Introduction. Aminoglycoside antibiotics such as amikacin and kanamycin are important components in the treatment of Mycobacterium tuberculosis (Mtb) infection. However, more and more clinical strains are found to be aminoglycoside antibiotic-resistant. Apramycin is another kind of aminoglycoside antibiotic that is commonly used to treat infections in animals.Hypothesis. Apramycin may have in vitro activity against Mtb.Aim. This study aims to evaluate the efficacy of apramycin against Mtb in vitro and determine its epidemiological cut-off (ECOFF) value.Methodology. One hundred Mtb isolates, including 17 pansusceptible and 83 drug-resistant tuberculosis (DR-TB) strains, were analysed for apramycin resistance using the MIC assay.Results. Apramycin exhibited significant inhibitory activity against Mtb clinical isolates, with an MIC50 of 0.5 μg ml-1 and an MIC90 of 1 μg ml-1. We determined the tentative ECOFF value as 1 µg ml-1 for apramycin. The resistant rates of multidrug-resistant tuberculosis (MDR-TB), pre-extensively drug-resistant (pre-XDR-TB) and extensively drug-resistant tuberculosis (XDR-TB) strains were 12.12 % (4/33), 20.69 % (6/29) and 66.67 % (14/21), respectively. The rrs gene A1401G is associated with apramycin resistance, as well as the cross-resistance between apramycin and other aminoglycosides.Conclusion. Apramycin shows high in vitro activity against the Mtb clinical isolates, especially the MDR-TB clinical isolates. This encouraging discovery calls for more research on the functions of apramycin in vivo and as a possible antibiotic for the treatment of drug-resistant TB.

OBJECTIVE: Bacillus anthracis clinical breakpoints, representing a systematic approach to guide clinicians in selecting the most appropriate antimicrobial treatments, are not part of the guidance from the European Committee on Antimicrobial Susceptibility Testing (EUCAST). This is because defined distributions of MIC values and of epidemiological cut-off values (ECOFFs) have been lacking. In this study, a Europe-wide network of laboratories in collaboration with EUCAST, aimed at establishing standardized antimicrobial susceptibility testing methods, wild-type MIC distributions, and ECOFFs for ten therapeutically relevant antimicrobials.
METHODS: About 335 B. anthracis isolates were tested by broth microdilution and disc diffusion methodologies. MIC and inhibition zone diameters were curated according to EUCAST SOP 10.2 and the results were submitted to EUCAST for ECOFFs and clinical breakpoint determination.
RESULTS: Broth microdilution and disc diffusion data distributions revealed putative wild-type distributions for the tested agents. For each antimicrobial agent, ECOFFs were defined. Three highly resistant strains with MIC values of 32 mg/L benzylpenicillin were found. MIC values slightly above the defined ECOFFs were observed in a few isolates, indicating the presence of resistance mechanisms to doxycycline, tetracycline, and amoxicillin.
CONCLUSIONS: B. anthracis antimicrobial susceptibility testing results were used by EUCAST to determine ECOFFs for ten antimicrobial agents. The MIC distributions were used in the process of determining clinical breakpoints. The ECOFFs can be used for the sensitive detection of isolates with resistance mechanisms, and for monitoring resistance development. Genetic changes causing phenotypic shifts in isolates displaying slightly elevated MICs remain to be investigated.

The characterization of wild-type minimum inhibitory concentration (MIC) and zone diameter distributions with the setting of epidemiological cut-off values (ECOFFs or ECVs) provides a reference for the otherwise relative MIC values in the international system for antimicrobial susceptibility testing. Distributions of MIC values for a species and an agent follow a log-normal distribution, which in the absence of resistance mechanisms is monomodal and designated wild type (WT). The upper end of the WT distribution, the ECOFF, can be identified with statistical methods. In the presence of phenotypically detectable resistance, the distribution has at least one more mode (the non-WT), but despite this, the WT is most often identifiable using the same methods. The ECOFF provides the most sensitive measure of resistance development in a species against an agent. The WT and non-WT modes are independent of the organism´s response to treatment, but when the European Committee on Antimicrobial Susceptibility Testing (EUCAST) determines the clinical breakpoints, the committee avoids breakpoints that split WT distributions of target species. This is to avoid the poorer reproducibility of susceptibility categorization when breakpoints split major populations but also because the EUCAST has failed to identify different clinical outcomes for isolates with different MIC values inside the wild-type distribution. In laboratory practice, the ECOFF is used to screen for and exclude resistance and allows the comparison of resistance between systems with different breakpoints from different breakpoint organizations, breakpoints evolving over time, and different breakpoints between human and animal medicine. The EUCAST actively encourages colleagues to question MIC distributions as presented on the website (https://www.eucast.org/mic_and_zone_distributions_and_ecoffs) and to contribute MIC and inhibition zone diameter data.

UNASSIGNED: Minimum inhibitory concentration (MIC) values are useful in guiding appropriate antimicrobial therapy however, routine provision and interpretation of MIC values to guide clinical decision-making is challenging.
UNASSIGNED: This proof of concept study aims to demonstrate the clinical utility and application of Vitek®-derived MIC values through categorisation of clinical isolates as wild type.
UNASSIGNED: A random selection of clinically relevant Gram negative isolates routinely tested on the Vitek® instrument were included. The Vitek® MIC values, for selected antimicrobials at the lowest calling range of that card, were compared to the broth microdilution reference method. The specified end-point was concordance between the two results if the reference MIC was less than or equal to the EUCAST-defined epidemiological cut-off value (ECOFF) for that drug-bug combination.
UNASSIGNED: A total of 525 isolates were included (468 Enterobacterales and 57 Pseudomonas aeruginosa), with an overall concordance rate of 96.4% (508/525). Correct ECOFF categorisation by the Vitek® was highest for ceftazidime and piperacillin (100%, n = 48 and n = 55, respectively) and lowest for cefepime (81.8%, n = 66).
UNASSIGNED: Vitek®-derived MIC values can be used to categorise organisms as wild-type if the MIC is reported at the card\'s lowest calling range (≤) as there is high likelihood that the MIC is at or below the ECOFF. This has important implications for antimicrobial management, assisting in choice of agent and in improving probability of target attainment for desired pharmacodynamic targets which can translate into improved clinical outcomes.
UNASSIGNED: Minimum inhibitory concentration data from an automated antimicrobial susceptibility testing instrument can be used to guide clinical decisions.

Campylobacter fetus is a pathogen, which is primarily associated with fertility problems in sheep and cattle. In humans, it can cause severe infections that require antimicrobial treatment. However, knowledge on the development of antimicrobial resistance in C. fetus is limited. Moreover, the lack of epidemiological cut-off values (ECOFFs) and clinical breakpoints for C. fetus hinders consistent reporting about wild-type and non-wild-type susceptibility. The aim of this study was to determine the phenotypic susceptibility pattern of C. fetus and to determine the C. fetus resistome [the collection of all antimicrobial resistance genes (ARGs) and their precursors] to describe the genomic basis of antimicrobial resistance in C. fetus isolates over time. Whole-genome sequences of 295 C. fetus isolates, including isolates that were isolated in the period 1939 till the mid 1940s, before the usage of non-synthetic antimicrobials, were analysed for the presence of resistance markers, and phenotypic antimicrobial susceptibility was obtained for a selection of 47 isolates. C. fetus subspecies fetus (Cff) isolates showed multiple phenotypic antimicrobial resistances compared to C. fetus subspecies venerealis (Cfv) isolates that were only intrinsic resistant to nalidixic acid and trimethoprim. Cff isolates showed elevated minimal inhibitory concentrations for cefotaxime and cefquinome that were observed in isolates from 1943 onwards, and Cff isolates contained gyrA substitutions, which conferred resistance to ciprofloxacin. Resistances to aminoglycosides, tetracycline and phenicols were linked to acquired ARGs on mobile genetic elements. A plasmid-derived tet(O) gene in a bovine Cff isolate in 1999 was the first mobile genetic element observed, followed by detection of mobile elements containing tet(O)-aph(3\')-III and tet(44)-ant(6)-Ib genes, and a plasmid from a single human isolate in 2003, carrying aph(3\')-III-ant(6)-Ib and a chloramphenicol resistance gene (cat). The presence of ARGs in multiple mobile elements distributed among different Cff lineages highlights the risk for spread and further emergence of AMR in C. fetus. Surveillance for these resistances requires the establishment of ECOFFs for C. fetus.

OBJECTIVE: For non-tuberculous mycobacteria (NTM), minimum inhibitory concentration (MIC) distributions of wild-type isolates have not been systematically evaluated despite their importance for establishing antimicrobial susceptibility testing (AST) breakpoints.
METHODS: We gathered MIC distributions for drugs used against the Mycobacterium avium complex (MAC) and Mycobacterium abscessus (MAB) obtained by commercial broth microdilution (SLOMYCOI and RAPMYCOI) from 12 laboratories. Epidemiological cut-off values (ECOFFs) and tentative ECOFFs (TECOFFs) were determined by EUCAST methodology including quality control (QC) strains.
RESULTS: The clarithromycin ECOFF was 16 mg/L for M. avium (n = 1271) whereas TECOFFs were 8 mg/L for M. intracellulare (n = 415) and 1 mg/L for MAB (n = 1014) confirmed by analysing MAB subspecies without inducible macrolide resistance (n = 235). For amikacin, the ECOFFs were 64 mg/L for MAC and MAB. For moxifloxacin, the WT spanned >8 mg/L for both MAC and MAB. For linezolid, the ECOFF and TECOFF were 64 mg/L for M. avium and M. intracellulare, respectively. Current CLSI breakpoints for amikacin (16 mg/L), moxifloxacin (1 mg/L) and linezolid (8 mg/L) divided the corresponding WT distributions. For QC M. avium and M. peregrinum, ≥95% of MIC values were well within recommended QC ranges.
CONCLUSIONS: As a first step towards clinical breakpoints for NTM, (T)ECOFFs were defined for several antimicrobials against MAC and MAB. Broad wild-type MIC distributions indicate a need for further method refinement which is now under development within the EUCAST subcommittee for anti-mycobacterial drug susceptibility testing. In addition, we showed that several CLSI NTM breakpoints are not consistent in relation to the (T)ECOFFs.

Ibrexafungerp is a novel triterpenoid antifungal that inhibits glucan synthase and thus fungal cell wall synthesis. We examined the in vitro activity against contemporary clinical yeast, investigated inter-laboratory and intra-laboratory variability, suggested wild-type upper-limit values (WT-UL), and compared in vitro activity of ibrexafungerp to five licensed antifungals. Susceptibility to ibrexafungerp and comparators was investigated prospectively for 1965 isolates (11,790 MICs) and repetitively for three QC strains (1764 MICs) following the EUCAST E.Def 7.3.2 method. Elevated ibrexafungerp/echinocandin MICs prompted FKS sequencing. Published ibrexafungerp EUCAST MIC-distributions were retrieved and aggregated for WT-UL determinations following EUCAST principles. Ibrexafungerp MICs were ≤2 mg/L except against C. pararugosa, Cryptococcus and some rare yeasts. Modal MICs (mg/L) were 0.06/0.125/0.25/0.5/0.5/0.5/0.5/1/2 for C. albicans/C. dubliniensis/C. glabrata/C. krusei/C. parapsilosis/C. tropicalis/S. cerevisiae/C. guilliermondii/C. lusitaniae and aligned within ±1 dilution with published values. The MIC ranges for QC strains were: 0.06-0.25/0.5-1/0.125-0.5 for CNM-CL-F8555/ATCC6258/ATCC22019. The WT-UL (mg/L) were: 0.25/0.5/1/1/2 for C. albicans/C. glabrata/C. krusei/C. parapsilosis/C. tropicalis. Adopting these, non-wild-type rates were 0.3%/0.6%/0%/8%/3% for C. albicans/C. glabrata/C. krusei/C. parapsilosis/C. tropicalis and overall lower than for comparators except amphotericin B. Five/six non-wild-type C. albicans/C. glabrata were echinocandin and Fks non-wild-type (F641S, F659del or F659L). Eight C. parapsilosis and three C. tropicalis non-wild-type isolates were echinocandin and Fks wild-type. Partial inhibition near 50% in the supra-MIC range may explain variable MICs. Ibrexafungerp EUCAST MIC testing is robust, although the significance of paradoxical growth for some species requires further investigation. The spectrum is broad and will provide an oral option for the growing population with azole refractory infection.

UNASSIGNED: Currently, antibiotic treatment is often a standard dosing regimen in bone and joint infections (BJI). However, it remains unknown if exposure at the target-site is adequate. The aim of this review is to gain more insight in the relationship between the target site concentration of antibiotic and the minimal inhibitory concentration to target the bacteria in bone and joint infections (BJI).
UNASSIGNED: A literature search was performed by Erasmus MC Medical library. Bone, bone tissue and synovial concentration of antibiotics were covered in humans. In addition, we reported number of patients, dose, sampling method, analytical method and tissue and plasma concentrations. We used the epidemiological cutoff value (ECOFF) values of the targeted micro-organisms. If more than 3 publications were available on the antibiotic, we graphically presented ECOFFS values against reported antibiotic concentrations.
UNASSIGNED: For most antibiotics, the literature is sparse. In addition, a lot of variable and total antibiotic concentrations are published. Ciprofloxacin, cefazolin, cefuroxime, vancomycin and linezolid seem to have adequate average exposure if correlating total concentration to ECOFF, when standard dosing is used. With regard to other antibiotics, results are inconclusive. More extensive pharmacokinetic/pharmacodynamic modeling in BJI is needed.

BACKGROUND: In the Netherlands, antimicrobial resistance (AMR) is monitored in commensal indicator Escherichia coli from healthy broilers at slaughter as part of a European monitoring programme. In a separate programme for poultry health, AMR is monitored in veterinary pathogens from diseased broilers. So far, it is unknown how the outcomes of these two AMR monitoring approaches in the same animal population are associated.
OBJECTIVE: This study aims to investigate the association between the outcomes of monitoring non-wildtype susceptibility (using epidemiological cut-off values, ECOFF, as prescribed by EU legislation) in commensal E. coli isolated from healthy broilers (i.e. active surveillance) with the outcomes of monitoring clinical resistance (using clinical breakpoints, to determine susceptibility for antibiotic treatment in veterinary practice) in E. coli isolated from diseased broilers (i.e. passive surveillance).
METHODS: Data acquired by broth microdilution was analysed for commensal indicator E. coli and clinical E. coli from the Netherlands, 2014-2019. A generalized linear multivariable model (Poisson regression) was used to determine time trends and identify differences in mean resistant proportions.
RESULTS: Observed resistant proportions of the monitored commensal E. coli and clinical E. coli were similar with overlapping confidence intervals for most time points for ampicillin, gentamicin, cefotaxime, tetracycline, colistin and trimethoprim/sulfonamide. The statistical analysis showed that only for cefotaxime and tetracycline, mean resistant proportions were different. In commensal E. coli, a decrease of resistant proportions over time was observed, except for gentamicin. In clinical E. coli, no time trend was detected in resistant proportions, except for cefotaxime and colistin.
CONCLUSIONS: Generally, the resistant proportions monitored in commensal and clinical E. coli were similar. However, some relevant differences were found, which can be explained by the type of monitoring approach, i.e. active or passive surveillance. The random sample of commensal E. coli isolated from healthy animals (active surveillance), was more suitable to monitor AMR time trends. The sample of clinical isolates from diseased animals (passive surveillance), resulted in a higher chance to detect low-prevalent resistance: i.e. cefotaxime and colistin. The clinical E. coli data showed more fluctuation over time, and data from a longer period of time would be needed to determine the association. This study shows the value of both an active and a passive surveillance component for AMR monitoring.

Bedaquiline and clofazimine are increasingly used to treat infections with Mycobacterium abscessus. We determined distributions of MICs by broth microdilution for bedaquiline and clofazimine for 61 M. abscessus clinical isolates using different media and incubation times. We show that incubation time and growth media critically influence the MIC. Our data will aid in defining future clinical breakpoints for in vitro susceptibility testing for bedaquiline and clofazimine in M. abscessus.