Transcriptomic investigations of polymyxins and colistin/sulbactam combination against carbapenem-resistant Acinetobacter baumannii.

Carbapenem-resistant Acinetobacter baumannii (CRAB) is a Priority 1 (Critical) pathogen urgently requiring new antibiotics. Polymyxins are a last-line option against CRAB-associated infections. This transcriptomic study utilized a CRAB strain to investigate mechanisms of bacterial killing with polymyxin B, colistin, colistin B, and colistin/sulbactam combination therapy. After 4 h of 2 mg/L polymyxin monotherapy, all polymyxins exhibited common transcriptomic responses which primarily involved disruption to amino acid and fatty acid metabolism. Of the three monotherapies, polymyxin B induced the greatest number of differentially expressed genes (DEGs), including for genes involved with fatty acid metabolism. Gene disturbances with colistin and colistin B were highly similar (89 % common genes for colistin B), though effects on gene expression were generally lower (0-1.5-fold in most cases) with colistin B. Colistin alone (2 mg/L) or combined with sulbactam (64 mg/L) resulted in rapid membrane disruption as early as 1 h. Transcriptomic analysis of this combination revealed that the effects were driven by colistin, which included disturbances in fatty acid synthesis and catabolism, and inhibition of nutrient uptake. Combination therapy produced substantially higher fold changes in 72 % of DEGs shared with monotherapy, leading to substantially greater reductions in fatty acid biosynthesis and increases in biofilm, cell wall, and phospholipid synthesis. This indicates synergistic bacterial killing with the colistin/sulbactam combination results from a systematic increase in perturbation of many genes associated with bacterial metabolism. These mechanistic insights enhance our understanding of bacterial responses to polymyxin mono- and combination therapy and will assist to optimize polymyxin use in patients.

Adult acute leukemia patients with gram-negative bacteria bloodstream infection: Risk factors and outcomes of antibiotic-resistant bacteria.

This study aims to analyze the risk factors for the development of multidrug-resistant (MDR) and carbapenem-resistant (CR) bacteria bloodstream infection (BSI) in a patient with acute leukemia (AL) and the mortality in gram-negative bacteria (GNB) BSI. This is a retrospective study conducted at West China Hospital of Sichuan University, which included patients diagnosed with AL and concomitant GNB BSI from 2016 to 2021. A total of 206 patients with GNB BSI in AL were included. The 30-day mortality rate for all patients was 26.2%, with rates of 25.8% for those with MDR GNB BSI and 59.1% for those with CR GNB BSI. Univariate and multivariate analyses revealed that exposure to quinolones (Odds ratio (OR) = 3.111, 95% confidence interval (95%CI): 1.623-5.964, p = 0.001) within the preceding 30 days was an independent risk factor for MDR GNB BSI, while placement of urinary catheter (OR = 6.311, 95%CI: 2.478-16.073, p < 0.001) and exposure to cephalosporins (OR = 2.340, 95%CI: 1.090-5.025, p = 0.029) and carbapenems (OR = 2.558, 95%CI: 1.190-5.497, p = 0.016) within the preceding 30 days were independently associated with CR GNB BSI. Additionally, CR GNB BSI (OR = 2.960, 95% CI: 1.016-8.624, p = 0.047), relapsed/refractory AL (OR = 3.035, 95% CI: 1.265-7.354, p = 0.013), septic shock (OR = 5.108, 95% CI: 1.794-14.547, p = 0.002), platelets < 30 × 109/L before BSI (OR = 7.785, 95% CI: 2.055-29.492, p = 0.003), and inappropriate empiric antibiotic therapy (OR = 3.140, 95% CI: 1.171-8.417, p = 0.023) were independent risk factors for 30-day mortality in AL patients with GNB BSI. Prior antibiotic exposure was a significant factor in the occurrence of MDR GNB BSI and CR GNB BSI. CR GNB BSI increased the risk of mortality in AL patients with GNB BSI.

Epidemiology of Citrobacter spp. infections among hospitalized patients: a systematic review and meta-analysis.

BACKGROUND: Infections due to Citrobacter species are increasingly observed in hospitalized patients and are often multidrug-resistant. Yet, the magnitude and burden of Citrobacter spp. resistance in the hospital setting have not been reported. We aimed to evaluate the epidemiology of Citrobacter spp. infections among hospitalized patients, their main resistance patterns and Citrobacter spp. involvement in hospital outbreaks. METHODS: We conducted a systematic review and meta-analysis of published literature (PROSPERO registration Jan-2023, CRD42023390084). We searched Embase, Medline and grey literature for studies on hospitalized patients diagnosed with Citrobacter spp. infections, and nosocomial outbreaks due to Citrobacter spp. published during the years 2000-2022. We included observational, interventional, surveillance studies and outbreak reports. Outcomes of interest were the frequency of Citrobacter spp. infections among hospitalized patients and 3rd generation cephalosporin and/or carbapenem resistance percentages in these infections. We used random-effects models to generate pooled outcome estimates and evaluated risk of bias and quality of reporting of outbreaks. RESULTS: We screened 1609 deduplicated publications, assessed 148 full-texts, and included 41 studies (15 observational, 13 surveillance and 13 outbreak studies). Citrobacter spp. urinary tract- and bloodstream infections were most frequently reported, with Citrobacter freundii being the main causative species. Hospital-acquired infection occurred in 85% (838/990) of hospitalized patients with Citrobacter infection. After 2010, an increasing number of patients with Citrobacter spp. infections was reported in observational studies. Pooled frequency estimates for Citrobacter spp. infections could not be generated due to lack of data. The pooled prevalence of ESBL and carbapenemase producers among Citrobacter isolates were 22% (95%CI 4-50%, 7 studies) and 18% (95%CI 0-63%, 4 studies), respectively. An increased frequency of reported Citrobacter outbreaks was observed after 2016, with an infection/colonization ratio of 1:3 and a case-fatality ratio of 7% (6/89 patients). Common outbreak sources were sinks, toilets, contaminated food and injection material. Implemented preventive measures included environmental cleaning, isolation of positive patients and reinforcement of hand hygiene. Only seven out of 13 outbreaks (54%) were definitively controlled. CONCLUSION: This review highlights the clinical importance of endemic and epidemic Citrobacter spp. in healthcare settings. As an emerging, multidrug­resistant nosocomial pathogen it requires heightened awareness and further dedicated surveillance efforts.

Clinical Characteristics and Molecular Insights of Carbapenem-Resistant Klebsiella pneumoniae Isolates from Patients in Intensive Care Units.

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

Clinical outcomes of carbapenem-resistant gram-negative bacterial bloodstream infection in patients with end-stage renal disease in intensive care units: a multicenter retrospective observational study.

BACKGROUND: Carbapenem-resistant gram-negative bacteria (CRGNB) present a considerable global threat due to their challenging treatment and increased mortality rates, with bloodstream infection (BSI) having the highest mortality rate. Patients with end-stage renal disease (ESRD) undergoing renal replacement therapy (RRT) face an increased risk of BSI. Limited data are available regarding the prognosis and treatment outcomes of CRGNB-BSI in patients with ESRD in intensive care units (ICUs). METHODS: This multi-center retrospective observational study included a total of 149 ICU patients with ESRD and CRGNB-BSI in Taiwan from January 2015 to December 2019. Clinical and microbiological outcomes were assessed, and multivariable regression analysis was used to evaluate the independent risk factors for day-28 mortality and the impact of antimicrobial therapy regimen on treatment outcomes. RESULTS: Among the 149 patients, a total of 127 patients (85.2%) acquired BSI in the ICU, with catheter-related infections (47.7%) and pneumonia (32.2%) being the most common etiologies. Acinetobacter baumannii (49.0%) and Klebsiella pneumoniae (31.5%) were the most frequently isolated pathogens. The day-28 mortality rate from BSI onset was 52.3%, and in-hospital mortality was 73.2%, with survivors experiencing prolonged hospital stays. A higher Sequential Organ Failure Assessment (SOFA) score (adjusted hazards ratio [aHR], 1.25; 95% confidence interval [CI] 1.17-1.35) and shock status (aHR, 2.12; 95% CI 1.14-3.94) independently predicted day-28 mortality. Colistin-based therapy reduced day-28 mortality in patients with shock, a SOFA score of ≥ 13, and Acinetobacter baumannii-related BSI. CONCLUSIONS: CRGNB-BSI led to high mortality in critically ill patients with ESRD. Day-28 mortality was independently predicted by a higher SOFA score and shock status. In patients with higher disease severity and Acinetobacter baumannii-related BSI, colistin-based therapy improved treatment outcomes.

Optimizing Treatment for Carbapenem-Resistant Acinetobacter baumannii Complex Infections: A Review of Current Evidence.

Carbapenem-resistant Acinetobacter baumannii complex (CRAB) poses a significant global health challenge owing to its resistance to multiple antibiotics and limited treatment options. Polymyxin-based therapies have been widely used to treat CRAB infections; however, they are associated with high mortality rates and common adverse events such as nephrotoxicity. Recent developments include numerous observational studies and randomized clinical trials investigating antibiotic combinations, repurposing existing antibiotics, and the development of novel agents. Consequently, recommendations for treating CRAB are undergoing significant changes. The importance of colistin is decreasing, and the role of sulbactam, which exhibits direct antibacterial activity against A. baumannii complex, is being reassessed. High-dose ampicillin-sulbactam-based combination therapies, as well as combinations of sulbactam and durlobactam, which prevent the hydrolysis of sulbactam and binds to penicillin-binding protein 2, have shown promising results. This review introduces recent advancements in CRAB infection treatment based on clinical trial data, highlighting the need for optimized treatment protocols and comprehensive clinical trials to combat the evolving threat of CRAB effectively.

Evolution of blaKPC Under the Pressure of Carbapenems and Ceftazidime/Avibactam in a Patient With Persistent Bacteremia Caused by Klebsiella pneumoniae.

A 30-year-old Korean man with myelodysplastic syndrome admitted hospital due to undifferentiated fever and recurrent skin lesions. He received combination therapy with high doses of meropenem, tigecycline and amikacin, yielding carbapenem resistant Klebsiella pneumoniae (CRKP) harboring K. pneumoniae carbapenemase (KPC)-2 from blood cultures on hospital day (HD) 23. Ceftazidime/avibactam was started at HD 37 and CRKP was eradicated from blood cultures after 5 days. However, ceftazidime/avibactam-resistant CRKP carrying KPC-44 emerged after 26 days of ceftazidime/avibactam treatment and then ceftazidime/avibactam-resistant, carbapenem-susceptible K. pneumoniae carrying KPC-135 was isolated on HD 65. The 3-D homology of KPC protein showed that hot spot changes in the omega loop could be attributed to ceftazidime/avibactam resistance and loss of carbapenem resistance. Whole genome sequencing of serial isolates supported that phenotypic variation was due to clonal evolution than clonal replacement. The treatment regimen was changed from CAZ/AVI to meropenem-based therapy (meropenem 1 g iv q 8 hours and amikacin 600 mg iv per day) starting with HD 72. CAZ/AVI-susceptible CRKP was presented again from blood cultures on HD 84, and the patient expired on HD 85. This is the first Korean report on the acquisition of ceftazidime/avibactam resistance through the emergence of blaKPC variants.

Investigation of Plasmid-Mediated Colistin Resistance Genes (mcr-1-8) in Enterobacterales Isolates.

Background The escalating global rise in multidrug-resistant gram-negative bacteria presents an increasingly substantial threat to patient safety. Over the past decade, carbapenem-resistant Enterobacterales (CRE) have emerged as one of the most critical pathogens in hospital-acquired infections, notably within intensive care units. Colistin has become one of the last-resort antimicrobial agents utilized to combat infections caused by CRE. However, the use of colistin has been accompanied by a notable increase in the prevalence of colistin-resistant bacteria. This study aimed to investigate plasmid-mediated colistin resistance genes ranging from mcr-1 to mcr-8 among members of the Enterobacterales order. Materials and methods This prospective study was conducted in the microbiology laboratory of Afyonkarahisar Health Sciences University Health Research and Practice Center between May 1, 2021 and July 31, 2022. A total of 2,646 Enterobacterales isolates were obtained from all culture-positive clinical samples sent from various clinics. Of these, 79 isolates exhibiting resistance to carbapenem antibiotics were included in the study. Among the 79 isolates, the presence of mcr-1 to mcr-8 genes was investigated in 27 isolates that were shown to be resistant to colistin. The identification of bacteria at the species level and antibiotic susceptibility tests were conducted using the VITEK 2 automated system (bioMérieux, USA). Colistin resistance among Enterobacterales strains exhibiting carbapenem resistance was evaluated using the broth microdilution technique (ComASP™ Colistin, Liofilchem, Italy), in accordance with the manufacturer's instructions. Results In our in vitro investigations, the minimum inhibitory concentration (MIC) values for meropenem were determined to be >8 µg/ml, whereas for colistin, the MIC50 value was >16 µg/ml and the MIC90 value was 8 µg/ml. A total of 27 colistin-resistant strains were identified among the 79 carbapenem-resistant Enterobacterales strains analyzed. The most prevalent agent among colistin-resistant strains was Klebsiella pneumoniae (K. pneumoniae), representing 66.7% of the isolates. This was followed by Proteus mirabilis (P. mirabilis) with 29.6% and Escherichia coli (E. coli) with 3.7%. The colistin resistance rate among carbapenem-resistant strains was found to be 34.2%, with colistin MIC values in strains tested by the broth microdilution method ranging from 4 to >16 µg/ml concentrations. In polymerase chain reaction (PCR) studies, the mcr-1 gene region was successfully detected by real-time PCR in the positive control isolate. Nevertheless, none of the gene regions from mcr-1 to mcr-8 were identified in our study investigating the presence of plasmid-mediated genes using a multiplex PCR kit. Conclusion Although our study demonstrated the presence of increased colistin resistance rates in carbapenem-resistant Enterobacterales isolates, it resulted in the failure to detect genes from mcr-1 to mcr-8 by the multiplex PCR method. Therefore, it is concluded that the colistin resistance observed in Enterobacteriaceae isolates in our region is not due to the mcr genes screened, but to different resistance development mechanisms.

Development and validation of a novel prediction model for Carbapenem-resistant organism infection in a large-scale hospitalized patients.

Carbapenem-resistant organism (CRO) are defined as gram-negative bacteria. The lack of safe and effective antibiotics has led to an increase in incidence rate. The purpose of this study is to establish and determine a risk nomogram to predict CRO infection in hospitalized patients. Hospitalized patients' information were collected from the electronic medical record system of hospital between January 2019 and December 2022. Based on the inclusion and exclusion criteria, we identified 131390 inpatients who met the criteria for this study. For the training cohort, the area under the curves (AUC) for predicting the CRO infection was 0.935. For the validation cohort, the AUC for predicting the CRO infection was 0.937. We have developed the first novel nomogram to predict CRO infection in hospitalized patients, which is reliable and high-performance. The nomogram performs well among hospitalized patients and has good predictive ability.

Co-colonization of different species harboring KPC or NDM carbapenemase in the same host gut: insight of resistance evolution by horizontal gene transfer.

Introduction: The dissemination of carbapenem-resistant Enterobacteriales (CRE) in nosocomial settings is primarily associated with the horizontal transfer of plasmids. However, limited research has focused on the in-host transferability of carbapenem resistance. In this study, ten isolates were collected from gut specimens of five individuals, each hosting two different species, including Escherichia coli, Klebsiella pneumoniae, Klebsiella aerogenes, Enterobacter cloacae, or Citrobacter koseri. Methods: Species identification and antimicrobial susceptibility were determined by MALDI-TOF MS and broth microdilution method. Carbapenemase genes were detected and localized using PCR, S1-PFGE and southern blot. The transferability of carbapenemase genes between species was investigated through filter mating experiments, and the genetic contexts of the plasmids were analyzed using whole genome sequencing. Results and discussion: Our results revealed that each of the ten isolates harbored a carbapenemase gene, including bla NDM-5, bla NDM-1, or bla KPC-2, on a plasmid. Five different plasmids were successfully transferred to recipient cells of E. coli, K. pneumoniae or A. baumannii by transconjugation. The genetic contexts of the carbapenemase gene were remarkably similar between the two CRE isolates from each individual. This study highlights the potential for interspecies plasmid transmission in human gut, emphasizing the colonization of CRE as a significant risk factor for the dissemination of carbapenemase genes within the host. These findings underscore the need for appropriate intestinal CRE screening and colonization prevention.

Tailoring Interventions for Control of Endemic Carbapenem-Resistant Acinetobacter baumannii: An Interrupted Time Series Analysis.

Background: We examined temporal trends in carbapenem-resistant Acinetobacter baumannii (CRAB) infections in a hospital with hyperendemic CRAB and assessed the efficacy of varied infection control strategies in different ward types. Methods: We retrospectively analyzed all CRAB clinical samples from 2006 to 2019 and categorized infections as hospital-onset (HO) or community-onset. We used interrupted time series analysis to assess the impact of interventions on the incidence of all HO-CRAB infections and bloodstream infections (BSIs) at the hospital and ward group levels. Results: Over 14 years, 4009 CRAB infections were identified (89.7% HO), with 813 CRAB BSI (93.2% HO). The incidence per 100 000 patient-days of CRAB infections peaked in 2008 at 79.1, and that of CRAB BSI peaked in 2010 at 16.2. These rates decreased by two-thirds by 2019. In the general intensive care unit (ICU), hand hygiene and environmental cleaning interventions were followed by a significant reduction in the level of HO-CRAB infections, with an additional decrease in the slope after the introduction of active surveillance and 2% chlorhexidine bathing. In the surgical ICU and surgical department, a reduction in slope or level of CRAB infection was observed after moving ventilated patients to single rooms. In medical wards, a multimodal intervention was followed by a reduction in the slope of HO-CRAB infections and BSIs. In wards where CRAB infections were uncommon, the incidence of HO-CRAB infections decreased throughout the study period. Conclusions: Ward-specific variables determine the success of interventions in reducing CRAB infections; therefore, interventions should be tailored to each setting.

In vivo emergence of cefiderocol and ceftazidime/avibactam cross-resistance in KPC-producing Klebsiella pneumoniae following ceftazidime/avibactam -based therapies.

We described the emergence of ceftazidime/avibactam and cefiderocol cross-resistance in patients with KPC-producing Klebsiella pneumoniae infections. All strains with ceftazidime/avibactam and cefiderocol cross-resistance showed point mutations on KPC Ω-loop. Taken together, our results indicate that prolonged exposure to ceftazidime/avibactam can confer cross-resistance to ceftazidime/avibactam and cefiderocol.

Comparison of ERIC carbapenem-resistant Enterobacteriaceae test, BD Phoenix CPO detect panel, and NG-test CARBA 5 for the detection of main carbapenemase types of carbapenem-resistant Enterobacterales.

BACKGROUND: This study aimed to assess the performance of three commercial panels, the ERIC Carbapenem-Resistant Enterobacteriaceae Test (ERIC CRE test), the NG-Test CARBA 5 (NG CARBA 5), and the BD Phoenix CPO Detect Panel (CPO panel), for the detection of main types of carbapenemases among carbapenem-resistant Enterobacterales (CRE). METHODS: We collected 502 isolates of carbapenem-resistant Enterobacterales (CRE) demonstrating intermediate or resistant profiles to at least one carbapenem antibiotic (ertapenem, imipenem, meropenem, or doripenem). Carbapenemase genes and their specific types were identified through multiplex PCR and sequencing methods. Subsequently, the ERIC CRE test, CPO panel, and NG CARBA 5 assay were conducted on these isolates, and the results were compared with those obtained from multiplex PCR. RESULTS: The results indicated that the ERIC CRE test exhibited an overall sensitivity and specificity of 98.1% and 93.6%, respectively, which were comparable to 99.1% and 90.6% for the NG CARBA 5. However, the CPO panel demonstrated a sensitivity of only 56.2% in identifying Ambler classes, exhibiting the poorest sensitivity for class A. Moreover, while the ERIC CRE test outperformed the NG CARBA 5 in identifying multi-gene isolates with multiple carbapenemase-encoding genes, the CPO panel failed to accurately classify these isolates. CONCLUSIONS: Our findings support the utilization of the ERIC CRE test as one of the methods for detecting carbapenemases in clinical laboratories. Nonetheless, further optimization is imperative for the CPO panel to enhance its accuracy in determining carbapenemase classification and address limitations in detecting multi-gene isolates.

Understanding the burden of antibiotic resistance: a decade of carbapenem-resistant Gram-negative bacterial infections in Italian intensive care units.

Introduction: In patients admitted to intensive care units (ICUs), Gram-negative bacteria (GNB) infections pose significant challenges due to their contribution to morbidity, mortality, and healthcare costs. During the SARS-CoV-2 pandemic, Italy witnessed a rise in healthcare-associated infections (HAIs), with GNBs involved in a substantial proportion of cases. Concerningly, carbapenem-resistant GNBs (CR-GNBs) have increased worldwide, posing therapeutic challenges. Methods: Retrospective multicentre study analysing data from over 299,000 patients admitted to Italian ICUs from 2013 to 2022. Results: The study revealed an average of 1.5 infections per patient, with HAIs peaking during the pandemic years. Ventilator associated pneumonia (VAP) emerged as the most common HAI, with Klebsiella spp. and Pseudomonas aeruginosa predominating. Alarmingly, CR-GNBs accounted for a significant proportion of infections, particularly in VAP, bloodstream infections, and intra-abdominal infections. Discussion: Our findings underscore the pressing need for enhanced infection control measures, particularly in the ICU setting, to mitigate the rising prevalence of CR-GNBs and their impact on patient outcomes. The study provides valuable insights into the epidemiology of HAIs in Italian ICUs and highlights the challenges posed by CR-GNBs, especially in the context of the SARS-CoV-2 pandemic, which exacerbated the issue and may serve as a crucial example for the management of future viral pandemics.

Molecular epidemiology and risk factors for carbapenem-resistant Enterobacteriaceae infections during 2020-2021 in Northwest China.

OBJECTIVES: Severe infection caused by Carbapenem-resistant Enterobacteriaceae (CRE) is a challenge for clinical anti-infective therapy, and clinical intervention to improve control of CRE is of great significance. The study aims to determine the molecular epidemiology and risk factors of CRE infections to provide evidence for effective control of nosocomial infection in patients with CRE. METHODS: A total of 192 non-repetitive CRE strains were collected from January 2020 to December 2021 in Northwest China. To explore the risk factors of CRE infection by univariate and Logistic regression analysis, 1:1 case-control study was used to select Carbapenem sensitive Enterobacteriaceae (CSE) infection patients at the same period as the control group. RESULTS: Among the 192 CRE strains, the most common isolates included Klebsiella pneumoniae (Kpn) and Enterobacter cloacae (Ecl). The CRE strain showed the lowest rate of resistance to amikacin at 58.3. 185 CRE strains carried carbapenemase resistance genes of concern in this study. KPC-2 (n=94) was the most common carbapenemase, followed by NDM-1 (n=69), NDM-5 (n=22) and IMP-4 (n=5). OXA-48 and VIM were not detected. And KPC-2 was the most common in all strains. Logistic regression analysis implicated days of invasive ventilator-assisted ventilation (OR=1.452; 95 % CI 1.250～1.686), antibiotic combination therapy (OR=2.149; 95 % CI 1.128～4.094), hypoalbuminemia (OR=6.137; 95 % CI 3.161～11.913), history of immunosuppressant use (OR=25.815; 95 % CI 6.821～97.706) and days of hospitalization (OR=1.020; 95 % CI 1.006～1.035) as independent risk factors associated with CRE infection. Age (OR=0.963; 95% CI 0.943～0.984) and history of hormone use (OR=0.119; 95 % CI 0.028～0.504) were protective factors for CRE infection (P < 0.05). CONCLUSIONS: The resistance of commonly used antibiotics in clinical is severe, and CRE strains mainly carry KPC-2 and NDM-1. Multiple risk factors for CRE infection and their control can effectively prevent the spread of CRE.

Occurrence of carbapenem-resistant hypervirulent Klebsiella pneumoniae in oysters in Egypt: a significant public health issue.

BACKGROUND: The global dissemination of critical-priority carbapenem-resistant hypervirulent Klebsiella pneumoniae (CR-hvKp) via food sources represents a significant public health concern. Epidemiological data on CR-hvKp in oysters in Egypt is limited. This study aimed to investigate the potential role of oysters sold in Egypt as a source for carbapenem-resistant K. pneumoniae (CRKP), hypervirulent K. pneumoniae (hvKp), and CR-hvKp and assess associated zoonotic risks. METHODS: A sample of 330 fresh oysters was randomly purchased from various retail fish markets in Egypt and divided into 33 pools. Bacteriological examination and the identification of Klebsiella pneumoniae were performed. Carbapenem resistance in K. pneumoniae isolates was determined by phenotypic and molecular methods. Additionally, the presence of hypervirulent K. pneumoniae was identified based on virulence gene markers (peg-344, rmpA, rmpA2, iucA, and iroB), followed by a string test. The clustering of CR-hvKp strains was carried out using R with the pheatmap package. RESULTS: The overall prevalence of K. pneumoniae was 48.5% (16 out of 33), with 13 isolates displaying carbapenem resistance, one intermediate resistance, and two sensitive. Both carbapenem-resistant K. pneumoniae and carbapenem-intermediate-resistant K. pneumoniae strains exhibited carbapenemase production, predominantly linked to the blaVIM gene (68.8%). HvKp strains were identified at a rate of 62.5% (10/16); notably, peg-344 was the most prevalent gene. Significantly, 10 of the 13 CRKP isolates possessed hypervirulence genes, contributing to the emergence of CR-hvKp. Moreover, cluster analysis revealed the clustering of two CR-hvKp isolates from the same retail fish market. CONCLUSION: This study provides the first insight into the emergence of CR-hvKp among oysters in Egypt. It underscores the potential role of oysters as a source for disseminating CR-hvKp within aquatic ecosystems, presenting a possible threat to public health.

Membrane-targeting, ultrashort lipopeptide acts as an antibiotic adjuvant and sensitizes MDR gram-negative pathogens toward narrow-spectrum antibiotics.

Globally, infections due to multi-drug resistant (MDR) Gram-negative bacterial (GNB) pathogens are on the rise, negatively impacting morbidity and mortality, necessitating urgent treatment alternatives. Herein, we report a detailed bio-evaluation of an ultrashort, cationic lipopeptide 'SVAP9I' that demonstrated potent antibiotic activity and acted as an adjuvant to potentiate existing antibiotic classes towards GNBs. Newly synthesized lipopeptides were screened against ESKAPE pathogens and cytotoxicity assays were performed to evaluate the selectivity index (SI). SVAP9I exhibited broad-spectrum antibacterial activity against critical MDR-GNB pathogens including members of Enterobacteriaceae (MIC 4-8 mg/L), with a favorable CC50 value of ≥100 mg/L and no detectable resistance even after 50th serial passage. It demonstrated fast concentration-dependent bactericidal action as determined via time-kill analysis and also retained full potency against polymyxin B-resistant E. coli, indicating distinct mode of action. SVAP9I targeted E. coli's outer and inner membranes by binding to LPS and phospholipids such as cardiolipin and phosphatidylglycerol. Membrane damage resulted in ROS generation, depleted intracellular ATP concentration and a concomitant increase in extracellular ATP. Checkerboard assays showed SVAP9I's synergism with narrow-spectrum antibiotics like vancomycin, fusidic acid and rifampicin, potentiating their efficacy against MDR-GNB pathogens, including carbapenem-resistant Acinetobacter baumannii (CRAB), a WHO critical priority pathogen. In a murine neutropenic thigh infection model, SVAP9I and rifampicin synergized to express excellent antibacterial efficacy against MDR-CRAB outcompeting polymyxin B. Taken together, SVAP9I's distinct membrane-targeting broad-spectrum action, lack of resistance and strong in vitro andin vivopotency in synergism with narrow spectrum antibiotics like rifampicin suggests its potential as a novel antibiotic adjuvant for the treatment of serious MDR-GNB infections.

Inhibitory Potential of Bifidobacterium longum FB1-1 Cell-Free Supernatant against Carbapenem-Resistant Klebsiella pneumoniae Drug Resistance Spread.

The widespread dissemination of carbapenem-resistant Klebsiella pneumoniae (CRKP) and its drug resistance transfer poses a global public health threat. While previous studies outlined CRKP's drug resistance mechanism, there is limited research on strategies inhibiting CRKP drug resistance spread. This study investigates the potential of Bifidobacterium longum (B. longum) FB1-1, a probiotic, in curbing the spread of drug resistance among CRKP by evaluating its cell-free supernatant (CFS) for antibacterial activity. Evaluating the inhibitory effect of FB1-1 CFS on CRKP drug resistance spread involved analyzing its impact on drug resistance and virulence gene expression; drug resistance plasmid transfer FB1-1 CFS exhibited an MIC range of 125 µL/mL against CRKP. After eight hours of co-culture, CFS achieved a 96% and 100% sterilization rate at two and four times the MIC, respectively. At sub-inhibitory concentrations (1/2× MIC), FB1-1 CFS reduced the expression of the bla_KPC gene, which is pivotal for carbapenem resistance, by up to 62.13% across different CRKP strains. Additionally, it markedly suppressed the expression of the uge gene, a key virulence factor, by up to 91%, and the fim_H gene, essential for bacterial adhesion, by up to 53.4%. Our study primarily focuses on determining the inhibitory effect of FB1-1 CFS on CRKP strains harboring the bla_KPC gene, which is a critical resistance determinant in CRKP. Furthermore, FB1-1 CFS demonstrated the ability to inhibit the transfer of drug resistance plasmids among CRKP strains, thus limiting the horizontal spread of resistance genes. This study highlights FB1-1 CFS's inhibitory effect on CRKP drug resistance spread, particularly in strains carrying the bla_KPC gene, thus offering a novel idea and theoretical foundation for developing antibacterial drugs targeting CRKP resistance.

Tracking emergence and outbreak of Klebsiella pneumoniae co-producing NDM-1 and KPC-2 after sulfamethoxazole-trimethoprim treatment: Insights from genetic analysis.

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

Synergistic Combination of Aztreonam and Ceftazidime-Avibactam-A Promising Defense Strategy against OXA-48 + NDM Klebsiella pneumoniae in Romania.

With the increasing burden of carbapenem-resistant Klebsiella pneumoniae (CR-Kp), including high rates of healthcare-associated infections, treatment failure, and mortality, a good therapeutic strategy for attacking this multi-resistant pathogen is one of the main goals in current medical practice and necessitates the use of novel antibiotics or new drug combinations. OBJECTIVES: We reviewed the clinical and microbiological outcomes of seven patients treated at the "Agrippa Ionescu" Clinical Emergency Hospital between October 2023 and January 2024, aiming to demonstrate the synergistic activity of the ceftazidime-avibactam (C/A) plus aztreonam (ATM) combination against the co-producers of blaNDM + blaOXA-48-like CR-Kp. MATERIAL AND METHODS: Seven CR-Kp with blaNDM and blaOXA-48 as resistance mechanisms were tested. Seven patients treated with C/A + ATM were included. The synergistic activity of C/A + ATM was proven through double-disk diffusion in all seven isolates. Resistance mechanisms like KPC, VIM, OXA-48, NDM, IMP, and CTX-M were assessed through immunochromatography. RESULTS: With a mean of nine days of treatment with the synergistic combination C/A + ATM, all patients achieved clinical recovery, and five achieved microbiological recovery. CONCLUSIONS: With the emerging co-occurrence of blaOXA-48 and blaNDM among Kp in Romania, the combination of C/A and ATM could be a promising therapeutic option.