Carbapenem-resistant Enterobacteriaceae Research Articles

A comparative evaluation of five phenotypic methods for identification of carbapenemase-producing Enterobacteriaceae: a modified carbapenemase detection test.

Rapid detection of carbapenemase-producing Enterobacteriaceae (CPE) is urgently needed to prevent their spread in healthcare settings. Here, we have evaluated the performance of the phenotypic methods for detection of carbapenemase production directly from bacterial cultures. A total of 99 clinical and rectal Enterobacteriaceae isolates were included (81 carrying known carbapenemase-encoding genes and 18 without carbapenemase production). All isolates were subjected to the five phenotypic tests including in-house Carba NP (iCarba NP), modified-Carba NP, E-Test MBL, modified Hodge test (MHT), and commercial combination disk test. Test results were read at different time points for iCarba NP and modified-Carba (1 min, 5 min, 15 min, 1 h and 2 h). The sensitivity and specificity of the iCarba NP were 78.87% and 100%, respectively, whereas those of the modified-Carba NP test were 95.06% and 94.44%, respectively. False-negative results were detected in four OXA-48 isolates with the use of modified-Carba NP, whereas one non-carbapenemase isolate had false-positive results. The sensitivity/specificity was 91.30%/100% and 80.25%/83.33% for the E-Test MBL and MHT, respectively. The sensitivity and specificity of the aminophenylboronic acid synergy test were 100% and 97.94%, respectively, whereas those of the dipicolinic acid synergy test were 82.61% and 96.23%, respectively. Rapid, simple, and reliable methods are needed for laboratory detection of CPE isolates to improve the detection and surveillance of these clinically relevant pathogens in an epidemiological context. We conclude that the modified-Carba NP test can be one of the reliable tests for the prediction of carbapenemase-producing bacteria.IMPORTANCEThe emergence of carbapenem resistance among Gram-negative bacteria is a serious global health threat. Here, we investigate the performance of the five phenotypic assays against carbapenemase-producing and carbapenemase-non-producing Enterobacteriaceae. Accurate and rapid detection of CPE isolates is critically required for clinical management and treatment of infections caused by these organisms. Among the five evaluated phenotypic tests, the mCNP test presented the highest sensitivity (95.06%) and, therefore, can be considered the best test to be used as a screening phenotypic methodology.

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

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. 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. 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). 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.

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

BackgroundThis 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). MethodsWe 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. ResultsThe 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. ConclusionsOur 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.

Cas12a/Guide RNA-Based Platform for Rapidly and Accurately Detecting blaKPC Gene in Carbapenem-Resistant Enterobacterales.

Accurate detection and identification of pathogens and their associated resistance mechanisms are essential prerequisites for implementing precision medicine in the management of Carbapenem-resistant Enterobacterales (CRE). Among the various resistance mechanisms, the production of KPC carbapenemase is the most prevalent worldwide. Consequently, this study aims to develop a convenient and precise nucleic acid detection platform specifically for the blaKPC gene. The initial phase of our research methodology involved developing a CRISPR/Cas12a detection framework, which was achieved by designing highly specific single-guide RNAs (sgRNAs) targeting the blaKPC gene. To enhance the sensitivity of this system, we incorporated three distinct amplification techniques-polymerase chain reaction (PCR), loop-mediated isothermal amplification (LAMP), and recombinase polymerase amplification (RPA)-into the CRISPR/Cas12a framework. Subsequently, we conducted a comparative analysis of the sensitivity and specificity of these three amplification methods when used in combination with the CRISPR/Cas12a system. Additionally, we assessed the clinical applicability of the methodologies by evaluating fluorescence readouts from 80 different clinical isolates. Furthermore, we employed lateral flow assay technology to provide a visual representation of the results, facilitating point-of-care testing. Following a comparative analysis of the sensitivity and specificity of the three methods, we identified the RPA-Cas12a approach as the optimal detection technique. Our findings demonstrated that the limit of detection (LoD) of the RPA-Cas12a platform was 1 aM (~1 copy/µL) for plasmid DNA and 5 × 10³ fg/µL for genomic DNA. Furthermore, both the sensitivity and specificity of the platform achieved 100% upon validation with 80 clinical isolates. These findings suggest that the developed RPA-Cas12a platform represents a promising tool for the cost-effective, convenient, and accurate detection of the blaKPC gene.

Mortality factors and antibiotic options in carbapenem-resistant Enterobacterales bloodstream infections: Insights from a high-prevalence setting with co-occurring NDM-1 and OXA-48.

Bloodstream infections (BSI) caused by carbapenem-resistant Enterobacterales (CRE) are associated with a high mortality rate. This study aimed to investigate factors associated with 14-day mortality and identify a potential treatment option. A retrospective cohort study was conducted on patients with CRE-BSI in Thailand from 2015 to 2020. The multivariate Cox proportional-hazards model was employed to identify factors influencing 14-day mortality. Out of 134 diagnosed cases of CRE-BSI, the all-cause 14-day mortality rate was 35.1%. The most prevalent organism isolated was Klebsiella pneumoniae (85.8%), followed by Escherichia coli (11.9%). Among the 60 isolates tested for carbapenemase genes, the majority exhibited co-occurring blaNDM-1 and blaOXA-48 (51.7%), followed by blaOXA-48 (31.7%) and blaNDM-1 (15.0%). In the multivariate analysis, neutropenia (adjusted hazard ratio [aHR] 2.55; 95% confidence interval [95%CI] 1.28-5.06; p = 0.008), sepsis/septic shock (aHR 3.02; 95%CI 1.33-6.86; p = 0.008), and previous metronidazole exposures (aHR 3.58; 95%CI 1.89-6.71; p < 0.001) were identified as independent factors for 14-day mortality. The fosfomycin-based regimen was found to be protective (aHR 0.37; 95%CI 0.15-0.92; p = 0.032). In patients with CRE-BSI, particularly in regions with a high occurrence of co-occurring blaNDM-1 and blaOXA-48, neutropenia, sepsis/septic shock, and previous metronidazole exposures emerged as independent risk factors for mortality. Moreover, the fosfomycin-based regimen showed an improvement in the survival rate.

Modified Carbapenem Inactivation Method and Ethylenediaminetetraacetic Acid (EDTA)-Carbapenem Inactivation Method for Detection of Carbapenemase-Producing Enterobacterales and Pseudomonas aeruginosa.

The rising incidence of carbapenem resistance in Enterobacterales and Pseudomonas aeruginosa is a concern. Since carbapenemase production is the primary resistance mechanism, detecting and identifying the genes responsible for it is crucial to effectively monitor its spread. This study aims to detect positivity for the modified carbapenem inactivation method (mCIM) and ethylenediaminetetraacetic acid (EDTA)-carbapenem inactivation method (eCIM) for the detection of carbapenemase-producing Enterobacterales and Pseudomonas aeruginosa. Methods:A cross-sectional study was carried out at a tertiary care hospital, including 250 clinical isolates of Enterobacterales and Pseudomonas aeruginosa. These isolates exhibited resistance to at least one of the carbapenems as determined by the VITEK AST 2 System (bioMérieux, USA). The isolates were subjected to mCIM testing, and those that tested positive were further tested using eCIM. The results were interpreted in accordance with the guidelines provided by the Clinical and Laboratory Standards Institute (CLSI) 2023. Out of the total 250 carbapenem-resistant Enterobacterales and Pseudomonas aeruginosa isolates, 151 (60.4%) were Klebsiella pneumonia, 44 (17.6%) were Escherichia coli, 10 (4.0%) were Enterobacter cloacae, 6 (2.4%) were Providencia spp., 4 (1.6%) were Serratia marcescens, 4 (1.6%) were Proteus mirabilis and 31 (12.4%) were Pseudomonas aeruginosa. Positivity for the mCIM was observed in 96% (240 out of 250) of the isolates. Of the mCIM-positive isolates, 234 (97.5%) also tested positive for eCIM, indicating metallo-β-Lactamase (MLB) production. A statistically significant association was found between both mCIM and eCIM positivity and the degree of resistance to carbapenem (p<0.05). Conclusion: This study shows that the inexpensive method, a combination of mCIM and eCIM assists in differentiating between serine carbapenemase producers and MLB producers, thereby guiding the selection of appropriate therapy and useful in infection control in resource-limited settings.

Ceftazidime-Avibactam in the Treatment of Patients with Bacteremia or Nosocomial Pneumonia: A Systematic Review and Meta-analysis.

Ceftazidime-avibactam (CAZ-AVI) is a combination of the third-generation cephalosporin ceftazidime and the novel, non-β-lactam β-lactamase inhibitor avibactam that is approved for the treatment of pediatric (≥ 3months) and adult patients with complicated infections including hospital-acquired and ventilator-associated pneumonia (HAP/VAP), and bacteremia. This systematic literature review and meta-analysis (PROSPERO registration: CRD42022362856) aimed to provide a quantitative and qualitative synthesis to evaluate the effectiveness of CAZ-AVI in treating adult patients with bacteremia or nosocomial pneumonia caused by carbapenem-resistant Enterobacterales (non metallo-β-lactamase-producing strains) and multi-drug resistant (MDR) Pseudomonas aeruginosa infections. The databases included in the search, until November 7, 2022, were Embase and PubMed. A total of 24 studies (retrospective: 22, prospective: 2) with separate outcomes for patients with bacteremia or pneumonia were included. The outcomes assessed were all-cause mortality, clinical cure, and microbiological cure. Qualitative (24 studies) and quantitative (8/24 studies) syntheses were performed. The quality of the studies was assessed using the MINORS checklist and the overall risk of bias was moderate to high. In studies included in the meta-analysis, lower all-cause mortality for patients with bacteremia (OR = 0.30, 95% CI 0.19-0.46) and improved rates of clinical cure for patients with bacteremia (OR = 4.90, 95% CI 2.60-9.23) and nosocomial pneumonia (OR = 3.20, 95% CI 1.55-6.60) was observed in the CAZ-AVI group compared with the comparator group. Data provided here may be considered while using CAZ-AVI for the treatment of patients with difficult-to-treat infections. PROSPERO CRD42022362856.

Dynamic evolution of ceftazidime-avibactam resistance from a single patient through the IncX3_NDM-5 plasmid transfer and blaKPC mutation

The rapid dissemination of carbapenem-resistant Enterobacterales (CRE) especially carbapenem-resistant Klebsiella pneumoniae (CRKP) poses a great threat to global public health. Ceftazidime-avibactam, a novel β-lactam/β-lactamase inhibitor combination, has been widely used due to its excellent antibacterial activity against KPC-producing K. pneumoniae. However, several resistance mechanisms have been reported since its use. Here, we conducted a series of in vitro experiments to reveal and demonstrate the dynamic evolution of ceftazidime-avibactam resistance including interspecies IncX3_NDM-5 plasmid transfer between Enterobacter cloacae and K. pneumoniae and blaKPC mutation from blaKPC-2 to blaKPC-33. Through the analysis of conjugation frequency and fitness cost, the IncX3_NDM-5 plasmid in this study showed strong transmissibility and stability in E. coli EC600 and clinical strain K. pneumoniae 5298 as recipient strain. With increasing ceftazidime-avibactam concentration, the conjugation frequency remained at 10-3-10-5, while the mutation frequency of K. pneumoniae 5298 was 10-6-10-8 at the same concentration. Further plasmid analysis (the IncX3_NDM plasmid from this study and other 658 plasmids from the NCBI database) revealed the diverse origin and genetic structure of blaNDM-5 carrying plasmids. E. coli (42.9%), China (43.9%), IncX3 (66.6%) are the most common strains, regions, and Inc types respectively. By analysing of genetic environment detected in IncX3 plasmids, the dominant structures (168/258, 65.1%) were identified: ISKox3-IS26-blaNDM-5-IS5-ISAba125-Tn3000-Tn3. In additon, several structural variations were found in the core gene structure. In conclusion, the high fitness and transmissibility of the IncX3_NDM-5 plasmids were noteworthy. More importantly, the diverse ceftazidime-avibactam resistance mechanisms including blaNDM-5 tranfer and blaKPC-2 mutation highlighted the importance of the continuous monitoring of antimicrobial susceptibility and carbapenemases subtype during ceftazidime-avibactam treatment.

Carbapenem-resistant Enterobacterales in wastewater resources and healthy carriers: A survey in Iran

ABSTRACT The carbapenem-resistant Enterobacterales (CRE) pose a pressing public health concern. Here, we investigated the frequency of CRE bacteria, carbapenemase-encoding genes, and the molecular epidemiology of carbapenemase-resistant Escherichia coli in wastewater resources and healthy carriers in Iran. Out of 617 Enterobacterales bacteria, 24% were carbapenem-resistant. The prevalence of CRE bacteria in livestock and poultry wastewater at 34% and hospital wastewater at 33% was significantly higher (P ≤ 0.05) than those in healthy carriers and municipal wastewater at 22 and 17%, respectively. The overall colonization rate of CRE in healthy individuals was 22%. Regarding individual Enterobacterales species, the following percentages of isolates were found to be CRE: E. coli (18%), Citrobacter spp. (24%), Klebsiella pneumoniae (28%), Proteus spp. (40%), Enterobacter spp. (25%), Yersinia spp. (17%), Hafnia spp. (31%), Providencia spp. (21%), and Serratia spp. (36%). The blaOXA-48 gene was detected in 97% of CRE isolates, while the blaNDM and blaVIM genes were detected in 24 and 3% of isolates, respectively. The B2 phylogroup was the most prominent group identified in carbapenem-resistant E. coli isolates, accounting for 80% of isolates. High prevalence of CRE with transmissible carbapenemase genes among healthy people and wastewater in Iran underscores the need for assertive measures to prevent further dissemination.

Development and implementation of a nationwide multidrug-resistant organism tracking and alert system for Veterans Affairs medical centers.

Develop and implement a system in the Veterans Health Administration (VA) to alert local medical center personnel in real time when an acute- or long-term care patient/resident is admitted to their facility with a history of colonization or infection with a multidrug-resistant organism (MDRO) previously identified at any VA facility across the nation. An algorithm was developed to extract clinical microbiology and local facility census data from the VA Corporate Data Warehouse initially targeting carbapenem-resistant Enterobacterales (CRE) and methicillin-resistant Staphylococcus aureus (MRSA). The algorithm was validated with chart review of CRE cases from 2010-2018, trialed and refined in 24 VA healthcare systems over two years, expanded to other MDROs and implemented nationwide on 4/2022 as "VA Bug Alert" (VABA). Use through 8/2023 was assessed. VABA performed well for CRE with recall of 96.3%, precision of 99.8%, and F1 score of 98.0%. At the 24 trial sites, feedback was recorded for 1,011 admissions with a history of CRE (130), MRSA (814), or both (67). Among Infection Preventionists and MDRO Prevention Coordinators, 338 (33%) reported being previously unaware of the information, and of these, 271 (80%) reported they would not have otherwise known this information. By fourteen months after nationwide implementation, 113/130 (87%) VA healthcare systems had at least one VABA subscriber. A national system for alerting facilities in real-time of patients admitted with an MDRO history was successfully developed and implemented in VA. Next steps include understanding facilitators and barriers to use and coordination with non-VA facilities nationwide.

#237 Carbapenemase producing Enterobacteriaceae rectal colonization in dialysis patients: a large monocentric retrospective analysis

Abstract Background and Aims Carbapenemase-producing Enterobacteriaceae (CPE) represent a growing global public health concern due to their increasing prevalence and resistance to carbapenems. Dialysis patients are particularly vulnerable to infections that represent the second cause of death in dialysis’ cohorts. Considering the immunosuppression, dialysis patients constitute a highly immunocompromised population and represent a unique setting similar to intensive care, hematology, or transplant recipients. Several studies have already been conducted in these populations, demonstrating the high infectious risk, increased mortality and morbidity burdened by CPE. Our study aims to analyze a large dialysis population and assess if there is an increased risk of mortality, sepsis, and/or hospitalizations within this cohort. Considering the frequency of evaluations for these patients, we have also decided to evaluate the treatments to which these patients are subjected, their antibiotic exposure, the number of hospitalizations and specific infections, while seeking predisposing or protective factors within this specific patient group. Method Our work is a retrospective, single-center cohort study that analyzes patients on HD and PD who were followed by the Nephrology and Dialysis Department of the Ravenna Hospital from 01/01/2015 to 31/12/2021 (674 patients). We analyzed demographic data (Date of birth, gender, blood type), comorbidities (Diabetes, hypertension), dialysis data (type of dialysis technique, dialytic age), blood tests (hemoglobin, white blood cells, platelets, albumin, ALT, lipase, C-reactive protein), serology and markers (HBV, HCV, HIV, EBV, CMV, Quantiferon, Helicobacter pylori colonization, SARS-COV2), therapies (antihypertensives, proton pump inhibitors, calcium-based phosphorus binders, non-calcium-based phosphorus binders, potassium binders, iron therapy, erythropoietin-stimulating agents), antibiotics (cephalosporins, penicillins, fluoroquinolones, macrolides, aminoglycosides, carbapenems, tetracyclines, vancomycin, linezolid, cefepime/avibactam), hospitalizations, colonization or lack thereof by CPE bacteria, infections (urinary tract infections and pneumonias) or sepsis and responsible microorganisms, survival. We exclude patients who had stomas or those who received chemotherapy or immunotherapy for a period longer than 3 months. Results Overall we considered for our analysis 528 patients and 51 presented CPE positive rectal swab during the period of observation. In our population CPE colonized patients present a higher risk to develop pneumonias (HR 2.048, p = 0.003). 178 patients presented 497 episodes of sepsis, 451 episodes in the non CPE colonized and 46 episodes in the CPE colonized (HR 2.057, p-value 0.001). Considering CPE-induced sepsis, a substantial difference was observed between the two groups. The colonized patient group had 7 (15.2%) cases of CPE sepsis, while the non-colonized group did not have any (0%). The result shows statistical significance, although the limited number of infections does not allow for definitive conclusions on this matter. Concerning the re-hospitalization rate we evidenced a significative difference between the two groups, displaying an involvement of CPE colonization (HR1.367, p-value = 0.038). Conclusion Our study highlighted that a colonization by CPE lead to a greater number of episodes of all bacterial sepsis, of CPE sepsis, of pneumonias and of re-hospitalizations. Colonization by CPE conferm an increased risk of CPE sepsis, and therefore, future research should focus on determining whether it is worthwhile to treat hyperpyrexia in colonized CPE dialysis patients as if it were CPE-related sepsis. This is a significant question that can guide our clinical practice.

The effect of silver nanoparticles and preparations of various pharmacological groups on the bactericidal activity of &lt;i&gt;Staphylococcus aureus&lt;/i&gt;

Multidrug-resistant bacteria have become one of the most serious threats to public health worldwide. The abuse of antibiotics has contributed to the emergence and transmission of resistance mechanisms among bacteria, jeopardizing the therapeutic potential of antibiotics. Uncontrolled use of drugs leads to the formation of antibiotic resistance due to mutations in chromosomal DNA, as well as the production of plasmids, integrons from other bacteria during horizontal gene transfer. In 2010, the BRICS countries (Brazil, Russia, India, China and South Africa) accounted for 76% of antibiotic consumption, with India consuming 12.9 billion units and China – 10 billion units. As of 2017, carbapenem-resistant Acetobacter baumannii and Enterobacteriaceae resulted in approximately US$ 281 million in healthcare costs in the United States. According to numerous reports from the Center for Disease Control and Prevention, approximately 2.3 million episodes of multidrug-resistant microbial diseases resulting in 25,000 deaths are recorded annually in the United States alone. In this regard, the world community of scientists has intensified the study of the combined use of various antibacterial drugs to achieve maximum bactericidal activity. Studies have been conducted to determine the synergistic effect when using combinations of drugs of various pharmacological groups and silver nanoparticles. A significant increase in bactericidal activity by 53.87 times (from 2.528 to 0.0098 mcg/ml) was found with the combined use of AgNPs and DSMO against the reference strain Staphylococcus aureus ATCC 25953. While the cultivation of St. aureus isolate with DSMO and silver nanoparticles revealed a sensitivity increase of 128.2 times (from 5.056 to 0.039 mcg/ml).

Global epidemiology and antimicrobial resistance of Enterobacterales harbouring genes encoding OXA-48-like carbapenemases: insights from the results of the Antimicrobial Testing Leadership and Surveillance (ATLAS) programme 2018-2021.

The recent emergence of carbapenem-resistant Enterobacterales poses a major and escalating threat to global public health. This study aimed to analyse the global distribution and antimicrobial resistance of Enterobacterales harbouring variant OXA-48-like carbapenemase-related genes. Enterobacterales isolates were collected from the Antimicrobial Testing Leadership and Surveillance (ATLAS) programme during 2018-2021. Comprehensive antimicrobial susceptibility testing and β-lactamase gene detection were also conducted, along with statistical analysis of the collected data. Among the 72 244 isolates, 1934 Enterobacterales isolates were identified to harbour blaOXA-48-like genes, predominantly Klebsiella spp. (86.9%). High rates of multidrug resistance were observed, with only ceftazidime/avibactam and tigecycline showing favourable susceptibility. A discrepancy between the genotype and phenotype of carbapenem resistance was evident: 16.8% (233 out of 1384) of the Enterobacterales isolates with blaOXA-48-like genes exhibited susceptibility to meropenem. Specifically, 37.4% (64/95) of Escherichia coli strains with blaOXA-48-like genes displayed meropenem susceptibility, while the corresponding percentages for Klebsiella pneumoniae and Enterobacter cloacae complex were 25.2% (160/1184) and 0% (0/36), respectively (P < 0.05). Geographical analysis revealed that the highest prevalence of blaOXA-48-like genes occurred in Asia, the Middle East and Eastern Europe. The proportion of K. pneumoniae isolates harbouring blaOXA-232 increased from 23.9% in 2018 to 56.0% in 2021. By contrast, the proportion of blaOXA-48 decreased among K. pneumoniae isolates during 2018-2021. This study underscores the widespread and increasing prevalence of blaOXA-48-like genes in Enterobacterales and emphasizes the need for enhanced surveillance, improved diagnostic methods and tailored antibiotic stewardship to combat the spread of these resistant pathogens.

Evaluation of a simple method for testing aztreonam and ceftazidime-avibactam synergy in New Delhi metallo-beta-lactamase producing Enterobacterales.

NDM-producing carbapenem-resistant bacterial infections became a challenge for clinicians. Combination therapy of aztreonam and ceftazidime-avibactam is a prudent choice for these infections. However, there is still no recommendation of a practically feasible method for testing aztreonam and ceftazidime-avibactam synergy. We proposed a simple method for testing aztreonam and ceftazidime-avibactam synergy and compared it with reference broth micro-dilution and other methods. Carbapenem-resistant Enterobacterales clinical isolates were screened for the presence of the NDM gene by the Carba R test. NDM harbouring isolates were tested for aztreonam and ceftazidime-avibactam synergy by broth microdilution (reference method), E strip-disc diffusion, double disc diffusion, and disc replacement methods. In the newly proposed method, the MHA medium was supplemented with ceftazidime-avibactam (corresponding to an aztreonam concentration of 4μg/ml). The MHA medium was then inoculated with the standard inoculum (0.5 McFarland) of the test organism. An AZT disc (30 μg) was placed on the supplemented MHA medium, and the medium was incubated overnight at 37°C. Aztreonam zone diameter on the supplemented MHA medium (in the presence of ceftazidime-avibactam) was compared with that from a standard disc diffusion plate (without ceftazidime-avibactam), performed in parallel. Interpretation of synergy was based on the restoration of aztreonam zone diameter (in the presence of ceftazidime-avibactam) crossing the CLSI susceptibility breakpoint, i.e., ≥ 21 mm. Of 37 carbapenem-resistant NDM-producing isolates, 35 (94.6%) were resistant to aztreonam and tested synergy positive by the proposed method. Its sensitivity and specificity were 97.14% and 100%, respectively. Cohen's kappa value showed substantial agreement of the reference method with the proposed method (κ = 0.78) but no other methods. The proposed method is simple, easily interpretable, and showed excellent sensitivity, specificity, and agreement with the reference method. Therefore, the new method is feasible and reliable for testing aztreonam synergy with avibactam in NDM-producing Enterobacterales.

Activity of propyl-propane-thiosulfinate and propyl-propane-thiosulfonate against carbapenem-resistant Gram-negative bacteria.

Organosulfur compounds derived from plants of the Allium genus, such as propyl-propane-thiosulfinate (PTS) and propyl-propane-thiosulfonate (PTSO), have been proposed as an alternative in antibiotic resistance. The aim of this study was to compare the activity of these substances with other antibiotics against clinical isolates of carbapenem-resistant (CAR-R) and carbapenem-susceptible (CAR-S) Gram-negative bacteria. A total of 126 clinical isolates of CAR-R and 155 CAR-S bacteria were selected, including Enterobacterales, A. baumannii and P. aeruginosa. The antibiotic susceptibility of all isolates was assessed using the microdilution and Kirby-Bauer methods for PTS, PTSO, amoxicillin/clavulanate, piperacillin/tazobactam, cefotaxime, ceftazidime, cefepime, imipenem, ciprofloxacin, and amikacin. Both PTS and PTSO demonstrated in vitro bactericidal activity against CAR-R Enterobacteriaceae and A. baumannii, with no significant difference in activity compared to their response against CAR-S isolates. However, both compounds were less active against P. aeruginosa than against any of the other bacteria, regardless of their resistance to carbapenems. In all cases, the minimum inhibitory concentration values of PTSO were significantly lower than those of PTS. These findings offer valuable information about the potential antibacterial use of these substances, particularly against infections that currently have limited therapeutic options.

Ironing out Persisters? Revisiting the Iron Chelation Strategy to Target Planktonic Bacterial Persisters Harboured in Carbapenem-Resistant Escherichia coli.

Antibiotic resistance is a global health crisis. Notably, carbapenem-resistant Enterobacterales (CRE) pose a significant clinical challenge due to the limited effective treatment options. This problem is exacerbated by persisters that develop upon antibiotic exposure. Bacteria persisters can tolerate high antibiotic doses and can cause recalcitrant infections, potentially developing further antibiotic resistance. Iron is a critical micronutrient for survival. We aimed to evaluate the utility of iron chelators, alone and in combination with antibiotics, in managing persisters. We hypothesized that iron chelators eradicate CRE persisters in vitro, when administered in combination with antibiotics. Our screening revealed three clinical isolates with bacteria persisters that resuscitated upon antibiotic removal. These isolates were treated with both meropenem and an iron chelator (deferoxamine mesylate, deferiprone or dexrazoxane) over 24 h. Against our hypothesis, bacteria persisters survived and resuscitated upon withdrawing both the antibiotic and iron chelator. Pursuing our aim, we next hypothesized that iron chelation is feasible as a post-antibiotic treatment in managing and suppressing persisters' resuscitation. We exposed bacteria persisters to an iron chelator without antibiotics. Flow cytometric assessments revealed that iron chelators are inconsistent in suppressing persister resuscitation. Collectively, these results suggest that the iron chelation strategy may not be useful as an antibiotic adjunct to target planktonic bacteria persisters.

Risk factors for in-hospital acquisition of carbapenem-resistant Enterobacterales in patients with severe burns

Although patients with severe burns are prone to severe infections with antibiotic-resistant bacteria and inevitably have some risk factors for carbapenem-resistant Enterobacterales (CRE) acquisition, risk factors for CRE infection or colonization in these patients have not been investigated. To identify the independent risk factors for CRE acquisition in patients with severe burns. Patients admitted to the burn intensive care unit (BICU) for acute burn care were categorized based on culture results during BICU care into the CRE group and non-CRE group, which included the carbapenem-susceptible Enterobacterales (CSE) and control groups. Clinical and microbiological factors were compared between the CRE and non-CRE groups, and between the CRE and CSE groups to identify independent risk factors for in-hospital CRE acquisition. Among the included 489 patients, 101 (20.7%) and 388 (79.3%) patients were classified in the CRE and non-CRE groups, respectively. The non-CRE group included 91 (18.6%) and 297 (60.7%) patients in the CSE and control groups, respectively. In multivariate analysis between the CRE and non-CRE groups, exposure to other CRE-acquired patients (P= 0.018), abbreviated burn severity index score ≥9 (P= 0.012), and mechanical ventilation (P < 0.001) were associated with CRE acquisition. In multivariate analysis between the CRE and CSE groups, exposure to other CRE-acquired patients was associated with CRE acquisition (P= 0.048). Considering the limitation of controlling the burn severity in hospitalized patients, enhanced infection control measures for preventing in-hospital CRE transmission among patients with severe burns should be emphasized.

A five-year retrospective antibiogram review in the paediatric burns unit at a tertiary South African Hospital

BackgroundThere is a global concern regarding increasing colonization and/or infection of paediatric burns patients with multi-drug-resistant organisms, especially in Africa. Surveillance of antimicrobial resistant patterns enables clinicians to opt for the most appropriate empiric antimicrobial agent according to the unit’s susceptibility profile. ObjectiveThis study sought to compile unit-specific antibiograms comparing the organism prevalence and their antimicrobial resistance trends in the paediatric burns unit, Chris Hani Baragwanath Academic Hospital (CHBAH) from January 2015 to December 2019. MethodsA retrospective, laboratory-based review was conducted of bacterial and fungal isolates including antimicrobial susceptibility testing (AST) data from blood cultures and tissue samples. Enterobacterales, non-fermenters, gram-positive organisms, and yeasts were analysed as a percentage of total positive blood cultures comparing 2015–2017 to 2018–2019 data. Analysis of antimicrobial resistance trends was performed for extended-spectrum beta-lactamases (ESBLs), carbapenem-resistant Enterobacterales (CRE), extensively-drug-resistant (XDR) Acinetobacter baumannii, and meticillin-resistant Staphylococcus aureus (MRSA) for the years 2015, 2017, and 2019. ResultsA total of 1243 blood culture isolates were studied with A.baumannii (208/1243;17 %), coagulase negative staphylococci (141/1243; 11 %) and Candida auris (53/1243; 4 %) representing the predominant gram-negative bacteria, gram-positive bacteria, and yeast, respectively. A significant decrease in ESBLs (p = 0.00) was noted with a rise in CREs (p = 0.00) during the study period. Majority of the A. baumannii were XDR (≥77 %) despite an appreciable decrease. A notable decrease in MRSA (p = 0.00) isolates was observed. Pseudomonas aeruginosa (72/275; 26 %) was the leading organism recovered from a total of 275 tissue isolates. ConclusionThe results of this study encourage review of the current antimicrobial choices in the unit for empiric antibacterial cover. Due to the low numbers of yeasts, we cannot comment on the institution of early antifungal strategies. We support reinforced infection prevention and control practices as well as antimicrobial stewardship initiatives.

Rapid colorimetric polymyxin B microelution directly from positive blood bottles: because patients with serious infections should not have to wait for results of culture-based methodologies.

To evaluate the performance of the rapid colorimetric polymyxin B microelution (RCPEm) in determining polymyxin B resistance directly from Enterobacterales-positive blood cultures. A set volume of positive blood culture bottles (diluted 1:10) was inoculated into a glucose-broth-phenol red solution (NP solution), where a polymyxin B disk was previously eluted (final concentration of 3µg/mL). Test was read each 1h for up to 4h. Color change from red/orange to yellow indicated resistant isolates. Results were compared to the reference method, broth microdilution (BMD), performed from colonies grown on solid media from the same blood culture bottle. One hundred fifty-two Enterobacterales-positive blood cultures were evaluated, 22.4% (34/152) of them resistant to polymyxin B (including 6.6% with borderline MICs). When performing directly from positive blood cultures (RCPEm-BC), specificity and sensitivity were 99.1% and 94.1%, respectively. Of note, 79.4% (27/34) of truly resistant isolates required 3h of incubation, compared to the 18 ± 2h incubation that microtiter plates of BMD demand before reading can be performed. RCPEm directly from blood cultures has great potential to be part of the routine of clinical microbiology laboratories to establish polymyxin B susceptibility, impacting outcome of patients with bloodstream infections caused by carbapenem-resistant Enterobacterales.

Surveillance of Enterobacteriaceae from Diabetic Foot Infections in a Tunisian Hospital: Detection of E. coli-ST131-blaCTX-M-15 and K. pneumoniae-ST1-blaNDM-1 Strains.

The study determined the prevalence, antimicrobial resistant (AMR) determinants, and genetic characteristics of Escherichia coli and Klebsiella pneumoniae isolates from patients with diabetic foot infection (DFI) in a Tunisian hospital. A total of 26 Escherichia spp. and Klebsiella spp. isolates were recovered and identified by MALDI-TOF-MS. Antimicrobial susceptibility testing, the detection of AMR determinants and Shiga-like toxin genes, phylogenetic grouping, and molecular typing were performed. Twelve E. coli, 10 K. pneumoniae, 3 K. oxytoca, and 1 E. hermanii were isolated. A multidrug-resistant phenotype was detected in 65.4% of the isolates. About 30.8% of isolates were extended-spectrum β-lactamase (ESBL) producers and mainly carried blaCTX-M-15 and blaCTX-M-14 genes. One blaNDM-1-producing K. pneumoniae-ST1 strain was identified. Class 1 integrons were detected in 11 isolates and 5 gene cassette arrangements were noted: dfrA1+aadA1 (n = 1), dfrA12+aadA2 (n = 3), and dfrA17+aadA5 (n = 1). Other non-β-lactam resistance genes detected were as follows (number of isolates): aac(3')-II (3), aac(6')-Ib-cr(8), qnrB (2), qnrS (4), cmlA (2), floR (4), sul1 (11), sul2 (11), and sul3 (2). The phylogroup B1 was the most frequent (41.7%) among E. coli, and two ESBL-producing isolates corresponded to the ST131-B2 lineage. The ESBL- and carbapenemase-producing Enterobacteriaceae in DFIs are described for the first time in Tunisia.