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ABSTRACT: Objectives: This study was performed to investigate the activity of the novel ß-lactam/ß-lactamase inhibitor combination cefepime/enmetazobactam, against recently circulating Enterobacterales isolates from Europe from 2019 to 2021. Methods: A total of 2627 isolates were collected, and antimicrobial susceptibility was determined according to the European Committee on Antimicrobial Susceptibility Testing guidelines. Isolates with phenotypic resistance to ceftriaxone and ceftazidime (but susceptible to meropenem) and isolates nonsusceptible to meropenem were screened for the presence of ß-lactamases. Results: Overall, susceptibility to third-generation cephalosporins was 77%, and 97.3% were susceptible to meropenem. Cefepime/enmetazobactam susceptibility was 97.9% (72% of these isolates were Klebsiella pneumoniae from Italy), compared with 80.0% susceptibility to piperacillin/tazobactam and 99.4% to ceftazidime/avibactam. A total of 320 isolates (12.2%) were resistant to third-generation cephalosporins but susceptible to meropenem, and virtually all (96.3%) carried an extended-spectrum ß-lactamase with or without an AmpC and these were all susceptible to cefepime/enmetazobactam. Most meropenem-nonsusceptible isolates carried a KPC (68%), which were not inhibited by cefepime/enmetazobactam but were inhibited by ceftazidime/avibactam. Additionally, most meropenem-nonsusceptible isolates carrying OXA-48 (9/12 isolates) were susceptible to cefepime/enmetazobactam. Conclusions: Cefepime/enmetazobactam was highly active against Enterobacterales isolates, especially those resistant to third-generation cephalosporins. These data suggest that cefepime/enmetazobactam could be used as a carbapenem-sparing agent to replace piperacillin/tazobactam.

Wastewaters belong among the most important sources of environmental pollution, including antibiotic-resistant bacteria. The aim of the study was to evaluate treated wastewaters as a possible transmission pathway for bacterial colonisation of gulls occupying the receiving river. A collection of antibiotic-resistant Escherichia coli originating both from treated municipal wastewaters discharged to the river Svratka (Czech Republic) and nestlings of Black-headed Gull (Chroicocephalus ridibundus) living 35 km downstream of the outlet was obtained using selective cultivation. Isolates were further characterised by various phenotyping and genotyping methods.From a total of 670 E. coli isolates (450 from effluents, 220 from gulls), 86 isolates (41 from effluents, 45 from gulls) showed identical antibiotic resistance phenotype and genotype and were further analysed for clonal relatedness using pulsed-field gel electrophoresis (PFGE). Despite the overall high diversity of the isolates, 21 isolates from both sources showed similar PFGE profiles. Isolates belonging to epidemiologically important sequence types (ST131, 15 isolates; ST23, three isolates) were subjected to whole-genome sequencing. Subsequent phylogenetic analysis did not reveal any close clonal relationship between the isolates from the effluents and gulls' nestlings with the closest strains showing 90 SNPs difference.Although our study did not provide direct evidence of transmission of antibiotic-resistant E. coli to wild gulls via treated wastewaters, we observed gull chicks as carriers of diverse multi-resistant E. coli, including high-risk clones, posing risk of further bacterial contamination of the surrounding environment.

Objectives: Sexual transmission of extended-spectrum β-lactamase-producing Enterobacterales (ESBL-E) is suspected. We aimed to identify clusters of sexual behavior associated with ESBL-E carriage among individuals at risk of sexually transmitted infections (STI). Methods: In this cross-sectional study, patients attending an STI-screening center and human immunodeficiency virus outpatient clinic in Paris, France between 2018 and 2019 were asked questions on the following sexual activities in the last 6 months: receptive/insertive anal intercourse, passive/active rimming, receptive/insertive fellatio, receptive/insertive fisting, receptive/insertive fingering, active/passive cunnilingus. ESBL-E carriage was determined from rectal swabs. Bayesian latent class analysis was used to identify clusters of sexual activity, which were then associated with ESBL-E carriage using logistic regression. Results: Among 1211 men who have sex with men (MSM), those belonging to two latent classes with higher prevalence of insertive fingering and active rimming (ESBL-E prevalence=15.3%, N = 124 and 16.0%, N = 100) and one class with higher proportions of all behaviors (24.3%, N = 70) had a higher risk of ESBL-E carriage compared to those in a class with few sexual behaviors (7.3%, N = 259) after adjustment. Among 439 other men and 479 women, no clear associations between sexual clusters and ESBL-E carriage were observed. Conclusions: Sexual behaviors are associated with varying degrees of ESBL-E carriage, particularly among MSM.

The occurrence of carbapenemases encoding genes in Providencia rettgeri is a critical public health concern since this species has intrinsic resistance to several antimicrobials, including polymyxins. The identification of this multidrug-resistant (MDR) pathogen outside the hospital setting has become increasingly frequent, and raises an alert for the global health agencies, as they indicate a possible spread of such pathogens. Herein, we described three MDR P. rettgeri isolates carrying a diversity of antimicrobial resistance genes (ARGs) isolated from stool samples of swine and bovine in Brazil. Molecular analysis revealed that all isolates belonged to the same clone. The whole genome sequencing (WGS) of a representative isolate (PVR-188) was performed by MiSeq Illumina® platform, while the assembling and annotation was achieved using SPAdes and Prooka, respectively. The WGS analyses indicated the presence of ARGs that confer resistance to β-lactams (blaNDM-1, blaCTX-M-2), quinolones (qnrD1), aminoglycosides (aadA2, aadA1, aph(3′)-Via), phenicol (catB2), sulfonamides (sul1, sul2), and trimethoprim (dfrA12, dfrA1). The presence of three plasmid replicons (Col3M, IncQ1, and IncT) was detected, but no phage sequences were found. The phylogenetic analyses confirmed the genomic relationship of the PVR-188 with P. rettgeri isolates recovered from animals and humans in the USA and Malaysia. In conclusion, we report the occurrence of MDR P. rettgeri clone colonizing the gut microbiota of food-producing animals in Brazil, revealing the spread of this pathogen beyond hospital boundaries.

Enterobacterales include foodborne pathogens of importance to public health and are often targeted in food surveillance programs as both safety and hygiene indicators. Furthermore, Enterobacterales are important in the context of antimicrobial resistance dissemination, also impacting infection treatment efficacy. In this study, the prevalence and characteristics of Enterobacterales in UK retail foods were examined. From 110 retail food samples, 253 Enterobacterales were recovered, with 16S rRNA sequencing revealing a diverse species community, including enteropathogens; the most common were Proteus mirabilis and Escherichia coli (18% each). Antimicrobial resistance was common, with 160/253 (63%) isolates being resistant to at least 1 antimicrobial. Resistance to all tested antimicrobials was observed. Thirteen percent of isolates were multidrug resistant, including 2 isolates each resistant to 8 or 9 of 9 antimicrobials tested. Klebsiella isolates possessed relatively higher levels of antimicrobial resistance to other species. Hafnia, Kluyvera, and Proteus isolates produced significantly higher biofilm biomass than Klebsiella (p = 0.038, 0.028, and 0.042, respectively) or Escherichia (p = 0.001, 0.008, and 0.001, respectively). Simultaneous curli fimbriae and cellulose production was noted in 7% of isolates at 37 °C, but not at 15 °C. This research demonstrates a high diversity of Enterobacterales within UK retail foods, alongside notable antimicrobial resistance phenotypes in enteropathogenic species, highlighting the need for effective surveillance and interventions.

Fresh produce is usually consumed raw or minimally processed, making it a potential vehicle for the transmission of antimicrobial-resistant (AMR) microorganisms to humans. The objective of the study was to assess the occurrence of extended-spectrum ß-lactamase (ESBL)- and carbapenemase-producing Enterobacterales (ESBL-E and CPE), respectively, in 118 fresh herbs and 101 bagged salads collected at retail level in Switzerland and to characterize the isolates’ phenotypic and genotypic properties using culture-based methods and whole genome sequencing (WGS).Of the fresh herbs, 6/118 contained ESBL-E and 7/118 yielded CPE. Of the salads, 13/101 contained ESBL-E and 1/101 CPE. Antimicrobial susceptibility testing (AST) identified 9/29 isolates as multidrug-resistant (MDR).ESBL-E were Escherichia coli (n = 6), Klebsiella pneumoniae (n = 4) Enterobacter chuandaensis (n = 1), and Kluyvera spp. (n = 1) carrying ß-lactamase (bla) genes belonging to the cefotaximase-München (blaCTX-M)-groups, Proteus spp. (n = 1) containing Hôpital-Universitaire-de-Genève-bla (blahugA), Raoultella ornithinolytica (n = 1) carrying sulfhydryl reagent variable bla (blaSHV), and Serratia fonticola (n = 7) carrying S. fonticula bla (blaFONA) genes. CPE were Enterobacter asburiae (n = 1) E. cloacae (n = 6) and E. vonholyi (n = 1) carrying imipenemase bla (blaIMI) genes.Several K. pneumoniae sequence types (STs) were identified (ST967, ST628, ST219, and ST1823), which have been linked to human disease and nosocomial outbreaks. They carried blaCTX-M-15 on plasmids detected globally in environmental and clinical samples. E. coli (ST10, ST48, ST609, ST2040, ST6215 and ST3580) and enterotoxigenic E. coli (ETEC) ST2040 carrying blaCTX-M-15 were found. E. cloacae (ST820 and ST1516) with blaIMI-1 have been found previously in clinical settings and community outbreaks.The occurrence and consumption of fresh produce containing MDR ESBL-E and CPE pose substantial public health risks and raise significant food safety concerns.

Background: The coronavirus disease 2019 (COVID-19) pandemic has contributed to the spread of antimicrobial resistance, including carbapenem-resistant Enterobacterales. Methods: This study utilized data from the Study for Monitoring Antimicrobial Resistance Trends (SMART) surveillance program in Taiwan. Enterobacterales from patients with bloodstream infections (BSIs) were collected and subjected to antimicrobial susceptibility testing and β-lactamase gene detection using a multiplex PCR assay. Statistical analysis was conducted to compare susceptibility rates and resistance genes between time periods before (2018–2019) and during the COVID-19 pandemic (2020–2021). Results: A total of 1231 Enterobacterales isolates were collected, predominantly Escherichia coli (55.6%) and Klebsiella pneumoniae (29.2%). The proportion of nosocomial BSIs increased during the COVID-19 pandemic (55.5% vs. 61.7%, p

Objectives: To report trends in carbapenem resistance and difficult-to-treat resistance (DTR) among clinical isolates of Gram-negative priority pathogens collected by the ATLAS global surveillance program from 2018 to 2022. Methods: Reference broth microdilution testing was performed in a central laboratory for 79,214 Enterobacterales, 30,504 Pseudomonas aeruginosa, and 13,500 Acinetobacter baumannii-calcoaceticus complex isolates collected by a constant set of 157 medical centres in 49 countries in Asia Pacific (APAC), Europe (EUR), Latin America (LATAM), Middle East-Africa (MEA), and North America (NA) regions. MICs were interpreted by 2023 CLSI M100 breakpoints. β-lactamase genes were identified for meropenem-nonsusceptible (MIC ≥2 mg/L) Enterobacterales isolates. Results: Carbapenem-resistant Enterobacterales (CRE) detection increased (P < 0.05) in APAC, EUR, LATAM, and MEA regions and decreased in NA, while annual DTR percentages increased in all five regions. Carbapenem-resistant P. aeruginosa (CRPA; decreased in MEA region) and carbapenem-resistant A. baumannii-calcoaceticus complex (CRAB; decreased in MEA region and increased in EUR) remained relatively stable over time in all regions, although notably, annual percentages of CRAB and DTR A. baumannii-calcoaceticus complex isolates were consistently >25 percentage points lower in NA than in other regions. For all regions except NA, the majority of changes in CRE percentages could be attributed to hospital-acquired infections. Among meropenem-nonsusceptible Enterobacterales, KPC was the most frequent carbapenemase in NA and EUR each year. NDM was the most prevalent carbapenemase detected in 2022 in other global regions. Conclusion: CRE, CRPA, CRAB, and DTR rates vary among global regions over time highlighting the need for continuing surveillance to inform treatment strategies and antimicrobial stewardship.

ABSTRACT: Background: The rapid and global spread of Escherichia coli carrying mcr-type genes at the human-animal-environmental interface has become a serious global public health problem. Objective: To perform a genomic investigation of a colistin-resistant E. coli strain (14005RM) causing urinary tract infection, using a hybrid de novo assembly of Illumina/Nanopore sequence data, presenting phylogenomic insights into the relationship with mcr-1-positive strains circulating at the human-animal-environmental interface, in Brazil. Methods: Genomic DNA was sequenced using both the Illumina NexSeq and Nanopore MinION platforms. De novo hybrid assembly was performed by Unicycler. Genomic data were assessed by in silico prediction and bioinformatic tools. Results: The genome assembly size was 5 333 039 bp. The mcr-1.5-positive E. coli strain 14005RM belongs to the sequence type ST354 and presented a broad resistome (antibiotics, heavy metals, disinfectants, and glyphosate) and virulome. The mcr-1.5 gene was carried by an IncI2 plasmid (p14005RM, sizing 65,458 kb). Full genome SNP-based phylogenetic analysis reveals that mcr-1.5-producing E. coli strain 14005RM is highly related (> 98% identity) to colistin-resistant mcr-1.1-positive ST354 lineages associated with urinary tract infections in Brazil since 2015. Conclusion: Mobile colistin resistance within the Brazilian One Health microbiosphere is mediated by mcr gene variants propagated by IncX4, IncHI2, and IncI2 plasmids, circulating among global clones of E. coli.

Summary: Members of the genus Phytobacter (order Enterobacterales) are isolated from the natural environment and clinical settings. Identification of Phytobacter strains based on biochemical characteristics is complicated due to taxonomic confusion, and they are often misidentified by automated identification systems in laboratories.In this study we describe the first three clinical cases associated with Phytobacter spp. reported in Argentina. We describe the identification, the molecular analysis using whole genome sequencing and the potential clinical relevance.

Objectives: The rise in Carbapenem-resistant Enterobacterales (CRE) is perturbing. To curb the menace of CRE, a comprehensive understanding of its prevalence and epidemiology is crucial. As varying reports abound, the true prevalence of CRE in Nigeria remains unknown. Here, we conducted a systematic review and meta-analysis following standard guidelines to assess the situation of CRE in Nigeria. Methods: We searched electronic databases including Pubmed, ScienceDirect, Scopus, Web of Science, and Google Scholar for articles providing information on CRE in Nigeria. The data gathered were analyzed using OpenMeta Analyst and Comprehensive Meta-Analysis software. The random-effect model was employed to calculate pooled resistance to carbapenem antibiotics. Results: From 321 retrieved records, 57 were finally included. The studies were predominantly from the South-West region (n = 19). Escherichia coli and Klebsiella pneumoniae were the most frequently tested Enterobacterales among the included studies. The pooled prevalence estimate for imipenem resistance among CRE was 11.2 % (95 % CI: 7.9–15.7). Meropenem resistance had an estimate of 13.5 % (95 % CI: 9.1–19.6), whereas ertapenem and doripenem were estimated at 17.0 % (95 % CI: 9.9–27.7) and 37.9 % (95 % CI: 15.0–67.8), respectively. High heterogeneity (I2>85 %, p

Ceftolozane, a novel cephalosporin, combined with tazobactam, a known β-lactamase inhibitor, shows robust antipseudomonal activity, although it doesn't cover carbapenemases. Our review of data from 2012 to 2021 in Taiwan highlights TOL/TAZ's in-vitro performance.TOL/TAZ is most effective against Pseudomonas aeruginosa (91.3–94.4 % susceptible, with an MIC

ABSTRACT: Background: Carbapenemase-producing Citrobacter freundii has been reported as a leading cause of healthcare-associated infections. Particularly, C. freundii belonging to the sequence type (ST) 18 is considered to be an emerging nosocomial clone. Objectives: To report the genomic background and phylogenomic analysis of a multidrug-resistant NDM-1-producing C. freundii ST18 (strain CF135931) isolated from an endangered green sea turtle affected by plastic pollution in Brazil. Methods: Genomic DNA was extracted and sequenced using the Illumina NextSeq platform. De novo assembly was performed by CLC Workbench, and in silico analysis accomplished by bioinformatics tools. For phylogenomic analysis, publicly available C. freundii (txid:546) genome assemblies were retrieved from the NCBI database. Results: The genome size was calculated at 5 290 351 bp, comprising 5263 total genes, 4 rRNAs, 77 tRNAs, 11ncRNAs, and 176 pseudogenes. The strain belonged to C. freundii ST18, whereas resistome analysis predicted genes encoding resistance to β-lactams (blaNDM-1, blaOXA-1, blaCMY-117, and blaTEM-1C), aminoglycosides (aph(3′')-Ib, aadA16, aph(3′)-VI, aac(6′)-Ib-cr, and aph(6)-Id), quinolones (aac(6′)-Ib-cr), macrolides (mph(A) and erm(B)), sulphonamides (sul1 and sul2), tetracyclines (tetA and tetD), and trimethoprim (dfrA27). The phylogenomic analysis revealed that CF135931 strain is closely related to international human-associated ST18 clones producing NDM-1. Conclusion: Genomic surveillance efforts are necessary for robust monitoring of the emergence of drug-resistant strains and WHO critical priority pathogens within a One Health framework. In this regard, this draft genome and associated data can improve understanding of dissemination dynamics of nosocomial clones of carbapenemase-producing C. freundii beyond hospital walls. In fact, the emergence of NDM-1-producing C. freundii of global ST18 in wildlife deserves considerable attention.

Objectives: Metallo-β-lactamase (MBL)-producing Enterobacterales are a major challenge worldwide due to limited treatment options. Aztreonam-avibactam (ATM-AVI), which is under clinical development, has shown activity against MBL-positive isolates. This study evaluated the prevalence of MBL producers and the nature of enzymes among a global collection of clinical isolates of Enterobacterales from the Antimicrobial Testing Leadership and Surveillance program (ATLAS) surveillance program (2016–2020), and the antimicrobial activity of ATM-AVI and comparators against this collection. Methods: Non-duplicate clinical isolates of Enterobacterales (N = 106 686) collected across 63 countries were analysed. Antimicrobial susceptibility was performed using broth microdilution. Minimum inhibitory concentrations (MICs) were interpreted using Clinical and Laboratory Standards Institute and European Committee on Antimicrobial Susceptibility Testing breakpoints. Provisional pharmacokinetic/pharmacodynamic breakpoint of ≤8 mg/L was considered for ATM-AVI. β-lactamase genes were characterized by polymerase chain reaction and sequencing. The Cochran Armitage Trend test was used to determine significant trends in percentage of isolates over time. Results: Overall, MBL-positive isolates were 1.6% of total Enterobacterales isolates globally, with a significant increasing trend observed over time, globally and across regions (P < 0.05). New Delhi MBL (NDM) was the most common MBL (83.3%). ATM-AVI demonstrated potent activity against MBL-positive isolates (MIC ≤8 mg/L: 99.4% isolates inhibited; MIC90, 1 mg/L). Consistent activity was also noted across different regions. Potent activity was demonstrated against different NDM variants and MBL-positive isolates co-carrying other carbapenemases (98.1% and 99.7% isolates inhibited at ≤8 mg/L, respectively). About 0.6% MBL-positive isolates (10/1707) had MICs >8 mg/L for ATM-AVI. Conclusion: ATM-AVI demonstrated potent activity against MBL-positive isolates, including NDM variants and MBL-positive isolates co-carrying other carbapenemases, and may represent a good option for treating infections caused by MBL-positive Enterobacterales.

ABSTRACT: Background: The global spread of extended-spectrum β-lactamase (ESβL)-producing Escherichia coli has been considered a One Health issue that demands continuous genomic epidemiology surveillance in humans and non-human hosts. Objectives: To report the occurrence and genomic data of ESβL-producing E. coli strains isolated from South American llamas inhabiting a protected area with public access in the Andean Highlands of Peru. Methods: Two ESβL-producing E. coli strains (E. coli L1LB and L2BHI) were identified by MALDI-TOF. Genomic DNAs were extracted and sequenced using the Illumina NextSeq platform. De novo assembly was performed by CLC Genomic Workbench and in silico prediction was accomplished by curated bioinformatics tools. SNP-based phylogenomic analysis was performed using publicly available genomes of global E. coli ST10. Results: Escherichia coli L1LB generated a total of 4 000 11 and L2BHI a total of 4 002 54 paired-end reads of ca.164 × and ca. 157 ×, respectively. Both E. coli strains were assigned to serotype O8:H4, fimH41, and ST10. The blaCTX-M-65 ESβL gene, along with other medically important antimicrobial resistance genes, was predicted. Broad virulomes, including the presence of the astA gene, were confirmed. The phylogenomic analysis revealed that E. coli L1LB and L2BHI strains are closely related to isolates from companion animals and human hosts, as well as environmental strains, previously reported in North America, South America, Africa, and Asia. Conclusion: Presence of ESβL-producing E. coli ST10 in South American camelids with historical and cultural importance supports successful expansion of international clones of priority pathogens in natural areas with public access.

Summary: Background: The presence of carbapenemase-producing carbapenem-resistant Enterobacterales (CP-CRE) around the world is increasing, particularly in healthcare settings. Surveillance testing for plasmid-mediated carbapenemase genes is necessary to tracking CP-CRE infections. Aim: In the state of Ohio, surveillance of carbapenem-resistant Enterobacterales (CRE) began in 2018, and to the authors' knowledge data on these cases has not been published to date. This study analyzed data on CRE from a large teaching hospital in Ohio, and by the Ohio Department of Health Laboratory (ODHL). Methods: Carbapenemase production was detected using mCIM, and plasmid-mediated carbapenemase genes were detected using rtPCR. Data was collected on 344 standard-of-care isolates from a large teaching hospital in Ohio, including data collected from chart review. Deidentified surveillance data on 4,391 CRE isolates was provided by the ODHL. Statistical analysis was performed using binary logistic regression. Findings: While KPC was the most common carbapenemase gene (n=1590), NDM (n=98), VIM (n=10), IMP (n=39) and OXA-48 (n=35) were also detected in the isolates studied. Klebsiella pneumoniae and Enterobacter cloacae were the most common CRE, and carbapenemase genes were most commonly detected in K. pneumoniae. Inpatient hospital stays and long-term care were associated with CP-CRE and were more common in women. Conclusion: Surveillance data shows that CP-CRE are present in Ohio, most commonly in Klebsiella pneumoniae. A better understanding of the prevalence of CRE, plasmid-mediated carbapenemase genes present, and the populations affected are important when tracking the spread of disease. Further study and surveillance of carbapenem-resistant organisms can provide a better understanding of their prevalence in the state.

Summary: Klebsiella pneumoniae is among the World Health Organization's list of priority pathogens, notorious for its role in causing healthcare-associated infections and neonatal sepsis globally. Containment of K. pneumoniae transmission depends on the continued effectiveness of antimicrobials and of biocides used for topical antisepsis and surface disinfection. Klebsiella pneumoniae is known to disseminate antimicrobial resistance (AMR) through a large auxiliary genome made up of plasmids, transposons and integrons, enabling it to evade antimicrobial killing through the use of efflux systems and biofilm development. Because AMR mechanisms are also known to impart tolerance to biocides, AMR is frequently linked with biocide resistance (BR). However, despite extensive research on AMR, there is a gap in knowledge about BR and the extent to which AMR and BR mechanisms overlap remains debatable. The aim of this paper is to review and summarise the current knowledge on the determinants of BR in K. pneumoniae and highlight content areas that require further inquiry.

Summary: Background: The burden of antimicrobial resistance (AMR) has been estimated to be the highest in sub-Saharan Africa (SSA). The current study estimated the proportion of drug-resistant Enterobacterales causing infections in SSA children. Methods: We searched MEDLINE/PubMed, Embase and the Cochrane Library to identify retrospective and prospective studies published from 01/01/2005 to 01/06/2022 reporting AMR of Enterobacterales causing infections in sub-Saharan children (0–18 years old). Studies were excluded if they had unclear documentation of antimicrobial susceptibility testing methods or fewer than ten observations per bacteria. Data extraction and quality appraisal were conducted by two authors independently. The primary outcome was the proportion of Enterobacterales resistant to antibiotics commonly used in paediatrics. Proportions were combined across studies using mixed-effects logistic regression models per bacteria and per antibiotic. Between-study heterogeneity was assessed using the I2 statistic. The protocol was registered with PROSPERO (CRD42021260157). Findings: After screening 1111 records, 122 relevant studies were included, providing data on more than 30,000 blood, urine and stool isolates. Escherichia coli and Klebsiella spp. were the predominant species, both presenting high proportions of resistance to third-generation cephalosporins, especially in blood cultures: 40.6% (95% CI: 27.7%–55%; I2: 85.7%, number of isolates (n): 1032) and 84.9% (72.8%–92.2%; I2: 94.1%, n: 2067), respectively. High proportions of resistance to other commonly used antibiotics were also observed. E. coli had high proportions of resistance, especially for ampicillin (92.5%; 95% CI: 76.4%–97.9%; I2: 89.8%, n: 888) and gentamicin (42.7%; 95% CI: 30%–56.5%; I2: 71.9%, n: 968). Gentamicin-resistant Klebsiella spp. were also frequently reported (77.6%; 95% CI: 65.5%–86.3%; I2: 91.6%, n: 1886). Interpretation: High proportions of resistance to antibiotics commonly used for empirical treatment of infectious syndromes were found for Enterobacterales in sub-Saharan children. There is a critical need to better identify local patterns of AMR to inform and update clinical guidelines for better treatment outcomes. Funding: No funding was received.

ABSTRACT: Objectives: Ceftazidime-avibactam (CAZ-AVI) combines ceftazidime and a reversible β-lactamase inhibitor that has shown activity against multidrug-resistant (MDR) Enterobacterales and P. aeruginosa. Using data from the Antimicrobial Testing Leadership and Surveillance program (ATLAS), this study examined the in vitro antimicrobial activity of CAZ-AVI and other antibiotics against Gram-negative bacteria collected from Chilean hospitals between 2015 and 2021. Methods: Clinical isolates of Enterobacterales and P. aeruginosa were collected from three medical centres in Chile. Blood, abdominal fluid, urine, soft tissues, and respiratory tract samples were obtained from infected patients. Minimum inhibitory concentrations using the broth microdilution method were determined for susceptibility testing, and the Clinical and Laboratory Standards Institute (CLSI) breakpoints were used for interpreting the results. Extended-spectrum β-lactamases (ESBL) and carbapenemase genes were also detected through polymerase chain reaction. Results: A total of 2600 Enterobacterales and 836 P. aeruginosa were analysed. CAZ-AVI was the antibiotic with the highest in vitro activity against Enterobacterales (99.72%). The incidence of carbapenem-resistant Enterobacterales (CRE) was 1.5% (n = 39), and the antibiotics with the best in vitro activity were tigecycline (92.31%), CAZ-AVI (88.57%), and amikacin (79.49%). CAZ-AVI was the antibiotic with the best activity against ESBL-producing Enterobacterales (99.34%) and MDR Enterobacterales (99.31%). For KPC-producing Enterobacterales, susceptibility to amikacin was 100%, whereas susceptibility to CAZ-AVI was 91.67%. Regarding MDR and difficult-to-treat resistance P. aeruginosa, 44.83% and 38.99% were susceptible to CAZ-AVI, respectively. Conclusion: CAZ-AVI shows excellent in vitro activity against Enterobacterales in general, CRE, ESBL-producing Enterobacterales, and KPC-producing Enterobacterales. CAZ-AVI is also an option against MDR P. aeruginosa.

ABSTRACT: Objectives: To report the in vitro susceptibility of Enterobacterales (n = 3905) and Pseudomonas aeruginosa (n = 1,109) isolates, collected from patients in sub-Saharan Africa (four countries) in 2017–2021, to a panel of 10 antimicrobial agents with a focus on ceftazidime-avibactam activity against resistant phenotypes and β-lactamase carriers. Methods: MICs were determined by CLSI broth microdilution and interpreted using both 2022 CLSI and EUCAST breakpoints. β-lactamase genes were identified in select β-lactam-nonsusceptible isolate subsets using multiplex PCR assays. Results: Among Enterobacterales, 96.2% of all isolates were ceftazidime-avibactam-susceptible (MIC90, 0.5 µg/mL), including all serine carbapenemase-positive (n = 127), 99.6% of ESBL-positive, carbapenemase-negative (n = 730), 91.9% of multidrug resistant (MDR; n = 1817), and 42.7% of DTR (difficult-to-treat resistance; n = 171) isolates. Metallo-β-lactamase (MBL) genes were identified in most (n = 136; 91.2%) ceftazidime-avibactam-resistant isolates (3.5% of all Enterobacterales isolates). Ceftazidime-avibactam percent susceptible values ranged from 99.5% (Klebsiella species other than Klebsiella pneumoniae) to 92.5% (K. pneumoniae) for the various Enterobacterial taxa examined. Greater than 90% of Enterobacterales isolates from each country (Cameroon, Ivory Coast, Nigeria, South Africa) were ceftazidime-avibactam-susceptible. Among P. aeruginosa, 88.9% of all isolates were ceftazidime-avibactam-susceptible (MIC90, 16 µg/mL). Most (88.5%) MBL-negative, meropenem-resistant (n = 78), 68.1% of MDR (n = 385), and 19.2% of DTR isolates (n = 99) were ceftazidime-avibactam-susceptible. MBL genes were identified in 43.1% of ceftazidime-avibactam-resistant isolates (n = 53; 4.8% of all P. aeruginosa isolates). Country-specific ceftazidime-avibactam percent susceptible values for P. aeruginosa ranged from 94.1% (Cameroon) to 76.2% (Nigeria). Conclusion: Reference in vitro antimicrobial susceptibility testing demonstrated that most recent Enterobacterales (96%) and P. aeruginosa (89%) clinical isolates from four sub-Saharan African countries were ceftazidime-avibactam susceptible.

ABSTRACT: Objectives: Antibiotic resistance (AR) is a global health issue with multidimensional repercussions. There is a paucity of data regarding the molecular epidemiology of multidrug-resistant Enterobacterales (MDR-E) and extended-spectrum ß-lactamase-producing Enterobacterales (ESBL-PE) in Africa, especially among people living with HIV (PLHIV). This study aimed at determining the prevalence, risk factors, phenotypic and genotypic profiles of MDR-E and ESBL-PE isolated from PLHIV in Yaoundé, Cameroon. Methods: In total, samples were collected from 185 PLHIV during a three-month period (April–June 2021) at the Yaoundé Central Hospital. Stool samples and rectal swabs were collected and cultured on MacConkey agar. The API 20E kit was used for the phenotypic identification of the isolates, whereas antibiotic susceptibility testing was performed using the Kirby-Bauer disk diffusion method. The ß-lactamase genes and genotypic relatedness were studied by PCR and ERIC-PCR, respectively. Results: The prevalence of MDR-E among PLHIV was 81%, of which 39% were ESBL-PE. A high level of resistance to fosfomycin (89%), chloramphenicol (63%), and gentamicin (56%) was observed. Escherichia coli was the predominant MDR non-ESBL-PE (80.8%) and MDR ESBL-PE (73.77%). The principal ß-lactamases genes in MDR non-ESBL and MDR ESBL-PE were blaTEM (62.90%) and blaCTX-M (40.86%), respectively. Genetic fingerprinting revealed high genetic relatedness among E. coli isolates. Conclusion: This study shows a high prevalence of MDR-E and ESBL-PE in the gut of PLHIV in Yaoundé, with blaTEM and blaCTX-M being the most prevalent. It demonstrates the need to strengthen real-time surveillance of these resistant bacteria in order to improve management of infection among PLHIV.

Background: Worldwide observations revealed increased frequencies of multi-resistant Enterobacterales and resistance genes in hospital wastewater compared to any other type of wastewater. Despite the description of clonal lineages possibly adapted to hospital wastewater, little is known about long term persistence as well as evolution of these lineages. Methods: In this study, wastewater isolates of different Enterobacterales species from a tertiary care hospital were investigated with 2.5 years distance. Whole Genome Sequencing (WGS) and resistance gene identification were performed for E. coli, C. freundii, S. marcescens, K. pneumoniae, K. oxytoca, and E. cloacae isolates (n = 59), isolated in 2022 and compared with strains isolated from the same wastewater pipeline in 2019 (n = 240). Results: Individual clonal lineages with highly related isolates could be identified in all species identified more than once in 2022 that appear to persist in the wastewater drainage. A common motif of all persistent clonal lineages was the carriage of mobile genetic elements encoding carbapenemase genes with hints for horizontal gene transfer in persistent clones in this environment observed over the 2.5-year period. Multiple plasmid replicons could be detected in both years. In 2022 isolates blaVIM-1 replaced blaOXA-48 as the most common carbapenemase gene compared to 2019. Interestingly, despite a similar abundance of carbapenemase genes (>80% of all isolates) at both time points genes encoding extended spectrum β-lactamases decreased over time. Conclusions: This data indicates that hospital wastewater continuously releases genes encoding carbapenemases to the urban wastewater system. The evolution of the resident clones as well as the reasons for the selection advantage in this specific ecological niche needs to be further investigated in the future.

Among the most problematic bacteria with clinical relevance are the carbapenem-resistant Enterobacterales (CRE), as there are very limited options for their treatment. Treated wastewater can be a route for the release of these bacteria into the environment and the population. The aim of this study was to isolate CRE from treated wastewater from the Zagreb wastewater treatment plant and to determine their phenotypic and genomic characteristics. A total of 200 suspected CRE were isolated, 148 of which were confirmed as Enterobacterales by MALDI-TOF MS. The predominant species was Klebsiella spp. (n = 47), followed by Citrobacter spp. (n = 40) and Enterobacter cloacae complex (cplx.) (n = 35). All 148 isolates were carbapenemase producers with a multidrug-resistant phenotype. Using multi-locus sequence typing and whole-genome sequencing (WGS), 18 different sequence types were identified among these isolates, 14 of which were associated with human-associated clones. The virulence gene analysis of the sequenced Klebsiella isolates (n = 7) revealed their potential pathogenicity. PCR and WGS showed that the most frequent carbapenemase genes in K. pneumoniae were blaOXA-48 and blaNDM-1, which frequently occurred together, while blaKPC-2 together with blaNDM-1 was mainly detected in K. oxytoca, E. cloacae cplx. and Citrobacter spp. Colistin resistance was observed in 40% of Klebsiella and 57% of Enterobacter isolates. Underlying mechanisms identified by WGS include known and potentially novel intrinsic mechanisms (point mutations in the pmrA/B, phoP/Q, mgrB and crrB genes) and acquired mechanisms (mcr-4.3 gene). The mcr-4.3 gene was identified for the first time in K. pneumoniae and is probably located on the conjugative IncHI1B plasmid. In addition, WGS analysis of 13 isolates revealed various virulence genes and resistance genes to other clinically relevant antibiotics as well as different plasmids possibly associated with carbapenemase genes. Our study demonstrates the important role that treated municipal wastewater plays in harboring and spreading enterobacterial pathogens that are resistant to last-resort antibiotics.

Background: Over the past few decades, antimicrobial resistance (AMR) has emerged as a global health challenge in human and veterinary medicine. Research on AMR genes in captive wild animals has increased. However, the presence and molecular characteristics of tet(X)-carrying bacteria in these animals remain unknown. Methods: Eighty-four samples were collected from captive wild animals. tet(X) variants were detected using polymerase chain reaction and the isolates were identified using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. All isolated strains were subjected to antimicrobial susceptibility testing and whole-genome sequencing. The virulence of an Escherichia coli strain carrying enterotoxin genes was assessed using a Galleria mellonella larval model. Results: We isolated two tet(X4)-positive E. coli strains and one tet(X4)-positive Raoultella ornithinolytica strain. Antimicrobial susceptibility tests revealed that all three tet(X4)-carrying bacteria were sensitive to the 13 tested antimicrobial agents, but exhibited resistance to tigecycline. Notably, one tet(X4)-carrying E. coli strain producing an enterotoxin had a toxic effect on G. mellonella larvae. Whole-genome sequencing analysis showed that the two tet(X4)-carrying E. coli strains had more than 95% similarity to tet(X4)-containing E. coli strains isolated from pigs and humans in China. Conclusion: The genetic environment of tet(X4) closely resembled that of the plasmid described in previous studies. Our study identified tet(X4)-positive strains in wildlife and provided valuable epidemiological data for monitoring drug resistance. The identification of enterotoxin-producing E. coli strains also highlights the potential risks posed by virulence genes.

Background: Presently, community-associated carbapenemase-producing Enterobacterales (CPE) remains largely unknown and require public attention. This study aimed to investigate the presence of CPE from outpatients in Thailand. Methods: Non-duplicate stool (n = 886) and urine (n = 289) samples were collected from outpatients with diarrhea and urinary tract infection, respectively. Demographic data and characteristics of patients were collected. Isolation of CPE was performed by plating enrichment culture on agar supplemented with meropenem. Carbapenemase genes were screened by PCR and sequencing. CPE isolates were phenotypically and genotypically characterized. Results: Fifteen samples (1.3%, 14 stool and 1 urine) yielded blaNDM-1-positive carbapenemase-producing Klebsiella pneumoniae (CPKP). Additional resistance to colistin and tigecycline was observed in 53.3% and 46.7% of isolates, respectively. Age >60 years was identified as a risk factor for patients with CPKP (P

ABSTRACT: Objectives: The aim of this study was to perform a genomic investigation of a multiple fluoroquinolone-resistant Leclercia adecarboxylata strain isolated from a synanthropic pigeon in São Paulo, Brazil. Methods: Whole-genome sequencing was performed using an Illumina platform, and in silico deep analyses of the resistome were performed. Comparative phylogenomics was conducted using a global collection of publicly available genomes of L. adecarboxylata strains isolated from human and animal hosts. Results: L. adecarboxylata strain P62P1 displayed resistance to human (norfloxacin, ofloxacin, ciprofloxacin, and levofloxacin) and veterinary (enrofloxacin) fluoroquinolones. This multiple quinolone-resistant profile was associated with mutations in the gyrA (S83I) and parC (S80I) genes and the presence of the qnrS gene within an ISKpn19-orf-qnrS1-ΔIS3-blaLAP-2 module, previously identified in L. adecarboxylata strains isolated from pig feed and faeces in China. Genes associated with arsenic, silver, copper, and mercury resistance were also predicted. Phylogenomic analysis revealed clustering (378–496 single nucleotide polymorphism differences) with two L. adecarboxylata strains isolated from human and fish sources in China and Portugal, respectively. Conclusions: L. adecarboxylata is a Gram-negative bacterium of the Enterobacterales order and is considered an emergent opportunistic pathogen. Since L. adecarboxylata has adapted to human and animal hosts, genomic surveillance is highly recommended, in order to identify the emergence and spread of resistant lineages and high-risk clones. In this regard, this study provides genomic data that can help clarify the role of synanthropic animals in the dissemination of clinically relevant L. adecarboxylata within a One Health context.

ABSTRACT: Objectives: The occurrence of extended-spectrum ß-lactamase (ESBL)-producing Enterobacterales in broilers represents a risk to public health because of the possibility of transmission of ESBL producers and/or blaESBL genes via the food chain or within settings where human-animal interfaces exist. Methods: This study assessed the occurrence of ESBL producers among faecal samples of broilers at slaughter. Isolates were characterised by multilocus sequence typing, antimicrobial susceptibility testing, and whole-genome sequencing. Results: The flock prevalence, determined by sampling crates of 100 poultry flocks, was 21%. The predominant blaESBL gene was blaSHV-12, identified in 92% of the isolates. A variety of Escherichia coli and Klebsiella pneumoniae sequence types (STs) were identified, including extraintestinal pathogenic E. coli ST38, avian pathogenic E. coli ST10, ST93, ST117, and ST155, and nosocomial outbreak clone K. pneumoniae ST20. Whole-genome sequencing was used to characterise a subset of 15 isolates, including 6 E. coli, 4 K. pneumoniae, 1 Klebsiella grimontii, 1 Klebsiella michiganensis, 1 Klebsiella variicola, and 1 Atlantibacter subterranea. Fourteen isolates carried identical or closely related 46338–54929 bp IncX3 plasmids encoding blaSHV-12 and qnrS1. One E. coli isolate carried a 46338 bp IncX3 plasmid, which was integrated chromosomally into ydbD. Conclusions: The blaSHV-12 gene has replaced the previously predominant blaCTX-M-1 in ESBL-producing Enterobacterales from broilers in Switzerland. Broilers may play a role in the dissemination of blaSHV-12 and qnrS1 associated with epidemic IncX3 plasmids, representing a risk to human and animal health.

ABSTRACT: Objective: Both ertapenem and other carbapenems, including imipenem, meropenem, and doripenem, are recommended in the treatment of extended-spectrum-β-lactamase (ESBL)-producing Enterobacterales infection. However, whether ertapenem is as effective as other carbapenems for ESBL-producing Enterobacterales remains unclear. Therefore, this meta-analysis was conducted to compare the clinical efficacy of ertapenem versus other carbapenems in the treatment of ESBL-producing Enterobacterales infection. Methods: PubMed, Web of Science, and Cochrane Library were searched from their inception to 29 November 2022. Only studies comparing ertapenem and other carbapenems in the treatment of patients with ESBL-producing Enterobacterales infections were included. Results: A total of six studies meeting selection criteria were identified. Overall, ertapenem was associated with a significantly lower 30-d mortality when compared with other carbapenems (10.7% [46/431] vs. 17.7% [104/586]; risk ratio [RR], 0.61; 95% CI: 0.40–0.91). The ertapenem group exhibited a significantly shorter length of hospital stay than the other carbapenem groups (mean differences, -6.02 d; 95% CI, -9.39 to -2.64). No significant differences were noted between ertapenem and other carbapenem groups in terms of rates of clinical cure or improvement (RR, 1.11; 95% CI: 0.97–1.25) and microbiological eradication (RR, 1.01; 95% CI: 0.97–1.06). Conclusions: Ertapenem could be as effective as other carbapenems in the treatment of patients with ESBL-producing Enterobacterales infections.

Background: New tetracycline derivatives exhibit broad-spectrum antimicrobial activities. This study aimed to assess the in vitro activity of eravacycline against common Enterobacterales. Methods: Clinical Enterobacterales isolates were collected between 2017 and 2021. The minimum inhibitory concentration (MIC) was determined using a broth microdilution test. Results: We identified Klebsiella pneumoniae (n = 300), Escherichia coli (n = 300), Klebsiella oxytoca (n = 100), Enterobacter cloacae complex (n = 100), Citrobacter freundii (n = 100), and Proteus mirabilis (n = 100). All P. mirabilis strains were resistant to eravacycline. Excluding P. mirabilis, the susceptibility rates to eravacycline, omadacycline, and tigecycline were 75.2%, 66.9%, and 73%, respectively. The MIC50 and MIC90 (mg/L) of eravacycline were 0.5 and 4 for K. pneumoniae, 0.5 and 1 for E. coli, 0.5 and 1 for K. oxytoca, 0.5 and 2 for E. cloacae complex, and 0.25 and 1 for C. freundii. In cefotaxime non-susceptible and meropenem susceptible Enterobacterales, excluding P. mirabilis, the susceptibility rates of eravacycline, omadacycline, and tigecycline were 69.7%, 57.1%, and 66.2%. We found decreased susceptibility rates of three new tetracycline derivatives against meropenem non-susceptible Enterobacterales (eravacycline: 47.1%, omadacycline: 39.4%, and tigecycline: 39.4%). Eravacycline showed a high susceptibility rate against cefotaxime non-susceptible and meropenem susceptible K. oxytoca (100%), C. freundii (93.2%), E. coli (85.9%), and meropenem non-susceptible E. coli (100%). Conclusion: This study provides the MIC and susceptibility rate of eravacycline for common Enterobacterales. Eravacycline could be a therapeutic choice for cefotaxime non-susceptible or meropenem non-susceptible Enterobacterales, especially K. oxytoca, C. freundii, and E. coli.

The spread of carbapenemase-producing Klebsiella pneumoniae beyond hospital settings is a global critical issue within a public health and One Health perspective. Another worrisome concern is the convergence of virulence and resistance in healthcare-associated lineages of K. pneumoniae leading to unfavorable clinical outcomes. During a surveillance study of WHO critical priority pathogens circulating in an impacted urban river in São Paulo, Brazil, we isolate two hypermucoviscous and multidrug-resistant K. pneumoniae strains (PINH-4250 and PINH-4900) from two different locations near to medical centers. Genomic investigation revealed that both strains belonged to the global high-risk sequence type (ST) ST11, carrying the blaKPC-2 carbapenemase gene, besides other medically important antimicrobial resistance determinants. A broad virulome was predicted and associated with hypervirulent behavior in the Galleria mellonella infection model. Comparative phylogenomic analysis of PINH-4250 and PINH-4900 along to an international collection of publicly available genomes of K. pneumoniae ST11 revealed that both environmental strains were closely related to hospital-associated K. pneumoniae strains recovered from clinical samples between 2006 and 2018, in São Paulo city. Our findings support that healthcare-associated KPC-2-positive K. pneumoniae of ST11 clone has successfully expanded beyond hospital settings. In summary, aquatic environments can become potential sources of international clones of K. pneumoniae displaying carbapenem resistance and hypervirulent behaviors, which is a critical issue within a One Health perspective.

Introdução/Objetivo: As Infecções de Corrente Sanguínea (ICS) causadas por Enterobacterales são eventos frequentes e relacionados à elevada morbimortalidade, principalmente quando causadas por isolados resistentes aos carbapenêmicos. No Brasil, o primeiro relato de Enterobacterales produtores de Carbapenemases (ERC) ocorreu em 2005, enquanto a detecção da carbapenemase do tipo KPC aconteceu pela primeira vez em 2009. A partir de então, outros relatos surgiram no país, mas para alguns estados brasileiros os dados ainda são escassos, como é o caso da Bahia. Assim, o objetivo deste trabalho é descrever características epidemiológicas e clínicas de pacientes com ICS por Enterobacterales, identificando as principais espécies isoladas e seu perfil de sensibilidade antimicrobiana. Métodos: Foi realizado um estudo do tipo corte transversal, sendo incluídos casos de ICS por Enterobacterales provenientes de dois hospitais terciários de Salvador-Bahia, entre 01/ 2016 e 12/2018. Dados epidemiológicos foram coletados através de revisão de prontuários e os isolados foram identificados com a tecnologia do MALDI-TOF® e pelo VITEK-2®. O perfil de sensibilidade antimicrobiana foi realizado através do sistema automatizado (VITEK-2®). Análises estatísticas descritivas e univariadas foram efetuadas através do programa Epi Info v 3.5.1. Resultados: Foram identificados 252 casos de ICS, dos quais 14,3% dos isolados foram resistentes a carbapenêmicos. Escherichia coli e Klebsiella pneumoniae foram as espécies mais frequentes, sendo todos (n=119) os isolados de E. coli sensíveis aos carbapenêmicos e à tigeciclina. A sensibilidade às cefalosporinas variou entre 72% e 76,6%. Entre os isolados de K. pneumoniae, a sensibilidade aos carbapenêmicos foi de 66,7–69,7% e a resistência à colistina foi identificada em 14 isolados. Resistência aos carbapenêmicos não foi identificada nas infecções da comunidade, enquanto 18,7% (n=26) dos isolados nas infecções relacionadas à assistência à saúde foram resistentes a estas drogas. A mortalidade geral foi de 37,3%, chegando a 75% no grupo de ICS por ERC. Conclusões: Diante dos achados, conclui-se que as ICS causadas Enterobacterales apresentam elevada letalidade, sobretudo quando causadas por ERC, sendo a compreensão de sua epidemiologia fundamental para o estabelecimento de melhores estratégias terapêuticas e na adoção de medidas preventivas.

Introdução: O objetivo deste estudo foi descrever o aumento de isolados clínicos de Enterobacterales produtoras de dupla carbapenemases (NDM e KPC) recuperados de pacientes internados em 22 centros de saúde, distribuídos em cinco estados brasileiros, durante e após o período da pandemia de COVID-19. Métodos: Entre julho de 2020 a junho de 2023, o setor de microbiologia clínica do laboratório detectou isolados de CRE com triagem positiva para dupla carbapenemase. A identificação bacteriana foi realizada por espectrometria de massa e a concentração inibitória mínima de antibióticos foi determinada por sistema automatizado, exceto para polimixina B, o qual foi utilizado microdiluição em caldo. A detecção dos genes de carbapenemases (blaKPC, blaNDM, blaIMP, blaVIM, blaGES e blaOXA-48-like) foi realizada por PCR. A relação genética das cepas foi caracterizada pelo método de PFGE. Resultados: Foram identificadas 40 enterobactérias produtoras de dupla carbapenemase nos anos 2020 (n= 3), 2021 (n= 05), 2022 (n=16) e 2023 (n=16). Klebsiella pneumoniae (KPN) foi o agente mais frequente (67,5%), seguido por Proteus mirabilis (12,5%), Klebsiella oxytoca (10%), Escherichia coli (5%), Klebsiella aerogenes (2,5%) e Citrobacter koseri (2,5%). Esses microrganismos foram isolados de urina (n=13), sangue (n=8), swab retal (n=8), ponta de cateter (n=4), pele e partes moles (n=3), secreção abdominal (n=2) e aspirado traqueal (n= 2). Em todos eles foram detectados níveis elevados de resistência a amoxicilina-ácido clavulânico, ceftazidima, ceftriaxona, cefepima, ertapenem, meropenem, aztreonam e ceftazidima-avibactam. Entre as KPN, 15%, 37% e 77% também foram resistentes à polimixina B, amicacina e gentamicina, respectivamente. Todas as cepas de Enterobacterales abrigaram as enzimas NDM-1 e KPC-2. A análise PFGE mostrou 11 clusters (padrão A à K) entre 16 cepas KPN isoladas de 14 hospitais. O mesmo clone (padrão A) foi encontrado em cinco cepas de KPN isoladas de um mesmo hospital. Para cepas de K. oxytoca, quatro padrões de PFGE foram encontrados em diferentes hospitais. Conclusão: O aumento de cepas de Enterobacterales coprodutoras de carbapenemases representam um grande desafio, não só pelas limitadas opções de tratamento, mas pela dificuldade de detecção por um laboratório de rotina. Esses achados podem estar subestimados, considerando tais dificuldades.

Introdução/objetivo: A pandemia de COVID-19 acelerou o avanço da resistência bacteriana devido as altas taxas de prescrições de antibióticos, internações prolongadas em UTIs, uso de dispositivos invasivos, além de falhas na aplicação de medidas de prevenção e controle de infecções em ambientes hospitalares. O objetivo desse trabalho foi avaliar o impacto da pandemia de COVID-19 no aumento de Enterobacterales Produtoras de Carbapenemases (EPC), em isolados clínicos de um hospital de alta complexidade de Salvador-BA. Métodos: No período de 2019 a 2022, foram avaliados 566 isolados consecutivos, não duplicados de EPC. A identificação dos isolados foi realizada pelo pelo Maldi-tof (Vitek-MS, bioMérieux) e a detecção de carbapenemases foi realizada com testes imunocromatogáficos O.K.N RESIST-3® ou NG-TEST CARBA-5®. Resultados: Do total de amostras analisadas, a maioria dos isolados foi proveniente de infecções do trato urinário (36%) e corrente sanguínea (26%), sendo K. pneumoniae o microrganismo de maior prevalência (73%). No ano de 2019 foram obtidos 115 isolados, sendo 81 produtores de KPC (70%), 29 NDM (25%), 4 KPC/NDM (3%) e 1 KPC/OXA-48 (1%). Em 2020 foram detectados 156 isolados, sendo 82 KPC (53%), 56 NDM (36%), 16 KPC/NDM (10%) e 2 KPC/OXA-48 (1%), já em 2021, foram 172 isolados, sendo 92 KPC (53%), 65 NDM (38%) e 15 KPC/NDM (9%). Em 2022 foram obtidos 123 isolados, sendo 70 KPC (57%), 49 NDM (40%) e 4 KPC/NDM (3%), se aproximando da detecção observada no período pré-pandêmico (2019). Comparando os períodos pré-pandêmico e pandêmico, foi observado um aumento de 36% na detecção de EPC entre 2019 e 2020 e de 10% entre 2020 e 2021. Já entre os anos de 2021 e 2022 observou-se uma queda de 28% na detecção. A frequência de KPC apresentou uma diminuição em 2022 em relação ao ano de 2019, caindo de 70% para 57%. Em contrapartida, a frequência de NDM aumentou de 25% em 2019 para 40% em 2022. Conclusão: A detecção de EPC aumentou substancialmente no período pandêmico. Com o maior controle da pandemia após a vacinação e adoção de medidas para minimizar as Infecções Relacionadas à Assistência à Saúde (IRAS), foi possível reduzir a detecção de EPC no ano de 2022, se aproximando aos índices observados em 2019. Entretanto, o aumento de NDM em relação as outras carbapenemases representa um alerta de saúde pública, devido às limitações terapêuticas para o tratamento de pacientes acometidos por essas infecções.

Inadequate cleaning and disinfection (C&D) in slaughterhouses can cause bacterial contamination of meat, resulting in foodborne disease and reduced meat quality. Different methods for monitoring the efficacy of C&D procedures are available, but few studies have assessed their reliability. This study examined C&D efficacy in slaughterhouses and evaluated the diagnostic performance of methods for measuring surface hygiene.One red meat and one poultry slaughterhouse in Sweden were each visited on six occasions before and six occasions after C&D. Sampling points were sampled with: swabbing and plating for total aerobic bacteria (TAB) and Enterobacterales (EB); dipslides for total viable count; and ATP-bioluminescence tests. To evaluate the diagnostic performance of the dipslide and ATP-bioluminescence methods, the results were compared with (TAB) as a reference.In total, 626 samples were collected. For the majority of samples, TAB was lower after than before C&D and EB were mainly detected before C&D, indicating C&D efficacy. Greater reductions in mean TAB were observed in processing areas (2.2 and 2.8 log CFU/100 cm2 in red meat and poultry slaughterhouse, respectively) than in slaughter areas (1.3 log CFU/100 cm2 in both slaughterhouses). Approximately half of all samples were assessed as non acceptably clean (52% for red meat and 46% for poultry slaughterhouse) according to previously published thresholds. Critical food contact surfaces that were insufficiently cleaned and disinfected were plucking fingers, shackles, and a post-dehairing table. Cleaning and disinfection of drains and floors were inadequate.The ATP-bioluminescence method showed low specificity compared with the reference (TAB) in both the red meat (0.30) and poultry slaughterhouses (0.64). The sensitivity of dipslides was low (0.26) in the red meat slaughterhouse compared with TAB. A combination of ATP-bioluminescence and dipslides could provide more accurate estimates of C&D efficacy.

ABSTRACT: Objectives: Ceftolozane-tazobactam (C/T) proved its efficacy for the treatment of infections caused by non-carbapenemase producing Pseudomonas aeruginosa and Enterobacterales. Here, we aimed to provide susceptibility data on a large series of Enterobacterales since the revision of EUCAST categorization breakpoints in 2020. Methods: First, C/T susceptibility was determined on characterized Enterobacterales resistant to third generation cephalosporins (3GCs) (extended spectrum β-lactamase [ESBL] production or different levels of AmpC overexpression) (n = 213) and carbapenem-resistant Enterobacterales (CRE) (n = 259), including 170 carbapenemase producers (CPE). Then, 1632 consecutive clinical Enterobacterales responsible for infection were prospectively collected in 23 French hospitals. C/T susceptibility was determined by E-test® (biomérieux) and broth microdilution (BMD) (Sensititre™, Thermo Scientific) to perform method comparison. Results: Within the collection isolates, 88% of 3GC resistant strains were susceptible to C/T, with important variation depending on the resistance mechanism: 93% vs. 13% susceptibility for CTX-M and SHV-ESBL producers, respectively. Only 20% of the CRE were susceptible to C/T. Among CPE, 80% of OXA-48-like producers were susceptible to C/T, whereas all metallo-β-lactamase producers were resistant. The prospective study revealed that 95.6% of clinical isolates were susceptible to C/T. Method comparison performed on these 1632 clinical isolates demonstrated 99% of categorization agreement between MIC to C/T determined by E-test® in comparison with the BMD (reference) and only 74% of essential agreement. Conclusion: Overall, C/T showed good activity against wild-type Enterobacterales, AmpC producers, and ESBL-producing Escherichia coli but is less active against ESBL-producing Klebsiella pneumoniae, and CRE. E-test® led to an underestimation of the MICs in comparison to the BMD reference.

Objectives: To assess whether having a pet in the home is a risk factor for community-acquired urinary tract infections associated with extended-spectrum β-lactamase (ESBL)- or AmpC β-lactamase (ACBL)- producing Enterobacterales. Methods: An unmatched case-control study was conducted between August 2015 and September 2017. Cases (n = 141) were people with community-acquired urinary tract infection (UTI) caused by ESBL- or ACBL-producing Enterobacterales. Controls (n = 525) were recruited from the community. A telephone questionnaire on pet ownership and other factors was administered, and associations were assessed using logistic regression. Results: Pet ownership was not associated with ESBL- or ACBL-producing Enterobacterales-related human UTIs. A positive association was observed for recent antimicrobial treatment, travel to Asia in the previous year, and a doctor's visit in the last 6 months. Among isolates with an ESBL-/ACBL-producing phenotype, 126/134 (94%) were Escherichia coli, with sequence type 131 being the most common (47/126). Conclusions: Companion animals in the home were not found to be associated with ESBL- or ACBL-producing Enterobacterales-related community-acquired UTIs in New Zealand. Risk factors included overseas travel, recent antibiotic use, and doctor visits.

Background: Multi-drug resistant Enterobacterales is a growing health threat. Imipenem/relebactam and meropenem/vaborbactam, are not clinically used in Taiwan and the susceptibility is lack from routine laboratory tests. Methods: Broth microdilution method was used to determine the minimum inhibitory concentrations (MICs) and interpreted according to the Clinical and Laboratory Standards Institute (CLSI) breakpoints criteria. Isolates that were not susceptible to imipenem (MIC ≥ 2 mg/L), imipenem/relebactam (MIC ≥ 2 mg/L), or ceftolozane-tazobactam (MIC ≥ 4 mg/L) were selected for further molecular testing for genes encoding extended-spectrum beta-lactamases (ESBLs), AmpC β-lactamases, and carbapenemases by multiplex PCR assays. Results: A total of 290 Enterobacterales isolates from 9 participating hospitals were collected in 2020. Escherichia coli (n = 135, 46.6%) and Klebsiella pneumoniae (n = 88, 30.3%) were two leading pathogens of all Enterobacterales isolates. The antimicrobial agents with susceptibility rates more than 90% included amikacin (99.3%, 288/290), ertapenem (90.0%, 261/290), meropenem (97.2%, 282/290), imipenem/relebactam (94.8%, 275/290) and meropenem/vaborbactam (99.3%, 288/290). K. pneumoniae isolates were less susceptible to ertapenem, imipenem, meropenem, piperacillin-tazobactam and ceftozolane/tazobactam than E. coli (all p

The use of antibiotics in fish production can induce bacterial populations to develop resistance to multiple antibiotics and transfer antibiotic resistance genes to other bacteria, including clinically relevant bacteria. This study evaluated the diversity of Enterobacterales in sediment from lagoons with fish farming activity and analyzed antibiotic resistance in the central region of Peru. Sediment samples were collected from four fish-active ponds and transported to the laboratory for analysis. Bacterial diversity was analyzed using DNA sequencing and antibiotic resistance was tested using the disk diffusion method. The results showed variability of bacterial diversity in the ponds with fish farming activity. Simpson's index indicated that the Habascocha lagoon is the most diverse in bacterial species of the order Enterobacterales (0.8), but the least dominant. The Shannon-Wiener index revealed that it is the most diverse (2.93) and the Margalef index revealed that species richness in this lagoon is high (5.72). Similarity percentage analysis (SIMPER) allowed the identification of the main Enterobacterales with the highest percentage contribution in the frequencies of individuals. In general, the Enterobacterales species isolated showed multi-resistance to the antibiotics used and Escherichia coli was the most resistant.

ABSTRACT: Objectives: Herein, we sequenced and assembled the genome of a Shigella sonnei isolate carrying several small plasmids using a hybrid approach that combined Oxford Nanopore Technologies and Illumina platforms. Methods: Whole-genome sequencing was conducted using the Illumina iSeq 100 and Oxford Nanopore MinION systems, and the resulting reads were used for hybrid genome assembly via Unicycler. Coding sequences were annotated using RASTtk, while genes involved in antimicrobial resistance and virulence were identified using AMRFinderPlus. Plasmid nucleotide sequences were aligned to the NCBI non-redundant database using BLAST, and replicons were identified using PlasmidFinder. Results: The genome consisted of 1 chromosome (4 801 657 bp), 3 major plasmids (212 849 bp, 86 884 bp, and 83 425 bp, respectively) and 12 small cryptic plasmids (ranging from 8390 bp to 1822 bp). BLAST analysis revealed that all plasmids were highly similar to previously deposited sequences. Genome annotation predicted 5522 coding regions, including 19 antimicrobial resistance genes and 17 virulence genes. Four of the antimicrobial resistance genes were located in small plasmids, and four of the virulence genes were located in a large virulence plasmid. Conclusion: The presence of antimicrobial resistance genes in small cryptic plasmids may represent an overlooked mechanism for the propagation of these genes among bacterial populations. Our work provides new data on these elements that may inform the development of new strategies to control the spread of extended spectrum β-lactamase–producing bacterial strains.

Purpose: In this study, we aimed to assess the geographic distribution and molecular characteristics of β-lactamases among Enterobacterales isolates causing intra-abdominal infections (IAIs) from 2015 to 2018 in the Asia–Pacific region. Method: Isolates were investigated for extended-spectrum β-lactamases (ESBLs), AmpC β-lactamases, and carbapenemases using multiplex PCR assays and full-gene DNA sequencing. Result: A total of 832 Enterobacterales isolates from 8 different countries with β-lactamase genes were analysed. Plasmid-mediated ESBLs and AmpC β-lactamases were encoded in 598 (71.9 %) and 314 (37.7 %) isolates, respectively. In 710 (85.3 %) carbapenemase-negative isolates, positivity for both AmpC β-lactamases and ESBLs was identified in 51 (8.5 %) Escherichia coli and 24 (3.4 %) Klebsiella pneumoniae isolates. The most prevalent countries were Taiwan and Vietnam, and the co-occurrence of CMY/CTX-M in E. coli and DHA-1/ESBLs in K. pneumoniae was predominant. All isolates showed high susceptibility to colistin, but susceptibility to carbapenems varied among different resistance mechanism combinations. Among 122 (14.7 %) isolates encoding carbapenemase, NDM (n = 67, including 64.2 % NDM-1) was the most common, followed by the OXA-48-type (n = 49), KPC (n = 24) and IMP (n = 4). The most prevalent country was Thailand (n = 44), followed by Vietnam (n = 35) and the Philippines (n = 21). Twenty-two isolates were found to encode multiple carbapenemases, 16 of which were collected from Thailand and harbored NDM-1, OXA-232 and CTX-M-15. Despite high susceptibility to amikacin, susceptibility to colistin was only 56 %. Conclusion: The emergence of carbapenem-non-susceptible AmpC/ESBL co-occurring Enterobacterales and colistin non-susceptible carbapenemases co-occurring K. pneumoniae highlights potential therapeutic challenges in the Asia–Pacific region.

ABSTRACT: Objectives: Ceftolozane-tazobactam (C-T) is an anti-pseudomonal cephalosporin combined with a well-described β-lactamase inhibitor. Ceftolozane-tazobactam has enhanced activity against Pseudomonas aeruginosa, and activity against Enterobacterales isolates that produce extended-spectrum β-lactamases (ESBLs) or AmpC cephalosporinases. In this study, we analysed the susceptibility of Gram-negative isolates to C-T and comparators collected in Australia and New Zealand from 2016 to 2018 as part of the Program to Assess Ceftolozane-Tazobactam Susceptibility (PACTS) surveillance. Methods: A total of 1693 nonduplicate Enterobacterales and 435 P. aeruginosa isolates were collected prospectively from hospitalized patients in six medical centres in Australia and two in New Zealand. Susceptibilities (S) to C-T and comparators were determined using broth microdilution. EUCAST breakpoints were used. Isolates with multi-drug resistant (MDR), extensively drug resistant (XDR), extended-spectrum β-lactamase non-carbapenem resistant (ESBL, non-CRE) phenotype, and CRE were analysed. Results: For P. aeruginosa, 97.5% were S to C-T while 89.9% were S to meropenem. According to EUCAST criteria, 86.4% were susceptible-increased exposure to piperacillin-tazobactam. MDR and XDR P. aeruginosa isolates had 76.7% and 65.4% S to C-T, respectively; 34.9% and 19.2% S to meropenem, respectively; and 23.3% and 15.4% were susceptible-increased exposure to piperacillin-tazobactam, respectively. Meropenem (99.8% S), amikacin (99.1% S), and C-T (96.5% S) were the most active against Enterobacterales. Susceptibilities to C-T were 94.3% for ESBL, non-CRE phenotype, and 78.4% for MDR isolates. Only three CRE and five XDR isolates were identified. Conclusions: These in vitro data indicate that C-T is a potent antimicrobial with activity against MDR and XDR P. aeruginosa, as well as ESBL, non-CRE phenotype isolates and MDR Enterobacterales.

Background and objective: The presence of carbapenemase enzymes among Enterobacterales is the main mechanism to reduce susceptibility to a wide range of antibiotics. Carbapenemase enzymes such as the Klebsiella pneumoniae carbapenemase (KPC) hydrolyse beta-lactam antibiotics group, which includes carbapenem, leads to fewer treatment options. We aim to describe the first report of carbapenem-resistant K. pneumoniae (CRKP) sequence type (ST) 11 harbouring KPC in Oman. Material and methods: Five confirmed CRKP isolates were isolated from clinical samples during the period of January 2019 till December 2019. Strains were genotyped by pulse field gel electrophoresis (PFGE) for genetic relatedness. Whole genome sequencing (WGS) was performed to observe relationships with global strains using multilocus sequence typing (MLST). Antimicrobial genes, capsular loci-K-types, plasmids types and virulence genes were also identified using whole genome sequence data. Results: All five CRKP were determined to have blaKPC-2 with or without blaOX-A48 and blaNDM-2. The molecular genotyping by PFGE showed 100% similarity among the five isolates. The MLST allelic profile analysis clonally clustered our strains with SL-258, CG-11 and ST11 mainly reported from South Asia. Further molecular characterization of the capsular K-locus and O-locus genes, revealed the strains to belong to KL-47 type and OL101 type respectively. The core genome typing suggests that our strains were clonally related to Chinese strains with less than five chromosomal nucleotides differences. Conclusion: Epidemiological and molecular analyses confirmed that these KPC-producing K. pneumoniae strains are from a single clone that caused multiple nosocomial infections in one health institution. This finding highlights the importance to sustain the surveillance and infection prevention efforts and to step up active screening to prevent the spread of nosocomial infection.

ABSTRACT: Objectives: To report reference method antimicrobial susceptibility testing results for recent clinical isolates of Gram-negative bacilli from Morocco. Methods: CLSI (Clinical and Laboratory Standards Institute) broth microdilution antimicrobial susceptibility testing was performed by a central laboratory for isolates of Enterobacterales (n = 810), Pseudomonas aeruginosa (n = 321), and Acinetobacter baumannii (n = 191) collected in 2018–2020 by three hospital laboratories in Morocco. MICs were interpreted using both CLSI (2021) and EUCAST (European Committee on Antimicrobial Susceptibility Testing) (2021) breakpoints. Molecular testing for β-lactamase genes was performed on isolates meeting defined screening criteria. Results: Most isolates of Enterobacterales were susceptible (CLSI/EUCAST breakpoints) to amikacin (98.0%/96.2%), ceftazidime-avibactam (94.8%/94.8%), and meropenem (92.5%/94.2%). Of Enterobacterales isolates eligible for β-lactamase gene screening (n = 210), 174 were ESBL-positive, 40 were metallo-β-lactamase-positive (all NDM), 39 were serine carbapenemase-positive (all OXA); and 7 isolates carried both OXA-48 and NDM-1. Amikacin (89.1%/89.1%) and ceftazidime-avibactam (88.2%/88.2%) were the most active agents tested against P. aeruginosa. Applying CLSI and EUCAST breakpoints, MDR rates were 21.9% and 29.3% for Enterobacterales and 18.4% and 21.8% for P. aeruginosa. Susceptible rates for amikacin, ceftazidime-avibactam, and meropenem were 93.2%/89.5%, 77.4%/82.3%, and 67.8%/80.2% for MDR Enterobacterales and 50.8%/57.1%, 40.7%/45.7%, and 27.1/32.9% for MDR P. aeruginosa. ≥70% of A. baumannii isolates were resistant to all agents tested (except colistin, EUCAST breakpoints only) including amikacin and meropenem. Conclusion: Newer β-lactam/β-lactamase inhibitor combinations such as ceftazidime-avibactam warrant testing and reporting for Enterobacterales and P. aeruginosa in Morocco given the presence of significant resistance to first-line β-lactams and fluoroquinolones, pervasive ESBLs and carbapenemases, and toxicity concerns associated with some second-line agents.

Objectives: To perform the first prospective surveillance evaluating the occurrence of genes encoding colistin resistance, fosfomycin resistance, carbapenemase, or extended-spectrum ß-lactamases (ESBLs) among Enterobacterial isolates recovered from the gut flora of pigs from Egypt. Methods: Between February and April 2020, 81 rectal swabs were collected from pigs in a slaughterhouse, Cairo, Egypt. Samples were screened for different resistance mechanisms using SuperPolymyxin, ChromID ESBL, SuperFOS, and SuperCarba selective agar plates. Antimicrobial susceptibility testing was performed for all isolates using disk diffusion and broth microdilution techniques. PCR screening was performed for ESBLs, carbapenemases, mcr, and fosA genes. Mating-out assays, multilocus sequence typing analysis, and plasmid typing were also performed. Results: A high prevalence of ESBLs, carbapenemases, fosfomycin, and colistin resistance genes was evidenced among those isolates. The predominant ESBL identified was blaCTX-M-15, followed by blaCTX-M-9. We also identified blaNDM-5 and blaOXA-244. fosA3, fosA4, and fosA6 were identified in E. coli isolates. In addition, 11 MCR-1 producers were recovered. Notably, co-occurrence of ESBL genes and mcr or fosA genes was observed. MLST analysis revealed a high clonal diversity, ruling out the dissemination of one major clone. IncFIB-type was predominantly present among ESBL and FosA producers. The blaNDM-5 gene was carried on an IncX4-type, although the blaOXA-244 gene was chromosomally located. The mcr-1 gene was carried on a diversity of plasmids (IncI2, IncX4, and IncHI2). Conclusion: These results raise serious public health concerns as Egyptian pig meat could serve as a reservoir for antimicrobial resistance genes (ARGs), leading to worldwide dissemination.

ABSTRACT: Objectives: There are scant primary clinical data on antimicrobial resistance (AMR) burden from low- and middle-income countries (LMICs). We adapted recent World Health Organization methodology to measure the effect of third-generation cephalosporin resistance (3GC-R) on mortality and excess length of hospital stay in Fiji. Methods: We conducted a prospective cohort study of inpatients with Enterobacterales bloodstream infections (BSIs) at Colonial War Memorial Hospital, Suva. We used cause-specific Cox proportional hazards models to estimate the effect of 3GC-R on the daily risk (hazard) of in-hospital mortality and being discharged alive (competing risks), and we used multistate modelling to estimate the excess length of hospital stay. Results: From July 2020 to February 2021 we identified 162 consecutive Enterobacterales BSIs; 3GC-R was present in 66 (40.7%). Crude mortality for patients with 3GC-susceptible and 3GC-R BSIs was 16.7% (16/96) and 30.3% (20/66), respectively. 3GC-R was not associated with the in-hospital mortality hazard rate (adjusted hazard ratio [aHR] 1.13, 95% confidence interval [CI] 0.51–2.53) or being discharged alive (aHR 0.99, 95% CI 0.65–1.50), whereas Charlson comorbidity index score (aHR 1.62, 95% CI 1.36–1.93) and Pitt bacteraemia score (aHR 3.57, 95% CI 1.31–9.71) were both associated with an increased hazard rate of in-hospital mortality. 3GC-R was associated with an increased length of stay of 2.6 days (95% CI 2.5–2.8). 3GC-R was more common among hospital-associated infections, but genomics did not identify clonal transmission. Conclusion: Patients with Enterobacterales BSIs in Fiji had high mortality. There were high rates of 3GC-R, which was associated with increased hospital length of stay but not with in-hospital mortality.

Objectives: Carbapenem-resistant Enterobacterales (CRE) remain an urgent public health priority in the United States. CRE poses a major threat to patients in healthcare and a potential risk to the community. This study examined the epidemiological trends, clinical, and microbiological data of CRE in the Greater Houston region of Texas. Methods: A multi-institutional retrospective observational study was conducted using surveillance data collected from 2015 to 2020. Predictors of incidence rates of CRE were determined by a negative binomial regression fit using a generalized estimation equation. Results: Over a 6-year period, 4236 CRE cases were reported, of which Klebsiella pneumoniae accounted for 84.8%. The results show a steady increase in CRE cases, with a sharp rise since 2018. The majority of carbapenemase-producing Enterobacterales were Klebsiella pneumoniae carbapenemase (KPC)-producing (77.2%), followed by other rare carbapenemases, which includes OXA-48, NDM, IMP, VIM, coproduction of KPC with OXA-48, KPC with NDM, and NDM with OXA-48. Acute care hospitals (ACH) accounted for 68.5% of the source of CRE cases. The incidence rate of CRE cases reported from ACH and long-term acute care (LTAC) facilities was 1.16 times that of long-term care facilities (adjusted rate ratio [ARR] = 1.16, 95% confidence interval [CI]:1.04–1.30). The incidence rate of CRE among patients with indwelling devices was 15% (ARR = 0.85, 95% CI: 0.79–0.92) lower than that of patients without indwelling devices. Conclusion: The rise in the rate of CRE cases despite aggressive infection prevention and control strategies in the region is alarming. Evaluating and improving the current infection control strategies may be warranted.

Objectives: : Infections caused by drug-resistant Enterobacterales including those producing metallo-β-lactamases (MBLs) are particularly challenging due to limited therapeutic options. The drug combination aztreonam/avibactam (ATM-AVI) is under clinical development for treating serious infections caused by these strains. This study assessed the in vitro activity of ATM-AVI against Enterobacterales isolates collected globally in the ATLAS surveillance programme in 2019. Methods: : Clinical isolates of Enterobacterales (N = 18 713) including Citrobacter freundii, Citrobacter koseri, Enterobacter cloacae complex, Escherichia coli, Klebsiella aerogenes, Klebsiella oxytoca, Klebsiella pneumoniae, Proteus mirabilis, and Serratia marcescens collected from 232 sites in 2019 were analysed. Antimicrobial susceptibility testing was performed by reference broth microdilution. A pharmacokinetic/pharmacodynamic based breakpoint of 8 mg/L was considered for ATM-AVI activity. Results: : ATM-AVI demonstrated potent antimicrobial activity against all Enterobacterales, with 99.9% isolates inhibited at MIC ≤8 mg/L (MIC90, 0.25 mg/L). MICs ≤8 mg/L (>99.0%) were noted for ATM-AVI across regions worldwide. Among other antimicrobials, amikacin, colistin, imipenem, meropenem, and tigecycline were also active (susceptibility >85.0%) against Enterobacterales. Activity of ATM-AVI was sustained against multidrug-resistant, extended-spectrum β-lactamase producing, and carbapenem-resistant isolates (susceptibility >99%; MIC90, 0.25–0.5 mg/L). Importantly, potent activity for ATM-AVI (>99.0%; MIC90, 0.5 mg/L) was noted among MBL-positive isolates and those producing other carbapenemases, such as KPC and OXA-48. Conclusion: : Our results demonstrated that ATM-AVI was highly active against a recent collection of Enterobacterales isolates, including those producing MBLs either alone or in combination with other carbapenemases. Thus, ATM-AVI represents a potential option for treating infections caused by antibiotic-resistant Enterobacterales including MBL-producing strains.

The Rcs sensor system, comprising the RcsB/RcsC/RcsD and RcsF proteins, is used by bacteria of the order Enterobacterales to withstand envelope damage. In non-stress conditions, Rcs is repressed by IgaA, a membrane protein with three cytoplasmic regions (cyt-1, cyt-2 and cyt-3). How the Rcs-IgaA axis evolved within Enterobacterales has not been yet explored. Here, we report phylogenetic data supporting co-evolution of IgaA with RcsC/RcsD. Functional exchange assays showed that IgaA from Shigella and Dickeya, but not from Yersinia or the endosymbionts Photorhabdus and Sodalis, repress the Rcs system of Salmonella. IgaA from Dickeya, however, repress only partially the Rcs system despite being produced at high levels in the complementation assay. The modelled structures of these IgaA variants uncovered one periplasmic and two cytoplasmic conserved β-rich architectures forming partially closed small β-barrel (SBB) domains. Conserved residues map in a connector linking cytoplasmic SSB-1 and SBB-2 domains (E180-R265); a region of cyt-1 facing cyt-2 (R188-E194-D309 and T191-H326); and between cyt-2 and cyt-3 (H293-E328-R686). These structures validated early in vivo studies in Salmonella that assigned a role in function to R188, T191 and G262, and in addition revealed a previously unnoticed “hybrid” SBB-2 domain to which cyt-1 and cyt-2 contribute. IgaA variants not functional or partially functional in Salmonella lack H192-P249 and R255-D313 interactions. Among these variants, only IgaA from Dickeya conserves the helix α6 in SSB-1 that is present in IgaA from Salmonella and Shigella. RcsF and RcsD, which interact directly with IgaA, failed to show structural features linked to specific IgaA variants. Altogether, our data provide new insights into IgaA by mapping residues selected differently during evolution and involved in function. Our data also infer contrasting lifestyles of Enterobacterales bacteria as source of variability in the IgaA-RcsD/IgaA-RcsF interactions.

Summary: Introduction: Acquired carbapenemase-producing Gram-negative bacteria are an increasing public health concern globally and have been mandatory to report in England since October 2020. However, in light of the COVID-19 (SARS-CoV-2) pandemic, the Royal College of Pathologists (RCPath) released new guidance “for reducing the need for screening of CRE (carbapenem-resistant Enterobacterales) […] in low-risk areas”, without defining “low risk”. Methods: To assess the impact of the RCPath recommendations on screening of carbapenemase-producing Enterobacterales (CPE), an online Select Survey was sent to all NHS acute hospitals in England. The initial survey distribution was between March and April 2021 and the survey was relaunched between November 2021 and March 2022. Results: In total, 54 hospitals completed the survey, representing 39.1% of 138 eligible Trusts. All hospitals had a CPE screening policy in place, and the majority of these reflect UKHSA's Framework of actions to contain CPE. Of the 23 hospitals who reported a reduction in CPE screening, only three (13.0%) indicated that this was due to the RCPath recommendations, with 21 (91.3%) indicating that there had been a natural reduction in the number of patients admitted to the Trust who would have previously been screened due to the COVID-19 pandemic. Conclusion: For most surveyed hospitals, CPE screening was not reduced due to the RCPath recommendations. However, the results highlighted that there is a large amount of individual variation in CPE screening practices and diagnostic testing between hospitals.