Impact of phenotypic rapid diagnostic assay on duration of empiric antibiotics for gram-negative bacteremia.

Objective: Rapid diagnostic tests (RDTs) are increasingly being implemented as antimicrobial stewardship tools to facilitate antibiotic modification and reduce complications related to their overutilization. We measured the clinical impact of a phenotypic RDT with antimicrobial stewardship (AMS) in the setting of gram-negative bacteremia. Setting and participants: In this single-center retrospective cohort study, we evaluated adult patients with gram-negative bacteremia who received at least 72 hours of an antibiotic. Methods: The primary outcome was the duration of empiric antibiotic therapy for gram-negative bacteremia. Secondary outcomes included time-to-directed therapy, proportion of modifications, hospital length of stay (LOS), and subsequent infection with a multidrug-resistant organism (MDRO) or C. difficile infection (CDI). Results: The duration of empiric antibiotics decreased in the RDT+AMS group (4 days vs 2 days; P < .01). Time to directed therapy decreased from 75.0 to 27.9 hours (P < .01). Conclusions: The clinical outcomes of LOS, MDRO, and CDI were reduced. The phenotypic RDT demonstrated an improvement in stewardship measures and clinical outcomes.

Systematic Evaluation of the Accelerate Pheno System for Susceptibility Testing of Gram-Negative Bacteria Isolated from Blood Cultures.

Bacteremia is a major cause of morbidity and mortality. Rapid identification of pathogens for early targeted antimicrobial therapy is crucial for detecting emergence of antibiotic resistance and improving outcomes. However, there are limited data regarding the analytical performance of a rapid identification (ID) and antimicrobial susceptibility testing (AST) method like Accelerate Pheno blood culture detection system compared with the conventional methods routinely used in microbiology laboratories. We undertook a systematic quality improvement (QI) study to compare AST results obtained with Accelerate Pheno system rapid ID/AST system with a standard reference method in a university hospital microbiology laboratory. This was a single center, retrospective (5/10/19 to 8/1/19) and prospective (8/1/19 to 1/31/20) study that evaluated all blood cultures growing Gram-negative rods (GNR). We compared AST results obtained using the reference disk diffusion (DD) susceptibility method with those obtained by the Accelerate Pheno system. We calculated the error rates and categorical agreement between the Accelerate Pheno system and DD for each organism and specific drug tested. We evaluated 355 blood cultures growing GNR, of which 284 met the inclusion criteria. We grouped all Enterobacterales (n = 263) for analysis (156 Escherichia coli, 60 Klebsiella spp., 20 Proteus mirabilis, 17 Enterobacter spp., and 10 Serratia marcescens). Twenty-one Pseudomonas aeruginosa isolates were analyzed separately. For Enterobacterales, categorical agreement (CA) was ≥90% for amikacin (AMK), aztreonam (ATM), cefepime (FEP), ceftriaxone (CRO), ertapenem (ETP), gentamicin (GEN), meropenem (MEM), and tobramycin (TOB); and very major error (VME) was <5% for ampicillin/sulbactam (SAM), GEN, MEM, TOB, CRO, and ceftazidime (CAZ). For ciprofloxacin (CIP), CA was 87% and VME was 8%. For P. aeruginosa, CA was ≥90% for AMK and TOB, and VME was ≥5% for AMK, CAZ, GEN, MEM, piperacillin-tazobactam (TZP), and TOB. Accelerate Pheno rapid ID/AST system for GNR isolated from blood culture (BCs) was reliable for some but not all agents in the panel. Based on the findings from this study, our laboratory reports Accelerate Pheno system AST results only for Enterobacterales, and we limit our reports to CRO, CAZ, TZP, CIP, ATM, and GEN. IMPORTANCE This was an 8-month retrospective and prospective study looking at the analytical performance of the Accelerate Pheno system on clinical isolates obtained from patients seen in our tertiary care hospital. Most of the published literature on the analytical performance of Accelerate Pheno System has been from clinical trials with limited data from clinical microbiology laboratories postimplementation of the system. Here we compare the AST results on 355 blood cultures growing Gram-negative bacteria in Accelerate Pheno system with the CLSI reference disk diffusion (DD) method. The findings from this study highlight the "real-world" performance of the Accelerate Pheno system for Gram-negative bacteria from blood cultures. We provide data to show the reliable susceptibility testing results of Enterobacterales for most of the commonly used antimicrobial agents and significant limitation for susceptibility testing results of Pseudomonas aeruginosa on the Accelerate Pheno system.

Comparing mortality in patients with carbapenemase-producing carbapenem resistant Enterobacterales and non-carbapenemase-producing carbapenem resistant Enterobacterales bacteremia.

Carbapenem-resistant Enterobacterales (CRE) are classified as either carbapenemase-producing CRE (CP-CRE) or non-carbapenemase-producing CRE (non-CP-CRE) based on their mechanism of carbapenem resistance. Few studies have compared outcomes associated with each type of infection. We attempted to determine if either CRE subset is associated with increased mortality. We performed a retrospective observational study to collect demographic, clinical and outcomes data to compare patients with CP-CRE and non-CP-CRE bacteremia. Of 146 cases analyzed, 88/146 (60%) were CP-CRE and 58/146 (40%) were non-CP-CRE. Patients with CP-CRE bacteremia were less likely to receive active empiric or targeted antibiotic therapy. Non-CP-CRE bacteremia was associated with a 2.4 times higher hazard of death at 30 days after bacteremia onset compared to CP-CRE (HR, 2.4; 95% CI, 1.2, 4.6). Patients with non-CP-CRE bacteremia had a higher hazard of death at 30 days after bacteremia onset compared to those with CP-CRE bacteremia.

Characterization of predictors of ESBL-producing enterobacteriaceae in urine cultures of emergency department patients.

STUDY OBJECTIVE: With increasing prevalence of extended-spectrum beta-lactamase-producing enterobacteriaceae (ESBLE), more reliable identification of predictors for ESBLE urinary tract infection (UTI) in the emergency department (ED) is needed. Our objective was to evaluate risk factors and their predictive ability for ED patients with ESBLE UTI. METHODS: This was a retrospective case-control study at an urban academic medical center. Microbiology reports identified adult ED patients with positive urine cultures from 2015-2018. Inclusion criteria were diagnosis of UTI with monomicrobial enterobacteriaceae culture growth. Exclusions were cultures with carbapenemase-resistant enterobacteriaceae or urinary colonization. Collected variables included demographics, comorbidities, and recent medical history. Patient disposition, urine culture susceptibilities, presence of ESBLE, empiric antibiotics, and therapy modifications were collected. Patients were stratified based on ESBLE status and analyzed via descriptive statistics. The data were divided into 2 parts: the first used to identify possible predictors of ESBLE UTI and the second used to validate an additive scoring system. RESULTS: Of 466 patients, 16.3% had ESBLE urine culture growth and 83.7% did not; 39.5% of ESBLE patients required antibiotic therapy modification, as compared to 6.4% of ESBLE negative patients (odds ratio [OR] 9.5; confidence interval [CI] 8.9-10.1). Independent predictors of ESBLE UTI were IV antibiotics within 1 year (OR 5.4; CI 2.1-12.8), surgery within 90 days (OR 6.4; CI 1.5-27.8), and current refractory UTI (OR 8.5; CI 2.0-36.6). CONCLUSION: Independent predictors of ESBLE UTI in emergency department patients included IV antibiotics within 1 year, surgery within 90 days, and current refractory UTI.

Point-Counterpoint: Differences between the European Committee on Antimicrobial Susceptibility Testing and Clinical and Laboratory Standards Institute Recommendations for Reporting Antimicrobial Susceptibility Results.

INTRODUCTIONAntibiotic susceptibility test results are among the most important results issued by clinical microbiology laboratories because they routinely guide critical treatment decisions. Interpretations of MIC or disk diffusion test results, such as "susceptible" or "resistant," are easily understood. Clinical laboratories also need to determine whether and how their reports will reflect more complex situations. Such situations include, first, whether there is need to administer higher or more frequent doses of antibiotic than usual for clinical efficacy; second, whether an antimicrobial is likely to be effective at a body site where it concentrates; and third, whether there is some uncertainty in the test results due to technical variability that cannot be eliminated. Two leading organizations that set standards for antimicrobial susceptibility testing, the European Committee on Antimicrobial Susceptibility Testing (EUCAST) and the Clinical and Laboratory Standards Institute (CLSI), have taken different strategies to deal with these challenges. In this Point-Counterpoint, Gunnar Kahlmeter and Christian Giske discuss how EUCAST is addressing these issues, and Thomas Kirn and Susan Sharp discuss the CLSI approach.

A Laboratory Medicine Best Practices Systematic Review and Meta-analysis of Nucleic Acid Amplification Tests (NAATs) and Algorithms Including NAATs for the Diagnosis of Clostridioides (Clostridium) difficile in Adults.

The evidence base for the optimal laboratory diagnosis of Clostridioides (Clostridium) difficile in adults is currently unresolved due to the uncertain performance characteristics and various combinations of tests. This systematic review evaluates the diagnostic accuracy of laboratory testing algorithms that include nucleic acid amplification tests (NAATs) to detect the presence of C. difficile The systematic review and meta-analysis included eligible studies (those that had PICO [population, intervention, comparison, outcome] elements) that assessed the diagnostic accuracy of NAAT alone or following glutamate dehydrogenase (GDH) enzyme immunoassays (EIAs) or GDH EIAs plus C. difficile toxin EIAs (toxin). The diagnostic yield of NAAT for repeat testing after an initial negative result was also assessed. Two hundred thirty-eight studies met inclusion criteria. Seventy-two of these studies had sufficient data for meta-analysis. The strength of evidence ranged from high to insufficient. The uses of NAAT only, GDH-positive EIA followed by NAAT, and GDH-positive/toxin-negative EIA followed by NAAT are all recommended as American Society for Microbiology (ASM) best practices for the detection of the C. difficile toxin gene or organism. Meta-analysis of published evidence supports the use of testing algorithms that use NAAT alone or in combination with GDH or GDH plus toxin EIA to detect the presence of C. difficile in adults. There is insufficient evidence to recommend against repeat testing of the sample using NAAT after an initial negative result due to a lack of evidence of harm (i.e., financial, length of stay, or delay of treatment) as specified by the Laboratory Medicine Best Practices (LMBP) systematic review method in making such an assessment. Findings from this systematic review provide clarity to diagnostic testing strategies and highlight gaps, such as low numbers of GDH/toxin/PCR studies, in existing evidence on diagnostic performance, which can be used to guide future clinical research studies.

Collaboration between an antimicrobial stewardship team and the microbiology laboratory can shorten time to directed antibiotic therapy for methicillin-susceptible staphylococcal bacteremia and to discontinuation of antibiotics for coagulase-negative staphylococcal contaminants.

BACKGROUND: Rapid identification of Gram-positive cocci in clusters (GPCC) in positive blood cultures (pBC) may limit exposure to unnecessary or inappropriate antibiotics. METHODS: Inpatients with pBC showing GPCC between October 2013 and December 2017 were included. In the baseline period (BL), final ID and susceptibility results were reported in the electronic medical record (EMR) within 48 h of telephoned Gram stain report. The laboratory introduced rapid phenotypic identification and direct susceptibility testing (INT1), later replaced by PCR (INT2). In the last Intervention (INT3), Antimicrobial Stewardship Response Team (ASRT) contacted providers with PCR results and recommendations. RESULTS: Time to directed therapy (TDT) for MSSA and coagulase-negative Staphylococci (CoNS) decreased from BL to INT3 (48.5-17.9 h, 50.3-16.4 h, respectively). Time to ID from BL to INT3 for MSSA and CoNS also decreased (23.2-1.9 h, 44.7-2.8, respectively). CONCLUSIONS: TDT can be improved by modification of reporting methods with utilization of an ASRT.

Assessing Clinical Microbiology Practice Guidelines: American Society for Microbiology Ad Hoc Committee on Evidence-Based Laboratory Medicine Practice Guidelines Assessment.

As part of the American Society for Microbiology (ASM) Evidence-Based Laboratory Medicine Practice Guidelines Committee of the Professional Practice Committee, an ad hoc committee was formed in 2014 to assess guidelines published by the committee using an assessment tool, Appraisal of Guidelines for Research Evaluation II (AGREE II). The AGREE II assessment helps reviewers determine whether published guidelines are robust, transparent, and clear in presenting practice recommendations in a standardized manner. Identifying strengths and weaknesses of practice guidelines by ad hoc assessments helps with improving future guidelines through the participation of key stakeholders. This minireview describes the development of the ad hoc committee and results from their review of several ASM best practices guidelines and a non-ASM practice guideline from the Emergency Nurses Association.

Automatic Digital Plate Reading for Surveillance Cultures.

The automation of specimen processing and culture workup has rapidly emerged in clinical microbiology laboratories throughout the world and more recently in the United States. While many U.S. laboratories have implemented some form of automated specimen processing and some have begun performing digital plate reading, automated colony analysis is just beginning to be utilized clinically. In this issue of the Journal of Clinical Microbiology, M. L. Faron et al. (J Clin Microbiol 54:2470-2475, 2016, http://dx.doi.org/10.1128/JCM.01040-16) report the results of their evaluation of the performance of the WASPLab Chromogenic Detection Module (CDM) for categorizing chromogenic agar plates as negative or "nonnegative" for vancomycin-resistant enterococci (VRE). Their major finding was 100% sensitivity for detection of "nonnegative" specimens using CDM compared to manual methods for specimens plated on two different types of VRE chromogenic agar plates. Additionally, utilization of digital plate reading in conjunction with automated colony analysis was predicted to result in significant savings based on greatly reduced labor costs.

Point-Counterpoint: A Nucleic Acid Amplification Test for Streptococcus pyogenes Should Replace Antigen Detection and Culture for Detection of Bacterial Pharyngitis.

Nucleic acid amplification tests (NAATs) have frequently been the standard diagnostic approach when specific infectious agents are sought in a clinic specimen. They can be applied for specific agents such as S. pyogenes, or commercial multiplex NAATs for detection of a variety of pathogens in gastrointestinal, bloodstream, and respiratory infections may be used. NAATs are both rapid and sensitive. For many years, S. pyogenes testing algorithms used a rapid and specific group A streptococcal antigen test to screen throat specimens, followed, in some clinical settings, by a throat culture for S. pyogenes to increase the sensitivity of its detection. Now S. pyogenes NAATs are being used with increasing frequency. Given their accuracy, rapidity, and ease of use, should they replace antigen detection and culture for the detection of bacterial pharyngitis? Bobbi Pritt and Robin Patel of the Mayo Clinic, where S. pyogenes NAATs have been used for well over a decade with great success, will explain the advantages of this approach, while Richard (Tom) Thomson and Tom Kirn of the NorthShore University HealthSystem will discuss their concerns about this approach to diagnosing bacterial pharyngitis.

Cost-Effectiveness of 30- Compared to 20-Milliliter Blood Cultures: a Retrospective Study.

The importance of blood culture (BC) volume for detection of bloodstream infections (BSIs) is documented. Recently, improved diagnostic sensitivity was demonstrated for 30- versus 20-ml BCs in adults (Cockerill FR, Wilson JW, Vetter EA, Goodman KM, Torgerson CA, Harmsen WS, Schleck CD, IIstrup DM, Washington JA, Wilson WR. Clin Infect Dis 38:1724-1730, 2004, http://dx.doi.org/10.1128/JCM.01314-11). Hospitals receive higher reimbursement for patients with documented septicemia. We determined the cost-effectiveness of 30-ml versus 20-ml BCs using results from our institution and previously published data. Positive BC results from 292 bacteremic episodes were reviewed. The costs of the reagents, equipment, phlebotomist, and technologist time were determined. The medical records department provided Medicare reimbursement (MR) data for patients with selected ICD-9 codes. These data provided an estimate of the annualized increase in MR versus costs associated with conversion to 30-ml BCs. MR for 464 annual primary BSIs was $24,808/episode. An expected 7.2% increase in BSIs detected using 30-ml BCs would add 34 additional cases annually and increase MR by $843,472. Comparative MR data for cases where septicemia complicated another diagnosis were available for 4 International Classification of Diseases, Ninth Revision (ICD-9) codes: laparoscopic cholecystectomy, biliary tract disorders, pneumonia, and cellulitis. The mean incremental MR was $9,667 per episode, which projected to a $483,350 revenue increase annually. The annual cost associated with conversion to 30-ml BCs was estimated to be $157,798. Thus, the potential net increase in hospital revenue would be $1,169,031 for 30-ml versus 20-ml BCs. Our results suggest that conversion to 30-ml BCs may not only improve patient care by detecting more BSIs but also increase hospital revenue substantially.

Air pollution particulate matter alters antimycobacterial respiratory epithelium innate immunity.

Inhalation exposure to indoor air pollutants and cigarette smoke increases the risk of developing tuberculosis (TB). Whether exposure to ambient air pollution particulate matter (PM) alters protective human host immune responses against Mycobacterium tuberculosis has been little studied. Here, we examined the effect of PM from Iztapalapa, a municipality of Mexico City, with aerodynamic diameters below 2.5 µm (PM2.5) and 10 µm (PM10) on innate antimycobacterial immune responses in human alveolar type II epithelial cells of the A549 cell line. Exposure to PM2.5 or PM10 deregulated the ability of the A549 cells to express the antimicrobial peptides human ß-defensin 2 (HBD-2) and HBD-3 upon infection with M. tuberculosis and increased intracellular M. tuberculosis growth (as measured by CFU count). The observed modulation of antibacterial responsiveness by PM exposure was associated with the induction of senescence in PM-exposed A549 cells and was unrelated to PM-mediated loss of cell viability. Thus, the induction of senescence and downregulation of HBD-2 and HBD-3 expression in respiratory PM-exposed epithelial cells leading to enhanced M. tuberculosis growth represent mechanisms by which exposure to air pollution PM may increase the risk of M. tuberculosis infection and the development of TB.

Serotype 10A in case patients with invasive pneumococcal disease: a pilot study of PCR-based serotyping in New Jersey.

In 2008, the New Jersey Department of Health (NJDOH) identified a 21.1% increase in reported invasive pneumococcal disease (IPD). In 2009, NJDOH piloted nucleic acid-based serotyping to characterize serotypes causing IPD. From April through September, NJDOH received specimens from 149 of 302 (49%) case patients meeting our case definition. An uncommon serotype, 10A, accounted for 25.2% of IPD overall and was identified in 12 counties, but it was associated with one county (rate ratio = 5.4, 95% confidence interval [CI] 2.1, 11.8). NJDOH subsequently conducted a case-control study to assess the presentation of and clinical risk factors for 10A IPD. Case patients with 10A IPD were more likely to have had immunosuppression, asthma, and multiple chronic medical conditions than control subjects had (odds ratio [OR] = 2.6, 95% CI 1.1, 6.3; OR=4.7, 95% CI 1.7, 13.2; and OR=2.3, 95% CI 1.0, 5.2, respectively). State-based pneumococcal serotype testing identified an uncommon serotype in New Jersey. Continued pneumococcal serotype surveillance might help the NJDOH identify and respond to future serotype-specific increases.

Systematic evaluation of commercial susceptibility testing methods for determining the in vitro activity of daptomycin versus Staphylococcus aureus and Enterococci.

We systematically evaluated 5 methods for testing daptomycin versus 48 Enterococcus faecalis, 51 Enterococcus faecium, and 50 Staphylococcus aureus isolates using (i and ii) broth microdilution (BMD) with 50-mg/liter calcium medium supplementation (reference method) and 30-mg/liter calcium medium supplementation (BMD30 method), (iii) Etest, and (iv and v) MicroScan panel 33 using 2 methods to prepare the bacterial inoculum (MicroScan turbidity and MicroScan Prompt). Isolates were categorized as susceptible (S) or nonsusceptible (NS) based on measured MICs. Essential (± 1 dilution) agreement (EA) and categorical (S/NS) agreement (CA) for each method were compared to the reference method. For E. faecium, categorical agreement was poor between the reference method and BMD30 as well as with the three commercial methods, with frequent false-NS results (30 for BMD30, 18 for Etest, 22 for MicroScan Prompt, and 25 for MicroScan turbidity). All E. faecalis isolates were judged to be S by the reference method; two of these isolates were categorized as NS using the BMD30 method, and one was categorized as NS by all three commercial methods. All S. aureus isolates were judged to be S using all five methods. MIC values determined by the comparator methods tended to be higher than those for the reference method, especially for E. faecium isolates. EAs between the reference BMD and BMD30, Etest, MicroScan Prompt, and MicroScan turbidity were 63%, 63%, 63%, and 56%, respectively, for E. faecium, 87%, 83%, 98%, and 80%, respectively, for E. faecalis, and all 100% for S. aureus.

Laboratory diagnosis of HIV in adults: a review of current methods.

Of the estimated 1.2 million people infected with human immunodeficiency virus (HIV) in the United States, 20% are unaware of their diagnosis. Improved methods of HIV testing could decrease this number, as well as identify those who have recently acquired HIV infection and are at the most critical stage of infectivity. People with acute HIV infection have demonstrated enhanced transmission of HIV in multiple epidemiologic and pathogenetic studies. More than 50 000 HIV infections occur annually in the United States, and 30%-50% have been attributed to persons with recent infection. The original HIV diagnostic testing algorithm was developed by the Centers for Disease Control and Prevention in 1989. Recently proposed alterations to the algorithm would incorporate advancements made in HIV diagnostic testing, thereby increasing sensitivity while reducing turnaround time and cost. Improved diagnosis of acute HIV, and HIV type 2 in particular, would be expected. Knowledge of the available laboratory methods for HIV diagnosis is essential in the fight against the spread of HIV.

Clostridium difficile MazF toxin exhibits selective, not global, mRNA cleavage.

Clostridium difficile is an important, emerging nosocomial pathogen. The transition from harmless colonization to disease is typically preceded by antimicrobial therapy, which alters the balance of the intestinal flora, enabling C. difficile to proliferate in the colon. One of the most perplexing aspects of the C. difficile infectious cycle is its ability to survive antimicrobial therapy and transition from inert colonization to active infection. Toxin-antitoxin (TA) systems have been implicated in facilitating persistence after antibiotic treatment. We identified only one TA system in C. difficile strain 630 (epidemic type X), designated MazE-cd and MazF-cd, a counterpart of the well-characterized Escherichia coli MazEF TA system. This E. coli MazF toxin cleaves mRNA at ACA sequences, leading to global mRNA degradation, growth arrest, and death. Likewise, MazF-cd expression in E. coli or Clostridium perfringens resulted in growth arrest. Primer extension analysis revealed that MazF-cd cleaved RNA at the five-base consensus sequence UACAU, suggesting that the mRNAs susceptible to cleavage comprise a subset of total mRNAs. In agreement, we observed differential cleavage of several mRNAs by MazF-cd in vivo, revealing a direct correlation between the number of cleavage recognition sites within a given transcript and its susceptibility to degradation by MazF-cd. Interestingly, upon detailed statistical analyses of the C. difficile transcriptome, the major C. difficile virulence factor toxin B (TcdB) and CwpV, a cell wall protein involved in aggregation, were predicted to be significantly resistant to MazF-cd cleavage.