Carbapenem Resistance Research Articles

First report of carbapenems encoding multidrug-resistant gram-negative bacteria from a pediatric hospital in Gaza Strip, Palestine

BackgroundThe worldwide prevalence of multi-drug resistance (MDR) in Gram-negative bacteria (GNB), particularly related to extended-spectrum beta-lactamases (ESBLs) and carbapenemases, poses significant global public health and clinical challenges.ObjectivesTo characterize ESBL-producing Gram-negative bacilli, within a pediatric hospital in Gaza using whole genome sequencing (WGS).MethodsA total of 158 clinical isolates of Gram-negative bacilli were collected from Al-Nasser Pediatric Hospital. These isolates were tested for ESBL production using the double disk synergy test. The antibiotic susceptibility profile was determined using the Kirby Bauer method following the Clinical and Laboratory Standard Institute guidelines. Selected 15 phenotypically MDR isolates were whole-genome sequenced and characterized for their genome-based species identity and antibiotic resistance gene profile.ResultsOf the 158 isolates, 93 (58.9%) were positive for ESBL production. The frequency of Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa, Acinetobacter baumannii, Proteus mirabilis, and Serratia marcescens was 50%, 22.7%, 22.7%, 1.8%, 1.2%, and 1.2% respectively. The prevalence of ESBL among urine, pus, blood, and sputum was 64%, 44%, 23%, and 63.6%, respectively. Chloramphenicol, Imipenem, and Meropenem were the most effective antibiotics against ESBL producers.In sequenced isolates, an average of six anti-microbial resistance (AMR) genes were noted per isolate, where one of them carried up to 13 antibiotic resistance genes. Carbapenem resistance genes such as blaKPC-2(6.6%), blaPDC-36/12 (6.6%), and blaPOM-1 (6.6%) were detected. All the sequenced E. coli isolates (n = 8) showed multiple resistance genes, mainly against β-lactamase (25.0%), aminoglycosides (37.5%), sulfonamides (37.5%), and genes conferring resistance to tetracyclines (25.0).ConclusionOur results showed a high prevalence of ESBL-producing GNB isolated from a pediatric hospital in the Gaza Strip. Various antibiotic resistance genes were identified, including those encoding ESBL and carbapenems. The results highlight the significant challenge posed by MDR in GNB and emphasize the need for effective antibiotic strategies. Given the high endemicity observed in various studies from Palestine, it is important to conduct clinical and molecular epidemiology research to identify risk factors, transmission patterns, and clinical outcomes associated with GNB strains that carry ESBL and carbapenem resistance genes.

The context of blaOXA-23 gene in Iraqi carbapenem-resistant Acinetobacter baumannii isolates belonging to global clone 1 and global clone 2.

Of the genes conferring resistance to carbapenems in Acinetobacter baumannii, the blaOXA-23 gene is the most widely found across the world. The gene carrying blaOXA-23 transposons in A. baumannii isolates of global clones GC1 and GC2 is found worldwide. Here, we examined whether transposons play a role in the dissemination of the blaOXA-23 in globally distributed clones, GC1 and GC2 A. baumannii isolates from Iraq. The 119 non-repetitive A. baumannii isolates including 94 recovered from clinical specimens and 25 isolates from hospital environment between September 2021 and April 2022 from different medical centers located at various regions in Baghdad, Iraq. The global clones (GC) and the genes encoding carbapenem resistance, including blaOXA-23, blaOXA-24, and blaOXA-58 were identified using multiplex PCR assays. Antibiotic susceptibility testing was performed by the Kirby-Bauer disk diffusion susceptibility method. The transposons carrying blaOXA-23 were examined using PCR mapping. In cases when carbapenem susceptible A. baumannii isolates were found, they were subjected to E test, full length sequencing of blaOXA-Ab (blaOXA-51-like) and Institut Pasteur multi-locus sequence typing scheme. All but two isolates (92 clinical and 25 environmental) were identified carbapenem-resistant A. baumannii (CRAB). Of 117 CRAB isolates, 20 belong to GC1, 19 contained blaOXA-23; of them, 17 isolates harbored the blaOXA-23 located on Tn2006. Among the 46 CRAB belonging to GC2, 39 contained blaOXA-23; of them, 34 carried the blaOXA-23 located on Tn2006. The remaining GC1 and GC2 isolates, one GC1 as well as one GC2 isolate, were susceptible to imipenem, doripenem, and meropenem and considered carbapenem-susceptible A. baumannii (CSAB). Full-length sequencing of the blaOXA-Ab and MLST for the two CSAB isolates belonging to GC1 and GC2 confirmed that the GC1 isolate belongs to ST 623 and contained an allele that encodes an blaOXA-69 variant of the blaOXA-Ab while the GC2 belong to ST2 and carried an blaOXA-66 variant. This study provides evidence for the dissemination of blaOXA-23 on the Tn2006 in CRAB isolates in Baghdad, Iraq. It appears that this transposon is widespread in GC1 and 2 isolates as in the other parts of the world. Interestingly, one GC1 and one GC2 isolate from Iraq were found to be susceptible to carbapenem while the isolates belonging to GC1 and GC2 have so far rarely been found to be susceptible to carbapenem globally.

The OprF porin as a potential target for the restoration of carbapenem susceptibility in Pseudomonas aeruginosa expressing acquired carbapenemases.

Carbapenem resistance in Pseudomonas aeruginosa is primarily due to the acquisition of carbapenemases and is often associated with a diminution of the membrane permeability. The outer membrane protein, OprD, is a well-known route, by which carbapenems, predominantly imipenem, can enter the cell, and its loss has been associated with reduced susceptibility to imipenem. In this study, we investigated the antibiotic susceptibility patterns of isogenic P. aeruginosa mutants containing various acquired β-lactamases, including carbapenemases, in a porin-depleted background. We identified that the deletion of oprF was associated with some recovery of susceptibility to carbapenems.

Association between the presence of CRISPR-Cas system genes and antibiotic resistance in Klebsiella pneumoniae isolated from patients admitted in Ahvaz teaching hospitals

BackgroundThis study aims to investigate the frequency of cas1 and cas3 and CRISPR1,2,3 genes in Klebsiella pneumoniae isolates, as well as their connection with antibiotic resistance.Materials and methods106 K. pneumoniae isolates were identified by biochemical assays and PCR. The susceptibility to antibiotics was determined by Kirby–Bauer disk diffusion method. Screening of ESBLs was undertaken by using double disk diffusion and standard disk diffusion methods. The E-test and mCIM techniques was used to confirm the disc diffusion-based carbapenem resistance profiles. CRISPR-Cas system genes were identified using PCR.ResultsESBL production was found in 19% of isolates. Carbapenemase production was found in 46% of the isolates. Furthermore, the bacteria were classified as multidrug (76%), extensively drug-resistant (4%), or pan-drug-resistant (2%). When CRISPR/Cas systems were present, antibiotic resistance was lower; conversely, when they were absent, resistance was higher.ConclusionsIf the CRISPR/Cas modules aren’t present, the bacteria can still acquire foreign DNA, including antibiotic resistance genes. K. pneumoniae isolates with a CRISPR-Cas system were less likely to carry antibiotic-resistance genes than those lacking this defense system.

Drug-resistant Pantoea agglomerans Causing Bacteremia at a Tertiary Care Hospital in Kolkata, India: First Report of Carbapenem Resistance Mediated by OXA-181.

The spread of antibiotic resistance (ABR) in uncommon human pathogens endangers global public health, escalating morbidity, death, and healthcare expenditures. Pantoea agglomerans, a member of the Erwiniaceae family that rarely infects humans, is emerging as a drug-resistant nosocomial pathogen. Seven P. agglomerans isolates were recovered from bacteremia patients at a tertiary care hospital in Kolkata, West Bengal, between March 2022 and October 2022. The isolates were evaluated for phenotypic resistance, β-lactamase and plasmid-mediated quinolone resistance (PMQR) genes, plasmid profiling, and clonality assessment. All isolates were resistant to fluoroquinolones and third-generation cephalosporins, with four resistant to carbapenems. The following β-lactamases and PMQR genes were identified: blaOXA-1 (n = 1), blaTEM (n = 1), blaCTX-M-1 (n = 2), blaNDM (n = 5), blaOXA-181 (n = 1), qnrB (n = 2), and qnrS (n = 4). Six isolates carried up to seven plasmids ranging in size from 2kb to > 212kb. IncFI, FII, HI, and X3 plasmid types were detected in three isolates, while the rest remained untypable. Four different genetic patterns were noted. Four isolates were clonally related, with three being clonal. The swap of environmental isolates to human pathogens exacerbates the ABR dilemma, periling patient care and outcomes. This is the first report in India of a carbapenem-resistant P. agglomerans blood isolate carrying blaOXA-181. In-depth genomic research of drug-resistant microbes adapted to the environment-human interfaces might underpin the source-route-containment of ABR.

Genomic epidemiology of carbapenem-resistant Enterobacterales at a New York City hospital over a 10-year period reveals complex plasmid-clone dynamics and evidence for frequent horizontal transfer of bla KPC.

Transmission of carbapenem-resistant Enterobacterales (CRE) in hospitals has been shown to occur through complex, multifarious networks driven by both clonal spread and horizontal transfer mediated by plasmids and other mobile genetic elements. We performed nanopore long-read sequencing on CRE isolates from a large urban hospital system to determine the overall contribution of plasmids to CRE transmission and identify specific plasmids implicated in the spread of bla KPC (the Klebsiella pneumoniae carbapenemase [KPC] gene). Six hundred and five CRE isolates collected between 2009 and 2018 first underwent Illumina sequencing for genome-wide genotyping; 435 bla KPC-positive isolates were then successfully nanopore sequenced to generate hybrid assemblies including circularized bla KPC-harboring plasmids. Phylogenetic analysis and Mash clustering were used to define putative clonal and plasmid transmission clusters, respectively. Overall, CRE isolates belonged to 96 multilocus sequence types (STs) encoding bla KPC on 447 plasmids which formed 54 plasmid clusters. We found evidence for clonal transmission in 66% of CRE isolates, over half of which belonged to four clades comprising K. pneumoniae ST258. Plasmid-mediated acquisition of bla KPC occurred in 23%-27% of isolates. While most plasmid clusters were small, several plasmids were identified in multiple different species and STs, including a highly promiscuous IncN plasmid and an IncF plasmid putatively spreading bla KPC from ST258 to other clones. Overall, this points to both the continued dominance of K. pneumoniae ST258 and the dissemination of bla KPC across clones and species by diverse plasmid backbones. These findings support integrating long-read sequencing into genomic surveillance approaches to detect the hitherto silent spread of carbapenem resistance driven by mobile plasmids.

A-288 Active Surveillance Cultures: Frequency of Microorganisms and Phenotypic Resistance Profile From Brazilian Public Hospitals After COVID-19 Pandemic Peak

Abstract Background Surveillance cultures are routinely used by public health authorities to screen for multidrug-resistant bacteria. Recently, the overuse of antibiotics intensified by the COVID-19 pandemic worldwide brought concerns about potential impacts on the acquisition and/or selective pressure of resistance genes, that in general carried by plasmids, which contribute to the rapid resistance spread within specific bacterial populations. The purpose of this study was to determine the prevalence of multidrug-resistant (MDR) organisms recovered from surveillance cultures (rectal, anal, nasal, and axillary) isolated from several Brazilian public hospitals. Methods Surveillance samples were analyzed from 1st January to 31st December 2023. All the swab samples were collected from patients admitted from high-risk settings or transferred from areas with high rates of MDR organisms. All clinical specimens were inoculated onto a selective media (ChromID media, bioMerieux) and incubated at 35±2°C for 18-24 h. The screening test was performed to Gram-negative bacilli (GNB) using the antibiotics ertapenem, imipenem and meropenem and for Gram-positive Cocci (GPC) screening was tested oxacillin (methicillin) and vancomycin using disc diffusion method. Bacteria identification was performed by MALDI-TOF mass spectrometry (Vitek-MS and bioMerieux) and as required, the minimal inhibitory concentrations (MIC) of antibiotics were determined using the Vitek 2 System (bioMerieux). Results A total of 71,064 rectal swab samples were processed. 4,652/71,064 (6.5%) samples showed positive results for the presence of 5.255 MDR microorganisms. Out of which, 4,477/5,255 (85.2%) were Gram-negative Bacilli (GNB) and 778/5,255 (14.8%) were Gram-positive Cocci (GPC). The majority of the isolated carbapenems resistance was Klebsiella spp. [2,791/5,255 (53.1%)], which the Klebsiella pneumoniae was the predominant species, followed by Acinetobacter baumannii [1,268/5,255 (24.1%)], Pseudomonas aeruginosa [263/5,255 (5%)], Enterobacter spp. [96/5255 (1.8%)], Serratia spp. [31/5,255 (0.6%)], Citrobacter spp. [22/5,255 (0.4%)], and others BGN [6/5,255 (0.2%)]. The resistance to vancomycin and methicillin were observed in 10% (525/5,255) of Enterococcus spp. and 4.8% (253/5,255) of Staphylococcus aureus, respectively. Conclusions The study found that K. pneumoniae carbapenem-resistant and Enterococcus spp. vancomycin-resistant were the main Gram-negative and Gram-positive organisms recovered from surveillance culture samples. However, our data draws attention to the increased cases of carbapenem-resistant Acinetobacter baumannii as the second most frequent pathogen isolated from surveillance samples after the last wave of COVID-19 pandemic. Active surveillance culture for target microorganisms followed by contact precautions to minimize the chance of transmitting MDR organisms from colonized to non-colonized patients consist of an important activity of infection control policy.

B-205 Performance of a Novel Fluorogenic Assay for Detection of Carbapenemase-producing Enterbacteriaceae

Abstract Background In recent years, under the selective challenge of antibiotics, the variety and number of drugresistant pathogenic microorganisms have increased significantly, which brings great challenges to clinical diagnosis and treatment, especially the infection caused by carbapenem resistant Enterobacteriaceae (CRE). Carbapenemases production is the main mechanism of drug resistance of Enterobacteriaceae to carbapenems. Drug resistance of Carbapenems can be caused by three mechanisms, resulting in the production of five major Carbapenemases. These are Klebsiella pneumoniae enzyme (KPC), New Delhi metal β-lactamase (NDM), carbapenem hydrolyzed oxalase (OXA-48 like), integrin-encoded metal β- lactamase (VIM) and IMP (Imipenemase). The real-time fluorescence quantitative PCR (qPCR) method based on molecular beacons was combined with the melting curve analysis to identify five drug resistance genes simultaneously by a single PCR reaction, with rapid detection, high sensitivity and strong specificity. Based on this principle, we developed the novel fluorogenic assay for rapid detection of Carbapenemases in multidrug-resistant Enterobacteriaceae (Dynamiker Biotechnology (Tianjin) Co., Ltd.). Methods We evaluated the performance of the novel fluorogenic assay for rapid detection of Carbapenemases in multidrug-resistant Enterobacteriaceae, including the limit of detection (LoD) and cross-reactivity, and compared it with the lateral flow immunochromatography assay (LFA). Results The LoD ranged from 75-450 CFU/mL for the five carbapenemase genes. The analytical specificity for target genes was 100%, as assessed with a panel of 15 pathogens, which indicated no cross-reactions. Comparison of qPCR and LFA results from twenty-three CRE clinical isolates with characterized carbapenemase content demonstrated a complete agreement (Table 1). Conclusions The novel fluorogenic assay for rapid detection of Carbapenemases in multidrug-resistant Enterobacteriaceae is an accurate and rapid method to identify KPC, NDM, VIM, IMP and OXA-48-like carbapenemases in the clinical microbiology laboratory, which can guide infection control programs to limit the spread of these organisms.

Prevalence, Risk Factors, and Outcome of Carbapenem-resistant Acinetobacter Infections in a Community Hospital in Madinah, Saudi Arabia.

Acinetobacter is a Gram-negative bacterium that causes nosocomial infections, increasing healthcare costs, patient morbidity, and mortality. The rate of carbapenem resistance among Acinetobacter species is rising in several countries, including Saudi Arabia. To determine the risk factors and compare the predictors of mortality in patients infected with carbapenem-susceptible and carbapenem-resistant Acinetobacter strains. This retrospective study included patients with Acinetobacter infection who were admitted to a community hospital in Madinah, Saudi Arabia, between January 2017 and June 2021. A logistic regression analysis was conducted to assess the risks of acquiring carbapenem-resistant Acinetobacter infections and the mortality risk associated with these infections. This study included 138 Acinetobacter-infected cases, of which 114 (82%) were carbapenem-resistant infections. Between 2017 and 2020, resistance rates increased from 75% to 87%. Patients with carbapenem-resistant Acinetobacter infections had higher 90-day mortality than those with carbapenem-susceptible infection (62% vs. 29%, P = 0.006). The risk factors for carbapenem-resistant Acinetobacter infections were prior antimicrobial therapy (aOR: 8.36 [1.69-41.29]; P = 0.009) and mechanical ventilation (aOR: 6.07 [1.82-20.20]; P = 0.003). Among all patients with Acinetobacter infections, significant predictors of 90-day mortality were carbapenem resistance (aOR: 3.26 [1.19-8.90]; P = 0.021) and Charlson comorbidity score (aOR: 1.19 [1.06-1.34]; P = 0.004). The increase in carbapenem-resistant Acinetobacter cases in this study was consistent with the findings of other studies from Saudi Arabia. This, together with the high associated mortality rates, indicates the urgent need for effective antimicrobials and infection prevention strategies to combat carbapenem-resistant Acinetobacter infections in hospitals.

Genotypic Characterisation of Carbapenemases in Clinical Isolates of Acinetobacter baumannii in a Tertiary Care Teaching Hospital of Kerala

Background: The multidrug-resistant and carbapenem-resistant Acinetobacter baumannii strains are an increasing global concern. The primary cause of carbapenem resistance in A. baumannii is production of various beta-lactamases with versatile hydrolytic capacities. The present study investigates the prevalence of different carbapenemases among clinical isolates of carbapenem-resistant A. baumannii from a tertiary care teaching hospital in Kerala, India. Methods: Non-duplicate isolates from sputum, endotracheal aspirate, pus, urine, and blood samples were included in the study. Isolate identification and antibiotic susceptibility testing of the isolates were done using vitek 2 system. A conventional polymerase chain reaction was used to detect the carbapenemase-encoding genes in the isolates. Results: A total of the 126 isolates studied, among these 105 (83.3%) isolates were from intensive care unit patients and 21(16.6%) were from non-ICU inpatients. Most of the isolates were obtained from respiratory specimens-78(61.9%) followed by pus-33(26.2%), urine-12(9.5%) and blood- 3(2.4%). The prevalence of carbapenemase genes were as follows:blaOXA-51(n=126, 100%) blaOXA-23 (n=98, 77.7%), blaOXA-58 (n=7, 5.6%), blaNDM (n=66, 52.4%), blaIMP (n=26, 20.6%), and blaVIM (n=20, 15.9%). Among the total isolates 78 (61.9%) isolates harbored metallo-beta-lactamase genes, 47(37.3%) isolates harboured a single carbapenemase gene, while 69(54.8%) isolates harbored two or more genes. Conclusion: This study reveals a higher prevalence of metallo-beta lactamases and the co-occurrence of multiple carbapenemase genes in Carbapenem resistant A. baumannii isolates.

Occurrence of blaOXA-116 Carbapenemase in Escherichia coli ST2519 of Clinical Origin: A Report from Northeast India.

Carbapenem-resistant Escherichia coli pose a significant threat to global public health due to the dearth of available treatment options, resulting in infections with high mortality and morbidity. The study aimed to investigate the mechanism of carbapenem resistance in a carbapenem non-susceptible E. coli isolate recovered from an urinary tract infection patient admitted to a tertiary referral hospital, through whole-genome sequencing using Illumina NovaSeq 6000 platform. Carbapenemase production followed by antibiotic susceptibility testing were performed following Clinical Laboratory Standard Institute guidelines.Polymerase chain reaction targeting carbapenemase genes was performed followed by an investigation of horizontal transferability. The Center for Genomic Epidemiology database was used to analyze the sequenced data. ST2519 E. coli BJD_EC1808 with a genome size of 5.8 Mb harbored Col440I plasmid and a chromosomally located blaOXA-116 gene with an IS18 element upstream, along with multiple antibiotic resistance genes conferring clinical resistance toward beta-lactams, aminoglycosides, amphenicols, sulfonamides, tetracyclines, trimethoprim, rifampin, macrolide, and streptogramin antibiotics and antiseptics. E. coli ST2519 harboring blaOXA-116 associated with a mobile genetic element exhibiting carbapenem resistance is a public health threat due to its limiting effect on the therapeutic usage of carbapenem and their dissemination into carbapenem non-susceptible phenotypes will contribute to carbapenem resistance burden and, therefore, warrants urgent monitoring and clinical intervention.

Detection of antibiogram pattern and prevalence of blaNDM and blaVIM genes among carbapenem resistant Eschericia coli isolates in a tertiary care hospital

Background: Clinical significance of Eschericia coli is due to its capacity to develop resistance against many antibiotics specially carbapenem by producing carbapenemase which limits treatment options. blaNDM and blaVIM genes are responsible for this multi-drug-resistance. This is an alarming condition for society as well as clinician because they are very difficult to treat due to limited options of antibiotics and its high virulence. Aims and Objectives: The study was conducted to detect antibiogram pattern and prevalence of blaNDM and blaVIM genes among carbapenem resistant E. coli isolates in a tertiary care hospital. Our study will also help to identify molecular basis of Carbapenem resistance including treatment guideline. Materials and Methods: Cross-sectional study was performed during January’2021 to June’2022 time period. Identification of E. coli was followed by standard protocol and antibiotic sensitivity was done by Kirby-Bauer disk-diffusion method following CLSI guideline, using VITEK-2D-SYSTEM. Detection of genes was done by polymerase chain reaction (PCR). Results: Out of 1867 samples, 161 isolates were identified as E. coli, among which 90 were resistant to carbapenem. Among those, 50 (31%) were resistant to all three carbapenems. Among those, 100% were resistant to ampicillin, amoxycillin-clavulanic acid, cefuroxime, cefuroxime-axetil, cefoperazone-sulbactam, ceftriaxone, cefepime, piperacillin-tazobactam, nalidixic acid, ciprofloxacin; 52% were resistant to amikacin; 54% were resistant to gentamicin; 86% were resistant to cotrimoxazole; 22% were resistant to nitrofurantoin and 96% were sensitive to tigecycline; 100% were intermediate sensitive to colistin. blaNDM gene is detected in 17 (34%) isolates and blaVIM gene is detected in 2 (4%) isolates by PCR. Conclusion: E. coli leads to common clinical presentations such as urinary tract infection, blood stream infection, pneumonia, and wound infection. High burden of carbapenem resistant E. coli was associated with urinary indoor samples. Rising incidence of multi-drug-resistance occurs due to lack of infection control measures, irrational use of antibiotics. Continuous surveillance will prevent development of multi-drug-resistance and help to improve treatment guideline.

Characteristics of Oral Acinetobacter spp. and Evolution of Plasmid-Mediated Carbapenem Resistance in Bacteremia Patients with Hematological Malignancies.

Patients with hematological malignancies are more susceptible to infections, leading to a poor prognosis. Acinetobacter colonization is a risk factor for secondary bacteremia. Antibiotic susceptibility phenotypes and genomic characteristics of 48 oral Acinetobacter spp. and one bloodstream Acinetobacter baumannii from patients with hematological malignancies were analyzed by antimicrobial susceptibility tests and whole-genome sequencing. We conducted comparative genomic analysis of oral and blood isolates from the same patient. A. baumannii was the most common (72.92%, 35/48) Acinetobacter species in oral Acinetobacter spp. isolates. Seventeen different A. baumannii sequence types were identified using the Pasteur MLST scheme; however, the dominant global clones GC1 and GC2 were not present. Among the isolates, 46 (95.8%) were carbapenem-susceptible Acinetobacter spp. One patient treated with meropenem for 15 days developed A. baumannii bacteremia 46 days after the isolation of oral A. baumannii AOR07. Oral and bloodstream isolates from the same patient were closely related to only four non-synonymous mutations on the chromosome. The bla OXA-58 gene was transferred between plasmids through XerCD-mediated recombination, leading to an elevated copy number, causing carbapenem resistance in bloodstream isolates. Oral Acinetobacter spp. may cause secondary bacteremia. The amplification and transfer of bla OXA-58 in the plasmids explained the increased carbapenem resistance in the bloodstream isolate.

Phenotypic and Genetic Characteristics of Carbapenemase-Producing Enterobacterales Isolates at Okayama University Hospital.

Spread of carbapenemase-producing Enterobacterales (CPE) is an ongoing public health issue worldwide, including in Japan. In this study, we investigated the phenotypic and genetic characteristics of CPE isolates at Okayama University Hospital over the 5 years (2013-2018) prior to the outbreak of the 2019 coronavirus pandemic. Of 24 carbapenem-resistant Enterobacterales isolated during the study period, we identified 8 CPE isolates harboring blaIMP-1 (5 isolates) and blaIMP-6 genes (3 isolates). Bacterial species and carbapenem susceptibility patterns exhibited diversity. Minimum inhibitory concentrations (MICs) of meropenem were generally higher than those of imipenem and biapenem. Results of pulsed-field gel electrophoresis demonstrated that neither clonal nor plasmid-mediated outbreaks of blaIMP-harboring CPE isolates have developed at our hospital. One Klebsiella oxytoca isolate showed a high MIC (128 μg/mL) of meropenem, which could be explained by the high plasmid copy number. Subsequent analysis of this isolate may elucidate the intricacies of carbapenem resistance profiles among CPE isolates. Collectively, our findings underscore the necessity for ongoing genetic surveillance of CPE, complemented by tailored approaches for infection prevention and control.

Molecular Detection of Carbapenemases in Acinetobacter baumannii Strains of Portugal and Association With Sequence Types, Capsular Types, and Virulence

Carbapenem-resistant Acinetobacter baumannii (CRAB) is an important nosocomial pathogen. The capsular type (K-type) is considered a major virulence factor, contributing to the evasion of host defenses. The global spread and dissemination dynamics between K-types, sequence types (ST), antibiotic resistance genes, and virulence factors remain largely unknown in Portugal. A collection of 96 CRAB clinical samples collected between 2005 and 2019 in the northern region of Portugal were tested for antimicrobial susceptibility profile and screened by polymerase chain reaction for resistance genetic determinants. A subset of 26 representative isolates was subjected to whole-genome sequencing to assess K types, ST types, and genomic relatedness. The pathogenicity of distinct K-types was also tested using Galleria mellonella model. For the 96 CRAB isolates analyzed, high antimicrobial resistance (>90%) was observed to the carbapenems, fluoroquinolones, and miscellaneous agents. Greater antimicrobial susceptibility (∼30%-57%) was observed for aminoglycosides, particularly tobramycin, and amikacin. Genotypically, 75 strains (78.5%) carried blaOXA-23-like, 18 strains (18.8%) carried blaIMP-like, and 11 strains (14.9%) carried blaOXA-40-like carbapenem resistance genes, respectively. Associations between OXA and ST/capsular locus (KL) types were observed over the years (eg, OXA-40-like/ST46Past/KL120 and OXA-23-like/ST2Past/KL2). ST2Past of clonal complex II was present in most strains, a dominant drug-resistant lineage in the United States and Europe. KL7 was also the most prevalent KL-type (38.5%), followed by KL2 (34.6%), KL120 (23.1%), and KL9 (3.8%). Virulence assessment for different K-types in a Galleria mellonella model revealed a significantly increased virulence for KL120 when compared with KL7, KL9, and KL2. There are specific CRAB serotypes circulating in Portugal, accounting by the low diversity of acquired carbapenemase genes (OXA-23-like and OXA-40-like), ST types (ST2 and ST46) and KL types (KL2, KL7, KL9, and KL120) identified. The high prevalent of ST2, especially when associated with KL2 and blaOXA-23-like, suggest that antibiotic resistance has been driven by clonal expansion of clonal complex II. Such findings provide useful information on the diversity of multidrug-resistant bacterium that might be relevant for antibacterial interventions.

Profile of Bacterial Resistance in Emergency Wards, Hospitalization, and Intensive Care Units in Mexican Hospitals with the Antimicrobial Stewardship Program

IntroductionBacterial infections are important causes of death. Some reports have shown that the COVID-19 pandemic may have had an impact on drug resistance. AimTo compare the resistance profiles between 2019 and 2020 emergency room, hospitalization non-intensive care units (ICU), and ICU areas in Mexican hospitals. MethodDatabases from 15 hospitals from 8 states in Mexico including susceptibility results for Escherichia coli, Klebsiella pneumoniae, and Pseudomonas aeruginosa collected from blood, urine, respiratory, and intra-abdominal secretions were analyzed. Different hospital areas were included: emergency room, hospitalization non-ICU, and ICU. The databases were converted using the BacLink2 tool and analyzed using the WHOnet 2022 platform. The percentage of antibiotic resistance was compared for each species and clinical specimen among years (2019 vs 2020) by a chi-square test using the criterion of p<0.05 for statistical significance. ResultsA statistically significant decrease in resistance to imipenem was observed for P. aeruginosa recovered from respiratory specimens in the ICU (55.6% vs 39.3%) and from urine samples from hospitalization non-ICU areas (52.9% vs 37.3%). Also, an increase in resistance was detected in blood isolates from hospitalization non-ICU areas for carbapenems in P. aeruginosa imipenem, (23.3 % vs 54.5%) and meropenem (27.4 % vs 49%) and in E. coli, ertapenem (2.4% vs 10.1%), imipenem (1.9% vs 13.2%), and meropenem (1.6% vs 6.8%). Also, an increase in resistance for imipenem (20.0 % vs 57.5%) and meropenem (17.6% vs 48.7 %) was detected in the emergency room for P. aeruginosa in respiratory isolates. ConclusionA decrease in carbapenem resistance for P. aeruginosa recovered from respiratory specimens in the ICU and from urine samples from hospitalization non-ICU areas was detected. In contrast, an increase in carbapenem resistance was detected for P. aeruginosa in blood isolates from hospitalization non-ICU areas and respiratory isolates from the emergency room. Regarding respiratory samples, a higher resistance for meropenem was detected in 2020 than in 2019 for K. pneumoniae (2.6 vs 10.6%, p = 0.05). These results underline the analysis of AMR in different hospital wards to focus efforts on AMR reduction.

Persistent carbapenem resistance in mechanically ventilated ICU patients: A before-and-after analysis of the COVID-19 surge

ObjectiveHow did the antimicrobial resistance profile of critically ill patients evolve before, during, and after COVID-19 surge periods? MethodsWe retrospectively analysed all critically ill mechanically ventilated adult patients admitted to eight Brazilian hospitals from January 1st, 2018, to April 30th, 2023. We stratified the patients into three periods based on their admission date: pre-surge (Jan 01/2018-Mar 01/2020), surge (Mar 01/2020 - Oct 01/2021), and post-surge (after Oct 01/2021). We compared the proportion of positive cultures, prevalence of pathogens, and resistance rates across periods using the rate ratios (RR) and their 95% confidence intervals (95% CI). ResultsWe analysed 9,780 ICU patients: 3,718 were in the pre-surge, 3,815 in the surge, and 2,247 in the post-surge period. Patients in the surge period were younger (median: 70 vs. 74 pre-surge vs. 75 post-surge) and presented a higher duration of invasive mechanical ventilation (median 7 vs. 5 days). The utilisation of blood and respiratory cultures increased throughout periods (56.9 pre-surge vs. 69.4 surge vs. 70.4 patients/1,000 patient days post-surge). The isolation of carbapenem-resistant gram-negative bacteria increased during the surge (RR [95% CI]: 1.8 [1.5-2.2], compared to pre-surge), decreased in post-surge (RR [95% CI]: 0.72 [0.6-0.9], and remained higher than pre-surge (RR [95% CI]: 1.3 [1.0-1.6]). Resistance rates for Pseudomonas aeruginosa reduced from 32% in pre- to 23% post-surge, whereas Klebsiella pneumoniae doubled during the surge, 26% to 52%, and remained higher than pre-surge. ConclusionCarbapenem resistance increased during the surge period. Although it decreased post-surge, it remained higher than the rates observed before the pandemic.

Metagenome mining divulges virulent and multidrug resistant Pseudomonas aeruginosa ST242 and Klebsiella michiganensis ST∗1b23 coinfecting an 8-month-old meningitis infant under ICU in Kampala, Uganda, East Africa

Pediatric meningitis is a global health problem, with insufficiently known pathogens and antibiotic resistance (AMR) especially in low-resource settings. Here, we sought to uncover the virulence and AMR of pathogens associated with infant meningitis, treated with ceftriaxone, in Kampala, Uganda. In a bid to isolate Klebsiella oxytoca, we coincidentally recovered a co-culture and challenged it with antibiotic susceptibility testing (AST) on a panel of 14 antibiotics. We then combined metagenome binning with antiSMASH/PRISM genome mining to unveil the pathogens, their virulence and molecular targets in relation to meningitis. From AST, the co-culture exhibited resistance to multiple aminoglycosides, fluroquinolones, and β-lactams, including ceftriaxone, the inherently used drug. From metagenome annotation, the first bin was identified as Pseudomonas aeruginosa ST242 and the second as Klebsiella michiganensis ST∗1b23. Among others, P. aeruginosa ST242 virulence factors include type 3 and type 6 secretion systems, biofilm, and nonribosomal peptides (NRPs) of the pyoverdine synthase operon, targeting claudin-5, a component of the tight junctions of the blood-brain barrier (BBB). The P. aeruginosa ST242 genome portrays intrinsic resistance to beta-lactamases (blaOXA-50 and blaPAO), aminoglycosides [aph(3′)-IIb)], fluoroquinolones (crpP), tetracycline (tmexD2) and fosfomycin (fosA), among others. From K. michiganensis ST∗1b23 genome mining we elucidated a yersiniabactin-related metabolite, targeting the ligand-binding domain of the human polymeric immunoglobulin receptor (pIgR) and other components of the BBB. The K. michiganensis ST∗1b23 chromosome encodes the genes blaOXY-1 and OqxA/B, conferring resistance to β-lactams, fluoroquinolones, and trimethoprim respectively. Notably, we found one frameshift and nine substitution mutations conferring carbapenem and cephalosporin resistance mechanisms. Overall, our findings strongly suggest coinfection and a mechanistic crosstalk between P. aeruginosa ST242 and K. michiganensis ST∗1b23 in the pathogenesis of meningitis in this case. Importantly, ceftriaxone could be an inappropriate treatment choice for these pathogens. Hence, serious surveillance and experimental studies will improve the management of pediatric meningitis.

Emerging Colistin Resistance: Experience from an Intensive Care Unit of a Tertiary Care Center in India.

Resistance to the currently available classes of antimicrobials has emerged as a major health concern, with a growing number of isolated organisms demonstrating resistance to available antimicrobials. With the escalation of carbapenem resistance, the use of polymyxin B/E (colistin) has increased over the years, which has, in turn, contributed to the worldwide emergence of colistin resistance. The available data on colistin resistance from Southeast Asian nations, including India, is limited, especially in intensive care unit (ICU) settings. The primary objective of our study was to analyze the clinical profile of patients with polymyxin B/E (colistin) resistant positive cultures and to study their outcome in terms of length of ICU stay and outcome at discharge. This observational, single-center, prospective study was conducted in a 20-bed adult ICU serving both medical and surgical patients at a 1,600-bed tertiary care institute in northern India between Jan 2020 and Dec 2021. In this study, all adult patients, defined as individuals older than 18 years of age, admitted to our ICU with cultures detecting polymyxin B/E (colistin) resistant organisms were included as cases, and all adult ICU patients (age >18 years) with polymyxin B/E (colistin) sensitive cultures were taken as controls. Clinical, laboratory, and demographic parameters, along with ICU variables like severity of illness, length of ICU stay, mechanical ventilation, days of shock, renal replacement therapy (RRT), etc., and outcome at discharge were collected. Identification of resistance to colistin and minimal inhibitory concentration (MIC) of colistin is based on Clinical and Laboratory Standards Institute (CLSI) guidelines. For statistical analysis, Mann-Whitney U test, Fisher's exact test, and binary logistic regression were used. Twenty-eight cases and 55 controls (n = 83) were included for analysis. Abdominal [gastrointestinal (GI)] sepsis was the most common diagnosis (41.8%) at admission to the ICU, and Klebsiella pneumoniae was the most common species isolated in both the cases (96.4%) and the controls (50.9%). The most common site of isolation of Gram-negative bacteria in both cases and controls was blood (71.4 vs 74.5%), followed by deep-seated pus (21.4 vs 23.64%). The most common class of drugs to which the cases were sensitive was tetracyclines in 60.7%, followed by ceftazidime-avibactam. Factors such as prior exposure to colistin, exposure to monobactams in the ICU, and ICU stay in days were identified as independent predictors for colistin resistance. Overall mortality was not statistically different between cases and controls (p -value 0.38). Factors like prolonged ICU stay, exposure to monobactams in the ICU, and a history of prior exposure to colistin in the previous 90 days are independent predictors of colistin resistance. The most common organism to develop colistin resistance identified was K. pneumoniae, and the most common site of isolation was blood. There was no difference in mortality between colistin-resistant and colistin-sensitive patients.

Acetylshikonin reduces the spread of antibiotic resistance via plasmid conjugation

The plasmid-mediated conjugative transfer of antibiotic resistance genes (ARGs) stands out as the primary driver behind the dissemination of antimicrobial resistance (AMR). Developing effective inhibitors that target conjugative transfer represents an efficient strategy for addressing the issue of AMR. Here, we studied the effect of acetylshikonin (ASK), a botanical derivative, on plasmid conjugation. The conjugative transfer of RP4-7 plasmid inter and intra species was notably reduced by ASK. The conjugation process of IncI2 and IncX4 plasmids harboring the mobile colistin resistance gene (mcr-1), IncX4 and IncX3 plasmids containing the carbapenem resistance gene (blaNDM-5), and IncFI and IncFII plasmids possessing the tetracycline resistance gene [tet(X4)] were also reduced by ASK. Importantly, the conjugative transfer frequency of mcr-1 positive IncI2 plasmid in mouse peritoneal conjugation model and gut conjugation model was reduced by ASK. The mechanism investigation showed that ASK disrupt the functionality of the bacterial cell membrane. Furthermore, the proton motive force (PMF) was dissipated. In addition, ASK blocked the electron transmission in bacteria's electron transport chain (ETC) through disturbing the quinone interaction, resulting in an insufficient energy supply for conjugation. Collectively, ASK is a potential conjugative transfer inhibitor, providing novel strategies to prevent the spread of AMR.