EUCAST Breakpoints Research Articles

Performance of disc diffusion, MIC gradient tests and Vitek 2 for ceftolozane/tazobactam and ceftazidime/avibactam susceptibility testing of Pseudomonas aeruginosa.

To assess performance of disc diffusion, gradient tests and Vitek 2 system to determine the susceptibility of clinical Pseudomonas aeruginosa strains to ceftolozane/tazobactam (C/T) and ceftazidime/avibactam (CZA). Two-hundred non-duplicate P. aeruginosa strains isolated by 47 French medical laboratories were selected to cover a wide range of C/T and CZA MICs. Performance of C/T disc (30/10 μg, Bio-Rad), CZA discs (10/4 μg) (Thermo Fisher and Bio-Rad), C/T and CZA gradient tests (Etest, BioMérieux; MIC Test Strip, Liofilchem), and AST-XN12 card of Vitek 2 system (BioMérieux) were compared with a broth microdilution (BMD) method (Thermo Fisher). MIC and disc results were interpreted using current EUCAST breakpoints. Twenty percent and 17% of strains were resistant to C/T and CZA, respectively. All the methods tested satisfactorily determined the susceptibility of P. aeruginosa to C/T [Category Agreement (CA) ≥95%] except the disc diffusion method. Because of the high rates of Major Errors (MEs) (12.5%), this latter method tends to overestimate the resistance. For CZA, only the gradient tests yielded more than 90% of CA. The Vitek 2 system and disc diffusion misclassified 18.1%, 10.1% (disc Bio-Rad) and 11.9% (disc Thermo Fisher) of susceptible strains, respectively. The gradient tests (MIC Test Strip and Etest) and Vitek 2 card XN12 performed the best to determine the susceptibility of P. aeruginosa to C/T, whereas gradient tests were an acceptable alternative to BMD to assess CZA susceptibility.

Antifungal activity of nitroxoline against Candida auris isolates

ObjectivesTo investigate the in vitro activity of nitroxoline against a molecularly characterized collection of clinical Candida auris isolates. MethodsThirty-five clinical isolates of C. auris from diverse sources representing all five different C. auris clades were included in the study. Nitroxoline activity was assessed using broth microdilution. Additionally, susceptibility testing by disc diffusion was assessed on RPMI-1640 and Müller–Hinton agar plates. Minimal inhibitory concentrations of the antifungals fluconazole, voriconazole, amphotericin B and anidulafungin were determined. ResultsNitroxoline MICs ranged from 0.125 to 1 mg/L (MIC50/90 0.25/0.5 mg/L). Compared with amphotericin B (MIC >1 mg/L in 4/35 isolates), anidulafungin (MIC >0.06 mg/L in 26/35 isolates) and fluconazole (MIC >4 mg/L in 31/35 isolates), in vitro activity of nitroxoline was high. Isolates belonging to clade I had marginally lower nitroxoline MICs (range 0.125–0.5 mg/L, mean MIC 0.375 mg/L) compared with clade III (range 0.5–1 mg/L, mean MIC 0.7 mg/L; p = 0.0094). The correlation of MIC and inhibition zones by disc diffusion was good when using RPMI-agar for disc diffusion, with a Pearson's correlation coefficient of –0.74 (95% CI –0.86 to –0.54). ConclusionsNitroxoline has excellent in vitro activity against C. auris isolates, with MICs of 0.125–1 mg/L (for comparison, the EUCAST breakpoint for uncomplicated urinary tract infection with Escherichia coli is ≤ 16 mg/L). It is an approved, well-tolerated antimicrobial that achieves high urinary concentrations after oral administration and could be a useful treatment option in C. auris candiduria.

In vitro antimicrobial susceptibility of clinical respiratory isolates to ceftazidime-avibactam and comparators (2016\u20132018)

BackgroundThis antimicrobial surveillance study reports in vitro antimicrobial activity and susceptibility data for a panel of agents against respiratory isolates of Enterobacterales and Pseudomonas aeruginosa.MethodsIsolates from respiratory specimens were collected in Africa/Middle East, Asia/South Pacific, Europe and Latin America between 2016 and 2018, as part of the Antimicrobial Testing Leadership and Surveillance (ATLAS) program. Broth microdilution methodology was used to quantify minimum inhibitory concentrations, from which rates of susceptibility were determined using EUCAST breakpoints (version 10). Rates of subsets with genes encoding β-lactamases (extended-spectrum β-lactamases [ESBLs], serine carbapenemases and metallo-β-lactamases [MBLs]) were also determined, as well as rates of multidrug-resistant (MDR) P. aeruginosa.ResultsAmong all respiratory Enterobacterales isolates, susceptibility to ceftazidime-avibactam, meropenem, colistin and amikacin was ≥94.4% in each region. For Enterobacterales isolates that were ESBL-positive or carbapenemase-positive/MBL-negative, ceftazidime-avibactam susceptibility was 93.6 and 98.9%, respectively. Fewer than 42.7% of MBL-positive Enterobacterales isolates were susceptible to any agents, except colistin (89.0% susceptible). Tigecycline susceptibility was ≥90.0% among Citrobacter koseri and Escherichia coli isolates, including all β-lactamase-positive subsets. ESBL-positive Enterobacterales were more commonly identified in each region than isolates that were ESBL/carbapenemase-positive; carbapenemase-positive/MBL-negative; or MBL-positive. Among all respiratory P. aeruginosa isolates, the combined susceptibility rates (susceptible at standard dosing regimen plus susceptible at increased exposure) were highest to ceftazidime-avibactam, colistin and amikacin (≥82.4% in each region). Susceptibility to colistin was ≥98.1% for all β-lactamase-positive subsets of P. aeruginosa. The lowest rates of antimicrobial susceptibility were observed among MBL-positive isolates of P. aeruginosa (≤56.6%), with the exception of colistin (100% susceptible). MDR P. aeruginosa were most frequently identified in each region (18.7–28.7%), compared with the subsets of ESBL-positive; carbapenemase-positive/MBL-negative; or MBL-positive isolates.ConclusionsRates of susceptibility among the collections of respiratory Enterobacterales and P. aeruginosa isolates were highest to ceftazidime-avibactam, colistin and amikacin in each region. Tigecycline was active against all subsets of C. koseri and E. coli, and colistin was active against all subsets of P. aeruginosa. The findings of this study indicate the need for continued antimicrobial surveillance among respiratory Gram-negative pathogens, in particular those with genes encoding MBLs.

Bacteremia caused by Anaerococcus SPP: Is this an underdiagnosed infection?

The objectives of this study were to report 10 episodes of clinically significant bacteremia caused by species of the genus Anaerococcus isolated between July 2018 and February 2021 from the microbiology laboratory of a tertiary hospital in Granada (Spain). None of the isolates were identified by MALDI-TOF MS, and the definitive species identification was performed by 16 S rRNA gene sequencing. No reference spectra of the Anaerococcus species were present in the MALDI-TOF MS database. Eight isolates were finally identified as A. octavius, one isolate as A. tetradius and the other as A. urinomassiliensis. The majority of these infections were seen in patients aged >70 years. Risk factors for anaerobic infection were observed in eight patients, especially diabetes mellitus, surgery, and the presence of cancer. Fever was present in all patients. Three patients died, but only one death was attributed to the infection. Mean detection time of positive blood cultures was 47.5 h (range 24–92 h). Antimicrobial susceptibility to penicillin, amoxicillin-clavulanate, imipenem, moxifloxacin, clindamycin, metronidazole, and piperacillin-tazobactam was tested using the gradient diffusion technique and EUCAST breakpoints (except for moxifloxacin). No resistance to amoxicillin-clavulanate, metronidazole, imipenem, or piperacillin-tazobactam was detected; however, the majority of isolates were resistant to clindamycin.When MALDI-TOF MS does not provide a correct identification at genus or species level, as in some isolates of Gram-positive anaerobic cocci, microbiologists should perform an additional confirmatory technique, such as gene sequencing analysis, to obtain a definitive diagnosis.

Continuous Infusion of Piperacillin/Tazobactam and Meropenem in ICU Patients Without Renal Dysfunction: Are Patients at Risk of Underexposure?

Morbidity and mortality from serious infections are common in intensive care units(ICUs). The appropriateness of the antibiotic treatment is essential to combat sepsis. We aimed to evaluate pharmacokinetic/pharmacodynamic target attainment of meropenem and piperacillin/tazobactam administered at standard total daily dose as continuous infusion in critically ill patients without renal dysfunction and to identify risk factors of non-pharmacokinetic/pharmacodynamic target attainment. We included 118 patients (149 concentrations), 47% had microorganism isolation. Minimum inhibitory concentration (MIC)[median (interquartile range, IQR) values in isolated pathogens were: meropenem: 0.05 (0.02-0.12) mg/l; piperacillin: 3 (1-4) mg/l]. Pharmacokinetic/pharmacodynamic target attainments (100%fCss≥1xMIC, 100%fCss≥4xMIC and 100%fCss ≥ 8xMIC, respectively) were: 100%, 96.15%, 96.15% (meropenem) and 95.56%, 91.11%, 62.22% (piperacillin) for actual MIC; 98.11%, 71.70%, 47.17% (meropenem, MIC 2 mg/l), 95.83%, 44.79%, 6.25% (piperacillin, MIC 8 mg/l), 83.33%, 6.25%, 1.04% (piperacillin, MIC 16 mg/l) for EUCAST breakpoint of Enterobacteriaceae spp. and Pseudomonas spp. Multivariable linear analysis identified creatinine clearance (CrCL) as a predictive factor of free antibiotic concentrations (fCss) of both therapies (meropenem [β = - 0.01 (95% CI - 0.02 to - 0.0; p = 0.043)] and piperacillin [β = - 0.01 (95% CI - 0.02 to 0.01, p < 0.001)]). Neurocritical status was associated with lower piperacillin fCss [β = - 0.36 (95% CI - 0.61 to - 0.11; p = 0.005)]. Standard total daily dose of meropenem allowed achieving pharmacokinetic/pharmacodynamic target attainmentsin ICU patients without renal dysfunction. Higher doses of piperacillin/tazobactam would be needed to cover microorganisms with MIC > 8 mg/l. CrCL was the most powerful factor predictive of fCss in both therapies.

Prevalence and Phenotypic and Genotypic Resistance Mechanisms of Multidrug-Resistant Pseudomonas aeruginosa Strains Isolated from Clinical, Environmental, and Poultry Litter Samples from the Ashanti Region of Ghana.

Background Antibiotic resistance in bacteria is a major global health challenge. Reports on the prevalence of multidrug-resistant P. aeruginosa, a common pathogenic bacterium implicated in nosocomial infections and poultry diseases, are limited in Ghana. This study therefore sought to determine the prevalence of P. aeruginosa from hospitals, poultry farms, and environmental samples from the Ashanti region of Ghana. Methodology. Stool, urine, and blood samples from 364 patients from two hospitals in the Ashanti region of Ghana were randomly sampled. P. aeruginosa was isolated and confirmed using routine selective media and PCR-based oprL gene amplification. The Kirby-Bauer disk diffusion method employing EUCAST breakpoint values was used to identify multidrug-resistant strains. The occurrence of common antibiotic inactivating enzymes and resistance encoding genes and the assessment of strain efflux capacity were investigated with double disc synergy test (DDST), imipenem-EDTA synergy test, phenylboronic acid test, D-test, routine PCR, and ethidium bromide agar-cartwheel method. Results A total of 87 (9.7%, n = 87/900) P. aeruginosa isolates were confirmed from the samples. 75% (n = 65/87) were resistant to more than one group of antipseudomonal agents, while 43.6% (n = 38/87) were multidrug-resistant (MDR). High prevalence of extended spectrum β-lactamases (84.2%), metallo-β-lactamases (34.1%), and AmpC inducible cephalosporinases (50%) was observed in the MDR strains. About 57.8% of the MDR strains showed moderate to very high efflux capacity. Class 1 integrons were detected in 89.4% of the MDR isolates but β-lactamase encoding genes (blaSHV, blaTEM, blaCTX-M, blaVIM, and blaIMP) were not detected. Conclusion Surveillance of antibiotic-resistant strains of bacteria should be routinely conducted in clinical and veterinary practice in Ghana to inform selection of antibiotics for therapeutic use.

Elimination of fosfomycin during dialysis with the Genius system in septic patients

To assess fosfomycin (FOS) elimination in patients with sepsis and acute kidney injury (AKI) undergoing slow-extended daily dialysis (SLEDD) with the Genius system in a prospective observational study. After ethics committee approval ten patients with sepsis and AKI stage 3 underwent daily SLEDD sessions of eight hours. FOS was applied i.v. at doses of 3 × 5 g per day. FOS serum levels were measured pre- and post hemofilter before, during, and after SLEDD sessions, and instantaneous clearance was calculated. In five of the patients, we analyzed FOS levels after the first dose, in the other five patients serum levels were measured during ongoing therapy. FOS was eliminated rapidly via the hemofilter. FOS clearance decreased from 152 ± 10 mL/min (start of SLEED session) to 43 ± 38 mL/min (end of SLEDD session). In 3/5 first-dose patients after 4–6 h of SLEDD the FOS serum level fell below the EUCAST breakpoint of 32 mg/L for Enterobacterales and Staphylococcus species. In all patients with ongoing fosfomycin therapy serum levels were high and above the breakpoint at all times. FOS toxicity or adverse effects were not observed. FOS serum concentrations exhibit wide variability in critically ill patients with sepsis and AKI. FOS is eliminated rapidly during SLEDD. A loading dose of 5 g is not sufficient to achieve serum levels above the EUCAST breakpoint for common bacteria in all patients, considering that T > MIC > 70% of the dosing interval indicates sufficient plasma levels. We thus recommend a loading dose of 8 g followed by a maintenance dose of 5 g after a SLEDD session in anuric patients. We strongly recommend therapeutic drug monitoring of FOS levels in critically ill patients with AKI and dialysis therapy.

Evaluation of the Vitek 2, Phoenix, and MicroScan for Antimicrobial Susceptibility Testing of Stenotrophomonas maltophilia.

Stenotrophomonas maltophilia causes high-mortality infections in immunocompromised hosts with limited therapeutic options. Many U.S. laboratories rely on commercial automated antimicrobial susceptibility tests (cASTs) and use CLSI breakpoints (BPs) for S. maltophilia. However, contemporary data on these systems are lacking. We assessed performance of Vitek 2, MicroScan WalkAway, and Phoenix relative to that of reference broth microdilution for trimethoprim-sulfamethoxazole (SXT), levofloxacin (LEV), minocycline (MIN), and ceftazidime (CAZ) with 109 S. maltophilia bloodstream isolates. Using CLSI breakpoints, categorical agreement (CA) was below 90% on all systems and drugs, with the exception of SXT by MicroScan (98.1%) and Phoenix (98.1%) and MIN by MicroScan (100%) and Phoenix (99.1%). For SXT, Vitek 2 yielded a 77.1% CA. LEV and CAZ CA ranged from 67% to 85%. Very major errors (VME) were >3% for SXT (MicroScan, Phoenix), LEV (MicroScan), and CAZ (all systems). Major errors (ME) were >3% for SXT (Vitek 2), LEV (Phoenix), and CAZ (MicroScan, Phoenix). Minor errors were >10% for CAZ and LEV on all systems. Data were analyzed with EUCAST pharmacokinetic/pharmacodynamic CAZ, LEV, ciprofloxacin (CIP), and tigecycline (TGC) breakpoints when possible. CA was <90% for all. VME were >3% for CAZ (all systems), LEV (MicroScan), and TGC (Vitek 2), and ME were >3% for LEV (MicroScan), CAZ (all systems), ciprofloxacin (Vitek 2 and MicroScan), and TGC (Vitek 2, Phoenix). Minor errors (MI) were >10% for all agents and systems, by EUCAST breakpoints with an intermediate category (LEV, CAZ, CIP). Laboratories should use caution with cASTs for S. maltophilia, as a high rate of errors may be observed.

In vitro activity of ceftaroline and comparators against bacterial isolates collected globally from patients with skin infections

This study reports the antimicrobial activities of ceftaroline and comparators against bacterial isolates from patients with skin and skin-structure infections (2015-2018). A central laboratory performed antimicrobial susceptibility testing according to CLSI broth microdilution methodology. EUCAST breakpoints were used. Isolates were collected in Europe (14 408 isolates; 53.9%), Asia/South Pacific (SP) (5317; 19.9%), Latin America (4268; 16.0%) and Africa/Middle East (ME) (2753; 10.3%). In all regions, all 7950 methicillin-susceptible Staphylococcus aureus (MSSA) isolates were susceptible to ceftaroline and vancomycin; susceptibility to daptomycin, linezolid, teicoplanin and tigecycline was ≥99.6%. Susceptibility of all 9174 methicillin-resistant S. aureus (MRSA) isolates to daptomycin, linezolid, teicoplanin, tigecycline and vancomycin was ≥97.7%, with 90.8-96.5% susceptible to ceftaroline. The ceftaroline MIC90 was 0.008 mg/L against Streptococcus pyogenes, 0.015-0.03 mg/L against Streptococcus agalactiae and 0.008-0.015 mg/L against Streptococcus dysgalactiae. All β-haemolytic streptococci were susceptible to vancomycin. Susceptibility of extended-spectrum β-lactamase (ESBL)-negative Escherichia coli to ceftaroline ranged from 67.0% in Asia/SP to 91.0% in Africa/ME; susceptibility to amikacin, meropenem and tigecycline was ≥96.7% in all regions. Susceptibility of ESBL-negative Klebsiella pneumoniae to ceftaroline ranged from 78.4% in Europe to 83.2% in Africa/ME, and among ESBL-negative Klebsiella oxytoca was 76.3% in Asia/SP and 89.0-93.5% in other regions. Among ESBL-negative K. pneumoniae and ESBL-negative K. oxytoca, susceptibility was highest to amikacin (93.7-96.4% and 95.7-100%, respectively) and meropenem (89.7-97.4% and 98.3-100%, respectively). Ceftaroline was active against the Gram-positive isolates collected. Susceptibility of ESBL-negative Gram-negative isolates showed regional variations.

In vitro activity of cefiderocol and comparators against isolates of Gram-negative pathogens from a range of infection sources: SIDERO-WT-2014–2018 studies in Italy

Antimicrobial resistance, particularly carbapenem resistance, in Gram-negative pathogens poses a significant healthcare threat. Carbapenem resistance rates in Italy are among the highest in Europe. We report the in vitro activity of cefiderocol, a novel siderophore cephalosporin, and comparator antibiotics against Gram-negative isolates from Italy as part of the SIDERO-WT studies. Isolates were collected between 2014 and 2018. Minimum inhibitory concentrations (MICs) were determined using International Organization for Standardization and EUCAST guidelines. Antimicrobial susceptibilities were interpreted using EUCAST breakpoints; pharmacodynamic/pharmacokinetic breakpoints were used if EUCAST breakpoints were not specified. The 2472 isolates [1545 (62.5%) Enterobacterales and 927 (37.5%) non-fermenters] represented a range of infection sources, including nosocomial pneumonia (902; 36.5%), complicated urinary tract infection (374; 15.1%), bloodstream infection (596; 24.1%), complicated intra-abdominal infection (257; 10.4%) and other infection sources (343; 13.9%). Cefiderocol was active against the majority of isolates, regardless of infection source (susceptibility, 94.2-97.3%). A high proportion of non-fermenters (97.6%) and Enterobacterales (95.6%) were cefiderocol-susceptible, although susceptibility was lower in Klebsiella pneumoniae (88.1%). Susceptibility to cefiderocol was significantly (P < 0.01) greater than comparators overall (96.4% vs. 71.3-81.6%) and in non-fermenters (97.6% vs. 44.3-90.3%) across infection sources. Overall 612/2472 isolates (24.8%) were meropenem-resistant (MIC > 8 mg/L), comprising 516/927 (55.7%) non-fermenters and 96/1545 (6.2%) Enterobacterales. Cefiderocol (499/516; 96.7%) activity was greater than colistin (440/516; 85.3%), ceftazidime/avibactam (123/516; 23.8%) and ceftolozane/tazobactam (89/516; 17.2%) in meropenem-resistant non-fermenter isolates. Susceptibility to cefiderocol was significantly greater than meropenem, colistin, ceftazidime/avibactam and ceftolozane/tazobactam overall, regardless of infection source.

Pharmacokinetic/pharmacodynamic evaluation of the antimicrobial therapy of pneumococcal invasive disease in adults in post-PCV13 vaccine period in Madrid, Spain.

The objective of our study was to evaluate by pharmacokinetic/pharmacodynamic (PK/PD) analysis, if the antimicrobials used for the treatment of invasive pneumococcal disease (IPD) in adults, including meningitis, are adequate considering the susceptibility profile of S. pneumoniae in Spain after the implantation of PVC13 vaccine. Pharmacokinetic parameters of benzylpenicillin and cefotaxime were obtained from the literature, and susceptibility data of invasive S. pneumoniae strains recovered in 2017 (post-PCV13 vaccination period) were provided by the Public Health Regional Laboratory of Madrid. We have also studied levofloxacin because it is used to treat pneumococcal pneumonia previously to be diagnosed as bacteremic pneumonia. Monte Carlo simulation was used to estimate the probability of target attainment (PTA) and the cumulative fraction of response (CFR). All doses of benzylpenicillin except 2 mU q6h provide a high probability of treatment success for MIC values ≤ 1 mg/L; 4 mU q4h is even useful for MIC values up to 4 mg/L. This high dose, used for the treatment of meningitis, also provides high probability of treatment success for MIC ≤ 0.5 mg/L. At the susceptibility EUCAST breakpoint (≤ 0.5 mg/L), cefotaxime provides a high rate of PD target achievement, even at the lowest dose (1 g q8h). For meningitis, 2 g q6h ensures probabilities of target attainment ≥90% for MIC up to 1 mg/L. Our study confirms that after the implementation of PCV13 vaccine, the treatment with benzylpenicillin and cefotaxime provides high probability of the therapy success of IPD, including meningitis.

Comparison of antimicrobial susceptibility interpretation among Enterobacteriaceae using CLSI and EUCAST breakpoints

PurposeTo determine the difference in antimicrobial susceptibility of various antibiotics using the CLSI & EUCAST breakpoints. MethodsIn this non interventional, retrospective observational study, we reviewed minimum inhibitory concentrations (MIC) of various antibiotics routinely reported for Enterobacteriaceae clinical isolates, from an automated microbiology identification system (VITEK-2). These MICs were then analysed using both CLSI 2019 and EUCAST 2019 guidelines and classified as per the breakpoints into various categories. ResultsThe concordance rates of the antimicrobial susceptibility for various drugs ranged from 78.2% to 100% among two breakpoints. Perfect agreement with κ = 1 (p < 0.001) was observed for only three antimicrobials ceftriaxone, levofloxacin and trimethoprim-sulfamethoxazole. The changes in antimicrobial susceptibility interpretation for cefepime, ciprofloxacin, amoxicillin clavulanic acid was majorly in Intermediate category. ConclusionThe change in interpretation of the susceptibility will lead to change in the usage of antibiotics especially due to recent change in definition of I by EUCAST. There is need of more studies in this aspect to ascertain clinical implication of change in antimicrobial susceptibility.

In vitro activity of cefiderocol and comparators against isolates of Gram-negative pathogens from a range of infection sources: SIDERO-WT-2014-2018 studies in France.

ObjectivesOver recent years, France has experienced an increase of infections caused by carbapenem-resistant Gram-negative (GN) pathogens. Cefiderocol is approved in Europe for the treatment of aerobic GN infections in adults with limited treatment options. This study evaluated the in vitro activity of cefiderocol and comparators against GN clinical isolates from France.MethodsMICs were determined by broth microdilution, according to International Organization for Standardization guidelines. Cefiderocol was tested using iron-depleted CAMHB. Susceptibility rates were based on EUCAST breakpoints. In the absence of a species-specific breakpoint, pharmacokinetic/pharmacodynamic breakpoints were used.ResultsOf 2027 isolates, 1344 (66.3%) were Enterobacterales and 683 (33.7%) were non-fermenters. The most common pathogen was Pseudomonas aeruginosa (16.8%), followed by Escherichia coli (16.0%), Klebsiella pneumoniae (13.1%), Acinetobacter baumannii (7.9%) and Stenotrophomonas maltophilia (5.1%). Isolates represented a range of infection sources including nosocomial pneumonia (33.6%), complicated urinary tract infection (24.3%), bloodstream infection (13.1%) and complicated intra-abdominal infection (18.0%). In total, 135/2027 (6.7%) isolates were meropenem resistant (MIC >8 mg/L); 133/135 (98.5%) were non-fermenters. Overall, 1330/1344 (99.0%) Enterobacterales and 681/683 (99.7%) non-fermenters were cefiderocol susceptible, including 100% of meropenem-resistant S. maltophilia (n = 98) and P. aeruginosa (n = 18) isolates. Susceptibility to cefiderocol was significantly higher (P < 0.01) in nosocomial pneumonia isolates (681/682 [99.9%]) than susceptibility to meropenem (586/682 [85.9%]), ceftolozane/tazobactam (593/682 [87.0%]), ceftazidime/avibactam (612/682 [89.7%]) and colistin (538/682 [78.9%]).ConclusionsCefiderocol demonstrated high in vitro susceptibility rates against a wide range of Gram-negative pathogens, including meropenem-resistant strains, and was significantly more active than comparators against pneumonia isolates.

In vitro activity of ceftazidime/avibactam against clinical isolates of Enterobacterales and Pseudomonas aeruginosa from Middle Eastern and African countries: ATLAS global surveillance programme 2015–18

ObjectivesTo assess the in vitro activity of ceftazidime/avibactam against a recent, 2015–18, collection of clinical isolates of Gram-negative bacilli from Middle Eastern and African countries with a focus on isolates from ICUs and with MDR and difficult-to-treat resistance (DTR) phenotypes.MethodsAntimicrobial susceptibility testing of 4608 isolates of Enterobacterales (997 isolates from ICU patients) and 1358 isolates of Pseudomonas aeruginosa (374 isolates from ICU patients) was performed by CLSI broth microdilution methodology in a central laboratory. MICs were interpreted using both CLSI (2020) and EUCAST (2020) MIC breakpoints.ResultsMost isolates of Enterobacterales (Middle East: ICU, 99.1% susceptible, non-ICU, 99.1%; Africa: ICU, 96.9% susceptible, non-ICU, 98.3%) and P. aeruginosa (Middle East: ICU, 93.4%, non-ICU, 92.1%; Africa: ICU, 89.8%; non-ICU, 94.1%) were susceptible to ceftazidime/avibactam. Applying CLSI and EUCAST breakpoints, MDR rates were similar for Enterobacterales (27.8%–36.0% of isolates) and P. aeruginosa (25.0%–36.4%) while DTR rates were lower for Enterobacterales (1.6%–1.8%) than for P. aeruginosa (5.2%–7.4%). Percentage susceptible rates for ceftazidime/avibactam for MDR Enterobacterales were 96.8%–97.5% (Middle East) and 92.5%–94.3% (Africa) while rates for P. aeruginosa were 70.1%–80.0% (Middle East) and 69.5%–78.2% (Africa). 60.5%–65.8% (Middle East) and 38.9%–52.2% (Africa) of isolates of Enterobacterales with DTR phenotypes were ceftazidime/avibactam susceptible as were 29.2%–31.1% (Middle East) and 28.2%–35.8% (Africa) of DTR P. aeruginosa.ConclusionsOverall, the isolates of Enterobacterales and P. aeruginosa tested from Middle Eastern and African countries were highly susceptible to ceftazidime/avibactam. Most MDR and many DTR isolates of Enterobacterales and P. aeruginosa were susceptible to ceftazidime/avibactam.

False amoxicillin/clavulanic acid susceptibility in Bacteroides fragilis using gradient strip tests

BackgroundRepeatedly, too low MIC results were obtained in Bacteroides fragilis quality assessment strains, using gradient strip tests with a ratio of amoxicillin:clavulanic acid of 2:1. We aimed to find the most accurate available gradient strip tests for susceptibility testing of amoxicillin/clavulanic acid in B. fragilis in comparison with agar dilution with EUCAST methodology and breakpoints. MethodsTwenty-seven clinical B. fragilis isolates were investigated using gold standard EUCAST amoxicillin/clavulanic acid agar dilution (fixed clavulanic acid concentration at 2 mg/L, with increasing amoxicillin concentrations) as well as three commercial gradient strip tests: XL (ratio), AUG (ratio) or AMC (fixed concentration). ResultsUsing agar dilution (fixed concentration), 19 isolates were susceptible, 1 isolate was susceptible increased exposure (I) and 7 isolates were resistant. Categorical agreement of the gradient strip tests with agar dilution (fixed concentration) was 70% for XL (ratio), 71% for AUG (ratio) and 89% for AMC (fixed concentration). Very major error rates in comparison with agar dilution (fixed concentration) were 100%, 0%, and 0%, respectively. ConclusionsEUCAST breakpoint usage in amoxicillin/clavulanic acid susceptibility tests for B. fragilis should be accompanied by EUCAST methodology. When using alternative methods such as gradient strip tests, a higher degree of alignment with EUCAST methodology, such as using fixed clavulanic acid concentrations, improves precision.

Antimicrobial susceptibility testing of Eggerthella lenta blood culture isolates at a university hospital in Belgium from 2004 to 2018

ObjectivesEggerthella lenta is a Gram-positive anaerobic bacillus that is an important cause of bloodstream infections. This study aims to test the susceptibility of Eggerthella lenta blood culture isolates to commonly used antibiotics for the empirical treatment of anaerobic infections. MethodsIn total, 49 positive blood cultures for Eggerthella lenta were retrospectively included from patients hospitalised at the Universitair Ziekenhuis Brussel, Belgium, between 2004 and 2018. Identification was done by matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) system. Antimicrobial susceptibility testing was performed using the reference agar dilution method according to Clinical and Laboratory Standards Institute (CLSI) guidelines with Brucella agar supplemented with 5 μg/mL hemin, 1 μg/mL vitamin K1 and 5% laked sheep blood.The minimal inhibitory concentrations were interpreted using the EUCAST breakpoints. Clinical characteristics were collected by reviewing the patient’s medical records. ResultsAll isolates were susceptible to amoxicillin-clavulanate, metronidazole and meropenem. Eighty-eight % of them were susceptible to clindamycin and 94% (20% S, 74% I) were susceptible to piperacillin-tazobactam. The mean age of the patients was 64 (±20) and they showed a 30-day mortality of 27%. The source of infection was in 65.3% of the cases abdominal, 20.4% were sacral pressure ulcers and 14.3% were unknown causes.While all isolates were fully susceptible at standard dosing regimen to amoxicillin-clavulanate, most were only susceptible at increased exposure or resistant to piperacillin-tazobactam. ConclusionsOur results suggest to be careful with the use of piperacillin-tazobactam and clindamycin in the empirical treatment of Eggerthella lenta infections.

Reporting antimicrobial susceptibilities and resistance phenotypes in Staphylococcus spp.: a nationwide proficiency study.

ObjectivesTo evaluate the proficiency of microbiology laboratories in Spain in antimicrobial susceptibility testing (AST) of Staphylococcus spp.Materials and methodsEight Staphylococcus spp. with different resistance mechanisms were selected: six Staphylococcus aureus (CC-01/mecA, CC-02/mecC, CC-03/BORSA, CC-04/MLSBi, CC-06/blaZ and CC-07/linezolid resistant, cfr); one Staphylococcus epidermidis (CC-05/linezolid resistant, 23S rRNA mutation); and one Staphylococcus capitis (CC-08/daptomycin non-susceptible). Fifty-one laboratories were asked to report: (i) AST system used; (ii) antimicrobial MICs; (iii) breakpoints used (CLSI or EUCAST); and (iv) clinical category. Minor, major and very major errors (mEs, MEs and VMEs, respectively) were determined.ResultsThe greatest MIC discrepancies found were: (i) by AST method: 19.4% (gradient diffusion); (ii) by antimicrobial agent: daptomycin (21.3%) and oxacillin (20.6%); and (iii) by isolate: CC-07/cfr (48.0%). The greatest error rates were: (i) by AST method: gradient diffusion (4.3% and 5.1% VMEs, using EUCAST and CLSI, respectively); (ii) by breakpoint: 3.8% EUCAST and 2.3% CLSI; (iii) by error type: mEs (0.8% EUCAST and 1.0% CLSI), MEs (1.8% EUCAST and 0.7% CLSI) and VMEs (1.2% EUCAST and 0.6% CLSI); (iii) by antimicrobial agent: VMEs (4.7% linezolid and 4.3% oxacillin using EUCAST); MEs (14.3% fosfomycin, 9.1% tobramycin and 5.7% gentamicin using EUCAST); and mEs (22.6% amikacin using EUCAST).ConclusionsClinical microbiology laboratories should improve their ability to determine the susceptibility of Staphylococcus spp. to some antimicrobial agents to avoid reporting false-susceptible or false-resistant results. The greatest discrepancies and errors were associated with gradient diffusion, EUCAST breakpoints and some antimicrobials (mEs for aminoglycosides; MEs for fosfomycin, aminoglycosides and oxacillin; and VMEs for linezolid and oxacillin).

Bir Üniversite Hastanesinde Kan Kültürlerinden İzole Edilen Enterobacterales Takımı Üyelerinin Dağılımının ve Antimikrobiyal Duyarlılıklarının İncelenmesi

Objective: Members of Enterobacterales can cause various diseases in humans. The objective of this study was to determine the genus/species distribution and antimicrobial susceptibilities of Enterobacterales isolated from blood cultures in Central Laboratory of Hacettepe University Hospital. Method: Enterobacterales isolated from blood between July-2014 and April-2018 were included in the study. MALDI-TOF MS was used for the identification of isolates. Antimicrobial susceptibilities were determined with automated system (VITEK 2 Compact for the isolates between 2014 and 2018; BD Phoenix for the isolates in 2018) and disk diffusion method. Results of antimicrobial susceptibility testing were interpreted according to EUCAST breakpoints. Results: In total, 1765 isolates belonging to the order Enterobacterales were isolated from blood cultures. The most common microorganisms were Escherichia coli (47.6%), Klebsiella (34.1%), Enterobacter (6%), Proteus (4.4%) and Serratia spp. (3.5%), respectively. The remaining isolates included Salmonella, Citrobacter, M. morganii, Pantoea, Raoultella and Providencia spp. The lowest resistance rates among E. coli, Klebsiella and Enterobacter spp. isolates were observed against meropenem and amikacin. However, 21.1% of Klebsiella isolates were resistant to meropenem. The most active antimicrobials against Proteus isolates were piperacillintazobactam and meropenem. Resistance was not observed against piperacillin-tazobactam and meropenem among Proteus isolates. The most active antimicrobial against Serratia isolates was trimethoprim/ sulfamethoxazole with a resistance rate of 0%. Resistance was not noted against ampicillin and trimethoprim/ sulfamethoxazole among Salmonella isolates, whereas 26.1% of isolates were resistant to ciprofloxacin. All Citrobacter isolates were susceptible to meropenem, amikacin and cefepime. Conclusion: Findings of our study may guide the selection of proper antimicrobials for the treatment of bacteremia caused by Enterobacterales. Furthermore, this study provides important epidemiological information regarding the distribution of members of Enterobacterales causing bacteremia.

Evaluation of a Commercial Broth Microdilution Panel for Colistin Susceptibility Testing of Clinical Isolates of Escherichia coli and Klebsiella pneumoniae.

Colistin is among the last resort antibiotics for the treatment of infections caused by multidrug-resistant Gram-negative pathogens. Antimicrobial susceptibility testing of colistin is challenging due to its physicochemical properties. Broth microdilution (BMD) is the recommended method for colistin susceptibility testing. However BMD is not practical for clinical microbiology laboratories as manual preparation of BMD plates is time-consuming and labor intensive. Recently, some more user-friendly BMD products with commercial panels have become available. Our objective was to evaluate the performance of a commercial broth microdilution (BMD) product [Sensititre (Thermo Fisher Scientific)] for colistin MIC determination by comparison with reference BMD method using a collection of E. coli and K. pneumoniae isolates. A total of 323 unique patient isolates (102 E. coli, 221 K. pneumoniae) were included in the study. Isolates were stored at -70°C and subcultured twice on sheep blood agar before testing. Colistin MICs of the isolates were determined using Sensititre (a premade BMD product with dried antibiotics) and an 'in-house prepared BMD panel prepared in accordance with CLSI guidelines' (reference method). MIC determination with Sensititre was performed according to manufacturer's instructions. The reference method was performed using untreated 96-well sterile polystyrene plates. Colistin MIC results were interpreted according to EUCAST breakpoints (susceptible, ≤ 2 mg/L; resistant, > 2 mg/L). Overall susceptibility rate of isolates to colistin by reference BMD was 75.9%. Overall categorical agreement (CA), essential agreement (EA), very major error (VME), and major error (ME) rates for Sensititre were 98.5%, 72.5%, 3.8%, and 0.8%, respectively. The CA and EA between Sensititre and reference BMD for the isolates with reference colistin MICs close to the susceptibility breakpoint (2 - 8 mg/L) was 94.2% and 48.1%, respectively. Sensititre yielded a VME rate of 15% and ME rate of 0%, respectively, for this subset of isolates. In conclusion, Sensititre showed high CA but low EA with reference BMD for entire collection of isolates. The VME rate was just slightly above 3% and ME rate was acceptable. The rates of CA and EA were decreased and the rate of VME was increased when a subset consisting of more challenging isolates was used.

1296. Efficacy of Human-Simulated Exposures of Meropenem/Vaborbactam (MVB) and Meropenem (MEM) against OXA-48 β-lactamase-producing Enterobacterales in the Neutropenic Murine Thigh Infection Model

BackgroundOXA-48 exhibits variable hydrolytic activity toward carbapenems, with imipenem and meropenem MICs, though increased, often reporting within the ‘susceptible’ or ‘intermediate’ ranges defined by CLSI and EUCAST. Although vaborbactam (VAB) does not enhance MEM activity against OXA-48, ~ 1/3 of OXA-48-producing Enterobacterales will test susceptible to MVB due to its higher breakpoint. Clinical implications of this discordance warrant investigation.Methods26 isolates harboring OXA-48 (n=24) and KPC (n=2) were evaluated in the neutropenic murine thigh model. MICs were determined in triplicate per CLSI. Human-simulated regimens of MVB (simulating doses of 2-2 g IV q 8 hours (h) over 3 h) and MEM (2 g IV q 8 h over 3 h) were administered resulting in similar f%T >MIC and fAUC as humans for MEM and VAB, respectively. Mice received MVB, MEM, or sham control for 24 h. Efficacy was assessed on the resulting overall change in mean± SD log10 CFU/thigh as well as the achievement of ≥ 1 log10 reduction as an established surrogate marker predictive of success for serious infections.ResultsMVB and MEM MICs ranged from 1- 64 and 2 - > 64 mg/L, respectively. Relative to 0 h control, the mean bacterial growth (mean ± SD, CFU/thigh) at 24 h in the untreated control mice was 2.69 ± 1.31. As anticipated for KPCs, MVB resulted in a mean bacterial reduction ≥ 1 log10 (-1.10 ± 0.26), whereas growth was observed on MEM (+1.45 ± 0.88). For all OXA-48 isolates, MVB resulted in variable bacterial densities ranging from -2.54 to +2.68, similarly MEM resulted in -2.18 to +2.66. Addition of vaborbactam did not enhance MEM activity for any isolate. For isolates with MVB MICs ≥ 16 (n=5), 8 (n=5, EUCAST breakpoint), 4 (n=9, CLSI breakpoint), and ≤ 2 (n=5) mg/L, 0%, 0%, 44%, and 60% of isolates treated with MVB or MEM achieved the target reduction of ≥ 1 log10 kill, respectively.ConclusionAcross the range of MICs evaluated, MVB and MEM humanized exposures in vivo resulted in similar reductions and growth in bacterial density for OXA-48-producing Enterobacterales. Moreover, these data highlight the poor efficacy of MVB for OXA-48 defined as susceptible using the current EUCAST and CLSI susceptibility criterion. Caution is therefore warranted when treating Enterobacterales testing susceptible to MVB without the genotypic profile.Disclosures David P. Nicolau, PharmD, Cepheid (Other Financial or Material Support, Consultant, speaker bureau member or has received research support.)Merck & Co., Inc. (Consultant, Grant/Research Support, Speaker’s Bureau)Wockhardt (Grant/Research Support)