Staphylococcus lugdunensis Bacteremia with an Infected Aortic Thrombus in a Preterm Infant

Staphylococcus lugdunensis is a rare cause of late-onset sepsis in preterm infants. To our best knowledge, we report the fourth case of a male preterm infant who developed fulminant late-onset sepsis due to Staphylococcus lugdunensis with persistent bacteremia secondary to an infected aortic thrombus confirmed with two positive blood cultures. Our patient was an extremely low birth weight growth-restricted infant born at 27 weeks gestation and initially required an umbilical arterial catheter for blood pressure and blood gas monitoring. The course of this neonate was complicated by severe respiratory distress syndrome that evolved into chronic lung disease along with multiple episodes of tracheitis. Hemodynamically, the infant had a significant patent ductus arteriosus, and an episode of medical necrotizing enterocolitis followed by Staphylococcus lugdunensis septicemia. He was diagnosed with an infected aortic thrombus, probably the occult focus responsible for the persistent bacteremia. After a 6-week course of intravenous antibiotics and 4-week course of anticoagulant therapy, the infant responded and recovered without complications.

Follow-Up Blood Cultures in Young Infants With Bacteremic Urinary Tract Infections

BACKGROUND AND OBJECTIVES Researchers in previous studies suggest that the clinical yield of follow-up blood cultures (FUBCs) is low in infants with bacteremic urinary tract infection (UTI) because persistent bacteremia is rare; however, no researchers have analyzed the practice of routinely obtaining FUBCs. In our study, we evaluate outcomes in infants with FUBCs, examine opportunities for improvement of blood culture practices, and add important information to inform both clinical practice as well as further study. DESIGN This retrospective cohort study included infants <90 days of age with bacteremia and UTI with the same pathogen at 22 hospitals that make up Intermountain Healthcare between 2002 to 2020. Infants with culture proven meningitis, osteomyelitis, central line infection, and infections occurring during NICU hospitalization were excluded. RESULTS Total number of patients with bacteremic UTI was 174, 153 (88%) patients had at least 1 FUBC, 14 of 153 (9%) had a positive FUBC with same organism, and 4 of 153 (3%) were contaminants. The length of stay was longer for patients with positive FUBCs. Patients with Escherichia coli are more likely to have a negative FUBC. Readmissions within 30 days were similar among infants with positive FUBCs, negative FUBCs, and no FUBCs. CONCLUSIONS FUBCs in infants with bacteremic UTI should not be routinely performed, especially for E coli, and it is unclear whether FUBCs improve outcomes.

128. Mutations that Inactivate the Tricarboxylic Acid Cycle in Staphylococcus aureus Arise During Persistent MRSA Bacteremia

Background Persistent MRSA bacteremia is common with high morbidity and mortality despite appropriate antibiotics. Persistent infections are associated with antibiotic tolerance and can arise from perturbations in cellular pathways. We performed whole genome sequencing of clinical isolates to identify the genetic bases of antibiotic tolerance. Methods Whole genomes of MRSA from patients with persistent bacteremia were sequenced, which identified 8 isolates harboring different citZ mutations. To assess the effect of the mutations directly on citrate synthase activity, purified recombinant enzymes were assayed using a commercially available kit. The same kit was used to measure activity in whole cell lysates of MRSA isolates harboring wild-type and mutant citZ alleles. Enzyme kinetic parameters were determined by fitting initial rate data to the Michaelis-Menten equation using nonlinear regression. Figure 1. Whole Genome Sequencing of Persistent MRSA Bacteremia Whole genome sequencing of 206 blood cultures from 20 patients with persistent MRSA bacteremia (defined as >7 days) under the hypothesis that continuous antibiotic exposure will give rise to - and enrich for - mutations that convey antibiotic tolerance through in-host evolution. Results Analysis of whole genomes from 206 blood cultures from 20 patients with persistent MRSA bacteremia identified citZ, which encodes for citrate synthase, the first step of the tricarboxylic acid cycle, as the most repeatedly mutated gene. The data revealed parallelism in its evolution, as citZ was mutated 8 independent times, a rate far greater than from chance alone. To characterize the impact of the mutations on enzyme activity, recombinant proteins were expressed in E. coli and purified for enzymatic assays. As compared to wildtype enzyme, two mutants had reduced activity, and four mutants had near-absent activity; two mutants were unable to be expressed due to destabilizing mutations. Michaelis-Menten analysis of the two mutants with residual activity show that, as compared to wildtype, both had lower affinity for its substrates, and lower maximum activity. Furthermore, we found the cell was unable to compensate for the loss of citrate synthase activity, as lysates harboring these mutations also displayed analogous reductions in activity. Figure 2. Purified citrate synthase mutant proteins have reduced enzyme activity Citrate synthase catalyzes the reaction between acetyl-CoA and oxaloacetic acid to form citric acid with CoA-SH as a byproduct. CoA-SH can interact with DTNB to form TNB, which can be measured spectrophotometrically at A412nm to indirectly measure the activity of citrate synthase. Wildtype citrate synthase and the D141N mutant have comparable activity whereas the remaining study mutants have either reduced (mutants A313P and A313V) or near-absent (mutants G7D, G7D + D141N, S201P, P354S) activity. The three active site mutants (H248G, D309G, H219G) also have near-absent activity and serve as negative controls. Positive control is citrate synthase provided in the Sigma Citrate Synthase Assay Kit. No enzyme control replaces enzyme with assay buffer. All error bars are ± 1 SD, calculated from at least three replicate experiments. Figure 3. A313P and A313V mutants have decreased substrate affinity and decreased maximal activity Panels A and B show plots of the initial rate of citrate synthase activity (Y-axis, in units of µmole/mL/sec) versus OAA/AcCoA substrate concentration (X-axis, in millimolar units), for wildtype citrate synthase and the A313P and A313V mutants. Panel (A) shows enzymatic parameters for wildtype, A313P mutant, and A313V mutant with variable OAA (0-0.625 mM) and fixed AcCoA (0.3 mM). Panel (B) shows enzymatic parameters for wildtype, A313P mutant, and A313V mutant with fixed OAA (0.5 mM) and variable AcCoA (0-2.5 mM). Three replicates were performed for each condition, with error bars showing ± 1 SD. Figure 4. Citrate synthase mutants disrupt functional dimerization and destabilize alpha-helix packing Identified citrate synthase mutants mapped onto the crystal structure of T. thermophilus citrate synthase (PDB 1IOM) show that the mutations are located either at dimerization interfaces or within the interior of hydrophobic packing interfaces. Conclusion Cellular metabolism and virulence regulation are interconnected in S. aureus, as alterations in TCA cycle activity lead to increased persister formation and host macrophage inactivation. Our findings that inactivating citZ mutations are enriched can provide a potential explanation for the mechanism of persistent bacteremia. Disclosures All Authors: No reported disclosures

89. Follow-Up Blood Cultures (FUBC) in the Management of Gram-Negative Bacilli (GNB) Bloodstream Infections (BSIs): Frequently Obtained and Rarely Helpful

Background While GNB BSIs remain a major cause of morbidity and mortality, no clear guidelines exist on the utilization of FUBCs to guide management. Despite the recognition of persistent bacteremia as a risk factor for increased mortality, early studies suggested FUBCs were low yield in this setting, and thus had low utility. More recently, some controversy has arisen with multivariate analyses suggesting FUBC acquisition may be associated with lower mortality. We sought to characterize the utilization and yield of FUBCs for GNB BSIs at our institution. Methods We performed a retrospective review of 514 episodes of consecutive blood cultures from unique adult inpatients with GNB BSI between July 2017-July 2019. Exclusion criteria included prior positive culture, polymicrobial Gram stain, or discharge, death, or comfort measures only within 24 hours of Gram stain. FUBCs were defined as blood cultures collected between 24 hours to 7 days after the index blood culture. Baseline clinical and microbiologic characteristics were compared between groups, as well as clinical outcomes. Results Of 514 episodes, 338 (66%) had FUBCs performed, with a median of 2 FUBCs/episode. The majority of FUBCs (322/338; 95%) were negative, with 9 (3%) yielding the same organism and 9 (3%) yielding a different organism. Most initial FUBCs were obtained prior to index antimicrobial susceptibility results (227/338; 67%). Patients with FUBCs performed had a higher median Pitt bacteremia score (2 vs 1; p = 0.015) and were more likely to have hospital onset (36% vs 22%; p = 0.002), severe neutropenia (16% vs 4%; p < 0.001) and a catheter-associated source (13% vs 4%; p = 0.001). 30-day mortality did not differ between patients with or without FUBCs (10% vs 11%; p = 0.84). Conclusion FUBCs were frequently obtained, but were of low yield even in comparison to recent similar studies. Though FUBCs were performed in more severe cases, a difference in mortality was not observed. Delaying the decision of whether to obtain FUBCs until after index antimicrobial susceptibility results are available would reduce unnecessary testing in most cases. Further study could better define where FUBCs after antimicrobial susceptibility testing would be most helpful. Disclosures All Authors: No reported disclosures

Changing characteristics of S. aureus bacteremia caused by PVL-negative, MRSA strain over 11 years

Community-acquired methicillin-resistant Staphylococcus aureus (MRSA) has emerged as an important cause of infection. We conducted a longitudinal study to evaluate changes in clinical and microbiological characteristics as well as outcomes of sequence type (ST) 72 MRSA bacteremia. We reviewed adult patients enrolled in a prospective cohort with ST72 MRSA bacteremia from August 2008 to December 2018 at Asan Medical Center, Seoul, South Korea. Changes in clinical characteristics, outcomes, and microbiological characteristics of patients over time were evaluated. Generalized linear and linear regression models were used to evaluate changes. Of the 1,760 isolates, 915 (62%) were MRSA bacteremia and 292 (31.9%) were ST72 MRSA. During the study period, the relative risk (RR) of MRSA bacteremia decreased annually by 3.7%; however, among MRSA bacteremia, RR of ST72 MRSA increased annually by 8.5%. Vancomycin minimum inhibitory concentration (MIC) decreased over the study period. Metastatic infection, persistent bacteremia, and recurrence of bacteremia within 12 weeks decreased significantly. There were no significant changes in 30-d and 12-week mortality. Antibiotic susceptibility of ST72 MRSA was evaluated, and the resistance rate to erythromycin decreased significantly. ST72 MRSA incidence increased annually; its vancomycin MIC and erythromycin resistance rate decreased over the 11 years.

Synergistic Meropenem/Vaborbactam Plus Fosfomycin Treatment of KPC Producing K. pneumoniae Septic Thrombosis Unresponsive to Ceftazidime/Avibactam: From the Bench to the Bedside

Gram-negative bacilli septic thrombosis (GNB-ST) represents a subtle and often misleading condition, potentially fatal if not recognized early and requiring prolonged antimicrobial therapy and anticoagulation. Herein, reported for the first time, is a very challenging case of Klebsiella producing carbapenemase (KPC)-producing K. pneumoniae (KPC-Kp) ST unresponsive to ceftazidime/avibactam (CZA) relapsed first with meropenem/vaborbactam (MVB) monotherapy and subsequently cured with MVB plus fosfomycin (FOS) combination. The present case highlights the possibility of CZA underexposure on the infected thrombus and the risk of in vivo emergence of CZA resistance in the setting of persistent bacteremia and sub-optimal anticoagulation. Pharmacokinetic analyses showed that both MVB and FOS were in the therapeutic range. In vitro studies demonstrated a high level of MVB + FOS synergism that possibly allowed definitive resolution of the endovascular infection.

Impact of initial vancomycin pharmacokinetic/pharmacodynamic parameters on the clinical and microbiological outcomes of methicillin-resistant Staphylococcus aureus bacteremia in children

Optimal vancomycin exposure is important to minimize treatment failure of methicillin-resistant Staphylococcus aureus (MRSA) bacteremia. We aimed to analyze the impact of initial vancomycin pharmacokinetic/pharmacodynamic (PK/PD) parameters, including the initial vancomycin C trough and the area under the curve (AUC)/minimal inhibitory concentration (MIC) on the outcomes of pediatric MRSA bacteremia. The study population consisted of hospitalized children aged between 2 months and 18 years with MRSA bacteremia, in whom C trough was measured at least one time within the time period of January 2010 to March 2018. Demographic profiles, underlying diseases, and clinical/microbiological outcomes were abstracted retrospectively. During the study period, 73 cases of MRSA bacteremia occurred in children with a median age of 12.4 months. Severe clinical outcomes leading to intensive care unit stay and/or use of mechanical ventilation occurred in 47.5% (35/73); all-cause 30-day mortality was 9.7% (7/72). The median dosage of vancomycin was 40.0 mg/kg/day. There was a weak linear relationship between C trough and the corresponding AUC/MIC (r = 0.235). ROC curves for achieving an AUC/MIC of 300 suggested that the initial C trough at 10 μg/mL could be used as a cut-off value with a sensitivity of 90.5% and a specificity of 44%. Although persistent bacteremia at 48–72 hours after vancomycin administration was observed more frequently when the initial C trough was < 10 μg/mL and initial AUC/MIC was < 300, initial AUC/MIC < 300 was the only risk factor associated with persistent bacteremia at 48–72 hours (adjusted OR 3.05; 95% CI, 1.07–8.68). Initial C trough and AUC/MIC were not associated with 30-day mortality. Although there was a weak relationship between C trough and AUC/MIC, initial AUC/MIC < 300 could be used as a predictor of persistent MRSA bacteremia at 48–72 hours. Further prospective data on optimal vancomycin dosing are necessary to improve clinical and microbiological outcomes in pediatric MRSA bacteremia.