outbreak. This plasmid was later seen in two other species and other KPC-Kpn (ST14,ST661) including clonal spread of KPC-Kpn (ST661) from a symptomatic case to nine ward contacts.ConclusionsWGS of outbreak KPC isolates demonstrated blaKPC dissemination via horizontal transposition (Tn4401a), plasmid spread (pKpQIL-D2) and clonal spread (K. pneumoniae ST661). Despite rapid outbreak control, considerable dissemination of blaKPC still occurred among K. pneumoniae and other Enterobacteriaceae, emphasizing its high transmission potential and the need for enhanced control efforts.

Risk factors and predictors of carbapenem-resistant Pseudomonas aeruginosa and Acinetobacter baumannii mortality in critically ill bacteraemic patients over a 6-year period (2010-15): antibiotics do matter.

Purpose. Acinetobacter baumannii and Pseudomonas aeruginosa provoke serious infections, especially in intensive care unit (ICU) patients.Methodology. The risk factors and predictors of mortality for P. aeruginosa (n=84; 46 carbapenem-resistant) and A. baumannii (n=129; all carbapenem-resistant) bloodstream infections (BSIs) in an ICU were evaluated. Antibiotic susceptibility testing was performed using the agar disk diffusion method according to EUCAST guidelines. The minimum inhibitory concentration was determined by a gradient method (Etest). Multilocus sequence typing (MLST) was performed for P. aeruginosa during the carbapenem-resistant outbreak in 2014. Epidemiological data were collected from the patients' chart reviews.Results/Key findings. Hospitalization during the summer months, prior KPC-producing Klebsiella pneumoniae (KPC-Kp) BSI, and the administration of tigecycline, aminoglycosides and cortisone were independently associated with P. aeruginosa BSIs. MLST revealed the dissemination of clone ST227, including carbapenem-resistant P. aeruginosa strains. Hospitalization during the summer months, prior KPC-Kp BSI, and the administration of antibiotics, carbapenem and cortisone were independently associated with A. baumannii BSIs. The 30-day mortality rate for P. aeruginosa and A. baumannii BSI was 45.2 and 39.5 %, respectively. Sequential organ failure assessment (SOFA) score at onset, septic shock, age, and prior KPC-Kp BSI were significantly associated with P. aeruginosa BSI mortality. The administration of at least one active antibiotic was identified as a predictor of a good prognosis. Septic shock and simplified acute physiology score (SAPS) II at onset were independently associated with A. baumannii BSI mortality. The administration of at least one active antibiotic and colistin-vancomycin co-administration were identified as predictors of a good prognosis.Conclusion. KPC-Kp infection predisposes ICU patients to BSI by either A. baumannii or P. aeruginosa. The administration of at least one active antibiotic leads to better survival rates.