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Data in the literature on the use of ceftazidime-avibactam (CAZ-AVI) in carbapenem-resistant Klebsiella pneumoniae bloodstream infections (CRKP-BSIs) are limited especially in OXA-48 (Oxacillinase-48) predominant regions. Our study aimed to evaluate the effect of CAZ-AVI use on outcomes in CRKP-BSIs in Turkey, where OXA-48 is endemic. A multicenter retrospective observational study was conducted between January 2017 and September 2021. The effects of clinical and treatment characteristics on 30-day mortality and relapse in CRKP-BSIs were analyzed. Predictors of outcomes were detected using a Cox regression model. The study enrolled 106 adults with CAZ-AVI-sensitive CRKP-BSIs who received CAZ-AVI for at least 72 h. Patients who received CAZ-AVI as initial therapy had lower mortality rates when compared to those who switched from last resort regimens [14.3% (n = 3/21) vs. 37.7% (n = 32/85), p = 0.04]. In multivariate analysis, older age and severe neutropenia were detected to be associated with higher mortality, significantly. Initiation of CAZ-AVI on the day of blood culture was obtained, was found to be significantly associated with lower mortality (HR: 0.25, CI: 0.07-0.84, p = 0.025). CAZ-AVI monotherapy is an important treatment option for CRKP-BSIs in OXA-48 endemic areas. Early initiation of CAZ-AVI should be preferred rather than switching from a last-resort regimen as it profoundly improves the survival rates., (© 2024. The Author(s).)

Background: Carbapenem-resistant Klebsiella pneumoniae (CRKP) infections poses global challenges, with limited options available for targeted therapy. Polymyxin was been regarded as one of the most important last-resort antimicrobial agents. Many factors could accelerate the resistance evolution of polymyxin. Insertion sequence (IS) inserted into mgrB is the main polymyxin resistance mechanism in K. pneumoniae. In this study, two CRKPs (KP31157 and KP31311) were isolated from the urine of a patient, shifting from susceptible to resistant as the mgrB inserted by ISkpn14. We intended to explore the origin of the IS and underlying mechanisms resulting in polymyxin resistance., Methods: The within-host evolution relationship and molecular features of both CRKPs were determined by pulsed-field gel electrophoresis (PFGE) and whole-genome sequencing (WGS). pKP31311&#95;KPC-2 plasmid genome structures contained in the above two CRKPs were aligned with the homologic plasmids, retrieved from the NCBI genome database via comparative genomic analysis. The plasmids encoding ISkpn14 elements flanked by direct repeat (DR) or not were analyzed. The mRNA expression, plasmid curing and in vitro antibiotics inducing experiment were employed to understand the potential mechanism of polymyxin resistance., Results: Both strains, sharing homology, exhibited polymyxin resistance due to the insertion of ISkpn14 into the mgrB gene, influenced by minocycline exposure. Minocycline and tigecycline could accelerate polymyxin resistance (P < 0.05), validated by an in vitro induction experiment. The ISkpn14 without DR flanked expressed about 4 times higher than that with DR. The frequency of the mgrB insertion induced by polymyxin was significantly reduced (0 strain detected) after the bla KPC-2 -carrying plasmid was eliminated., Conclusions: This study provides direct experimental evidence that the ISkpn14 element causing mgrB inactivation and polymyxin resistance in K. pneumoniae originates from bla KPC-2 -carrying plasmids. Minocycline exposure will accelerate the evolution of polymyxin resistance. Understanding the dynamics of IS transposition and its association with antibiotic exposure is crucial for developing effective strategies to reduce the emergence of polymyxin resistance in CRKP., (© 2024. The Author(s).)

Background: Carbapenem-resistant Klebsiella pneumoniae (CRKP), a significant worldwide public health threat, is common in patients in intensive care units. Methods: A retrospective study was conducted over a period of 22 months to assess the risk factors associated with infection caused by CRKP isolates. Strain identification was performed using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS), and antimicrobial sensitivity was assessed using the micro broth dilution method and Kirby-Bauer test. The genes bla KPC , bla OXA-48 , bla NDM , bla VIM , and bla GES were amplified using polymerase chain reaction (PCR), followed by sequencing of the PCR products. The polymerase hypermucoviscosity phenotype was determined using the string test. Capsular serotypes (K1, K2) and presence of the virulence gene ( rmpA ) in positive isolates were investigated using phenotypic tests followed by PCR. Results: Length of hospitalization and use of carbapenems were associated with CRKP infection. CRKP isolates exhibited extensive drug resistance, but retained sensitivity to colistin and ceftazidime-avibactam (CZA). The main gene detected in 35 CRKP isolates was bla KPC-2 . In addition, 11 strains were positive in the string test, and two of these strains carried rmpA . Conclusions: Prolonged hospitalization and carbapenem exposure increased the risk of CRKP infection in intensive care unit (ICU) patients. The prevalence of CRKP carrying the blaKPC-2 gene was high, and suspected hypervirulent carbapenem-resistant K. pneumoniae isolates were scattered.

The global rise of antibiotic resistance has renewed interest in phage therapy, as an alternative to antibiotics to eliminate multidrug-resistant (MDR) bacterial pathogens. However, optimizing the broad-spectrum efficacy of phage therapy remains a challenge. In this study, we addressed this issue by employing strategies to improve antimicrobial efficacy of phage therapy against MDR Klebsiella pneumoniae strains, which are notorious for their resistance to conventional antibiotics. This includes the selection of broad host range phages, optimization of phage formulation, and combinations with last-resort antibiotics. Our findings unveil that having a broad host range was a dominant trait of isolated phages, and increasing phage numbers in combination with antibiotics significantly enhanced the suppression of bacterial growth. The decreased incidence of bacterial infection was explained by a reduction in pathogen density and emergence of bacterial resistance. Furthermore, phage-antibiotic synergy (PAS) demonstrated considerable broad-spectrum antibacterial potential against different clades of clinical MDR K. pneumoniae pathogens. The improved treatment outcomes of optimized PAS were also evident in a murine model, where mice receiving optimized PAS therapy demonstrated a reduced bacterial burden in mouse tissues. Taken together, these findings offer an important development in optimizing PAS therapy and its efficacy in the elimination of MDR K. pneumoniae pathogens., Importance: The worldwide spread of antimicrobial resistance (AMR) has posed a great challenge to global public health. Phage therapy has become a promising alternative against difficult-to-treat pathogens. One important goal of this study was to optimize the therapeutic efficiency of phage-antibiotic combinations, known as phage-antibiotic synergy (PAS). Through comprehensive analysis of the phenotypic and genotypic characteristics of a large number of CRKp-specific phages, we developed a systematic model for phage cocktail combinations. Crucially, our finding demonstrated that PAS treatments not only enhance the bactericidal effects of colistin and tigecycline against multidrug-resistant (MDR) K. pneumoniae strains in in vitro and in vivo context but also provide a robust response when antibiotics fail. Overall, the optimized PAS therapy demonstrates considerable potential in combating diverse K. pneumoniae pathogens, highlighting its relevance as a strategy to mitigate antibiotic resistance threats effectively., Competing Interests: The authors declare no conflict of interest.

Objectives: The emergence of carbapenem-resistant Klebsiella pneumoniae presents significant health challenges. Here, we present the structural genome sequence of an NDM-5-producing K. pneumoniae (HZKP2) in China., Methods: Antimicrobial susceptibility tests were conducted via broth microdilution. Whole-genome sequencing was performed for genomic analysis. Wzi and capsular polysaccharide (KL) were analysed using Kaptive. Resistance genes, virulence factors, and comparative genomics analyses were also conducted. Multilocus sequence typing (MLST), replicons type, and core genome MLST analysis were further conducted using BacWGSTdb server., Results: HZKP2 was resistant to cefepime, ceftazidime, ciprofloxacin, ciprofloxacin, meropenem, and ertapenem. It harboured fosA, bla SHV-187 , oqxA, oqxB, sul1, dfrA1, tet(A), floR, aph(6)-Id, aph(3'')-Ib, sul2, bla CTX-M-55 , and bla NDM-5 . Based on the RAST results, 5563 genes that belonged to 398 subsystems were annotated. The complete genome sequence of HZKP2 was characterized as ST1, wzi 19, and KL19, 5 five contigs totalling 5 654 446 bp, including one chromosome and four plasmids. Further analysis found that bla NDM-5 was located in a 46 161 bp IncX3 plasmid (pHZKP2-3). The genetic structure of bla NDM-5 gene was ISKox3-IS26-ble MBL -bla NDM-5 -IS5-ISAb125-IS3000. Further analysis revealed that insertion sequences mediated the dissemination of bla NDM-5 from other species of Enterobacterales. Phylogenetic analysis showed that the closest relative was from a human stool specimen in China, which differed by 53 core genome MLST alleles., Conclusions: Our study provides the first structural perspective of the ST1 K. pneumoniae isolate producing NDM-5 in China. These results could provide valuable insights into the genetic characteristics, antimicrobial resistance mechanisms, and transmission dynamics of carbapenem-resistant K. pneumoniae in clinical settings., Competing Interests: Competing interests None declared., (Copyright © 2024. Published by Elsevier Ltd.)

Purpose: Infection by carbapenem-resistant Klebsiella pneumoniae (CRKP) has high mortality. There is no clear optimal therapeutic choice for pneumonia caused by CRKP. The aim of this study was to compare the clinical outcomes and safety of the standard doses of polymyxin B-based regimens vs tigecycline-based regimens and to identify risk factors for mortality., Methods: This retrospective cohort study included patients with pneumonia caused by CRKP between January 1, 2020 and December 31, 2022. The primary outcomes were 7-day bacterial eradication rate and 14- and 28-day all-cause mortality. The secondary outcome was incidence of acute kidney injury., Results: Seventy-three patients were included in this study, 29 in the polymyxin B-based combination therapy group and 44 in tigecycline-based combination therapy group. There were no significant differences between the two groups in terms of the 7-day bacterial eradication rate (31.03% vs 20.45%, p  = 0.409), the 14-day all-cause mortality (37.93% vs 22.73%, p  = 0.160), and the incidence of acute kidney injury (14.29% vs 6.82%, p  = 0.526). The 28-day all-cause mortality in the polymyxin B-based therapy group was higher than in the tigecycline-based group (75.86% vs 45.45%, p  = 0.010). Binary logistic regression analysis revealed that male and previous use of carbapenems were independent factors associated with 28-day all-cause mortality for patients treated with polymyxin B ( p  < 0.05)., Conclusions: Polymyxin B-based combination therapy at the standard dose should be used with caution for patients with CRKP-induced pneumonia, especially for men who used carbapenems prior to CRKP detection.

Background: Klebsiella pneumoniae (KP) is the second most prevalent Gram-negative bacterium causing bloodstream infections (BSIs). In recent years, the management of BSIs caused by KP has become increasingly complex due to the emergence of carbapenem-resistant Klebsiella pneumoniae (CRKP). Although numerous studies have explored the risk factors for the development of CRKP-BSIs, the mortality of patients with KP-BSIs, and the molecular epidemiological characteristics of CRKP, the variability in data across different populations, countries, and hospitals has led to inconsistent conclusions. In this single-center retrospective observational study, we utilized logistic regression analyses to identify independent risk factors for CRKP-BSIs and factors associated with mortality in KP-BSI patients. Furthermore, a risk factor-based prediction model was developed. CRKP isolates underwent whole-genome sequencing (WGS), followed by an evaluation of microbiological characteristics, including antimicrobial resistance and virulence genes, as well as epidemiological characteristics and phylogenetic analysis., Results: Our study included a total of 134 patients with KP-BSIs, comprising 50 individuals infected with CRKP and 84 with carbapenem-susceptible Klebsiella pneumoniae (CSKP). The independent risk factors for CRKP-BSIs were identified as gastric catheterization (OR = 9.143; CI = 1.357-61.618; P = 0.023), prior ICU hospitalization (OR = 4.642; CI = 1.312-16.422; P = 0.017), and detection of CRKP in non-blood sites (OR = 8.112; CI = 2.130-30.894; P = 0.002). Multivariate analysis revealed that microbiologic eradication after 6 days (OR = 3.569; CI = 1.119-11.387; P = 0.032), high Pitt bacteremia score (OR = 1.609; CI = 1.226-2.111; P = 0.001), and inappropriate empirical treatment after BSIs (OR = 6.756; CI = 1.922-23.753; P = 0.003) were independent risk factors for the 28-day mortality in KP-BSIs. The prediction model confirmed that microbiologic eradication after 6.5 days and a Pitt bacteremia score of 4.5 or higher were significant predictors of the 28-day mortality. Bioinformatics analysis identified ST11 as the predominant CRKP sequence type, with bla KPC-2 as the most prevalent gene variant. CRKP stains carried multiple plasmid-mediated resistance genes along with some virulence genes. Phylogenetic analysis indicated the presence of nosocomial transmission of ST11 CRKP within the ICU., Conclusions: The analysis of risk factors for developing CRKP-BSIs and the association between KP-BSIs and 28-day mortality, along with the development of a risk factor-based prediction model and the characterization of CRKP strains, enhances clinicians' understanding of the pathogens responsible for BSIs. This understanding may help in the timely administration of antibiotic therapy for patients with suspected KP-BSIs, potentially improving outcomes., (© 2024. The Author(s).)

Background: Klebsiella pneumoniae is the most commonly encountered pathogen in clinical practice. Widespread use of broad-spectrum antibiotics has led to the current global dissemination of carbapenem-resistant K. pneumoniae, which poses a significant threat to antibacterial treatment efficacy and public health. Outer membrane vesicles (OMVs) have been identified as carriers capable of facilitating the transfer of virulence and resistance genes. However, the role of OMVs in carbapenem-resistant K. pneumoniae under external pressures such as antibiotic and phage treatments remains unclear., Methods: To isolate and purify OMVs under the pressure of phages and tigecycline, we subjected K. pneumoniae 0692 harboring plasmid-mediated bla NDM-1 and bla KPC-2 genes to density gradient separation. The double-layer plate method was used to isolate MJ1, which efficiently lysed K. pneumoniae 0692 cells. Transmission electron microscopy (TEM) was used to characterize the isolated phages and extract OMV groups for relevant morphological identification. Determination of protein content of each OMV group was conducted through bicinchoninic acid assay (BCA) and proteomic analysis., Results: K. pneumoniae 0692 released OMVs in response to different environmental stimuli, which were characterized through TEM as having the typical structure and particle size of OMVs. Phage or tigecycline treatment alone resulted in a slight increase in the mean protein concentration of OMVs secreted by K. pneumoniae 0692 compared to that in the untreated group. However, when phage treatment was combined with tigecycline, there was a significant reduction in the average protein concentration of OMVs compared to tigecycline treatment alone. Proteomics showed that OMVs encapsulated numerous functional proteins and that under different external stresses of phages and tigecycline, the proteins carried by K. pneumoniae 0692-derived OMVs were significantly upregulated or downregulated compared with those in the untreated group., Conclusions: This study confirmed the ability of OMVs to carry abundant proteins and highlighted the important role of OMV-associated proteins in bacterial responses to phages and tigecycline, representing an important advancement in microbial resistance research., (© 2024. The Author(s).)

The co-production of KPC and NDM carbapenemases in carbapenem-resistant Klebsiella pneumoniae (CRKP) complicates clinical treatment and increases mortality rates. The emergence of KPC-NDM CRKP is believed to result from the acquisition of an NDM plasmid by KPC CRKP, especially under the selective pressure of ceftazidime-avibactam (CZA). In this study, a CRKP-producing KPC-2 (JNP990) was isolated from a patient at a tertiary hospital in Shandong Province, China. Following sulfamethoxazole-trimethoprim (SXT) treatment, the isolate evolved into a strain that co-produces KPC and NDM (JNP989), accompanied by resistance to SXT (minimum inhibitory concentration >2/38 µg/mL) and CZA (dd ≤14 mm). Whole-genome sequencing and S1 nuclease pulsed-field gel electrophoresis revealed that JNP989 acquired an IncC plasmid (NDM plasmid) spanning 197 kb carrying sul1 and bla NDM-1 genes. The NDM plasmid could be transferred successfully into Escherichia coli J53 at a conjugation frequency of (8.70±2.47) × 10 -4 . The IncFⅡ/IncR plasmid carrying the bla KPC-2 gene in JNP990 could only be transferred in the presence of the NDM plasmid at a conjugation frequency of (1.93±0.41) × 10 -5 . Five CRKP strains with the same resistance pattern as JNP989, belonging to the same clone as JNP989, with sequence type 11 were isolated from other patients in the same hospital. Two strains lost resistance to CZA due to the loss of the bla NDM-1 -carrying fragment mediated by insertion sequence 26. Plasmid stability testing indicated that the IncC plasmid was more stable than the bla NDM-1 genes in the hosts. This study describes the evolution of KPC-NDM CRKP and its spread in hospitalized patients following antibiotic treatment, highlighting the severity of the spread of resistance., (Copyright © 2024 The Authors. Published by Elsevier Ltd.. All rights reserved.)

This study aimed to assess the in vitro efficacy of ceftazidime-avibactam (CZA) in combination with various antimicrobial agents against carbapenem-resistant Klebsiella pneumoniae (CRKP). We selected 59 clinical CRKP isolates containing distinct drug resistance mechanisms. The minimum inhibitory concentrations (MICs) of meropenem (MEM), colistin (COL), eravacycline (ERA), amikacin (AK), fosfomycin (FOS), and aztreonam (ATM), both individually and in combination with CZA, were tested using the checkerboard method. The interactions of antimicrobial agent combinations were assessed by fractional inhibitory concentration index (FICI) and susceptible breakpoint index (SBPI). The time-kill curve assay was employed to dynamically evaluate the effects of these drugs alone and in combination format. In the checkerboard assay, the combination of CZA+MEM showed the highest level of synergistic effect against both KPC-producing and carbapenemase-non-producing isolates, with synergy rates of 91.3% and 100%, respectively. Following closely was the combination of FOS+CZA . For metallo-beta-lactamases (MBLs) producing strains, ATM+CZA displayed complete synergy, while the combination of MEM+CZA showed a synergy rate of only 57.14% for NDM-producing strains and 91.67% for IMP-producing strains. In the time-kill assay, MEM+CZA also demonstrated significant synergistic effects against the two KPC-2-producing isolates (Y070 and L70), the two carbapenemase-non-producing isolates (Y083 and L093), and the NDM-1-producing strain L13, with reductions in log 10 CFU/mL exceeding 10 compared to the control. Against the IMP-producing strain Y047, ATM+CZA exhibited the highest synergistic effect, resulting in a log 10 CFU/mL reduction of 10.43 compared to the control. The combination of CZA and MEM exhibited good synergistic effects against KPC-producing and non-enzyme-producing strains, followed by the FOS+CZA combination. Among MBL-producing strains, ATM+CZA demonstrated the most pronounced synergistic effect. However, the combinations of CZA with ERA, AK, and COL show irrelevant effects against the tested clinical isolates., Importance: Our study confirmed the efficacy of the combination CZA+MEM against KPC-producing and non-carbapenemase-producing strains. For metalloenzyme-producing strains, CZA+ATM demonstrated the most significant synergy. Additionally, CZA exhibited a notable synergy effect when combined with FOS. These combination therapies present promising new options for the treatment of CRKP infection., Competing Interests: The authors declare no conflict of interest.

Carbapenem-resistant Klebsiella pneumoniae (CRKP) poses immense threats to the health of infected patients worldwide, especially children. This study reports the infection caused by CRKP in a paediatric intensive care unit (PICU) child and its drug-resistant mutation during the treatment. Twelve Klebsiella pneumoniae carbapenemase (KPC)-producing K. pneumoniae strains were isolated from the child. Broth microdilution method, plasmid transformation assay, and whole genome sequencing (WGS) were performed to investigate the antimicrobial susceptibility, resistance mechanisms, and genetic structural features of CRKPs. The results showed that 12 strains were highly resistant to most available antimicrobial agents. Among them, K. pneumoniae FD11 and K. pneumoniae FD12 were resistant to ceftazidime-avibactam (CZA, MIC >64 mg/L) and restored the carbapenem susceptibility (Imipenem, MIC =0.25 mg/L; Meropenem, MIC =2 mg/L). The patient improved after treatment with CZA in combination with aztreonam. Plasmid transformation assay demonstrated that the bla KPC-33 -positive transformant increased MICs of CZA by at least 33-fold and 8-fold compared with the recipient Escherichia coli DH5α and bla KPC-2 -positive transformants. WGS analysis revealed that all strains belonged to the ST11-KL64 type and showed highly homologous (3-26 single nucleotide polymorphisms [SNPs]). A single base mutation (G532T) of bla KPC-2 resulted in a tyrosine to aspartic acid substitution at Ambler amino acid position 179 (D179Y), which conferred CZA resistance in K. pneumoniae. This is the first report of a drug-resistant mutation evolving into bla KPC-33 during the treatment of bla KPC-2 -positive CRKP in paediatric-infected patients. It advises clinicians that routine sequential antimicrobial susceptibility testing and KPC genotyping are critical during CZA therapy in children infected with CRKP., (Copyright © 2024 Elsevier Ltd and International Society of Antimicrobial Chemotherapy. All rights reserved.)

Objectives: To investigate the clinical characteristics and molecular epidemiology of CRKP infection in neonatal patients in a children's hospital in China from 2017 to 2021., Methods: Species identification and antibiotic susceptibilities were tested with matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) and VITEK 2 systems. The clinical data were collected from medical records. Carbapenem-resistant Klebsiella pneumoniae (CRKP) isolates were investigated by antimicrobial susceptibility testing, carbapenemase genes and multilocus sequence typing., Results: Six kinds of resistant genes and 23 STs were detected. Bla NDM-1 (n=83, 55.3%) was the predominant carbapenemase gene, followed by bla KPC-2 (n=45, 30.0%), bla NDM-5 (n=7, 4.7%), bla IMP-38 (n=6, 4.0%). Bla NDM-1 was predominant in 2017 and 2018, whereas bla KPC-2 increased in 2019 and became the predominant gene from 2020 to 2021. ST11 accounted for most infections (n=35, 23.3%), followed by ST278 (n=23, 15.3%), ST17 (n=17, 11. 3%) and ST2735 (n=16, 10.7%). ST278 and ST17 were predominant in 2017 and 2018, whereas ST11 increased in 2019 and became the predominant sequence type from 2020 to 2021. Compared with bla NDM-1 , the CRKP strains producing bla KPC-2 were characterized by high resistance to gentamicin, amikacin and levofloxacin and the change trend of drug resistance rate before and after COVID-19 was consistent with that of bla NDM-1 and bla KPC-2 ., Conclusions: The main sequence type of CRKP infection changed dynamically from ST278-NDM-1 to ST11-KPC-2 during the years 2017-2021 in the newborns. Antibiotic exposure and the prevalence of COVID-19 since 2020 may have led to changes in hospital population and lead to the changes., (© 2023. The Author(s), under exclusive licence to Springer Nature Switzerland AG.)

Background: Ceftazidime-avibactam is a treatment option for carbapenem-resistant gram-negative bacilli (CR-GNB) infections. However, the risk factors associated with ceftazidime-avibactam (CAZ-AVI) treatment failure in kidney transplant (KT) recipients and the need for CAZ-AVI-based combination therapy remain unclear., Methods: From June 2019 to December 2023, a retrospective observational study of KT recipients with CR-GNB infection treated with CAZ-AVI was conducted, with the primary outcome being 30-day mortality and secondary outcomes being clinical cure, microbiological cure, and safety. Risk factors for 30-day mortality and clinical failure were also investigated., Results: A total of 81 KT recipients treated with CAZ-AVI were included in this study. Forty recipients (49.4%) received CAZ-AVI monotherapy, with a 30-day mortality of 22.2%. The clinical cure and microbiological cure rates of CAZ/AVI therapy were 72.8% and 66.7%, respectively. CAZ-AVI alone or in combination with other medications had no effect on clinical cure or 30-day mortality. Multivariate logistic regression analysis revealed that a higher Acute Physiology and Chronic Health Evaluation (APACHE) II score (odds ratio [OR]: 4.517; 95% confidence interval [CI]: 1.397-14.607; P = 0.012) was an independent risk factor for 30-day mortality. Clinical cure was positively associated with the administration of CAZ-AVI within 48 hours of infection onset (OR: 11.009; 95% CI: 1.344-90.197; P =0.025) and negatively associated with higher APACHE II scores (OR: 0.700; 95% CI: 0.555-0.882; P =0.002). Four (4.9%) recipients experienced recurrence within 90 days after the initial infection, 3 (3.7%) recipients experienced CAZ-AVI-related adverse events, and no CAZ-AVI resistance was identified., Conclusion: CAZ-AVI is an effective medication for treating CR-GNB infections following kidney transplantation, even as monotherapy. Optimization of CAZ/AVI therapy (used within 48 hours of infection onset) is positively associated with potential clinical benefit. Further larger-scale studies are needed to validate these findings., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Zhang, Li, Zhong, Ding, Liao and Liang.)

Aims: Carbapenem-resistant Klebsiella pneumonia (CRKP) is a global threat that varies by region. The global distribution, evolution, and clinical implications of the ST11 CRKP clone remain obscure., Methods: We conducted a multicenter molecular epidemiological survey using isolates obtained from 28 provinces and municipalities across China between 2011 and 2021. We integrated sequences from public databases and performed genetic epidemiology analysis of ST11 CRKP., Results: Among ST11 CRKP, KL64 serotypes exhibited considerable expansion, increasing from 1.54% to 46.08% between 2011 and 2021. Combining our data with public databases, the phylogenetic and phylogeography analyses indicated that ST11 CRKP appeared in the Americas in 1996 and spread worldwide, with key clones progressing from China's southeastern coast to the inland by 2010. Global phylogenetic analysis showed that ST11 KL64 CRKP has evolved to a virulent, resistant clade with notable regional spread. Single-nucleotide polymorphism (SNP) analysis identified BMPPS (bmr3, mltC, pyrB, ppsC, and sdaC) as a key marker for this clade. The BMPPS SNP clade is associated with high mortality and has strong anti-phagocytic and competitive traits in vitro., Conclusions: The high-risk ST11 KL64 CRKP subclone showed strong expansion potential and survival advantages, probably owing to genetic factors., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Carbapenem-resistant Klebsiella pneumoniae (CR-Kp) is a significant threat to public health worldwide. The primary reservoir for CR-Kp is the intestinal tract. There, the bacterium is usually present at low density but can bloom following antibiotic treatment, mostly in hospital settings. The impact of disturbances in the intestinal environment on the fitness, survival, expansion, and drug susceptibility of this pathogen is not well-understood, yet it may be relevant to devise strategies to tackle CR-Kp colonization and infection. Here, we adopted an in vivo model to examine the transcriptional adaptation of a CR-Kp clinical isolate to immune activation in the intestine. We report that as early as 6 hours following host treatment with anti-CD3 antibody, CR-Kp underwent rapid transcriptional changes including downregulation of genes involved in sugar utilization and amino acid biosynthesis and upregulation of genes involved in amino acid uptake and catabolism, antibiotic resistance, and stress response. In agreement with these findings, treatment increased the concentration of oxidative species and amino acids in the mouse intestine. Genes encoding for proteins containing the domain of unknown function (DUF) 1471 were strongly upregulated, however their deletion did not impair CR-Kp fitness i n vivo upon immune activation. Transcription factor enrichment analysis identified the global regulator cAMP-Receptor Protein, CRP, as a potential orchestrator of the observed transcriptional signature. In keeping with the recognized role of CRP in regulating utilization of alternative carbon sources, crp deletion in CR-Kp resulted in strongly impaired gut colonization, although this effect was not amplified by immune activation. Thus, following intestinal colonization, which occurs in a CRP-dependent manner, CR-Kp can rapidly respond to immune cues by implementing a well-defined and complex transcriptional program whose direct relevance toward bacterial fitness warrants further investigation. Additional analyses utilizing this model may identify key factors to tackle CR-Kp colonization of the intestine.

Objective: To investigate the prevalence of a tet(A) gene variant and its role in developing high-level tigecycline resistance among carbapenem-resistant Klebsiella pneumoniae (CRKP) clinical isolates., Methods: The mechanism of high-level tigecycline resistance in CRKP mediated by a tet(A) variant was explored by induction experiments, antimicrobial susceptibility testing, whole-genome sequencing and bioinformatics analysis. The amplification and overexpression of the tet(A) variant were measured by the determination of sequencing depth, gene copy numbers, and qRT-PCR., Results: A high rate (62.1%, 998/1607) of tet(A) variant carriage was observed among 1607 CRKP clinical isolates from Henan Province, China. High-level tigecycline resistance could rapidly develop by the amplification of the tet(A) variant in these isolates. The analysis of the raw sequencing data and the plasmid mapping depth revealed that the ΔtnpA homologous sequence of Tn1721 supports the amplification of the region that harbours the tet(A) variant by forming a large number of repeat arrays through translocatable units (TUs). Moreover, the epidemiological analysis of tet(A) variant-carrying structures among 1607 clinical CRKPs showed that the TU structure is widely present., Conclusion: The presence of a tigecycline resistance-mediating tet(A) variant in CRKP clinical isolates represents a greater health concern than initially thought and should be monitored consistently., (Copyright © 2023 European Society of Clinical Microbiology and Infectious Diseases. Published by Elsevier Ltd. All rights reserved.)

Objective: This study (NCT05588531) aimed to evaluate the safety and pharmacokinetics of cefepime-avibactam (YK-1169) in healthy Chinese subjects and explore the optimal regimen for treating carbapenem-resistant Klebsiella pneumoniae (CRKP) based on the pharmacokinetic/pharmacodynamic evaluation., Methods: YK-1169 single-ascending doses (0.5, 1.25, 2.5 or 3.75 g, 2-h infusion) and multiple doses (2.5 or 3.75 g every 8 h [q8h], 2-h infusion) given for 7 days were evaluated in pharmacokinetic studies. Subjects were randomized to receive cefepime (2 g), avibactam (0.5 g) or YK-1169 (2.5 g) to assess drug-drug interactions. The minimum inhibitory concentrations (MICs) of YK-1169 were determined by the broth microdilution method. Monte Carlo simulation was used to evaluate 10 different dose regimens., Results: Cefepime and avibactam both showed a linear pharmacokinetic profile. No accumulation was found after multiple doses. The cefepime C max,ss and AUC 0-∞,ss were 9.20 and 16.0 μg/mL, 407.2 and 659.45 μg·h/mL in the 2.5 and 3.75 g multiple-dose groups, respectively. The avibactam C max,ss and AUC 0-∞,ss were 0.545 and 0.837 μg/mL, 53.31 and 79.55 μg·h/mL in the 2.5 and 3.75 g multiple-dose groups, respectively. Cefepime and avibactam did not affect each other's pharmacokinetics. No serious adverse events occurred. All regimens achieved 90% probability of target attainment (PTA) goals when the MIC was ≤8 mg/L. The regimens of 2.5 (q8h, 2-h infusion), 3.75 (q8h, 2-, 3- and 4-h infusions) and 7.5 g (24-h continuous infusion) reached a 90% cumulative fraction of response., Conclusion: YK-1169 had good antibacterial activity against CRKP and could be an option for CRKP infections. The regimen of 2.5 g q8h intravenously guttae (ivgtt) 2 h should be considered in future clinical trials., (© 2023 British Pharmacological Society.)

Background & Objectives: To determine the effect of Wen Run Fei Ning formula (WRFNF) intervention in class I integron-mediated carbapenem-resistant Klebsiella pneumoniae., Methods: A drug-susceptibility test and PCR amplification were used to screen for carbapenem-resistant K. pneumoniae containing class I integrons. Following nasal drip and tail vein injection to infect healthy male rats with carbapenem-resistant K. pneumoniae, three models were created: control (group A); model (group B, tail vein injection); and model-WRFNF treatment group (group C, by tail vein injection). Rats in Group C were gavaged with pre-warmed WRFNF extract. On the third, fifth, and seventh days after the experiment, the rats in groups A and B were gavaged with an equal quantity of saline and killed in batches., Results: Group C showed considerably higher serum IL-6 and TNF- levels on days 3, 5, and 7 compared to group A, as well as a significant increase in peripheral blood leukocyte count and a histopathologic inflammatory cell infiltration of the lungs. As the WRFNF delivery duration was prolonged, group C's histopathologic inflammatory cell infiltration gradually improved in contrast to group B, with the biggest improvement occurring on day 7. Compared to group B, group C's serum IL-6 and TNF- levels were lower. When the trial's duration was increased to 7 days, the levels of IL-6 and TNF- in group C decreased on day 7 compared to on day 5., Interpretation & Conclusion: WRFNF decreased inflammatory cell infiltration as well as IL-6 and TNF expression in the lung of the rats infected with carbapenem-resistant K. pneumoniae., Competing Interests: None

Background: Carbapenem-resistant Klebsiella pneumoniae (CRKP) is one of the serious forms of health care-associated infection. Pan-drug resistant (PDR) CRKP infections can cause severe infections. Mortality and treatment costs in the pediatric intensive care unit (PICU) are high. This study aims to share our experience regarding the treatment of oxacillinase (OXA)-48-positive PDR-CRKP infection in our 20-bed tertiary PICU with isolated rooms and 1 nurse for every 2-3 patients. Methods: Patient demographic characteristics, underlying diseases, previous infections, source of infection PDR-CRKP, treatment modalities, measures used, and outcomes were recorded. Findings: Eleven patients (eight men and three women) were found to have PDR OXA-48-positive CRKP. Because of the simultaneous detection of PDR-CRKP in three patients and the rapid spread of the disease, it was classified as a clinical outbreak, and strict infection control measures were taken. Combination therapy with double carbapenemase (meropenem and imipenem), amikacin, colistin, and tigecycline was used for treatment. The mean duration of treatment and isolation was 15.7 and 65.4 days, respectively. No treatment-related complication was observed, only one patient died, and the mortality rate was 9%. Conclusions: This severe clinical outbreak can be successfully treated with effective treatment with combined antibiotics and strict adherence to infection control measures. ClinicalTrial.gov ID: 28/01/2022 - 1/5.

Concerns regarding carbapenem-resistant Klebsiella pneumoniae (CR-Kp), especially in bloodstream infections (BSIs), are continuing to increase worldwide. Several novel agents with activity against BSI CR-Kp have been approved or are in late-stage clinical development. In this study, the antibacterial effects of ceftazidime/avibactam (CZA), aztreonam/avibactam (AZA), meropenem/vaborbactam (MEV), imipenem-cilastatin/relebactam (ICR) and eravacycline (ERV) against three colistin-resistant CR-Kp (COLR-Kp) and four CZA-resistant CR-Kp (CZAR-Kp) were tested by time-kill assay. Klebsiella pneumoniae ATCC® BAA-1705TM was used as a control strain. Two COLR-Kp isolates carried the blaKPC-2 gene and four CAZR-Kp isolates carried metallo-β-lactamase genes. The results revealed that ERV resulted in re-growth of seven tested isolates. CZA and MEV showed a bactericidal effect against isolates harbouring blaKPC-2. ICR reduced the population of six isolates to >5 log10 CFU/mL compared with the initial count. AZA showed a bactericidal effect (>5 log10 CFU/mL) against seven isolates and a bacteriostatic effect (<3 log10 CFU/mL) against one CZAR-Kp isolate. Therefore, AZA and ICR are effective therapeutic candidates for COLR-Kp and CZAR-Kp isolates., Competing Interests: Declaration of Competing Interests None declared., (Copyright © 2021 The Authors. Published by Elsevier Ltd.. All rights reserved.)

We assessed the prevalence of polymyxin B (PMB)- and tigecycline (TGC)-heteroresistant Klebsiella pneumoniae isolates and investigated the combined effect of PMB and TGC against dual-heteroresistant K. pneumoniae. Ninety-five nonduplicated carbapenem-resistant K. pneumoniae (CRKP) clinical isolates were collected from a tertiary-care teaching hospital in China. PCR was used to detect the resistant genes among the CRKP isolates. Population analysis profiling (PAP) was carried out to evaluate the existence of heteroresistance. A time-kill assay of PMB combined with TGC was conducted against heteroresistant K. pneumoniae strains. Real-time PCR was performed to determine the pmrA , phoP , and acrB expression levels. Among them, 74 isolates (77.9%) were susceptible to TGC, and 90 isolates (94.7%) were susceptible to PMB. In addition, of the TGC-susceptible isolates, 49 strains (66.2%) exhibited heteroresistant phenotypes. All of the PMB-susceptible isolates showed heteroresistant phenotypes. Forty-six isolates (48.4%) were heteroresistant to both TGC and PMB. All of the isolates carried the bla KPC gene, and one strain carried both bla KPC and bla NDM genes. The time-kill assay revealed in four isolates that early bactericidal activity could be triggered by the combination of PMB and TGC, and there was no regrowth, even at a relatively lower concentration (0.125 mg/liter PMB with 1 mg/liter TGC). Upregulated expression of pmrA , phoP , and acrB indicated that heteroresistance could be related to two-component systems and the AcrAB-TolC efflux pump. The combination of PMB and TGC may be a treatment strategy for those infected with CRKP heteroresistant to PMB and/or TGC. IMPORTANCE Tigecycline and colistin are two of the last treatment options remaining for carbapenem-resistant Enterobacteriaceae . Unfortunately, tigecycline resistance and colistin heteroresistance are also increasing rapidly. In the current study, we identified a high prevalence of heteroresistance to both PMB and TGC among clinical isolates of carbapenem-resistant K. pneumoniae (CRKP). The resistant subpopulations could survive pressure from TGC or PMB but were killed by the combination at a relatively low dose. It is proposed that the combination of PMB and TGC may be a treatment strategy for patients who are infected with CRKP heteroresistant to PMB or TGC.

Background: Hypermucoviscous carbapenem-resistant Klebsiella pneumoniae (hmCRKp) is emerging globally and approaching the worst-case scenario in health care system. Aims: The main objective in this study was to determine the hypermucoviscous characteristics among the carbapenem-resistant K. pneumoniae (CRKp) isolated from a teaching hospital in Malaysia. The association of hypermucoviscous phenotype with the virulence traits and clinical presentations were also investigated. Methods: A retrospective study was conducted in University Malaya Medical Centre (UMMC). The presence of hypermucoviscous K. pneumoniae was identified among a collection of CRKp clinical isolates (first isolate per patient) from 2014 to 2015 using string test. Correlation between clinical and microbial characteristics of the hmCRKp was investigated. Results: A total of nine (7.5%) hmCRKp were detected among 120 CRKp isolates. Majority of the isolates were hospital acquired or health care-associated infections. None of the patients had typical pyogenic liver abscess. All of the hmCRKp isolates harbored carbapenemase genes and were multidrug resistant. K1/K serotype, peg-344 , allS , and magA were not identified among hmCRKp isolates, whereas aerobactin siderophore receptor gene ( iutA ), iroB , rmpA , and rmpA2 were detected. Only three hmCRKp isolates were resistant to serum bactericidal. Conclusions: All the isolates presented inconclusive evidence for the interpretation of hypervirulence. Therefore, more study should be performed in the future to have a better understanding of the virulence mechanisms in correlation with the clinical and microbial determinants.

Tigecycline is an alternative antibiotic for managing carbapenem-resistant Gram-negative bacterial infections. However, disk diffusion and automated testing often show false-intermediate or false-resistant results in tigecycline susceptibility, misleading clinical antimicrobial therapy. Broth microdilution (BMD) is the reference method for testing tigecycline susceptibility, but it is labor intensive and time consuming to perform in clinical laboratories. Therefore, a simple and accurate method is urgently needed. We evaluated the performance of VITEK 2, E-test, Kirby-Bauer disk diffusion (KB), and modified KB disk diffusion (mKB) versus BMD in testing tigecycline susceptibility of 372 strains of carbapenem-resistant Klebsiella pneumoniae (CRKP) and 346 strains of carbapenem-resistant Acinetobacter baumannii (CRAB). BMD confirmed that 96.8% of CRKP and 91% of CRAB strains were susceptible to tigecycline. E-test, VITEK 2, KB, and mKB yielded categorical agreement of 96.7/59.3%, 69.9/54.3%, 78.5/87.3%, and 96.5%/91% for CRKP/CRAB, respectively. No very major error was found for either CRKP or CRAB by any method. No major error was found for CRKP or CRAB by the mKB method. The mKB method enhanced by R-buffer is simple, accurate, and inexpensive for clinical laboratories to test the susceptibility of CRKP and CRAB isolates to tigecycline.

Capsular polysaccharide (CPS) heterogeneity within carbapenem-resistant Klebsiella pneumoniae (CR- Kp ) strain sequence type 258 (ST258) must be considered when developing CPS-based vaccines. Here, we sought to characterize CPS-specific antibody responses elicited by CR- Kp -infected patients. Plasma and bacterial isolates were collected from 33 hospital patients with positive CR- Kp cultures. Isolate capsules were typed by wzi sequencing. Reactivity and measures of efficacy of patient antibodies were studied against 3 prevalent CR- Kp CPS types ( wzi29 , wzi154 , and wzi50 ). High IgG titers against wzi154 and wzi50 CPS were documented in 79% of infected patients. Patient-derived (PD) IgGs agglutinated CR- Kp and limited growth better than naive IgG and promoted phagocytosis of strains across the serotype isolated from their donors. Additionally, poly-IgG from wzi50 and wzi154 patients promoted phagocytosis of nonconcordant CR- Kp serotypes. Such effects were lost when poly-IgG was depleted of CPS-specific IgG. Additionally, mice infected with wzi50 , wzi154 , and wzi29 CR- Kp strains preopsonized with wzi50 patient-derived IgG exhibited lower lung CFU than controls. Depletion of wzi50 antibodies (Abs) reversed this effect in wzi50 and wzi154 infections, whereas wzi154 Ab depletion reduced poly-IgG efficacy against wzi29 CR- Kp We are the first to report cross-reactive properties of CPS-specific Abs from CR- Kp patients through both in vitro and in vivo models. IMPORTANCE Carbapenem-resistant Klebsiella pneumoniae is a rapidly emerging public health threat that can cause fatal infections in up to 50% of affected patients. Due to its resistance to nearly all antimicrobials, development of alternate therapies like antibodies and vaccines is urgently needed. Capsular polysaccharides constitute important targets, as they are crucial for Klebsiella pneumoniae pathogenesis. Capsular polysaccharides are very diverse and, therefore, studying the host's capsule-type specific antibodies is crucial to develop effective anti-CPS immunotherapies. In this study, we are the first to characterize humoral responses in infected patients against carbapenem-resistant Klebsiella pneumoniae expressing different wzi capsule types. This study is the first to report the efficacy of cross-reactive properties of CPS-specific Abs in both in vitro and in vivo models.

The narrow therapeutic window of polymyxin B constrains its clinical use against the multidrug-resistant organisms (MDRO). A 45-year-old patient was suffering with bloodstream infection with high fever and received a combined treatment with polymyxin B and tigecycline. Therapeutic drug monitoring (TDM) was applied to polymyxin B to develop a personalized medication against MDRO. The dose adjustment of polymyxin B with TDM successfully alleviated the infection and reduced the incident of acute kidney injury as caused in case of the original doses of polymyxin B. TDM of polymyxin B represents a valid treatment to ensure the efficiency and safety.

Background: The emergence and transmission of tigecycline- and carbapenem-resistant Klebsiella pneumoniae (TCRKP) have become a major concern to public health globally. Here, we investigated the molecular epidemiology and mechanisms of tigecycline resistance in carbapenem-resistant K pneumoniae (CRKP) isolates., Methods: Forty-five non-duplicate CRKP isolates were collected from January 2017 to June 2019. We performed antimicrobial susceptibility tests, multilocus sequence typing (MLST), and pulsed-field gel electrophoresis (PFGE). PCR and DNA sequencing were performed for the detection and mutation analysis of acrR, oqxR, ramR, rpsJ, tet(A), and tet(X) genes, which are related to tigecycline resistance. The expression levels of efflux pump genes acrB and oqxB and their regulator genes rarA, ramA, soxS, and marA were assessed by quantitative real-time PCR., Results: The resistance rate to tigecycline in CRKP isolates was 37.8% (17/45). K pneumoniae ST307 was a predominant clone type (70.6%, 12/17) among the TCRKP isolates. The expression levels of acrB (P < .001) and marA (P = .009) were significantly higher in the tigecycline-resistant group than in the tigecycline-intermediate and tigecycline-susceptible groups. Increased expression of acrB was associated with marA expression (r = 0.59, P = .013)., Conclusions: We found that the activated MarA-induced overexpression of AcrAB efflux pump plays an important role in the emergence of tigecycline resistance in CRKP isolates., (© 2020 The Authors. Journal of Clinical Laboratory Analysis Published by Wiley Periodicals LLC.)

An emerging multidrug-resistant Klebsiella pneumoniae high-risk clone of sequence type 307 (ST307) has been increasingly reported worldwide. Here, we described the genomic characteristics of an IMP-38-producing ST307 K. pneumoniae strain and investigated the prevalence of bla IMP-38 among carbapenem-resistant Klebsiella pneumoniae isolates from a tertiary care hospital in central China. A total of 14 IMP-38-producing ST307 K. pneumoniae strains were identified from 2013 to 2016, with 13 strains isolated from patients with neonatal sepsis in the neonatal ward. PacBio and Illumina whole-genome sequencing analysis performed on a representative IMP-38-producing K. pneumoniae strain, WCGKP294, showed that it contained a circular chromosome and two plasmids. Carbapenemase gene bla IMP-38 is colocated with bla CTX-M-3 in transposon Tn 6382 on an IncHI5 plasmid (pWCGKP294-2). WCGKP294 harbors another IncFIB plasmid, pWCGKP294-1, carrying three copies of tandem-repeated IS 26 - bla SHV-2A - deoR - ygbJ - ygbK - fucA -IS 26 composite transposon elements. Phylogenetic analysis placed WCGKP294 in the global ST307 cluster, distant from the U.S. (Texas) and South Africa clusters. Nevertheless, WCGKP294 does not contain the chromosomal fluoroquinolone resistance-associated mutations and IncFIIK/IncFIBK plasmid-associated bla CTX-M-15 gene that are frequently found in other global ST307 strains. IMPORTANCE We described the genome and resistome characterization of a carbapenem-resistant Klebsiella pneumoniae ST307 strain carrying bla IMP-38 in China. This report highlights that the high-risk ST307 clone continues to acquire different antimicrobial resistance genes, posing significant challenges to clinical practice, and should be closely monitored., (Copyright © 2020 Wang et al.)

Background: Carbapenem-resistant Klebsiella pneumoniae (CRKP) is considered as a serious global threat. CRKPs occurred only sporadically in the Second Hospital of Shanxi Medical University. Our study aimed to investigate and control the first outbreak of CRKP in our hospital occurred between October 2017 and August 2019., Methods: The antimicrobial stewardship (AMS) workers have been implemented control measures properly. Clinical and epidemiological data were retrospectively collected from medical records. Carbapenemase genes were detected by modified carbapenem inactivation method (mCIM) test and the EDTA-modified carbapenem inactivation method (eCIM) test. Resistance genes were identified by polymerase chain reaction (PCR) and sequencing. Genetic relatedness was studied by multilocus sequence typing (MLST)., Results: During the outbreak, 31 patients were infected with CRKP isolates. 20 (64.5%) patients were infected with KPC-2 and/or NDM-1 producing K. pneumoniae. Mostly MLST-sequence types belonged to ST11 (21/31). The outbreak was two major K. pneumoniae clusters present in epidemiologically linked patients., Conclusions: Setting up AMS workers is potentially a highly efficient strategy for the successful control of the outbreak. A multimodal and multidisciplinary infection control strategy proved to be crucial. The emergence of CRKP in our hospital emphasizes the importance of continuous monitoring of these isolates, which helps to limit the spread of CRKPs and improve the level of management.

A preliminary study found that eugenol expressed an antibacterial activity against Klebsiella pneumoniae. However, the mechanism of action of eugenol against K. pneumoniae still remains unexplored. The aim of this study was to gain further insight into the antibacterial effect of eugenol against carbapenem-resistant Klebsiella pneumoniae (CRKP) and possible mode of action. Here, minimum inhibitory concentration (MIC) of eugenol against CRKP strains was determined using the agar dilution method. Moreover, variations in intracellular ATP concentration, intracellular pH (pH in ), membrane potential and membrane integrity were measured to evaluate the effect of eugenol on cell membrane. Besides, changes in cell structure and biofilm formation of CRKP as well as biofilm-associated cell damage were determined using field emission scanning electron microscope (FESEM), transmission electron microscope (TEM) and confocal laser scanning microscopy (CLSM). Finally, gene expression of biofilm-related biosynthesis was investigated. The results showed that MICs of eugenol against four tested CRKP were 0.2 mg/mL. Eugenol damaged the cell membrane of CRKP, as evidenced by decreased intracellular ATP concentration, reduced pH in and cell membrane hyperpolarization, coupled with enhanced membrane permeability. Furthermore, eugenol compromised cell structure and induced loss of intracellular components of CRKP. Additionally, eugenol inhibited biofilm formation and inactivated biofilm CRKP cells. Finally, eugenol presented strong inhibitory effects on biofilm formation and biofilm-associated gene expression, and inactivated CRKP cells growing in biofilms. These findings suggest that eugenol exhibits antimicrobial effect against CRKP strains and could be potentially used to control CRKP-related infections., Competing Interests: Declaration of competing interest The authors declare no conflict of interest., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

Aim: To evaluate the antimicrobial effect of paeoniflorin against carbapenem-resistant Klebsiella pneumoniae (CRKP)., Methods and Results: Minimum inhibitory concentration (MIC) of paeoniflorin against CRKP was determined by agar dilution method. Changes in intracellular ATP concentration, intracellular pH (pHin), cell membrane potential and membrane integrity were investigated to assess the influence of paeoniflorin on cell membrane damage. Additionally, alterations in cell structure of CRKP cells and cell damage within biofilms were examined. The results indicated that paeoniflorin was effective against CRKP at MIC of 1·2 mg ml -1 . Paeoniflorin destroyed the integrity of CRKP cell membrane, as was confirmed by decrease of intracellular ATP, pHin, membrane potential, as well as distinctive alteration in cell morphology, resulting in leakage of CRKP intracellular components. Moreover, paeoniflorin displayed a markedly inhibitory influence on biofilm formation of CRKP and inactivated CRKP cells within biofilms., Conclusions: Paeoniflorin shows promise as an effective antibiotic against CRKP with the potential to be an alternative therapeutic agent., Significance and Impact of the Study: This study provides information about the potential use of paeoniflorin to reduce the infection of CRKP in clinical practice and food production., (© 2019 The Society for Applied Microbiology.)

Carbapenem-resistant Klebsiella pneumoniae (CRKP) has become a serious challenge for clinical treatment and public health. We found that both KPC-2-producing K. pneumoniae (KPC-KP) and NDM-1-producing K. pneumoniae (NDM-KP) are epidemic in a teaching hospital in Northeast China. The main aim of the present study was to compare antimicrobial susceptibility differences between KPC-KP and NDM-KP and elucidate complex resistant genotypes of the KPC-KP and NDM-KP by PCR and sequencing. Among 82 CRKP isolated between January 2015 and December 2016, 59 isolates were KPC-KP and 23 isolates were NDM-KP. All 59 KPC-KP had no susceptibility to gentamicin, tobramycin, levofloxacin, and ciprofloxacin, had very low susceptibility to amikacin (3.39%) and fosfomycin (8.47%), whereas the susceptibility of NDM-KP to the above antibiotics was 21.74%, 13.04%, 17.39%, 17.39%, 69.57%, and 73.91%, respectively. Although the susceptibility of NDM-KP to tigecycline (95.65%) and polymyxin B (73.91%) was higher than that of KPC-KP (84.75% and 69.49%, respectively), the difference was not statistically significant. The MIC 90 of KPC-KP and NDM-KP to aztreonam-avibactam were 4 and 2 μg/mL, respectively. All 82 CRKP carried 2 or 3 Extended Spectrum Beta-Lactamase (ESBL) genes, and 79/82 CRKP carried the AmpC gene bla FOX . The aminoglycoside resistance gene rmtB was detected in 96.61% of KPC-KP and in 21.74% of NDM-KP. It seems that KPC-KP was more resistant to antibiotics than NDM-KP in this study, so that available therapeutic regimens against KPC-KP are very limited. Aztreonam-avibactam may be a promising and valuable option against both KPC-KP and NDM-KP.

Objectives: The aim was to investigate the activity of ceftazidime/avibactam (CAZ/AVI) against carbapenem-resistant Klebsiella pneumoniae (CRKP) and identify the resistance mechanisms before CAZ/AVI coming to Chinese market., Methods: Clinical CRKP isolates were continuously collected from 36 tertiary hospitals in China from 1 March 2017 to 31 July 2017. CAZ/AVI MICs were determined by agar dilution method. CAZ/AVI resistant isolates were submitted to whole genome sequencing. The copy number and relative expression of bla KPC were determined by quantitative PCR., Results: A total of 872 CRKP isolates were collected, and MIC 50 and MIC 90 of CAZ/AVI were 4 and 8 mg/L. The resistant rate of CAZ/AVI was 3.7% (32/872). Among the resistant isolates, 53.1% (17/32) were metallo-β-lactamase-producing K. pneumoniae (MBL-KP), 40.6% (13/32) were Klebsiella pneumoniae carbapenemase (KPC)-producing K. pneumoniae (KPC-KP) and 6.3% (2/32) produced both MBL and KPC. One of the KPC-KP with high level CAZ/AVI resistance (>128 mg/L) harboured mutated bla KPC-2 (D179Y). In 12 wild-type bla KPC-2 isolates, the relative copy number and expression of bla KPC-2 gene were 2.5-fold and 2.7-fold higher than that in the CAZ/AVI MIC ≤0.5 mg/L group (p < 0.05), and when added avibactam at a fixed concentration of 8 mg/L, 91.7% (11/12) isolates could restore susceptibility., Conclusions: Resistance against CAZ/AVI in CRKP emerged before clinical use of CAZ/AVI in China, although most of the CRKP isolates maintained the susceptibility. MBL production, bla KPC-2 point mutation and high KPC expression played an important role in CAZ/AVI resistance., (Copyright © 2019 European Society of Clinical Microbiology and Infectious Diseases. Published by Elsevier Ltd. All rights reserved.)

The purpose of this study was to investigate the synergistic and bactericidal effects of combinations of colistin with meropenem or amikacin in vitro and provide laboratory data needed for development of therapeutic strategies for the treatment of carbapenem-resistant Klebsiella pneumoniae (CRKP) infection. We found that minimum inhibitory concentration (MIC) of colistin, meropenem and amikacin were 2~32, 4~256, and 1~16384 μg/ml, respectively. The minimum bactericidal concentration of the antibiotics was either 1× or 2×MIC. Treatments of 6 CRKP isolates at 1 μg/ml colistin completely killed 2 of them and suppressed 4 others growth. 4 CRKP isolates at 16 μg/ml meropenem or amikacin completely killed and suppressed 2 others growth. 2 CRKP isolates showed synergic effects in all colistin combination and 3 CRKP isolates showed synergic effects in part of colistin combination. Our data suggest that colistin in combination with either meropenem or amikacin could be a valid therapeutic option against colistin-resistant CRKP isolates. Moreover, the combination of colistin-amikacin is less expensive to treat CRKP infections in Eastern Heilongjiang Province., (Copyright © 2019 Yu, Zhang, Fu, Zhao, Wang, Zhao, Guo, Li and Zhang.)

Objective: To evaluate the efficacy and safety of different antimicrobial regimens in patients with bloodstream infections caused by carbapenem-resistant Klebsiella pneumoniae (CRKP). Methods: The clinical date of patients with CRKP bloodstream infections were retrospectively analyzed at the First Affiliated Hospital of Zhejiang University Medical College between January 2017 and January 2018. All subjects were separated into three groups based on antibiotics regimens over 72 hours, including meropenem 2.0 g every 8 hours, tigecycline 200 mg as initial dose and 100 mg every 12 hours, and polymyxin B 1.25 mg/kg every 12 hours as salvage treatment of tigecycline. Results: A total of 86 patients were finally recruited, including 14, 52 and 20 patients in groups of meropenem, tigecycline and polymyxin B salvage, respectively. All of the strains were resistant to meropenem and susceptible to tigecycline and polymyxin B initially, while 2 of them became resistant to tigecycline during treatment. The 28-day mortality was significantly higher in meropenem group (13/14) than that in tigecycline group and polymyxin B salvage group (61.5%, 32/52) and (12/20), respectively ( P< 0.01), while as no significant difference was seen in the last two groups (χ(2)=0.014, P> 0.05). The incidences of hepatic impairment [3.8%(2/52) vs. 1/20] and renal dysfunction (0 vs . 1/20) between tigecycline group and polymyxin B salvage group were both comparable ( P> 0.05). Conclusion: The meropenem-based therapy is not recommended for CRKP-related bloodstream infections. Tigecycline-based therapy is still disappointing despite salvage use of polymyxin B after 72 hours. Hepatic and nephretic toxicities caused by additional polymyxin B are acceptable.

Purpose: To highlight the clinical significance of carbapenem-resistant Klebsiella pneumoniae (CRKP) rectal colonization by examining the risk factors for CRKP rectal colonization and subsequent bloodstream infection (BSI) in critically ill patients., Methodology: Prospective study of CRKP rectal colonization in an intensive care unit (ICU) during a 39-month period. CRKP strains isolated from both the blood cultures and corresponding rectal specimens (n=96) of patients were screened by PCR for the presence of antibiotic resistance-associated genes. Molecular analyses were conducted to investigate the clonal relatedness of CRKP strains from the rectal and blood specimens., Results: Among the 498 patients, 226 were rectally colonized by CRKP, 48 of whom developed a CRKP BSI. The median time from hospital admission to the detection of CRKP rectal colonization was 8 days, while the median time from colonization to BSI was 4 days. The duration of ICU stay, patient/nurse ratio and prior use of antianaerobic antimicrobials were associated with CRKP rectal colonization. No specific factor was associated with BSIs in the colonized patients. The blaKPC-2 gene was detected in all 96 strains, which were all classified as sequence type ST-258. Representative pairs (n=48) of CRKP strains colonizing and infecting the same patient shared the same pulsotype., Conclusion: Our results indicate that hospitalized patients become infected with their colonizing strains, supporting the strong association between colonization and BSI. Limiting antianaerobic antimicrobial administration, reducing the duration of ICU stay and maintaining a low patient/nurse ratio are possible strategies to restrict rectal CRKP colonization in ICUs.

Objective: To determine whether infection-prevention and control (IPC) interventions can reduce the colonisation and infection of intensive care unit (ICU)-acquired carbapenem-resistant Klebsiella pneumoniae (CRKP) in a general ICU ward in China., Methods: We used a quasi-experimental before-and-after study design. The study was conducted in 4 stages: baseline period, January 2013-June 2013; IPC interventions period including de-escalation and targeted bundle interventions, July 2013-June 2014; modified IPC interventions period, July 2014-June 2015; and follow-up period, July 2015-June 2016. We used modified de-escalation interventions according to patient-risk assessments to prevent the transmission of CRKP., Results: A total of 629 patients were enrolled in study. The incidence of ICU-acquired CRKP colonisation/infection was 10.08 (4.43-16.43) per 1000 ICU patient-days during the baseline period, and significantly decreased early during the IPC interventions, but the colonisation/infections reappeared in April 2014. During the modified IPC intervention and follow-up periods, the incidence of ICU-acquired CRKP colonisations/infections reduced to 5.62 (0.69-6.34) and 2.84 (2.80-2.89), respectively, with ongoing admission of cases with previously acquired CRKP. The incidence of ICU-acquired CRKP catheter-related bloodstream infections decreased from 2.54 during the baseline period to 0.41 during the follow-up period. The incidence of ventilator-associated pneumonia and skin and soft tissue infections showed a downward trend from 2.84 to 0.41 and from 3.4 to 0.47, respectively, with slight fluctuations., Conclusions: Comprehensive IPC interventions including de-escalation and targeted bundle interventions showed a significant reduction in ICU-acquired CRKP colonisations/infections, despite ongoing admission of patients colonised/infected with CRKP., Competing Interests: The study protocol was approved by the Institutional Ethic Committees of the Shanghai Jiao Tong University School of Medicine and Ruijin Hospital (2013 NO. 29) and written consent was obtained from all participating patients.Not applicable.The authors declare that they have no competing interests.Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Background: Carbapenem-resistant Klebsiella pneumoniae (CR-KP) infections in solid organ transplant patients are progressively increasing and are associated with worse outcomes, although potential risk factors and therapeutic strategies are still not well defined., Methods: We conducted a retrospective matched-pair analysis in which we compared 26 recipients CR-KP-positive after kidney transplantation (KT) with 52 CR-KP-negative patients transplanted in the same period, during a CR-KP outbreak that occurred in our hospital. Twenty-one patients (80%) received a combined antibiotic treatment. At the end of the follow-up, of the 26 CR-KP infected patients, 11 (42.3%) experienced at least one episode of re-infection, 9 (34.6%) remained colonized, and 6 (23.0%) had a symptomatic infection. Two of the 11 patients with re-infection died, while 9 were colonized at the end of the study., Results: A significantly better patient (P = .043) and graft (P < .001) survival was observed in CR-KP-negative patients. Univariate analysis identified the following variables as potential risk factors associated with CR-KP infection after KT: lower body mass index (P = .020); higher creatinine levels at post-transplant days 7 (P = .009), 15 (P = .026), and 30 (P = .019); longer hospital stay (P = .007); longer cold ischemia time (P = .004); delayed graft function (P = .020); and higher Clavien-Dindo score (P = .006)., Conclusion: The study confirmed that a CR-KP positivity may affect the outcome of a kidney transplant population. In severe CR-KP infections with sepsis, a combined antibiotic treatment seems to be advisable., (© 2017 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Background: The emergence of carbapenem-resistant Klebsiella pneumoniae strains is threatening antimicrobial treatment., Methods: Sixty-eight carbapenemase-producing K. pneumoniae strains isolated at Luigi Sacco University Hospital-ASST Fatebenefratelli Sacco (Milan, Italy) between 2012 and 2014 were characterised microbiologically and molecularly. They were tested for drug susceptibility and carbapenemase phenotypes, investigated by means of repetitive extra-genic palindromic polymerase chain reaction (REP-PCR), and fully sequenced by means of next-generation sequencing for the in silico analysis of multi-locus sequence typing (MLST), their resistome, virulome and plasmid content, and their core single nucleotide polymorphism (SNP) genotypes., Results: All of the samples were resistant to carbapenems, other β-lactams and ciprofloxacin; many were resistant to aminoglycosides and tigecycline; and seven were resistant to colistin. Resistome analysis revealed the presence of blaKPC genes and, less frequently blaSHV, blaTEM, blaCTX-M and blaOXA, which are related to resistance to carbapenem and other β-lactams. Other genes conferring resistance to aminoglycoside, fluoroquinolone, phenicol, sulphonamide, tetracycline, trimethoprim and macrolide-lincosamide-streptogramin were also detected. Genes related to AcrAB-TolC efflux pump-dependent and pump-independent tigecycline resistance mechanisms were investigated, but it was not possible to clearly correlate the genomic features with tigecycline resistance because of the presence of a common mutation in susceptible, intermediate and resistant strains. Concerning colistin resistance, the mgrB gene was disrupted by an IS5-like element, and the mobile mcr-1 and mcr-2 genes were not detected in two cases. The virulome profile revealed type-3 fimbriae and iron uptake system genes, which are important during the colonisation stage in the mammalian host environment. The in silico detected plasmid replicons were classified as IncFIB(pQil), IncFIB(K), ColRNAI, IncX1, IncX3, IncFII(K), IncN, IncL/M(pMU407) and IncFIA(HI1). REP-PCR showed five major clusters, and MLST revealed six different sequence types: 512, 258, 307, 1519, 745 and 101. Core SNP genotyping, which led to four clusters, correlated with the MLST data. Isolates of the same sequencing type often had common genetic traits, but the SNP analysis allowed greater strain tracking and discrimination than either the REP-PCR or MLST analysis., Conclusion: Our findings support the importance of implementing bacterial genomics in clinical medicine in order to complement traditional methods and overcome their limited resolution.

Background: Carbapenem-resistant Klebsiella pneumoniae (CRKP) spread and infections in patients with haematological malignancies are a serious concern especially in endemic areas. Treatment failures and delay in appropriate therapy for CRKP infections are frequent and the mortality rate associated with CRKP bacteremia in neutropenic haematological patients is reported about 60%., Methods: Haematological patients harboring CRKP hospitalized between February 2012 and May 2013 in an Italian Teaching hospital were examined. Conditions favouring CRKP spread in a haematological unit, risk factors for bacteremia in CRKP-carriers and for CRKP bacteremia-related death were evaluated in this observational retrospective study., Results: CRKP was isolated in 22 patients, 14 (64%) had bacteremia. Control measures implementation, particularly the weekly rectal screening for CRKP performed in all hospitalized patients and contact precautions for CRKP-carriers and newly admitted patients until proved CRKP-negative, reduced significantly the CRKP spread (14 new carriers identified of 131 screened patients vs 5 of 242 after the intervention, p = 0.001). Fifty-eight percent of carriers developed CRKP bacteremia, and acute myeloid leukemia (AML) resulted independently associated with the bacteremia occurrence (p = 0.02). CRKP bacteremias developed mainly during neutropenia (86%) and in CRKP-carriers (79%). CRKP bacteremias were breakthrough in 10 cases (71%). Ten of 14 patient with CRKP bacteremias died (71%) and all had AML. The 70% of fatal bacteremias occurred in patients not yet recognized as CRKP-carriers and 80% were breakthrough. Initial adequate antibiotic therapy resulted the only independent factor able to protect against death (p = 0.02)., Conclusions: The identification of CRKP-carriers is confirmed critical to prevent CRKP spread. AML patients colonized by CRKP resulted at high risk of CRKP-bacteremia and poor outcome and the adequacy of the initial antibiotic therapy may be effective to improve survival. To limit the increase of resistance, the extensive use of antibiotics active against CRKP should be avoided, but in the setting of high CRKP pressure and high-risk CRKP-colonized haematological patients, timely empiric antibiotic combinations active against CRKP could be suggested as treatment of febrile neutropenia.

Carbapenem-resistant Klebsiella pneumoniae (CRKP) has been known as a nosocomial pathogen, both for the last 10 years in Turkey and for 20 years worldwide. Due to limited treatment options and high mortality rates, despite improvements in the field of medicine at the present time, CRKP is still a big threat for public health. This study was carried out between the dates of January 2010 and September 2014. Patients ≥18 who were hospitalized for at least 72 h and who also had CRKP growth were included in the study as a case group. In the same period patients, who were hospitalized in the same ward and did not have CRKP growth were selected as the control group. It was determined that no glycopeptides and steroids use nor tracheostomy as protective factors would be employed in terms of non-development of CRKP. Mechanical ventilation, tracheostomy, urinary catheter presence, central venous catheterization, nasogastric tube placement, advanced age, acute renal insufficiency, total parenteral nutrition, carbapenem, glycopeptide, and piperacillin tazobactam were all detected as risk factors in terms of CRKP infection development. As a result, rational usage of antibiotics for preventing infections developing with CRKP should be targeted.

Available therapeutic options against carbapenem-resistant Klebsiella pneumoniae (CR-Kp) are limited because of the high level of resistance to other antimicrobial classes including polymyxins. The double-carbapenem regimen has been recently considered a possible therapeutic strategy. In the present study, we evaluated the in vitro bactericidal and synergistic activity of a double-carbapenem regimen consisting of ertapenem plus high-dose meropenem in a series of patients with healthcare-associated CR-Kp infections in whom the use of colistin was not indicated because of potential nephrotoxicity and/or resistance. In vitro synergy was evaluated using checkerboard and killing studies. A total of 15 patients were included in the study, with sepsis, severe sepsis and septic shock found in two (13.3%), five (33.3%) and one (6.7%) patients, respectively. Overall, the clinical/microbiological response was 12/15 (80%). Synergy was observed in 11/14 (78.6%) isolates using the checkerboard method whereas in killing studies 12/14 (85.7%) and 14/14 (100%) strains were synergistic and bactericidal at 24 h at concentrations of 1 × MIC MEM+1 × MIC ERT and 2 × MEM+1 × MIC ERT, respectively, with a significant decrease of log CFU/mL compared with other combinations (p <0.0001). The double-carbapenem regimen showed clinical and in vitro effectiveness in patients with CR-Kp infections., (Copyright © 2015 European Society of Clinical Microbiology and Infectious Diseases. Published by Elsevier Ltd. All rights reserved.)

Carbapenem-resistant Klebsiella pneumoniae (CRKP) is emerging as a public health problem worldwide. In Italy, a remarkable increase in CRKP cases has been reported since 2010. In this study, CRKP diffusion, distribution and in-hospital transmission trends were evaluated in a university hospital in Milan, Italy, from January 2012 to December 2013. Isolates from 63 newly detected CRKP-positive patients were genotyped, and possible transmission was determined by combining the molecular results with data concerning the patients' admission and in-hospital transfers. Most of the cases (90.4%) were from general medical and surgery wards, and the remaining 9.6% were from the intensive care unit. Fifteen of the 46 hospital-associated cases (32.6%) were attributable to in-hospital transmission. After the introduction of targeted and hospital-wide control measures, the transmission index significantly decreased from 0.65 to 0.13 (p=0.01). There was also a decrease in the overall nosocomial case incidence, from 0.37 to 0.17 per 1000 person-days (p=0.07). Our findings indicate that the spread of CRKP in Northern Italy hospitals may go far beyond high-risk settings (i.e., intensive care units) and that strict surveillance should be extended to general areas of care., (Copyright © 2015 King Saud Bin Abdulaziz University for Health Sciences. Published by Elsevier Ltd. All rights reserved.)

Background: Although high mortality associated with carbapenem-resistant Klebsiella pneumoniae (CRKP) bacteremia has been well described, the epidemiology and outcomes of nonbacteremic infection are unknown., Methods: Medical charts of adults hospitalized for CRKP pneumonia or urinary tract infection between January 2011 and December 2013 were reviewed retrospectively for relevant demographic and clinical details. Cases were matched to controls (non-carbapenem-resistant, non-extended-spectrum beta-lactamase [ESBL]-producing K pneumoniae [NRKP]) by the primary site of infection and year of isolation and compared in terms of risk of acquisition and outcomes., Results: The CRKP and NRKP arms (n = 48 each) were elderly (median age, 74 years). Compared with controls, more patients in the CRKP arm resided in skilled nursing/long-term acute care facilities (77% vs 29%; P < .01), had a chronic tracheostomy (29% vs 0%; P < .001), decubitus ulcers (69% vs 17%; P < .01), higher Acute Physiology and Chronic Health Evaluation (APACHE) II scores (median, 21.5 vs 14; P = .02), and required intensive care unit admission (54% vs 31%; P = .04). More patients in the CRKP arm had previous ESBL infection (23% vs 6%; P = .04), and this arm had at least a 10-fold greater risk of coinfection with other carbapenem-resistant pathogens (44% vs 4%; P < .01), as well as a 7-fold greater likelihood of previous carbapenem therapy (23% vs 4%; P = .01). Patients in the CRKP arm had prolonged hospitalization (median, 13 days) and a 32% rate of readmission within 30 days of discharge., Conclusions: CRKP nonbacteremic infections occur in debilitated patients and are associated with frequent previous carbapenem exposure and high resource utilization, underscoring the need to focus efforts on antimicrobial stewardship and infection control., (Copyright © 2015 Association for Professionals in Infection Control and Epidemiology, Inc. Published by Elsevier Inc. All rights reserved.)