Search

Background Surveillance is imperative to successful antibiotic stewardship, but active surveillance is resource-intensive. Regional antibiograms provide a supplementary method of surveillance while conserving resources. We developed the first statewide antibiograms to track susceptibility across the state of Minnesota and compared 2019 and 2020 data. Methods Hospital antibiograms for 2019 and 2020 were submitted to the Minnesota Department of Health in January 2022. Twelve organisms (Enterococcus faecalis, Enterococcus faecium, methicillin-resistant Staphylococcus aureus, methicillin-susceptible S. aureus, S. aureus, Acinetobacter spp., Enterobacter spp., Escherichia coli, Klebsiella aerogenes, K. oxytoca, K. pneumoniae, and Pseudomonas aeruginosa) were selected for analysis against 32 drugs, leading to a total of 142 unique organism/agent combinations. The number of isolates and percent susceptibility from each antibiogram were used to calculate a weighted average percent susceptibility as the state-level susceptibility. Chi-square analysis compared the proportion of susceptible to non-susceptible isolates between the years. Results Submitted antibiograms (n=40) represented 30% of Minnesota hospitals. The total number of isolates reported decreased by 5% from 2019 (n=85,010) to 2020 (n=80,781), with E. coli having the greatest proportion (2019=41.7%, 2020=39.5%) of the total isolates. Significant changes in susceptibility proportions occurred in 33% of analyzed organism/agent combinations, with 68% of these showing increased susceptibility in 2020 compared to 2019. All antibiotics tested against P. aeruginosa demonstrated a statistical increase in susceptibility from 2019 to 2020 (Figure 1). Figure 1 Statewide susceptibility profile of P. aeruginosa in Minnesota, 2019 and 2020. Antimicrobial agents grouped by class from narrow spectrum to broad spectrum, cephalosporins ordered by generation. *denotes statistical significance (p Conclusion Analysis of statewide antibiograms for 2019 and 2020 demonstrated overall stability in antimicrobial susceptibility, with two-thirds of organism/agent combinations showing no significant changes. In those with significant changes, the majority exhibited an increase in susceptibility. Susceptibility changed more frequently in gram-negative organisms than gram-positive organisms. Ongoing collection of antibiograms will be used to assess regional and statewide susceptibility trends across Minnesota. Disclosures Elizabeth B. Hirsch, PharmD, FCCP, FIDSA, Melinta: Advisor/Consultant|MeMed: Advisor/Consultant|Merck: Advisor/Consultant|Merck: Grant/Research Support.

Fosfomycin has been shown to have a wide spectrum of activity against multidrug-resistant gram-negative bacteria; however, breakpoints have been established only for Escherichia coli or Enterobacterales per the Clinical and Laboratory Standards Institute (CLSI) and the European Committee on Antimicrobial Susceptibility Testing (EUCAST), respectively. Lack of additional organism breakpoints limits clinical use of this agent and has prompted extrapolation of these interpretive categories to other organisms like Pseudomonas aeruginosa without supporting evidence. Further complicating the utility of fosfomycin is the specified method for minimal inhibitory concentration determination, namely agar dilution, which is not widely available and is both labor- and time-intensive. We therefore sought to determine the susceptibility of a large international collection of P. aeruginosa isolates (n = 198) to fosfomycin, and to compare testing agreement rates across four methods: agar dilution, broth microdilution, disk diffusion, and Etest. Results were interpreted according to CLSI E. coli breakpoints with 49.0-85.8% considered susceptible, dependent upon the testing method used. Epidemiological cutoff values were calculated and determined to be 256 μg/mL or 512 μg/mL for agar dilution and broth microdilution, respectively. Agreement rates were analyzed using both agar dilution and broth microdilution with a resulting high essential agreement rate of 91.3% between the two susceptibility testing methods. These results indicate that broth microdilution may be a reliable method for fosfomycin susceptibility testing against P. aeruginosa and stress the need for P. aeruginosa-specific breakpoints.

Antimicrobial peptides may be alternatives to traditional antibiotics with reduced bacterial resistance. The antimicrobial peptide GL13K was derived from the salivary protein BPIFA2. This study determined the relative activity of the L-and D-enantiomers of GL13K to wild-type and drug-resistant strains of three gram-negative species and against Pseudomonas aeruginosa biofilms. DGL13K displayed in vitro activity against ESBL and KPC-producing Klebsiella pneumoniae (MICs 16-32 μg/ml), MDR and XDR P. aeruginosa, and XDR Acinetobacter baumannii carrying metallo-beta-lactamases (MICs 8-32 μg/ml). P. aeruginosa showed low inherent resistance to DGL13K and the increased metabolic activity and growth caused by sub-MIC concentrations of GL13K peptides did not result in acquired bacterial resistance. Daily dosing for approximately two weeks did not increase the MIC of DGL13K or cause cross-resistance between LGL13K and DGL13K. These data suggest that DGL13K is a promising candidate antimicrobial peptide for further development.

Background Fosfomycin combination therapy is a potential approach for treatment of multidrug-resistant (MDR) PA infections despite a lack of approved susceptibility breakpoints for this organism. While DD testing is commonly used for fosfomycin, growth of discrete inner colonies (IC) within the zone of inhibition has been observed for multiple organisms following DD. Criteria recommended by CLSI and EUCAST are contradictory for interpreting these inner colonies. We therefore sought to determine the frequency of inner colonies and MIC differences between PA parent-inner colony pairs from an international isolate collection. Methods A convenience collection of 198 clinical PA isolates from three U.S institutions (n = 82), two Australian institutions (n = 72), and the CDC & FDA Antibiotic Resistance Isolate Bank (n = 44) were included. Fosfomycin MIC values were determined in duplicate on separate days by DD and broth microdilution (BMD) testing. For parent isolates with discrete IC observed during DD, IC isolates were subcultured and MIC values were determined and then compared to their corresponding parent isolates. MIC values were interpreted using CLSI Escherichia coli (EC) breakpoints (susceptible: MIC ≤ 64 μg/mL, intermediate: MIC = 128, resistant: MIC ≥ 256 μg/mL). Results Parent isolate BMD MIC values ranged from < 4 to > 256 μg/mL while IC isolate BMD MIC values ranged from 128 to > 1024 μg/mL. MIC50/90 values were 128/256 μg/mL and > 1024/ > 1024 μg/mL for the parent and IC isolates, respectively. A high frequency of 45% (89/198) of parent isolates displayed discrete IC which also demonstrated a higher frequency of resistance (97.8%) compared to the parent isolates (23.7%). Conclusion IC MIC values were higher overall compared to parent MIC values, with an average fold difference of ~18 between the parent-inner colony pairs. The frequency of IC found in this study (45%) is considerably higher than previously observed in EC clinical isolates. These data highlight the need to further investigate the importance of these IC and warrant caution for extrapolation of EC breakpoints for fosfomycin susceptibility testing against PA. Disclosures All Authors: No reported disclosures

Background FOF has been used in the treatment of multidrug-resistant (MDR) KP infections despite established susceptibility breakpoints. At present, agar dilution (AD) is considered the reference method for FOF while broth microdilution (BMD) is specifically recommended against despite its convenience over AD. We therefore sought to assess FOF activity against KP, along with essential and categorical agreement between AD and BMD methods to determine if BMD could be used as a reliable testing method. Methods Minimal inhibitory concentration (MIC) values were determined for a convenience collection of 69 KP isolates (59.4% MDR) from three US institutions. MIC testing was conducted in duplicate on separate days using AD and BMD methods; essential and categorical agreement were calculated using AD as the reference method. Fourteen isolates were also analyzed using high-inoculum AD (105.3-5.9 CFU/mL) similar to the BMD method. MIC values were categorized using Clinical and Laboratory Standards Institute (CLSI) interpretive criteria for Escherichia coli (≤ 64 mg/L, susceptible). ECVs were determined according to CLSI methodology. Results MIC values varied between methods, withMIC50/MIC90 values being 32/256 mg/L for AD and 128/256 mg/L for BMD. Using E. coli criteria, susceptible/intermediate/resistant rates were 82.6/2.9/14.5% (AD) and 44.9/21.7/33.3% (BMD). Essential agreement was 44.9% and categorical agreement was 60.8%. When using high-inoculum AD, MIC values were on average three-fold higher compared to standard-inoculum AD, with 10 of the 14 (71.4%) isolates brought into essential agreement with BMD. Calculated ECVs were 128 mg/L for standard-inoculum AD and 1024 mg/L for BMD. Conclusion Our collection of KP displayed high MIC values to FOF, in addition to substantial discrepancies between AD and BMD methods. Essential agreement increased with the use of high-inoculum AD testing, which better correlated with BMD results. ECV for BMD was three dilutions higher than that for standard-AD ECV. Based on these results, we recommend further investigation of BMD for FOF testing using a larger isolate collection, along with optimization of currently recommended testing methods. In light of these results, KP-specific breakpoints should also be examined. Disclosures Elizabeth B. Hirsch, PharmD, Merck (Grant/Research Support)Nabriva Therapeutics (Advisor or Review Panel member)

Background In the United States, interpretive criteria for FOF are established only for EC, yet those criteria are often extrapolated to KP. Recent studies have highlighted both inferior clinical outcomes after FOF treatment and difficulties in interpretation of inner colony subpopulations, the presence of which may affect clinical efficacy. We sought to compare FOF activity against EC and KP and to determine the prevalence of inner colony subpopulations following disk diffusion (DD) testing of the two species. Methods A convenience collection of 73 KP and 42 EC isolates from 3 U.S. institutions were included. Minimal inhibitory concentration (MIC) testing was performed in duplicate on separate days using agar dilution (AD) and DD as recommended by the Clinical and Laboratory Standards Institute guidelines, with application of EC susceptibility (≤ 64mg/L) breakpoints. The frequency and counts of inner colonies observed during DD testing was calculated, and colonies were subcultured for use in future studies. Results MIC50/90 values were 1/16 mg/L and 32/256 mg/L for EC and KP respectively. All EC isolates were considered susceptible and therefore categorical agreement was 100%. The majority of KP isolates were considered susceptible (83.6% with AD and 86.3% with DD) and categorical agreement between the methods was 84.9%. Inner colonies were observed during DD testing in 88.1% of EC isolates and 80.8% of KP isolates during at least one replicate, with 47.6% of EC isolates and 39.7% of KP isolates showing inner colony growth during both DD test replicates. More than 10 inner colonies were observed in 50% of EC isolates compared to 12.3% of KP isolates. Conclusion KP isolates demonstrated considerably higher FOF MIC values compared to EC, as evidenced by MIC50/90 values 4-5 dilutions higher than those for EC. The categorical agreement rate was higher among EC than KP, highlighting concerns regarding the practice of extrapolating FOF susceptibility breakpoints for EC to KP. The high frequency of inner colonies observed in DD for both species necessitates further studies to determine best practices for interpreting their relevance, fitness, and resistance in order to identify potential impacts to clinical efficacy of FOF. Disclosures Elizabeth B. Hirsch, PharmD, Merck (Grant/Research Support)Nabriva Therapeutics (Advisor or Review Panel member)

Background FOF has been used clinically for the treatment of PA infections in the absence of established interpretive criteria. A recent study identified a low frequency of nonsusceptible inner colony mutants during disk diffusion (DD) testing of Escherichia coli; however, the frequency of this phenomenon in PA isolates is not well characterized. We sought to determine FOF activity against an international collection of PA isolates and the frequency of inner colony mutants observed during Etest and DD testing. Methods Minimal inhibitory concentration (MIC) values were determined for a convenience collection of 109 PA ([70/94] 64.2% MDR) isolates from 4 institutions in the United States and Australia. MIC testing was conducted in duplicate on separate days utilizing agar dilution (AD), broth microdilution (BMD), DD, and Etest as recommended per Clinical and Laboratory Standards Institute (CLSI). CLSI E.coli interpretive criteria (≤ 64 mg/L susceptible) were used for MIC interpretations. The proportion of isolates containing inner colonies was determined using DD and Etest. Inner colony mutants were subcultured and retested using BMD with comparison to the parent isolate MICs. Results FOF MICs varied widely and ranged from 1024 mg/L with MIC50/MIC90 values of 64/256 (AD), 64/512 (Etest), and 64/256 (BMD) mg/L. Using E. coli criteria, susceptible/resistant rates were: 60.5/17.4% for AD; 60.5/22.0% for Etest; 86.2/7.3% for DD; and 53.2/17.4% for BMD. Inner colonies were frequently observed in 38.5% and 35.8% of DD and Etest inhibition zones, respectively. After repeat testing, mutant MIC values ranged from 64 to > 1024 mg/L and a majority (85.9%) had MIC values ≥ 512 mg/L. Conclusion Observed MIC values of this (64% MDR) collection varied widely with MIC50/90 values commonly at or above the E. coli susceptibility breakpoint. Inner colony mutants were frequently observed and highly resistant. Whole-genome sequencing is currently underway for a subset of parent/mutant pairs to determine whether specific genetic alterations are attributed to the increased MICs. Based on these results, caution should be warranted in extrapolating E. coli breakpoints to other organisms, and treatment of PA with FOF should be further evaluated. Disclosures Elizabeth B. Hirsch, PharmD, Merck: Grant/Research Support, Research Grant; Nabriva Therapeutics: Advisory Board; Paratek Pharmaceuticals: Advisory Board.