Evaluation of treatment of methicillin-resistant Staphylococcus aureus biofilms with intermittent electrochemically generated H2O2 or HOCl.

Chronic wound infections can be difficult to treat and may lead to impaired healing and worsened patient outcomes. Novel treatment strategies are needed. This study evaluated the effects of intermittently produced hydrogen peroxide (H2O2) and hypochlorous acid (HOCl), generated via an electrochemical bandage (e-bandage), against methicillin-resistant Staphylococcus aureus biofilms in an agar membrane biofilm model. By changing the working electrode potential, the e-bandage generated either HOCl (1.5 VAg/AgCl) or H2O2 (-0.6 VAg/AgCl). The degree of biocidal activity of intermittent treatment with HOCl and H2O2 correlated with HOCl treatment time; HOCl treatment durations of 0, 1.5, 3, 4.5, and 6 hours (with the rest of the 6-hour total treatment time devoted to H2O2 generation) resulted in mean biofilm reductions of 1.36 ± 0.2, 2.22 ± 0.16, 3.46 ± 0.38, 4.63 ± 0.74, and 7.66 ± 0.5 log CFU/cm2, respectively, vs. non-polarized controls, respectively. However, application of H2O2 immediately after HOCl treatment was detrimental to biofilm removal. For example, 3 hours HOCl treatment followed by 3 hours H2O2 resulted in a 1.90 ± 0.84 log CFU/cm2 lower mean biofilm reduction than 3 hours HOCl treatment followed by 3 hours non-polarization. HOCl generated over 3 hours exhibited biocidal activity for at least 7.5 hours after e-bandage operation ceased; 3 hours of HOCl generation followed by 7.5 hours of non-polarization resulted in a biofilm cell reduction of 7.92 ± 0.12 log CFU/cm2 vs. non-polarized controls. Finally, intermittent treatment with HOCl (i.e., interspersed with periods of e-bandage non-polarization) for various intervals showed similar effects (approximately 6 log CFU/cm2 reduction vs. non-polarized control) to continuous treatment with HOCl for 3 hours, followed by 3 hours of non-polarization. These findings suggest that timing and sequencing of HOCl and H2O2 treatments are crucial for maximizing biofilm control when using an e-bandage strategy.

Phage K exposure generates phage-neutralizing activity in a rabbit model-humoral receptiveness may impact efficacy of phage therapy.

Phage therapy has not been established in the clinical routine, in part due to uncertainties concerning efficacy and immunogenicity. Here, three rabbits were immunized against staphylococcal phage K to assess viral potency in the presence of immunized serum. Three rabbits received weekly intramuscular injections of ~1010±1 pfu/mL phage K. Phage K-specific IgG formation was measured by an enzyme-linked immunosorbent assay (ELISA); phage inactivation was assessed by calculating K-rates. Using transmission electron microscopy (TEM) and immunogold labeling, antibody binding to phage K was visualized. This was numerically assessed by objective imaging analysis comparing the relative distances of each gold particle to the nearest phage head and tail structure. Immunization led to a strong IgG response, plateauing 7 days after the last phage injection. There was no significant correlation between K-rate and antibody titer over time. TEM showed IgG binding to the head structure of phage K. Image analysis showed a significant reduction in relative distances between antibodies and phage head structures when comparing samples from day 0 and day 28 (P < 0.0001). These results suggest that while individual serum analysis for antibodies against therapeutic phage bears consideration prior to and with prolonged therapy, during phage application, the formation of specific antibodies against phage may only partially explain decreased phage potency in the presence of immunized serum. Instead, other factors may contribute to an individual's "humoral receptiveness" to phage therapy. Future investigations should be directed toward the identification of the humoral factors that have the most significant predictive value on phage potency in vivo.

Evaluation of Sonicate Fluid Culture Cutoff Points for Periprosthetic Joint Infection Diagnosis.

Introduction: Implant sonication is useful for recovery of periprosthetic joint infection (PJI) pathogens in culture, but exact cutoff points for definition of clinically significant sonicate fluid culture results vary from study to study. The aim of this study was to define ideal sonicate fluid culture cutoff points for PJI diagnosis. Methods: Sonicate fluid cultures from hip and knee prosthesis components removed between February 2007 and December 2020 were studied. Prosthesis components were placed in solid containers in the operating room; in the clinical microbiology laboratory, 400 mL Ringer's solution was added, and containers subjected to vortexing, sonication and then vortexing, followed by centrifugation. Concentrated sonicate fluid was plated on aerobic and anaerobic solid media, and culture results reported semiquantitatively, as no growth, <20, 20-50, 51-100, or >100 CFU/10 mL sonicate fluid. Sonicate cultures from cement spacers and cultures yielding more than 1 microorganism were excluded. Sensitivity and specificity of each cutoff point was evaluated. Results: A total of 1448 sonicate fluid cultures were evaluated, 68% from knees and 32% from hips. PJI was present in 644 (44%) cases. Sensitivity of sonicate culture was 75.0% at <20 CFU/10 mL, 55.3% at ≥20 CFU/10 mL, 46.9% at >51 CFU/10 mL, and 39.8% at >100 CFU/10 mL. Specificity was 78.2%, 99.8%, 100%, and 100%, at the 4 cutoff points, respectively. Conclusions: A cutoff point for sonicate fluid culture positivity of ≥20 CFU/10 mL is suitable for PJI diagnosis.

Comparative transcriptomic analysis of Staphylococcus epidermidis associated with periprosthetic joint infection under in vivo and in vitro conditions.

Staphylococcus epidermidis is part of the commensal microbiota of the skin and mucous membranes, though it can also act as a pathogen in certain scenarios, causing a range of infections, including periprosthetic joint infection (PJI). Transcriptomic profiling may provide insights into mechanisms by which S. epidermidis adapts while in a pathogenic compared to a commensal state. Here, a total RNA-sequencing approach was used to profile and compare the transcriptomes of 19 paired PJI-associated S. epidermidis samples from an in vivo clinical source and grown in in vitro laboratory culture. Genomic comparison of PJI-associated and publicly available commensal-state isolates were also compared. Of the 1919 total transcripts found, 145 were from differentially expressed genes (DEGs) when comparing in vivo or in vitro samples. Forty-two transcripts were upregulated and 103 downregulated in in vivo samples. Of note, metal sequestration-associated genes, specifically those related to staphylopine activity (cntA, cntK, cntL, and cntM), were upregulated in a subset of clinical in vivo compared to laboratory grown in vitro samples. About 70% of the total transcripts and almost 50% of the DEGs identified have not yet been annotated. There were no significant genomic differences between known commensal and PJI-associated S. epidermidis isolates, suggesting that differential genomics may not play a role in S. epidermidis pathogenicity. In conclusion, this study provides insights into phenotypic alterations employed by S epidermidis to adapt to infective and non-infected microenvironments, potentially informing future therapeutic targets for related infections.

Evaluation of Treatment of Methicillin-Resistant Staphylococcus aureus Biofilms with Intermittent Electrochemically-Generated H2O2 or HOCl.

Chronic wound infections can be difficult to treat and may lead to impaired healing and worsened patient outcomes. Novel treatment strategies are needed. This study evaluated effects of intermittently produced H2O2 and HOCl, generated via an electrochemical bandage (e-bandage), against methicillin-resistant Staphylococcus aureus biofilms in an agar membrane biofilm model. By changing the working electrode potential, the e-bandage generated either HOCl (1.5 VAg/AgCl) or H2O2 (-0.6 VAg/AgCl). The degree of biocidal activity of intermittent treatment with HOCl and H2O2 correlated with HOCl treatment time; HOCl treatment durations of 0, 1.5, 3, 4.5, and 6 hours (with the rest of the 6 hour total treatment time devoted to H2O2 generation) resulted in mean biofilm reductions of 1.36±0.2, 2.22±0.16, 3.46±0.38, 4.63±0.74 and 7.66±0.5 log CFU/cm2, respectively vs. non-polarized controls, respectively. However, application of H2O2 immediately after HOCl treatment was detrimental to biofilm removal. For example, 3-hours HOCl treatment followed by 3-hours H2O2 resulted in a 1.90±0.84 log CFU/cm2 lower mean biofilm reduction than 3-hours HOCl treatment followed by 3-hours non-polarization. HOCl generated over 3-hours exhibited biocidal activity for at least 7.5-hours after e-bandage operation ceased; 3-hours of HOCl generation followed by 7.5-hours of non-polarization resulted in a biofilm cell reduction of 7.92±0.12 log CFU/cm2 vs. non polarized controls. Finally, intermittent treatment with HOCl (i.e., interspersed with periods of e-bandage non-polarization) for various intervals showed similar effects (approximately 6 log CFU/cm2 reduction vs. non-polarized control) to continuous treatment with HOCl for 3-hours, followed by 3-hours of non-polarization. These findings suggest that timing and sequencing of HOCl and H2O2 treatments are crucial for maximizing biofilm control.

DUAL ACTION ELECTROCHEMICAL BANDAGE OPERATED by a PROGRAMMABLE MULTIMODAL WEARABLE POTENTIOSTAT.

Electrochemical bandages (e-bandages) can be applied to biofilm-infected wounds to generate reactive oxygen species, such as hypochlorous acid (HOCl) or hydrogen peroxide (H 2 O 2 ). The e-bandage-generated HOCl or H 2 O 2 kills biofilms in vitro and in infected wounds on mice. The HOCl-generating e-bandage is more active against biofilms in vitro , although this distinction is less apparent in vivo . The H 2 O 2 -generating e-bandage, more than the HOCl-generating e-bandage, is associated with improved healing of infected wounds. A strategy in which H 2 O 2 and HOCl are generated alternately-for dual action-was explored. The goal was to develop a programmable multimodal wearable potentiostat (PMWP) that could be programmed to generate HOCl or H 2 O 2 , as needed. An ultralow-power microcontroller unit managed operation of the PMWP. The system was operated with a 260-mAh capacity coin battery and weighed 4.6 grams, making it suitable for small animal experiments or human use. The overall cost of a single wearable potentiostat was $6.50 (USD). The device was verified using established electrochemical systems and functioned comparably to a commercial potentiostat. To determine antimicrobial effectiveness, PMWP-controlled e-bandages were tested against clinical isolates of four prevalent chronic wound bacterial pathogens, methicillin-resistant Staphylococcus aureus (MRSA), Pseudomonas aeruginosa, Acinetobacter baumannii , and Enterococcus faecium , and one fungal pathogen of emerging concern, Candida auris . PMWP-controlled e-bandages exhibited broad-spectrum activity against biofilms of all study isolates tested when programmed to deliver HOCl followed by H 2 O 2 . These results show that the PMWP operates effectively and is suitable for animal testing.

HOCl-producing Electrochemical Bandage is Active in Murine Polymicrobial Wound Infection.

Wound infections, exacerbated by the prevalence of antibiotic-resistant bacterial pathogens, necessitate innovative antimicrobial approaches. Polymicrobial infections, often involving Pseudomonas aeruginosa and methicillin-resistant Staphylococcus aureus (MRSA), present formidable challenges due to biofilm formation and antibiotic resistance. Hypochlorous acid (HOCl), a potent antimicrobial agent produced naturally by the immune system, holds promise as an alternative therapy. An electrochemical bandage (e-bandage) that generates HOCl in situ was evaluated for treatment of murine wound biofilm infections containing both MRSA and P. aeruginosa with "difficult-to-treat" resistance. Previously, the HOCl-producing e-bandage was shown to reduce wound biofilms containing P. aeruginosa alone. Compared to non-polarized e-bandage (no HOCl production) and Tegaderm only controls, the polarized e-bandages reduced bacterial loads in wounds infected with MRSA plus P. aeruginosa (MRSA: vs Tegaderm only - 1.4 log10 CFU/g, p = 0.0015, vs. non-polarized - 1.1 log10 CFU/g, p = 0.026. P. aeruginosa: vs Tegaderm only - 1.6 log10 CFU/g, p = 0.0015, vs non-polarized - 1.6 log10 CFU/g, p = 0.0032), and MRSA alone (vs Tegaderm only - 1.3 log10 CFU/g, p = 0.0048, vs. non-polarized - 1.1 log10 CFU/g, p = 0.0048), without compromising wound healing or causing tissue toxicity. Addition of systemic antibiotics did not enhance the antimicrobial efficacy of e-bandages, highlighting their potential as standalone therapies. This study provides additional evidence for the HOCl-producing e-bandage as a novel antimicrobial strategy for managing wound infections, including in the context of antibiotic resistance and polymicrobial infections.

Guide to Utilization of the Microbiology Laboratory for Diagnosis of Infectious Diseases: 2024 Update by the Infectious Diseases Society of America (IDSA) and the American Society for Microbiology (ASM).

The critical nature of the microbiology laboratory in infectious disease diagnosis calls for a close, positive working relationship between the physician and the microbiologists who provide enormous value to the health care team. This document, developed by experts in both adult and pediatric laboratory and clinical medicine, provides information on which tests are valuable and in which contexts, and on tests that add little or no value for diagnostic decisions. Sections are divided into anatomic systems, including Bloodstream Infections and Infections of the Cardiovascular System, Central Nervous System Infections, Ocular Infections, Soft Tissue Infections of the Head and Neck, Upper Respiratory Infections, Lower Respiratory Tract infections, Infections of the Gastrointestinal Tract, Intraabdominal Infections, Bone and Joint Infections, Urinary Tract Infections, Genital Infections, and Skin and Soft Tissue Infections; or into etiologic agent groups, including arboviral Infections, Viral Syndromes, and Blood and Tissue Parasite Infections. Each section contains introductory concepts, a summary of key points, and detailed tables that list suspected agents; the most reliable tests to order; the samples (and volumes) to collect in order of preference; specimen transport devices, procedures, times, and temperatures; and detailed notes on specific issues regarding the test methods, such as when tests are likely to require a specialized laboratory or have prolonged turnaround times. In addition, the pediatric needs of specimen management are also addressed. There is redundancy among the tables and sections, as many agents and assay choices overlap. The document is intended to serve as a reference to guide physicians in choosing tests that will aid them to diagnose infectious diseases in their patients.

Practical Guidance for Clinical Microbiology Laboratories: Microbiologic diagnosis of implant-associated infections.

SUMMARYImplant-associated infections (IAIs) pose serious threats to patients and can be associated with significant morbidity and mortality. These infections may be difficult to diagnose due, in part, to biofilm formation on device surfaces, and because even when microbes are found, their clinical significance may be unclear. Despite recent advances in laboratory testing, IAIs remain a diagnostic challenge. From a therapeutic standpoint, many IAIs currently require device removal and prolonged courses of antimicrobial therapy to effect a cure. Therefore, making an accurate diagnosis, defining both the presence of infection and the involved microorganisms, is paramount. The sensitivity of standard microbial culture for IAI diagnosis varies depending on the type of IAI, the specimen analyzed, and the culture technique(s) used. Although IAI-specific culture-based diagnostics have been described, the challenge of culture-negative IAIs remains. Given this, molecular assays, including both nucleic acid amplification tests and next-generation sequencing-based assays, have been used. In this review, an overview of these challenging infections is presented, as well as an approach to their diagnosis from a microbiologic perspective.

Infectious Diseases Society of America Guidelines on the Diagnosis of COVID-19: Serologic Testing.

BACKGROUND: The role of serologic testing for SARS-CoV-2 has evolved during the pandemic as seroprevalence in global populations has increased. The Infectious Diseases Society of America (IDSA) convened an expert panel to perform a systematic review of the coronavirus disease 2019 (COVID-19) serology literature and construct updated best practice guidance related to SARS-CoV-2 serologic testing. This guideline is an update to the fourth in a series of rapid, frequently updated COVID-19 guidelines developed by IDSA. OBJECTIVE: To develop evidence-based recommendations and identify unmet research needs pertaining to the use of anti-SARS-CoV-2 antibody tests for diagnosis, decisions related to vaccination and administration of monoclonal antibodies or convalescent plasma in immunocompromised patients, and identification of a serologic correlate of immunity. METHODS: A multidisciplinary panel of infectious diseases clinicians, clinical microbiologists and experts in systematic literature reviewed, identified, and prioritized clinical questions related to the use of SARS-CoV-2 serologic tests. Grading of Recommendations Assessment, Development and Evaluation (GRADE) methodology was used to assess the certainty of evidence and make testing recommendations. RESULTS: The panel recommends against serologic testing to diagnose SARS-CoV-2 infection in the first two weeks after symptom onset (strong recommendations, low certainty of evidence). Serologic testing should not be used to provide evidence of COVID-19 in symptomatic patients with a high clinical suspicion and repeatedly negative nucleic acid amplification test results (strong recommendation, very low certainty of evidence). Serologic testing may assist with the diagnosis of multisystem inflammatory syndrome in children (strong recommendation, very low certainty of evidence). To seek evidence for prior SARS-CoV-2 infection, the panel suggests testing for IgG, IgG/IgM, or total antibodies to nucleocapsid protein three to five weeks after symptom onset (conditional recommendation, low certainty of evidence). In individuals with previous SARS-CoV-2 infection or vaccination, we suggest against routine serologic testing given no demonstrated benefit to improving patient outcomes (conditional recommendation, very low certainty of evidence.) The panel acknowledges further that a negative spike antibody test may be a useful metric to identify immunocompromised patients who are candidates for immune therapy. CONCLUSIONS: The high seroprevalence of antibodies against SARS-CoV-2 worldwide limits the utility of detecting anti-SARS CoV-2 antibody. The certainty of available evidence supporting the use of serology for diagnosis was graded as very low to low. Future studies should use serologic assays calibrated to a common reference standard.

Genotypic and phenotypic characterization of Enterococcus faecalis isolates from periprosthetic joint infections.

Over 2.5 million prosthetic joint implantation surgeries occur annually in the United States. Periprosthetic joint infections (PJIs), though occurring in only 1-2% of patients receiving replacement joints, are challenging to diagnose and treat and are associated with significant morbidity. The Gram-positive bacterium Enterococcus faecalis, which can be highly antibiotic resistant and is a robust biofilm producer on indwelling medical devices, accounts for 2-11% of PJIs. E. faecalis PJIs are understudied compared to those caused by other pathogens, such as Staphylococcus aureus. This motivates the need to generate a comprehensive understanding of E. faecalis PJIs to guide future treatments for these infections. To address this, we describe a panel of E. faecalis strains isolated from the surface of prosthetic joints in a cohort of individuals treated at Mayo Clinic in Rochester, MN. Here, we present the first complete genome assemblage of E. faecalis PJI isolates. Comparative genomics shows differences in genome size, virulence factors, antimicrobial resistance genes, plasmids, and prophages, underscoring the genetic diversity of these strains. These isolates have strain-specific differences in in vitro biofilm biomass, biofilm burden, and biofilm morphology. We measured robust changes in biofilm architecture and aggregation for all isolates when grown in simulated synovial fluid (SSF). Lastly, we evaluated antibiotic efficacy of these isolates and found strain specific changes across all strains when grown in SSF. Results of this study highlight the existence of genetic and phenotypic heterogeneity among E. faecalis PJI isolates which will provide valuable insight and resources for future E. faecalis PJI research.

Testing for Helicobacter pylori in an era of antimicrobial resistance.

Antimicrobial resistance in Helicobacter pylori has reached alarming levels and is compromising traditional empiric treatment of H. pylori. Antimicrobial susceptibility testing is routinely performed for infectious diseases when there is a risk of resistance and is now recommended to guide therapy for H. pylori. This mini-review overviews the current diagnostics for H. pylori with a focus on tests that enable susceptibility-guided treatment, including molecular tests performed directly on stool and endoscopically collected specimens.

In vitro activity of sulbactam-durlobactam against colistin-resistant and/or cefiderocol-non-susceptible, carbapenem-resistant Acinetobacter baumannii collected in U.S. hospitals.

The activity of a novel ß-lactamase inhibitor combination, sulbactam-durlobactam (SUL-DUR), was tested against 87 colistin-resistant and/or cefiderocol-non-susceptible carbapenem-resistant Acinetobacter baumannii clinical isolates collected from U.S. hospitals between 2017 and 2019. Among them, 89% and 97% were susceptible to SUL-DUR and imipenem plus SUL-DUR, with MIC50/MIC90 values of 2 µg/mL/8 µg/mL and 1 µg/mL/4 µg/mL, respectively. The presence of amino acid substitutions in penicillin-binding protein 3, including previously reported A515V or T526S, was associated with SUL-DUR non-susceptibility.

HOCl-producing electrochemical bandage for treating Pseudomonas aeruginosa-infected murine wounds.

The growing threat of antibiotic-resistant bacterial pathogens necessitates the development of alternative antimicrobial approaches. This is particularly true for chronic wound infections, which commonly harbor biofilm-dwelling bacteria. A novel electrochemical bandage (e-bandage) delivering low-levels of hypochlorous acid (HOCl) was evaluated against Pseudomonas aeruginosa murine wound biofilms. 5 mm skin wounds were created on the dorsum of mice and infected with 106 colony-forming units (CFU) of P. aeruginosa. Biofilms were formed over 2 days, after which e-bandages were placed on the wound beds and covered with Tegaderm. Mice were administered Tegaderm-only (control), non-polarized e-bandage (no HOCl production), or polarized e-bandage (using an HOCl-producing potentiostat), with or without systemic amikacin. Purulence and wound areas were measured before and after treatment. After 48 hours, wounds were harvested for bacterial quantification. Forty-eight hours of polarized e-bandage treatment resulted in mean biofilm reductions of 1.4 log10 CFUs/g (P = 0.0107) vs non-polarized controls and 2.2 log10 CFU/g (P = 0.004) vs Tegaderm-only controls. Amikacin improved CFU reduction in Tegaderm-only (P = 0.0045) and non-polarized control groups (P = 0.0312) but not in the polarized group (P = 0.3876). Compared to the Tegaderm-only group, there was less purulence in the polarized group (P = 0.009). Wound closure was neither impeded nor improved by either polarized or non-polarized e-bandage treatment. Concurrent amikacin did not impact wound closure or purulence. In conclusion, an HOCl-producing e-bandage reduced P. aeruginosa in wound biofilms with no impairment in wound healing, representing a promising antibiotic-free approach for addressing wound infection.

Clinical Outcomes and Bacterial Characteristics of Carbapenem-resistant Acinetobacter baumannii Among Patients From Different Global Regions.

BACKGROUND: Carbapenem-resistant Acinetobacter baumannii (CRAb) is 1 of the most problematic antimicrobial-resistant bacteria. We sought to elucidate the international epidemiology and clinical impact of CRAb. METHODS: In a prospective observational cohort study, 842 hospitalized patients with a clinical CRAb culture were enrolled at 46 hospitals in five global regions between 2017 and 2019. The primary outcome was all-cause mortality at 30 days from the index culture. The strains underwent whole-genome analysis. RESULTS: Of 842 cases, 536 (64%) represented infection. By 30 days, 128 (24%) of the infected patients died, ranging from 1 (6%) of 18 in Australia-Singapore to 54 (25%) of 216 in the United States and 24 (49%) of 49 in South-Central America, whereas 42 (14%) of non-infected patients died. Bacteremia was associated with a higher risk of death compared with other types of infection (40 [42%] of 96 vs 88 [20%] of 440). In a multivariable logistic regression analysis, bloodstream infection and higher age-adjusted Charlson comorbidity index were independently associated with 30-day mortality. Clonal group 2 (CG2) strains predominated except in South-Central America, ranging from 216 (59%) of 369 in the United States to 282 (97%) of 291 in China. Acquired carbapenemase genes were carried by 769 (91%) of the 842 isolates. CG2 strains were significantly associated with higher levels of meropenem resistance, yet non-CG2 cases were over-represented among the deaths compared with CG2 cases. CONCLUSIONS: CRAb infection types and clinical outcomes differed significantly across regions. Although CG2 strains remained predominant, non-CG2 strains were associated with higher mortality. Clinical Trials Registration. NCT03646227.

Comparison of Blood-Based Shotgun and Targeted Metagenomic Sequencing for Microbiological Diagnosis of Infective Endocarditis.

Background: Shotgun and targeted metagenomic sequencing have been shown in separate studies to be potentially useful for culture-free pathogen identification in blood and/or plasma of patients with infective endocarditis (IE). However, the 2 approaches have not been directly compared. The aim of this study was to compare shotgun metagenomic sequencing with targeted metagenomic sequencing (tMGS) for organism identification in blood or plasma of patients with IE. Methods: Patients with possible or definite IE were prospectively enrolled from October 2020 to July 2021. Shotgun metagenomic sequencing was performed with the Karius test, which uses microbial cell-free DNA (mcfDNA) sequencing to detect, identify, and quantitate DNA-based pathogens in plasma. tMGS was performed using a 16S ribosomal RNA (rRNA) polymerase chain reaction assay targeting the V1 to V3 regions of the 16S rRNA gene. Results were compared using the McNemar test of paired proportions. Results: Samples from 34 patients were investigated. The Karius test was positive in 24/34 (71%), including 3/6 (50%) with blood culture-negative endocarditis (BCNE), which was not significantly different from the positivity rate of tMGS (P = .41). Results of the Karius test were concordant with tMGS in 75% of cases. The Karius test detected 2 cases of methicillin-resistant Staphylococcus aureus among the 7 S. aureus detections, in accordance with results of phenotypic susceptibility testing. The combination of blood cultures, the Karius test, and tMGS found a potential causative pathogen in 33/34 (97%), including 5/6 with BCNE. Conclusions: The Karius test and tMGS yielded comparable detection rates; however, beyond organism identification, the Karius test generated potentially useful antibiotic resistance data.

The Infectious Diseases Society of America Guidelines on the Diagnosis of Coronavirus Disease 2019 (COVID-19): Molecular Diagnostic Testing.

Accurate molecular diagnostic tests are necessary for confirming a diagnosis of coronavirus disease 2019 (COVID-19) and for identifying asymptomatic carriage of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). The number of available SARS-CoV-2 nucleic acid detection tests continues to increase as does the COVID-19 diagnostic literature. Thus, the Infectious Diseases Society of America (IDSA) developed an evidence-based diagnostic guideline to assist clinicians, clinical laboratorians, patients, and policymakers in decisions related to the optimal use of SARS-CoV-2 nucleic acid amplification tests. In addition, we provide a conceptual framework for understanding molecular diagnostic test performance, discuss nuances of test result interpretation in a variety of practice settings, and highlight important unmet research needs related to COVID-19 diagnostic testing. IDSA convened a multidisciplinary panel of infectious diseases clinicians, clinical microbiologists, and experts in systematic literature review to identify and prioritize clinical questions and outcomes related to the use of SARS-CoV-2 molecular diagnostics. Grading of Recommendations Assessment, Development and Evaluation (GRADE) methodology was used to assess the certainty of evidence and make testing recommendations. The panel agreed on 12 diagnostic recommendations. Access to accurate SARS-CoV-2 nucleic acid testing is critical for patient care, hospital infection prevention, and the public health response to COVID-19 infection. Information on the clinical performance of available tests continues to grow, but the quality of evidence of the current literature to support this updated molecular diagnostic guideline remains moderate to very low. Recognizing these limitations, the IDSA panel weighed available diagnostic evidence and recommends nucleic acid testing for all symptomatic individuals suspected of having COVID-19. In addition, testing is suggested for asymptomatic individuals with known or suspected contact with a COVID-19 case when the results will impact isolation/quarantine/personal protective equipment (PPE) usage decisions. Evidence in support of rapid testing and testing of upper respiratory specimens other than nasopharyngeal swabs, which offer logistical advantages, is sufficient to warrant conditional recommendations in favor of these approaches.

Interlaboratory comparison of Pseudomonas aeruginosa phage susceptibility testing.

Standardized approaches to phage susceptibility testing (PST) are essential to inform selection of phages for study in patients with bacterial infections. There is no reference standard for assessing bacterial susceptibility to phage. We compared agreement between PST performed at three centers: two centers using a liquid assay standardized between the sites with the third, a plaque assay. Four Pseudomonas aeruginosa phages: PaWRA01ø11 (EPa11), PaWRA01ø39 (EPa39), PaWRA02ø83 (EPa83), PaWRA02ø87 (EPa87), and a cocktail of all four phages were tested against 145 P. aeruginosa isolates. Comparisons were made within measurements at the two sites performing the liquid assay and between these two sites. Agreement was assessed based on coverage probability (CP8), total deviation index, concordance correlation coefficient (CCC), measurement accuracy, and precision. For the liquid assay, there was satisfactory agreement among triplicate measurements made on different days at site 1, and high agreement based on accuracy and precision between duplicate measurements made on the same run at site 2. There was fair accuracy between measurements of the two sites performing the liquid assay, with CCCs below 0.6 for all phages tested. When compared to the plaque assay (performed once at site 3), there was less agreement between results of the liquid and plaque assays than between the two sites performing the liquid assay. Similar findings to the larger group were noted in the subset of 46 P. aeruginosa isolates from cystic fibrosis. Results of this study suggest that reproducibility of PST methods needs further development.

3D Printed Materials for Combating Antimicrobial Resistance.

Three-dimensional (3D) printing is a rapidly growing technology with a significant capacity for translational applications in both biology and medicine. 3D-printed living and non-living materials are being widely tested as a potential replacement for conventional solutions for testing and combating antimicrobial resistance (AMR). The precise control of cells and their microenvironment, while simulating the complexity and dynamics of an in vivo environment, provides an excellent opportunity to advance the modeling and treatment of challenging infections and other health conditions. 3D-printing models the complicated niches of microbes and host-pathogen interactions, and most importantly, how microbes develop resistance to antibiotics. In addition, 3D-printed materials can be applied to testing and delivering antibiotics. Here, we provide an overview of 3D printed materials and biosystems and their biomedical applications, focusing on ever increasing AMR. Recent applications of 3D printing to alleviate the impact of AMR, including developed bioprinted systems, targeted bacterial infections, and tested antibiotics are presented.

HOCl-producing Electrochemical Bandages for Treating Pseudomonas aeruginosa -Infected Murine Wounds.

A novel electrochemical bandage (e-bandage) delivering low-level hypochlorous acid (HOCl) was evaluated against Pseudomonas aeruginosa murine wound biofilms. 5 mm skin wounds were created on the dorsum of Swiss-Webster mice and infected with 10 6 colony forming units (CFU) of P. aeruginosa . Biofilms were formed over two days, after which e-bandages were placed on the wound beds and covered with Tegaderm™. Mice were administered Tegaderm-only (control), non-polarized e-bandage (no HOCl production), or polarized e-bandage (using an HOCl-producing potentiostat), with or without concurrently administered systemic amikacin. Purulence and wound areas were measured before and after treatment. After 48 hours, animals were sacrificed, and wounds were harvested for bacterial quantification. Forty-eight hours of polarized e-bandage treatment resulted in mean biofilm reductions of 1.4 log 10 CFUs/g (9.0 vs 7.6 log 10 ; p = 0.0107) vs non-polarized controls, and 2.2 log 10 CFU/g (9.8 vs 7.6 log 10 ; p = 0.004) vs Tegaderm only controls. Systemic amikacin improved CFU reduction in Tegaderm-only (p = 0.0045) and non-polarized control groups (p = 0.0312), but not in the polarized group (p = 0.3876). Compared to the Tegaderm only group, there was more purulence reduction in the polarized group (p = 0.009), but not in the non-polarized group (p = 0.064). Wound closure was not impeded or improved by either polarized or non-polarized e-bandage treatment. Concurrent amikacin did not impact wound closure or purulence. In conclusion, an HOCl-producing e-bandage reduced P. aeruginosa in wound biofilms with no impairment in wound healing, representing a promising antibiotic-free approach for addressing wound infections.