Search

Klebsiella pneumoniae is among the most important Gram-negative pathogens that can cause serious nosocomial infections. The emergence and prevalence of hypervirulent carbapenem-resistant K. pneumoniae (Hv-CRKP) pose a significant challenge to public health. In this study, we characterized thirty carbapenem-resistant K. pneumoniae (CRKP) strains from a tertiary care hospital in Sichuan province, China, by whole-genome sequencing and genome analysis. These strains were all highly resistant to carbapenem but remained susceptible to tigecycline. Of the 30 tested CRKP strains, 23 were positive for blaKPC−2 and seven for blaNDM−5. These blaKPC−2-positive strains all belonged to ST11, while blaNDM−5-positive strains belonged to five distinct STs. Phylogenetic analysis revealed a predominant intra-hospital transmission of ST11-KL64 in KPC-2-producing CRKP, and that both clonal and horizontal transmission of blaNDM−5 have occurred among NDM-5-producing CRKP strains in this hospital. Hypervirulence genes were commonly detected in the CRKP. The prevalent pLVKP-like plasmid and ICEKp seem to have contributed largely to the transmission of virulence genes in them. blaNDM−5 was located on highly similar IncX3 plasmids in the collected strains, and its truncated vision was highlighted. blaKPC−2 was primarily carried by IncFII/IncR plasmids in our collection. At least two IncFII/IncR plasmid subtypes were identified, exhibiting high similarity to many previously reported blaKPC−2-bearing plasmids from different parts of China. The findings provide an expanded knowledge of the genetic characteristics of CRKP, the transmission pattern of carbapenem-resistance genes, and also the convergence of Hv-CRKP. [ABSTRACT FROM AUTHOR]

Abstract Klebsiella pneumoniae is among the most important Gram-negative pathogens that can cause serious nosocomial infections. The emergence and prevalence of hypervirulent carbapenem-resistant K. pneumoniae (Hv-CRKP) pose a significant challenge to public health. In this study, we characterized thirty carbapenem-resistant K. pneumoniae (CRKP) strains from a tertiary care hospital in Sichuan province, China, by whole-genome sequencing and genome analysis. These strains were all highly resistant to carbapenem but remained susceptible to tigecycline. Of the 30 tested CRKP strains, 23 were positive for bla KPC−2 and seven for bla NDM−5. These bla KPC−2-positive strains all belonged to ST11, while bla NDM−5-positive strains belonged to five distinct STs. Phylogenetic analysis revealed a predominant intra-hospital transmission of ST11-KL64 in KPC-2-producing CRKP, and that both clonal and horizontal transmission of bla NDM−5 have occurred among NDM-5-producing CRKP strains in this hospital. Hypervirulence genes were commonly detected in the CRKP. The prevalent pLVKP-like plasmid and ICEKp seem to have contributed largely to the transmission of virulence genes in them. bla NDM−5 was located on highly similar IncX3 plasmids in the collected strains, and its truncated vision was highlighted. bla KPC−2 was primarily carried by IncFII/IncR plasmids in our collection. At least two IncFII/IncR plasmid subtypes were identified, exhibiting high similarity to many previously reported bla KPC−2-bearing plasmids from different parts of China. The findings provide an expanded knowledge of the genetic characteristics of CRKP, the transmission pattern of carbapenem-resistance genes, and also the convergence of Hv-CRKP.

Background: Carbapenem-resistant Klebsiella pneumoniae (CRKP) is a great public health problem and is associated with many disease outbreaks and high mortality rates. Alarmingly, K. pneumoniae has been isolated from food in several recent studies. This study aimed to investigate the prevalence and characteristics of CRKP in food samples from Egypt. Methods: A total of 311 food samples (including 116 minced meat, 92 chicken meat, 75 diced meat, and 28 mutton) were collected from local markets in Egypt and were screened for CRKP with the determination of their antimicrobial resistance profiles. The whole genome sequence was done for 23 CRKP isolates to clarify the relationship between CRKP from food and human cases in Egypt using the SNP core genome. The conjugation probability of the blaNDM-5 harboring plasmid was identified using oriTfinder Results: CRKP was isolated from 11% (35/311) of the samples, with 45.71% (16/35) of them showing resistance to colistin, one of the last-resort options for treating CRKP-mediated infections. In addition to the carbapenem and colistin resistance, the CRKP isolates frequently exhibited resistance to multiple antimicrobials including β-lactams, fluoroquinolones, aminoglycosides, tetracyclines, and chloramphenicol. In addition, most of the CRKP were potentially hypervirulent K. pneumoniae (HvKP) identified as phylogroup Kp1 and of high-risk groups as detected in STs reported in many human outbreaks globally, such as ST383 and ST147. The core-genome phylogeny showed similarities between the isolates from this study and those previously isolated from clinical human samples in Egypt. In addition, analysis of the plasmid on which blaNDM is encoded revealed that several antimicrobial resistance genes such as blaOXA-9, blaCTX-M-15, aac(6′)-Ib, qnrS1, and several virulence genes are encoded on the same plasmid. Conclusions: This study is significant for food safety and public health and is important to further identify the change in the epidemiology of CRKP infections, especially the consumption of contaminated food products.

BACKGROUND: K.pneumoniae is a gram-negative bacterium belonging to the Enterobacteriaceae family. This microorganism is a part of the healthy microbiome of individuals and colonizes many parts of the body. The incidence of carbapenemresistant Klebsiella pneumoniae (CRKP) infections is increasing globally. In India, around 65% of K. pneumoniae isolates are resistant to carbapenem antibiotics. The blaNDM is the predominant carbapenem-resistant gene in CRKP isolates. AIM AND OBJECTIVE: To Study the Molecular Characterization of blaNDM-1 Resistant genes in Carbapenem Resistance Klebsiella Pneumonia Clinical isolates. MATERIAL AND METHODS:This was a cross sectional study carried out for a period of 1 year i.e, during 2023 to 2024at K.G.M.U, Lucknow . Samples like urine, sputum, ET tube, pleural fluid, pus, CSF, blood and Ascitic fluid were included in this study. Carbapenemase resistance was detected phenotypically by MHT, mCIM and eCIM methods and genotypically by using PCR where the DNA extraction was done and the resistant gene blaNDM-1 was confirmed by PCR assay. RESULT:A total of 350 isolates of K. pneumoniae were collected from patients, with 127(36.28%) identified as CRKP. There were 30 (57.69%) isolates obtained from male and 22(42.30%) were obtained from female, while the maximum number of patients belong to the age group <50 years and maximum patients were engaged as Student/Housewife/Retired (44.09%) followed by Clerical/Shopkeeper/Farmer (36.22%) and others. In our study,it was also observed that, in phenotypic test 81.88% were positive for M-CIM while 77.95% were positive for E-CIM and in genotypic test there were 86.61 % positive for NDM-1. CONCLUSION: The study highlighted the existence of carbapenemase-producing K. pneumoniae, particularly in blaNDM-1, in patients with comorbidities. Our findings emphasize the importance of the molecular characterization of resistance-determinant-carrying bacterial pathogens as a part of infection control and prevention in hospital settings. [ABSTRACT FROM AUTHOR]

Carbapenem-resistant Klebsiella pneumoniae (CRKP) is a great public health problem and is associated with many disease outbreaks and high mortality rates. Alarmingly, K. pneumoniae has been isolated from food in several recent studies. This study aimed to investigate the prevalence and characteristics of CRKP in food samples from Egypt. A total of 311 food samples (including 116 minced meat, 92 chicken meat, 75 diced meat, and 28 mutton) were collected from local markets in Egypt and were screened for CRKP with the determination of their antimicrobial resistance profiles. The whole genome sequence was done for 23 CRKP isolates to clarify the relationship between CRKP from food and human cases in Egypt using the SNP core genome. The conjugation probability of the bla NDM-5 harboring plasmid was identified using oriTfinder CRKP was isolated from 11% (35/311) of the samples, with 45.71% (16/35) of them showing resistance to colistin, one of the last-resort options for treating CRKP-mediated infections. In addition to the carbapenem and colistin resistance, the CRKP isolates frequently exhibited resistance to multiple antimicrobials including β-lactams, fluoroquinolones, aminoglycosides, tetracyclines, and chloramphenicol. In addition, most of the CRKP were potentially hypervirulent K. pneumoniae (HvKP) identified as phylogroup Kp1 and of high-risk groups as detected in STs reported in many human outbreaks globally, such as ST383 and ST147. The core-genome phylogeny showed similarities between the isolates from this study and those previously isolated from clinical human samples in Egypt. In addition, analysis of the plasmid on which bla NDM is encoded revealed that several antimicrobial resistance genes such as bla OXA-9 , bla CTX-M-15 , aac(6′)-Ib, qnrS1, and several virulence genes are encoded on the same plasmid. This study is significant for food safety and public health and is important to further identify the change in the epidemiology of CRKP infections, especially the consumption of contaminated food products. [ABSTRACT FROM AUTHOR]

ABSTRACT Klebsiella pneumoniae is a common pathogen of nosocomial infection, which can cause pneumonia, urinary tract infection, cystitis, and bloodstream infections (BSIs). Here, we genetically characterize a novel carbapenem-resistant K. pneumoniae (CRKP) strain recovered from the blood of a 44-year-old male patient with severe acute necrotizing pancreatitis and septic shock in China. The strain is a ST45 K. pneumoniae with a novel serotype of K43, named 18SHX166. The susceptibility testing results showed that 18SHX166 was resistant to cephalosporin and carbapenems but still susceptible to ceftazidime-avibactam, quinolones, colistin, and amikacin. Genomic sequencing revealed that 18SHX166 contains three plasmids, namely pSHX166-Hv, pSHX166-KPC, and pSHX166-3. pSHX166-Hv harbored the iucABCD operon, encoding the siderophore of aerobactin. pSHX166-KPC harbored blaKPC-2 gene and possessed complete conjugative regions. The conjugation experiment verified pSHX166-KPC as a self-transmissible plasmid mediating the dissemination of antibiotic resistance, with a conjugation rate of 2.21 × 10−5. Additionally, the growth curve showed that 18SHX166 demonstrates a higher growth rate than the control strains. The characteristics of 18SHX166 indicate a potential high risk of clinical transmission.IMPORTANCEST45-K43 carbapenem-resistant Klebsiella pneumoniae isolate, 18SHX166, carries a carbapenem resistance plasmid and virulence plasmid. It has the characteristics of multidrug resistance, high transmissibility, and a fast growth rate, which could pose a threat to the control of antimicrobial resistance and clinical transmission, causing a severe challenge to public health.

ABSTRACT Klebsiella pneumoniae carbapenemase (KPC) variants can contribute to resistance to ceftazidime–avibactam (CZA) in Klebsiella pneumoniae (KP). However, two-copy KPC variant-mediated resistance to CZA has rarely been reported to date. Here, we aimed to clarify the evolutionary trajectory of CZA resistance driven by mutations in double-copy blaKPC-2 to blaKPC-189 carried by the tandem core structure (ISKpn6-blaKPC-ISKpn27-tnpR-IS26) during treatment of ST11 carbapenem-resistant K. pneumoniae (CRKP). The CZA-resistant KP strain carried double-copy blaKPC-189, a variant with alanine–threonine and aspartate–tyrosine substitutions at Ambler amino acid positions 172 (A172T) and 179 (D179Y) of blaKPC-2. Clone experiments confirmed that, compared with that of the wild-type blaKPC-2 clone strain, the minimum inhibitory concentration of CZA increased 16-fold in the blaKPC-189-mutant strain. Furthermore, protein structure analysis revealed the A172T and D179Y mutations of blaKPC-189 can have a direct effect on the binding affinity of CAZ and AVI for KPC. Sequence comparison revealed that blaKPC-189 was mutated in a double-copy format upon CZA exposure, which was carried by the IS26-mediated tandem core structure ISKpn27-blaKPC-ISKpn6. This tandem core structure apparently evolves in vivo during infection, although not by self-transferring, and multiple ISKpn27-blaKPC-ISKpn6 copy numbers could mediate transferable CZA resistance upon mobilization. In addition, compared with the wild-type blaKPC-2 gene, the blaKPC-189 gene had no fitness cost. In summary, our study highlighted the emergence of CZA-resistant blaKPC-189 variants in the ST11 clone and the presence of a double-copy blaKPC-189 in the IncFII-type plasmid, which is carried by a tandem core structure (IS26-ISKpn6-blaKPC-189-ISKpn27-tnpR-IS26).IMPORTANCETo date, ceftazidime–avibactam (CZA) resistance caused by double-copy Klebsiella pneumoniae carbapenemase (KPC) variants has not been elucidated. The multicopy forms of carbapenem resistance genes carried by the same plasmid are relatively rare in most carbapenem-resistant Enterobacteriaceae. In this study, we elucidate the evolutionary trajectory of CZA resistance in ST11 carbapenem-resistant K. pneumoniae harboring a double-copy blaKPC and provide new insights into the mechanisms of acquired resistance to CZA.

Carbapenems are effective and reliable β-lactam antibiotics often used as the last option for treating severe infections caused by MDR Gram-negative bacteria, including K. pneumoniae. Among the Enterobacteriaceae group, clinically significant carbapenemases are the Ambler molecular class A (blaKPC), class B (blaVIM, blaIMP, blaNDM-1) and class D (blaOXA-48-like) types. These types are commonly found in K. pneumoniae and are frequently associated with serious nosocomial infections and outbreaks leading to the emergence and distribution of multidrug-resistant (MDR) and carbapenemresistant Klebsiella pneumoniae (CRKP) that has become a global health threat. To study the molecular characterization of blaOXA-48 and blaNDM-1 Resistant genes in Carbapenem Resistance Klebsiella pneumonia clinical isolates. This was a cross sectional study carried out for a period of 1 year i.e, November 2022 to November 2023 period of at Index Medical College, Hospital & Research Centre (IMCHRC), Indore (M.P.), India. The study included all the patients of OPD and IPD. Samples like urine, sputum, ET tube, pleural fluid, pus, CSF, blood and ascitic fluid were included in this study. Carbapenemase resistance was detected phenotypically by MHT, mCIM and eCIM methods and genotypically by using RTPCR where the DNA extraction was done using the Qiagen DNA extraction kit and the resistant gene blaOXA-48 and blaNDM-1 was confirmed by RTPCR assay. A total of 430 clinical samples were screened, out of which 160 (37.20 %) samples showed growth of K. pneumoniae, while 270 (62.8 %) samples showed growth of bacteria other than K. pneumoniae or sterile samples. There were 69.37% (n=111) of total Klebsiella isolates, which were carbapenem resistant (suggestive of Carbapenemase producers) by Kirby-Bauer disc diffusion, but 56.76% (63/111) of Klebsiella isolates were confirmed for Carbapenemase production by real time PCR. In conclusion, prevalence of carbapenem resistance in the current study was observed to be 56% as confirmed by real time PCR. Male patients were predominant in our study, the most common age group being above 60 years in both the sexes. In this study our results indicate the phenotypic detection mCIM combined with eCIM showed high sensitivity and specificity to detect carbapenemase producing K.pneumoniae compared with MHT. [ABSTRACT FROM AUTHOR]

Introduction: Carbapenem-resistant hypervirulent Klebsiella pneumoniae (CR-HvKP) strains combining virulence and multidrug resistance (MDR) features pose a great public health concern. The aimof this study is to explore the evolutionary characteristics of virulence in CR-HvKP by investigating the genetic features of resistance and virulence hybrid plasmids. Methods: The resistance and virulence phenotypes were determined by using antimicrobial susceptibility testing and the mouse bacteremia infection model, respectively. Plasmid profiles were investigated by S1 nuclease pulsed-field gel electrophoresis (S1-PFGE) and Southern blotting, conjugation assay, and whole genome sequencing (WGS). Bioinformatics tools were used to uncover the genetic features of the resistance and virulence hybrid plasmids. Results: Two ST11-KL64 CRKP clinical isolates (KP18-3-8 and KP18-2079), which exhibited enhanced virulence compared with the classic CRKP, were detected positive for blaKPC-2 and rmpA2. The virulence level of the hypermucoviscous strain KP18-3-8 was higher than that of KP18-2079. S1-PFGE, Southern hybridization and WGS analysis identified two novel hybrid virulence plasmids in KP18-3-8 (pKP1838-KPC-vir, 228,158 bp) and KP18-2079 (pKP1838-KPC-vir, 182,326 bp), respectively. The IncHI1B/repB-type plasmid pKP1838-KPC-vir co-harboring blaKPC-2 and virulence genes (rmpA2, iucABCD and iutA) but lacking type IV secretion system could transfer into non-hypervirulent ST11 K. pneumoniae with the assistance of a helper plasmid in conjugation. The IncFII/IncR-type virulence plasmid pKP18-2079-vir may have been generated as a result of recombination between a typical pLVPK-like virulence plasmid and an MDR plasmid. Conclusion: Our studies further highlight co-evolution of the virulence and resistance plasmids in ST11-CRKP isolates. Close surveillance of such hybrid virulence plasmids in clinical K. pneumoniae should be performed. [ABSTRACT FROM AUTHOR]

Abstract Background The escalating challenge of Carbapenem-resistant Klebsiella pneumoniae (CRKP) in hospital-acquired pneumonia (HAP) is closely linked to the blaNDM-1 gene. This study explores the regulatory mechanisms of blaNDM-1 expression and aims to enhance antibacterial tactics to counteract the spread and infection of resistant bacteria. Methods KP and CRKP strains were isolated from HAP patients' blood samples. Transcriptomic sequencing (RNA-seq) identified significant upregulation of blaNDM-1 gene expression in CRKP strains. Bioinformatics analysis revealed blaNDM-1 gene involvement in beta-lactam resistance pathways. CRISPR-Cas9 was used to delete the blaNDM-1 gene, restoring sensitivity. In vitro and in vivo experiments demonstrated enhanced efficacy with Imipenem and Thanatin or Subatan combination therapy. Results KP and CRKP strains were isolated with significant upregulation of blaNDM-1 in CRKP strains identified by RNA-seq. The Beta-lactam resistance pathway was implicated in bioinformatics analysis. Knockout of blaNDM-1 reinstated sensitivity in CRKP strains. Further, co-treatment with Imipenem, Thanatin, or Subactam markedly improved antimicrobial effectiveness. Conclusion Silencing blaNDM-1 in CRKP strains from HAP patients weakens their Carbapenem resistance and optimizes antibacterial strategies. These results provide new theoretical insights and practical methods for treating resistant bacterial infections.

Background: Carbapenem-resistant Klebsiella pneumoniae (CRKP) has become a real threat to hospitals worldwide with significant morbidity and mortality. Aim: to determine the rate of CRKP in ICUs of Zagazig University hospitals (ZUHs) by confirmatory phenotypic and molecular methods Subjects and Methods: This prospective cross-sectional study included 86 K. pneumoniae isolates from hospitalized patients in ICUs. Carbapenem resistance was identified by VITEK2 COMPACT system and Carbapenemase activities were identified by Modified Hodge Test, (MHT), modified Carbapenem Inactivation Method (mCIM) and KPC/MBL and OXA-48 Confirm Kit (KMOC test). Carbapenemase-encoding genes (blaKPC, blaNDM, blaVIM, blaIMP, and blaOXA-48) were detected by Multiplex PCR. Results: 66.3% (57/86) of K. pneumoniae isolates were CRKP. Hospital acquired infection (HAI) accounts for 84.2% among CRKP isolates. The rate of resistance by MHT, mCIM and KMOC test for carbapenems was 49.1%, 86% and 80.7% for CRKP isolates respectively. The blaNDM-1, blaOXA-48 and blaKPC genes were detected at a rate of 31.6%, 24.6%, and 12.3% by PCR respectively. Coexistence of carbapenemases was detected in 15.7% of isolates. Resistance pattern of Eravacycline, Cefiderocol, Ceftazidime/avibactam and Ceftolozane/tazobactam was 52%, 65%, 82.2% and 98% respectively. Conclusion: CRKP was responsible for a significant number of HAI cases in the ICUs of ZUHs. Antibiotic resistance was shown to be widespread in our study. The identification of carbapenemases classes will be useful for the improvement of patient’s treatment and prognosis and for infections control measures in ICUs.

Background: The escalating resistance of Klebsiella pneumoniae, a prevalent pathogen in healthcare settings, especially its carbapenem-resistant K. pneumoniae (CRKP), to a wide array of antibiotics, notably ß-lactams, constitutes a formidable challenge for healthcare and global public health management. Methods: This research compared the resistance phenotypes and genomic profiles of CRKP and Non-CRKP isolates in a Beijing hospital, focusing on high-risk blaKPC-2 gene-bearing CRKP clones and the structure of mobile genetic elements facilitating their spread across hospital departments. Forty K. pneumoniae isolates were collected from various departments of the hospital and subjected to antimicrobial susceptibility testing and whole-genome sequencing to analyze their resistance phenotypes and genomic features. Results: The study revealed that among the 31 CRKP isolates, ST11 is the most common sequence type, with K47 and OL101 being the dominant capsule types, primarily observed in the respiratory department. In terms of antimicrobial susceptibility: 87.5% of the isolates exhibited multidrug resistance (MDR), with a high resistance rate of 30% against tigecycline. All CRKP isolates demonstrated resistance to multiple drug classes (=5 CLSI classes). Non-CRKP isolates also showed high resistance rates to minocycline and doxycycline (77.8%). the ST11-KL47-OL101 type emerged as the predominant clone among the CRKP isolates carrying the blaKPC-2 gene. This dominance appears to be mediated by the pKpnR03_2 plasmid, which harbors not only blaKPC-2 and rmtb but also gene clusters pertinent to iron transport and arsenic resistance. These isolates, clustering in the C3 clade of the phylogenetic tree, exhibited minor genetic variations and close evolutionary relationships, suggesting a plasmid-driven spread across various hospital departments. Conclusion: In summary, our study highlights the extensive spread of antibioticresistant K. pneumoniae across various departments in our hospital, with a particular emphasis on the dominant clonal proliferation of the ST11-KL47-OL101 CRKP strain. This finding underscores the significant role of plasmidmediated gene transfer in the evolution and dissemination of resistant strains within hospital environments. The study emphasizes the necessity for ongoing surveillance of antibiotic resistance and genomic analysis in hospital settings to effectively monitor and manage these challenges. [ABSTRACT FROM AUTHOR]

Background: The escalating challenge of Carbapenem-resistant Klebsiella pneumoniae (CRKP) in hospital-acquired pneumonia (HAP) is closely linked to the blaNDM-1 gene. This study explores the regulatory mechanisms of blaNDM-1 expression and aims to enhance antibacterial tactics to counteract the spread and infection of resistant bacteria. Methods: KP and CRKP strains were isolated from HAP patients' blood samples. Transcriptomic sequencing (RNA-seq) identified significant upregulation of blaNDM-1 gene expression in CRKP strains. Bioinformatics analysis revealed blaNDM-1 gene involvement in beta-lactam resistance pathways. CRISPR-Cas9 was used to delete the blaNDM-1 gene, restoring sensitivity. In vitro and in vivo experiments demonstrated enhanced efficacy with Imipenem and Thanatin or Subatan combination therapy. Results: KP and CRKP strains were isolated with significant upregulation of blaNDM-1 in CRKP strains identified by RNA-seq. The Beta-lactam resistance pathway was implicated in bioinformatics analysis. Knockout of blaNDM-1 reinstated sensitivity in CRKP strains. Further, co-treatment with Imipenem, Thanatin, or Subactam markedly improved antimicrobial effectiveness. Conclusion: Silencing blaNDM-1 in CRKP strains from HAP patients weakens their Carbapenem resistance and optimizes antibacterial strategies. These results provide new theoretical insights and practical methods for treating resistant bacterial infections. [ABSTRACT FROM AUTHOR]

Objectives: The aim of this study was to investigate the clinical and molecular characteristics of Klebsiella pneumoniae infection from a tertiary general hospital in Wuhan, China. Methods: From December 2019 to August 2022, 311 non-duplicate isolates of K. pneumoniae were collected from a tertiary hospital in Wuhan. These comprised 140 carbapenem-resistant K. pneumoniae (CRKP) isolates and 171 carbapenem-susceptible K. pneumoniae (CSKP) isolates. The clinical characteristics of patients with K. pneumoniae infection were retrospectively collected. Polymerase chain reaction (PCR) assays were used to identify the main carbapenem resistance genes, virulence genes and multi-locus sequence typing (MLST) profiles of the isolates, and the Galleria mellonella infection model was used to determine their virulence phenotypes. Results: Independent risk factors for CRKP infection were hypertension, neurological disorders, being admitted to the intensive care unit (ICU) and prior use of antibiotics. Patient with CRKP infection had higher mortality than those with CSKP infection (23.6% vs 14.0%, P < 0.05). One hundred and two sequence types (STs) were identified among the K. pneumoniae isolates, and the most prevalent ST type was ST11 (112/311, 36.0%). All of the ST11 isolates were CRKP. Among the 112 ST11 isolates, 105 (93.8%) harboured the carbapenem resistance gene blaKPC-2 (ST11-KPC-2), and of these isolates, 78 (74.3%, 78/105) contained all of the four virulence genes, namely rmpA, rmpA2, iroN and iucA, suggesting that these genes were widespread among the isolates responsible for K. pneumoniae infections. Conclusion: In this study, ST11-KPC-2 was responsible for most of the K. pneumoniae infection cases. Carbapenem resistance rather than the co-occurrence of the virulence genes rmpA, rmpA2, iroN and iucA was associated with K. pneumoniae infection-related mortality during hospitalisation. Furthermore, a high proportion of ST11-KPC-2 isolates carried all of the four virulence genes. [ABSTRACT FROM AUTHOR]

Background: Carbapenem-resistant Klebsiella pneumoniae (CRKP) has become a real threat to hospitals worldwide with significant morbidity and mortality. Aim: to determine the rate of CRKP in ICUs of Zagazig University hospitals (ZUHs) by confirmatory phenotypic and molecular methods Subjects and Methods: This prospective crosssectional study included 86 K. pneumoniae isolates from hospitalized patients in ICUs. Carbapenem resistance was identified by VITEK2 COMPACT system and Carbapenemase activities were identified by Modified Hodge Test, (MHT), modified Carbapenem Inactivation Method (mCIM) and KPC/MBL and OXA-48 Confirm Kit (KMOC test). Carbapenemase-encoding genes (blaKPC, blaNDM, blaVIM, blaIMP, and blaOXA-48) were detected by Multiplex PCR. Results: 66.3% (57/86) of K. pneumoniae isolates were CRKP. Hospital acquired infection (HAI) accounts for 84.2% among CRKP isolates. The rate of resistance by MHT, mCIM and KMOC test for carbapenems was 49.1%, 86% and 80.7% for CRKP isolates respectively. The blaNDM-1, blaOXA-48 and blaKPC genes were detected at a rate of 31.6%, 24.6%, and 12.3% by PCR respectively. Coexistence of carbapenemases was detected in 15.7% of isolates. Resistance pattern of Eravacycline, Cefiderocol, Ceftazidime/avibactam and Ceftolozane/tazobactam was 52%, 65%, 82.2% and 98% respectively. Conclusion: CRKP was responsible for a significant number of HAI cases in the ICUs of ZUHs. Antibiotic resistance was shown to be widespread in our study. The identification of carbapenemases classes will be useful for the improvement of patient's treatment and prognosis and for infections control measures in ICUs. [ABSTRACT FROM AUTHOR]

Objective　To analyze the epidemiological characteristics of an outbreak of carbapenem resistant Klebsiella pneumoniae（ CRKP） hospital infection in the neonatal intensive care unit（ NICU） ， and to explore possible causes of the outbreak. Methods Case analysis，epidemiological studies and environmental detection methods were used to investigate the detection of CRKP in the NICU of a hospital from March 11，2022 to May 3， 2022，and the causes and corresponding prevention and control measures were analyzed. Results This CRKP outbreak in the NICU occurred in two phases. The first phase was from March 11，2022，to March 30，2022, during which 3 people contracted CRKP hospital infections. The second phase was from April 13，2022，to May 3， 2022，during which 4 people contracted CRKP hospital infections. The source of the hospital infection outbreak was patient No.1. All 13 CRKP cases showed sensitivity only to amikacin and tigecycline, with intermediate sensitivity to levofloxacin, and high resistance to other antibiotics. All 13 CRKP cases carried resistant genes for metalloenzymes. After the outbreak, the ward environment was sampled three times，with 35 samples taken in total, and 14 of them tested positive for CRKP. After strict environmental cleaning, disinfection, and multiple control measures, no new hospital infection cases similar to this outbreak were detected after May 13，2022. Conclusion　The outbreak of NICU hospital infections may be due to the contamination of the ward environment， medical equipment, and inadequate hand hygiene of medical staff, leading to its spread. Therefore, strict cleaning and disinfection of the environmental surfaces， strengthening hand hygiene of medical staff， and implementing multiple measures can effectively control hospital infections of CRKP. [ABSTRACT FROM AUTHOR]

Background: Klebsiella pneumoniae is a causative agent of bacterial meningitis in adults. However, there is little information regarding this infection. Therefore, this study comprehensively analyzed the clinical characteristics and prognosis of Klebsiella pneumoniae meningitis (KPM) patients. Methods: The clinical data of adult hospitalized patients with KPM were retrospectively collected from January 2015 to December 2022. The clinical characteristics and antibiotic resistance of KPM were evaluated. Meanwhile, a set of logistic regression models was constructed to identify prognostic factors for death. These prognostic factors were subsequently combined to develop a nomogram for predicting the risk of in-hospital mortality in individual patients. Finally, the receiver operating characteristic curve and calibrate plot were utilized to verify the performance of the nomogram. Results: This study included 80 adult patients with KPM, 58 (72.5%) of whom were males. The mortality rate was 45%. Among them, 74 (92.5%) were diagnosed with healthcare-associated meningitis. Thirty-seven carbapenem-resistant Klebsiella pneumoniae (CRKP) strains were susceptible to tigecycline, polymyxin, and ceftazidime/avibactam. CRKP (OR = 9.825, 95%CI = 2.757–35.011, P

Chaoe Zhou,1 Liying Sun,2 Haixia Li,2 Lei Huang,2,&ast; Xinmin Liu1,&ast; 1Department of Geriatrics, Peking University First Hospital, Beijing, People’s Republic of China; 2Department of Clinical Laboratory, Peking University First Hospital, Beijing, People’s Republic of China&ast;These authors contributed equally to this workCorrespondence: Xinmin Liu, Department of Geriatrics, Peking University First Hospital, Beijing, 100034, People’s Republic of China, Tel +86-10-8357-2128, Email lxm2128@163.com Lei Huang, Department of Clinical Laboratory, Peking University First Hospital, Beijing, 100034, People’s Republic of China, Tel +86-10-8357-5546, Email leihuang2031@bjmu.edu.cnPurpose: Hospital-acquired pneumonia (HAP) caused by carbapenem-resistant K. pneumoniae (CRKP), especially in elderly patients, results in high morbidity and mortality. Studies on risk factors, mortality, and antimicrobial susceptibility of CRKP pulmonary infection among elderly patients are lacking.Patients and Methods: A retrospective case–control study was conducted from January 2019 to December 2021. The elderly inpatients (≥ 65 years) who were diagnosed with HAP caused by K. pneumoniae were enrolled. Clinical data were collected. Univariate and multivariate logistic regression analyses were used to identify risk factors. Propensity score matching was used to minimize the effect of potential confounding variables. Kaplan–Meier analysis was used to compare survival.Results: A total of 115 patients with CRKP infection and 78 patients with carbapenem-susceptible K. pneumoniae (CSKP) infection were recruited. There were four independent risk factors for CRKP infection: history of intensive care unit (ICU) stays from hospital admission to positive respiratory specimen culture for K. pneumoniae (odds ratio (OR)=2.530), Charlson comorbidity index score ≥ 3 (OR = 2.420), prior exposure to carbapenems (OR = 5.280), and prior K. pneumoniae infection or colonization in the preceding 3 years (OR = 18.529). The all-cause 30-day mortality was 22.3%, the mortality of CRKP and CSKP infection was 28.7% and 12.8%, respectively. Independent risk factors for mortality included: older age (OR = 1.107), immunocompromised patients (OR = 8.632), severe pneumonia (OR = 51.244), quick Sepsis-related Organ Failure Assessment (qSOFA) score ≥ 2 (OR = 6.187), exposure to tigecycline before infection (OR = 24.702), and prolonged ICU stay (OR = 0.987). Thirty-day mortality was significantly lower in patients receiving ceftazidime-avibactam (CAZ-AVI) containing regimens than patients receiving polymyxin B sulfate (PB) containing regimens (P = 0.048). qSOFA score had a good prognostic effect [area under receiver operating characteristic curve (AUROC) of 0.838].Conclusion: Active screening of CRKP for the high-risk populations, especially elderly patients, is significant for early detection and successful management of CRKP infection.Keywords: elderly, hospital-acquired pneumonia, CRKP, CSKP, ceftazidime-avibactam, qSOFA

Abstract To fight the global epidemic of drug-resistant bacteria, essential oils have gained increasing attention as a new source of antibiotics. The antimicrobial activity of Monarda didyma essential oils (MDEO) for the Carbapenem-resistant Klebsiella pneumoniae (CRKP) strains were determined by agar disc diffusion assay and broth microdilution assay. To further understand MDEO efficacy, a time-growth curve was performed. The biofilm formation of CRKP were determined by crystalline violet staining method, additionally, changes in intracellular Adenosine triphosphate (ATP), protein, Alkaline phosphatase (AKP) activities, and membrane integrity were investigated to assess the influence of MDEO on cell membrane damage. Finally, the activities of key enzymes in the tricarboxylic acid (TCA) pathways and pentose phosphate (PPP) pathways were examined to determine the effect of MDEO on the respiratory metabolism of CRKP. This study presents the antibacterial mechanism of MDEO against CRKP with a minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of 1.25 mg/ml. To understand MDEO efficacy, a time-kill kinetics approach was performed. The bactericidal effect of MDEO was evident at 2 h compared to the control at its MIC and 2MIC. Surface electron microscopic and ATP assay studies provided evidence for the multi-target action of MDEO against CRKP. MDEO could inhibit CRKP biofilm formation. MDEO could also cause irreversible damage to the CRKP cell membrane, resulting in the leakage of biological macromolecules (protein, ATP) and the reduction of intracellular enzymes (AKP) activities. Finally, MDEO affected the pathways of respiratory metabolism, such as PPP and TCA pathways. MDEO could reduce the activity of key enzymes (Glucose-6-phosphate dehydrogenase, citrate synthase, isocitrate dehydrogenase, and α-ketoglutarate dehydrogenase) in the PPP and TCA pathways to exert its biological effects against CRKP. These results suggest MDEO can exert inhibitory effects on CRKP, and potential mechanisms of action including inhibition of biofilm formation, damage of cell membrane structure and inhibition of energy metabolism.

Yibo Zhang,1,&ast; Yichen Wang,1,&ast; Zike Sheng,2,&ast; Qun Wang,1 Dake Shi,1 Shirui Xu,3 Yaping Ai,4 Erzhen Chen,5 Yumin Xu1,2 1Department of Hospital Infection Management, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, People’s Republic of China; 2Department of Infectious Diseases, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, People’s Republic of China; 3Department of Clinical Laboratory Medicine, Shanghai Fenglin Clinical Laboratory Co. Ltd, Shanghai, People’s Republic of China; 4Health Economics and Outcome Research, Becton & Dickinson Medical Device (Shanghai) Ltd, Shanghai, People’s Republic of China; 5Department of Emergency Medicine, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, People’s Republic of China&ast;These authors contributed equally to this workCorrespondence: Erzhen Chen, Department of Emergency Medicine, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, People’s Republic of China, Tel +86 021-64370045-665599, Email chenerzhen@hotmail.com Yumin Xu, Department of Infectious Diseases, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, People’s Republic of China, Tel +86 021-64370045-600653, Email xym121@163.comPurpose: Indwelling central venous catheters (CVCs) can cause catheter related bloodstream infection (CRBSI). CRBSI occurring in intensive care unit (ICU) patients may lead to the worse outcomes and extra medical costs. The present study aimed to assess the incidence and incidence density, pathogens and economic burden of CRBSI in ICU patients.Patients and Methods: A case-control study was retrospectively carried out in six ICUs of one hospital between July 2013 and June 2018. The Department of Infection Control performed routinely surveillance for CRBSI on these different ICUs. Data of the clinical and microbiological characteristics of patients with CRBSI, the incidence and incidence density of CRBSI in ICUs, the attributable length of stay (LOS), and the costs among patients with CRBSI in ICU were collected and assessed.Results: A total of 82 ICU patients with CRBSI were included into the study. The CRBSI incidence density was 1.27 per 1000 CVC-days in all ICUs, in which the highest was 3.52 per 1000 CVC-days in hematology ICU and the lowest was 0.14 per 1000 CVC-days in Special Procurement ICU. The most common pathogen causing CRBSI was Klebsiella pneumoniae (15/82, 16.67%), in which 12 (80%) were carbapenem resistant. Fifty-one patients were successfully matched with control patients. The average costs in the CRBSI group were &dollar; 67,923, which were significantly higher (P < 0.001) than the average costs in the control group. The total average costs attributable to CRBSI were &dollar;33, 696.Conclusion: The medical costs of ICU patients were closely related to the incidence of CRBSI. Imperative measures are needed to reduce CRBSI in ICU patients.Keywords: CRBSI, prolonged length of stay, cost, CRKP, ICU

BackgroundThe escalating resistance of Klebsiella pneumoniae, a prevalent pathogen in healthcare settings, especially its carbapenem-resistant K. pneumoniae (CRKP), to a wide array of antibiotics, notably β-lactams, constitutes a formidable challenge for healthcare and global public health management.MethodsThis research compared the resistance phenotypes and genomic profiles of CRKP and Non-CRKP isolates in a Beijing hospital, focusing on high-risk blaKPC-2 gene-bearing CRKP clones and the structure of mobile genetic elements facilitating their spread across hospital departments. Forty K. pneumoniae isolates were collected from various departments of the hospital and subjected to antimicrobial susceptibility testing and whole-genome sequencing to analyze their resistance phenotypes and genomic features.ResultsThe study revealed that among the 31 CRKP isolates, ST11 is the most common sequence type, with K47 and OL101 being the dominant capsule types, primarily observed in the respiratory department. In terms of antimicrobial susceptibility: 87.5% of the isolates exhibited multidrug resistance (MDR), with a high resistance rate of 30% against tigecycline. All CRKP isolates demonstrated resistance to multiple drug classes (≥5 CLSI classes). Non-CRKP isolates also showed high resistance rates to minocycline and doxycycline (77.8%). the ST11-KL47-OL101 type emerged as the predominant clone among the CRKP isolates carrying the blaKPC-2 gene. This dominance appears to be mediated by the pKpnR03_2 plasmid, which harbors not only blaKPC-2 and rmtb but also gene clusters pertinent to iron transport and arsenic resistance. These isolates, clustering in the C3 clade of the phylogenetic tree, exhibited minor genetic variations and close evolutionary relationships, suggesting a plasmid-driven spread across various hospital departments.ConclusionIn summary, our study highlights the extensive spread of antibiotic-resistant K. pneumoniae across various departments in our hospital, with a particular emphasis on the dominant clonal proliferation of the ST11-KL47-OL101 CRKP strain. This finding underscores the significant role of plasmid-mediated gene transfer in the evolution and dissemination of resistant strains within hospital environments. The study emphasizes the necessity for ongoing surveillance of antibiotic resistance and genomic analysis in hospital settings to effectively monitor and manage these challenges.

ABSTRACTCarbapenem-resistant Klebsiella pneumoniae (CRKP) is an important multidrug resistance (MDR) pathogen that threatens human health and is the main source of hospital-acquired infection. Outer membrane vesicles (OMVs) are extracellular vesicles derived from Gram-negative bacteria and contain materials involved in bacterial survival and pathogenesis. They also contribute to cellular communication to nearby or distant recipient cells and influence their functions and phenotypes. In this study, we sought to understand the mechanism of bacterial response to meropenem pressure and explore the relationship between pathogenic proteins and the high pathogenicity of bacteria. We performed whole-genome PacBio sequencing on a clinical CRKP strain, and its OMVs were characterized using nanoparticle tracking analysis, transmission electron microscopy, and proteomic analysis. Thousands of vesicle proteins have been identified in mass spectrometry-based high-throughput proteomics analyses of K. pneumoniae OMVs. Protein functionality analysis showed that the OMVs were predominantly involved in metabolic, intracellular compartments, nucleic acid binding, survival, defense, and antibiotic resistance, such as Chromosome partition protein MukB, 3-methyl-2-oxobutanoate hydroxymethyltransferase, methionine—tRNA ligase, Heat shock protein 60 family chaperone GroEL, and Gamma-glutamyl phosphate reductase. Additionally, a protein-protein interaction network demonstrated that OMVs from meropenem-treated K. pneumoniae showed the highest connectivity in DNA polymerase I, phenylalanine–tRNA ligase beta subunit, DNA-directed RNA polymerase subunit beta, methionine–tRNA ligase, DNA-directed RNA polymerase subunit beta, and DNA-directed RNA polymerase subunit alpha. The OMVs proteome expression profile indicates increased secretion of stress proteins released from meropenem-treated K. pneumoniae, which provides clues for revealing the biogenesis and pathophysiological functions of Gram-negative bacteria OMVs. The significant differentially expressed proteins identified in this study are of great significance for exploring effective control strategies for CRKP infection.IMPORTANCEMeropenem is one of the main antibiotics used in the clinical treatment of carbapenem-resistant Klebsiella pneumoniae (CRKP). This study demonstrated that some important metabolic changes occurred in meropenem-induced CRKP-outer membrane vesicles (OMVs), The OMVs proteome expression profile indicates increased secretion of stress proteins released from meropenem-induced Klebsiella pneumoniae. Furthermore, this is the first study to discuss the protein-protein interaction network of the OMVs released by CRKP, especially under antibiotic stress.

ABSTRACTGut microbiota plays a crucial role in providing colonization resistance against multi-drug resistant organisms including carbapenem-resistant Klebsiella pneumoniae (CRKP). However, the impact of different carbapenems on intestinal colonization resistance against CRKP remains poorly understood. In this study, we aimed to investigate the effects of three commonly used carbapenems (meropenem, imipenem, and ertapenem) administered at single-day doses, which are typically employed in emergency departments for managing infected patients, on CRKP gut colonization. We conducted experiments in mice by intravenously administering each carbapenem using human-simulated one-day regimens. The composition of the gut microbiota was analyzed using 16S rRNA amplicon sequencing before and after carbapenem administration. Our results revealed that all three carbapenems, when administered at a single-day dose significantly altered the abundance and diversity of the gut microbiota, leading to compromised colonization resistance against CRKP. Notably, the ertapenem and imipenem groups showed an increase in Allobaculum, Bifidobacterium, Enterobacteriaceae, while the meropenem and imipenem groups exhibited a decrease in Ruminococcaceae, S24-7, and Akkermania. Additionally, the Shannon index exhibited a negative correlation with both the number of days of CRKP colonization CFUs and the duration of bacteria shedding. Furthermore, the count of CRKP in mice after ertapenem administration on the first day and the duration of CRKP shedding were significantly lower compared to meropenem and imipenem. Predictive metabolic pathway analysis demonstrated that the three carbapenems similarly affected a range of metabolic pathways of gut microflora, including carbohydrate metabolism and vitamin B. Our findings emphasize that ertapenem, with its relatively narrow spectrum, minimizes perturbations of the gut microbiota and has a relatively less impact on gut colonization resistance against CRKP.IMPORTANCEThe intestinal colonization of carbapenem-resistant Klebsiella pneumoniae (CRKP) is an important source of clinical infection. Our research showed that even single-day dose use of carbapenems caused CRKP colonization and continuous bacterial shedding, which reminds clinical doctors to prescribe carbapenems cautiously. Whenever possible, ertapenem should be the preferred choice over other carbapenems especially when the identified or highly suspected pathogens can be effectively targeted by ertapenem.

Background: Carbapenem-resistant Klebsiella pneumoniae (CRKP) is epidemically transmitted globally, but few studies focused on the prevalence in district-level hospitals. In this study, we investigated CRKP strains collected from nine district hospitals from September 2019 to September 2020, aiming to determine the resistance mechanisms, virulence profiles, and molecular epidemiological characteristics of CRKP in district hospitals in Southwest China. Methods: A total of 51 CRKP strains were collected from 9 district-level hospitals. Matrix-assisted laser desorption/ionization-time of flight mass spectrometer was used for strain identification review, and the micro-broth dilution method was used for antibiotic sensitivity detection. Molecular epidemiological investigation of strains was performed by multilocus sequence typing (MLST) and pulsedfield gel electrophoresis (PFGE) methods. PCR and efflux pump inhibition tests were used to detect CRKP resistance mechanisms. PCR and serum killing tests were used to detect capsular serotype, virulence-related genes, and virulence validation. Results: The CRKP strains in district hospitals presented high levels of MIC50 and MIC90 in carbapenem antibiotics especially ertapenem and meropenem. A total of 90.2% (46/51) CRKP strains were detected as carbapenemase producers, and the proportion of strains co-expressing carbapenemases was 11.8% (6/51). All CRKP strains were grouped into eight MLST types, and ST11 was the most prevalent genotype. A total of 11.8% (6/51) CRKP isolates were positive for the string test, and three strains of hypervirulent and carbapenem-resistant K. pneumoniae (HV-CRKP) were positive in serum killing test. The molecular typing of all the CRKP isolates was grouped into 29 different PFGE patterns, and 40 ST11 isolates belonged to 20 different PFGE clusters. Conclusion: CRKP strains showed high-level antibiotic resistance and virulence phenotype in district hospitals in Southwest China, which suggested that we should immediately pay attention to the rapid dissemination of the CRKP in regional hospitals. Our study will provide new insights into the epidemiology of CRKP in regional hospitals, which will help regional hospitals develop nosocomial infection prevention and control policies tailored to local conditions. [ABSTRACT FROM AUTHOR]

To fight the global epidemic of drug-resistant bacteria, essential oils have gained increasing attention as a new source of antibiotics. The antimicrobial activity of Monarda didyma essential oils (MDEO) for the Carbapenem-resistant Klebsiella pneumoniae (CRKP) strains were determined by agar disc diffusion assay and broth microdilution assay. To further understand MDEO efficacy, a time-growth curve was performed. The biofilm formation of CRKP were determined by crystalline violet staining method, additionally, changes in intracellular Adenosine triphosphate (ATP), protein, Alkaline phosphatase (AKP) activities, and membrane integrity were investigated to assess the influence of MDEO on cell membrane damage. Finally, the activities of key enzymes in the tricarboxylic acid (TCA) pathways and pentose phosphate (PPP) pathways were examined to determine the effect of MDEO on the respiratory metabolism of CRKP. This study presents the antibacterial mechanism of MDEO against CRKP with a minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of 1.25 mg/ml. To understand MDEO efficacy, a time-kill kinetics approach was performed. The bactericidal effect of MDEO was evident at 2 h compared to the control at its MIC and 2MIC. Surface electron microscopic and ATP assay studies provided evidence for the multi-target action of MDEO against CRKP. MDEO could inhibit CRKP biofilm formation. MDEO could also cause irreversible damage to the CRKP cell membrane, resulting in the leakage of biological macromolecules (protein, ATP) and the reduction of intracellular enzymes (AKP) activities. Finally, MDEO affected the pathways of respiratory metabolism, such as PPP and TCA pathways. MDEO could reduce the activity of key enzymes (Glucose-6-phosphate dehydrogenase, citrate synthase, isocitrate dehydrogenase, and α-ketoglutarate dehydrogenase) in the PPP and TCA pathways to exert its biological effects against CRKP. These results suggest MDEO can exert inhibitory effects on CRKP, and potential mechanisms of action including inhibition of biofilm formation, damage of cell membrane structure and inhibition of energy metabolism. [ABSTRACT FROM AUTHOR]

Abstract Background Hospitals with their high antimicrobial selection pressure represent the presumably most important reservoir of multidrug-resistant human pathogens. Antibiotics administered in the course of treatment are excreted and discharged into the wastewater system. Not only in patients, but also in the sewers, antimicrobial substances exert selection pressure on existing bacteria and promote the emergence and dissemination of multidrug-resistant clones. In previous studies, two main clusters were identified in all sections of the hospital wastewater network that was investigated, one K. pneumoniae ST147 cluster encoding NDM- and OXA-48 carbapenemases and one VIM-encoding P. aeruginosa ST823 cluster. In the current study, we investigated if NDM- and OXA-48-encoding K. pneumoniae and VIM-encoding P. aeruginosa isolates recovered between 2014 and 2021 from oncological patients belonged to those same clusters. Methods The 32 isolates were re-cultured, whole-genome sequenced, phenotypically tested for their antimicrobial susceptibility, and analyzed for clonality and resistance genes in silico. Results Among these strains, 25 belonged to the two clusters that had been predominant in the wastewater, while two others belonged to a sequence-type less prominently detected in the drains of the patient rooms. Conclusion Patients constantly exposed to antibiotics can, in interaction with their persistently antibiotic-exposed sanitary facilities, form a niche that might be supportive for the emergence, the development, the dissemination, and the maintenance of certain nosocomial pathogen populations in the hospital, due to antibiotic-induced selection pressure. Technical and infection control solutions might help preventing transmission of microorganisms from the wastewater system to the patient and vice versa, particularly concerning the shower and toilet drainage. However, a major driving force might also be antibiotic induced selection pressure and parallel antimicrobial stewardship efforts could be essential.

Background: All b-lactamase enzymes, or carbapenemases, are able to hydrolyzing the beta-lactam antibiotics. The ability of Klebsiella pneumoniae to produce carbapenemases is the primary cause of carbapenem resistance in this organism. Objective: To define the prevalence as well as characterization of carbapenem resistance with genes coding for the carbapenemase in carbapenem-resistant Klebsiella pneumoniae (CRKP) isolates that was taken from patients in the intensive care units (ICUs) of Zagazig University Hospitals. Patients and Methods: In a cross-sectional study we included patients who were diagnosed with K. pneumoniae infections from different ICUs of Zagazig University Hospitals, which are referral hospitals serving many patients in eastern Egypt. Results: In this study 120 out of the 190 participants who had a confirmed K. pneumoniae infection were found to have CRKP infections. The XpertCarba-R assay was used to test the isolates for the presence of blaNDM, blaIMP, blaVIM, blaKPC, and blaOXA-48. The blaNDM (60%), blaOXA-48 (47.6%), and blaKPC (8.4%) genes were the most common. None of the isolates had either blaVIM or blaIMP present. Conclusion: These current data indicate spread of CRKP isolates in our institution poses a serious health threat. Limiting transmission depends on early detection of isolates that are resistant to carbapenem. In order to prevent the spread of the CRKP, it is necessary to pay more attention to rationalizing the use of antibiotics and strengthening the application of infection control precautions, such as hand hygiene, patient isolation, environmental cleaning and antibiotics stewardship programs. [ABSTRACT FROM AUTHOR]

The rise of antimicrobial resistance is a global challenge that requires a coordinated effort to address. In this study, we examined the genetic similarity of carbapenem-resistant Klebsiella pneumoniae (CRKP) in countries belonging to the Gulf Cooperation Council (GCC) to gain a better understanding of how these bacteria are spreading and evolving in the region. We used in silico genomic tools to investigate the occurrence and prevalence of different types of carbapenemases and their relationship to specific sequence types (STs) of CRKP commonly found in the region. We analyzed 720 publicly available genomes of multi-drug resistant K. pneumoniae isolates collected from six GCC countries between 2011 and 2020. Our findings showed that ST-14 and ST-231 were the most common STs, and 51.7% of the isolates carried blaOXA-48-like genes. Additionally, we identified rare carbapenemase genes in a small number of isolates. We observed a clonal outbreak of ST-231 in Oman, and four Saudi isolates were found to have colistin resistance genes. Our study offers a comprehensive overview of the genetic diversity and resistance mechanisms of CRKP isolates in the GCC region that could aid in developing targeted interventions to combat this pressing global issue. [ABSTRACT FROM AUTHOR]

Background: The aim of the study was to provide a clinical treatment reference for acute pancreatitis (AP) with infection, we analyzed the clinical and genomic characteristic of carbapenem-resistant Klebsiella pneumoniae (CRKP) isolates from AP with infection in China. Methods: Our clinical database was retrospectively analyzed with focus on the carbapenem-resistant characteristics among AP with infection in our Intensive Care Unit (ICU). Whole-genome sequencing (WGS) was used to analyze the antibiotic resistance gene, and antimicrobial susceptibility testing (AST) was performed to study the relevant phenotype in vitro. The CRISPR-Cas9 system was used to verify the relevant phenotype. Results: Based on 2,211 AST data of 627 AP patients with infection, CRKP had the highest proportion among carbapenem- resistant Enterobacteriaceae (CRE), at 37.8% for imipenem and 45.3% for meropenem. WGS revealed key β-lactamase genes, specifically blaCTX-M-15, blaCTX-M-65, blaKPC-2, blaLAP-2, blaNDM-5, blaTEM-181, blaOXA-1, and blaSHV. A total of 31.3% of CRKP were NDM-5-KPC-2-producing strains, and NDM-5-producing CRKP was resistant to imipenem/meropenem combined with avibactam, with an MIC of 512 mg/L. In addition, after knocking out blaKPC-2 and blaNDM-5, NDM-5-producing and KPC-2-producing CRKP had the same resistance level to imipenem/ meropenem. Conclusions: We first provided key insights into the clinical and genomic characteristic of CRKP in AP with infection and then made it clear that NDM-5 and KPC-2 had the same resistance level to carbapenems. [ABSTRACT FROM AUTHOR]

Background: Carbapenem-resistant gram-negative bacteria pose a serious threat worldwide, and some patients even have rapidly aggravated life-threatening infection. However, as a result of the complexities of clinical therapy, antibiotic options against carbapenem-resistant pathogens have not yet been fully standardized. It should be individualized to control carbapenem-resistant pathogens in accordance with the different region. Methods: In this study, we conducted a retrospective study in 65,000 inpatients over a 2-year period that involved a total of 86 patients from whom carbapenem-resistant gram-negative bacteria were isolated. Results: Monotherapy using trimethoprim/ sulfamethoxazole, amikacin, meropenem, and/or doxycycline in our hospital exhibited a clinical success rate of 83.3% for carbapenem-resistant Klebsiella pneumoniae, monotherapy using moxifloxacin, piperacillin/tazobactam, cefepime, and/or ceftazidime for carbapenem-resistant Pseudomonas aeruginosa exhibited a clinical success rate of 77.7%, and monotherapy using cefoperazone/sulbactam or combination therapy with tigecycline and cefoperazone/sulbactam for carbapenem-resistant Acinetobacter baumannii exhibited a clinical success rate of 62.1%. Conclusions: Taken together, our findings highlight the clinical strategies used in our hospital to successfully treat carbapenem-resistant gram-negative bacterial infections. [ABSTRACT FROM AUTHOR]

We report here a fatal case of carbapenem-resistant Klebsiella pneumoniae (CRKP) infections in a renal transplant patient without a travel history in the prior year, from whom 2 genetically different CRKP (sequence type 14 [ST14] and ST2497) strains carrying the same plasmids and antimicrobial resistance genes, including blaNDM-1, blaOXA-232, blaCTX-M-15, armA, and tet(D), were isolated from blood and the abdominal cavity. The isolates were susceptible to colistin, tigecycline, eravacycline, and cefiderocol, which was used to treat the CRKP in combination with ceftazidime-avibactam and polymyxin B and resulted in bacterial clearance. Despite the aggressive treatment, the patient died of ischemic colitis and multiorgan failure.

Ning Du,1 En-Qiang Mao,2 Zhi-Tao Yang,2 Hong-Ping Qu,3 Xian Qian,4 Ying Shi,5 Xiao-Lan Bian,6 Juan He,6,&ast; Er-Zhen Chen2,&ast; 1National Cancer Center/National Clinical Research Center for Cancer/Cancer Hospital & Shenzhen Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, Shenzhen, People’s Republic of China; 2Emergency Intensive Care Unit, Ruijin Hospital Affiliated to Shanghai Jiao Tong University School of Medicine, Shanghai, People’s Republic of China; 3Department of Critical Care Medicine, Ruijin Hospital Affiliated to Shanghai Jiao Tong University School of Medicine, Shanghai, People’s Republic of China; 4Department of Pharmacy, Shanghai Eastern Hepatobiliary Surgery Hospital, Shanghai, People’s Republic of China; 5Department of Clinical Pharmacy, Shaoxing Hospital of Traditional Chinese Medicine, Zhejiang, People’s Republic of China; 6Department of Pharmacy, Ruijin Hospital Affiliated to Shanghai Jiao Tong University School of Medicine, Shanghai, People’s Republic of China&ast;These authors contributed equally to this workCorrespondence: Er-Zhen Chen, Emergency Intensive Care Unit, Ruijin Hospital Affiliated to Shanghai Jiao Tong University School of Medicine, Shanghai, People’s Republic of China, Email chenerzhen@hotmail.com Juan He, Department of Pharmacy, Ruijin Hospital Affiliated to Shanghai Jiao Tong University School of Medicine, Shanghai, People’s Republic of China, Email hejuanwin@126.comPurpose: Infection with carbapenem-resistant Klebsiella pneumoniae (CRKP) is a great challenge. Central nervous system (CNS) infection caused by CRKP is rarely reported, and effective treatment is limited. Thus, this study aimed to assess intrathecal (IT) or intraventricular (IVT) injection of tigecycline for clearing infection with CRKP in CNS.Patients and Methods: Two patients who had intracranial infection with CRKP after craniotomy were treated in our institution and analyzed retrospectively, summarizing their therapeutic schedules.Results: They all had a fever with the positive results of cerebrospinal fluid (CSF) test, and CSF culture showed positive for CPKP, which was sensitive only to tigecycline. In addition, the MIC of polymyxin B was not tested due to the limited laboratory conditions. After IT or IVT injection of tigecycline treatment, the temperature of the patients became normal in 3 days, with normal levels of white blood cells, protein, glucose and chlorine concentrations in the CSF. Crucially, twice CSF cultures also became negative with no clinical symptoms of intracranial infection after IT or IVT injection of tigecycline treatment. Moreover, there were no adverse drug reactions observed.Conclusion: IT or IVT injection of tigecycline may be a bright choice to control intracranial infection with CRKP.Keywords: central nervous system infection, CNS, carbapenem-resistant Klebsiella pneumoniae, CRKP, tigecycline, treatment

Rania Abd El-Hamid El-Kady,1,2 Mohamed Ali Elbaiomy,3 Rasha Mokhtar Elnagar1 1Department of Medical Microbiology and Immunology, Faculty of Medicine, Mansoura University, Mansoura, Egypt; 2Department of Pathological Sciences, Fakeeh College for Medical Sciences, Jeddah, Kingdom of Saudi Arabia; 3Medical Oncology Unit, Oncology Center, Mansoura University, Mansoura, EgyptCorrespondence: Rania Abd El-Hamid El-Kady, Department of Pathological Sciences, Fakeeh College for Medical Sciences, P.O. Box 2537, Jeddah, 21461, Kingdom of Saudi Arabia, Tel +966 569849897, Email raniael_kady@yahoo.comBackground: A growing body of evidence suggests that ceftazidime/avibactam (CZA) is a potential therapeutic option for carbapenem-resistant Klebsiella pneumoniae (CRKP) infections; however, resistant strains are increasingly emerged worldwide. Herein, we deemed to investigate the susceptibility profile of CRKP isolates from cancer patients to CZA and to identify the underlying resistance mechanisms.Methods: Clinical samples were obtained from adult patients admitted to the Oncology Center of Mansoura University, Mansoura, Egypt. The antibiotic susceptibility pattern of K. pneumoniae isolates to different antibiotics was tested by the modified Kirby Bauer’s disc diffusion method. Minimum inhibitory concentrations of CZA were assessed using broth microdilution method. Screening for carbapenemase-producing strains was achieved by the modified Hodge test. Multiplex polymerase chain reactions (PCRs) were conducted for uncovering of carbapenemase-encoding genes (blaKPC, blaVIM, blaIMP, blaNDM-1, and blaOXA-48), and outer membrane porin genes (ompK35 and ompK36).Results: A total of 12 CZA-resistant isolates were identified out of 47 CRKP isolates (25.5%). The MIC50 and MIC90 of CZA against CRKP were 1 and 64 μg/mL, respectively. Risk factors for CZA resistance included chronic kidney disease, mechanical ventilation, longer length of hospital stay, and ICU admission. The multivariate logistic regression demonstrated that longer length of hospital stay (P=0.03) was the only independent predictor for acquisition of CZA-resistant isolates. The leading mechanism for CZA resistance was sustained by blaKPC (50%), meanwhile 16.7% and 8.3% of the CZA-resistant isolates harbored blaOXA-48 and blaOXA-48/blaNDM-1, respectively. The MBL-encoding genes blaNDM-1 and blaIMP were detected in 16.7% and 8.3% of the isolates, respectively. Absence of both ompK35 and ompK36 was observed in 58.3% of the CZA-resistant isolates.Conclusion: CZA has displayed superior in vitro activity against CRKP isolates in comparison to other antibiotics; however, thorough molecular characterization of resistant strains is highly recommended in future studies to detect and monitor the emergence of further tackling strains.Keywords: CRKP, ceftazidime/avibactam, blaKPC, OmpK35, OmpK36, cancer

Background: Hospitals with their high antimicrobial selection pressure represent the presumably most important reservoir of multidrug-resistant human pathogens. Antibiotics administered in the course of treatment are excreted and discharged into the wastewater system. Not only in patients, but also in the sewers, antimicrobial substances exert selection pressure on existing bacteria and promote the emergence and dissemination of multidrug-resistant clones. In previous studies, two main clusters were identified in all sections of the hospital wastewater network that was investigated, one K. pneumoniae ST147 cluster encoding NDM- and OXA-48 carbapenemases and one VIM-encoding P. aeruginosa ST823 cluster. In the current study, we investigated if NDM- and OXA-48-encoding K. pneumoniae and VIM-encoding P. aeruginosa isolates recovered between 2014 and 2021 from oncological patients belonged to those same clusters. Methods: The 32 isolates were re-cultured, whole-genome sequenced, phenotypically tested for their antimicrobial susceptibility, and analyzed for clonality and resistance genes in silico. Results: Among these strains, 25 belonged to the two clusters that had been predominant in the wastewater, while two others belonged to a sequence-type less prominently detected in the drains of the patient rooms. Conclusion: Patients constantly exposed to antibiotics can, in interaction with their persistently antibiotic-exposed sanitary facilities, form a niche that might be supportive for the emergence, the development, the dissemination, and the maintenance of certain nosocomial pathogen populations in the hospital, due to antibiotic-induced selection pressure. Technical and infection control solutions might help preventing transmission of microorganisms from the wastewater system to the patient and vice versa, particularly concerning the shower and toilet drainage. However, a major driving force might also be antibiotic induced selection pressure and parallel antimicrobial stewardship efforts could be essential. [ABSTRACT FROM AUTHOR]

ABSTRACT Colistin has been considered a last-line option for the treatment of infections caused by carbapenem-resistant Klebsiella pneumoniae (CRKP). Heterogeneous resistance to colistin leads to unexplained clinical colistin treatment failure for CRKP. Our study aimed to investigate the extent of colistin heteroresistance among CRKP strains in China. A total of 455 colistin-susceptible strains, collected from six tertiary care hospitals in China, were characterized. The overall rate of colistin heteroresistance was 6.2%, as determined by the population analysis profiles (PAPs). Genomic analysis revealed that 60.7% of the colistin-heteroresistant isolates belonged to the epidemic sequence type 11 (ST11) clone. Single-nucleotide polymorphisms (SNPs) suggested that 6 ST5216 strains shared the same origin. Each of the subpopulations had a ≥8-fold decrease in colistin MIC in the presence of carbonyl cyanide m-chlorophenylhydrazone (CCCP), which indicated that heteroresistance could be suppressed by an efflux pump inhibitor. In addition, our results suggested that the PhoPQ pathway plays an important role in the mechanisms of heteroresistance. IMPORTANCE The problem of CRKP has raised alarms concerning global health. Our study enriches the epidemiological study of colistin heteroresistance among CRKP strains in China, where the prevalence of this phenomenon was previously unknown. Importantly, colistin-heteroresistant strains may cause the failure of clinical treatment with colistin, even if the clinical laboratory reports that the strains are sensitive. The commonly used broth microdilution method is unable to detect this special phenomenon. Additionally, our results indicate that efflux pumps play a major role in colistin heteroresistance, and inhibitors can effectively reverse it. Our study is the first to provide a detailed analysis of the prevalence of colistin heteroresistance in China, as well as an analysis of the genetic mechanisms of this phenomenon.

Introduction: Polymyxin B is a last-line therapy for carbapenem-resistant microorganisms. However, a lack of clinical pharmacokinetic/pharmacodynamic (PK/PD) data has substantially hindered dose optimization and breakpoint setting. Methods: A prospective, multi-center clinical trial was undertaken with polymyxin B [2.5 mg/kg loading dose (3-h infusion), 1.25 mg/kg/12 h maintenance dose (2-h infusion)] for treatment of carbapenem-resistant K. pneumoniae (CRKP) bloodstream infections (BSI). Safety, clinical and microbiological efficacy were evaluated. A validated liquid chromatographytandem mass spectrometry (LC-MS/MS) method was applied to determine the concentrations of polymyxin B in blood samples. Population pharmacokinetic (PK) modeling and Monte Carlo simulations were conducted to examine the susceptibility breakpoint for polymyxin B against BSI caused by CRKP. Results: Nine patients were enrolled and evaluated for safety. Neurotoxicity (5/9), nephrotoxicity (5/9), and hyperpigmentation (1/9) were recorded. Blood cultures were negative within 3 days of commencing therapy in all 8 patients evaluated for microbiological efficacy, and clinical cure or improvement occurred in 6 of 8 patients. Cmax and Cmin following the loading dose were 5.53 ± 1.80 and 1.62 ± 0.41 mg/L, respectively. With maintenance dosing, AUCss,24 h was 79.6 ± 25.0 mg h/L and Css,avg 3.35 ± 1.06 mg/L. Monte Carlo simulations indicated that a 1 mg/kg/12-hourly maintenance dose could achieve >90% probability of target attainment (PTA) for isolates with minimum inhibitory concentration (MIC) =1 mg/L. PTA dropped substantially for MICs =2 mg/L, even with a maximally recommended daily dose of 1.5 mg/kg/12-hourly. Conclusion: This is the first clinical PK/PD study evaluating polymyxin B for BSI. These results will assist to optimize polymyxin B therapy and establish its breakpoints for CRKP BSI. [ABSTRACT FROM AUTHOR]

Abstract Background Extensive drug resistance in Klebsiella pneumoniae (K. pneumoniae) causing major outbreaks in large hospitals is an emerging challenge. We describe a near fatal outbreak of colistin resistant, carbapenem resistant K. pneumoniae (CRKp) producing metallo beta-lactamases (bla NDM) and bla OXA-48 in the neonatal intensive care unit (NICU) at the background of a larger outbreak involving multiple parts of the hospital and the challenges in its containment. Methods Following identification of an outbreak due to colistin resistant CRKp between April to June 2017 in the NICU, a thorough surveillance of similar cases and the hospital environment was performed to trace the source. All the isolated K. pneumoniae were tested for susceptibility to standard antibiotics by disc diffusion and microbroth dilution methods. Molecular detection of extended spectrum beta lactamases (ESBLs) and carbapenemases (classes A, B, D) genes was done. Enterobacterial repetitive intergenic consensus (ERIC) PCR and multi-locus sequence typing (MLST) was done to determine the genetic relatedness of the isolates. Characteristics of different sequence types were statistically compared (Student’s t-test). Results A total of 45 K. pneumoniae isolates were studied from NICU (14 cases of neonatal sepsis), ICU (18 cases), other wards (7 cases) along with 6 isolates from hospital environment and human colonizers. The primary case was identified in the ICU. All the K. pneumoniae from NICU and 94.4% from the ICU were colistin resistant CRKp. Majority (59.37% and 56.25%) harbored bla SHV/bla CTXM and bla OXA-48 genes, respectively. Two distinct sequence types ST5235 and ST5313 were noted with colistin resistance, distribution within the NICU and mortality as significant attributes of ST5235 (p

ABSTRACT We conducted a 6-month prospective study in a newly opened ICU for high-resolution tracking of carbapenem-resistant Klebsiella pneumoniae (CRKP) through environmental surveillance, patient screening, and genome sequencing. Among all ICU patients (n = 348) screened, 3.5% carried CRKP on admission and 16.3% acquired CRKP thereafter. CRKP was not detected in the environment until 10 weeks and was then isolated from 98 of 2,989 environmental samples (3.3%). The first CRKP isolate from rectal swabs (n = 37) and the first clinical isolate (n = 8) of each patient as well as the 98 isolates from environmental were subjected to whole-genome sequencing. The 143 CRKP isolates from patients and environment samples were assigned to four sequence types, with ST11 dominating (95.8%) and further divided into 14 clones, suggesting introduction of multiple clones. Subsequent CRKP transmission was complex and dynamic with 10 clones found in multiple patients and seven also detected in the environment. Two particular ST11 clones caused extensive (≥5 rooms) and persistent (≥10 weeks) environmental contamination. Both clones were associated with patients who carried CRKP throughout their prolonged ICU stay. Such “super-contaminators” are a priority for isolation and environmental surveillance. IMPORTANCE Carbapenem-resistant Klebsiella pneumoniae (CRKP) is a global challenge for human health. In health care settings, patients have frequent interactions with other patients and the environment, rendering challenges for untangling the introduction and transmission of CRKP. We conducted a prospective surveillance study in a newly opened ICU for high-resolution tracking of CRKP. Our study demonstrated the dynamic, complicated transmission of CRKP and has important findings that may help to curb its spread in health care settings. First, compliance with basic measures such as routine environment cleaning and postdischarge terminal cleaning is needed to minimize the environmental contamination-driven spread. Second, active screening could demonstrate the scale of the problem, and room transfer of patients with CRKP should be prohibited whenever possible. Third, the priority for single-room isolation should be given to patients with prolonged carriage of CRKP, especially in resource-limited settings. Good infection control practice lays a foundation for tackling multidrug-resistant organisms like CRKP.

Carbapenem-resistant Klebsiella pneumoniae (CRKP) seriously threaten the efficacy of modern medicine with a high associated mortality rate and unprecedented transmission rate. In this study, we isolated a clinical K. pneumoniae strain DY1928 harboring blaNDM-1 from a neonate with blood infection. Antimicrobial susceptibility testing indicated that DY1928 was resistant to various antimicrobial agents, including meropenem, imipenem, ceftriaxone, cefotaxime, ceftazidime, cefepime, piperacillin-tazobactam, and amoxicillin-clavulanate. S1 nuclease-pulsed field gel electrophoresis (S1-PFGE), southern blot and conjugation experiment revealed that the blaNDM-1 gene was located on a conjugative plasmid of IncA/C2 type with a 147.9 kb length. Whole-genome sequencing showed that there was a conservative structure sequence (blaNDM-1-ble-trpF-dsbD) located downstream of the blaNDM-1 gene. Multilocus sequence typing (MLST) classified DY1928 as ST25, which was a hypervirulent K. pneumoniae type. Phylogenetic analysis of genomic data from all ST25 K. pneumoniae strains available in the NCBI database suggested that all blaNDM-1 positive strains were isolated in China and had clinical origins. A mouse bloodstream infection model was constructed to test the virulence of DY1928, and 11 K. pneumoniae strains homologous to DY1928 were isolated from the feces of infected mice. Moreover, we found that DY1928 had a tendency to flow from the blood into the intestine in mice and caused multiple organ damage. To our knowledge, this is the first study to report an infection caused by blaNDM-1-positive ST25 K. pneumoniae in the neonatal unit. Our findings indicated that stricter surveillance and more effective actions were needed to reduce the risk of disseminating such K. pneumoniae strains in clinical settings, especially in neonatal wards. [ABSTRACT FROM AUTHOR]

Objectives: To characterize one OXA-232-producing wzi93-KL112-O1 carbapenem-resistant Klebsiella pneumoniae (CRKP) co-harboring chromosomal blaCTX-M-15 and one rmpA2-associated virulence plasmid. Methods: Minimum inhibitory concentrations (MICs) were measured via broth microdilution method. Conjugation, chemical transformation, string test and Galleria mellonella infection model experiments were also conducted. Wholegenome sequencing (WGS) was performed on the Illumina and Nanopore platforms. Antimicrobial resistance determinants were identified using ABRicate program with ResFinder database. Insertion sequences (ISs) were identified using ISfinder. Bacterial virulence factors were identified using virulence factor database (VFDB). Wzi, capsular polysaccharide (KL) and lipoolygosaccharide (OCL) were analyzed using Kleborate with Kaptive. Phylogenetic analysis of 109 ST15 K. pneumoniae strains was performed using core genome multilocus sequence typing (cgMLST) on the Ridom SeqSphere+ server. MLST, replicons type, SNP strategies and another cgMLST analysis for 45 OXA-232-producing K. pneumoniae strains were further conducted using BacWGSTdb server. Results: K. pneumoniae KPTCM strain belongs to ST15 with wzi93, KL112 and O1. It possessed a multidrug-resistant (MDR) profile and was resistant to carbapenems (meropenem and ertapenem), ciprofloxacin and amikacin. Virulence assays demonstrated KPTCM strain possesses a low virulence phenotype. WGS revealed it contained one circular chromosome and nine plasmids. The carbapenemase-encoding gene blaOXA-232 was located in a 6141-bp ColKP3-type non-conjugative plasmid and flanked by DISEcp1 and DlysR-DereA. Interestingly, blaCTX-M-15 was located in the chromosome mediated by ISEcp1-based transposon Tn2012. Importantly, it harbored a rmpA2-associated pLVPK-like virulence plasmid with iutA-iucABCD gene cluster and one IS26-mediated MDR fusion plasmid according to 8-bp (AGCTGCAC or GGCCTTTG) target site duplications (TSD). Based on the cgMLST and SNP analysis, data showed OXA-232-producing ST15 K. pneumoniae isolates were mainly isolated from China and have evolved in recent years. Conclusions: Early detection of CRKP strains carrying chromosomal blaCTX-M-15, OXA-232 carbapenemase and pLVPK-like virulence plasmid is recommended to avoid the extensive spread of this high-risk clone. [ABSTRACT FROM AUTHOR]

Klebsiella pneumoniae is a common pathogen of nosocomial infection, which can cause pneumonia, urinary tract infection, cystitis, and bloodstream infections (BSIs). Here, we genetically characterize a novel carbapenem-resistant K. pneumoniae (CRKP) strain recovered from the blood of a 44-year-old male patient with severe acute necrotizing pancreatitis and septic shock in China. The strain is a ST45 K. pneumoniae with a novel serotype of K43, named 18SHX166. The susceptibility testing results showed that 18SHX166 was resistant to cephalosporin and carbapenems but still susceptible to ceftazidime-avibactam, quinolones, colistin, and amikacin. Genomic sequencing revealed that 18SHX166 contains three plasmids, namely pSHX166-Hv, pSHX166-KPC, and pSHX166-3. pSHX166-Hv harbored the iucABCD operon, encoding the siderophore of aerobactin. pSHX166-KPC harbored bla KPC-2 gene and possessed complete conjugative regions. The conjugation experiment verified pSHX166-KPC as a self-transmissible plasmid mediating the dissemination of antibiotic resistance, with a conjugation rate of 2.21 × 10 -5 . Additionally, the growth curve showed that 18SHX166 demonstrates a higher growth rate than the control strains. The characteristics of 18SHX166 indicate a potential high risk of clinical transmission.IMPORTANCEST45-K43 carbapenem-resistant Klebsiella pneumoniae isolate, 18SHX166, carries a carbapenem resistance plasmid and virulence plasmid. It has the characteristics of multidrug resistance, high transmissibility, and a fast growth rate, which could pose a threat to the control of antimicrobial resistance and clinical transmission, causing a severe challenge to public health., Competing Interests: The authors declare no conflict of interest.

Purpose: The rise of carbapenem-resistant Klebsiella pneumoniae (CRKP) has led to increased morbidity and mortality in clinical patients, highlighting the urgent need for effective antibacterial agents., Methods: We obtained a synthetic compound, MTEBT-3, using hydrophobic triphenylamine as the skeleton and hydrophilic ammonium salts. We determined the MIC of MTEBT-3 using the macro-broth susceptibility testing method. We isolated a clinical CRKP strain ST3984 and performed synergistic antibiotic sensitivity tests, time-kill assays, and resistance evolution studies. Biofilm formation under sub-MIC conditions was evaluated using crystal violet staining and CLSM. Additionally, biofilm proteins and polysaccharides were quantified. We assessed the bactericidal mechanism of MTEBT-3 by examining the integrity of CRKP bacterial cell membranes and analyzing the transcription of virulence-regulating genes via quantitative real-time PCR., Results: MTEBT-3 exhibited broad-spectrum antibacterial activity with a low resistance rate, achieving an MIC of 8 μg/mL. The compound displayed additive effects with meropenem and imipenem and synergistic effects with tigecycline. It maintained its efficacy over multiple bacterial generations, with no significant increase in resistance observed. Under sub-MIC conditions, the biomass of biofilms was significantly reduced, and the levels of proteins and polysaccharides within the biofilms were markedly lowered in a concentration-dependent manner. The bactericidal mechanism of MTEBT-3 involved disrupting the integrity of CRKP bacterial cell membranes, leading to increased permeability. Quantitative real-time PCR results showed that MTEBT-3 effectively suppressed the expression of key virulence genes, including fimH, wbbM, rmpA, and rmpA2, which are associated with biofilm formation and bacterial adhesion., Conclusion: The significant antimicrobial activity of MTEBT-3 against clinically isolated CRKP, along with its synergistic or additive effects with commonly used antibiotics, positions it as a promising candidate for treatment. Its ability to disrupt biofilm formation and reduce virulence factor expression further underscores its potential in managing CRKP infections., Competing Interests: Declaration of Competing Interest ☒ The authors declare the following financial interests/personal relationships which may be considered as potential competing interestsJinju Duan reports financial support was provided by Second Hospital of Shanxi Medical University. Jinju Duan reports a relationship with Natural Science Foundation of Shanxi Province that includes: funding grants. If there are other authors, they 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. Published by Elsevier Ltd.)

Klebsiella pneumoniae carbapenemase (KPC) variants can contribute to resistance to ceftazidime-avibactam (CZA) in Klebsiella pneumoniae (KP). However, two-copy KPC variant-mediated resistance to CZA has rarely been reported to date. Here, we aimed to clarify the evolutionary trajectory of CZA resistance driven by mutations in double-copy bla KPC-2 to bla KPC-189 carried by the tandem core structure (IS Kpn6-bla KPC (CRKP). The CZA-resistant KP strain carried double-copy Kpn27-tnpR -IS 26 ) during treatment of ST11 carbapenem-resistant K. pneumoniae (CRKP). The CZA-resistant KP strain carried double-copy bla KPC-189 , a variant with alanine-threonine and aspartate-tyrosine substitutions at Ambler amino acid positions 172 (A172T) and 179 (D179Y) of bla KPC-2 . Clone experiments confirmed that, compared with that of the wild-type bla KPC-2 clone strain, the minimum inhibitory concentration of CZA increased 16-fold in the bla KPC-189 -mutant strain. Furthermore, protein structure analysis revealed the A172T and D179Y mutations of bla KPC-189 can have a direct effect on the binding affinity of CAZ and AVI for KPC. Sequence comparison revealed that bla KPC-189 was mutated in a double-copy format upon CZA exposure, which was carried by the IS 26 -mediated tandem core structure IS Kpn27-bla KPC -IS Kpn6 . This tandem core structure apparently evolves in vivo during infection, although not by self-transferring, and multiple IS Kpn27-bla KPC -IS Kpn6 copy numbers could mediate transferable CZA resistance upon mobilization. In addition, compared with the wild-type bla KPC-2 gene, the bla KPC-189 gene had no fitness cost. In summary, our study highlighted the emergence of CZA-resistant bla KPC-189 variants in the ST11 clone and the presence of a double-copy bla KPC-189 in the IncFII-type plasmid, which is carried by a tandem core structure (IS 26- IS Kpn6-bla KPC-189 -IS Kpn27-tnpR -IS 26 )., Importance: To date, ceftazidime-avibactam (CZA) resistance caused by double-copy Klebsiella pneumoniae carbapenemase (KPC) variants has not been elucidated. The multicopy forms of carbapenem resistance genes carried by the same plasmid are relatively rare in most carbapenem-resistant Enterobacteriaceae . In this study, we elucidate the evolutionary trajectory of CZA resistance in ST11 carbapenem-resistant K. pneumoniae harboring a double-copy blaKPC and provide new insights into the mechanisms of acquired resistance to CZA., Competing Interests: The authors declare no conflict of interest.

The rise of antimicrobial resistance is a global challenge that requires a coordinated effort to address. In this study, we examined the genetic similarity of carbapenem-resistant Klebsiella pneumoniae (CRKP) in countries belonging to the Gulf Cooperation Council (GCC) to gain a better understanding of how these bacteria are spreading and evolving in the region. We used in silico genomic tools to investigate the occurrence and prevalence of different types of carbapenemases and their relationship to specific sequence types (STs) of CRKP commonly found in the region. We analyzed 720 publicly available genomes of multi-drug resistant K. pneumoniae isolates collected from six GCC countries between 2011 and 2020. Our findings showed that ST-14 and ST-231 were the most common STs, and 51.7% of the isolates carried blaOXA-48-like genes. Additionally, we identified rare carbapenemase genes in a small number of isolates. We observed a clonal outbreak of ST-231 in Oman, and four Saudi isolates were found to have colistin resistance genes. Our study offers a comprehensive overview of the genetic diversity and resistance mechanisms of CRKP isolates in the GCC region that could aid in developing targeted interventions to combat this pressing global issue.

The advent of carbapenem-resistant Klebsiella pneumoniae (CRKP) poses a significant challenge to public health, as carbapenems are typically employed as a last resort to treat nosocomial infections caused by such organisms, especially in intensive care units (ICUs). This study aims to characterize the CRKP isolated from patients admitted to the Zagazig University Hospitals (ZUHs) ICU in Egypt. About 56.2%, 41.0%, and 32.4% of the isolates indicated the presence of blaNDM, blaOXA-48, and blaKPC, respectively. Carbapenemase-encoding genes were found in many isolates, and blaNDM was the most predominant gene. Nevertheless, this situation has become a heavy burden in developing countries, including Egypt, and is associated with substantial morbidity, mortality, and increased healthcare expenses. [ABSTRACT FROM AUTHOR]

Abstract Background The rate of fluoroquinolone (FQ) resistance among carbapenem-resistant Klebsiella pneumoniae (CRKP) is high. The present study aimed to investigate the distribution of fluoroquinolone resistance determinants in clinical CRKP isolates associated with bloodstream infections (BSIs). Results A total of 149 BSI-associated clinical CRKP isolates collected from 11 Chinese teaching hospitals from 2015 to 2018 were investigated for the prevalence of fluoroquinolone resistance determinants, including plasmid-mediated quinolone resistance (PMQR) genes and spontaneous mutations in the quinolone resistance-determining regions (QRDRs) of the gyrA and parC genes. Among these 149 clinical CRKP isolates, 117 (78.5%) exhibited resistance to ciprofloxacin. The GyrA substitutions (Ser83 → IIe/Phe) and (Asp87 → Gly/Ala) were found among 112 (75.2%) of 149 isolates, while the substitution (Ser80 → IIe) of ParC was found in 111 (74.5%) of the 149 isolates. In total, 70.5% (105/149) of the CRKP isolates had at least two mutations within gyrA as well as a third mutation in parC. No mutations in the QRDRs were found in 31 ciprofloxacin susceptible CRKP isolates. Eighty-nine (56.9%) of 149 were found to carry PMQR genes including qnrS1 (43.0%), aac(6′)-Ib-cr (16.1%), qnrB4 (6.0%), qnrB2 (2.7%), and qnrB1 (1.3%). Nine isolates contained two or more PMQR genes, with one carrying four [aac(6′)-Ib-cr, qnr-S1, qnrB2, and qnrB4]. The co-existence rate of PMQR determinants and mutations in the QRDRs of gyrA and parC reached 68.5% (61/89). Seventy-four (83.1%, 74/89) PMQR-positive isolates harbored extended-spectrum beta-lactamase (ESBL)-encoding genes. Multilocus sequence typing (MLST) analysis demonstrated that the ST11 was the most prevalent STs in our study. Conclusions Mutations in the QRDRs of gyrA and parC were the key factors leading to the high prevalence of fluoroquinolone resistance among BSI-associated CRKP. The co-existence of PMQR genes and mutations in the QRDRs can increase the resistance level of CRKP to fluoroquinolones in clinical settings. ST11 CRKP isolates with identical QRDR substitution patterns were found throughout hospitals in China.

Hypervirulence and carbapenem resistance have emerged as two distinct evolutionary directions for Klebsiella pneumoniae, which pose a great threat in clinical settings. Multiple virulence factors contribute to hypervirulence, and the mechanisms of carbapenem resistance are complicated. However, more and more K. pneumoniae strains have been identified in recent years integrating both phenotypes, resulting in devastating clinical outcomes. Hypervirulent and carbapenem-resistant K. pneumoniae (CR-hvKP) emerged in the early 2010s and thereafter have become increasingly prevalent. CR-hvKP are primarily prevalent in Asia, especially China, but are reported all over the world. Mechanisms for the emergence of CR-hvKP can be summarised by three patterns: (i) carbapenem-resistant K. pneumoniae (CRKP) acquiring a hypervirulent phenotype; (ii) hypervirulent K. pneumoniae (hvKP) acquiring a carbapenem-resistant phenotype; and (iii) K. pneumoniae acquiring both a carbapenem resistance and hypervirulence hybrid plasmid. With their global dissemination, continued surveillance of the emergence of CR-hvKP should be more highly prioritised.

Enbo Liu, 1,* Peiyao Jia, 1, 2,* Xue Li, 1, 3,* Menglan Zhou, 1, 2 Timothy Kudinha, 4, 5 Chuncai Wu, 1 Yingchun Xu, 1 Qiwen Yang 1 1Department of Clinical Laboratory, State Key Laboratory of Complex Severe and Rare Diseases, Peking Union Medical College Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, 100730, People’s Republic of China; 2Graduate School, Peking Union Medical College, Chinese Academy of Medical Sciences, Beijing, People’s Republic of China; 3Department of Clinical Laboratory, Beijing Anzhen Hospital, Capital Medical University, Beijing, People’s Republic of China; 4School of Biomedical Sciences, Charles Sturt University, Orange, 2800, Australia; 5Pathology West, NSW Health Pathology, Orange, 2800, Australia*These authors contributed equally to this workCorrespondence: Qiwen Yang Tel +86-10-6915-9763Fax +86-10-6915-9766Email yangqiwen81@vip.163.comObjective: This study aimed to evaluate the in vitro and in vivo effects of different combinations of antimicrobial agents against carbapenemase-producing and non-producing Klebsiella pneumoniae from China.Methods: A checkerboard assay of meropenem (MEM), amikacin (AK), tigecycline (TGC), colistin (COL) and their combinations was carried out against 58 clinical carbapenem-resistant K. pneumoniae (CRKp) isolates, including 11 carbapenemase-non-producing K. pneumoniae isolates and 21 isolates producing KPC-2 enzyme, 11 NDM-1, 13 IMP, one VIM-1 and one OXA-48. The checkerboard assay was analyzed by the fractional inhibitory concentration index (FICI). A time–kill assay and Galleria mellonella infection model were conducted to evaluate the in vitro and in vivo effects of the four drugs alone and in combination.Results: In the checkerboard assay, TGC+AK and MEM+AK combinations showed the highest synergistic effect against KPC-2 and NDM-1 carbapenemase-producing isolates, with synergy+partial synergy (defined as FICI < 1) rates of 76.2% and 71.4% against KPC-2 producers, and 54.5% and 81.8% against NDM-1 producers. TGC+AK and MEM+COL combinations showed the highest rate of synergistic effect against IMP-producing isolates. Against carbapenemase-non-producing isolates, TGC+COL and TGC+AK combinations showed the highest rate of synergy effect (63.6% and 54.5%). MEM+AK showed a synergistic effect against one VIM-1 producer (FICI=0.31) and an additivite effect (FICI=1) against one OXA-48 producer. In the time–kill assay, COL+AK, COL+TGC, COL+MEM and AK+TGC showed good synergistic effects against the KPC-2-producing isolate D16. COL+MEM and COL+TGC combinations showed good effects against the NDM-1-producing isolate L13 and IMP-4-producing isolate L34. Against the carbapenemase-non-producing isolate Y105, MEM+TGC and COL+AK showed high synergistic effects, with log 10CFU/mL decreases of 6.2 and 5.5 compared to the most active single drug. In the G. mellonella survival assay, MEM-based combinations had relatively high survival rates, especially when combined with colistin, against KPC-2 producers (90% survival rate) and with amikacin against metallo-beta-lactamase producers (95– 100% survival rate).Conclusion: Our study suggests that different antimicrobial agent combinations should be considered against CRKp infections with different resistance mechanisms.Keywords: resistance mechanisms, time–kill curve assay, carbapenem-resistant Klebsiella pneumoniae, CRKP, antimicrobial agent combinations, Galleria mellonella infection model