Multidrug-resistant Gram-negative Bacteria Research Articles

Daunorubicin resensitizes Gram-negative superbugs to the last-line antibiotics and prevents the transmission of antibiotic resistance

Although meropenem, colistin, and tigecycline are recognized as the last-line antibiotics for multidrug-resistant Gram-negative bacteria (MDR-GN), the emergence of mobile resistance genes such as blaNDM, mcr, and tet(X) severely compromises their clinical effectiveness. Developing novel antibiotic adjuvants to restore the effectiveness of existing antibiotics provides a feasible approach to address this issue. Herein, we discover that a Food and Drug Administration (FDA)-approved drug daunorubicin (DNR) drastically potentiates the activity of last-resort antibiotics against MDR-GN pathogens and biofilm-producing bacteria. Furthermore, DNR effectively inhibits the evolution and spread of colistin and tigecycline resistance. Mechanistically, DNR and colistin combination exacerbates membrane disruption, induces DNA damage and the massive production of reactive oxygen species (ROS), ultimately leading to bacterial cell death. Importantly, DNR restores the effectiveness of colistin in Galleria mellonella and murine models of infection. Collectively, our findings provide a potential drug combination strategy for treating severe infections elicited by Gram-negative superbugs.

Exploring Cefiderocol Resistance Mechanisms in Burkholderia pseudomallei.

Cefiderocol is a siderophore cephalosporin designed mainly for treatment of infections caused by β-lactam and multidrug-resistant Gram-negative bacteria. Burkholderia pseudomallei clinical isolates are usually highly cefiderocol susceptible, with in vitro resistance found in a few isolates. Resistance in clinical B. pseudomallei isolates from Australia is caused by a hitherto uncharacterized mechanism. We show that, like in other Gram-negatives, the PiuA outer membrane receptor plays a major role in cefiderocol nonsusceptibility in isolates from Malaysia.

The Antimicrobial Effects of Colistin Encapsulated in Chelating Complex Micelles for the Treatment of Multi-Drug-Resistant Gram-Negative Bacteria: A Pharmacokinetic Study.

Background: Infections caused by multi-drug-resistant Gram-negative bacteria (MDR-GNB) are an emerging problem globally. Colistin is the last-sort antibiotic for MDR-GNB, but its toxicity limits its clinical use. We aimed to test the efficacy of colistin-loaded micelles (CCM-CL) against drug-resistant Pseudomonas aeruginosa and compare their safety with that of free colistin in vitro and in vivo. Materials and methods: We incorporated colistin into chelating complex micelles (CCMs), thus producing colistin-loaded micelles (CCM-CL), and conducted both safety and efficacy surveys to elucidate their potential uses. Results: In a murine model, the safe dose of CCM-CL was 62.5%, which is much better than that achieved after the intravenous bolus injection of 'free' colistin. With a slow drug infusion, the safe dose of CCM-CL reached 16 mg/kg, which is double the free colistin, 8 mg/kg. The area under the curve (AUC) levels for CCM-CL were 4.09- and 4.95-fold higher than those for free colistin in terms of AUC0-t and AUC0-inf, respectively. The elimination half-lives of CCM-CL and free colistin groups were 12.46 and 102.23 min, respectively. In the neutropenic mice model with carbapenem-resistant Pseudomonas aeruginosa pneumonia, the 14-day survival rate of the mice treated with CCM-CL was 80%, which was significantly higher than the 30% in the free colistin group (p < 0.05). Conclusions: Our results showed that CCM-CL, an encapsulated form of colistin, is safe and effective, and thus may become a drug of choice against MDR-GNB.

A spectrum of non-spore-forming fermentative and non-fermentative Gram-negative bacteria: multi-drug resistance, extended-spectrum beta-lactamase, and carbapenemase production

BackgroundIn developing countries, the co-existence of a high burden of infectious diseases caused by Gram-negative bacteria and the rapid increase and spread of multidrug-resistant bacteria have become a serious health threat.ObjectiveProfiling of Gram-negative bacteria and determining the magnitude of their antimicrobial resistance among patients.ResultsA total of 175 non-spore-forming Gram-negative bacteria were isolated from 873 different clinical samples. Of a total of 175 bacteria, 154 (88%) were fermentative Gram-negative bacteria, while 21 (12%) were non-fermentative Gram-negative bacteria. E. coli with a frequency of 58.3% and K. pneumoniae with a frequency of 18.3% were the predominant fermentative Gram-negative bacteria, while P. aeruginosa 9 (5.1%) and A. baumannii 6 (3.4%) were the predominant non-fermentative Gram-negative bacteria. The highest percentage level of antibiotic resistance was seen against ampicillin (86%), and the lowest against meropenem (9.8). About 49 (28%) Gram-negative bacilli were positive for ESBLase. The overall prevalence rate of MDR bacteria was 80.5%, of which 100% of A. baumannii, 90.6% of K. pneumonia. Sixteen isolates were resistant to meropenem, out of which 11 tested for carbapenemase production. Five of the nine were metallo-lactamase producers, with the remaining four being serine carbapenemase producers.ConclusionThe prevalence of Gram-negative bacterial infection was found to be 20%, with a significant proportion (80.0%) due to fermentative Gram-negative bacteria and the remaining 20% due to non-fermentative Gram-negative bacteria. The study has also demonstrated a high prevalence rate of MDR, ESBLase, and carbapenemase-producing Gram-negative bacteria. Antimicrobial resistance of Gram-negative bacteria should be monitored on a regular basis, and an effective infection control program should be implemented.

Antimicrobial Resistance in Romania: Updates on Gram-Negative ESCAPE Pathogens in the Clinical, Veterinary, and Aquatic Sectors.

Multidrug-resistant Gram-negative bacteria such as Acinetobacter baumannii, Pseudomonas aeruginosa, and members of the Enterobacterales order are a challenging multi-sectorial and global threat, being listed by the WHO in the priority list of pathogens requiring the urgent discovery and development of therapeutic strategies. We present here an overview of the antibiotic resistance profiles and epidemiology of Gram-negative pathogens listed in the ESCAPE group circulating in Romania. The review starts with a discussion of the mechanisms and clinical significance of Gram-negative bacteria, the most frequent genetic determinants of resistance, and then summarizes and discusses the epidemiological studies reported for A. baumannii, P. aeruginosa, and Enterobacterales-resistant strains circulating in Romania, both in hospital and veterinary settings and mirrored in the aquatic environment. The Romanian landscape of Gram-negative pathogens included in the ESCAPE list reveals that all significant, clinically relevant, globally spread antibiotic resistance genes and carrying platforms are well established in different geographical areas of Romania and have already been disseminated beyond clinical settings.

Polymyxin-based fluorescent probes to combat Gram-negative antimicrobial resistance

Reliable diagnostic approaches especially those targeting critical Gram-negative bacteria are urgently needed for the prevention of antimicrobial resistance. Polymyxin B (PMB) which specifically targets the outer membrane of Gram-negative bacteria is the last-line antibiotic against life-threatening multidrug-resistant Gram-negative bacteria. However, increasing number of studies have reported the spread of PMB-resistant strains. With the aim to specifically detect Gram-negative bacteria and potentially reduce the irrational use of antibiotics, we herein rationally designed two Gram-negative bacteria specific fluorescent probes based on our previous activity-toxicity optimization of PMB. The in vitro probe PMS-Dns showed fast and selective labeling of Gram-negative pathogens in complex biological cultures. Subsequently, we constructed the caged in vivo fluorescent probe PMS-Cy-NO2 by conjugating bacterial nitroreductase (NTR)-activatable positive charged hydrophobic near-infrared (NIR) fluorophore with polymyxin scaffold. Significantly, PMS-Cy-NO2 exhibited excellent Gram-negative bacterial detection capability with the differentiation between Gram-positive and Gram-negative in a mouse skin infection model.

In Vitro Activity of Cefiderocol on Multiresistant Bacterial Strains and Genomic Analysis of Two Cefiderocol Resistant Strains.

Cefiderocol is a new siderophore cephalosporin that is effective against multidrug-resistant Gram-negative bacteria, including carbapenem-resistant strains. The aim of this study was to evaluate the activity of this new antimicrobial agent against a collection of pathogens using broth microdilution assays and to analyze the possible mechanism of cefiderocol resistance in two resistant Klebsiella pneumoniae isolates. One hundred and ten isolates were tested, comprising 67 Enterobacterales, two Acinetobacter baumannii, one Achromobacter xylosoxidans, 33 Pseudomonas aeruginosa and seven Stenotrophomonas maltophilia. Cefiderocol showed good in vitro activity, with an MIC < 2 μg/mL, and was able to inhibit 94% of the tested isolates. We observed a resistance rate of 6%. The resistant isolates consisted of six Klebsiella pneumoniae and one Escherichia coli, leading to a resistance rate of 10.4% among the Enterobacterales. Whole-genome sequencing analysis was performed on two cefiderocol-resistant Klebsiella pneumoniae isolates to investigate the possible mutations responsible for the observed resistance. Both strains belonged to ST383 and harbored different resistant and virulence genes. The analysis of genes involved in iron uptake and transport showed the presence of different mutations located in fhuA, fepA, iutA, cirA, sitC, apbC, fepG, fepC, fetB, yicI, yicJ, and yicL. Furthermore, for the first time, to the best of our knowledge, we described two Klebsiella pneumoniae isolates that synthesize a truncated fecA protein due to the transition from G to A, leading to a premature stop codon in the amino acid position 569, and a TonB protein carrying a 4-amino acid insertion (PKPK) after Lysine 103. In conclusion, our data show that cefiderocol is an effective drug against multidrug-resistant Gram-negative bacteria. However, the higher resistance rate observed in Enterobacterales underlines the need for active surveillance to limit the spread of these pathogens and to avoid the risks associated with the emergence of resistance to new drugs.

Synthesis and Antibiotic Activity of Chitosan-Based Comb-like Co-Polypeptides

Infections caused by multidrug-resistant Gram-negative bacteria have been named one of the most urgent global health threats due to antimicrobial resistance. Considerable efforts have been made to develop new antibiotic drugs and investigate the mechanism of resistance. Recently, Anti-Microbial Peptides (AMPs) have served as a paradigm in the design of novel drugs that are active against multidrug-resistant organisms. AMPs are rapid-acting, potent, possess an unusually broad spectrum of activity, and have shown efficacy as topical agents. Unlike traditional therapeutics that interfere with essential bacterial enzymes, AMPs interact with microbial membranes through electrostatic interactions and physically damage cell integrity. However, naturally occurring AMPs have limited selectivity and modest efficacy. Therefore, recent efforts have focused on the development of synthetic AMP analogs with optimal pharmacodynamics and an ideal selectivity profile. Hence, this work explores the development of novel antimicrobial agents which mimic the structure of graft copolymers and mirror the mode of action of AMPs. A family of polymers comprised of chitosan backbone and AMP side chains were synthesized via the ring-opening polymerization of the N-carboxyanhydride of l-lysine and l-leucine. The polymerization was initiated from the functional groups of chitosan. The derivatives with random- and block-copolymer side chains were explored as drug targets. These graft copolymer systems exhibited activity against clinically significant pathogens and disrupted biofilm formation. Our studies highlight the potential of chitosan-graft-polypeptide structures in biomedical applications.

Phentolamine Significantly Enhances Macrolide Antibiotic Antibacterial Activity against MDR Gram-Negative Bacteria.

Multidrug-resistant (MDR) Gram-negative bacterial infections have limited treatment options due to the impermeability of the outer membrane. New therapeutic strategies or agents are urgently needed, and combination therapies using existing antibiotics are a potentially effective means to treat these infections. In this study, we examined whether phentolamine can enhance the antibacterial activity of macrolide antibiotics against Gram-negative bacteria and investigated its mechanism of action. Synergistic effects between phentolamine and macrolide antibiotics were evaluated by checkerboard and time-kill assays and in vivo using a Galleria mellonella infection model. We utilized a combination of biochemical tests (outer membrane permeability, ATP synthesis, ΔpH gradient measurements, and EtBr accumulation assays) with scanning electron microscopy to clarify the mechanism of phentolamine enhancement of macrolide antibacterial activity against Escherichia coli. In vitro tests of phentolamine combined with the macrolide antibiotics erythromycin, clarithromycin, and azithromycin indicated a synergistic action against E. coli test strains. The fractional concentration inhibitory indices (FICI) of 0.375 and 0.5 indicated a synergic effect that was consistent with kinetic time-kill assays. This synergy was also seen for Salmonella typhimurium, Klebsiella pneumoniae, and Actinobacter baumannii but not Pseudomonas aeruginosa. Similarly, a phentolamine/erythromycin combination displayed significant synergistic effects in vivo in the G. mellonella model. Phentolamine added singly to bacterial cells also resulted in direct outer membrane damage and was able to dissipate and uncouple membrane proton motive force from ATP synthesis that, resulted in enhanced cytoplasmic antibiotic accumulation via reduced efflux pump activity. Phentolamine potentiates macrolide antibiotic activity via reducing efflux pump activity and direct damage to the outer membrane leaflet of Gram-negative bacteria both in vitro and in vivo.

Prevalence and Genomic Characteristics of mcr-Positive Escherichia coli Strains Isolated from Humans, Pigs, and Foods in China.

Colistin is one of the last-resort antibiotics for treating infections caused by multidrug-resistant (MDR) Gram-negative bacteria. However, mcr genes conferring resistance to colistin have been widely identified, which is considered a global threat to public health. Here, we investigated the prevalence and characteristics of mcr-harboring Escherichia coli strains isolated from humans, animals, and foods in China by PCR, antimicrobial susceptibility testing, conjugation experiments, molecular typing, genome sequencing, and bioinformatics analysis. In total, 135 mcr-1-harboring E. coli isolates were acquired from 847 samples, and 6 isolates carried mcr-3. Among them, 131 isolates were MDR bacteria. Sixty-five resistance genes conferring resistance to multiple antimicrobials were identified in 135 isolates. The diverse pulsed-field gel electrophoresis (PFGE) patterns and sequence types (STs) of mcr-1-carrying isolates demonstrated that clonal dissemination was not the dominant mode of mcr-1 transmission. Seven types of plasmids were able to carry mcr-1 in this study, including IncI2, IncX4, IncHI2, p0111, IncY, and two hybrid plasmids. The genetic structures carrying mcr-1 of 60 isolates were successfully transferred into the recipient, including 25 IncI2 plasmids, 23 IncX4 plasmids, and an IncHI2 plasmid. mcr-1-pap2 was the dominant mcr-1-bearing structure, followed by ISApl1-mcr-1-pap2-ISApl1 (Tn6330) and ISApl1-mcr-1-pap2, among 7 mcr-1-bearing structures of 135 isolates. In conclusion, IncI2, IncX4, and IncHI2 plasmids were the major vectors spreading mcr-1 from different geographical locations and sources. The prevalence of Tn6330 may accelerate the transmission of mcr-1. Continuous surveillance of mcr-1 and variants in bacteria is vital for evaluating the public health risk posed by mcr genes. IMPORTANCE The spread of polymyxin-resistant Enterobacteriaceae poses a significant threat to public health and challenges the therapeutic options for treating infections on a global level. In this study, mcr-1-bearing ST10 E. coli was isolated from pigs, pork, and humans simultaneously, which demonstrated that ST10 E. coli was an important vehicle for the spread of mcr-1 among animals, foods, and humans. The high prevalence of mcr-1-positive E. coli strains in pigs and pork and the horizontal transmission of mcr-1-bearing plasmids in diverse E. coli strains suggest that pigs and pork are important sources of mcr-1-positive strains in humans and pose a potential threat to public health. Additional research on the prevalence and characteristics of mcr-1-positive E. coli is still required to facilitate early warning to improve polymyxin management in hospitals.

Risk Factors for Infection With Multidrug-Resistant Gram-Negative Bacteria in a Tertiary Care Hospital in Saudi Arabia: A Case-Control Study.

Background The increase in the incidence of multidrug-resistant (MDR) organisms especially Gram-negative bacteria (GNB) in healthcare facilities is a serious cause of concern. This study identified risk factors for the infection with these MDR GNB, such asAcinetobacterbaumannii, Klebsiella pneumoniae, Pseudomonas aeruginosa, and Escherichia coli to inform healthcare workers about strategies for their containment. Methods A case-control study was carried out at a tertiary care hospital where 100 patients with healthcare-associated infections (infections arising 48 hours after admission) caused by MDR GNB were compared with two control groups, i.e., 100 patients with healthcare-associated infections caused by non-MDR GNB (not meeting the criteria of MDR) and 100 patients without infection caused by GNB. MDR bacteria were defined as the ones that were non-susceptible to at least one antibiotic in three or more classes of antibiotics. The data were analyzed using descriptive statistics (frequency and percentage of categorical variables). Multivariate regression analysis was undertaken to identify significant predictors of MDR GNB. Odds ratios with 95% confidence intervals were calculated, and the level of significance was determined at p-value < 0.05. Results A total of 388 organisms were isolated during four months (January-April 2015) from 332 patients. Fifty-six (17%) of the patients were infected with more than one organism. Among the MDR bacteria, the most dominant MDR organism wasA. baumannii(38%), followed byK.pneumoniae(31%),P. aeruginosa(20%), andE. coli(11%). Among the non-MDR organisms, the most dominant wasP. aeruginosa(47%), followed byE. coli (32%),K.pneumoniae (18%), andA. baumannii(3%). Patients with MDR organisms compared with the first control group (patients with non-MDR organisms) showed that prior antibiotic use (p-value: 0.001), intensive care unit (ICU) admission (p-value: 0.001), and indwelling medical devices (p-value: 0.005) were significant risk factors for MDR infections. It was also found that the risk factors for MDR GNB infection were the same in the second control group (patients without infection): prior antibiotic use (p-value: 0.002), ICU admission (p-value: 0.001), and indwelling medical devices (p-value: 0.03). Based on the comparison of the two control groups, prolonged hospital stays of more than five days (p-value: 0.001), immunosuppressive therapy (p-value: 0.02), and over 60 years of age (p-value: 0.02) were significant risk factors for non-MDR infection. Conclusion The risk factors identified in our study provide guidance to healthcare workers for the prevention and containment of MDR GNB.

Bloodstream infections due to Gram-negative bacteria in patients with hematologic malignancies: updated epidemiology and risk factors for multidrug-resistant strains in an Italian perspective survey

Bloodstream infections (BSI) caused by Gram-negative bacteria (GNB) in patients with hematological malignancies (HM) have been associated with high mortality rates, particularly with infections caused by antibiotic-resistant strains.A multicenter cohort study including all consecutive episodes of GNB BSI in HM patients was conducted to update the epidemiology and antibiotic resistance patterns (compared to our previous survey conducted between 2009 and 2012) and investigate risk factors for GNB BSI due to multidrug-resistant (MDR) isolates.A total of 834 GNB were recovered in 811 BSI episodes from January 2016 to December 2018. Compared to the previous survey, there was a significant reduction in use of fluoroquinolone prophylaxis and a significant recovery in susceptibility rates to ciprofloxacin among Pseudomonas aeruginosa, Escherichia coli and Enterobacter cloacae isolates. In addition, there was a shift to a significantly increased susceptibility of P. aeruginosa isolates to ceftazidime, meropenem, and gentamicin. A total of 256/834 (30.7%) isolates were MDR. In multivariable analysis, MDR bacteria culture-positive surveillance rectal swabs, previous therapy with aminoglycosides and carbapenems, fluoroquinolone prophylaxis, and time at risk were independently associated with MDR GNB BSI.In conclusion, despite the persistence of a high prevalence of MDR GNB, there was a shift to a reduced use of fluoroquinolone prophylaxis and increased rates of susceptibility to fluoroquinolones in almost all isolates and to almost all antibiotics tested among P. aeruginosa isolates, compared to our previous survey. Fluoroquinolone prophylaxis and previous rectal colonization by MDR bacteria were independent risk factors for MDR GNB BSI in the present study.

Contamination of Automated Teller Machines Surfaces with Multi-drug Resistance Gram-negative Bacteria in Dar es Salaam, Tanzania

Background: In Tanzania, little is known about the proportion of Multi-drug resistance (MDR) Gram-negative bacteria contamination on Automated Teller Machine (ATMs) surfaces. The study aimed to determine the proportion of MDR Gram-negative bacteria contamination on ATMs surfaces, antimicrobial resistance patterns as well as associated factors. Methodology: A cross-sectional study was conducted between January and March -2021 in Dar es Salaam, involving 298 ATMs. Cultures were performed on Mac-Conkey agar while antimicrobial susceptibility was done using the Kirby Bauer disc diffusion method with Klebsiella pneumoniae ATCC 700603 and Escherichia coli ATCC 25922 used as controls. Data analysis was done using STATA version 15.1. Chi-square and Modified Poisson regression was used to assess factors associated with MDR contamination. Data was presented as prevalence ratio (PR) and 95% Confidence Interval. A p-value of &lt;.05 was considered statistically significant. Results: More than half (55.4%) of ATMs in Dar es Salaam are contaminated with Gram negative bacteria, mostly by Klebsiella pneumoniae 18.5% (31/168). The highest level of resistance was observed against ampicillin (68.9%). About one-third (34.5%) of the isolates were MDR. About 35.7% were Extended-Spectrum Beta-Lactamases (ESBL) producers while 19.6% were quinolone/ fluoroquinolones-resistance. Risk factors for contamination of ATMs included highly populated location such as; Ubungo (PR adjusted = 3.62, 95%CI = 1.58-8.30, P=.002), Kigamboni (PR adjusted = 2.78, 95%CI = 1.20-6.42, P=.017), and Temeke (PR adjusted = 2.75, 95%CI = 1.04-3.72, P=.023), and less frequent cleaned ATMs (PR adjusted = 1.98, 95%CI = 1.04-3.73, P=.04). Conclusions: More than half of ATMs in Dar es Salaam are contaminated with Gram-negative and one-third of them with MDR bacteria, especially those located in highly populated areas and those that are less frequently cleaned. This calls for interventional measures regarding public awareness of ATMs as potential vehicles for the transmission of infectious agents.

The Challenge of Bacteremia Treatment due to Non-Fermenting Gram-Negative Bacteria.

Nosocomial infections caused by non-fermenting Gram-negative bacteria are a real challenge for clinicians, especially concerning the accuracy of empirical treatment. This study aimed to describe the clinical characteristic, empirical antibiotic therapy, accuracy of these prescriptions for appropriate coverage and risk factor for clinical failure of bloodstream infections due to non-fermenting Gram-negative bacilli. This retrospective, observational cohort study was conducted between January 2016 and June 2022. Data were collected from the hospital's electronic record. The statistic tests corresponding to each objective were applied. A multivariate logistic regression was performed. Among the total 120 patients included in the study, the median age was 63.7 years, and 79.2% were men. Considering the appropriate empirical treatment rate by species, inappropriate treatment for S. maltophilia was 72.4% (p = 0.088), for A. baumanii 67.6% and 45.6% for P. aeruginosa. Clinical success was achieved in 53.3%, and overall, 28-day mortality was 45.8%. ICU admission, sepsis or shock septic, age, previous antibiotic treatment and contact with healthcare facilities were independently associated with clinical failure. In conclusion, bloodstream infection produced by multidrug-resistant non-fermenting Gram-negative bacteria is a significant therapeutic management challenge for clinicians. The accuracy of empirical treatment is low due to the fact that it is not recommended to cover these microorganisms empirically, especially S. maltophilia and A. baumanii.

Gram-negative ESKAPE bacteria bloodstream infections in patients during the COVID-19 pandemic.

Bloodstream infections due to bacteria are a highly consequential nosocomial occurrences and the organisms responsible for them are usually multidrug-resistant. The aims of this study were to describe the incidence of bacteremia caused by Gram-negative ESKAPE bacilli during the COVID-19 pandemic and characterize the clinical and microbiological findings including antimicrobial resistance. A total of 115 Gram-negative ESKAPE isolates were collected from patients with nosocomial bacteremia (18% of the total bacteremias) in a tertiary care center in Mexico City from February 2020 to January 2021. These isolates were more frequently derived from the Respiratory Diseases Ward (27), followed by the Neurosurgery (12), Intensive Care Unit (11), Internal Medicine (11), and Infectious Diseases Unit (7). The most frequently isolated bacteria were Acinetobacter baumannii (34%), followed by Klebsiella pneumoniae (28%), Pseudomonas aeruginosa (23%) and Enterobacter spp (16%). A. baumannii showed the highest levels of multidrug-resistance (100%), followed by K. pneumoniae (87%), Enterobacter spp (34%) and P. aeruginosa (20%). The bla CTX-M-15 and bla TEM-1 genes were identified in all beta-lactam-resistant K. pneumoniae (27), while bla TEM-1 was found in 84.6% (33/39) of A. baumannii isolates. The carbapenemase gene bla OXA-398 was predominant among carbapenem-resistant A. baumannii (74%, 29/39) and bla OXA-24was detected in four isolates. One P. aeruginosa isolate was bla VIM-2 gene carrier, while two K. pneumoniae and one Enterobacter spp were bla NDM gene carriers. Among colistin-resistant isolates mcr-1 gene was not detected. Clonal diversity was observed in K. pneumoniae, P. aeruginosa and Enterobacter spp. Two outbreaks caused by A. baumannii ST208 and ST369 were detected, both belonging to the clonal complex CC92 and IC2. A. baumannii was associated with a death rate of 72% (28/32), most of them (86%, 24/28) extensively drug-resistant or pandrug-resistant isolates, mainly in patients with COVID-19 (86%, 24/28) in the Respiratory Diseases Ward. A. baumannii isolates had a higher mortality rate (72%), which was higher in patients with COVID-19. There was no statistically significant association between the multidrug-resistant profile in Gram-negative ESKAPE bacilli and COVID-19 disease. The results point to the important role of multidrug-resistant Gram-negative ESKAPE bacteria causing bacteremia in nosocomial settings before and during the COVID-19 epidemic. Additionally, we were unable to identify a local impact of the COVID-19 pandemic on antimicrobial resistance rates, at least in the short term.

Population pharmacokinetic analysis and dosing optimization of polymyxin B in critically ill patients.

Objectives: Since the global broadcast of multidrug-resistant gram-negative bacteria is accelerating, the use of Polymyxin B is sharply increasing, especially in critically ill patients. Unsatisfactory therapeutic effects were obtained because of the abnormal physiological function in critically ill patients. Therefore, the determination of optimal polymyxin B dosage becomes highly urgent. This study aimed to illustrate the polymyxin B pharmacokinetic characteristics by defining the influencing factors and optimizing the dosing regimens to achieve clinical effectiveness. Methods: Steady-state concentrations of polymyxin B from twenty-two critically ill patients were detected by a verified liquid chromatography-tandem mass spectrometry approach. The information on age, weight, serum creatinine, albumin levels, and Acute Physiology and Chronic Health Evaluation-II (APACHE-II) score was also collected. The population PK parameters were calculated by the non-parametric adaptive grid method in Pmetrics software, and the pharmacokinetic/pharmacodynamics target attainment rate was determined by the Monte Carlo simulation method. Results: The central clearance and apparent volume of distribution for polymyxin B were lower in critically ill patients (1.24 ± 0.38Lh-1 and 16.64 ± 12.74 L, respectively). Moreover, albumin (ALB) levels can be used to explain the variability in clearance, and age can be used to describe the variability in the apparent volume of distribution. For maintaining clinical effectiveness and lowering toxicity, 75mg q12h is the recommended dosing regimen for most patients suffering from severe infections. Conclusion: This study has clearly defined that in critically ill patients, age and ALB levels are potentially important factors for the PK parameters of polymyxin B. Since older critically ill patients tend to have lower ALB levels, so higher dosages of polymyxin B are necessary for efficacy.

Polymyxin B exerts nephrotoxicity effects via inhibition of the Nrf2/NQO1 pathway-mediated antioxidant response.

Polymyxin B (PMB) is a polypeptide antibiotic widely used in treating multidrug-resistant Gram-negative bacteria. However, nephrotoxicity is a serious adverse effect that limits its clinical use. Therefore, clarification of the molecular mechanism of PMB-induced renal injury is essential. Our study aimed to explore possible mechanisms of PMB-induced nephrotoxicity in vivo and in vitro. Mice were treated with PMB to construct the kidney injury model. The antioxidant capacity was assessed by measuring the superoxide dismutase (SOD) and catalase (CAT) activities and the glutathione (GSH)and malondialdehyde (MDA) contents. The pathway of the nuclear factor erythroid 2-related factor 2/NADH quinone oxidoreductase 1 (Nrf2/NQO1) was examined after PMB treatment in NRK-52E cells and mice. Finally, the expressions of genes and proteins (Bax, Bcl-2, Caspase-3, Caspase-9) related to apoptosis were evaluated through quantitative polymerasechain reaction and western blot assay. The study verified PMB-induced nephrotoxicity in mice and NRK-52E cells in a dose- and time-dependent manner. PMB treatment significantly decreased the expression of Nrf2 and its downstream target gene NQO1 and increased the apoptosis-related proteins expression. In summary, our results suggested that PMB-induced oxidative stress damage by inhibiting the Nrf2/NQO1 pathway and promoting apoptosis in kidney tissues.

Flufenamic Acid, a Promising Agent for the Sensitization of Colistin-Resistant Gram-Negative Bacteria to Colistin.

The continuous development of multidrug-resistant (MDR) Gram-negative bacteria poses a serious risk to public health on a worldwide scale. Colistin is used as the last-line antibiotic for the treatment of MDR pathogens, and colistin-resistant (COL-R) bacterial emergence thus has the potential to have a severe adverse impact on patient outcomes. In this study, synergistic activity was observed when colistin and flufenamic acid (FFA) were combined and used for the in vitro treatment of clinical COL-R Pseudomonas aeruginosa, Escherichia coli, Klebsiella pneumoniae, and Acinetobacter baumannii strains, as shown by checkerboard and time-kill assays. Crystal violet staining and scanning electron microscopy revealed the synergistic action of colistin-FFA against biofilms. When used to treat murine RAW264.7 macrophages, this combination did not induce any adverse toxicity. Strikingly, the survival rates of bacterially infected Galleria mellonella larvae were improved by such combination treatment, which was also sufficient to reduce the measured bacterial loads in a murine thigh infection model. Mechanistic propidium iodide (PI) staining analysis further demonstrated the ability of these agents to alter bacterial permeability in a manner that enhanced the efficacy of colistin treatment. Together, these data thus demonstrate that colistin and FFA can be synergistically combined to combat the spread of COL-R Gram-negative bacteria, providing a promising therapeutic tool with the potential to protect against COL-R bacterial infections and improve patient outcomes. IMPORTANCE Colistin is a last-line antibiotic used for the treatment of MDR Gram-negative bacterial infections. However, increasing resistance to it has been observed during clinical treatment. In this work, we assessed the efficacy of the combination of colistin and FFA for the treatment of COL-R bacterial isolates, demonstrating that the combined treatment has effective antibacterial and antibiofilm activities. Due to its low cytotoxicity and good therapeutic effects in vitro, the colistin-FFA combination may be a potential candidate for research into a resistance-modifying agent to combat infections caused by COL-R Gram-negative bacteria.

Antibiotic susceptibility and biofilm formation of multi-drug resistant Gram-negative bacteria

Antibiotic susceptibility and biofilm formation of multi-drug resistant Gram-negative bacteria

Limiting spread of VIM-positive Pseudomonas aeruginosa from colonized sink drains in a tertiary care hospital: A before-and-after study.

In healthcare environments, sinks are being increasingly recognized as reservoirs for multidrug-resistant Gram-negative bacteria. In our hospital, carbapenemase-producing, Verona Integron-encoded Metallo-beta-lactamase (VIM)-positive Pseudomonas aeruginosa (VIM-PA) was detected at low endemicity in patients, and environmental culturing revealed that sink drains were primary reservoirs. Therefore, an intervention was initiated in several wards to install sink drain plugs as physical barriers against splashing to prevent transmission of VIM-PA from drain reservoirs to the surrounding sink environment. To assess the efficacy of the intervention on limiting spread of VIM-PA. Swabs were taken from inner sink environments (i.e. drains), and outer sink environments (i.e. wash basins, faucet aerators, and countertops) twice before and three times after the intervention. Siphon water and drain wells were also sampled before and at the moment of the intervention, respectively. All samples were screened for VIM-PA, and isolates were typed with multiple-locus variable-number tandem repeat analysis (MLVA). There was a significant reduction in VIM-PA positivity in both inner (P-value <0.001) and outer (P-value 0.001) sink environments after the intervention. However, VIM-PA recolonization was observed in the inner sink environments of patient rooms, and also in rooms exclusive to healthcare personnel, over time. Surfaces in the outer sink environment were rarely positive for VIM-PA after the intervention. MLVA revealed three genetic clusters, with one found in all wards and room types during the study period. Drain plugs are a simple and effective infection prevention and control measure to contain spread of VIM-PA from drain reservoirs.