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Carbapenem-resistant Acinetobacter baumannii-calcoaceticus complex (CRAB) is one of the top-priority pathogens for new antibiotic development. Unlike other antibiotic-resistant threats, none of the available therapies have been shown to consistently reduce mortality or improve patient outcomes in clinical trials. Antibiotic combination therapy is routinely used in clinical practice; however, the preferred combination has not been defined. This narrative review focuses on evidence-based solutions for the treatment of invasive CRAB infections. We dissect the promise and perils of traditional agents used in combination, such as colistin, sulbactam, and the tetracyclines, and offer clinical pearls based on our interpretation of the available data. Next, we investigate the merits of newly developed β-lactam agents like cefiderocol and sulbactam-durlobactam, which have demonstrated contrasting results in recent randomized clinical trials. The review concludes with the authors’ perspective on the evolving treatment landscape for CRAB infections, which is complicated by limited clinical data, imperfect treatment options, and a need for future clinical trials. We propose that effective treatment for CRAB infections requires a personalized approach that incorporates host factors, the site of infection, pharmacokinetic-pharmacodynamic principles, local molecular epidemiology of CRAB isolates, and careful interpretation of antibiotic susceptibility testing results. In most clinical scenarios, a dose-optimized, sulbactam-based regimen is recommended with the addition of at least one other in vitro active agent. Should sulbactam-durlobactam receive regulatory approval, recommendations will need to be re-evaluated with the most recent evidence.

Objectives We evaluated the clinical characteristics and outcomes of patients with COVID-19 who received three-drug combination regimens for treatment of carbapenem-resistant Acinetobacter baumannii (CRAB) infections during a single-centre outbreak. Our objective was to describe the clinical outcomes and molecular characteristics and in vitro synergy of antibiotics against CRAB isolates. Materials and methods Patients with severe COVID-19 admitted between April and July 2020 with CRAB infections were retrospectively evaluated. Clinical success was defined as resolution of signs/symptoms of infection without need for additional antibiotics. Representative isolates underwent whole-genome sequencing (WGS) and in vitro synergy of two- or three-drug combinations was assessed by checkerboard and time-kill assays, respectively. Results Eighteen patients with CRAB pneumonia or bacteraemia were included. Treatment regimens included high-dose ampicillin-sulbactam, meropenem, plus polymyxin B (SUL/MEM/PMB; 72%), SUL/PMB plus minocycline (MIN; 17%) or other combinations (12%). Clinical resolution was achieved in 50% of patients and 30-day mortality was 22% (4/18). Seven patients had recurrent infections, during which further antimicrobial resistance to SUL or PMB was not evident. PMB/SUL was the most active two-drug combination by checkerboard. Paired isolates collected before and after treatment with SUL/MEM/PMB did not demonstrate new gene mutations or differences in the activity of two- or three-drug combinations. Conclusions Use of three-drug regimens for severe CRAB infections among COVID-19 resulted in high rates of clinical response and low mortality relative to previous studies. The emergence of further antibiotic resistance was not detected phenotypically or through WGS analysis. Additional studies are needed to elucidate preferred antibiotic combinations linked to the molecular characteristics of infecting strains.

Objectives Infections due to carbapenem-resistant Enterobacterales are considered urgent public health threats and often treated with a β-lactam/β-lactamase inhibitor combination. However, clinical treatment failure and resistance emergence have been attributed to inadequate dosing. We used a novel framework to provide insights of optimal dosing exposure of ceftazidime/avibactam. Methods Seven clinical isolates of Klebsiella pneumoniae producing different KPC variants were examined. Ceftazidime susceptibility (MIC) was determined by broth dilution using escalating concentrations of avibactam. The observed MICs were characterized as response to avibactam concentrations using an inhibitory sigmoid Emax model. Using the best-fit parameter values, %fT>MICi was estimated for various dosing regimens of ceftazidime/avibactam. A hollow-fibre infection model (HFIM) was subsequently used to ascertain the effectiveness of selected regimens over 120 h. The drug exposure threshold associated with bacterial suppression was identified by recursive partitioning. Results In all scenarios, ceftazidime MIC reductions were well characterized with increasing avibactam concentrations. In HFIM, bacterial regrowth over time correlated with emergence of resistance. Overall, suppression of bacterial regrowth was associated with %fT>MICi ≥ 76.1% (100% versus 18.2%; P Conclusions An optimal drug exposure to suppress KPC-producing bacteria was identified. The novel framework is informative and may be used to guide optimal dosing of other β-lactam/β-lactamase inhibitor combinations. Further in vivo investigations are warranted.

In response to the global burden of antimicrobial resistance (AMR) and the critical role antimicrobial stewardship plays in optimizing antibiotic use and reducing the subsequent emergence of AMR, Antimicrobial Agents and Chemotherapy is excited to add a new section to the journal focused on antimicrobial stewardship studies. Combatting the devastating burden of AMR requires novel, multipronged approaches from clinicians and scientists alike.

ObjectivesThe availability of new β-lactam/β-lactamase inhibitors ceftazidime/avibactam, meropenem/vaborbactam and imipenem/relebactam have redefined contemporary treatment of Klebsiella pneumoniae carbapenemase-producing Klebsiella pneumoniae (KPC-Kp) infections. We aimed to characterize and contrast the in vitro activity of these agents against genetically diverse KPC-Kp clinical isolates.MethodsWe analysed genomes of 104 non-consecutive KPC-Kp isolates and compared the in vitro antibiotic activity by KPC subtype and ompK36 genotype. MICs were determined in triplicate by CLSI methods. Twenty representative isolates were selected for time–kill analyses against physiological steady-state and trough concentrations, as well as 4× MIC for each agent.ResultsFifty-eight percent and 42% of isolates harboured KPC-2 and KPC-3, respectively. OmpK36 mutations were more common among KPC-2- compared with KPC-3-producing Kp (P ConclusionsOur investigation identified key genotypes that attenuate, to varying degrees, the in vitro activity for each of the new β-lactam/β-lactamase inhibitors. Additional studies are needed to translate the importance of these observations into clinical practice.

ObjectivesTo characterize ablaCMYvariant associated with ceftazidime-avibactam (CZA) resistance from a serially collectedEscherichia coliisolate.MethodsA patient with an intra-abdominal infection due to recurrentE. coliwas treated with CZA. On day 48 of CZA therapy,E. coliwith a CZA MIC of >256 mg/L was identified from abdominal drainage. Illumina WGS was performed on all isolates to identify potential resistance mechanisms. Site-directed mutants of CMY β-lactamase were constructed to identify amino acid residues responsible for CZA resistance.ResultsWGS revealed that all three isolates wereE. coliST410. The CZA-resistant strain uniquely acquired a novel CMY β-lactamase gene, herein calledblaCMY-185, harbored on an IncIγ-type conjugative plasmid. The CMY-185 enzyme possessed four amino acid substitutions relative to CMY-2 including A114E, Q120K, V211S, and N346Y and conferred high-level CZA resistance with an MIC of 32 mg/L. Single CMY-2 mutants did not confer reduced CZA susceptibility. However, double and triple mutants containing N346Y previously associated with CZA resistance in other AmpC enzymes, conferred CZA MICs ranging between 4 and 32 mg/L as well as reduced susceptibility to the newly developed cephalosporin, cefiderocol. Molecular modelling suggested that the N346Y substitution confers the reduction of avibactam inhibition due to the steric hindrance between the side chain of Y346 and the sulfate group of avibactam.ConclusionWe identified CZA resistance inE. coliassociated with a novel CMY variant. Unlike other AmpC enzymes, CMY-185 appears to require an additional substitution on top of N346Y to confer CZA resistance.

BackgroundThe objectives of this study were to describe the changing epidemiology of gram-negative infective endocarditis (GNIE) and to identify factors associated with treatment failure and death.MethodsAdult patients with GNIE were included if they met modified Duke criteria for definitive infective endocarditis (IE) between April 2010 and December 2021. Patients were identified using Boolean search terms. Clinical failure was a defined as a composite of all-cause 42-day mortality or microbiologic failure. All analyses were performed using Stata, version 15.1.ResultsOne-hundred twenty-three patients were included. The most common pathogens were Serratia spp. (43%), Pseudomonas aeruginosa (21%), and Klebsiella spp. (14%). Fifty-two percent of cases were among persons who injection drugs (PWID; n = 64), for whom Serratia spp. (70%) was the most common cause of GNIE. Overall, patients infected with P. aeruginosa had higher microbiologic failure rates than other patients (23% vs 6%; P = .004). Patients who received combination therapy (n = 53) had comparable median lengths of stay (23 vs 19.5 days; P = .412), microbiologic failure rates (11.3% vs 7.1%; P = .528), clinical failure rates (18.9% vs 22.9%; P = .592), and 90-day mortality rates (13.2% vs 25.7%; P = .088) as those treated with monotherapy. After applying stepwise logistic regression, male gender, Pitt Bacteremia Score, and not receiving surgical intervention despite a surgical indication were associated with clinical failure.ConclusionsThis is the first study to identify Serratia spp. as the most common etiology of GNIE, which was particularly true among PWID. Microbiologic failures occurred most commonly among P. aeruginosa, and use of combination antimicrobial therapy did not improve clinical outcomes.

The purpose of this narrative review is to bring together the most recent epidemiologic, preclinical, and clinical findings to offer our perspective on best practices for managing patients with A. baumannii infections with an emphasis on carbapenem-resistant A. baumannii (CRAB). To date, the preferred treatment for CRAB infections has not been defined. Traditional agents with retained in vitro activity (aminoglycosides, polymyxins, and tetracyclines) are limited by suboptimal pharmacokinetic characteristics, emergence of resistance, and/or toxicity. Recently developed and US Food and Drug Administration (FDA)-approved β-lactam/β-lactamase inhibitor agents do not provide enhanced activity against CRAB. On balance, cefiderocol and eravacycline demonstrate potent in vitro activity and are well tolerated, but clinical data for patients with CRAB infections do not yet support widespread use. Given that CRAB has the capacity to infect vulnerable patients and preferred regimens have not been identified, we advocate for combination therapy. Our preferred regimen for critically ill patients infected, or considered to be at high risk for CRAB, includes meropenem, polymyxin B, and ampicillin/sulbactam. Importantly, site of infection, severity of illness, and local epidemiology are essential factors to be considered in selecting combination therapies. Molecular mechanisms of resistance may unveil preferred combinations at individual centers; however, such data are often unavailable to treating clinicians and have not been linked to improved clinical outcomes. Combination strategies may also pose an increased risk for antibiotic toxicity and Clostridioides difficile infection, and should therefore be balanced by understanding patient goals of care and underlying health conditions. Promising therapies that are in clinical development and/or under investigation include durlobactam–sulbactam, cefiderocol combination regimens, and bacteriophage therapy, which may over time eliminate the need for the continued use of polymyxins. Future goals for CRAB management include pathogen-focused treatment paradigms that are based on molecular mechanisms of resistance, local susceptibility rates, and the availability of well-tolerated, effective treatment options.

Carbapenem-resistant Klebsiella pneumoniae (CRKP) are major pathogens globally. It is unknown whether bloodstream infections (BSIs) by CRKP and other bacteria are commonly caused by single organisms or mixed microbial populations. We hypothesized that contemporaneous CRKP from blood cultures of individual patients are genetically and phenotypically distinct. We determined short-read whole genome sequences of 10 strains from single colonies from CRKP-positive blood cultures in each of 6 patients (Illumina HiSeq). All strains were sequence type (ST)-258 K. pneumoniae that were unique by core genome single nucleotide polymorphism phylogeny, antibiotic resistance and virulence genes, capsular polysaccharide (CPS) gene mutations, and/or plasmid loss. Strains from each of 3 patients that differed in antibiotic resistance, virulence and/or CPS gene content underwent long-read sequencing for genome completion (Oxford Nanopore), and were tested for phenotypes in vitro and pathogenicity during mouse BSIs. Genetically distinct strains within individual patients exhibited significant differences in carbapenem, beta-lactam/beta-lactamase inhibitor and other antibiotic responses, CPS production, mucoviscosity, and susceptibility to serum killing. In 2 patients, strains differed significantly in their ability to infect organs and cause mortality in mice. In conclusion, we identified genotypic and phenotypic variant ST258 K. pneumoniae strains from blood cultures of individual patients, which were not detected by the clinical laboratory at time of BSI diagnosis. The data support a new paradigm of CRKP population diversity during BSIs. If validated for other BSIs, within-host bacterial diversity may have profound implications for medical, microbiology laboratory and infection prevention practices, and for understanding emergence of antibiotic resistance and pathogenesis.IMPORTANCEIn processing positive microbiologic cultures, standard clinical laboratory practice is to test a single bacterial strain from each morphologically distinct colony. We performed comprehensive whole genome sequence analyses on 10 carbapenem-resistant Klebsiella pneumoniae (CRKP) strains from positive blood cultures from each of 6 patients. Our findings that all strains were genetically unique and that genetic variants manifested differences in phenotypes like antibiotic responsiveness and virulence suggest that CRKP bloodstream infections may be commonly caused by mixed bacterial populations. Results raise questions about laboratory protocols and treatment decisions that are directed against a single strain. The observation that pan-genome analyses revealed inter-strain differences that were not evident by studying core genomes has important implications for investigating nosocomial outbreaks and transmission. Data also suggest a model of pathogenesis of CRKP infections, in which environmental pressures in vivo may select for outgrowth of variants that manifest antibiotic resistance, tolerance or specific virulence attributes.

Background The optimal metronidazole dose for the treatment of Bacteroides spp. has not been defined. The purpose of this study was to evaluate the utility of a twice (BID), rather than thrice (TID) daily metronidazole dosing strategy among patients with bacteremia due to Bacteroides spp. Methods Adult, hospitalized patients with bacteremia secondary to Bacteroides spp. between October 2010 and June 2021 were assessed across 11 hospitals. The primary endpoint was clinical failure which was a composite of all-cause 30-day mortality, escalation of antimicrobial therapy, 30-day readmission or recurrence due to an anaerobic infection, positive repeat blood cultures for Bacteroides spp., or failure to resolve leukocytosis or fever. Results 208 patients were included; 68 received metronidazole 500mg BID and 140 patients received metronidazole 500mg TID (Figure). Patient age, Charlson comorbidity index, and Pitt Bacteremia score were similar among patients receiving BID vs TID dosing (Table 1). On balance, patients who received BID dosing were more likely to receive oral metronidazole and had shorter lengths of hospitalization prior to bacteremia. Overall, there was no significant difference between rates of clinical failure or other outcomes between patients who received BID versus TID metronidazole dosing (Table 2). In the multivariate model, neither the use of TID dosing (OR = 0.74; 95% CI = 0.33–1.65; P=0.457), time to treatment initiation (OR = 1; 95% CI = 0.81–1.22; P=0.968), days of initial non-metronidazole anaerobic therapy (OR = 0.91; 95% CI = 0.59–1.34; P=0.646), pre-infection length of stay (OR = 1.02; 95% CI = 0.99–1.05; P=0.106), admission prior to 2016 (OR = 1.09; 95% CI = 0.49–2.39; P=0.829), or initial oral metronidazole use (OR = 0.45; 95% CI = 0.18–1.03; P=0.066) were significantly associated with clinical failure. Figure 1:Inclusion and exclusion Non-metronidazole anaerobic coverage consisted of a beta-lactam/beta-lactamase inhibitor, cefoxitin, or a carbapenem Table 1:Demographics and baseline characteristicsTable 2:Clinical outcomes*Patients were considered to have escalated antimicrobial therapy if, in the setting of ongoing signs of infections, antimicrobial therapy was either broadened or the frequency of metronidazole was increased from twice daily to thrice daily. Conclusion In the largest study to date of patients with Bacteroides spp. bacteremia treated with metronidazole, there was no significant difference between BID and TID metronidazole dosing strategies. In the absence of a clear benefit, metronidazole 500mg BID is a reasonable dosing strategy in lieu 500mg TID for infections due to Bacteroides spp. Disclosures Ryan K. Shields, PharmD, MS, Infectious Disease Connect: Advisor/Consultant|Merck: Advisor/Consultant|Merck: Grant/Research Support|Roche: Grant/Research Support.

Background An ASP is mandated for all hospitals and requires extensive resources with multidisciplinary collaboration. We measured the impact of implementing real-time decision support software (ILUM Insight) within our ASP. Methods Our ASP has relied on prior authorization since 2002 and focused audit and feedback since 2015. In August 2021 we implemented to bring actionable data to front-line stewards. ILUM provides real-time notifications, organizes communications, and tracks patient-and provider-level data. We hypothesized that ILUM would increase the efficiency of ASP workflow and result in decreased antimicrobial utilization. We compared data 6 months before (8/20 – 1/21) and after (8/21 – 1/22) implementation. There were no significant staffing changes during either period. Results Existing notifications within ILUM were tailored to local practices, including alerts with intervention for positive blood cultures, antibiotic de-escalation, and bug-drug mismatches. New notifications were built for restricted antimicrobials, antibiotic timeouts, and MRSA screening. ASP pharmacists and physicians received training in July and November, respectively. A breakdown of all notifications received during the post-implementation period is provided in Fig 1. With increased ILUM usage, the number of interventions made by our ASP increased while missed opportunities decreased (Fig 2.). During the same time period, ASP communications rose from 205 to 1200 per month. Comparing pre- and post-implementation periods, antimicrobial days of therapy (DOT) per 1,000 patient days (PD) decreased by 14.5% from a median of 969 to 846 per month (Fig 3;P=0.002). Antimicrobial expenditures were decreased by a median 21% per month during the post-intervention period compared to baseline. Among patients prescribed antimicrobials during an index admission, 30-day re-admissions decreased from 330 to 262 and re-admissions associated with re-ordering of antimicrobials decreased from 235 to 182 (Fig 4). Conclusion Custom-designed, task-specific software improves the efficiency of daily ASP workflow and significantly decreased antimicrobial utilization without the need for additional ASP team members. Disclosures Ryan K. Shields, PharmD, MS, Infectious Disease Connect: Advisor/Consultant|Merck: Advisor/Consultant|Merck: Grant/Research Support|Roche: Grant/Research Support J Ryan Bariola, MD, Infectious Disease Connect: Salary support|Merck: Grant/Research Support Caley Yakemowicz, n/a, Infectious Disease Connect: Employee Courtney Simonick, n/a, Infectious Disease Connect: Stocks/Bonds Riaan Erwee, na, Infectious Disease Connect: Employee Erin K. McCreary, PharmD, Infectious Disease Connect: Advisor/Consultant Rima Abdel-Massih, MD, Infectious Disease Connect: Co founder and Chief Medical Officer|Infectious Disease Connect: Ownership Interest.

Background Endoscopic Endonasal Surgery (EES) is an innovative surgical technique to remove brain tumors and lesions. Post-operative central nervous system (CNS) infections following EES are poorly described. The objective of this study was to define the epidemiology and characteristics of post-EES CNS infections. Methods Adult patients who underwent EES between 1/2010 and 7/2021 were evaluated and included if microbiologically confirmed CNS infection occurred within 30 days of EES. Suspected contaminants, ventricular drain colonization, and pre-EES CNS infections were excluded. Results Overall, 2005 patients underwent EES; 1.8% (37/2005) developed CNS infection. The median [IQR] age was 51 [42-60] years, 32.4% (12/37) were female, and 54% (20/37) had a prior EES. The most common indications for EES were tumor resection [67.6% (25/37)] and cerebrospinal fluid (CSF) leak repair [24.3% (9/37)]. Post-operative CSF leaks were documented in 70.3% (26/37) of patients and 24.3% (9/37) had an extra-ventricular drain or shunt in place for >48 hours at the time of infection. Ceftriaxone prophylaxis was prescribed in 64.9% (24/37) of cases and other regimens varied. The median [IQR] time from EES to diagnosis of CNS infection was 12 [6-19] days. The most common pathogens were S. aureus, Enterobacterales, and P. aeruginosa(Fig 1). Among 20 patients with prior EES, pathogens included S. aureus (5/20), Enterobacterales (3/20), Enterococcus spp. (3/20) and polymicrobic infections (3/20). Overall, 35.1% (13/37) of patients developed CNS infection due to a pathogen susceptible to pre-EES prophylaxis. Among those colonized with MRSA at time of EES, 75% (3/4) developed MRSA CNS infection compared to 6.1% (2/33) of non-colonized MRSA patients (P=0.005). The overall 30-day mortality rate was 2.7% (1/37). Figure:Microbiology A polymicrobic case was defined as >1 pathogen isolated from CSF (n=1) or from rhinocerebral tissue if CSF cultures were negative (n=11). Among polymicrobic cases (n=12), P. aeruginosa (n=5), Enterococcus spp. (n=4). and S. aureus (n=3) were predominant. Cases labeled as other consisted of Trichoderma spp, A. xylosoxidans, P. acnes, S. epidermidis, Peptostreptococcus spp. Conclusion CNS infection post-EES is rare and causative pathogens vary. Given the predominance of S. aureus, antimicrobial prophylaxis should ensure adequate coverage of this pathogen in addition to sinus flora, and programs may benefit from screening patients for MRSA colonization pre-EES. Our data also suggest that prophylaxis should target Gram-negative and other colonizing bacteria among patients with prior EES. Disclosures Ryan K. Shields, PharmD, MS, Infectious Disease Connect: Advisor/Consultant|Merck: Advisor/Consultant|Merck: Grant/Research Support|Roche: Grant/Research Support.

Background Invasive candidiasis (IC) is associated with significant morbidity and mortality. Echinocandins (ECH; caspofungin (CAS), micafungin (MFG), anidulafungin (AFG)) are first line therapy for IC. ECH resistance has emerged widely, most commonly among Candida glabrata. At some centers, > 10% of C. glabrata are ECH resistant. We reported ECH resistance in 8% of C. glabrata at our hospital from 2008–2014, which prompted stewardship strategies to limit prolonged ECH treatment courses. We evaluated trends of ECH resistance from Jan 2010-Dec 2020 among C. glabrata clinical isolates, and assayed isolates for FKS hot spot (HS) mutations. Methods We mined electronic medical records for ECH MIC data against C. glabrata clinical isolates at UPMC. We defined ECH resistance using Clinical and Laboratory Standards Institute clinical breakpoints. Isolates resistant to any ECH underwent FKS Sanger sequencing. Statistical analysis was performed using Stata. Results Overall CAS, AFG and MFG resistance rates among 516 isolates were 4%, 3%, and 2.5%, respectively. There was no significant difference in resistance rates between blood and non-blood sites (Table 1). There was a significant decrease in MFG resistance over the 12-year period (Fig 1). Time series regression analysis showed a decrease in rate of MFG resistance of 0.73% per year (p=.007). Trends for decreases in AFG and CAS resistance rates were also noted (p=.1 and p=.17). Similar analyses showed annual decreases in MFG MICs (0.019 μg/mL per year; p=.07). The average percentage of AFG resistant isolates was significantly lower in 2017–2020 than in 2012–2016 (p=.04); similar trends were evident for MFG (p=.07) and CAS (p=.08) (Fig 2). Sanger sequencing of FKS1/2 was performed for 27 isolates resistant to ≥1 ECH. Mutations were found in 44% (12/27) of isolates: 5 mutations in HS1 FKS1, 1 in HS2 FKS1, 5 in HS1 FKS2 and 1 in HS2 FKS2. Using both CAS and AFG MICs was most predictive of FKS mutations (Fig. 3). Conclusion ECH resistance among C. glabrata stabilized or decreased over 11-years at our hospital, in keeping with stewardship interventions to reduce prolonged use of ECHs. These results demonstrate that it is feasible to conserve ECH susceptibility. Disclosures Cornelius J. Clancy, MD, receives research funding paid to his institution from Astellas and Merck: Grant/Research Support|serves as an advisory Board member for Astellas, Cidara, and Scynexis, served on the advisory board for Merck, Qpex Biopharma, and Shionogi: Advisor/Consultant|Venatorx and Needham & Associates: Advisor/Consultant Minh-Hong Nguyen, MD, QPX: Grant/Research Support.

Background Antibiotic resistant ENT are a growing threat worldwide. QPX is an ultra-broad-spectrum β-lactamase inhibitor (BLI) with potent inhibition of classes A and D, and many metallo-β-lactamases, which has potential to fill the void of currently available BL-BLIs. Methods We evaluated QPX (fixed concentration of 8 µg/mL) in combination with meropenem (MEM), cefepime (FEP) and aztreonam (ATM) against a diverse group of ENT clinical isolates selected from a worldwide repository. Antibiotic resistance determinants were assessed by mining whole-genome sequencing data generated using Illumina MiSeq. Results Klebsiella spp, Enterobacter spp, and Escherichia coli were most common among the 90 isolates tested (Fig. 1). 71% (64) produced carbapenemases (Fig. 2): 3 isolates produced both class A and B carbapenemases. 11% of isolates harbored mutations in ompK 36/ompC genes. 57% (51) were resistant to MEM, 31% to imipenem-relebactam, 17% (15) to ceftazidime-avibactam (CZA), and 9% to MEM-vaborbactam (MVB). ENT-resistant to CZA were due to production of class B, D or KPC-3 variants, or ompC mutation in a K. aerogenes isolate. ENT-resistant to MVB were due to class B or D enzymes, or class A with ompK36 or ompC mutations. MIC distributions of BL with or without QPX against all isolates tested are presented in Fig. 3. Addition of QPX to MEM, FEP and AMT reduced MIC50s and MIC90s of these agents (Fig. 3; p< 0.0001). Of note, addition of QPX reduced MEM MICs more than did vaborbactam (median 533- versus 24-fold; p=0.0006). QPX reduced MICs of MEM, FEP and ATM to levels below respective breakpoints for non-susceptibility (Fig. 3). MIC distribution of ENT isolates stratified by the types of carbapenemases are presented in Fig. 4. QPX potentiated activity of MEM, FEP or ATM against ENT, regardless of whether they produced class A, B or D carbapenemases. Conclusion QPX enhanced activity of MEM, FEP and ATM against carbapenemase-producing or other carbapenem-resistant ENT (CRE), regardless of species or other resistance determinants. Most remarkably, QPX rendered each BL equally active against CRE. The expanded spectrum of QPX against class B and D carbapenemases addresses a major unmet need against CRE. Disclosures Cornelius J. Clancy, MD, receives research funding paid to his institution from Astellas and Merck: Grant/Research Support|serves as an advisory Board member for Astellas, Cidara, and Scynexis, served on the advisory board for Merck, Qpex Biopharma, and Shionogi: Advisor/Consultant|Venatorx and Needham & Associates: Advisor/Consultant.

Background The longstanding paradigm is that almost all bloodstream infections (BSIs) stem from a single, clonal organism. We hypothesized that carbapenem resistant K. pneumoniae (CRKP) from individual patients (pts) with BSIs are genetically diverse and manifest phenotypic differences that are not typically recognized by the clinical microbiology laboratory at time of diagnosis. Methods We streaked blood culture (BC) broth from 6 pts with CRKP BSI onto blood agar plates, and randomly picked 9 colonies (strains) for HiSeq (Illumina) whole genome sequence. Single BSI isolates recovered by the clinical micro lab from individual pts underwent short- and long-read MinION (ONT) sequencing. Results 2 pts were infected with clade 1 (B, G), and 4 with clade 2 (A, D, F, J) ST258 KPC-producing KP. Strains from individual pts clustered by cgSNP phylogeny (Fig. 1A-B). BC bottles from each pt harbored genetically heterogenous KP populations, with strains differing from each other by cgSNPs (Fig. 1B), presence/absence of specific antibiotic resistance genes (Fig. 1A), mutations of capsular genes (Fig 2) and at other loci involved in host interactions, and/or loss of plasmids or plasmid-borne genes (Fig. 1A). Differences in capsular gene composition were observed in KL107 capsule type strains from pts A, D and J (Fig. 2). Pangenome analyses showed accessory gene composition diversity among strains from all pts (Fig. 3). Intra-pt genetically diverse strains exhibited differences in antibiotic resistance (Fig. 4), viscosity and mucosity, capsular content, and resistance to serum and macrophage killing. Various strains from pts A and J differed in ability to cause target organ infections or mortality in a mouse model of intravenous disseminated infection (Fig. 5). Conclusion We identified genotypic and phenotypic variant strains of ST258 KP from BSIs of individual patients that were not recognized at time of diagnosis. Our data suggest a new, population-based paradigm for BSIs by CRKP. The findings potentially have profound implications for medical, microbiology laboratory and infection prevention practices, and for understanding emergence of antibiotic resistance and pathogenesis. Disclosures Cornelius J. Clancy, MD, receives research funding paid to his institution from Astellas and Merck: Grant/Research Support|serves as an advisory Board member for Astellas, Cidara, and Scynexis, served on the advisory board for Merck, Qpex Biopharma, and Shionogi: Advisor/Consultant|Venatorx and Needham & Associates: Advisor/Consultant.

Background Pseudomonas aeruginosa (PA) is an important pathogen notorious for antibiotic resistance. Xeruborbactam (QPX) is a potent ultra-broad-spectrum boronic acid β-lactam inhibitor (BLI) that, in combination with selected BL antibiotics, has excellent in vitro activity against carbapenem-resistant Enterobacterales (CRE), and Acinetobacter baumanii or PA producing class A/B or D carbapenemases. We evaluated QPX in combination with anti-pseudomonal BLs against clinical PA isolates from tertiary care US hospitals. Methods We tested PA clinical isolates resistant to ≥1 BL (imipenem (IMP), meropenem (MEM), cefepime (FEP), piperacillin-tazobactam (PIP-TAZ), aztreonam (ATM), ceftolozane-tazobactam (TOL-TZP), ceftazidime-avibactam (CZA), IMI-relebactam (IMI-REL), MEM-vaborbactam (MVB)) against QPX (in fixed concentration of 8 µg/mL) combined with anti-pseudomonal BLs. We performed whole-genome sequencing on isolates using the MiSeq platform (Illumina). Results Antibiograms and resistance determinants of 77 isolates tested to date are summarized in Figs 1, 2. 91% of isolates were CR; 43%, 58% and 61% were resistant to CZA, IMI-REL and MVB, respectively. No isolates produced class A/B/D carbapemases. All except 2 isolates carried PDC variants. 92% either had oprD porin single nucleotide polymorphism (SNP) or deletions. mutS mutations (present in 23% of isolates) were associated with resistance to IPM/QPX (p=.04), but not TOL-QPX or PIP-QPX. Addition of QPX significantly reduced MIC50 of IPM (32-fold), PIP (16-fold), FEP and ATM (4-fold) (all p< .0001, Fig 3). Addition of QPX to MEM or TOL reduced MIC50 by 2-fold (p=.02, p< .0001). QPX reduced IMP and PIP MICs more than did REL (32 vs 8-fold, p< .0001) or TZP (16 vs nil, p=.0007), respectively. Addition of QPX reduced BL resistance, especially for TOL, PIP and IMP (Fig 4). We identified factors associated with BL/QPX resistance by logistic regression; for IMP/QPX: IMP resistance (p=.001), SNP in mexB (p=.01); for TOL/QPX: TOL resistance (p=.02); for PIP/QPX (SNP in mexR and mexB (p=.03). Conclusion QPX enhanced the activity of BLs, especially TOL and PIP, against PA with baseline resistance to BL, more so than other BLIs. TOL/QPX and PIP/QPX are less impacted by PA efflux and porin mutations than IMP/QPX. Disclosures Cornelius J. Clancy, MD, receives research funding paid to his institution from Astellas and Merck: Grant/Research Support|serves as an advisory Board member for Astellas, Cidara, and Scynexis, served on the advisory board for Merck, Qpex Biopharma, and Shionogi: Advisor/Consultant|Venatorx and Needham & Associates: Advisor/Consultant.

Enterococcus faeciumis a member of the human gastrointestinal (GI) tract microbiota but can also cause invasive infections, especially in immunocompromised hosts. Enterococci display intrinsic resistance to many antibiotics, and most clinicalE. faeciumisolates have acquired vancomycin resistance, leaving clinicians with a limited repertoire of effective antibiotics. As such, vancomycin-resistantE. faecium(VREfm) has become an increasingly difficult to treat nosocomial pathogen that is often associated with treatment failure and recurrent infections. We followed a patient with recurrentE. faeciumbloodstream infections (BSIs) of increasing severity that ultimately became unresponsive to antibiotic combination therapy over the course of 7 years. Whole genome sequencing (WGS) showed that the patient was colonized with closely relatedE. faeciumstrains for at least two years, and that invasive isolates likely emerged from a largeE. faeciumpopulation in the patient’s GI tract. The addition of bacteriophage (phage) therapy to the patient’s antimicrobial regimen was associated with several months of clinical improvement and reduced intestinal burden of VRE andE. faecium. Eventual recurrence ofE. faeciumBSI was not associated with the development of antibiotic or phage resistance in post-treatment isolates. However, an anti-phage neutralizing antibody response occurred simultaneously with an increased relative abundance of VRE in the GI tract, both of which may have contributed to clinical failure. Taken together, these findings highlight the potential utility and limitations of phage therapy to treat antibiotic-resistant enterococcal infections.ImportancePhage therapy is an emerging therapeutic approach for treating bacterial infections that do not respond to traditional antibiotics. The addition of phage therapy to systemic antibiotics to treat a patient with recurrentE. faeciuminfections that were non-responsive to antibiotics alone resulted in fewer hospitalizations and improved the patient’s quality of life. Combination phage and antibiotic therapy reducedE. faeciumand VRE abundance in the patient’s stool. Eventually an anti-phage antibody response emerged that was able to neutralize phage activity, which may have limited clinical efficacy. This study demonstrates the potential of phages as an additional option in the antimicrobial toolbox for treating invasive enterococcal infections and highlights the need for further investigation to ensure phage therapy can be deployed for maximum clinical benefit.

Background Multidrug-resistant Enterobacterales (MDR-E), including carbapenem-resistant and third-generation cephalosporin-resistant Enterobacterales (CRE, CefR-E), are major pathogens following solid organ transplantation (SOT). Methods We prospectively studied patients who underwent lung, liver, and small bowel transplant from February 2015 through March 2017. Weekly perirectal swabs (up to 100 days post-transplant) were cultured for MDR-E. Whole-genome sequencing (WGS) was performed on gastrointestinal (GI) tract–colonizing and disease-causing isolates. Results Twenty-five percent (40 of 162) of patients were MDR-E GI-colonized. Klebsiella pneumoniae was the most common CRE and CefR-E. Klebsiella pneumoniae carbapenemases and CTX-M were leading causes of CR and CefR, respectively. Thirty-five percent of GI colonizers developed MDR-E infection vs 2% of noncolonizers (P < .0001). The attack rate was higher among CRE colonizers than CefR-E colonizers (53% vs 21%, P = .049). GI colonization and high body mass index were independent risk factors for MDR-E infection (P ≤ .004). Thirty-day mortality among infected patients was 6%. However, 44% of survivors developed recurrent infections; 43% of recurrences were late (285 days to 3.9 years after the initial infection). Long-term survival (median, 4.3 years post-transplant) did not differ significantly between MDR-E–infected and MDR-E–noninfected patients (71% vs 77%, P = .56). WGS phylogenetic analyses revealed that infections were caused by GI-colonizing strains and suggested unrecognized transmission of novel clonal group-258 sublineage CR-K. pneumoniae and horizontal transfer of resistance genes. Conclusions MDR-E GI colonization was common following SOT and predisposed patients to infections by colonizing strains. MDR-E infections were associated with low short- and long-term mortality, but recurrences were frequent and often occurred years after initial infections. Findings provide support for MDR-E surveillance in our SOT program.

We report on 11 critically ill burn patients treated with cefiderocol for carbapenem-resistant Acinetobacter baumannii infections. Clinical success was achieved in 36% and complicated by treatment-emergent resistance and interpatient transmission of cefiderocol-resistant A. baumannii. Resistant isolates harbored disrupted pirA and piuA genes that were not disrupted among susceptible isolates.

Severe infections caused by methicillin-resistant Staphylococcus aureus (MRSA) are often complicated by persistent bacteremia (PB) despite active antibiotic therapy. Antibiotic resistance rarely contributes to MRSA-PB, suggesting an important role for antibiotic tolerance pathways. To identify bacterial factors associated with PB, we sequenced the whole genomes of 206 MRSA isolates derived from 20 patients with PB and looked for genetic signatures of adaptive within-host evolution. We found that genes involved in the tricarboxylic acid cycle (

Treatment options for Achromobacter xylosoxidans are limited. Eight cystic fibrosis patients with A. xylosoxidans were treated with 12 cefiderocol courses. Pretreatment in vitro resistance was seen in 3 of 8 cases. Clinical response occurred after 11 of 12 treatment courses. However, microbiologic relapse was observed after 11 of 12 treatment courses, notably without emergence of resistance.

Background Carbapenem-resistant gram-negative bacteria (CRGNB) continue to present a global healthcare crisis. We aimed to identify emerging trends of CRGNB over nearly 2 decades and describe the impact of CRGNB on patient outcomes. Methods Patients from whom CRGNB were isolated between 2000 and 2017 were included in the study. Carbapenem resistance was defined by the most recent breakpoints and applied across the study period. Patient demographics, clinical characteristics, and outcomes were retrieved from the electronic health record. Results A total of 94 888 isolates from 64 422 patients were identified; 9882 (10%) isolates from 4038 patients were carbapenem-resistant. Pseudomonas aeruginosa was the most common CRGNB each year. The second most common CRGNB emerged in waves over time. Carbapenem daily defined doses increased in parallel with CRGNB rates (R2 = 0.8131). The overall 30-day mortality rate was 19%, which decreased from 24% in 2000 to 17% in 2017 (P = .003; R2 = .4330). Among patients with CRGNB bloodstream infections (n = 319), overall 30- and 90-day mortality rates were 27% and 38%, respectively. Charlson score (adjusted odds ratio [aOR], 1.11 per point), intensive care unit residence (aOR, 7.32), and severe liver disease (aOR, 4.8.4) were independent predictors of 30-day mortality, while receipt of transplantation was associated with lower rates of death (aOR, 0.39). Among patients admitted between 2011 and 2017 (n = 2230), 17% died during hospitalization, 32% were transferred to long-term care facilities, and 38% were discharged home. Conclusions CRGNB emerged in waves over time, causing high rates of mortality. Despite increasing rates of CRGNB, overall patient outcomes have improved, suggesting that recognition and novel therapeutics have made a major impact.

Background Invasive fungal infections (IFIs) are common following lung transplantation. Isavuconazole is unstudied as prophylaxis in organ transplant recipients. We compared effectiveness and tolerability of isavuconazole and voriconazole prophylaxis in lung transplant recipients. Methods A single-center, retrospective study of patients who received isavuconazole (September 2015–February 2018) or voriconazole (September 2013–September 2015) for antifungal prophylaxis. IFIs were defined by EORTC/MSG criteria. Results Patients received isavuconazole (n = 144) or voriconazole (n = 156) for median 3.4 and 3.1 months, respectively. Adjunctive inhaled amphotericin B (iAmB) was administered to 100% and 41% of patients in the respective groups. At 1 year, 8% of patients receiving isavuconazole or voriconazole developed IFIs. For both groups, 70% and 30% of IFIs were caused by molds and yeasts, respectively, and breakthrough IFI (bIFI) rate was 3%. Outcomes did not significantly differ for patients receiving or not receiving iAmB. Independent risk factors for bIFI and breakthrough invasive mold infection (bIMI) were mold-positive respiratory culture and red blood cell transfusion >7 units at transplant. Bronchial necrosis >2 cm from anastomosis and basiliximab induction were also independent risk factors for bIMI. Isavuconazole and voriconazole were discontinued prematurely due to adverse events in 11% and 36% of patients, respectively (P = .0001). Most common causes of voriconazole and isavuconazole discontinuation were hepatotoxicity and lack of oral intake, respectively. Patients receiving ≥90 days prophylaxis had fewer IFIs at 1 year (3% vs 9%, P = .02). IFIs were associated with increased mortality (P = .0001) and longer hospitalizations (P = .0005). Conclusions Isavuconazole was effective and well tolerated as antifungal prophylaxis following lung transplantation.

Background and Purpose— Because of unique attributes of mechanical thrombectomy performed between 6 and 24 hours after symptom onset in acute ischemic stroke patients, it is not known if predictors of angiographic recanalization and favorable outcome in patients treated with thrombectomy in the late (6–24 hour) time window are similar to those treated in the early time window. Methods— We analyzed data from the DAWN trial (DWI or CTP Assessment With Clinical Mismatch in the Triage of Wake-Up and Late Presenting Strokes Undergoing Neurointervention With Trevo) which enrolled patients with symptom onset 6 to 24hours after last known well and occlusion of the intracranial internal carotid artery or proximal middle cerebral artery with a mismatch between severity of clinical deficit and infarct core volume as identified by computed tomography–perfusion or diffusion magnetic resonance imaging. We evaluated the effect of tandem occlusions, periprocedural heparin use, procedural speed (from puncture to procedure completion), general anesthesia, balloon-guide catheters, thrombectomy device size, and number of passes on substantial reperfusion (modified Thrombolysis in Cerebral Infarction 2b/3) and on likelihood of obtaining a modified Rankin Scale at 3 months indicating functional independence. Results— Of 107 patients who underwent MT in the interventional arm of DAWN, substantial reperfusion and modified Rankin Scale score 0 to 2 at 3 months was seen in 90 (84%) and 52 (49%), respectively. In univariate analysis, general anesthesia (odds ratio [OR] 0.27; P =0.042) and ≥3 passes with stent retriever (OR, 0.17; P =0.002) were inversely associated with substantial reperfusion. In multivariate analyses, only ≥3 passes were associated with lack of revascularization (OR, 0.17; P =0.002). in univariate analysis ≥3 passes (OR, 0.24; P =0.003) and baseline National Institutes of Health Stroke Scale score >17 (OR, 0.19; P P =0.003) and National Institutes of Health Stroke Scale score >17 (OR, 0.19; P Conclusions— Patients requiring ≥3 thrombectomy passes had reduced substantial reperfusion and favorable outcome at 3 months in DAWN. Whether or not additional thrombectomy techniques beyond ≥3 thrombectomy passes with the Trevo stent retriever are beneficial for patient outcomes in this patient population remains to be clarified by future studies. Clinical Trial Registration— URL: https://www.clinicaltrials.gov . Unique identifier: NCT02142283.

β-Lactamase-mediated resistance to ceftazidime-avibactam (CZA) is a serious limitation in the treatment of Gram-negative bacteria harboring Klebsiella pneumoniae carbapenemase (KPC). Herein, the basis of susceptibility to carbapenems and resistance to ceftazidime (CAZ) and CZA of the D179Y variant of KPC-2 and -3 was explored. First, we determined that resistance to CZA in a laboratory strain of Escherichia coli DH10B was not due to increased expression levels of the variant enzymes, as demonstrated by reverse transcription PCR (RT-PCR). Using timed mass spectrometry, the D179Y variant formed prolonged acyl-enzyme complexes with imipenem (IMI) and meropenem (MEM) in KPC-2 and KPC-3, which could be detected up to 24 h, suggesting that IMI and MEM act as covalent β-lactamase inhibitors more than as substrates for D179Y KPC-2 and -3. This prolonged acyl-enzyme complex of IMI and MEM by D179Y variants was not observed with wild-type (WT) KPCs. CAZ was studied and the D179Y variants also formed acyl-enzyme complexes (1 to 2 h). Thermal denaturation and differential scanning fluorimetry showed that the tyrosine substitution at position 179 destabilized the KPC β-lactamases (KPC-2/3 melting temperature [T(m)] of 54 to 55°C versus D179Y T(m) of 47.5 to 51°C), and the D179Y protein was 3% disordered compared to KPC-2 at 318 K. Heteronuclear (1)H/(15)N-heteronuclear single quantum coherence (HSQC) nuclear magnetic resonance (NMR) spectroscopy also revealed that the D179Y variant, compared to KPC-2, is partially disordered. Based upon these observations, we discuss the impact of disordering of the Ω loop as a consequence of the D179Y substitution. These conformational changes and disorder in the overall structure as a result of D179Y contribute to this unanticipated phenotype.

We report the emergence of imipenem-relebactam nonsusceptible Pseudomonas aeruginosa in 5 patients treated for nosocomial pneumonia for 10–28 days. Genome sequence analysis identified treatment-emergent mutations in MexAB-OprM and/or MexEF-OprN efflux operons that arose independently in each patient across distinct P. aeruginosa sequence types. Testing with efflux-inhibitor PAβN restored imipenem-relebactam susceptibility.

The objective of this study was to characterize isolates with reduced susceptibility to cefiderocol in patients receiving cefiderocol for nosocomial pneumonia or carbapenem-resistant infections in the Phase 3 APEKS-NP and CREDIBLE-CR studies. Susceptibility testing of isolates was conducted at a central laboratory, and post-treatment changes were evaluated according to available breakpoints for cefiderocol. Whole-genome sequencing and multilocus sequence typing were performed for isolates to confirm their origin and identify mutations. Five (APEKS-NP) and nine (CREDIBLE-CR) isolates demonstrated a ≥ 4-fold minimum inhibitory concentration (MIC) increase compared with genetically related baseline isolates; most remained susceptible to cefiderocol despite the ≥4-fold MIC increase. Mutations in β-lactamases or penicillin-binding protein (PBP) were identified in 4/14 isolates: one

By serially exposing an NDM-producing Klebsiella pneumoniae clinical strain to cefiderocol, we obtained a mutant with cefiderocol MIC of >128 μg/ml. The mutant contained an early stop codon in the iron transporter gene cirA, and its complementation fully restored susceptibility. The cirA-deficient mutant was competed out by the parental strain in vitro, suggesting reduced fitness. IMPORTANCE Cefiderocol, a newly approved cephalosporin agent with an extensive spectrum of activity against Gram-negative bacteria, is a siderophore cephalosporin that utilizes iron transporters to access the bacterial periplasm. Loss of functional CirA, an iron transporter, has been associated with cefiderocol resistance. Here, we show that such genetic change can be selected under selective pressure and cause high-level cefiderocol resistance, but with a high fitness cost. Whether these resistant mutants can survive beyond selective pressure will inform stewardship of this agent in the clinic.

Background Bloodstream infections caused by MSSA are associated with significant morbidity and mortality. Traditional treatment of MSSA bacteraemia includes an IV antistaphylococcal β-lactam and surgical source control when indicated. Objectives To evaluate the time to blood culture clearance as well as in-hospital and 90 day mortality in patients with persistent MSSA bacteraemia treated with combination antistaphylococcal penicillin plus carbapenem therapy. Methods Consecutive patients with persistent MSSA bacteraemia treated with combination therapy were identified by study investigators and reviewed by independent clinicians. The decision to initiate combination therapy was made by the consulting clinician or by the institution’s multidisciplinary endocarditis team. Results Among 10 patients with a median of 5 days of persistent MSSA bacteraemia, treatment with an antistaphylococcal penicillin plus carbapenem led to sterilization of blood cultures in all patients. Blood culture clearance occurred in a median of 1 day and patients received a median of 6 days of combination treatment. Four of seven patients who underwent source control of their primary site of infection cleared their bacteraemia on combination therapy prior to the surgical intervention. All patients survived to hospital discharge and 90 days post-discharge. Conclusions These data extend prior findings and provide further evidence that suggests the potential benefits of combination therapy among patients with persistent MSSA bacteraemia.

Pseudomonas aeruginosa infections can be difficult to treat and new therapeutic approaches are needed. Bacteriophage therapy is a promising alternative to traditional antibiotics, but large numbers of isolated and characterized phages are lacking. We collected 23 genetically and phenotypically diverse P. aeruginosa isolates from people with cystic fibrosis (CF) and clinical infections, and characterized their genetic, phenotypic, and prophage diversity. We then used these isolates to screen and isolate 14 new P. aeruginosa-targeting phages from hospital wastewater. Phages were characterized with genome sequencing, comparative genomics, and lytic activity screening against all 23 bacterial host isolates. For four different phages, we evolved bacterial mutants that were resistant to phage infection. We then used genome sequencing and functional analysis of the resistant mutants to study their mechanisms of phage resistance as well as changes in virulence factor production and antibiotic resistance, which differed from corresponding parent bacterial isolates. Finally, we tested two phages for their ability to kill P. aeruginosa grown in biofilms in vitro, and observed that both phages reduced viable bacteria in biofilms by least one order of magnitude. One of these phages also showed activity against P. aeruginosa biofilms grown on CF airway epithelial cells. Overall, this study demonstrates how systematic genomic and phenotypic characterization can be deployed to develop bacteriophages as precision antibiotics.

Acinetobacter baumannii is currently classified as one of six pathogens that contribute to increased patient mortality. Thus, exploratory studies navigating alternative treatment strategies are of supreme interest. Herein, we completed minimum inhibitory concentration (MIC) testing, and time-kill analyses (TKA) on 50 carbapenem-resistant Acinetobacterbaumannii isolates including 28 colistin-resistant isolates. Upon testing of MEM or TGC in the presence of sub-inhibitory COL against the 50 isolates, there was a median 2-fold reduction in MEM and TGC MICs. In the TKAs, the COL+MEM combination was synergistic in 45 (90%) isolates and bactericidal in 43 (86%) isolates at 24 hours, whereas the COL+TGC combination TKAs demonstrated synergy in 32 (64%) isolates and bactericidal activity was shown in 28 (56%) isolates. Additionally, sulbactam (SUL) and TGC were added to the COL+MEM dual therapy regimen to assess the possible utility of a triple therapy regimen against five non-responsive isolates. The COL+MEM+SUL and COL+MEM+TGC regimens effectively restored synergy in (5/5) 100% of the isolates. The results of this study demonstrate the potential utility of COL combinations in the treatment of carbapenem-resistant isolates.

Background and Purpose— It is unknown whether noncontrast computed tomography (NCCT) can identify patients who will benefit from intra-arterial treatment (IAT) in the extended time window. We sought to characterize baseline Alberta Stroke Program Early CT Score (ASPECTS) in DAWN (DWI or CTP Assessment With Clinical Mismatch in the Triage of Wake-Up and Late Presenting Strokes Undergoing Neurointervention With Trevo) and to assess whether ASPECTS modified IAT effect. Methods— Core lab adjudicated ASPECTS scores were analyzed. The trial cohort was divided into 2 groups by qualifying imaging (computed tomography versus magnetic resonance imaging). ASPECTS-by-treatment interaction was tested for the trial coprimary end points (90-day utility-weighted modified Rankin Scale (mRS) score and mRS, 0–2), mRS 0 to 3, and ordinal mRS. ASPECTS was evaluated separately as an ordinal and a dichotomized (0–6 versus 7–10) variable. Results— Of 205 DAWN subjects, 123 (60%) had NCCT ASPECTS, and 82 (40%) had diffusion weighted imaging ASPECTS. There was a significant ordinal NCCT ASPECTS-by-treatment interaction for 90-day utility-weighted mRS (interaction P =0.04) and mRS 0 to 2 (interaction P =0.02). For both end points, IAT effect was more pronounced at higher NCCT ASPECTS. The dichotomized NCCT ASPECTS-by-treatment interaction was significant only for mRS 0 to 2 (interaction P =0.04), where greater treatment benefit was seen in the ASPECTS 7 to 10 group (odds ratio, 7.50 [2.71–20.77] versus odds ratio, 0.48 [0.04–5.40]). A bidirectional treatment effect was observed in the NCCT ASPECTS 0 to 6 group, with treatment associated with not only more mRS 0 to 3 outcomes (50% versus 25%) but also more mRS 5 to 6 outcomes (40% versus 25%). There was no significant modification of IAT effect by diffusion weighted imaging ASPECTS. Conclusions— Baseline NCCT ASPECTS appears to modify IAT effect in DAWN. Higher NCCT ASPECTS was associated with greater benefit from IAT. No treatment interaction was observed for diffusion weighted imaging ASPECTS.

Background OXA-2 is a class D β-lactamase that confers resistance to penicillins, as well as narrow-spectrum cephalosporins. OXA-2 was recently reported to also possess carbapenem-hydrolysing activity. Here, we describe a KPC-2-encoding Klebsiella pneumoniae isolate that demonstrated reduced susceptibility to ceftazidime and ertapenem due to production of OXA-2. Objectives To elucidate the role of OXA-2 production in reduced ceftazidime and ertapenem susceptibility in a K. pneumoniae ST258 clinical isolate. Methods MICs were determined by the agar dilution method. WGS was conducted to identify and compare resistance genes between isolates. Expression of KPC-2 was quantified by quantitative RT–PCR and immunoblotting. OXA-2 was expressed in Escherichia coli TOP10, as well as in K. pneumoniae ATCC 13883, to define the relative contribution of OXA-2 in β-lactam resistance. Kinetic studies were conducted using purified OXA-2 enzyme. Results K. pneumoniae 1761 belonged to ST258 and carried both blaKPC-2 and blaOXA-2. However, expression of blaKPC-2 was substantially reduced due to an IS1294 insertion in the promoter region. K. pneumoniae 1761, K. pneumoniae ATCC 13883 and E. coli TOP10 carrying blaOXA-2-harbouring plasmids showed reduced susceptibility to ertapenem and ceftazidime, but meropenem, imipenem and cefepime were unaffected. blaOXA-2 was carried on a 2910 bp partial class 1 integron containing aacA4-blaOXA-2-qacEΔ1-sul1 on an IncA/C2 plasmid, which was not present in the earlier ST258 isolates possessing blaKPC-2 with intact promoters. Hydrolysis of ertapenem by OXA-2 was confirmed using purified enzyme. Conclusions Production of OXA-2 was associated with reduced ceftazidime and ertapenem susceptibility in a K. pneumoniae ST258 isolate.

Enterococcus faecalis is a leading cause of infective endocarditis (IE), especially among older patients with comorbidities. Here we investigated the genomic diversity and antimicrobial susceptibility of 33 contemporary E. faecalis isolates from definite or probable IE cases at the University of Pittsburgh Medical Center (UPMC) between 2018 and 2020. Isolates belonging to two multi-locus sequence types (STs), ST6 and ST179, were isolated from nearly 40% of IE patients. Both of these dominant STs carried known beta-lactam resistance-associated mutations affecting the low-affinity penicillin-binding protein 4 (PBP4). We assessed the ability of ampicillin and ceftriaxone (AC) both alone and in combination to inhibit genetically diverse E. faecalis IE isolates in checkerboard synergy assays and an in vitro one-compartment pharmacokinetic-pharmacodynamic (PK-PD) model of AC treatment. ST6 isolates as well as an isolate with a mutation in the PP2C-type protein phosphatase IreP had higher ceftriaxone MICs compared to other isolates, and showed diminished in vitro synergy of AC. Additionally, both ST6 and ST179 isolates exhibited regrowth after 48 hours of humanized exposures to AC. Overall, we found evidence for diminished in vitro AC activity among E. faecalis IE isolates with PBP4 and IreP mutations. This study highlights the need to evaluate alternate antibiotic combinations in clinical practice against diverse contemporary E. faecalis IE isolates.

Acinetobacter baumannii is currently classified as one of six pathogens that contribute to increased patient mortality. Thus, exploratory studies navigating alternative treatment strategies are of supreme interest. Herein, we completed minimum inhibitory concentration (MIC) testing, and time-kill analyses (TKA) on 50 carbapenem-resistant Acinetobacter baumannii isolates including 28 colistin-resistant isolates. Upon testing of MEM or TGC in the presence of sub-inhibitory COL against the 50 isolates, there was a median 2-fold reduction in MEM and TGC MICs. In the TKAs, the COL+MEM combination was synergistic in 45 (90%) isolates and bactericidal in 43 (86%) isolates at 24 hours, whereas the COL+TGC combination TKAs demonstrated synergy in 32 (64%) isolates and bactericidal activity was shown in 28 (56%) isolates. Additionally, sulbactam (SUL) and TGC were added to the COL+MEM dual therapy regimen to assess the possible utility of a triple therapy regimen against five non-responsive isolates. The COL+MEM+SUL and COL+MEM+TGC regimens effectively restored synergy in (5/5) 100% of the isolates. The results of this study demonstrate the potential utility of COL combinations in the treatment of carbapenem-resistant isolates.

Background Urinary tract infections (UTIs) are the most common infections caused by Gram-negative bacteria and represent a major healthcare burden. Carbapenem-resistant (CR) strains of Enterobacterales and non-lactose fermenting pathogens further complicate treatment approaches. Methods We conducted a retrospective analysis of the US Premier Healthcare Database (2014–2019) in hospitalised adults with a UTI to estimate the healthcare burden of Gram-negative CR UTIs among patients with or without concurrent bacteraemia. Results Among the 47,496 patients with UTI analysed, CR infections were present in 2076 (4.4%). Bacteraemia was present in 24.5% of all UTI patients, and 1.7% of these were caused by a CR pathogen. The most frequent CR pathogens were Pseudomonas aeruginosa (49.4%) and Klebsiella pneumoniae (14.2%). Patients with CR infections had a significantly longer hospital length of stay (LOS) (median [range] 8 [5–12] days vs 6 [4–10] days, P P P P P Conclusions Among hospitalised patients with UTIs, the presence of a CR organism and bacteraemia increased the burden of disease, with worse outcomes and higher hospitalisation charges than disease associated with CS pathogens and those without bacteraemia.

Background: Multidrug-resistant (MDR) Pseudomonas aeruginosa is a major public health threat. Treatment with ceftolozane-tazobactam improves patient outcomes compared to salvage therapy; however, resistance has emerged in ~15% of patients following courses ranging from 7 to 53 days. Understanding the development and mechanisms of resistance in these difficult to treat MDR P. aeruginosa has public health importance. Our objective was to study the in vitro activity of alternative β-lactams in the setting of ceftolozane-tazobactam resistance. Methods: Isolates from 23 patients in whom ceftolozane-tazobactam resistance emerged were selected for analysis. Minimum inhibitory concentrations (MICs) were determined by standard broth microdilution in triplicate and interpreted by CLSI breakpoints. Mechanisms of resistance and relatedness of isolates were explored through whole-genome sequence (WGS) analysis in 15 patients from whom baseline and post-treatment isolates were available. Results: 23 baseline and 32 post-treatment isolates were included. The median baseline ceftolozane-tazobactam MIC was 2 µg/mL (range: 0.5 – 8 µg/mL). 75%, 25%, 82.6%, and 83.3% of baseline isolates were non-susceptible to ceftazidime, ceftazidime-avibactam, imipenem, and piperacillin-tazobactam respectively. Following a median 16 (range: 3- 60) days of therapy, the median post-exposure ceftolozane-tazobactam MIC was 64 µg/mL (range: 8 – >256 µg/mL). 100%, 72.7%, 69.6%, and 79.2% of post-treatment isolates were resistant to ceftazidime, ceftazidime-avibactam, imipenem, and piperacillin-tazobactam. The corresponding MIC foldchanges were 4, 8, -2, and 0, respectively. Median imipenem-relebactam MICs did not change before or after treatment with ceftolozane-tazobactam (median= 2 µg/mL for both) and 16.7% were classified as resistant. WGS data revealed several mutations in ampC and ampR sequences. Discussion: Our findings show that resistance to ceftolozane-tazobactam impacts the susceptibility of alternative β-lactams. Cross resistance occurs with ceftazidime and ceftazidimeavibactam (median 4 and 8 fold MIC increase, respectively). Imipenem MICs are decreased 2fold potentially demonstrating collateral sensitivity. Piperacillin-tazobactam MICs were unchanged and isolates remained resistant. Importantly, imipenem-relebactam MICs were unchanged suggesting the mechanism of ceftolozane-tazobactam resistance may be due to structural changes in ampC. WGS data showed a number of different mutations in both ampC and ampR. Certain mutations, such as F147L and mutations found in positions 234-244, were found to promote resistance to ceftolozane-tazobactam.

We describe a case of a 48 years old male with left sided endocarditis and septic emboli secondary to a Pseudomonas aeruginosa strain that developed resistance to other β-lactam antibiotics during therapy resulting in prolonged bacteremia. Blood cultures sterilized within 1 day of initiating ceftolozane/tazobactam 3 g every 8 hours in combination with ciprofloxacin. Steady state free ceftolozane plasma Cmax and Cmin concentrations were calculated to be 122.2μg/mL and 24.3μg/mL, respectively. The multidrug-resistant strain harbored chromosomal β-lactamases OXA-486 and PDC-3, mutations in ampD and dacB predicted to lead to ampC over-expression, and mutations in OprD predicted to decrease outer membrane permeability. Following completion of a 42 day course and aortic valve replacement, the patient was deemed clinically cured without recurrence of infection at follow up 2 years later. To our knowledge, this is the first reported case to measure ceftolozane concentrations during the treatment of endocarditis which supports dose optimization approaches of severe endovascular disease due to multidrug resistant pathogens.

Background Cefepime (FEP) induced neurotoxicity (NT) may have serious implications for patients (pts). Retrospective studies have employed variable definitions of NT, finding renal impairment and FEP trough concentrations (Cmin) > 20 mg/L as risk factors. Prospective studies comparing antibiotics have not been performed. Methods We conducted a prospective study of pts receiving FEP or meropenem (MEM) with neurologic evaluation and therapeutic drug monitoring (TDM). A NT advisory board (NTAB) was established to develop standardized definitions of possible, probable and definitive NT (Fig 1). Cases of potential NT were adjudicated by the NTAB who were blinded to study treatment. FEP and MEM midpoint and Cmin concentrations were measured at steady-state by validated methods. Results 127 patients were included (70 FEP, 57 MEM). Demographics and treatment characteristics were similar between groups (Fig 2); 63% were in the ICU. FEP and MEM Cmin varied from 1.9 – 140.5 and 0.6 – 31.3 mg/L, respectively. Median FEP Cmin and total exposures (AUC) were 23.1 mg/L and 347.6 hr*mg/L, respectively. Corresponding MEM values were 5.9 mg/L and 124.8 hr*mg/L, respectively. Cmin values were inversely correlated with renal function for both FEP and MEM (P< 0.001). Rates of possible, probable, or definitive NT were 10% and 5% for FEP and MEM, respectively (P=0.51; Fig 3). 16% and 3% of pts with FEP Cmin > or < 20 mg/L had NT, respectively (P=0.11; Fig 4). Median MEM Cmin were 12.3 and 5.4 mg/L among pts with and without NT, respectively (P=0.09; Fig 4). Rates of NT did not vary by infusion length or dose. FEP and MEM exposures were similar between patients with (17%) or without (83%) microbiologic recurrence due to the same pathogen. FEP was discontinued in 4 pts due to NT; no pts stopped MEM due to NT. Conclusion Our study is the first to evaluate FEP NT prospectively and compare rates of NT to pts receiving MEM. We established criteria that were applied by a blinded NTAB. In doing so we found rates of NT to be lower than previously reported and not statistically different between FEP and MEM. Cmin values were highly variable and associated with numerically, but not statistically higher rates of NT for both agents. These findings serve as the basis for larger, multicenter studies and justify use of routine TDM to limit NT among high-risk pts. Disclosures Brandon Smith, MD, PharmD, Shionogi (Consultant, Advisor or Review Panel member) Alexandra Urban, MD, Neuropace (Consultant) Ryan K. Shields, PharmD, MS, Shionogi (Consultant, Research Grant or Support)