Daptomycin Nonsusceptibility Research Articles

Combinations of (lipo)glycopeptides with β-lactams against MRSA: susceptibility insights.

Increasing application of vancomycin due to the high prevalence of MRSA infections has led to the emergence of vancomycin intermediate-resistant Staphylococcus aureus (VISA) and heterogeneous VISA (hVISA). Consequently, the need for alternative therapies that target MRSA has become evident. To evaluate the synergy between (lipo)glycopeptides (LGP/GPs) (vancomycin, teicoplanin, telavancin, dalbavancin and oritavancin) and β-lactams (ceftaroline, cefepime, cefazolin and oxacillin) against MRSA, hVISA, VISA and daptomycin non-susceptible (DNS) phenotypes. Twenty randomly selected clinical MRSA strains (i.e. 5 MRSA, 5 hVISA, 5 VISA and 5 DNS) were assessed versus LGP/GPs alone and LGP/GPs in combination with β-lactams for MICs. Although verification of antibiotic potency against bacterial strains is assessed by the microbroth dilution (MBD) MIC method recommended by the CLSI, some antibiotics need modified assay conditions in order to demonstrate their optimal activity. Addition of β-lactams reduced MIC values of LGP/GPs against all strains (up to 160-fold reduction). In general, LGPs (dalbavancin, oritavancin and telavancin) were more active (significant differences in MIC values, up to 8-fold) compared with vancomycin and teicoplanin. The majority of these combinations were bactericidal and superior to any single agent. This report has examined the susceptibility patterns of LGP/GPs and their combination with β-lactams. Of interest, the impact of susceptibility tests (in terms of MIC plates and their surface area) on the synergistic activity in 24 h time-kill experiments was apparent for LGPs. Further clinical research is required to investigate synergy with LGP/GPs and β-lactams against these Staphylococcus strains.

Metabolic changes associated with adaptive resistance to daptomycin in Streptococcus mitis-oralis

BackgroundViridans group streptococci of the Streptococcus mitis-oralis subgroup are important endovascular pathogens. They can rapidly develop high-level and durable non-susceptibility to daptomycin both in vitro and in vivo upon exposure to daptomycin. Two consistent genetic adaptations associated with this phenotype (i.e., mutations in cdsA and pgsA) lead to the depletion of the phospholipids, phosphatidylglycerol and cardiolipin, from the bacterial membrane. Such alterations in phospholipid biosynthesis will modify carbon flow and change the bacterial metabolic status. To determine the metabolic differences between daptomycin-susceptible and non-susceptible bacteria, the physiology and metabolomes of S. mitis-oralis strains 351 (daptomycin-susceptible) and 351-D10 (daptomycin non-susceptible) were analyzed. S. mitis-oralis strain 351-D10 was made daptomycin non-susceptible through serial passage in the presence of daptomycin.ResultsDaptomycin non-susceptible S. mitis-oralis had significant alterations in glucose catabolism and a re-balancing of the redox status through amino acid biosynthesis relative to daptomycin susceptible S. mitis-oralis. These changes were accompanied by a reduced capacity to generate biomass, creating a fitness cost in exchange for daptomycin non-susceptibility.ConclusionsS. mitis-oralis metabolism is altered in daptomycin non-susceptible bacteria relative to the daptomycin susceptible parent strain. As demonstrated in Staphylococcus aureus, inhibiting the metabolic changes that facilitate the transition from a daptomycin susceptible state to a non-susceptible one, inhibits daptomycin non-susceptibility. By preventing these metabolic adaptations in S. mitis-oralis, it should be possible to deter the formation of daptomycin non-susceptibility.

Daptomycin susceptibility in enterococci isolated from urinary samples in a tertiary care hospital

are gram positive cocci encountered in various human infections.The most striking feature of is their relative and absolute resistance to variety of antibiotics.Vancomycin resistant (VRE) were first described in late 1980 and the incidence of nosocomial infections by VRE being 11.9% in Asia Pacific and 35.5% in U.S. Daptomycin is a lipopeptide antibiotic with bactericidal activity . To study the daptomycin susceptibility in isolated in urinary samples. A total of 76 strains of were isolated during the study period.25 (33%) were identified as and 51 (67%) as Most of the Enterococcal isolates (82.8%) were resistant to Norfloxacin followed by 71% being resistant to tetracycline and 76% of isolates were sensitive to nitrofurantoin. Most of the isolates were multidrug resistant. Of the 76 isolated only 2(2.63%) were VRE and both the VRE strains isolated were . There was no resistance to linezolid including the VRE strains. All the isolates 76 (100%) were sensitive to daptomycin including the VRE strains. Our study mainly focused on susceptibility of enterococcal isolates in urinary samples to daptomycin which has bactericidal activity against gram positive organisms including multidrug resistant strains. This study shows growing concern for increasing antibiotic resistance in enterococcal isolates and treatment of these infections with daptomycin is invited as there are only few reports of daptomycin non susceptible (DNSE).

In-Host Evolution of Daptomycin Resistance and Heteroresistance in Methicillin-Resistant Staphylococcus aureus Strains From Three Endocarditis Patients.

Daptomycin is considered an important alternative for the treatment of methicillin-resistant Staphylococcus aureus (MRSA). However, treatment failures associated with daptomycin nonsusceptibility isolates have been reported in recent years. In this study, we investigated serial MRSA strains from 3 endocarditis patients who had breakthrough bacteremia, despite treatment with daptomycin. The strains were analyzed by whole-genome sequencing, molecular typing, and mutation screening. Population analysis and growth curves were also applied to evaluate heteroresistance and fitness cost. This series of MRSA strains belonged to ST5, ST59, and ST4513. The daptomycin minimum inhibitory concentrations for these MRSA strains increased after daptomycin exposure, whereas daptomycin-resistant strains emerged with mutations in mprF and yycH. Population analysis profiling results demonstrated the presence of a daptomycin-heteroresistant subpopulation among daptomycin-susceptible MRSA strains, and no significant fitness cost was observed within these heteroresistant MRSA clones. We confirmed that daptomycin heteroresistance and resistance could emerge rapidly in MRSA strains of different lineages after daptomycin exposure. Further studies to fully understand the mechanism(s) underlying daptomycin resistance in MRSA are required.

Detection of Antimicrobial Resistance-Related Changes in Biochemical Composition of Staphylococcus aureus by Means of Atomic Force Microscopy-Infrared Spectroscopy

The development of antimicrobial resistance (AMR) resulting from widespread antibiotic usage is occurring at an alarming pace, much faster than our understanding of the mechanisms behind resistance. Knowledge about resistance-related phenotypic and genotypic changes is critical for the development of new drugs. Here, we identify changes in the chemical composition of Staphylococcus aureus associated with the development of resistance to last resort drugs, vancomycin and daptomycin, using a novel, single cell, nanoscale technique, atomic force microscopy-infrared spectroscopy (AFM-IR), combined with chemometric analysis. We utilized paired clinical isolates, with the parent (susceptible) strain isolated prior to treatment and the daughter (resistant) strain obtained from the same patient after drug admission and clinical failure. We observed an increase in the amount of nonintracellular carbohydrates, indicating thickening or changes in the packing of the cell wall, as well as changes in the phospholipid content in relation to vancomycin resistance and daptomycin nonsusceptibility, respectively.

627. Molecular Epidemiology of Daptomycin-resistant Staphylococcus aureus Causing Bloodstream Infections

BackgroundDaptomycin (Dap) has become increasingly important to combat infections caused by Staphylococcus aureus (SA). However, reports of daptomycin nonsusceptible (DNS) SA strains have emerged over the recent years, specifically with alteration of the MprF gene. Mutations in this gene lead to an increase in lysyl-phosphatidyl glycerol production resulting in a lower susceptibility to Dap and cationic antimicrobial peptides. Point mutations in the cls2 gene have also been associated with DNS SA through similar mechanisms of action. This study describes the gene mutations involved in our patients with SA bloodstream infections (BSIs) that developed Dap resistance.MethodsThis is a retrospective study in a tertiary healthcare system in southeast Michigan. SA blood isolates were sequenced through a gene-by-gene approach using Ridom SeqSphere. Charts were reviewed for clinical and laboratory data. Dap minimum inhibitory concentrations (MICs) were determined by E-test. DNS was defined as an MIC > 1.0 µg/mL.ResultsFrom December 3, 2015 to May 8, 2017, 7 patients were identified with DNS SA BSI. Mean age of the patients was 59 (SD = 13.7) and 86% were male. Mean bacteremia duration was 7 days (SD = 6.3), mean hospital length of stay was 13 days (SD = 9.8), 5 isolates were methicillin-resistant SA, and 2 were methicillin-sensitive SA. When comparing molecular sequencing of all seven patients with their Dap-sensitive strains vs. resistant strains, all patients, except for patient 3, had a point mutation at different allele loci in the MprF gene (Table 1). Additionally, two patients had point mutations in the cls2 gene. When comparing all 27 strains from the seven patients, each cluster was attributed to an individual patient with the exception of cluster one that included patients 5 and 6 suggesting transmission (Figure 1).ConclusionDNS SA is infrequent; however, the present study shows that it can emerge both from methicillin-resistant and -susceptible SA most commonly when developing MprF and cls2 gene mutations as seen in our patients. Patient 3 was the exception in our analysis and likely developed DNS SA through another gene mutation. Further studies are necessary to know the frequency of each mutation with all genes associated with DNS SA, and epidemiological risk factors of transmission. Disclosures All authors: No reported disclosures

Daptomycin resistance in methicillin-resistant Staphylococcus aureus is conferred by IS256 insertion in the promoter of mprF along with mutations in mprF and walK

Mechanisms underlying the emergence of daptomycin resistance in Staphylococcus aureus remain unclear. In this study, Staphylococcus aureus strain 3d0, isolated from a patient with bloodstream infection and belonging to the predominant Chinese hospital-associated methicillin-resistant S. aureus (MRSA) clone ST239, was serially passaged on gradient broth containing daptomycin for 34 days. The whole genomes of 3d0 and its serial passage strains were sequenced and compared. Five single nucleotide polymorphisms, four IS256 insertions, and one 39-bp insert occurred in the progress of daptomycin resistance acquisition. IS256 insertion in the mprF promoter region resulted in mprF overexpression. Two novel point mutations in mprF and walK, leading to amino acid substitutions in MprF (G299V and L473I) and WalK (L7Q and Y225N), were shown by allelic replacement experiments to increase the minimum inhibitory concentration (MIC) of daptomycin by 2-4 times. Allelic replacement of both mprF and walK in strain 3d0 increased the daptomycin MIC by 4-8-fold, indicating that mprF and walK mutations synergistically contribute to daptomycin non-susceptibility. Notably, these mutants acquired resistance without losing fitness and exhibited decreased expression of cell wall degradation-related genes. In conclusion, this study revealed novel mutations of MRSA daptomycin resistance acquisition in vitro as well as several novel mutations in walK and mprF, and includes the first in-depth analysis of the mprF promoter. This study sheds light on how MRSA may acquire daptomycin resistance during daptomycin treatment.

Metabolic interventions for the prevention and treatment of daptomycin non-susceptibility in Staphylococcus aureus.

A major developing problem in the treatment of Staphylococcus aureus infections is the emergence of resistance during treatment with daptomycin. Previous metabolomic analyses of isogenic S. aureus strains prior to and after evolution into a daptomycin non-susceptible (DapNS) state provided important metabolic information about this transition (e.g. perturbations of the tricarboxylic acid cycle). To assess the significance of these metabolic changes, in vitro susceptibility to daptomycin was determined in daptomycin-susceptible (DapS) and DapNSS. aureus strains cultivated with metabolic inhibitors targeting these changes. Only inhibitors that are approved for use in humans were chosen (i.e. fosfomycin, valproate, trimetazidine and 6-mercaptopurine) to assess the importance of metabolic pathways for daptomycin non-susceptibility. The ability of these inhibitors to forestall the emergence of DapNS strains was also assessed. The combination of daptomycin and fosfomycin synergistically killed both DapS and DapNS strains in vitro and enhanced the in vivo outcome against a DapNS strain in experimental endocarditis. Interestingly, fosfomycin acts on the peptidoglycan biosynthetic enzyme UDP-N-acetylglucosamine enolpyruvyl transferase (MurA); however, it also had a significant effect on the enzymatic activity of enolase, an essential enzyme in S. aureus. While fosfomycin acted synergistically with daptomycin, it failed to prevent the in vitro evolution of daptomycin non-susceptibility. In contrast, trimetazidine, an anti-angina drug that stimulates glucose oxidation, abolished the ability of DapSS. aureus strains to transition to a DapNS state. These data reveal that metabolic adaptations associated with DapNS strains can be targeted to prevent the emergence of and/or reverse pre-existing resistance to daptomycin.

Dalbavancin Alone and in Combination with Ceftaroline against Four Different Phenotypes of Staphylococcus aureus in a Simulated Pharmacodynamic/Pharmacokinetic Model.

Glycopeptides such as vancomycin have been used as the first-line therapy against MRSA infections for over half a century. Reduced susceptibility and emergence of resistance to first-generation glycopeptides has led to development of second-generation lipoglycopeptide derivatives such as dalbavancin which hold broader ranges of activity and enhanced pharmacokinetic properties. We evaluated the MIC values for a total of 100 isolates, including 25 methicillin-resistant Staphylococcus aureus (MRSA), 25 heterogeneus vancomycin-intermediate S. aureus, 25 daptomycin nonsusceptible (DNS), and 25 vancomycin-intermediate S. aureus strains against dalbavancin, ceftaroline, and vancomycin alone and in combination. Dalbavancin was highly active against hVISA, DNS, and MRSA strains, achieving 96 to 100% susceptibility and 72% susceptibility against VISA strains. The combination of dalbavancin plus ceftaroline reduced dalbavancin MICs 62.5-fold and demonstrated enhanced killing against all four phenotypes in pharmacokinetic/pharmacodynamic models. Four strains of the aforementioned phenotypes were randomly chosen for pharmacodynamic/pharmacokinetic simulation models. Of interest, while both dalbavancin and vancomycin in combination with ceftaroline demonstrated significant improvement in glycopeptide fAUC/MIC values against these four phenotypes, the dalbavancin-ceftaroline combinations exhibited a 44- to 11,270-fold higher fAUC/MIC value in comparison to vancomycin-ceftaroline combinations. In addition, the time to detection limit was reduced for this combination (24 to 32 h) versus the vancomycin-ceftaroline combination (24 to 72h). To our knowledge, this is the first comprehensive study of dalbavancin and vancomycin combinations with ceftaroline. These data provide a novel approach for combating recalcitrant MRSA infections.

Insights Into the Evolution of Staphylococcus aureus Daptomycin Resistance From an in vitro Bioreactor Model.

The extensive use of daptomycin for treating complex methicillin-resistant Staphylococcus aureus infections has led to the emergence of daptomycin-resistant strains. Although genomic studies have identified mutations associated with daptomycin resistance, they have not necessarily provided insight into the evolution and hierarchy of genetic changes that confer resistance, particularly as antibiotic concentrations are increased. Additionally, plate-dependent in vitro analyses that passage bacteria in the presence of antibiotics can induce selective pressures unrelated to antibiotic exposure. We established a continuous culture bioreactor model that exposes S. aureus strain N315 to increasing concentrations of daptomycin without the confounding effects of nutritional depletion to further understand the evolution of drug resistance and validate the bioreactor as a method that produces clinically relevant results. Samples were collected every 24 h for a period of 14 days and minimum inhibitory concentrations were determined to monitor the acquisition of daptomycin resistance. The collected samples were then subjected to whole genome sequencing. The development of daptomycin resistance in N315 was associated with previously identified mutations in genes coding for proteins that alter cell membrane charge and composition. Although genes involved in metabolic functions were also targets of mutation, the common route to resistance relied on a combination of mutations at a few key loci. Tracking the frequency of each mutation throughout the experiment revealed that mutations need not arise progressively in response to increasing antibiotic concentrations and that most mutations were present at low levels within populations earlier than would be recorded based on single-nucleotide polymorphism (SNP) filtering criteria. In contrast, a serial-passaged population showed only one mutation in a gene associated with resistance and provided limited detail on the changes that occur upon exposure to higher drug dosages. To conclude, this study demonstrates the successful in vitro modeling of antibiotic resistance in a bioreactor and highlights the evolutionary paths associated with the acquisition of daptomycin non-susceptibility.

Utilizing genomic analyses to investigate the first outbreak of van A vancomycin-resistant Enterococcus in Australia with emergence of daptomycin non-susceptibility

The majority of vancomycin-resistant Enterococcus faecium (VREfm) in Australia is of the vanB genotype. An outbreak of vanA VREfm emerged in our haematology/oncology unit between November 2014 and May 2015. The first case of daptomycin non-susceptible E. faecium (DNSEfm) detected was a patient with vanA VREfm bacteraemia who showed clinical failure of daptomycin therapy, prompting microbiologic testing confirming daptomycin non-susceptibility. To describe the patient profiles, antibiotic susceptibility and genetic relatedness of vanA VREfm isolates in the outbreak. Chart review of vanA VREfm colonized and infected patients was undertaken to describe the demographics, clinical features and outcomes of therapy. Whole genome sequencing of vanA VREfm isolates involved in the outbreak was conducted to assess clonality. In total, 29 samples from 24 patients tested positive for vanA VREfm (21 screening swabs and 8 clinical isolates). Five isolates were DNSEfm (four patients colonized, one patient with bacteraemia), with only one patient exposed to daptomycin previously. In silico multi-locus sequence typing of the isolates identified 25/26 as ST203, and 1/26 as ST796. Comparative genomic analysis revealed limited core genome diversity amongst the ST203 isolates, consistent with an outbreak of a single clone of vanA VREfm. Here we describe an outbreak of vanA VREfm in a haematology/oncology unit. Genomic analysis supports transmission of an ST203 vanA VRE clone within this unit. Daptomycin non-susceptibility in 5/24 patients left linezolid as the only treatment option. Daptomycin susceptibility cannot be assumed in vanA VREfm isolates and confirmatory testing is recommended.

1222. Risk Factors and Outcomes for Daptomycin Nonsusceptible Methicillin-Resistant Staphylococcus aureus Bloodstream Infections

BackgroundDaptomycin (dap) has been approved and successfully used for the treatment of methicillin-resistant Staphylococcus aureus (MRSA) infections. However, reports of daptomycin nonsusceptible (DNS) MRSA strains have emerged over the recent years. This study describes the clinical characteristics of patients with DNS MRSA bloodstream infections (BSIs) with the objective of identifying risk factors and outcomes.MethodsThis is a retrospective case–control study in a tertiary healthcare system in southeast Michigan. Cases included 34 patients with DNS MRSA BSI between September 24, 2005 and March 31, 2018. Cases were matched with controls with MRSA BSI based on age, source of BSI, and time-period of BSI in a 1:1 ratio. Charts were reviewed for clinical and laboratory data. Vancomycin (van) and dap minimum inhibitory concentrations (MICs) were determined by E-test. DNS was defined as an MIC >1.0 µg/mL. Chi-square test, Fisher’s exact test, and t-test were used to determine statistical significance.ResultsIn the case cohort, the source of BSI was endovascular in 11(32%) patients, central-line associated in 3(9%), secondary BSI in 13(38%), and unknown in 7(21%). Table 1 is a summary of the results.Table 1. Clinical Characteristics and Outcomes of Cases and ControlsCasesControls N = 34(%) N = 34(%) P-valueMean age (SD)63.5 (12.0)61.9 (11.2)0.572Male18 (52.9)21 (61.8)0.462Mean bacteremia duration in days (SD)4.4 (3.2)5.9 (4.9)0.195Mean LOS in days (SD)19.5 (13.6)18.4 (14.6)0.751Mean van MIC (SD)2.04 (1.19)1.39 (0.36)0.003Mean dap MIC (SD)2.69 (1.32)0.57 (0.24)<0.0001Epidemiologic acquisitionCommunity-acquired0 (0)9 (26.5)0.002Healthcare-associated21 (63.6)22 (64.7)0.927Hospital-acquired12 (36.4)3 (8.8)0.00790-day prior dap exposure23 (82.1)3 (9.7)<0.0001Mean dap exposure in days23.6 (21.0)2.68 (10.6)<0.000190-day prior van exposure25 (89.3)9 (29)<0.0001Mean van exposure in days13.0 (14.7)4.19 (12.7)0.02030-day mortalitya10 (32.3)6 (18.8)0.218Mean Charlson Comorbidity Index (SD)5.7 (3.07)4.4 (2.9)0.07790-day MRSA BSI recurrencea8 (44.4)2 (9.5)0.025 aFrom date of index BSI.ConclusionPrior exposure to dap and van, and higher van MIC in MRSA isolates are risk factors for DNS MRSA BSI. DNS is associated with significantly higher risk of 90-day MRSA BSI recurrence.Disclosures All authors: No reported disclosures.

1798. Phenotypic and Genotypic Impact of Antibiotic Stewardship Intervention on Daptomycin-Nonsusceptible Enterococcus faecium (DNSE) Clinical Isolates

BackgroundSince its introduction in 2009, use of daptomycin for treatment of enterococcal infections has resulted in the emergence of DNSE. Between 2009 and 2013, daptomycin nonsusceptibility among E. faecium was closely associated with emergence of a unique and dominant clone ST736 in our institution. In 2014, we instituted targeted measures to optimize the use of daptomycin. In this study, we describe the significant phenotypic and genotypic impact of reduced daptomycin use on clinical enterococcal isolates.MethodsEnterococcal clinical isolates were recovered from January 2014 through December 2017. Daptomycin susceptibility was determined by MicroScan WalkAway™ System and confirmed by E-test. Selected DNSE and vancomycin-resistant E. faecium (VREfm) clinical isolates were analyzed by next-generation sequencing (NGS) using the Illumina systems to provide multilocus sequencing type (MLST). Daptomycin utilization data were extracted from pharmacy records.ResultsTargeted antibiotic stewardship initiatives consisted of preapproval, daily review for optimization of dose and duration, rapid de-escalation, consideration for appropriate alternative antibiotics for select disease syndromes and stopping of inappropriate daptomycin therapy. Over 4 years, this lead to a 39% reduction in overall use of daptomycin. Besides direct cost saving, this reduced use was associated with significant reduction in daptomycin nonsusceptibility from 12% to 4%, lowering of MIC90 from 8 to 4 μg/mL, and a clonal shift from dominant ST736 to ST117.Daptomycin Usage Days of Therapy /1,000 Days (Monthly Average)Phenotypic ChangesGenotypic ChangesMIC50 (μg/mL)MIC90 (μg/mL)% DNSEDominant Genotype Among VREfm20147424812 ST736 (38%)ST18 (21%)ST412 (12%)ST117 (11%)2015633461020165764472017455444ST736 (5%)ST18 (4%)ST412 (11%)ST117 (41%)ConclusionA targeted antibiotic stewardship initiative to address rising rate of daptomycin nonsusceptibility among E. faecium, resulted in significant phenotypic and genotypic changes among clinical isolates. This study also shows successful integration of NGS in a clinical microbiology lab to validate phenotypic changes of daptomycin nonsusceptibility and to help design future infection control and antibiotic stewardship endeavors.Disclosures A. Dhand, Merck: Speaker’s Bureau, Speaker honorarium. Astellas: Scientific Advisor, Consulting fee.

2452. Treatment and Outcomes of Daptomycin-Nonsusceptible Methicillin-Resistant Staphylococcus aureus Bloodstream Infections

BackgroundDaptomycin (dap) is approved as an alternative to vancomycin (van) for therapy of methicillin-resistant Staphylococcus aureus (MRSA) bloodstream infection (BSI). Cases of therapy failure associated with the emergence of daptomycin-nonsusceptible (DNS) MRSA strains have been documented. Information on the treatment and outcome of DNS MRSA BSI is scarce. This study describes the treatment and outcome of patients with DNS MRSA BSI at our healthcare center.MethodsThis is a retrospective review of patients with DNS (E-test MIC >1.0 µg/mL) MRSA BSI at a tertiary healthcare center in Detroit, Michigan between September 24, 2005 and March 31, 2018. The variables collected were: BSI source, inpatient and discharge antibiotic therapy, BSI duration, in-hospital and 90-day mortality, and 90-day MRSA BSI recurrence. Inpatient therapy was defined as the treatment used for the most consecutive days from index DNS MRSA blood culture during hospitalization. Discharge therapy is the treatment used post-discharge or on the expiration date. Antibiotics used for ≤2 days were excluded.ResultsA total of 32 nonduplicate patients with DNS MRSA BSI were identified. One patient with an inaccessible chart was excluded. The source of BSI was endovascular in 9 (29%) patients, secondary BSI in 14 (45%), central-line associated in 3 (10%), and unknown in 5 (16%). A total of 24 different antibiotic regimens were used to treat DNS MRSA BSI. Van monotherapy was the most commonly used regimen for inpatient and discharge therapy, followed by dap + ceftaroline (cef). Table 1 is a summary of the results.Table 1:Treatment and Outcomes of Patients with DNS MRSA BSIInpatient TherapyDischarge Therapy (n)In-Hospital Mortality, n(%)90-Day Mortality, n(%)Mean BSI Duration (days)90-Day BSI Recurrence, n(%)van (10)van (8) cef (1) dap + cef (1)3(30)4(40)2.93(30)*dap + cef (5)cef + dap (3) cef + van (1) van (1)0(0)0(0)*4.41(20)**lin ± gen ± rif (5)lin (3) van + sxt (1) quin/dal (1)1(20)3(60)6.81(20)other (11)4(36)4(36)3.52(22)*Totals6(26)11(35)4.47(23)gen = gentamycin; rif = rifampin; lin = linezolid; sxt = TMP-SMX; quin/dal = quinupristin/dalfopristin.*1 pt with unknown status.** 3 patients with unknown status.ConclusionA variety of therapeutic regimens was used to treat DNS MRSA BSI in our cohort. However, van monotherapy was the most common inpatient and discharge regimen.Disclosures All authors: No reported disclosures.

1019. Treatment Outcomes for Enterococcus faecium Bacteremia in Solid-Organ Transplant Patients: Implications for Daptomycin

BackgroundOptimal antimicrobial therapy for Enterococcus faecium (EFM) bacteremia in the solid-organ transplant (SOT) population is not well defined. Antimicrobial resistance, immunosuppression, and high mortality associated with EFM infections all pose serious threats. The purpose of this study was to describe the pharmacotherapy and outcomes of EFM bacteremia in SOT patients.MethodsThis was a single-center retrospective cohort of SOT patients with EFM bloodstream infection from 2013 to 2018. Susceptibility of ampicillin (AMP), vancomycin (VAN), linezolid (LZD), and daptomycin (DAP) against EFM were reported as MIC90 when available. The primary outcome, 30-day all-cause mortality, was assessed in bivariate analysis to identify potential risk factors. Secondary outcomes included inpatient mortality and development of DAP nonsusceptibility (DNS).ResultsForty-four unique cases representing 40 patients were included in the analysis. The median age was 62.5 years and liver (65.9%), intestine (20.5%), and kidney (11.4%) were the most common organs transplanted. The MIC90 of VAN, DAP, and LZD of initial isolates collected were >32 mg/L, 4 mg/L, and 2 mg/L, respectively; all were AMP resistant. The median durations of hospitalization and intensive care stay were 29 days and 17.5 days, respectively. Most patients had indwelling central lines (81.8%) at the time of bacteremia; intra-abdominal abscesses/fluid collections were present in 45.5% of patients and 9.1% had endocarditis. The most common definitive antimicrobial regimens were DAP plus β-lactam (45.5%), DAP monotherapy (18.2%), and LZD 600 mg Q12H (25.0%). The mean initial and definitive DAP doses were 8.1 ± 1.6 and 8.9 ± 1.7 mg/kg actual body weight, respectively. Among subjects that received DAP, 21.9% developed DNS. Inpatient mortality was 39.5% and 30-day mortality was 27.3%. Mortality at 30-days was greater in patients with high-grade bacteremia (40.7 vs. 5.9%, P = 0.01) and receipt of DAP <10 mg/kg as the first active antibiotic (42.9 vs. 13.0%, P = 0.03).ConclusionInadequate DAP dosing for EFM bacteremia may be associated with mortality in the SOT population. Larger, matched analyses are necessary to determine the impact of optimized pharmacodynamics. Disclosures S. L. Davis, Achaogen: Scientific Advisor, Consulting fee. Allergan: Scientific Advisor, Consulting fee. Melinta: Scientific Advisor, Consulting fee. Nabriva: Scientific Advisor, Consulting fee. Zavante: Scientific Advisor, Consulting fee.

1225. High Rate of Linezolid (LZD) Nonsusceptibility (LNS) Among Enteric Vancomycin-Resistant Enterococci (VRE) Recovered From Hospitalized Patients Actively Screened for VRE Rectal Colonization (VREC)

BackgroundSelect hospitalized patients are actively screened for VREC but VRE isolates may not undergo antibiotic susceptibility testing. We sought to identify predictors of daptomycin (DAP) nonsusceptibility (DNS, MIC > 4) and LNS (MIC > 2) among enteric VRE isolates recovered from patients actively screened for VREC for which antibiotic susceptibility testing was not preformed.MethodsThis was a retrospective study of consecutive adults admitted to a surgical intensive care unit (ICU) or associated medical unit between June 1, 2017 and March 1, 2018 who had a VRE isolate from active screening. Only index isolates were included. DAP and LZD MICs were determined by Etest. Patient- and antimicrobial-level data, including ambulatory prescriptions, dating back to January 1, 2016 were collected. Multivariable logistic regression models were used to determine predictors of DNS and LNS VRE.ResultsIn total, 64 patients’ VRE rectal isolates were included. Fifty-nine (92.2%) were E. faecium and 50 (78.1%) were from ICU patients. Thirty-seven patients (57.8%) were female and the mean age ± SD was 60 ± 13 years. Five (7.8%) and 20 (31.3%) patients had previous abdominal transplant and VRE infection, respectively. DAP and LZD MIC distributions are shown in the table below. Forty-one (64.1%) VRE isolates were LNS, including five LZD-resistant isolates. Only one (1.6%) isolate was DNS precluding an analysis of DNS predictors; 12 (18.8%) isolates had a DAP MIC > 2 mg/L. Common antimicrobial exposures prior to index VRE isolate included: vancomycin (62.5%), ceftriaxone (64.1%), cefepime (53.1%), metronidazole (50%), and ciprofloxacin (50%). Previous LZD (17.2%) and DAP (15.6%) exposure were less common. In a multivariable model, number of previous cefazolin doses (adjusted odds ratio (aOR) 0.74 95% confidence interval (CI) 0.55–0.95), and previous tobramycin exposure (aOR 0.15, 95% CI 0.02–0.81) were inversely associated with LNS. Previous LZD exposure was not associated with LNS.MIC50MIC90DAP1.5 mg/L3 mg/LLZD4 mg/L4 mg/LConclusionLNS was common amongst VRE isolates in this cohort. Previous LZD exposure was infrequent and not associated with LNS. LZD susceptibility testing among VRE isolates recovered from patients actively screened for VREC warrants clinical consideration.Disclosures All authors: No reported disclosures.

Adaptations of Vancomycin-Intermediate Sequence Type 72 Methicillin-Resistant Staphylococcus aureus for Daptomycin Nonsusceptibility.

In Korea, the major clonal type of community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA) is sequence type 72 (ST72) with staphylococcal cassette chromosome mec (SCCmec) type IV (ST72-MRSA-IV). In this study, we used a previously well-characterized isogenic pair of ST72 vancomycin (VAN) susceptible-and VAN intermediate-MRSA strains (VSSA303 and VISA072) and several VSSA strains complemented with plasmids expressing single-point mutated genes (dprAG196C, femAF92C, vraRE127K, and vraSRE127K) identified in the VISA strain. Using the strain set, we assessed the (1) susceptibilities to daptomycin (DAP) and cationic antimicrobial peptides (CAMPs), (2) alterations in cell envelope phenotypes, such as cell wall autolysis, surface positive charge, and membrane potential (ΔΨ), (3) transcriptional expression profiles of genes involved in surface charge regulation and changes of ΔΨ, and (4) cytokine stimulation profiles in murine macrophages. The vraRE127K mutation could enhance surface positive charge through mprF- and dltABCD-independent mechanisms with thickened cell wall. However, none of the single-point mutated genes increased DAP resistance. The DAP nonsusceptible (DAP-NS) phenotype observed in VISA072 strain likely resulted from the combined effects of low ΔΨ and increased positive surface charge. These results suggest that physicochemical alterations in cell envelope are involved in the survival response of DAP-NS VISA072 in sites of infections.

Characterization of genetic changes associated with daptomycin nonsusceptibility in Staphylococcus aureus.

The extensive use of daptomycin (DAP) for treatment of methicillin-resistant Staphylococcus aureus (MRSA) infections in the last decade has led to the emergence of DAP non-susceptible (DNS) Staphylococcus aureus strains. A better understanding of the molecular changes underlying DAP-non-susceptibility is required for early diagnosis and intervention with alternate combination therapies. The phenotypic changes associated with DNS strains have been well established. However, the genotypic changes—especially the kinetics of expression of the genes responsible for DAP-non-susceptibility are not well understood. In this study, we used three clinically derived isogenic pairs of DAP-susceptible (DAP-S) and DNS S. aureus strains to study gene expression profiles with the objective of identifying the potential genotypic changes associated with DAP-nonsusceptibility. We determined the expression profiles of genes involved in cell membrane (CM) charge, autolysis, cell wall (CW) synthesis, and penicillin binding proteins in DAP-S and DNS isogenic pairs. Our results demonstrate characteristic expression profiles for mprF, dltABCD, vraS, femB, and pbp2a genes, which are common to all the DNS S. aureus strains tested. Whole genome sequencing of DAP-S and DNS clinical isolates of S. aureus showed non-synonymous mutations in all DNS strains in genes involved in CM charge, CM composition, CW thickness and CW composition. To conclude, this study unravels some of the complex molecular changes involved in the development of DAP-nonsusceptibility by demonstrating distinct differences in gene expression profiles and mutations in the DNS S. aureus strains. This knowledge will aid in rapid identification of DNS S. aureus in clinical settings.

The Combination of Daptomycin and Fosfomycin Has Synergistic, Potent, and Rapid Bactericidal Activity against Methicillin-Resistant Staphylococcus aureus in a Rabbit Model of Experimental Endocarditis.

We investigated whether the addition of fosfomycin or cloxacillin to daptomycin provides better outcomes in the treatment of methicillin-resistant Staphylococcus aureus (MRSA) experimental aortic endocarditis in rabbits. Five MRSA strains were used to perform in vitro time-kill studies using standard (106) and high (108) inocula. Combined therapy was compared to daptomycin monotherapy treatment in the MRSA experimental endocarditis model. A human-like pharmacokinetics model was applied, and the equivalents of cloxacillin at 2 g/4 h, fosfomycin at 2 g/6 h, and daptomycin at 6 to 10 mg/kg/day were administered intravenously. A combination of daptomycin and either fosfomycin or cloxacillin was synergistic in the five strains tested at both inocula. A bactericidal effect was detected in four of five strains tested with both combinations. The MRSA-277 strain (vancomycin MIC, 2 μg/ml) was used for the experimental endocarditis model. Daptomycin plus fosfomycin significantly improved the efficacy of daptomycin monotherapy at 6 mg/kg/day in terms of both the proportion of sterile vegetations (100% versus 72%, P = 0.046) and the decrease in the density of bacteria within the vegetations (P = 0.025). Daptomycin plus fosfomycin was as effective as daptomycin monotherapy at 10 mg/kg/day (100% versus 93%, P = 1.00) and had activity similar to that of daptomycin plus cloxacillin when daptomycin was administered at 6 mg/kg/day (100% versus 88%, P = 0.48). Daptomycin nonsusceptibility was not detected in any of the isolates recovered from vegetations. In conclusion, for the treatment of MRSA experimental endocarditis, the combination of daptomycin plus fosfomycin showed synergistic and bactericidal activity.

Daptomycin selects for genetic and phenotypic adaptations leading to antibiotic tolerance in MRSA.

Daptomycin non-susceptibility in Staphylococcus aureus can emerge via the accumulation of single or multiple mutations, each resulting in a slight increase in the daptomycin MIC. The daptomycin-non-susceptible phenotype may include other features such as daptomycin tolerance. This study identifies S. aureus genomic regions that frequently develop mutations following prolonged daptomycin exposure but have not been previously associated with daptomycin non-susceptibility. Sequence variations in the same eight loci independently observed following 28 day parallel serial passages of S. aureus J01 in daptomycin were introduced in isolation into S. aureus J01. MICs were determined by microbroth dilution. Daptomycin killing and tolerance were determined by kill curve analysis. Single mutations in snoF, hmp1, sspA, rimP, hepT, rsh, map1 and amaP had only a modest impact on the daptomycin MIC (≤2-fold). In contrast, individual mutation in several of these regions resulted in pronounced changes to daptomycin tolerance. This study demonstrates that less characterized mutations in S. aureus following daptomycin exposure do not result in significant daptomycin susceptibility changes, but rather allow for enhanced survival characteristics during treatment. This sheds new light on genetic adaptations that may play a role in persistent infection. Further studies are needed to elucidate the prevalence of these mutations in clinical isolates.