Development and validation of LC-MS/MS for quantifying omadacycline from stool for gut microbiome studies.

This study developed and validated a new liquid chromatography-tandem mass spectrometry (LC-MS/MS) method to quantify omadacycline and its epimerization in stool to facilitate microbiome studies. Omadacycline was extracted in a methanol-water-ethylenediaminetetraacetic acid (ETDA) solvent containing deuterated omadacycline as internal standard, followed by dilution. In an optimal gradient elution mode, omadacycline and its C4 epimer were separated within 5 min on reversed-phase C18 column. The method showed a broad working range of 0.1-200 ng/ml with a limitation of detection (LOD) of 0.03 ng/ml, little fecal matrix effect, good intra-day and inter-day accuracy (90-101 %), precision (2-15 %), and recovery rate (99-105 %). The method was sufficiently sensitive to quantify omadacycline in human fecal samples (n = 82) collected during a 10-day therapy course and at follow-up (day 13 and day 30) that ranged from 1 to 4785 µg/g. Further analysis revealed that ∼9 % of omadacycline was epimerized in fecal matrix control while, on average, 37.4 % was epimerized in human fecal samples. This study developed and validated a novel, simple, sensitive, and accurate method utilizing LC-MS/MS to quantify omadacycline its epimerization in the human gut. This has important implications for future studies of omadacycline and other tetracycline-class antibiotics as part of gut microbiome studies.

Antibacterial activity of ibezapolstat against antimicrobial-resistant clinical strains of Clostridioides difficile.

Antimicrobial resistance is emerging in clinical strains of Clostridioides difficile. Ibezapolstat (IBZ) is a DNA polymerase IIIC inhibitor that has completed phase II clinical trials. IBZ has potent in vitro activity against wild-type, susceptible strains but its effect on C. difficile strains with reduced susceptibility to metronidazole (MTZ), vancomycin (VAN), or fidaxomicin (FDX) has not been tested. The primary objective of this study was to test the antibacterial properties of IBZ against multidrug-resistant C. difficile strains. The in vitro activity, bactericidal, and time-kill activity of IBZ versus comparators were evaluated against 100 clinical strains of which 59 had reduced susceptibility to other C. difficile antibiotics. Morphologic changes against a multidrug resistance strain were visualized by light and scanning electron microscopy. The overall IBZ MIC50/90 values (µg/mL) for evaluated C. difficile strains were 4/8, compared with 2/4 for VAN, 0.5/1 for FDX, and 0.25/4 for MTZ. IBZ MIC50/90 values did not differ based on non-susceptibility to antibiotic class or number of classes to which strains were non-susceptible. IBZ bactericidal activity was similar to the minimum inhibitory concentration (MIC) and maintained in wild-type and non-susceptible strains. Time-kill assays against two laboratory wild-type and two clinical non-susceptible strains demonstrated sustained IBZ activity despite reduced killing by comparator antibiotics for IBZ and VAN non-susceptible strains. Microscopy visualized increased cell lengthening and cellular damage in multidrug-resistant strains exposed to IBZ sub-MIC concentrations. This study demonstrated the potent antibacterial activity of IBZ against a large collection of C. difficile strains including multidrug-resistant strains. This study highlights the therapeutic potential of IBZ against multidrug-resistant strains of C. difficile.

Fecal Pharmacokinetics and Gut Microbiome Effects of Oral Omadacycline Versus Vancomycin in Healthy Volunteers.

BACKGROUND: Clostridioides difficile infection (CDI) is a common healthcare-associated infection with limited treatment options. Omadacycline, an aminomethylcycline tetracycline, has potent in vitro activity against C difficile and a low propensity to cause CDI in clinical trials. We aimed to assess fecal pharmacokinetics and gut microbiome effects of oral omadacycline compared to oral vancomycin in healthy adults. METHODS: This was a phase 1, nonblinded, randomized clinical trial conducted in healthy volunteers aged 18-40 years. Subjects received a 10-day course of omadacycline or vancomycin. Stool samples were collected at baseline, daily during therapy, and at follow-up visits. Omadacycline and vancomycin stool concentrations were assessed, and microbiome changes were compared. RESULTS: Sixteen healthy volunteers with a mean age of 26 (standard deviation [SD], 5) years were enrolled; 62.5% were male, and participants' mean body mass index was 23.5 (SD, 4.0) kg/m2. Omadacycline was well tolerated with no safety signal differences between the 2 antibiotics. A rapid initial increase in fecal concentrations of omadacycline was observed compared to vancomycin, with maximum concentrations achieved within 48 hours. A significant difference in alpha diversity was observed following therapy in both the omadacycline and vancomycin groups (P < .05). Bacterial abundance and beta diversity analysis showed differing microbiome changes in subjects who received omadacycline versus vancomycin. CONCLUSIONS: Subjects given omadacycline had high fecal concentrations with a distinct microbiome profile compared to vancomycin. CLINICAL TRIALS REGISTRATION: NCT06030219.

Assessment of Quality of Life Among Patients With Recurrent Clostridioides difficile Infection Treated with Investigational Oral Microbiome Therapeutic SER-109: Secondary Analysis of a Randomized Clinical Trial.

Importance: Recurrent Clostridioides difficile infection (CDI) is a debilitating disease leading to poor health-related quality of life (HRQOL), loss of productivity, anxiety, and depression. The potential association of treatment with HRQOL has not been well evaluated. Objectives: To explore the association of SER-109 compared with placebo on HRQOL in patients with recurrent CDI up to week 8. Design, Setting, and Participants: This study was a secondary analysis of a randomized, double-blind, placebo-controlled trial that took place at 56 sites in the US and Canada from July 2017 to April 2020 and included 182 patients randomized to SER-109 or placebo groups. Interventions: SER-109 or placebo (4 capsules once daily for 3 days) following antibiotics for CDI. Main Outcomes and Measures: Exploratory analysis of HRQOL using the disease specific Clostridioides difficile Quality of Life Survey (Cdiff32) assessed at baseline, week 1, and week 8. Results: In this study, 182 patients (109 [59.9%] female; mean age, 65.5 [16.5] years) were randomized to SER-109 (89 [48.9%]) or placebo (93 [51.1%]) groups and were included in the primary and exploratory analyses. Baseline Cdiff32 scores were similar between patients in the SER-109 and placebo groups (52.0 [18.3] vs 52.8 [18.7], respectively). The proportion of patients with overall improvement from baseline in the Cdiff32 total score was higher in the SER-109 arm than placebo at week 1 (49.4% vs 26.9%; P = .012) and week 8 (66.3% vs 48.4%; P = .001).Greater improvements in total and physical domain and subdomain scores were observed in patients in the SER-109 group compared with placebo as early as week 1, with continued improvements observed at week 8. Among patients in the placebo group, improvements in HRQOL were primarily observed in patients with nonrecurrent CDI while patients in the SER-109 group reported improvements in HRQOL, regardless of clinical outcome. Conclusions and Relevance: In this secondary analysis of a phase 3 clinical trial, SER-109, an investigational microbiome therapeutic was associated with rapid and steady improvement in HRQOL compared with placebo through 8 weeks, an important patient-reported outcome. Trial Registration: ClinicalTrials.gov Identifier: NCT03183128.

Significant Publications on Infectious Diseases Pharmacotherapy in 2021.

Purpose: To summarize the most noteworthy infectious diseases (ID) pharmacotherapy articles published in peer-reviewed literature in 2021. Summary: Members of the Houston Infectious Diseases Network (HIDN) nominated articles that were deemed to have significant contributions to ID pharmacotherapy in 2021. These nominations included articles pertaining to both general ID, including coronavirus disease 2019 (COVID-19), and human immunodeficiency virus/acquired immunodeficiency syndrome (HIV/AIDS) pharmacotherapy. A total of 35 articles were nominated by HIDN: 30 articles pertaining to general ID pharmacotherapy and 5 articles with HIV/AIDS focus. To select the most influential articles of 2021, a survey was created and distributed to members of the Society of Infectious Diseases Pharmacists (SIDP). Of the 239 SIDP members who responded to the survey, there were 192 recorded votes for the top 10 general ID pharmacotherapy articles and 47 recorded votes for the top HIV/AIDS article, respectively. The top publications are summarized. Conclusion: Antimicrobial stewardship and the optimal management of infectious disease states continues to be a priority in the midst of the ongoing coronavirus disease 2019 (COVID-19) global pandemic. In light of the sheer volume of ID-related articles published in the past year, this review aims to aid clinicians in remaining up-to-date on key practice-changing ID pharmacotherapy publications from 2021.

Development of the invasive candidiasis discharge [I Can discharge] model: a mixed methods analysis.

Patients with invasive candidiasis (IC) have complex medical and infectious disease problems that often require continued care after discharge. This study aimed to assess echinocandin use at hospital discharge and develop a transition of care (TOC) model to facilitate discharge for patients with IC. This was a mixed method study design that used epidemiologic assessment to better understand echinocandin use at hospital discharge TOC. Using grounded theory methodology focused on patients given echinocandins during their last day of hospitalization, a TOC model for patients with IC, the invasive candidiasis [I Can] discharge model was developed to better understand discharge barriers. A total of 33% (1405/4211) echinocandin courses were continued until the last day of hospitalization. Of 536 patients chosen for in-depth review, 220 (41%) were discharged home, 109 (20%) were transferred, and 207 (39%) died prior to discharge. Almost half (46%, 151/329) of patients discharged alive received outpatient echinocandin therapy. Independent predictors for outpatient echinocandin use were osteomyelitis (OR, 4.1; 95% CI, 1.1-15.7; p = 0.04), other deep-seated infection (OR, 4.4; 95% CI, 1.7-12.0; p = 0.003), and non-home discharge location (OR, 3.9, 95% CI, 2.0-7.7; p < 0.001). The I Can discharge model was developed encompassing four distinct themes which was used to identify potential barriers to discharge. Significant echinocadin use occurs at hospital discharge TOC. The I Can discharge model may help clinical, policy, and research decision-making processes to facilitate smoother and earlier hospital discharges.

Recurrent Clostridioides difficile infection worsens anxiety-related patient-reported quality of life.

BACKGROUND: Clostridioides difficile infection (CDI) is associated with high recurrence rates impacting health-related quality of life (HrQOL). However, patient-reported data are lacking particularly in the outpatient setting. We assessed changes in HrQOL over time in patients treated with bezlotoxumab at US infusion centers and determined clinical factors associated with HrQOL changes. METHODS: The HrQOL survey was conducted in adult patients with CDI, who received bezlotoxumab in 25 US outpatient infusion centers. The survey was adapted from the Cdiff32 instrument to assess anxiety-related changes to HrQOL and completed on the day of infusion (baseline) and at 90 days post bezlotoxumab (follow-up). Demographics, disease history, CDI risk factors, and recurrence of CDI (rCDI) at 90-day follow-up were collected. Changes in HrQOL scores were calculated and outcomes assessed using a multivariable linear regression model with P < 0.05 defined as statistically significant. RESULTS: A total of 144 patients (mean age: 68 ± 15 years, 63% female, median Charlson index: 4, 15.9% rCDI) were included. The overall mean baseline and follow-up HrQOL scores were 26.4 ± 11.5 and 56.4 ± 25.0, respectively. At follow-up, this score was significantly higher for patients who had primary CDI (34.5 ± 21.7) compared to those with multiple rCDI (24.7 ± 21.0; P = 0.039). The mean HrQOL change at follow-up was significantly higher for patients without rCDI (34.1 ± 28.8 increase) compared to patients with rCDI (6.7 ± 19.5 increase; P < 0.001), indicating improvement in anxiety. CONCLUSIONS: Using the Cdiff32 instrument, we demonstrated that HrQOL worsened significantly in patients with further rCDI. These findings support the use of Cdiff32 in assessing CDI-related humanistic outcomes.

Multi-country surveillance of Clostridioides difficile demonstrates high prevalence of spores in non-healthcare environmental settings.

BACKGROUND: C. difficile spores are frequently isolated from hospital and non-healthcare settings but a worldwide analysis has not been done. The study objectives were to assess C. difficile spore contamination in the hospital and non-healthcare environments across a variety of countries. METHODS: Field studies assessed hospital vs. non-healthcare C. difficile spore contamination in hospitals, non-healthcare buildings, outdoor environments, and shoes. Swabs were cultured anaerobically for C. difficile and typed using PCR-fluorescent ribotyping. C. difficile contamination by swabbing area and geographic locations were compared. FINDINGS: A total of 7,857 unique samples were collected primarily from the USA (89%) in addition to 9 other countries. The global prevalence of C difficile from environmental samples was 25.3% and did not differ between countries. In USA based studies, C. difficile contamination rates were similar for healthcare buildings (23.2%), non-healthcare buildings (23.4%), and outdoor spaces (24.7%). Floor samples had significantly higher (p < 0.001) C. difficile contamination rate (46.5%) followed by non-floor samples (21.1%), and bathrooms (15.3%). In a comparison of USA to other country samples, C. difficile contamination rates were similar for USA samples (21.5%) compared to rest of world samples (22.3%; p = 0.61). The most common ribotypes included F014-020 (15.7%), F106 (12.6%), F010 (8.9%), F027 (8.8%), and F002 (8.1%) and did not differ significantly between USA and non-USA samples. Finally, 546 of 1,218 (44.8%) shoe soles swabbed from the USA were contaminated with C. difficile spores. INTERPRETATION: This large surveillance study of several countries demonstrated high prevalence of toxigenic C. difficile in non-healthcare environments with high contamination rates from floors and shoe soles.

Efficacy, Safety, Pharmacokinetics, and Microbiome Changes of Ibezapolstat in Adults with Clostridioides difficile Infection: A Phase 2a Multicenter Clinical Trial.

BACKGROUND: This study was the first human validation of the gram-positive bacterial DNA polymerase IIIC target in patients with Clostridioides difficile infection. The primary objectives were to assess clinical cure rates and adverse events (AEs). Secondary objectives were to evaluate plasma/fecal pharmacokinetics, microbiologic eradication, microbiome and bile acid effects, and sustained clinical cure (SCC) with ibezapolstat. METHODS: This single-arm, open-label, phase 2a study enrolled adults with C. difficile infection at 4 US centers. Patients received ibezapolstat 450 mg orally every 12 hours for 10 days and followed for an additional 28 days to assess study objectives. RESULTS: Ten patients with a mean (standard deviation [SD]) age of 49 [15] years were enrolled. Seven AEs were reported classified as mild-moderate. Plasma levels of ibezapolstat ranged from 233 to 578 ng/mL while mean (SD) fecal levels were 416 (494) µg/g stool by treatment day 3 and >1000 µg/g stool by days 8-10. A rapid increase in alpha diversity in the fecal microbiome was noted after starting ibezapolstat therapy, which was maintained after completion of therapy. A proportional decrease in Bacteroidetes phylum was observed (mean change [SD], -10.0% [4.8%]; P = .04) with a concomitantly increased proportion of Firmicutes phylum (+14.7% [5.4%]; P = .009). Compared with baseline, total primary bile acids decreased by a mean (SD) of 40.1 (9.6) ng/mg stool during therapy (P < .001) and 40.5 (14.1) ng/mg stool after completion of therapy (P = .007). Rates of both initial clinical cure and SCC at 28 days were 100% (10 of 10 patients). CONCLUSIONS: In this phase 2a study, 10 of 10 patients achieved SCC, demonstrated favorable pharmacokinetics, minimal AEs, and beneficial microbiome and bile acids results. These results support continued clinical development.

Real-world, Multicenter Experience With Meropenem-Vaborbactam for Gram-Negative Bacterial Infections Including Carbapenem-Resistant Enterobacterales and Pseudomonas aeruginosa.

BACKGROUND: We aimed to describe the clinical characteristics and outcomes of patients treated with meropenem-vaborbactam (MEV) for a variety of gram-negative infections (GNIs), primarily including carbapenem-resistant Enterobacterales (CRE). METHODS: This is a real-world, multicenter, retrospective cohort within the United States between 2017 and 2020. Adult patients who received MEV for ≥72 hours were eligible for inclusion. The primary outcome was 30-day mortality. Classification and regression tree analysis (CART) was used to identify the time breakpoint (BP) that delineated the risk of negative clinical outcomes (NCOs) and was examined by multivariable logistic regression analysis (MLR). RESULTS: Overall, 126 patients were evaluated from 13 medical centers in 10 states. The most common infection sources were respiratory tract (38.1%) and intra-abdominal (19.0%) origin, while the most common isolated pathogens were CRE (78.6%). Thirty-day mortality and recurrence occurred in 18.3% and 11.9%, respectively. Adverse events occurred in 4 patients: nephrotoxicity (n = 2), hepatoxicity (n = 1), and rash (n = 1). CART-BP between early and delayed treatment was 48 hours (P = .04). MEV initiation within 48 hours was independently associated with reduced NCO following analysis by MLR (adusted odds ratio, 0.277; 95% CI, 0.081-0.941). CONCLUSIONS: Our results support current evidence establishing positive clinical and safety outcomes of MEV in GNIs, including CRE. We suggest that delaying appropriate therapy for CRE significantly increases the risk of NCOs.

H2O2 biosensor consisted of hemoglobin-DNA conjugate on nanoporous gold thin film electrode with electrochemical signal enhancement.

In this research, we developed electrochemical biosensor which was composed of hemoglobin (Hb)-DNA conjugate on nanoporous gold thin film (NPGF) for hydrogen peroxide (H2O2) detection. For the first time, Hb and DNA was conjugated as a sensing platform for uniform orientation of Hb on electrode. The newly developed Hb-DNA conjugate was designed to prevent Hb from aggregation on electrode. DNA hybridization of Hb-DNA conjugate and complementary DNA (cDNA) on NPGF electrode induced uniformly assembled biosensor. Furthermore, NPGF electrode fabrication method was introduced to the increment of the surface area. To confirm the conjugation of Hb-DNA conjugate, sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and ultraviolet-visible spectroscopy (UV-VIS) were used. Formation of the NPGF electrode was verified by scanning electron microscope (SEM). Atomic force microscopy (AFM) was operated for the confirmation of Hb-DNA immobilization on electrode. The electrochemical property of fabricated electrode was investigated by cyclic voltammetry (CV). Also, H2O2 sensing performance of fabricated electrode was investigated by amperometric i-t curve technique. This sensor showed a wide linear range from 0.00025 to 5.00 mM and a correlation coefficient of R2 = 0.9986. The detection limit was 250 nM. Proposed biosensor can be utilized as a sensing platform for development of biosensor.

Electrochemical Biosensor Composed of Silver Ion-Mediated dsDNA on Au-Encapsulated Bi2 Se3 Nanoparticles for the Detection of H2 O2 Released from Breast Cancer Cells.

A newly developed electrochemical biosensor composed of a topological insulator (TI) and metallic DNA (mDNA) is fabricated. The bismuth selenide nanoparticle (Bi2 Se3 NP) is synthesized and sandwiched between the gold electrode and another Au-deposited thin layer (Bi2 Se3 @Au). Then, eight-silver-ion mediated double-stranded DNA (mDNA) is immobilized onto the substrate (Bi2 Se3 @Au-mDNA) for the further detection of hydrogen peroxide. The Bi2 Se3 NP acts as the electrochemical-signal booster, while unprecedentedly its encapsulation by the Au thin layer keeps the TI surface states protected, improves its electrochemical-signal stability and provides an excellent platform for the subsequent covalent immobilization of the mDNA through Au-thiol interaction. Electrochemical results show that the fabricated biosensor represents much higher Ag+ redox current (≈10 times) than those electrodes prepared without Bi2 Se3 @Au. The characterization of the Bi2 Se3 @Au-mDNA film is confirmed by atomic force microscopy, scanning tunneling microscopy, and cyclic voltammetry. The proposed biosensor shows a dynamic range of 00.10 × 10-6 m to 27.30 × 10-6 m, very low detection limit (10 × 10-9 m), unique current response (1.6 s), sound H2 O2 recovery in serum, and substantial capability to classify two breast cancer subtypes (MCF-7 and MDA-MB-231) based on their difference in the H2 O2 generation, offering potential applications in the biomedicine and pharmacology fields.

Silver Nanoparticle Modified Electrode Covered by Graphene Oxide for the Enhanced Electrochemical Detection of Dopamine.

Several neurological disorders such as Alzheimer's disease and Parkinson's disease have become a serious impediment to aging people nowadays. One of the efficient methods used to monitor these neurological disorders is the detection of neurotransmitters such as dopamine. Metal materials, such as gold and platinum, are widely used in this electrochemical detection method; however, low sensitivity and linearity at low dopamine concentrations limit the use of these materials. To overcome these limitations, a silver nanoparticle (SNP) modified electrode covered by graphene oxide for the detection of dopamine was newly developed in this study. For the first time, the surface of an indium tin oxide (ITO) electrode was modified using SNPs and graphene oxide sequentially through the electrochemical deposition method. The developed biosensor provided electrochemical signal enhancement at low dopamine concentrations in comparison with previous biosensors. Therefore, our newly developed SNP modified electrode covered by graphene oxide can be used to monitor neurological diseases through electrochemical signal enhancement at low dopamine concentrations.

Electrochemical H2O2 biosensor composed of myoglobin on MoS2 nanoparticle-graphene oxide hybrid structure.

In this research, the electrochemical biosensor composed of myoglobin (Mb) on molybdenum disulfide nanoparticles (MoS2 NP) encapsulated with graphene oxide (GO) was fabricated for the detection of hydrogen peroxide (H2O2). Hybrid structure composed of MoS2 NP and GO (GO@MoS2) was fabricated for the first time to enhance the electrochemical signal of the biosensor. As a sensing material, Mb was introduced to fabricate the biosensor for H2O2 detection. Formation and immobilization of GO@MoS2 was confirmed by transmission electron microscopy, ultraviolet-visible spectroscopy, scanning electron microscopy, and scanning tunneling microscopy. Immobilization of Mb, and electrochemical property of biosensor were investigated by cyclic voltammetry and amperometric i-t measurements. Fabricated biosensor showed the electrochemical signal enhanced redox current as -1.86µA at an oxidation potential and 1.95µA at a reduction potential that were enhanced relative to those of electrode prepared without GO@MoS2. Also, this biosensor showed the reproducibility of electrochemical signal, and retained the property until 9 days from fabrication. Upon addition of H2O2, the biosensor showed enhanced amperometric response current with selectivity relative to that of the biosensor prepared without GO@MoS2. This novel hybrid material-based biosensor can suggest a milestone in the development of a highly sensitive detecting platform for biosensor fabrication with highly sensitive detection of target molecules other than H2O2.