Dynamics of cytomegalovirus-specific T-cell recovery in allogeneic hematopoietic cell transplant recipients using a commercially available flow cytometry assay: A pilot study.

BACKGROUND: Cytomegalovirus-specific T-cell-mediated immunity (CMV-CMI) protects from CMV infection in allogeneic hematopoietic cell transplantation (allo-HCT), but to date, there is no validated measure of CMV immunity for this population. METHODS: In this prospective, observational, pilot study, CMV T-cell responses were evaluated monthly and at onset of graft-versus-host disease (GVHD) or CMV infection in CMV-seropositive allo-HCT recipients using a commercial flow cytometry assay, the CMV inSIGHT T-Cell Immunity Panel (CMV-TCIP). The primary endpoint was the time to first positive CMV-TCIP, defined as percentage of interferon-γ-producing CD4+ or CD8+ CMV-specific T cells >0.2%. Letermovir was prescribed from day +10 to ≥100. RESULTS: Twenty-eight allo-HCT recipients were enrolled. The median time to first positive CMV-TCIP result was earlier for CD4+ (60 days [interquartile range, IQR 33‒148]) than for CD8+ T cells (96 days [IQR 33‒155]) and longer for haploidentical and mismatched transplant recipients (77 and 96 days, respectively) than for matched donors (45 and 33 days, respectively). CD4+ and CD8+ CMV-CMI recovery was sustained in 10/10 (100%) and 10/11 (91%) patients, respectively, without GVHD, whereas CD4+ and/or CD8+ CMV-CMI was lost in 4/6 and 2/6 patients, respectively, with GVHD requiring steroids. As a predictor of clinically significant CMV infection in patients with low-level CMV reactivation, the sensitivity and negative predictive value of CMV-TCIP were 90% and 87.5%, respectively, for CD4+ CMV-TCIP and 66.7% and 62.5%, respectively, for CD8+ CMV-TCIP. CONCLUSIONS: There was significant variability in time to CMV-CMI recovery post-HCT, with slower recovery after haploidentical and mismatched HCT. CD4+ CMV-CMI may protect against CS-CMVi, but immunity may be lost with GVHD diagnosis and treatment.

Reappraisal of p16 for Determining HPV Status of Head and Neck Carcinomas Arising in HPV Hotspots.

In an era of head and neck oncology where HPV status will soon dictate patient management, reliable HPV detection is critical. P16 immunohistochemistry (IHC) is currently recommended as the test of choice for oropharyngeal squamous cell carcinomas (OPSCCs). The purpose of this study was to determine the performance characteristics of p16 IHC based on a large clinical experience of squamous cell carcinomas (SCC) arising from HPV hot-spot regions of the head and neck. Consecutive OPSCCs, sinonasal SCCs, and metastatic SCCs of unknown primary sites were evaluated for the presence of HPV by p16 IHC and PCR-based HPV DNA testing as part of clinical care. For discrepant cases, high-risk HPV E6/E7 mRNA in situ hybridization (ISH) and, when possible, matrix-assisted laser desorption/ionization-time of flight (MALDI-TOF) mass spectrometry (MassArray) genotyping were performed. 746 cancers underwent HPV testing by p16 IHC and DNA PCR genotyping. There was a 95.6% concordance between the 2 assays. Of the 33 discrepant cases, 32 cases (4.3%) were p16 positive but HPV DNA negative. In these cases, 68% were positive for mRNA ISH, invariably related to a non-16 HPV genotype. P16 IHC had an overall accuracy of 98.8%, a sensitivity of 99.8%, and a specificity of 92.1%. P16 IHC is a sensitive and specific assay for determining HPV status. HPV DNA PCR appears vulnerable to HPV genotype diversity and is prone to missing rare non-16 genotypes. HPV mRNA ISH is a practical and reliable direct measure of HPV that may help eliminate the small number of false-positive p16 cases and avoid potential patient harm related to erroneous HPV classification.

mEnrich-seq: methylation-guided enrichment sequencing of bacterial taxa of interest from microbiome.

Metagenomics has enabled the comprehensive study of microbiomes. However, many applications would benefit from a method that sequences specific bacterial taxa of interest, but not most background taxa. We developed mEnrich-seq (in which 'm' stands for methylation and seq for sequencing) for enriching taxa of interest from metagenomic DNA before sequencing. The core idea is to exploit the self versus nonself differentiation by natural bacterial DNA methylation and rationally choose methylation-sensitive restriction enzymes, individually or in combination, to deplete host and background taxa while enriching targeted taxa. This idea is integrated with library preparation procedures and applied in several applications to enrich (up to 117-fold) pathogenic or beneficial bacteria from human urine and fecal samples, including species that are hard to culture or of low abundance. We assessed 4,601 bacterial strains with mapped methylomes so far and showed broad applicability of mEnrich-seq. mEnrich-seq provides microbiome researchers with a versatile and cost-effective approach for selective sequencing of diverse taxa of interest.

Clinical, microbiological and genomic characterization of Gram-negative bacteria with dual carbapenemases as identified by rapid molecular testing.

Objective: Dual carbapenemase-producing organisms (DCPOs) are an emerging threat that expands the spectrum of antimicrobial resistance. There is limited literature on the clinical and genetic epidemiology of DCPOs. Methods: DCPO isolates were identified by Xpert® Carba-R PCR testing of routine diagnostic cultures performed from 2018 to 2021 at a New York City health system. WGS was performed by Illumina and/or PacBio. Medical records of patients were reviewed for clinical and epidemiological data. Results: Twenty-six DCPO isolates were obtained from 13 patients. Klebsiella pneumoniae (n = 22) was most frequent, followed by Pseudomonas aeruginosa (n = 2), Escherichia coli (n = 1) and Enterobacter cloacae (n = 1). The most common DCPO combination was blaNDM/blaOXA-48-like (n = 16). Notably, 1.05% (24/2290) of carbapenem-resistant Enterobacterales isolates were identified as DCPOs. The susceptibility profiles matched the identified resistance genes, except for a K. pneumoniae (blaKPC/blaOXA-48-like) isolate that was phenotypically susceptible to meropenem. Eleven patients were hospitalized within the year prior to admission, and received antibiotic(s) 1 month prior. Seven patients were originally from outside the USA. Hypertension, kidney disease and diabetes were frequent comorbidities. Death in two cases was attributed to DCPO infection. WGS of eight isolates showed that carbapenemases were located on distinct plasmids, except for one K. pneumoniae isolate where NDM and KPC carbapenemases were located on a single IncC-type plasmid backbone. Conclusions: Here we characterized a series of DCPOs from New York City. Foreign travel, prior hospitalization, antibiotic usage and comorbidities were common among DCPO cases. All carbapenemases were encoded on plasmids, which may facilitate horizontal transfer.

Screening for Latent Tuberculosis Infection in Solid Organ Transplant Recipients to Predict Active Disease: A Systematic Review and Meta-Analysis of Diagnostic Studies.

Background: This is a systematic review and meta-analysis of diagnostic test accuracy studies to assess the predictive value of both tuberculin skin test (TST) and interferon-gamma release assays (IGRA) for active tuberculosis (TB) among solid organ transplantation (SOT) recipients. Methods: Medline, Embase, and the CENTRAL databases were searched from 1946 until June 30, 2022. Two independent assessors extracted data from studies. Sensitivity analyses were performed to investigate the effect of studies with high or low risk of bias. Methodological quality of each publication was assessed using QUADAS-2. Results: A total of 43 studies (36 403 patients) with patients who were screened for latent TB infection (LTBI) and who underwent SOT were included: 18 were comparative and 25 noncomparative (19 TST, 6 QuantiFERON-TB Gold In-Tube [QFT-GIT]). For IGRA tests taken together, positive predictive value (PPV) and negative predictive value (NPV) were 1.2% and 99.6%, respectively. For TST, PPV was 2.13% and NPV was 95.5%. Overall, PPV is higher when TB burden is higher, regardless of test type, although still low in absolute terms. Incidence of active TB was similar between studies using LTBI prophylaxis (mean incidence 1.22%; 95% confidence interval [CI], .2179-2.221) and those not using prophylaxis (mean incidence 1.045%; 95% CI, 0.2731-1.817; P = .7717). Strengths of this study include the large number of studies available from multiple different countries; limitations include absence of gold standard for diagnosis of latent TB and low incidence of active TB. Conclusions: We found both TST and IGRA had a low PPV and high NPV for the development of active TB posttransplant. Further studies are needed to better understand how to prevent active TB in the SOT population.

Methicillin-resistant Staphylococcus aureus bacteremia with elevated vancomycin minimum inhibitory concentrations.

This case-control study of 25 cases with methicillin-resistant Staphylococcus aureus (MRSA) bacteremia with vancomycin minimum inhibitory concentration (MIC) ≥ 2 µg/mL and 391 controls (MIC < 2 µg/mL) characterized the clinical characteristics, treatments, and outcomes associated with elevated vancomycin MIC. Elevated vancomycin MIC was associated with baseline hemodialysis, prior MRSA colonization, and metastatic infection.

Going local: Evaluating guideline adherence and appropriateness of antibiotic prescribing in patients with febrile neutropenia at an academic teaching hospital.

Background: Febrile neutropenia (FN) is a medical emergency with significant morbidity and mortality for oncology patients, requiring comprehensive workup and timely antibiotic administration. We evaluated concordance with locally developed FN guidelines and outcomes of cancer patients admitted to general internal medicine at an academic teaching hospital. Methods: We conducted a retrospective observational cohort study of patients admitted between July 1, 2016, and June 30, 2017, for FN. Patients were classified as having low-risk or high-risk FN according to their malignancy and chemotherapy. Primary outcome was the proportion of patients receiving guideline-concordant antibiotics within 48 hours of admission to general internal medicine. Secondary outcomes were the proportion of patients in whom empirical antibiotics were active against pathogens isolated, rate of antibiotic-associated adverse events, and in-hospital mortality. We used logistic regression to model relationship between FN risk and guideline-concordant antibiotics. Results: Among 100 patients included, 34 (34%) were low-risk FN and 66 (66%) were high-risk. Proportion of guideline-concordant empirical antibiotics was significantly lower among low-risk FN patients than high-risk patients: 12 (35%) of 34 versus 47 (71%) of 66 (P = .001). Empirical antibiotics were active against 17 (94%) of 18 isolated pathogens. The mortality rate was 3%, and 16% of patients experienced antibiotic-associated adverse events. Hematological malignancy and infectious diseases-trained physician involvement were associated with guideline-concordant prescribing, with adjusted odds ratios of 3.76 (95% CI, 1.46-9.70; P = .006) and 3.71 (95% CI, 1.49-9.23; P = .005), respectively. Conclusions: Guideline concordance was low compared to published reports. Factors influencing appropriate antimicrobial prescribing in patients with FN warrant further exploration.

Implementation of a Routine Screening Program for Latent Tuberculosis Infection among Patients with Acute Leukemia at a Canadian Cancer Center.

BACKGROUND: Screening for latent tuberculosis infection (LTBI) in patients with hematological malignancy is recommended because of their increased risk of tuberculosis (TB). We assessed the utility of tuberculin skin test (TST) screening in patients with acute leukemia and subsequent outcomes of LTBI treatment. METHODS: We retrospectively evaluated patients ≥16 years of age with acute leukemia from 2013-2014 with a TST planted and read prior to the initiation of antineoplastic chemotherapy treatment. Demographics, clinical information and treatment outcomes of LTBI therapy were compared between patients with positive TST (≥10 mm induration) and negative TST. RESULTS: A total of 389 patients with acute leukemia were included in the cohort. Of them, 37/389 (9.5%) had a positive TST. Only 3.4% (8/235) of individuals originating from North and South America as well as the Caribbean were TST positive, while 21% (20/95) of individuals from Asia were TST positive. Diagnostic imaging findings consistent with prior tuberculosis infection were higher in TST positive patients compared to TST negative ones (29.7% versus 9.4%, p < 0.0001). Furthermore, 31/38 patients (81.6%) who were TST positive received LTBI therapy, which was well tolerated. There was no significant difference in overall survival among those who received LTBI therapy compared to those who did not. No patients developed active TB. CONCLUSIONS: Universal screening with TST may be of low yield in individuals with acute leukemia unless patients originate from a TB endemic country. When therapy for LTBI is prescribed, patients with acute leukemia do not experience drug-induced liver toxicity and are likely to complete the intended duration of therapy, thus preventing the development of active tuberculosis.

Utility of liquid biopsy in diagnosing isolated cerebral phaeohyphomycosis: illustrative case.

BACKGROUND: Cladophialophora bantiana is a dematiaceous, saprophytic fungus and a rare but reported cause of intracranial abscesses due to its strong neurotropism. Although it predominantly affects immunocompetent individuals with environmental exposure, more recently, its significance as a highly lethal opportunistic infection in transplant recipients has been recognized. Successful treatment requires timely but often challenging diagnosis, followed by complete surgical excision. Next-generation sequencing of microbial cell-free DNA (cfDNA) from plasma is a novel diagnostic method with the potential to identify invasive fungal infections more rapidly and less invasively than conventional microbiological testing, including brain biopsy. OBSERVATIONS: The authors described the case of a recipient of a liver transplant who presented with seizures and was found to have innumerable ring-enhancing intracranial lesions. The Karius Test, a commercially available method of next-generation sequencing of cfDNA, was used to determine the causative organism. Samples from the patient's plasma identified C. bantiana 6 days before culture results of the surgical specimen, allowing optimization of the empirical antifungal regimen, which led to a reduction in the size of the abscesses. LESSONS: The authors' findings suggest that microbial cfDNA sequencing may be particularly impactful in improving the management of brain abscesses in which the differential diagnosis is wide because of immunosuppression.

A Robust, Highly Multiplexed Mass Spectrometry Assay to Identify SARS-CoV-2 Variants.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants are characterized by differences in transmissibility and response to therapeutics. Therefore, discriminating among them is vital for surveillance, infection prevention, and patient care. While whole-genome sequencing (WGS) is the "gold standard" for variant identification, molecular variant panels have become increasingly available. Most, however, are based on limited targets and have not undergone comprehensive evaluation. We assessed the diagnostic performance of the highly multiplexed Agena MassARRAY SARS-CoV-2 Variant Panel v3 to identify variants in a diverse set of 391 SARS-CoV-2 clinical RNA specimens collected across our health systems in New York City, USA and Bogotá, Colombia (September 2, 2020 to March 2, 2022). We demonstrated almost perfect levels of interrater agreement between this assay and WGS for 9 of 11 variant calls (κ ≥ 0.856) and 25 of 30 targets (κ ≥ 0.820) tested on the panel. The assay had a high diagnostic sensitivity (≥93.67%) for contemporary variants (e.g., Iota, Alpha, Delta, and Omicron [BA.1 sublineage]) and a high diagnostic specificity for all 11 variants (≥96.15%) and all 30 targets (≥94.34%) tested. Moreover, we highlighted distinct target patterns that could be utilized to identify variants not yet defined on the panel, including the Omicron BA.2 and other sublineages. These findings exemplified the power of highly multiplexed diagnostic panels to accurately call variants and the potential for target result signatures to elucidate new ones. IMPORTANCE The continued circulation of SARS-CoV-2 amid limited surveillance efforts and inconsistent vaccination of populations has resulted in the emergence of variants that uniquely impact public health systems. Thus, in conjunction with functional and clinical studies, continuous detection and identification are quintessential to informing diagnostic and public health measures. Furthermore, until WGS becomes more accessible in the clinical microbiology laboratory, the ideal assay for identifying variants must be robust, provide high resolution, and be adaptable to the evolving nature of viruses like SARS-CoV-2. Here, we highlighted the diagnostic capabilities of a highly multiplexed commercial assay to identify diverse SARS-CoV-2 lineages that circulated from September 2, 2020 to March 2, 2022 among patients seeking care in our health systems. This assay demonstrated variant-specific signatures of nucleotide/amino acid polymorphisms and underscored its utility for the detection of contemporary and emerging SARS-CoV-2 variants of concern.

A consensus conference to define the utility of advanced infectious disease diagnostics in solid organ transplant recipients.

The last decade has seen an explosion of advanced assays for the diagnosis of infectious diseases, yet evidence-based recommendations to inform their optimal use in the care of transplant recipients are lacking. A consensus conference sponsored by the American Society of Transplantation (AST) was convened on December 7, 2021, to define the utility of novel infectious disease diagnostics in organ transplant recipients. The conference represented a collaborative effort by experts in transplant infectious diseases, diagnostic stewardship, and clinical microbiology from centers across North America to evaluate current uses, unmet needs, and future directions for assays in 5 categories including (1) multiplex molecular assays, (2) rapid antimicrobial resistance detection methods, (3) pathogen-specific T-cell reactivity assays, (4) next-generation sequencing assays, and (5) mass spectrometry-based assays. Participants reviewed and appraised available literature, determined assay advantages and limitations, developed best practice guidance largely based on expert opinion for clinical use, and identified areas of future investigation in the setting of transplantation. In addition, attendees emphasized the need for well-designed studies to generate high-quality evidence needed to guide care, identified regulatory and financial barriers, and discussed the role of regulatory agencies in facilitating research and implementation of these assays. Findings and consensus statements are presented.

A robust, highly multiplexed mass spectrometry assay to identify SARS-CoV-2 variants.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants are characterized by differences in transmissibility and response to therapeutics. Therefore, discriminating among them is vital for surveillance, infection prevention, and patient care. While whole viral genome sequencing (WGS) is the "gold standard" for variant identification, molecular variant panels have become increasingly available. Most, however, are based on limited targets and have not undergone comprehensive evaluation. We assessed the diagnostic performance of the highly multiplexed Agena MassARRAY ® SARS-CoV-2 Variant Panel v3 to identify variants in a diverse set of 391 SARS-CoV-2 clinical RNA specimens collected across our health systems in New York City, USA as well as in Bogotá, Colombia (September 2, 2020 - March 2, 2022). We demonstrate almost perfect levels of interrater agreement between this assay and WGS for 9 of 11 variant calls (κ ≥ 0.856) and 25 of 30 targets (κ ≥ 0.820) tested on the panel. The assay had a high diagnostic sensitivity (≥93.67%) for contemporary variants (e.g., Iota, Alpha, Delta, Omicron [BA.1 sublineage]) and a high diagnostic specificity for all 11 variants (≥96.15%) and all 30 targets (≥94.34%) tested. Moreover, we highlight distinct target patterns that can be utilized to identify variants not yet defined on the panel including the Omicron BA.2 and other sublineages. These findings exemplify the power of highly multiplexed diagnostic panels to accurately call variants and the potential for target result signatures to elucidate new ones. Importance: The continued circulation of SARS-CoV-2 amidst limited surveillance efforts and inconsistent vaccination of populations has resulted in emergence of variants that uniquely impact public health systems. Thus, in conjunction with functional and clinical studies, continuous detection and identification are quintessential to inform diagnostic and public health measures. Furthermore, until WGS becomes more accessible in the clinical microbiology laboratory, the ideal assay for identifying variants must be robust, provide high resolution, and be adaptable to the evolving nature of viruses like SARS-CoV-2. Here, we highlight the diagnostic capabilities of a highly multiplexed commercial assay to identify diverse SARS-CoV-2 lineages that circulated at over September 2, 2020 - March 2, 2022 among patients seeking care at our health systems. This assay demonstrates variant-specific signatures of nucleotide/amino acid polymorphisms and underscores its utility for detection of contemporary and emerging SARS-CoV-2 variants of concern.

Candida auris Pan-Drug-Resistant to Four Classes of Antifungal Agents.

Candida auris is an urgent antimicrobial resistance threat due to its global emergence, high mortality, and persistent transmissions. Nearly half of C. auris clinical and surveillance cases in the United States are from the New York and New Jersey Metropolitan area. We performed genome, and drug-resistance analysis of C. auris isolates from a patient who underwent multi-visceral transplantation. Whole-genome comparisons of 19 isolates, collected over 72 days, revealed closed similarity (Average Nucleotide Identity > 0.9996; Aligned Percentage > 0.9764) and a distinct subcluster of NY C. auris South Asia Clade I. All isolates had azole-linked resistance in ERG11(K143R) and CDR1(V704L). Echinocandin resistance first appeared with FKS1(S639Y) mutation and then a unique FKS1(F635C) mutation. Flucytosine-resistant isolates had mutations in FCY1, FUR1, and ADE17. Two pan-drug-resistant C. auris isolates had uracil phosphoribosyltransferase deletion (FUR1[1Δ33]) and the elimination of FUR1 expression, confirmed by a qPCR test developed in this study. Besides ERG11 mutations, four amphotericin B-resistant isolates showed no distinct nonsynonymous variants suggesting unknown genetic elements driving the resistance. Pan-drug-resistant C. auris isolates were not susceptible to two-drug antifungal combinations tested by checkerboard, Etest, and time-kill methods. The fungal population pattern, discerned from SNP phylogenetic analysis, was consistent with in-hospital or inpatient evolution of C. auris isolates circulating locally and not indicative of a recent introduction from elsewhere. The emergence of pan-drug-resistance to four major classes of antifungals in C. auris is alarming. Patients at high risk for drug-resistant C. auris might require novel therapeutic strategies and targeted pre-and/or posttransplant surveillance.

RT-PCR/MALDI-TOF Diagnostic Target Performance Reflects Circulating SARS-CoV-2 Variant Diversity in New York City.

As severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) continues to circulate, multiple variants of concern have emerged. New variants pose challenges for diagnostic platforms because sequence diversity can alter primer/probe-binding sites (PBSs), causing false-negative results. The MassARRAY SARS-CoV-2 Panel (Agena Bioscience) uses RT-PCR and mass spectrometry to detect five multiplex targets across N and ORF1ab genes. Herein, we use a data set of 256 SARS-CoV-2-positive specimens collected between April 11, 2021, and August 28, 2021, to evaluate target performance with paired sequencing data. During this time frame, two targets in the N gene (N2 and N3) were subject to the greatest sequence diversity. In specimens with N3 dropout, 69% harbored the Alpha-specific A28095U polymorphism that introduces a 3'-mismatch to the N3 forward PBS and increases risk of target dropout relative to specimens with 28095A (relative risk, 20.02; 95% CI, 11.36 to 35.72; P < 0.0001). Furthermore, among specimens with N2 dropout, 90% harbored the Delta-specific G28916U polymorphism that creates a 3'-mismatch to the N2 probe PBS and increases target dropout risk (relative risk, 11.92; 95% CI, 8.17 to 14.06; P < 0.0001). These findings highlight the robust capability of MassARRAY SARS-CoV-2 Panel target results to reveal circulating virus diversity, and they underscore the power of multitarget design to capture variants of concern.

Everybody nose: molecular and clinical characteristics of nasal colonization during active methicillin-resistant Staphylococcus aureus bloodstream infection.

BACKGROUND: Healthcare-associated infections pose a potentially fatal threat to patients worldwide and Staphylococcus aureus is one of the most common causes of healthcare-associated infections. S. aureus is a common commensal pathogen and a frequent cause of bacteremia, with studies demonstrating that nasal and blood isolates from single patients match more than 80% of the time. Here we report on a contemporary collection of colonizing isolates from those with methicillin-resistant S. aureus (MRSA) bloodstream infections to evaluate the diversity within hosts, and detail the clinical features associated with concomitant nasal colonization. METHODS: Swabs of the bilateral anterior nares were obtained from patients diagnosed with MRSA bacteremia. A single colony culture from the blood and an average of 6 colonies from the nares were evaluated for MRSA growth. For the nares cultures, we typed multiple isolates for staphylococcal protein A (spa) and derived the clonal complexes. Demographic and clinical data were obtained retrospectively from the electronic medical record system and analysed using univariate and multivariable regression models. RESULTS: Over an 11-month period, 68 patients were diagnosed with MRSA bloodstream infection, 53 were swabbed, and 37 (70%) were colonized with MRSA in the anterior nares. We performed molecular typing on 213 nasal colonies. Spa types and clonal complexes found in the blood were also detected in the nares in 95% of the cases. We also found that 11% of patients carried more than one clone of MRSA in the nares. Male sex and history of prior hospitalization within the past 90 days increased odds for MRSA colonization. CONCLUSION: The molecular epidemiological landscape of colonization in the setting of invasive disease is diverse and defining the interplay between colonization and invasive disease is critical to combating invasive MRSA disease.

Correlation between Identification of ß-Lactamase Resistance Genes and Antimicrobial Susceptibility Profiles in Gram-Negative Bacteria: a Laboratory Data Analysis.

We reported the frequency of resistance gene detection in Gram-negative blood culture isolates and correlated these findings with corresponding antibiograms. Data were obtained from 1045 isolates tested on the GenMark Dx ePlex Blood Culture Identification Gram-Negative Panels at the Mount Sinai Hospital Clinical Microbiology Laboratory in New York from March 2019 to February 2021. Susceptibilities were performed using Vitek 2 (bioMérieux Clinical Diagnostics) or Microscan (Beckman Coulter Inc.). blaCTX-M was detected in 26.4% Klebsiella pneumoniae, 23.5% Escherichia coli, and 16.4% Proteus mirabilis isolates. As would be expected, both blaCTX-M and blaCTX-M negative isolates were likely to be susceptible to newer agents while blaCTX-M positive isolates were more likely to be resistant to earlier generations of beta-lactam antibiotics. 3/204 blaCTX-M-positive isolates were found to be ceftriaxone-susceptible. Conversely, 2.8% ceftriaxone nonsusceptible strains were negative for all ß-lactamase genes on the ePlex BCID-GN panel, including blaCTX-M. The prevalence of CTX-M-producing Enterobacterales remains high in the United States. A small number of blaCTX-M-positive isolates were susceptible to ceftriaxone, and a small number of ceftriaxone nonsusceptible isolates were negative for blaCTX-M. Further studies are needed to determine the optimal management when an isolate is phenotypically susceptible to ceftriaxone, but blaCTX-M is detected. IMPORTANCE There is limited literature on corresponding results obtained from rapid molecular diagnostics with the antibiotic susceptibility profile. We reported a correlation between the results obtained from ePlex and the antibiograms against a large collection of Gram-negative bacteria. We reported that there can be a discrepancy in a small number of cases, but the clinical significance of that is unknown.

Creating Surveillance Data Infrastructure Using Laboratory Analytics: Leveraging Visiun and Epic Systems to Support COVID-19 Pandemic Response.

BACKGROUND: Pandemics are unpredictable and can rapidly spread. Proper planning and preparation for managing the impact of outbreaks is only achievable through continuous and systematic collection and analysis of health-related data. We describe our experience on how to comply with required reporting and develop a robust platform for surveillance data during an outbreak. MATERIALS AND METHODS: At Mount Sinai Health System, New York City, we applied Visiun, a laboratory analytics dashboard, to support main response activities. Epic System Inc.'s SlicerDicer application was used to develop clinical and research reports. We followed World Health Organization (WHO); federal and state guidelines; departmental policies; and expert consultation to create the framework. RESULTS: The developed dashboard integrated data from scattered sources are used to seamlessly distribute reports to key stakeholders. The main report categories included federal, state, laboratory, clinical, and research. The first two groups were created to meet government and state reporting requirements. The laboratory group was the most comprehensive category and included operational reports such as performance metrics, technician performance assessment, and analyzer metrics. The close monitoring of testing volumes and lab operational efficiency was essential to manage increasing demands and provide timely and accurate results. The clinical data reports were valuable for proper managing of medical surge requirements, such as healthcare workforce and medical supplies. The reports included in the research category were highly variable and depended on healthcare setting, research priorities, and available funding. We share a few examples of queries that were included in the designed framework for research projects. CONCLUSION: We reviewed here the key components of a conceptual surveillance framework required for a robust response to COVID-19 pandemics. We demonstrated leveraging a lab analytics dashboard, Visiun, combined with Epic reporting tools to function as a surveillance system. The framework could be used as a generic template for possible future outbreak events.

Robust clinical detection of SARS-CoV-2 variants by RT-PCR/MALDI-TOF multitarget approach.

The coronavirus disease 2019 (COVID-19) pandemic has sparked the rapid development of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) diagnostics. However, emerging variants pose the risk for target dropout and false-negative results secondary to primer/probe binding site (PBS) mismatches. The Agena MassARRAY® SARS-CoV-2 Panel combines reverse-transcription polymerase chain reaction and matrix-assisted laser desorption/ionization time-of-flight mass-spectrometry to probe for five targets across N and ORF1ab genes, which provides a robust platform to accommodate PBS mismatches in divergent viruses. Herein, we utilize a deidentified data set of 1262 SARS-CoV-2-positive specimens from Mount Sinai Health System (New York City) from December 2020 to April 2021 to evaluate target results and corresponding sequencing data. Overall, the level of PBS mismatches was greater in specimens with target dropout. Of specimens with N3 target dropout, 57% harbored an A28095T substitution that is highly specific for the Alpha (B.1.1.7) variant of concern. These data highlight the benefit of redundancy in target design and the potential for target performance to illuminate the dynamics of circulating SARS-CoV-2 variants.

Catheter-related bloodstream infection due to biofilm-producing Capnocytophaga sputigena.

Capnocytophaga sputigena is a facultatively-anaerobic bacterium that is part of the human oropharyngeal microflora. Although C. sputigena bacteremia is uncommon, systemic infections have been reported in both immunocompetent and immunocompromised patients. We report a case of catheter-related bloodstream infection by C. sputigena and highlight its enhanced biofilm-forming capacity in vitro.

Modified methicillin-resistant Staphylococcus aureus detected in neonatal intensive care patients.

OBJECTIVES: As part of an active MRSA surveillance programme in our neonatal ICU, we identified nares surveillance cultures from two infants that displayed heterogeneity in methicillin resistance between isolated subclones that lacked mecA and mecC. METHODS: The underlying mechanism for the modified Staphylococcus aureus (MODSA) methicillin-resistance phenotype was investigated by WGS. RESULTS: Comparison of finished-quality genomes of four MODSA and four MSSA subclones demonstrated that the resistance changes were associated with unique truncating mutations in the gene encoding the cyclic diadenosine monophosphate phosphodiesterase enzyme GdpP or a non-synonymous substitution in the gene encoding PBP2. CONCLUSIONS: These two cases highlight the difficulty in identifying non-mecA, non-mecC-mediated MRSA isolates in the clinical microbiology laboratory, which leads to difficulties in implementing appropriate therapy and infection control measures.