AmpC Β-lactamase Research Articles

Prevalence of Antibiotic Resistant E. coli Strains Isolated from Farmed Broilers and Hens in Greece, Based on Phenotypic and Molecular Analyses

The use of antimicrobials is beneficial for livestock health; however, their overuse and misuse may increase resistance to these compounds. Thus, the aim of the present study was the phenotypic and molecular examination of the presence of Escherichia coli antibiotic-resistant strains in broiler and laying hen farms. The resistance of E. coli strains was examined against various antibiotics, including several families of compounds such as penicillin class medications (ampicillin), cephalosporins (cefotaxime, cefoxitin, cefpodoxime and ceftazidime), sulfonamides (co-trimoxazole), quinolones (enrofloxacin and nalidixic acid), aminoglycosides (gentamicin), β-lactams (imipenem), aminoglycoside (streptomycin), and polymyxin (colistin). In total, 106 strains were investigated, sampled during the years 2016–2019 from 91 poultry farms, including 75 broiler farms and 16 laying hen farms, originating from three Regional Units in Greece. The examined isolates revealed the highest resistance rates to sulfamethoxazole (81.1%), nalidixic acid (73.6%), tetracyclin (70.8%), and streptomycin (70.8%). On the other hand, the resistance of the isolates to third generation cephalosporins was found to be at lower levels for ceftazidime (2.8%), ceftriaxone (3.7%) cefoxitin (4.7%), and cefotaxime (4.7%). Phenotypic tests showed that 13.6% and 10.2% of the isolates produced ESBL, while 2.7% and 1% produced AmpC b-lactamase, for broiler and laying hens, respectively. The prevalence of the mcr-1 gene was found to be 22.7%, detected only in broiler isolates. Based on our results, E. coli antibiotic resistance represents a critical control point in poultry production that, apart from farm animals, may affect public health as well.

Comparison of In Vitro Activity of Ceftazidime-Avibactam and Imipenem-Relebactam against Clinical Isolates of Pseudomonas aeruginosa.

The role of novel β-lactam/β-lactamase inhibitor combinations in ceftazidime-nonsusceptible (CAZ-NS) and imipenem-nonsusceptible (IPM-NS) Pseudomonas aeruginosa has not been fully elucidated. This study evaluated the in vitro activity of novel β-lactam/β-lactamase inhibitor combinations against Pseudomonas aeruginosa clinical isolates, determined how avibactam restored ceftazidime activity, and compared the activity of ceftazidime-avibactam (CZA) and imipenem-relebactam (IMR) against KPC-producing P. aeruginosa. Similar high susceptibility rates for CZA, IMR, and ceftolozane-tazobactam (88.9% to 89.8%) were found for 596 P. aeruginosa clinical isolates from 11 hospitals in China, and a higher susceptibility rate to ceftazidime than imipenem was observed (73.5% versus 63.1%). For CAZ-NS and IPM-NS isolates, susceptibility rates for CZA, ceftolozane-tazobactam, and IMR were 61.5% (75/122), 54.9% (67/122), and 51.6% (63/122), respectively. For CAZ-NS, IPM-NS but CZA-susceptible isolates, 34.7% (26/75) harbored acquired β-lactamases with KPC-2 predominant (n = 19), and 45.3% (34/75) presented overexpression of chromosomal β-lactamase ampC. Among 22 isolates carrying KPC-2 carbapenemase alone, susceptibility rates to CZA and IMR were 86.4% (19/22) and 9.1% (2/22), respectively. Notably, 95% (19/20) of IMR-nonsusceptible isolates had an inactivating mutation of oprD gene. In conclusion, CZA, ceftolozane-tazobactam, and IMR exhibit high activity against P. aeruginosa, and CZA is more active than IMR against CAZ-NS and IPM-NS isolates as well as KPC-producing P. aeruginosa. Avibactam overcomes ceftazidime resistance engendered by KPC-2 enzyme and overexpressed AmpC. IMPORTANCE The emergence of antimicrobial resistance poses a particular challenge globally, and the concept of P. aeruginosa with "difficult-to-treat" resistance (DTR-P. aeruginosa) was proposed. Here, P. aeruginosa clinical isolates were highly susceptible to three β-lactamase inhibitor combinations, CZA, IMR, and ceftolozane-tazobactam. The combination of KPC-2 enzyme and nonfunctional porin OprD contributed to IMR resistance in P. aeruginosa, and CZA was more active than IMR in fighting against KPC-2-producing P. aeruginosa. CZA also showed good activity against CAZ-NS and IPM-NS P. aeruginosa, primarily by inhibiting KPC-2 enzyme and overproduced AmpC, supporting the clinical use of CZA in the treatment of infections caused by DTR-P. aeruginosa.

Phenotypic and genetic screening of Klebsiella pneumoniae isolates from human UTI patients for beta-lactamases and their genetic diversity analysis by ERIC and REP PCRs.

Klebsiella pneumoniae is one of the major nosocomial pathogens responsible for pneumoniae, septicaemia, liver abscesses, and urinary tract infections. Coordinated efforts by antibiotic stewardship and clinicians are underway to curtail the emergence of antibiotic-resistant strains. The objective of the present study is to characterize K. pneumoniae strains through antibiotic resistance screening for production of beta-lactamases (β-lactamases) such as extended spectrum beta lactamases (ESBLs), AmpC β-lactamases, and carbapenemases by phenotypic and genotypic methods and genetic fingerprinting by enterobacterial repetitive intergenic consensus-polymerase chain reaction (ERIC-PCR) and repetitive element palindromic PCR (REP-PCR). A total of 85 K. pneumoniae strains isolated from 504 human urinary tract infections (UTI) were used in this study. Only 76 isolates showed positive in phenotypic screening test (PST), while combination disc method (CDM) as phenotypic confirmatory test (PCT) confirmed 72 isolates as ESBL producers. One or more β-lactamase genes were detected by PCR in 66 isolates (91.66%, 66/72) with blaTEM gene being the most predominant (75.75%, 50/66). AmpC genes could be detected in 21 isolates (31.8%, 21/66) with FOX gene being the predominant (24.24%, 16/66), whereas NDM-I was detected in a single strain (1.51%, 1/66). Genetic fingerprinting using ERIC-PCR and REP-PCR revealed wide heterogeneity among β-lactamase producing isolates with discriminatory power of 0.9995 and 1, respectively.

Phylogenetic Analysis of Escherichia coli Isolated from Australian Feedlot Cattle in Comparison to Pig Faecal and Poultry/Human Extraintestinal Isolates.

The similarity of commensal Escherichia coli isolated from healthy cattle to antimicrobial-resistant bacteria causing extraintestinal infections in humans is not fully understood. In this study, we used a bioinformatics approach based on whole genome sequencing data to determine the genetic characteristics and phylogenetic relationships among faecal Escherichia coli isolates from beef cattle (n = 37) from a single feedlot in comparison to previously analysed pig faecal (n = 45), poultry extraintestinal (n = 19), and human extraintestinal E. coli isolates (n = 40) from three previous Australian studies. Most beef cattle and pig isolates belonged to E. coli phylogroups A and B1, whereas most avian and human isolates belonged to B2 and D, although a single human extraintestinal isolate belonged to phylogenetic group A and sequence type (ST) 10. The most common E. coli sequence types (STs) included ST10 for beef cattle, ST361 for pig, ST117 for poultry, and ST73 for human isolates. Extended-spectrum and AmpC β-lactamase genes were identified in seven out of thirty-seven (18.9%) beef cattle isolates. The most common plasmid replicons identified were IncFIB (AP001918), followed by IncFII, Col156, and IncX1. The results confirm that feedlot cattle isolates examined in this study represent a reduced risk to human and environmental health with regard to being a source of antimicrobial-resistant E. coli of clinical importance.

ACT-107, a novel variant of AmpC β-lactamase from Enterobacter huaxiensis isolated from Neotropical leaf frog (Phyllomedusa distincta) inhabiting the Brazilian Atlantic Forest

The aim of this study was to characterise a broad-spectrum cephalosporin-resistant AmpC-positive Enterobacter huaxiensis colonising the skin of a Neotropical frog (Phyllomedusa distincta) inhabiting the Brazilian Atlantic Forest. During a genomic surveillance study of antimicrobial resistance, we screened skin samples from P. distincta. Gram-negative bacteria growing on MacConkey agar plates supplemented with 2 µg/mL ceftriaxone were identified by matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry. A cephalosporin-resistant E. huaxiensis was sequenced using the Illumina NextSeq platform. Genomic data were analysed using bioinformatics tools, whereas AmpC β-lactamase was characterised in depth by comparative analysis of amino acids, in silico modelling, and analysis of susceptibility to β-lactam antibiotics and combinations of β-lactamase inhibitors. Whole-genome sequencing analysis revealed a novel variant of AmpC β-lactamase belonging to the ACT family, designated ACT-107 by NCBI. This variant contains 12 novel amino acid mutations within the ACT family, 5 in the signal peptide sequence (Ile2, Met14, Tyr16, Gly18 and Thr20), and 7 in the mature protein (Gln22, His43, Cys60, Thr157, Glu225, Ala252 and Asn310). In silico modelling showed that substitutions occurring in the mature chain are localised in the solvent-accessible surface of the protein, where they are not expected to affect the β-lactamase activity, as observed in the resistance profile. Strikingly, 'not designated' ACT variants from E. huaxiensis were clustered (> 96% identity) with ACT-107. Since E. huaxiensis has been isolated from human infection, ACT-107 requires surveillance and the attention of clinicians.

Collateral Changes in Cell Physiology Associated with ADC-7 β-Lactamase Expression in Acinetobacter baumannii.

The ADC (AmpC) β-lactamase is universally present in the Acinetobacter baumannii chromosome, suggesting it may have a yet-to-be-identified cellular function. Using peptidoglycan composition analysis, we show that overexpressing the ADC-7 β-lactamase in A. baumannii drives changes consistent with altered l,d-transpeptidase activity. Based on this, we tested whether cells overexpressing ADC-7 would exhibit new vulnerabilities. As proof of principle, a screen of transposon insertions revealed that an insertion in the distal 3' end of canB, encoding carbonic anhydrase, resulted in a significant loss of viability when the adc-7 gene was overexpressed. A canB deletion mutant exhibited a more pronounced loss of viability than the transposon insertion, and this became amplified when cells overexpressed ADC-7. Interestingly, overexpression of the OXA-23 or TEM-1 β-lactamases also led to a pronounced loss of viability in cells with reduced carbonic anhydrase activity. In addition, we demonstrate that reduced CanB activity led to increased sensitivity to peptidoglycan synthesis inhibitors and to the carbonic anhydrase inhibitor ethoxzolamide. Furthermore, this strain exhibited a synergistic interaction with the peptidoglycan inhibitor fosfomycin and ethoxzolamide. Our results highlight the impact of ADC-7 overexpression on cell physiology and reveal that the essential carbonic anhydrase CanB may represent a novel target for antimicrobial agents that would exhibit increased potency against β-lactamase-overexpressing A. baumannii. IMPORTANCE Acinetobacter baumannii has become resistant to all classes of antibiotics, with β-lactam resistance responsible for the majority of treatment failures. New classes of antimicrobials are needed to treat this high-priority pathogen. This study had uncovered a new genetic vulnerability in β-lactamase-expressing A. baumannii, where reduced carbonic anhydrase activity becomes lethal. Inhibitors of carbonic anhydrase could represent a new method for treating A. baumannii infections.

Prevalence and Characterization of Extended-Spectrum β-Lactamase-Producing Escherichia coli Isolated from Dogs and Cats in South Korea.

Overall, 836 Escherichia coli isolates (695 isolates from dogs and 141 from cats) were recovered from the diarrhea, skin/ear, urine, and genitals of dogs and cats between 2018 and 2019. Cefovecin and enrofloxacin resistance were noted in 17.1% and 21.2% of E. coli isolates, respectively. The cefovecin and enrofloxacin resistance rates were higher in dog isolates (18.1% and 22.9%) compared with the rates in cat isolates (12.1%, 12.8%). Interestingly, resistance to both antimicrobials was noted in 10.8% (90/836) of the isolates, predominantly in isolates from dogs. blaCTX-M-14, blaCTX-M-15, and blaCMY-2 were the most frequent extended-spectrum β-lactamase/plasmid-mediated AmpC β-lactamase (ESBL/AmpC)- gene types. The co-existence of blaCTX-M andblaCMY-2 was noted in six E. coli isolates from dogs. Sequencing analysis demonstrated that S83L and D87N in gyrA and S80I in parC were the most frequent point mutations in the quinolone resistance-determining regions of the cefovecin and enrofloxacin-resistant isolates. A total of 11 isolates from dogs carried the plasmid-mediated quinolone resistance genes (six aac(6')-Ib-cr, four qnrS, and one qnrB), while only two cat isolates carried the qnrS gene. Multilocus sequence typing of the cefovecin and enrofloxacin-resistant isolates revealed that sequence type (ST)131 E. coli carrying blaCTX-M-14 and blaCTX-M-15 genes and ST405 E. coli carrying blaCMY-2 gene were predominant among the isolated E. coli strains. The majority of the ESBL/AmpC-producing isolates displayed diverse pulsed-field gel electrophoresis profiles. This study demonstrated that third-generation cephalosporin- and fluoroquinolone-resistant E. coli were widely distributed in companion animals. The detection of the pandemic ST131 clone carrying blaCTX-M-14/15 in companion animals presented a public health threat.

In vitro activity of imipenem/relebactam, meropenem/vaborbactam and comparators against Enterobacterales causing urinary tract infection in Taiwan: results from the Study for Monitoring Antimicrobial Resistance Trends (SMART), 2020

ObjectivesAntimicrobial resistance due to β-lactamase production is a worldwide issue, and β-lactamase inhibitors have been developed to overcome the growing problem. This study aimed to evaluate the in vitro activities of two recently introduced carbapenem/β-lactamase inhibitor combinations – imipenem/relebactam and meropenem/vaborbactam – and their comparators against Enterobacterales from patients with urinary tract infections (UTIs). MethodsEnterobacterales isolates from patients with UTIs in Taiwan and participating in the Study for Monitoring Antimicrobial Resistance Trends (SMART) in 2020 were included. Minimum inhibitory concentrations (MICs) for various antibiotics were determined using the broth microdilution method. Susceptibility was interpreted based on the MIC breakpoints of the Clinical and Laboratory Standards Institute 2022. Genes encoding common β-lactamases, including extended-spectrum β-lactamases, AmpC β-lactamases and carbapenemases, were detected using multiplex polymerase chain reaction. ResultsA total of 309 Enterobacterales isolates were included, against which both imipenem/relebactam and meropenem/vaborbactam exerted excellent efficacy (275 of 309, 95% and 288 of 309, 99.3%, respectively). Among imipenem non-susceptible isolates, 17 of 43 (39.5%) and 39 of 43 (90.7%) were susceptible to imipenem/relebactam and meropenem/vaborbactam, respectively. ConclusionImipenem/relebactam and meropenem/vaborbactam may be appropriate choices for treating UTIs due to Enterobacterales resistant to commonly used antibiotics. Continuous monitoring of antimicrobial resistance is crucial.

High frequency of NDM-1 and OXA-48 carbapenemase genes among Klebsiella pneumoniae isolates in central Iran

BackgroundThe emergence and distribution of multidrug-resistant (MDR) and carbapenem-resistant Klebsiella pneumoniae (CRKP) has become a global health threat. Therefore, this study aimed to investigate the frequency and antibiotic resistance patterns of MDR, extensively drug-resistant (XDR), and CRKP, as well as the antibiotic resistance genes of Klebsiella pneumoniae (K. pneumoniae) isolates from patients’ infectious samples from central Iran.MethodsThis study examined 546 clinical samples of patients to identify K. pneumoniae. The isolates were investigated for their antibiotic resistance profile, extended-spectrum β-lactamase (ESBL), AMPC β-lactamase, carbapenemase resistance, sulfonamide, tetracycline, plasmid-mediated quinolone resistance (PMQR) along with their resistance genes, integrase, and quaternary ammonium compounds (qac) by polymerase chain reaction (PCR).ResultsOut of 546 clinical samples, 121 (22.1%) cases of K. pneumoniae were identified using culture and PCR methods. The highest antibiotic resistance rates were found for ampicillin (119/121; 98.3%), cotrimoxazole (78/121; 64.4%), and cefixime, cefotaxime, ceftriaxone, and ceftazidime as a group (77/121; 63.6%). Tigecycline, colistin, and fosfomycin were the most effective antimicrobial agents with 98.4%, 96.7%, and 95.9% susceptibility, respectively. The amount of CRKP was 51 (42.1%). All CRKP isolates were MDR. The most abundant genes were blaTEM (77/77; 100%), blaCTX−M1 (76/77; 98.7%), blaSHV (76/77; 98.7%), blaCTX−M15 (73/77; 94.8%) for ESBL; blaCIT 28 (48.3%) and blaCMY−2 26 (44.8%) for AMPC β-lactamase; and blaOXA−48 46 (90.1%) and blaNDM 36 (70.5%) for carbapenemase. Among the PMQR determinants, qnrB (25/52; 48%), qnrS (19/52; 36.5%), and qnrA (11/52; 21.1%) were positive from the isolates. TetA and tetB were recognized in 25 (44.6%) and 17 (30.3%) isolates, respectively. Class 1 and 2 integrons were recognized in 97 (80.1%) and 53 (43.8%) isolates, respectively.ConclusionsDue to the high prevalence of MDR and CRKP in central Iran, tracking and immediate intervention are necessary for control and inhibition of K. pneumoniae resistant isolates. Tigecycline, colistin, and fosfomycin are the best treatment options for treatment of patients with CRKP in this geographical area.

Molecular Characterization and Epidemiology of Antibiotic Resistance Genes of β-Lactamase Producing Bacterial Pathogens Causing Septicemia from Tertiary Care Hospitals

Septicemia is a systematic inflammatory response and can be a consequence of abdominal, urinary tract and lung infections. Keeping in view the importance of Gram-negative bacteria as one of the leading causes of septicemia, the current study was designed with the aim to determine the antibiotic susceptibility pattern, the molecular basis for antibiotic resistance and the mutations in selected genes of bacterial isolates. In this study, clinical samples (n = 3389) were collected from potentially infected male (n = 1898) and female (n = 1491) patients. A total of 443 (13.07%) patients were found to be positive for bacterial growth, of whom 181 (40.8%) were Gram-positive and 262 (59.1%) were Gram-negative. The infected patients included 238 males, who made up 12.5% of the total number tested, and 205 females, who made up 13.7%. The identification of bacterial isolates revealed that 184 patients (41.5%) were infected with Escherichia coli and 78 (17.6%) with Pseudomonas aeruginosa. The clinical isolates were identified using Gram staining biochemical tests and were confirmed using polymerase chain reaction (PCR), with specific primers for E. coli (USP) and P. aeruginosa (oprL). Most of the isolates were resistant to aztreonam (ATM), cefotaxime (CTX), ampicillin (AMP) and trimethoprim/sulfamethoxazole (SXT), and were sensitive to tigecycline (TGC), meropenem (MEM) and imipenem (IPM), as revealed by high minimum inhibitory concentration (MIC) values. Among the antibiotic-resistant bacteria, 126 (28.4%) samples were positive for ESBL, 105 (23.7%) for AmpC β-lactamases and 45 (10.1%) for MBL. The sequencing and mutational analysis of antibiotic resistance genes revealed mutations in TEM, SHV and AAC genes. We conclude that antibiotic resistance is increasing; this requires the attention of health authorities and clinicians for proper management of the disease burden.

Characterization and Comparative Genomic Analysis of Three Virulent E. coli Bacteriophages with the Potential to Reduce Antibiotic-Resistant Bacteria in the Environment.

The emerging global crisis of antibiotic resistance demands new alternative antibacterial solutions. Although bacteriophages have been used to combat bacterial infections for over a century, a dramatic boost in phage studies has recently been observed. In the development of modern phage applications, a scientific rationale is strongly required and newly isolated phages need to be examined in detail. In this study, we present the full characterization of bacteriophages BF9, BF15, and BF17, with lytic activity against extended-spectrum β-lactamases (ESBLs)- and AmpC β-lactamases (AmpC)-producing Escherichia coli, the prevalence of which has increased significantly in livestock in recent decades, representing a great hazard to food safety and a public health risk. Comparative genomic and phylogenetic analysis indicated that BF9, BF15, and BF17 represent the genera Dhillonvirus, Tequatrovirus, and Asteriusvirus, respectively. All three phages significantly reduced in vitro growth of their bacterial host and retained the ability to lyse bacteria after preincubation at wide ranges of temperature (-20-40 °C) and pH (5-9). The results described herein indicate the lytic nature of BF9, BF15, and BF17, which, along with the absence of genes encoding toxins and bacterial virulence factors, represents an undoubted asset in terms of future phage application.

Building an International One Health Strain Level Database to Characterise the Epidemiology of AMR Threats: ESBL—AmpC Producing E. coli as An Example—Challenges and Perspectives

Antimicrobial resistance (AMR) is one of the top public health threats nowadays. Among the most important AMR pathogens, Escherichia coli resistant to extended spectrum cephalosporins (ESC-EC) is a perfect example of the One Health problem due to its global distribution in animal, human, and environmental sources and its resistant phenotype, derived from the carriage of plasmid-borne extended-spectrum and AmpC β-lactamases, which limits the choice of effective antimicrobial therapies. The epidemiology of ESC-EC infection is complex as a result of the multiple possible sources involved in its transmission, and its study would require databases ideally comprising information from animal (livestock, companion, wildlife), human, and environmental sources. Here, we present the steps taken to assemble a database with phenotypic and genetic information on 10,763 ESC-EC isolates retrieved from multiple sources provided by 13 partners located in eight European countries, in the frame of the DiSCoVeR Joint Research project funded by the One Health European Joint Programme (OH-EJP), along with its strengths and limitations. This database represents a first step to help in the assessment of different geographical and temporal trends and transmission dynamics in animals and humans. The work performed highlights aspects that should be considered in future international efforts, such as the one presented here.

YgfB increases β-lactam resistance in Pseudomonas aeruginosa by counteracting AlpA-mediated ampDh3 expression

YgfB-mediated β-lactam resistance was recently identified in multi drug resistant Pseudomonas aeruginosa. We show that YgfB upregulates expression of the β-lactamase AmpC by repressing the function of the regulator of the programmed cell death pathway AlpA. In response to DNA damage, the antiterminator AlpA induces expression of the alpBCDE autolysis genes and of the peptidoglycan amidase AmpDh3. YgfB interacts with AlpA and represses the ampDh3 expression. Thus, YgfB indirectly prevents AmpDh3 from reducing the levels of cell wall-derived 1,6-anhydro-N-acetylmuramyl-peptides, required to induce the transcriptional activator AmpR in promoting the ampC expression and β-lactam resistance. Ciprofloxacin-mediated DNA damage induces AlpA-dependent production of AmpDh3 as previously shown, which should reduce β-lactam resistance. YgfB, however, counteracts the β-lactam enhancing activity of ciprofloxacin by repressing ampDh3 expression and lowering the benefits of this drug combination. Altogether, YgfB represents an additional player in the complex regulatory network of AmpC regulation.

Genetic relatedness of third-generation cephalosporin-resistant Escherichia coli among livestock, farmers, and patients in Japan

ObjectivesThe third-generation cephalosporin (3GC)-resistant E. coli strains have been detected worldwide in humans and animals. Hence, in this study, we evaluated the prevalence and genetic characteristics of 3GC-resistant E. coli in livestock, farmers, and patients to further analyse if livestock serves as a potential reservoir of antimicrobial-resistant bacteria. MethodsFaecal samples were collected from 330 healthy livestock (216 cattle and 114 swine), 61 healthy livestock farmers (52 cattle farmers and 9 swine farmers), and 68 non-duplicate 3GC-resistant E. coli isolates were also obtained from the clinical specimens of patients in Japan between 2013 and 2015. Genes associated with resistance in 3GC-resistant E. coli were identified using polymerase chain reaction (PCR) and DNA sequencing. Genotypic diversity was determined by multilocus sequence typing (MLST) and pulsed-field gel electrophoresis (PFGE). ResultsWe obtained 39 and 17 non-duplicated 3GC-resistant E. coli strains from healthy livestock (33 cattle and six swine) and livestock farmers, respectively. All isolates carried either CTX-M-type extended-spectrum β-lactamase or plasmid-mediated AmpC β-lactamase genes, with CTX-M-14 being the most frequent. CTX-M producers from livestock and patients belonged to 22 and 19 different sequence types (STs), respectively, and only three STs were the same. Among the 3GC-resistant E. coli from livestock and farmers, three types of CTX-M producers have shown similar characteristics (CTX-M genotype, ST, PFGE patterns, and antimicrobial susceptibilities) and were identified as clonal isolates shared among their farms. ConclusionsOur study findings indicate that CTX-M-14 is predominant in Japan. No distinct relationship was observed between the 3GC-resistant E. coli isolated from livestock and patients; however, some clonal relatedness was observed between the isolates from livestock and farmers due to their close contact.

Activity of aztreonam/avibactam against metallo-β-lactamase-producing Enterobacterales from the UK: Impact of penicillin-binding protein-3 inserts and CMY-42 β-lactamase in Escherichia coli

Aztreonam/avibactam is being developed on the rationale that aztreonam evades metallo-β-lactamases (MBLs) whilst avibactam protects against co-produced serine β-lactamases. We measured the activity of aztreonam/avibactam against MBL-producing Enterobacterales referred to the UKHSA in 2015, 2017, 2019. MICs were determined by broth microdilution, genome sequences with Illumina technology. For Klebsiella and Enterobacter spp. with NDM, IMP or VIM enzymes, the MICs of aztreonam/avibactam were unimodally distributed, with >90% of isolates inhibited at 1+4 mg/L, and all inhibited at 8+4 mg/L. Over 85% of Escherichia coli with NDM carbapenemases were inhibited at 8+4 mg/L, but their MIC distribution was multimodal, with major peaks at 0.12 and 8 mg/L. Forty-eight of 50 NDM E. coli with 'high' aztreonam/avibactam MICs (defined as ≥8 mg/L) had YRIK inserted after amino-acid 333 of penicillin-binding protein (PBP)3 or had a YRIN insert plus an acquired AmpC β-lactamase, commonly CMY-42. Ten of 15 E. coli with 'moderately-raised' aztreonam/avibactam MICs (0.5-4 mg/L) had YRIN inserts without acquired AmpC. Twenty-two of 24 E. coli isolates with 'normal' MICs (0.03-0.25 mg/L) lacked PBP3 inserts. YRIK inserts were associated with E. coli ST405 and YRIN with ST167; however, many isolates with high or moderately-raised MICs were clonally diverse. No substantive MIC distribution shifts occurred across the 3 survey years; ST405 isolates with YRIK comprised more high-MIC organisms in 2019 versus earlier years, but the apparent excess lacked significance (P >0.05).

The European Union Summary Report on Antimicrobial Resistance in zoonotic and indicator bacteria from humans, animals and food in 2020/2021.

Antimicrobial resistance (AMR) data on zoonotic and indicator bacteria from humans, animals and food are collected annually by the EU Member States (MSs) and reporting countries, jointly analysed by EFSA and ECDC and presented in a yearly EU Summary Report. This report provides an overview of the main findings of the 2020-2021 harmonised AMR monitoring in Salmonella spp., Campylobacter jejuni and C. coli in humans and food-producing animals (broilers, laying hens and turkeys, fattening pigs and bovines under 1year of age) and relevant meat thereof. For animals and meat thereof, indicator E. coli data on the occurrence of AMR and presumptive Extended spectrum β-lactamases (ESBL)-/AmpC β-lactamases (AmpC)-/carbapenemases (CP)-producers, as well as the occurrence of methicillin-resistant Staphylococcus aureus are also analysed. In 2021, MSs submitted for the first time AMR data on E. coli isolates from meat sampled at border control posts. Where available, monitoring data from humans, food-producing animals and meat thereof were combined and compared at the EU level, with emphasis on multidrug resistance, complete susceptibility and combined resistance patterns to selected and critically important antimicrobials, as well as Salmonella and E. coli isolates exhibiting ESBL-/AmpC-/carbapenemase phenotypes. Resistance was frequently found to commonly used antimicrobials in Salmonella spp. and Campylobacter isolates from humans and animals. Combined resistance to critically important antimicrobials was mainly observed at low levels except in some Salmonella serotypes and in C. coli in some countries. The reporting of a number of CP-producing E.coli isolates (harbouring bla OXA-48, bla OXA-181, and bla NDM-5 genes) in pigs, bovines and meat thereof by a limited number of MSs (4) in 2021, requests a thorough follow-up. The temporal trend analyses in both key outcome indicators (rate of complete susceptibility and prevalence of ESBL-/AmpC- producers) showed that encouraging progress have been registered in reducing AMR in food-producing animals in several EU MSs over the last years.

A Representative Collection of Commensal Extended-Spectrum- and AmpC-β-Lactamase-Producing Escherichia coli of Animal Origin for Phage Sensitivity Studies.

Extended-spectrum β-lactamase (ESBL)- and AmpC β-lactamase (AmpC)-producing Escherichia coli from livestock and meat represent a zoonotic risk and biocontrol solutions are needed to prevent transmission to humans. In this study, we established a representative collection of animal-origin ESBL/AmpC E. coli as target to test the antimicrobial potential of bacteriophages. Bioinformatic analysis of whole-genome sequence data of 198 ESBL/AmpC E. coli from pigs, broilers, and broiler meat identified strains belonging to all known E. coli phylogroups and 65 multilocus sequence types. Various ESBL/AmpC genes and plasmid types were detected with expected source-specific patterns. Plaque assay using 15 phages previously isolated using the E. coli reference collection demonstrated that Warwickvirus phages showed the broadest host range, killing up to 26 strains. 154/198 strains were resistant to infection by all phages tested, suggesting a need for isolating phages specific for ESBL/AmpC E. coli. The strain collection described in this study is a useful resource fulfilling such need.

First Report Occurrence of CIT and DHA AmpC β-lactamase Gene in Escherichia coli and Klebsiella pnuemoniae from Clinical Sample in South Eastern, Nigeria

Background and Objectives: Over time, the enzymes AmpC β-lactamases have become more significant, due to their roles in antibiotic resistance among enterobacteriaceace especially in Escherichia coli and Klebsiella pnuemoniae. Due to increase multidrug resistant express by AmpC β-lactamases producing bacteria strain, the patients care in several hospital has been severely hampered. Hence, this study was designed to assess the occurrence of CIT and DHA AmpC β-lactamase gene in Escherichia coli and Klebsiella pnuemoniae from clinical sample in south eastern, Nigeria
 Methodology: This study was conducted over an 8-month period on sixteen (16) non-repetitive clinical isolates of Escherichia coli and Klebsiella pnuemoniae collected from medical microbiology laboratory unit of Alex Ekweume Federal University Teaching Hospital in Abakaliki, Nigeria. The isolates were further identified using Standard microbiological Techniques and screened for cefoxitin resistance using a disc diffusion assay, followed by phenotypic tests using phenyl boronic acid assays for confirmation of AmpC β-lactamases production. Escherichia coli and Klebsiella pneumoniae strains were further screen for AmpC β-lactamase CIT and DHA genotype by polymerase chain reactions
 Result: Of the sixteen (16) confirmed phenotypic AmpC β-lactamase producing bacteria, 100% of the AmpC β-lactamase genes (DHA and CIT) were detected in E. coli from wound and urine samples from both male and female patients. The overall proportion of AmpC β-lactamases gene in Klebsiella pneumoniae were DHA (100 %) and CIT (100 %), in both male and female.
 Conclusion: This study indicate the occurrence of CIT and DHA AmpC genotype. The detection of AmpC β-lactamases in this study is of clinically importance as such bacteria are often MDR. Thus, being aware of the presence of AmpC β-lactamase-producing bacteria could be very beneficial for achieving more accurate epidemiological results as well as controlling their spread, while surveillance is required to track any further dissemination and emergence of other AmpC β-lactamase genotypes.

β-Lactamase Producing Escherichia coli Encoding blaCTX-M and blaCMY Genes in Chicken Carcasses from Egypt.

Escherichia coli with multidrug resistance and β-lactamase genes may constitute a great public health hazard due to the potential for their transmission to humans through the food chain. This study determined the prevalence, antibiotic resistance profiles, phylogroups, and β-lactamase genes of E. coli isolates from chicken carcasses marketed in Mansoura, Egypt. Interestingly, E. coli was detected in 98% (98/100) of the chicken carcasses examined, which seemed among the highest contamination rates by E. coli worldwide. From the 425 genetically verified uidA gene-positive E. coli, 85 isolates were further studied for antimicrobial resistance profiles, phylogroups, and β-lactamase genes. Interestingly, 89.41% of E. coli (76/85) strains tested against 24 different antibiotics were multidrug-resistant. Of the examined 85 E. coli isolates, 22 (25.88%) isolates harbored blaCTX-M and were resistant to ampicillin, cefazoline, and ceftriaxone, while three of them were resistant to ceftazidime besides. Nine (10.59%) E. coli strains harbored AmpC- β-lactamase blaCMY and were resistant to ampicillin. One isolate co-carried blaCMY and blaCTX-M genes, though it was negative for the blaTEM gene. Of the 35 isolates that harbored either extended-spectrum β-lactamase (ESBL) and/or AmpC β-lactamase genes, six strains (17.14%) were assigned to pathogenic phylogroup F and one to phylogroup E, whereas 28 (80%) isolates belonged to commensal phylogenetic groups.

In vitro activity of imipenem/relebactam, meropenem/vaborbactam and comparators against Enterobacterales from patients with intra-abdominal infections: Results of the study for Monitoring Antimicrobial Resistance Trends (SMART) in Taiwan, 2020

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. 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. 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 <0.05). ESBL, AmpC, and carbapenemase were detected in 40.5% (17/42), 45.2% (19/42) and 11.9% (5/42) among tested isolates, respectively. The 5 carbapenemase genes included 4 bla KPC and 1 bla IMP . The imipenem-non-susceptible isolates (n = 30) had higher susceptibility rates to meropenem/vaborbactam (93.3%, 28/30) than imipenem/relebactam (50%, 12/30) ( p < 0.05). Imipenem/relebactam and meropenem/vaborbactam had excellent efficacy against Enterobacterales isolates . Meropenem/vaborbactam allowed better salvage therapy for carbapenem-resistant Enterobacterales infections.