Lymphoma Driver Mutations in the Pathogenic Evolution of an Iconic Human Autoantibody.

Pathogenic autoantibodies arise in many autoimmune diseases, but it is not understood how the cells making them evade immune checkpoints. Here, single-cell multi-omics analysis demonstrates a shared mechanism with lymphoid malignancy in the formation of public rheumatoid factor autoantibodies responsible for mixed cryoglobulinemic vasculitis. By combining single-cell DNA and RNA sequencing with serum antibody peptide sequencing and antibody synthesis, rare circulating B lymphocytes making pathogenic autoantibodies were found to comprise clonal trees accumulating mutations. Lymphoma driver mutations in genes regulating B cell proliferation and V(D)J mutation (CARD11, TNFAIP3, CCND3, ID3, BTG2, and KLHL6) were present in rogue B cells producing the pathogenic autoantibody. Antibody V(D)J mutations conferred pathogenicity by causing the antigen-bound autoantibodies to undergo phase transition to insoluble aggregates at lower temperatures. These results reveal a pre-neoplastic stage in human lymphomagenesis and a cascade of somatic mutations leading to an iconic pathogenic autoantibody.

The CD8+ T cell tolerance checkpoint triggers a distinct differentiation state defined by protein translation defects.

Peripheral CD8+ T cell tolerance is a checkpoint in both autoimmune disease and anti-cancer immunity. Despite its importance, the relationship between tolerance-induced states and other CD8+ T cell differentiation states remains unclear. Using flow cytometric phenotyping, single-cell RNA sequencing (scRNA-seq), and chromatin accessibility profiling, we demonstrated that in vivo peripheral tolerance to a self-antigen triggered a fundamentally distinct differentiation state separate from exhaustion, memory, and functional effector cells but analogous to cells defectively primed against tumors. Tolerant cells diverged early and progressively from effector cells, adopting a transcriptionally and epigenetically distinct state within 60 h of antigen encounter. Breaching tolerance required the synergistic actions of strong T cell receptor (TCR) signaling and inflammation, which cooperatively induced gene modules that enhanced protein translation. Weak TCR signaling during bystander infection failed to breach tolerance due to the uncoupling of effector gene expression from protein translation. Thus, tolerance engages a distinct differentiation trajectory enforced by protein translation defects.

STAT3 gain-of-function mutations connect leukemia with autoimmune disease by pathological NKG2Dhi CD8+ T cell dysregulation and accumulation.

The association between cancer and autoimmune disease is unexplained, exemplified by T cell large granular lymphocytic leukemia (T-LGL) where gain-of-function (GOF) somatic STAT3 mutations correlate with co-existing autoimmunity. To investigate whether these mutations are the cause or consequence of CD8+ T cell clonal expansions and autoimmunity, we analyzed patients and mice with germline STAT3 GOF mutations. STAT3 GOF mutations drove the accumulation of effector CD8+ T cell clones highly expressing NKG2D, the receptor for stress-induced MHC-class-I-related molecules. This subset also expressed genes for granzymes, perforin, interferon-γ, and Ccl5/Rantes and required NKG2D and the IL-15/IL-2 receptor IL2RB for maximal accumulation. Leukocyte-restricted STAT3 GOF was sufficient and CD8+ T cells were essential for lethal pathology in mice. These results demonstrate that STAT3 GOF mutations cause effector CD8+ T cell oligoclonal accumulation and that these rogue cells contribute to autoimmune pathology, supporting the hypothesis that somatic mutations in leukemia/lymphoma driver genes contribute to autoimmune disease.

Characterisation and reproducibility of the HumanMethylationEPIC v2.0 BeadChip for DNA methylation profiling.

BACKGROUND: The Illumina family of Infinium Methylation BeadChip microarrays has been widely used over the last 15 years for genome-wide DNA methylation profiling, including large-scale and population-based studies, due to their ease of use and cost effectiveness. Succeeding the popular HumanMethylationEPIC BeadChip (EPICv1), the recently released Infinium MethylationEPIC v2.0 BeadChip (EPICv2) claims to extend genomic coverage to more than 935,000 CpG sites. Here, we comprehensively characterise the reproducibility, reliability and annotation of the EPICv2 array, based on bioinformatic analysis of both manifest data and new EPICv2 data from diverse biological samples. RESULTS: We find a high degree of reproducibility with EPICv1, evidenced by comparable sensitivity and precision from empirical cross-platform comparison incorporating whole genome bisulphite sequencing (WGBS), and high correlation between technical sample replicates, including between samples with DNA input levels below the manufacturer's recommendation. We provide a full assessment of probe content, evaluating genomic distribution and changes from previous array versions. We characterise EPICv2's new feature of replicated probes and provide recommendations as to the superior probes. In silico analysis of probe sequences demonstrates that probe cross-hybridisation remains a significant problem in EPICv2. By mapping the off-target sites at single nucleotide resolution and comparing with WGBS we show empirical evidence for preferential off-target binding. CONCLUSIONS: Overall, we find EPICv2 a worthy successor to the previous Infinium methylation microarrays, however some technical issues remain. To support optimal EPICv2 data analysis we provide an expanded version of the EPICv2 manifest to aid researchers in understanding probe design, data processing, choosing appropriate probes for analysis and for integration with methylation datasets from previous versions of the Infinium Methylation BeadChip.

Transient exposure to miR-203 enhances the differentiation capacity of established pluripotent stem cells.

Full differentiation potential along with self-renewal capacity is a major property of pluripotent stem cells (PSCs). However, the differentiation capacity frequently decreases during expansion of PSCs in vitro. We show here that transient exposure to a single microRNA, expressed at early stages during normal development, improves the differentiation capacity of already-established murine and human PSCs. Short exposure to miR-203 in PSCs (miPSCs) induces a transient expression of 2C markers that later results in expanded differentiation potency to multiple lineages, as well as improved efficiency in tetraploid complementation and human-mouse interspecies chimerism assays. Mechanistically, these effects are at least partially mediated by direct repression of de novo DNA methyltransferases Dnmt3a and Dnmt3b, leading to transient and reversible erasure of DNA methylation. These data support the use of transient exposure to miR-203 as a versatile method to reset the epigenetic memory in PSCs, and improve their effectiveness in regenerative medicine.

Isolation and characterisation of PR3-specific B cells and their immunoglobulin sequences.

BACKGROUND: PR3 autoantibodies are essential to the diagnosis and monitoring of granulomatosus with polyangiitis, but to date no PR3 autoantibody sequences have been published. OBJECTIVES: To identify and characterise PR3-specific B cells from the peripheral blood of patients with PR3 autoantibodies. METHODS: Peripheral blood mononuclear cells from seven patients with PR3 autoantibodies were stained with PR3. B cells that bound PR3 underwent single cell sorting, transcriptome sequencing, and their immunoglobulin sequences expressed as antibodies and tested for PR3-specificity by ELISA. RESULTS: We identified 19 PR3-specific B cells from only one PR3-seropositive patient at a low frequency (0.0075 % of B cells) in the peripheral blood. These were polyclonal, IgG+ and enriched for IgG4, lambda pairing, IGHJ6 gene usage, CDRH3 length, IGHE and CD71 expression. They demonstrated relatively low levels of somatic hypermutation and variably reduced PR3 binding when reverted to germline. CONCLUSIONS: Identifying PR3-specific B cells in the peripheral blood is possible but challenging and those we did identify exhibited features suggesting that PR3-self reactivity may occur early in B-cell development.

Calling differentially methylated regions from whole genome bisulphite sequencing with DMRcate.

Whole genome bisulphite sequencing (WGBS) permits the genome-wide study of single molecule methylation patterns. One of the key goals of mammalian cell-type identity studies, in both normal differentiation and disease, is to locate differential methylation patterns across the genome. We discuss the most desirable characteristics for DML (differentially methylated locus) and DMR (differentially methylated region) detection tools in a genome-wide context and choose a set of statistical methods that fully or partially satisfy these considerations to compare for benchmarking. Our data simulation strategy is both biologically informed-employing distribution parameters derived from large-scale consortium datasets-and thorough. We report DML detection ability with respect to coverage, group methylation difference, sample size, variability and covariate size, both marginally and jointly, and exhaustively with respect to parameter combination. We also benchmark these methods on FDR control and computational time. We use this result to backend and introduce an expanded version of DMRcate: an existing DMR detection tool for microarray data that we have extended to now call DMRs from WGBS data. We compare DMRcate to a set of alternative DMR callers using a similarly realistic simulation strategy. We find DMRcate and RADmeth are the best predictors of DMRs, and conclusively find DMRcate the fastest.

Evaluation of an Advanced Practice Provider Onboarding and Development Program.

OBJECTIVE: To evaluate the impact of implementing a comprehensive secondary onboarding and development program within a hospital medicine group at a large tertiary academic medical institution. METHODS: This was a mixed-methods study with physician and advanced practice providers (APPs) at an academic medical institution. RESULTS: For quantitative methods, improvement in competencies was determined using a pre-and posttest for APPs and pre- and postevaluation scores from collaborating physicians. APPs also participated in a pre- and post-self-assessment. For qualitative methods, experiences in the secondary onbo.arding and development program were assessed using semistructured interviews. CONCLUSIONS: For quantitative results, there were a total of 25 APPs who completed the pre- and posttest and were evaluated by at least 9 physicians. The average pretest score for APPs was 71.7% and the average posttest score was 83.0%. The average score for physicians' evaluations of APPs was 4.24/5 and increased to 4.38/5 in the postprogram evaluations. The average score for APP self-assessment pretraining was 3.52/5 and after the 12-month onboarding training, average scores increased to 3.84/5. For qualitative results, 4 APPs and 6 physicians were interviewed. Three of the APPs reported having more confidence in treating patients, whereas 1 APP viewed the program as a refresher course. All of the APPs mentioned that they would recommend the program to other APPs. Physicians reported that the program was beneficial in standardizing the care provided among the different types of APPs (physician assistants and nurse practitioners). All of the physicians also would recommend the program to other physicians and APPs.

STAT5B restrains human B-cell differentiation to maintain humoral immune homeostasis.

BACKGROUND: Lymphocyte differentiation is regulated by coordinated actions of cytokines and signaling pathways. IL-21 activates STAT1, STAT3, and STAT5 and is fundamental for the differentiation of human B cells into memory cells and antibody-secreting cells. While STAT1 is largely nonessential and STAT3 is critical for this process, the role of STAT5 is unknown. OBJECTIVES: This study sought to delineate unique roles of STAT5 in activation and differentiation of human naive and memory B cells. METHODS: STAT activation was assessed by phospho-flow cytometry cell sorting. Differential gene expression was determined by RNA-sequencing and quantitative PCR. The requirement for STAT5B in B-cell and CD4+ T-cell differentiation was assessed using CRISPR-mediated STAT5B deletion from B-cell lines and investigating primary lymphocytes from individuals with germline STAT5B mutations. RESULTS: IL-21 activated STAT5 and strongly induced SOCS3 in human naive, but not memory, B cells. Deletion of STAT5B in B-cell lines diminished IL-21-mediated SOCS3 induction. PBMCs from STAT5B-null individuals contained expanded populations of immunoglobulin class-switched B cells, CD21loTbet+ B cells, and follicular T helper cells. IL-21 induced greater differentiation of STAT5B-deficient B cells into plasmablasts in vitro than B cells from healthy donors, correlating with higher expression levels of transcription factors promoting plasma cell formation. CONCLUSIONS: These findings reveal novel roles for STAT5B in regulating IL-21-induced human B-cell differentiation. This is achieved by inducing SOCS3 to attenuate IL-21 signaling, and BCL6 to repress class switching and plasma cell generation. Thus, STAT5B is critical for restraining IL-21-mediated B-cell differentiation. These findings provide insights into mechanisms underpinning B-cell responses during primary and subsequent antigen encounter and explain autoimmunity and dysfunctional humoral immunity in STAT5B deficiency.

Student, Faculty, and Coach Perspectives on Why Athletes Excel in Medical School: A Qualitative Analysis.

Phenomenon Medical schools are tasked with selecting applicants who will excel in a rigorous curriculum and successfully perform as future physicians. While many studies have assessed quantitative prematriculation data for predicting success in medical school, fewer studies have assessed for qualitative prematriculation factors influencing medical school performance. A recent study revealed that medical students with at least one year of varsity level college athletics participation outperformed their peers on United States Medical Licensing board exams and clinical clerkships. The current study sought to explore medical student, medical school faculty, and college coach perspectives about factors explaining why medical students with collegiate athletic experience succeed in medical school. Approach: In 2019, the authors conducted semi-structured interviews with medical students with collegiate athletic experience, medical school faculty with experience educating student athletes, and college coaches with experience training student athletes who matriculated into medical school. The interview transcripts were systematically coded and analyzed for themes using a grounded theory approach. Participants were recruited and interviewed until saturation of data was reached. Findings: Fifteen medical students with collegiate athletic experience, five medical school faculty, and three collegiate coaches participated in the study. Six themes were identified as important factors explaining the academic success of these students in medical school and each of these themes appeared in student, faculty, and coach interviews: goal setting, goal pursuit, and performance appraisal; development of time management, planning, and organizational skills; development of team values and teamwork skills; development of communication and interpersonal skills; acceptance of, coping strategies for, and resilient response to stress and adversity; and prioritization of personal wellness. Participants described meaningful connections between these attributes and skills, suggesting the students' development, transfer, and application of them is interrelated. Insights: In this study, academic success of medical students with collegiate athletic experience was attributed to specific skills and attributes developed during college. The grounded theory life skills transfer model can explain transfer of these attributes and skills from college to the medical school setting. Theoretical frameworks and empirical study findings from the sociology, educational psychology, sports psychology, and medical education literature provide helpful lenses for understanding why these skills and attributes confer success among student athletes in medical school. These findings offer important insights on skill development that may support the academic success of all medical students.

Enduring epigenetic landmarks define the cancer microenvironment.

The growth and progression of solid tumors involves dynamic cross-talk between cancer epithelium and the surrounding microenvironment. To date, molecular profiling has largely been restricted to the epithelial component of tumors; therefore, features underpinning the persistent protumorigenic phenotype of the tumor microenvironment are unknown. Using whole-genome bisulfite sequencing, we show for the first time that cancer-associated fibroblasts (CAFs) from localized prostate cancer display remarkably distinct and enduring genome-wide changes in DNA methylation, significantly at enhancers and promoters, compared to nonmalignant prostate fibroblasts (NPFs). Differentially methylated regions associated with changes in gene expression have cancer-related functions and accurately distinguish CAFs from NPFs. Remarkably, a subset of changes is shared with prostate cancer epithelial cells, revealing the new concept of tumor-specific epigenome modifications in the tumor and its microenvironment. The distinct methylome of CAFs provides a novel epigenetic hallmark of the cancer microenvironment and promises new biomarkers to improve interpretation of diagnostic samples.

Evaluation of cross-platform and interlaboratory concordance via consensus modelling of genomic measurements.

MOTIVATION: A synoptic view of the human genome benefits chiefly from the application of nucleic acid sequencing and microarray technologies. These platforms allow interrogation of patterns such as gene expression and DNA methylation at the vast majority of canonical loci, allowing granular insights and opportunities for validation of original findings. However, problems arise when validating against a "gold standard" measurement, since this immediately biases all subsequent measurements towards that particular technology or protocol. Since all genomic measurements are estimates, in the absence of a "gold standard" we instead empirically assess the measurement precision and sensitivity of a large suite of genomic technologies via a consensus modelling method called the row-linear model. This method is an application of the American Society for Testing and Materials Standard E691 for assessing interlaboratory precision and sources of variability across multiple testing sites. Both cross-platform and cross-locus comparisons can be made across all common loci, allowing identification of technology- and locus-specific tendencies. RESULTS: We assess technologies including the Infinium MethylationEPIC BeadChip, whole genome bisulfite sequencing (WGBS), two different RNA-Seq protocols (PolyA+ and Ribo-Zero) and five different gene expression array platforms. Each technology thus is characterised herein, relative to the consensus. We showcase a number of applications of the row-linear model, including correlation with known interfering traits. We demonstrate a clear effect of cross-hybridisation on the sensitivity of Infinium methylation arrays. Additionally, we perform a true interlaboratory test on a set of samples interrogated on the same platform across twenty-one separate testing laboratories. AVAILABILITY AND IMPLEMENTATION: A full implementation of the row-linear model, plus extra functions for visualisation, are found in the R package consensus at https://github.com/timpeters82/consensus. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

Development and implementation of a COVID-19 near real-time traffic light system in an acute hospital setting.

Common causes of death in COVID-19 due to SARS-CoV-2 include thromboembolic disease, cytokine storm and adult respiratory distress syndrome (ARDS). Our aim was to develop a system for early detection of disease pattern in the emergency department (ED) that would enhance opportunities for personalised accelerated care to prevent disease progression. A single Trust's COVID-19 response control command was established, and a reporting team with bioinformaticians was deployed to develop a real-time traffic light system to support clinical and operational teams. An attempt was made to identify predictive elements for thromboembolism, cytokine storm and ARDS based on physiological measurements and blood tests, and to communicate to clinicians managing the patient, initially via single consultants. The input variables were age, sex, and first recorded blood pressure, respiratory rate, temperature, heart rate, indices of oxygenation and C-reactive protein. Early admissions were used to refine the predictors used in the traffic lights. Of 923 consecutive patients who tested COVID-19 positive, 592 (64%) flagged at risk for thromboembolism, 241/923 (26%) for cytokine storm and 361/923 (39%) for ARDS. Thromboembolism and cytokine storm flags were met in the ED for 342 (37.1%) patients. Of the 318 (34.5%) patients receiving thromboembolism flags, 49 (5.3% of all patients) were for suspected thromboembolism, 103 (11.1%) were high-risk and 166 (18.0%) were medium-risk. Of the 89 (9.6%) who received a cytokine storm flag from the ED, 18 (2.0% of all patients) were for suspected cytokine storm, 13 (1.4%) were high-risk and 58 (6.3%) were medium-risk. Males were more likely to receive a specific traffic light flag. In conclusion, ED predictors were used to identify high proportions of COVID-19 admissions at risk of clinical deterioration due to severity of disease, enabling accelerated care targeted to those more likely to benefit. Larger prospective studies are encouraged.

Inbreeding in the last ruling dynasty of Portugal: The house of Braganza.

OBJECTIVES: To determine whether: (1) there are high levels of inbreeding in a European royal dynasty that continues until the 20th century, and (2) whether inbreeding is negatively associated with pre-reproductive survival and longevity. METHODS: Genealogical information of all Braganza monarchs (1640-1910) was used to compute the individual's inbreeding coefficient (F) and the coefficient of kinship (θ) of the marriage which were examined in relation to two life-history traits. RESULTS: Mean F of the monarchs was 0.0530, close to that corresponding to the progeny of first cousins (F = 0.0625). A statistically significant effect of maternal inbreeding on offspring longevity (P = 0.037) and a significant effect of individual's F on survival from birth to 10 years (P = 0.023) were detected. CONCLUSIONS: Another European royal dynasty besides the Habsburgs had high levels of inbreeding, especially high during a period after the early modern age. The lineage showed evidence of inbreeding depression. Results support the hypothesis that an increase in maternal homozygosity affects the lifespan of the progeny.

2015-2016 Vaccine Effectiveness of Live Attenuated and Inactivated Influenza Vaccines in Children in the United States.

Background: In the 2015-2016 season, quadrivalent live attenuated influenza vaccine (LAIV) and both trivalent and quadrivalent inactivated influenza vaccine (IIV) were available in the United States. Methods: This study, conducted according to a test-negative case-control design, enrolled children aged 2-17 years presenting to outpatient settings with fever and respiratory symptoms for <5 days at 8 sites across the United States between 30 November 2015 and 15 April 2016. A nasal swab was obtained for reverse-transcriptase polymerase chain reaction (RT-PCR) testing for influenza, and influenza vaccination was verified in the medical record or vaccine registry. Influenza vaccine effectiveness (VE) was estimated using a logistic regression model. Results: Of 1012 children retained for analysis, most children (59%) were unvaccinated, 10% received LAIV, and 31% received IIV. Influenza A (predominantly antigenically similar to the A/California/7/2009 strain) was detected in 14% and influenza B (predominantly a B/Victoria lineage) in 10%. For all influenza, VE was 46% (95% confidence interval [CI], 7%-69%) for LAIV and 65% (48%-76%) for IIV. VE against influenza A(H1N1)pdm09 was 50% (95% CI, -2% to 75%) for LAIV and 71% (51%-82%) for IIV. The odds ratio for vaccine failure with RT-PCR-confirmed A(H1N1)pdm09 was 1.71 (95% CI, 0.78-3.73) in LAIV versus IIV recipients. Conclusions: LAIV and IIV demonstrated effectiveness against any influenza among children aged 2-17 years in 2015-2016. When compared to all unvaccinated children, VE against influenza A(H1N1)pdm09 was significant for IIV but not LAIV. Clinical Trials Registration: NCT01997450.

Is weight associated with severity of acute respiratory illness?

BACKGROUND/OBJECTIVES: Obesity was an independent risk factor for severe disease in hospitalized adults during the 2009 pandemic H1N1 influenza season. Few studies have investigated the association between weight and severity of acute respiratory illnesses in children or in adults seeking care in the emergency department (ED) during other winter respiratory seasons. SUBJECTS/METHODS: We prospectively and systematically enrolled patients ≥2 years of age who presented to the ED or inpatient setting in a single geographic region with fever/acute respiratory illness over four consecutive winter respiratory seasons (2010-2014). We collected demography, height and weight, and high risk co-morbid conditions. Multivariable logistic regression was used for prediction of hospital admission (primary outcome), length of stay and supplemental oxygen requirement among those hospitalized, and antibiotic prescription (secondary outcomes). RESULTS: We enrolled 3560 patients (N = 749 children, 2811 adults), 1405 (39%) with normal weight, 860 (24%) with overweight, and 1295 (36%) with obesity. Following multivariable logistic regression, very young or very old age (p < 0.001) and high-risk conditions (p < 0.001) predicted hospitalization. Risk of hospitalization was decreased for adults with overweight [aOR 0.8 (95% CI 0.6-1.0)], class 1 obesity [aOR 0.7 (95% CI 0.5-1.0)], and class 2 obesity [aOR 0.6 (95% CI 0.4-0.8)] compared to normal-weight. Class 3 obesity was associated with supplemental oxygen requirement in adults [aOR 1.6 (95% CI 1.1-2.5)]. No association was seen in children. CONCLUSION: Overweight and obesity were not associated with increased risk of hospitalization during winter respiratory seasons in children or adults.

Widespread promoter methylation of synaptic plasticity genes in long-term potentiation in the adult brain in vivo.

BACKGROUND: DNA methylation is a key modulator of gene expression in mammalian development and cellular differentiation, including neurons. To date, the role of DNA modifications in long-term potentiation (LTP) has not been explored. RESULTS: To investigate the occurrence of DNA methylation changes in LTP, we undertook the first detailed study to describe the methylation status of all known LTP-associated genes during LTP induction in the dentate gyrus of live rats. Using a methylated DNA immunoprecipitation (MeDIP)-array, together with previously published matched RNA-seq and public histone modification data, we discover widespread changes in methylation status of LTP-genes. We further show that the expression of many LTP-genes is correlated with their methylation status. We show that these correlated genes are enriched for RNA-processing, active histone marks, and specific transcription factors. These data reveal that the synaptic activity-evoked methylation changes correlates with pre-existing activation of the chromatin landscape. Finally, we show that methylation of Brain-derived neurotrophic factor (Bdnf) CpG-islands correlates with isoform switching from transcripts containing exon IV to exon I. CONCLUSIONS: Together, these data provide the first evidence of widespread regulation of methylation status in LTP-associated genes.

'Details on the Establishment of Doctor Willis, for the Cure of Lunatics' (1796).

The 'mad-doctor' Dr Francis Willis achieved national and international celebrity following his successful treatment of King George III's first major episode of insanity in 1788-9. At the time of his summons to attend the King, Willis was a well-established provincial practitioner and madhouse proprietor. An anonymous French visitor published a description of Willis's Lincolnshire madhouse and his therapeutic practices in 1796. The translated text of the full article provides a unique insight into the work of a key figure in the historical development of psychological medicine. The accompanying Introduction summarizes Francis Willis's career as a mad-doctor and uses salient information from the original text to place him and his madhouse practice within a contemporary context.

Estimating the Burden of Pandemic Infectious Disease: The Case of the Second Wave of Pandemic Influenza H1N1 in Forsyth County, North Carolina.

BACKGROUND: Understanding the burden of influenza A(H1N1)pdm09 virus during the second wave of 2009-2010 is important for future pandemic planning. METHODS: Persons who presented to the emergency department (ED) or were hospitalized with fever and/or acute respiratory symptoms at the academic medical center in Forsyth County, North Carolina were prospectively enrolled and underwent nasal/throat swab testing for influenza A(H1N1)pdm09. Laboratory-confirmed cases of influenza A(H1N1)pdm09 virus identified through active surveillance were compared by capture-recapture analysis to those identified through independent, passive surveillance (physician-ordered influenza testing). This approach estimated the number of total cases, including those not captured by either surveillance method. A second analysis estimated the total number of influenza A(H1N1)pdm09 cases by multiplying weekly influenza percentages determined via active surveillance by weekly counts of influenza-associated discharge diagnoses from administrative data. Market share adjustments were used to estimate influenza A(H1N1)pdm09 virus ED visits or hospitalizations per 1,000 residents. RESULTS: Capture-recapture analysis estimated that 753 residents (95% confidence interval [CI], 424-2,735) with influenza A(H1N1)pdm09 virus were seen in the academic medical center from September 2009 through mid-April 2010; this result yielded an estimated 4.7 (95% CI, 2.6-16.9) influenza A(H1N1)pdm09 virus ED visits or hospitalizations per 1,000 residents. Similarly, 708 visits were estimated using weekly influenza percentages and influenza-associated discharge diagnoses, yielding an estimated 4.4 influenza A(H1N1)pdm09 virus ED visits or hospitalizations per 1,000 residents. CONCLUSION: This study demonstrates that the burden of influenza A(H1N1)pdm09 virus in ED and inpatient settings by capture-recapture analysis was 4-5 per 1,000 residents; this rate was approximately 8-fold higher than that detected by physician-ordered influenza testing.

Predictors of Influenza Diagnosis Among Patients With Laboratory-Confirmed Influenza.

OBJECTIVE: This study was performed to determine predictors of clinical influenza diagnosis among patients with laboratory-confirmed influenza. METHODS: Prospective, laboratory-confirmed surveillance for influenza was conducted among patients of all ages who were hospitalized or presented to the emergency department with fever and respiratory symptoms during 2009-2013. We evaluated all enrolled persons who had influenza confirmed by viral culture and/or polymerase chain reaction and received any discharge diagnosis. The primary outcome, clinical influenza diagnosis, was defined as (1) a discharge diagnosis of influenza, (2) a prescription of neuraminidase inhibitor, or (3) a rapid test positive for influenza virus. Bivariate analyses and multiple logistic regression modeling were performed. RESULTS: Influenza was diagnosed for 29% of 504 enrolled patients with laboratory-confirmed influenza and for 56% of 236 patients with high-risk conditions. Overall, clinical influenza diagnosis was predicted by race/ethnicity, insurance status, year, being hospitalized, having high-risk conditions, and receiving no diagnosis of bacterial infection. Being diagnosed with a bacterial infection reduced the odds of receiving an influenza diagnosis by >3-fold for all patients and for patients with high-risk conditions. CONCLUSIONS: Many influenza virus-positive patients, including those with high-risk conditions, do not receive a clinical diagnosis of influenza. The pattern of clinical diagnoses among influenza virus-positive patients suggests preferential consideration of bacterial diseases as a diagnosis.