Genomic characterization of extended-spectrum ß-lactamase-producing Enterobacterales isolated from abdominal surgical patients.

Rectal swabs of 104 patients who underwent abdominal surgery were screened for ESBL producers. Sequence types (STs) and resistance genes were identified by whole-genome sequencing of 46 isolates from 17 patients. All but seven isolates were assigned to recognized STs. While 18 ESBL-producing E. coli (EPEC) strains were of unique STs, ESBL-producing K. pneumoniae (EPKP) strains were mainly ST14 or ST15. Eight patients harboured strains of the same ST before and after abdominal surgery. The most prevalent resistant genes in E. coli were blaEC (69.57%), blaCTX-M (65.22%), and blaTEM (36.95%), while blaSHV was present in only K. pneumoniae (41.30%). Overall, genes encoding ß-lactamases of classes A (blaCTX-M, blaTEM, blaZ), C (blaSHV, blaMIR, and blaDHA), and D (blaOXA) were identified, the most prevalent variants being blaCTX-M-15, blaTEM-1B, blaSHV-28, and blaOXA-1. Interestingly, blaCMY-2, the most common pAmpC ß-lactamase genes reported worldwide, and mobile colistin resistance genes, mcr-10-1, were also identified. The presence of blaCMY-2 and mcr-10-1 is concerning as they may constitute a potentially high risk of pan-resistant post-surgical infections. It is imperative that healthcare professionals monitor intra-abdominal surgical site infections rigorously to prevent transmission of faecal ESBL carriage in high-risk patients.

High prevalence of plasmid-mediated quinolone resistance in escherichia coli strains producing extended-spectrum beta-lactamases isolated from faeces and urine of pregnant women with acute cystitis.

BACKGROUND: Escherichia coli is the most common etiological agent of urinary tract infections (UTIs). Meanwhile, plasmid-mediated quinolone resistance (PMQR) is reported in E. coli isolates producing extended-spectrum ß-lactamases (ESBLs). Furthermore, the reservoirs and mechanisms of acquisition of uropathogenic Escherichia coli (UPEC) strains are poorly understood. On the other hand, UTIs are common in pregnant women and the treatment challenge is alarming. METHODS AND RESULTS: In the present study, 54 pregnant women with acute cystitis were included. A total of 108 E. coli isolates, 54 isolates from UTI and 54 isolates from faeces of pregnant women (same host) were collected. In the antimicrobial susceptibility test, the highest rate of antibiotic resistance was to nalidixic acid (77%, 83/108) and the lowest rate was to imipenem (9%, 10/108). Among the isolates, 44% (48/108) were ESBLs producers. A high frequency of PMQR genes was observed in the isolates. The frequency of PMQR genes qnrS, qnrB, aac(6')-Ib-cr, and qnrA was 58% (63/108), 21% (23/108), 9% (10/108), and 4% (4/108), respectively. Meanwhile, PMQR genes were not detected in 24% (20/85) of isolates resistant to nalidixic acid and/or fluoroquinolone, indicating that other mechanisms, i.e. chromosomal mutations, are involved in resistance to quinolones, which were not detected in the present study. In ESBL-producing isolates, the frequency of PMQR genes was higher than that of non-ESBL-producing isolates (81% vs. 53%). Meanwhile, UTI and faeces isolates mainly belonged to phylogenetic group B2 (36/54, 67% and 25/54, 46%, respectively) compared to other phylogenetic groups. In addition, virulence factors and multidrug-resistant (MDR) were mainly associated with phylogenetic group B2. However, predominant clones in faeces were not found in UTIs. Rep-PCR revealed the presence of 85 clones in patients. Among the clones, 40 clones were detected only in faeces (faeces-only), 35 clones only in UTI (UTI-only) and 10 clones in both faeces and UTI (faeces-UTI). We found that out of 10 faeces-UTI clones, 5 clones were present in the host's faeces flora. CONCLUSION: This study revealed a high rate of resistance to the quinolone nalidixic acid and a widespread distribution of PMQR genes in MDR E. coli strains producing ESBLs. The strains represented virulence factors and phylogenetic group B2 are closely associated with abundance in UTI and faeces. However, the predominant clones in faeces were not found in UTIs and it is possible that rep-PCR is not sufficiently discriminating clones.

Clinical manifestations, antimicrobial resistance and genomic feature analysis of multidrug-resistant Elizabethkingia strains.

BACKGROUND: Elizabethkingia is emerging as an opportunistic pathogen in humans. The aim of this study was to investigate the clinical epidemiology, antimicrobial susceptibility, virulence factors, and genome features of Elizabethkingia spp. METHODS: Clinical data from 71 patients who were diagnosed with Elizabethkingia-induced pneumonia and bacteremia between August 2019 and September 2021 were analyzed. Whole-genome sequencing was performed on seven isolates, and the results were compared with a dataset of 83 available Elizabethkingia genomes. Genomic features, Kyoto Encyclopedia of Genes and Genomes (KEGG) results and clusters of orthologous groups (COGs) were analyzed. RESULTS: The mean age of the patients was 56.9 ± 20.7 years, and the in-hospital mortality rate was 29.6% (21/71). Elizabethkingia strains were obtained mainly from intensive care units (36.6%, 26/71) and emergency departments (32.4%, 23/71). The majority of the strains were isolated from respiratory tract specimens (85.9%, 61/71). All patients had a history of broad-spectrum antimicrobial exposure. Hospitalization for invasive mechanical ventilation or catheter insertion was found to be a risk factor for infection. The isolates displayed a high rate of resistance to cephalosporins and carbapenems, but all were susceptible to minocycline and colistin. Genomic analysis identified five ß-lactamase genes (blaGOB, blaBlaB, blaCME, blaOXA, and blaTEM) responsible for ß-lactam resistance and virulence genes involved in stress adaptation (ureB/G, katA/B, and clpP), adherence (groEL, tufA, and htpB) and immune modulation (gmd, tviB, cps4J, wbtIL, cap8E/D/G, and rfbC). Functional analysis of the COGs revealed that "metabolism" constituted the largest category within the core genome, while "information storage and processing" was predominant in both the accessory and unique genomes. The unique genes in our 7 strains were mostly enriched in KEGG pathways related to microRNAs in cancer, drug resistance (ß-lactam and vancomycin), ABC transporters, biological metabolism and biosynthesis, and nucleotide excision repair mechanisms. CONCLUSION: The Elizabethkingia genus exhibits multidrug resistance and carries carbapenemase genes. This study presents a comparative genomic analysis of Elizabethkingia, providing knowledge that facilitates a better understanding of this microorganism.

Emergence of eravacycline heteroresistance in carbapenem-resistant Acinetobacter baumannii isolates in China.

Carbapenem-resistant Acinetobacter baumannii (CRAB) is resistant to almost all antibiotics. Eravacycline, a newer treatment option, has the potential to treat CRAB infections, however, the mechanism by which CRAB isolates develop resistance to eravacycline has yet to be clarified. This study sought to investigate the features and mechanisms of eravacycline heteroresistance among CRAB clinical isolates. A total of 287 isolates were collected in China from 2020 to 2022. The minimum inhibitory concentration (MIC) of eravacycline and other clinically available agents against A. baumannii were determined using broth microdilution. The frequency of eravacycline heteroresistance was determined by population analysis profiling (PAP). Mutations and expression levels of resistance genes in heteroresistant isolates were determined by polymerase chain reaction (PCR) and quantitative real-time PCR (qRT-PCR), respectively. Antisense RNA silencing was used to validate the function of eravacycline heteroresistant candidate genes. Twenty-five eravacycline heteroresistant isolates (17.36%) were detected among 144 CRAB isolates with eravacycline MIC values ≤4 mg/L while no eravacycline heteroresistant strains were detected in carbapenem-susceptible A. baumannii (CSAB) isolates. All eravacycline heteroresistant strains contained OXA-23 carbapenemase and the predominant multilocus sequence typing (MLST) was ST208 (72%). Cross-resistance was observed between eravacycline, tigecycline, and levofloxacin in the resistant subpopulations. The addition of efflux pump inhibitors significantly reduced the eravacycline MIC in resistant subpopulations and weakened the formation of eravacycline heteroresistance in CRAB isolates. The expression levels of adeABC and adeRS were significantly higher in resistant subpopulations than in eravacycline heteroresistant parental strains (P < 0.05). An ISAba1 insertion in the adeS gene was identified in 40% (10/25) of the resistant subpopulations. Decreasing the expression of adeABC or adeRS by antisense RNA silencing significantly inhibited eravacycline heteroresistance. In conclusion, this study identified the emergence of eravacycline heteroresistance in CRAB isolates in China, which is associated with high expression of AdeABC and AdeRS.

Cobalt complexes modulate plasmid conjugation in Escherichia coli and Klebsiella pneumoniae.

Antimicrobial resistance genes (ARG), such as extended-spectrum ß-lactamase (ESBL) and carbapenemase genes, are commonly carried on plasmids. Plasmids can transmit between bacteria, disseminate globally, and cause clinically important resistance. Therefore, targeting plasmids could reduce ARG prevalence, and restore the efficacy of existing antibiotics. Cobalt complexes possess diverse biological activities, including antimicrobial and anticancer properties. However, their effect on plasmid conjugation has not been explored yet. Here, we assessed the effect of four previously characterised bis(N-picolinamido)cobalt(II) complexes lacking antibacterial activity on plasmid conjugation in Escherichia coli and Klebsiella pneumoniae. Antimicrobial susceptibility testing of these cobalt complexes confirmed the lack of antibacterial activity in E. coli and K. pneumoniae. Liquid broth and solid agar conjugation assays were used to screen the activity of the complexes on four archetypical plasmids in E. coli J53. The cobalt complexes significantly reduced the conjugation of RP4, R6K, and R388 plasmids, but not pKM101, on solid agar in E. coli J53. Owing to their promising activity, the impact of cobalt complexes was tested on the conjugation of fluorescently tagged extended-spectrum ß-lactamase encoding pCTgfp plasmid in E. coli and carbapenemase encoding pKpQILgfp plasmid in K. pneumoniae, using flow cytometry. The complexes significantly reduced the conjugation of pKpQILgfp in K. pneumoniae but had no impact on pCTgfp conjugation in E. coli. The cobalt complexes did not have plasmid-curing activity, suggesting that they target conjugation rather than plasmid stability. To our knowledge, this is the first study to report reduced conjugation of clinically relevant plasmids with cobalt complexes. These cobalt complexes are not cytotoxic towards mammalian cells and are not antibacterial, therefore they could be optimised and employed as inhibitors of plasmid conjugation.

A practical approach to screening for carbapenemase-producing Enterobacterales- views of a group of multidisciplinary experts from English hospitals.

INTRODUCTION: Carbapenemase-producing Enterobacterales (CPE) are an important public health threat, with costly operational and economic consequences for NHS Integrated Care Systems and NHS Trusts. UK Health Security Agency guidelines recommend that Trusts use locally developed risk assessments to accurately identify high-risk individuals for screening, and implement the most appropriate method of testing, but this presents many challenges. METHODS: A convenience sample of cross-specialty experts from across England met to discuss the barriers and practical solutions to implementing UK Health Security Agency framework into operational and clinical workflows. The group derived responses to six key questions that are frequently asked about screening for CPE. KEY FINDINGS: Four patient groups were identified for CPE screening: high-risk unplanned admissions, high-risk elective admissions, patients in high-risk units, and known positive contacts. Rapid molecular testing is a preferred screening method for some of these settings, offering faster turnaround times and more accurate results than culture-based testing. It is important to stimulate action now, as several lessons can be learnt from screening during the COVID-19 pandemic, as well as from CPE outbreaks. CONCLUSION: Further decisive and instructive information is needed to establish CPE screening protocols based on local epidemiology and risk factors. Local management should continually evaluate local epidemiology, analysing data and undertaking frequent prevalence studies to understand risks, and prepare resources- such as upscaled screening- to prevent increasing prevalence, clusters or outbreaks. Rapid molecular-based methods will be a crucial part of these considerations, as they can reduce unnecessary isolation and opportunity costs.

Multi-drug Resistant Acinetobacter baumannii: Phenotypic and Genotypic Resistance Profiles and the Associated Risk Factors in Teaching Hospital in Jordan.

BACKGROUND: This study aimed to determine the prevalence of Antimicrobial Resistance Genes (ARGs), with a focus on colistin resistance in clinical A. baumannii, as well as the risk factors associated with A. baumannii infection in Jordanian patients. METHODS: In total, 150 A. baumannii isolates were obtained from patients at a teaching hospital. The isolates were tested for antimicrobial susceptibility using disc diffusion and microdilution methods. PCR amplification was used to detect ARGs, and statistical analysis was conducted to evaluate the influence of identified risk factors on the ARGs acquisition. RESULTS: More than 90% of A. baumannii isolates were resistant to monobactam, carbapenem, cephalosporins, Fluoroquinolones, penicillin, and ß-lactam agents. Moreover, 20.6% of the isolates (n = 31) were colistin-resistant. Several ARGs were also detected in A. baumannii isolates. Univariate analysis indicated that risk factors and the carriage of ARGs were significantly associated P ≤ (0.05) with gender, invasive devices, immunodeficiency, systemic diseases, tumors, and covid-19. Logistic regression analysis indicated seven risk factors, and three protective factors were associated with the ARGs (armA, strA, and strB) P ≤ (0.05). In contrast, tetB and TEM were associated with 2 risk factors each P ≤ (0.05). CONCLUSION: Our study indicates a high prevalence of MDR A. baumannii infections in ICU patients, as well as describing the case of colistin-resistant A. baumannii for the first time in Jordan. Additionally, the risk factors associated with ARGs-producing A. baumannii infections among ICU patients suggest a rapid emergence and spread of MDR A. baumannii without adequate surveillance and control measures.

High rates of intestinal colonization with carbapenemase producing Enterobacteriaceae in hematopoietic stem cell transplant recipients.

Carbapenem resistant Enterobacteriaceae (CRE) are major human pathogens because, these cause high number of difficult-to-treat infections. Allogeneic hematopoietic stem cell transplant (AHSCT) recipients are highly exposed to these type of bacteria. The aim of our study was to investigate prevalence of CRE colonization in AHSCT patients and to determine genes encoding carbapenem resistance. A retrospective study conducted between January 2015 and December 2019, involved 55 patients colonized with CRE strains. We determined the rate of antibiotic resistance according to European Committee on Antimicrobial Susceptibility Testing (EUCAST) and the carbapenem resistance genes by PCR assays for genes encoding most frequent ß-lactamases namely, blaGES, blaKPC, blaIMI, blaNDM, blaVIM, blaIMP and blaOXA-48. Eighty-one episodes of CRE colonization were recorded in 55 patients, mainly suffering from acute leukaemia (30%) and aplastic anemia (26%). History of hospitalization was noted in 80 episodes. Prior antibiotic treatment, severe neutropenia and corticosteroid therapy were respectively found in 94%, 76% and 58% of cases. Among the 55 patients, six patients (11%) developed a CRE infection. The CRE responsible for colonization were carbapenemase producers in 90% of cases. They belonged mostly to Klebsiella pneumoniae (61/81) and Escherichia coli species (10/81). Antibiotic resistance rates were 100% for ertapenem, 53% for imipenem, 42% for amikacin, 88% for ciprofloxacin and 27% for fosfomycin. Molecular study showed that blaOXA-48 gene was the most frequent (60.5%), followed by blaNDM (58%). Thirty-five (43%) strains were co-producers of carbapenemases. In our study, we report a high rate of CRE intestinal colonization in AHSCT recipients of our center.

Hidden threats in the plastisphere: Carbapenemase-producing Enterobacterales colonizing microplastics in river water.

Carbapenem resistance poses a significant burden on healthcare systems worldwide. Microplastics (MPs) have emerged as potential contributors to antibiotic resistance spread in the environment. However, the link between MPs and carbapenem resistance remains unexplored. We investigated the prevalence of carbapenem-resistant bacteria colonizing MPs placed in a river. Three replicates of a mixture of polypropylene (PP), polyethylene (PE) and polyethylene terephthalate (PET) and of PET alone were placed both upstream and downstream a wastewater treatment plant (WWTP) discharge. Carbapenem-resistant Enterobacterales (CRE) were further characterized by phenotypic tests and whole-genome sequencing. The abundance of carbapenem-resistant bacteria on MPs increased significantly downstream the WWTP. Their prevalence was higher in the MPs mixture compared to PET alone. CRE strains colonizing MPs included Klebsiella pneumoniae (n = 3), Klebsiella quasipneumoniae (n = 3), Raoultella ornithinolytica (n = 2), Enterobacter kobei (n = 1) and Citrobacter freundii (n = 1), most (n = 8) recovered after the WWTP discharge. All strains exhibited at least one of the tested virulence traits (biofilm formation at 37 °C, haemolytic activity and siderophore production), were multi-drug resistant and carried carbapenemase-encoding genes [blaKPC-3 (n = 5), blaGES-5 (n = 2) or blaKPC-3 + blaGES-5 (n = 3)]. Uncommon phenotypes of resistance to imipenem/relebactam (n = 3) and ceftazidime/avibactam (n = 2) were observed. Two blaKPC-3-positive K. pneumoniae successfully transfer this gene trough conjugation. Genome analysis predicted all strains as human pathogens. The blaKPC-3 was associated with the Tn4401d transposon on a pBK30683-like plasmid in most of the isolates (n = 7). The blaGES-5 was mostly linked to class 3 integrons. A K. pneumoniae strain belonging to the outbreak-causing high-risk clone ST15 carried both blaKPC-3 and blaCTX-M-15. Two K. quasipneumoniae isolates carried the plasmid-mediated colistin resistance gene mcr-9. Our results underscore the role of MPs as vectors for CRE dissemination, particularly following WWTPs discharges. MPs may act as carriers, facilitating the dissemination of carbapenemase-encoding genes and potentially contributing to increased CRE incidence in the environment.

Levofloxacin-induced MexS mutation triggers imipenem-relebactam resistance in a KPC-producing Pseudomonas aeruginosa.

OBJECTIVES: Imipenem-relebactam (IMR), a novel ß-lactam/ß-lactamase inhibitor combination, is recommended for infections caused by difficult-to-treat Pseudomonas aeruginosa. This study aimed to investigate the evolution trajectory of IMR resistance under the selection of levofloxacin in P. aeruginosa. METHODS: Antimicrobial susceptibility testing, complete genome sequencing and gene manipulation experiments were performed. Quantitative reverse transcription PCR for specific genes and porin levels were detected. Evolution trajectory was simulated in vitro by induction assay. RESULTS: P. aeruginosa HS347 and HS355 were isolated from abdominal drainage of two neighbouring patients (S and Z) undergoing surgery of colon carcinoma in Shanghai, China, with the latter patient having received levofloxacin. They were closely related ST16 strains, and both carried blaKPC-2 plasmids highly similar to those of P. aeruginosa endemic clones from Zhejiang province, where patient Z had received enteroscopy before this admission. Acquisition of resistance was observed for both IMR and fluoroquinolones in HS355, likely prompted by treatment with levofloxacin. The T274I substitution in MexS (putative oxidoreductase), upregulated efflux pump operon mexEF-oprN and decreased production of porin OprD leading to cross-resistance to fluoroquinolones and IMR, which was also verified by in vitro mutant selection under levofloxacin selection. CONCLUSIONS: The emergence of a rare blaKPC-2-plasmid-bearing ST16 clone implies the horizonal spread and inter-regional dissemination of a high-risk plasmid-clone combination, representing a public health challenge. Levofloxacin exposure can select for mexS inactivating mutation, which in turn leads to IMR resistance phenotype, implicating the role of an unrelated, widely used antimicrobial agent in insidiously triggering the development of cross resistance to a latest ß-lactam/ß-lactamase inhibitor combination.

Evolution of ceftazidime-avibactam and cefiderocol resistance in ST131-H30R1-Escherichia coli isolates with KPC-3 mutants and application of FTIR biotyping.

Ceftazidime-avibactam and cefiderocol represent two of the few alternatives for infections by KPC-producing Enterobacterales. We reported the emergence of resistance to both ceftazidime-avibactam and cefiderocol in a KPC-producing ST131-Escherichia coli (KPC-ST131-Ec) clinical isolate. Antimicrobial susceptibility testing, Fourier-transform infrared (FTIR) spectroscopy, whole-genome sequencing, and cloning experiments were performed. A KPC-49-Ec isolate resistant to ceftazidime-avibactam (MICCZA > 16/4 mg/L) and susceptible to cefiderocol (MICFDC: 2 mg/L) was recovered in a blood sample from an oncologic patient hospitalized in the medical ICU (June 2019) during ceftazidime-avibactam treatment. After 44 days, a KPC-31-Ec resistant to both ceftazidime-avibactam and cefiderocol (MICCZA > 16/4 mg/L, MICFDC: 8 mg/L) was found in a rectal sample during a second cycle of ceftazidime-avibactam treatment. Both KPC-49 (R163S) and KPC-31 (D179Y) were detected in the epidemic ST131-H30R1-Ec high-risk clone and showed a phenotype resembling that of ESBL producers. FTIR spectroscopy managed to differentiate cefiderocol-susceptible and resistant ST131-Ec isolates, and these from others belonging to different clones. After cloning and transformation experiments, KPC-49 and KPC-31 were responsible for ceftazidime-avibactam resistance (MICCZA > 16/4 mg/L) and decreased carbapenem MICs (MICMER ≤ 0.12 mg/L, MICIMI ≤ 1 mg/L). KPC-31 was also shown to be associated with increased MICs of cefiderocol (twofold and threefold dilutions over KPC-3 and KPC-49, respectively). However, mutations in proteins participating in outer membrane stability and integrity, such as TolR, could have a more relevant role in cefiderocol resistance. The effects of ceftazidime-avibactam and cefiderocol co-resistance in clinical isolates of Enterobacterales producing KPC mutants make their identification challenging for clinical laboratories.IMPORTANCEThroughout four admissions in our hospital of a single patient, different KPC-3 variants (KPC-3, KPC-49, and KPC-31) were found in surveillance and clinical ST131-Escherichia coli isolates, after prolonged therapies with meropenem and ceftazidime-avibactam. Different patterns of resistance to cefiderocol and ceftazidime-avibactam emerged, accompanied by restored carbapenem susceptibility. The inability to detect these variants with some phenotypic methods, especially KPC-31 by immunochromatography, and the expression of a phenotype similar to that of ESBL producers, posed challenge to identify these variants in the clinical microbiology laboratory. Molecular methods and whole-genome sequencing are necessary and new techniques able to cluster or differentiate related isolates could also be helpful; this is the case of Fourier-transform infrared spectroscopy, which managed in our study to discriminate isolates by cefiderocol susceptibility within ST131, and those from the non-ST131 ones.

Characteristics of non-carbapenemase producing carbapenem-resistant Klebsiella pneumoniae from a tertiary hospital in China.

INTRODUCTION: The spread of carbapenem-resistant Klebsiella pneumoniae (CRKP) is a substantial severe global public health burden. Non-carbapenemase-producing CRKP (non-CP-CRKP) is increasingly recognized as the source of severe infections. METHODOLOGY: We analyzed the genotypic, and phenotypic profiles of non-CP-CRKP strains with the whole-genome sequences isolated between 2017 and 2019 and the clinical characterization of non-CP-CRKP infection. RESULTS: A total of 91 CRKP strains were collected, of which 5 (5.49%) strains were non-CP-CRKP. Four strains were from male patients; three strains were isolated from the bile of patients who underwent biliary interventional surgery and four had a history of antibiotic exposure. Three strains were sequence type (ST)11, one was ST1, and one was ST5523. The non-CP-CRKP strains were insusceptible to ertapenem. Three strains were susceptible to amikacin. All the strains were susceptible to imipenem, meropenem, tigecycline, ceftazidime/avibatam and polymyxin B. The ß-lactamases of non-CP-CRKP predominantly included blaCTX-M, blaSHV, and blaTEM subtypes. Two site mutations in ompK36 (p.A217S and p.N218H) and four in ompK37 (p.I70M, p.I128M, p.N230G, and m233_None234insQ) were detected accounting for carbapenem resistance. Plasmids IncFI and IncFII were found in most strains. Genes encoding aerobactin, yersiniabactin and allantoin utilization were not detected in several isolates, and all non-CP-CRKP strains did not carry rmpA gene. CONCLUSIONS: Non-CP-CRKP infected patients had a history of previous antibiotic exposure or invasive procedures. Non-CP-CRKP strains were insusceptible to ertapenem. The mechanism of resistance includes ß-lactamases production and the site mutations in ompK36 and ompK37. Several virulence genes were not detected in non-CP-CRKP.

Global spread characteristics of CTX-M-type extended-spectrum ß-lactamases: A genomic epidemiology analysis.

BACKGROUND: Extended-spectrum ß-lactamases (ESBLs) producing bacteria have spread worldwide and become a global public health concern. Plasmid-mediated transfer of ESBLs is an important route for resistance acquisition. METHODS: We collected 1345 complete sequences of plasmids containing CTX-Ms from public database. The global transmission pattern of plasmids and evolutionary dynamics of CTX-Ms have been inferred. We applied the pan-genome clustering based on plasmid genomes and evolution analysis to demonstrate the transmission events. FINDINGS: Totally, 48 CTX-Ms genotypes and 186 incompatible types of plasmids were identified. The geographical distribution of CTX-Ms showed significant differences across countries and continents. CTX-M-14 and CTX-M-55 were found to be the dominant genotypes in Asia, while CTX-M-1 played a leading role in Europe. The plasmids can be divided into 12 lineages, some of which forming distinct geographical clusters in Asia and Europe, while others forming hybrid populations. The Inc types of plasmids are lineage-specific, with the CTX-M-1_IncI1-I (Alpha) and CTX-M-65_IncFII (pHN7A8)/R being the dominant patterns of cross-host and cross-regional transmission. The IncI-I (Alpha) plasmids with the highest number, were presumed to form communication groups in Europe-Asia and Asia-America-Oceania, showing the transmission model as global dissemination and regional microevolution. Meanwhile, the main kinetic elements of blaCTX-Ms showed genotypic preferences. ISEcpl and IS26 were most frequently involved in the transfer of CTX-M-14 and CTX-M-65, respectively. IS15 has become a crucial participant in mediating the dissemination of blaCTX-Ms. Interestingly, blaTEM and blaCTX-Ms often coexisted in the same transposable unit. Furthermore, antibiotic resistance genes associated with aminoglycosides, sulfonamides and cephalosporins showed a relatively high frequency of synergistic effects with CTX-Ms. CONCLUSIONS: We recognized the dominant blaCTX-Ms and mainstream plasmids of different continents. The results of this study provide support for a more effective response to the risks associated with the evolution of blaCTX-Ms-bearing plasmids, and lay the foundation for genotype-specific epidemiological surveillance of resistance, which are of important public health implications.

Emerging of multidrug-resistant Pseudomonas guariconensis with blaVIM-2 in an asymptomatic bacteriuria patient: A rare clinical presentation.

This case presents the clinical and genomic aspects of a rare and multidrug-resistant Pseudomonas guariconensis isolate carrying blaVIM-2 and highlights the need for heightened awareness in healthcare facilities. A 63-year-old woman underwent surgery for the diagnosis of a paraspinal abscess and infectious spondylitis. During hospitalization, the patient was diagnosed with heart failure exacerbation. The patient had no symptoms of urinary tract infection and met the criteria for asymptomatic bacteriuria. In urine culture, colonies of the organism grew >105 CFU/mL on blood agar and on MacConkey agar. The Bruker Biotyper mass spectrometry showed P. guariconensis. Based on the 16S rRNA gene sequence showed that a 99.79 % match with as P. guariconensis LMG 27394T. The average nucleotide identity with P. guariconensis LMG 27394T was 91.53 %. Antimicrobial susceptibility testing showed that the isolate was not susceptible to most of the antibiotics. Antimicrobial resistance genes identified were aph(6)-Id, aph(3″)-Ib, aac(6')-Ib3, aac(3)-If, gyrA mutation (T83I) and blaVIM-2.

OXCT1 functions as a succinyltransferase, contributing to hepatocellular carcinoma via succinylating LACTB.

Metabolic reprogramming is an important feature of cancers that has been closely linked to post-translational protein modification (PTM). Lysine succinylation is a recently identified PTM involved in regulating protein functions, whereas its regulatory mechanism and possible roles in tumor progression remain unclear. Here, we show that OXCT1, an enzyme catalyzing ketone body oxidation, functions as a lysine succinyltransferase to contribute to tumor progression. Mechanistically, we find that OXCT1 functions as a succinyltransferase, with residue G424 essential for this activity. We also identified serine beta-lactamase-like protein (LACTB) as a main target of OXCT1-mediated succinylation. Extensive succinylation of LACTB K284 inhibits its proteolytic activity, resulting in increased mitochondrial membrane potential and respiration, ultimately leading to hepatocellular carcinoma (HCC) progression. In summary, this study establishes lysine succinyltransferase function of OXCT1 and highlights a link between HCC prognosis and LACTB K284 succinylation, suggesting a potentially valuable biomarker and therapeutic target for further development.

Recovery and genetic characterization of clinically-relevant ST2 carbapenem-resistant Acinetobacter baumannii isolates from untreated hospital sewage in Zhejiang Province, China.

The global transmission of carbapenem-resistant Acinetobacter baumannii (CRAB) poses a significant and grave threat to human health. To investigate the potential relationship between hospital sewage and the transmission of CRAB within healthcare facilities, isolates of Acinetobacter spp. obtained from untreated hospital sewage samples were subjected to antimicrobial susceptibility tests, genome sequencing, and bioinformatic and phylogenetic tree analysis, and that data were matched with those of the clinical isolates. Among the 70 Acinetobacter spp. sewage isolates tested, A. baumannii was the most prevalent and detectable in 5 hospitals, followed by A. nosocomialis and A. gerneri. Worryingly, 57.14 % (40/70) of the isolates were MDR, with 25.71 % (18/70) being resistant to carbapenem. When utilizing the Pasteur scheme, ST2 was the predominant type among these CRAB isolates, with Tn2006 (ΔISAba1-blaOXA-23-ATPase-yeeB-yeeA-ΔISAba1) and Tn2009 (ΔISAba1-blaOXA-23-ATPase-hp-parA-yeeC-hp-yeeB-ΔISAba1) being the key mobile genetic elements that encode carbapenem resistance. Seven A. gerneri isolates which harbored Tn2008 (ISAba1-blaOXA-23 -ATPase) and the blaPER-1 gene were also identified. Besides, an A. soil isolate was found to exhibit high-level of meropenem resistance (MIC ≥128 mg/L) and harbor a blaNDM-1 gene located in a core genetic structure of ISAba125-blaNDM-1-ble-trpF-dsbC-cutA. To investigate the genetic relatedness between isolates recovered from hospital sewage and those collected from ICUs, a phylogenetic tree was constructed for 242 clinical isolates and 9 sewage isolates. The results revealed the presence of two evolutionary clades, each containing isolates from both ICU and sewage water, suggesting that CRAB isolates in untreated sewage water were also the transmission clones or closely related evolutionary isolates recoverable in hospital settings. Findings in this work confirm that hospital sewage is a potential reservoir of CRAB.

Carbapenem-resistant Klebsiella pneumoniae: the role of plasmids in emergence, dissemination, and evolution of a major clinical challenge.

INTRODUCTION: Klebsiella pneumoniae is a major agent of healthcare-associated infections and a cause of some community-acquired infections, including severe bacteremic infections associated with metastatic abscesses in liver and other organs. Clinical relevance is compounded by its outstanding propensity to evolve antibiotic resistance. In particular, the emergence and dissemination of carbapenem resistance in K. pneumoniae has posed a major challenge due to the few residual treatment options, which have only recently been expanded by some new agents. The epidemiological success of carbapenem-resistant K. pneumoniae (CR-Kp) is mainly linked with clonal lineages that produce carbapenem-hydrolyzing enzymes (carbapenemases) encoded by plasmids. AREAS COVERED: Here, we provide an updated overview on the mechanisms underlying the emergence and dissemination of CR-Kp, focusing on the role that plasmids have played in this phenomenon and in the co-evolution of resistance and virulence in K. pneumoniae. EXPERT OPINION: CR-Kp have disseminated on a global scale, representing one of the most important contemporary public health issues. These strains are almost invariably associated with complex multi-drug resistance (MDR) phenotypes, which can also include recently approved antibiotics. The heterogeneity of the molecular bases responsible for these phenotypes poses significant hurdles for therapeutic and diagnostic purposes.

Novel Antimicrobial Peptide SAAP Mutant as a Better Adjuvant to Sulbactam-Based Treatments Against Clinical Strains of XDR Acinetobacter baumannii.

The production of extended spectrum ß-lactamases (ESBLs) in extensively drug-resistant (XDR) strains of Acinetobacter baumannii has created havoc amongst clinicians making the treatment procedure challenging. Carbapenem-resistant strains have displayed total ineffectiveness towards newer combinations of ß-lactam-ß-lactamase inhibitors (ßL-ßLI) in tertiary healthcare settings. Therefore, the present study was aimed to design potential ß-lactamase antimicrobial peptide (AMP) inhibitors against ESBLs produced by the strains. We have constructed an AMP mutant library with higher antimicrobial efficacy (range: ~ 15 to 27%) than their parent peptides. The mutants were thoroughly screened based on different physicochemical and immunogenic properties revealing three peptides, namely SAAP-148, HFIAP-1, myticalin-C6 and their mutants with safe pharmacokinetics profile. Molecular docking highlighted SAAP-148_M15 displaying maximum inhibitory potential with lowest binding energies against NDM1 (- 1148.7 kcal/mol), followed by OXA23 (- 1032.5 kcal/mol) and OXA58 (- 925.3 kcal/mol). The intermolecular interaction profiles displayed SAAP-148_M15 exhibiting hydrogen bonds and van der Waals hydrophobic interactions with the crucial residues of metallo ß-lactamase [IPR001279] and penicillin-binding transpeptidase [IPR001460] domains. Coarse-grained clustering and molecular dynamics simulations (MDS) further validated the stable backbone profile and minimal residue-level fluctuations of the protein-peptide complex that were maintained throughout the simulation timeframe. The present study hypothesised that the combination of sulbactam (ßL) with SAAP-148_M15 (ßLI) holds immense potential in inhibiting the ESBLs alongside restoration of sulbactam activity. The current in silico findings upon further experimental validations can pave path towards designing of successful therapeutic strategy against XDR strains of A. baumannii.

Global distribution and genomic characteristics of carbapenemase-producing Escherichia coli among humans, 2005-2023.

Carbapenem-resistant Escherichia coli (CREC) has become a major public health problem worldwide. To date, there is a limited understanding of the global distribution of CREC. In this study, we performed a comprehensive genomic analysis of 7, 731 CRECs of human origin collected from different countries worldwide between 2005 and 2023. Our results showed that these CRECs were distributed in 75 countries, mainly from the United States (17.49%), China (14.88%), and the United Kingdom (14.73%). Eight carbapenemases were identified among the CRECs analyzed, including KPC, IMP, NDM, VIM, OXA, FRI, GES, and IMI. NDM was the most predominant carbapenemase (52.15%), followed by OXA (30.09%) and KPC (14.72%). Notably, all CRECs carried multiple antibiotic resistance genes (ARGs), with 178 isolates carrying mcr-1 and 9 isolates carrying tet(X). The CREC isolates were classified into 465 known sequence types (STs), with ST167 being the most common (11.5%). Correlation analysis demonstrated the significant role of mobile genetic elements in facilitating the transfer of carbapenem resistance genes. Furthermore, some CRECs from different countries showed high genetic similarity, suggesting clonal transmission exists. According to the GWAS results, the genetic difference of blaNDM-positive CRECs from China were mainly enriched in bacterial Type IV secretion system pathways compared with those from the United Kingdom and the United States. Therefore, continuous global surveillance of CRECs is imperative in the future.

Evaluation of antibiotic resistance, toxin-antitoxin systems, virulence factors, biofilm-forming strength and genetic linkage of Escherichia coli strains isolated from bloodstream infections of leukemia patients.

BACKGROUND: One of the most common complications in patients with febrile neutropenia, lymphoma, leukemia, and multiple myeloma is a bloodstream infection (BSI). OBJECTIVE: This study aimed to evaluate the antibiotic resistance patterns, virulence factors, biofilm-forming strength, and genetic linkage of Escherichia coli strains isolated from bloodstream infections (BSIs) of leukemia patients. METHODS: The study conducted in Iran from June 2021 to December 2022, isolated 67 E. coli strains from leukemia patients' bloodstream infections in hospitals in two different areas. Several techniques including disk diffusion and broth microdilution were used to identify patterns of antibiotic resistance, microtiter plate assay to measure biofilm formation, and PCR to evaluate the prevalence of different genes such as virulence factors, toxin-antitoxin systems, resistance to ß-lactams and fluoroquinolone antibiotics of E. coli strains. Additionally, the genetic linkage of the isolates was analyzed using the Enterobacterial Repeat Intergenic Consensus Polymerase Chain Reaction (ERIC-PCR) method. RESULTS: The results showed that higher frequency of BSI caused by E. coli in man than female patients, and patients with acute leukemia had a higher frequency of BSI. Ampicillin and Amoxicillin-clavulanic acid showed the highest resistance, while Imipenem was identified as a suitable antibiotic for treating BSIs by E. coli. Multidrug-resistant (MDR) phenotypes were present in 22% of the isolates, while 53% of the isolates were ESBL-producing with the blaCTX-M gene as the most frequent ß-lactamase gene. The fluoroquinolone resistance genes qnrB and qnrS were present in 50% and 28% of the isolates, respectively. More than 80% of the isolates showed the ability to form biofilms. The traT gene was more frequent than other virulence genes. The toxin-antitoxin system genes (mazF, ccdAB, and relB) showed a comparable frequency. The genetic diversity was detected in E. coli isolates. CONCLUSION: Our results demonstrate that highly diverse, resistant and pathogenic E. coli clones are circulating among leukemia patients in Iranian hospitals. More attention should be paid to the treatment and management of E. coli bloodstream infections in patients with leukemia.