bluuebunny/crossref_metadata_2025_split huggingface dataset:
Subclonal Populations Detected by Flow Cytometry in Chronic Lymphocytic Leukemia
Authors: Yip James , Simonson Paul 
Published: October 2024

Abstract

Introduction
Chronic lymphocytic leukemia/small lymphocytic lymphoma (CLL/SLL) is a mature, small B-cell lymphoma that typically co-expresses CD5 and CD23. Molecular-targeted therapies have produced remarkable therapeutic benefits in CLL, yet many patients are resistant to treatment. This study aims to identify subpopulations in CLL by conventional flow cytometry and clustering analysis.

Methods
One-hundred-and-forty-one CLL cases were taken from a public database and analyzed by uniform manifold approximation and projection (UMAP). Differences in size, granularity, and intensities of CD19, CD20, CD79a, CD5, CD23, CD10, and FMC7 were plotted with BD FACSDiva Software.
Eighteen cases with a new diagnosis of CLL at our institution between January 1, 2021 to December 31, 2022 were analyzed. Only peripheral blood samples were included. Markers including forward scatter, side scatter, and fluorescent intensities of CD19, CD20, CD5, CD23, CD10, surface kappa, and surface lambda were used to identify discrete CLL clusters in UMAP plots.

Results: Public Dataset
Clustering analysis of conventional flow cytometry data from 141 cases from a public database reveals discrete populations of cells exhibiting varying sizes, granularity and fluorescent intensities. Among the 141 cases, 33 (23.4%) had 1 cluster, 64 (45.4%) with 2 clusters, 42 (29.8%) with 3 clusters and 2 (1.4%) with 4 clusters. Among the cases with 2 clusters, a smaller population of cells deviated from the main neoplastic population. This smaller population had a higher forward and side scatter, and brighter intensities in CD45 and CD19, consistent with prolymphocytes. Among the cases with 3 clusters, a third population recurrently had a smaller size by forward scatter. A few cases additionally had varying intensities by CD23 and CD19. Among the 2 cases with 4 clusters, only slight deviations by UMAP were seen and may be artifactual.

Institutional Dataset
Clustering analysis of 18 cases at our institution revealed complex patterns of cell clusters exhibiting varying sizes, granularity and surface receptor expression intensity. Among the 18 cases, 1 (5.6%) had 1 cluster, 1 (5.6%) with 2 clusters, 10 (55.6%) with 3 clusters, 3 (16.7%) with 4 clusters, and 3 (16.7%) with 5 clusters. Similar to the public dataset, the third population, in addition to the predominant population and prolymphocytes, consistently varied by lower forward scatter. Additional clusters, such as a population with varying CD23 signal intensities, were identified in some cases.

Conclusions
In our study, multiple clonal subsets in CLL/SLL are detected by conventional flow cytometry analysis. These subclonal populations may result in therapy resistance for these patients, and identifying them by flow cytometry analysis may provide additional prognostic information in the clinical setting. Furthermore, the ability to detect these subclonal populations may allow for isolation of specific clones for future investigation in research settings.
Clinical Utility of Metagenomic Next-Generation Sequencing of Plasma Microbial Cell-Free DNA: a Single-Center Retrospective Study
Authors: Almeida Guilherme Gomide , Dumm Rebekah 
Published: October 2024

Abstract
Infectious diseases account for significant morbidity and mortality in healthcare, and metagenomic next-generation sequencing of plasma microbial cell-free DNA has emerged as a promising non-invasive diagnostic tool in this scenario. However, the available data on the real impact of this test in clinical practice is limited and conflicting. In this context, we performed a retrospective evaluation of the utilization of this test at our institution since February 17, 2019. A total of 76 tests were performed in this period in 68 different patients. The mean age of the patients was 9 years old, with 40 being male (58.82%) and 28 female (41.18%). The ordering specialties included infectious diseases (82.90%), hematology/oncology (9.21%), neurology (3.95%), pulmonology (2.63%), and dermatology (1.31%). The vast majority of the tests were ordered as inpatient (73 tests, 96.05%). 45 tests (59.21%) resulted as positive, of which 19 (42.22%) were polymicrobial. The tests were concordant with conventional microbiological studies 47 times (61.84%). Most tests (67 tests, 88.16%) had no significant clinical impact, 8 tests (10.53%) had a positive impact, and one test (1.31%) had an indeterminate impact. No tests were determined to have a negative clinical impact. Overall, our data is consistent with most studies which assessed the clinical utility of metagenomic next-generation sequencing of plasma microbial cell-free DNA in the literature, showing that testing did not have a significant impact on patient care for the majority of cases. There were some cases, however, in which testing showed a positive impact in escalating or de-escalating antibiotic therapy. In conclusion, despite being mostly a bystander, in selected cases and in the right clinical context, the test can be useful to guide antimicrobial treatment without causing significant harm. More studies are needed to further investigate the real utility of the test.
Development of a Novel and Rapid LC-MS/MS Method for Measuring Phosphatidylethanol in Whole Blood
Authors: Nielsen Jacob , Yang Qingwen , Lahorewala Sarrah , Yi Xin 
Published: October 2024

Abstract
Alcohol consumption can lead to serious health repercussions, including motor impairments and organ failure. Testing for short-term alcohol use is routinely conducted using blood ethanol tests in clinical laboratories. However, there is an emergent need to identify heavy alcohol users, especially crucial for patients undergoing organ transplant evaluations. With the advent of various biomarkers to detect heavy alcohol use, phosphatidyl ethanol (PEth), a group of abnormal phospholipids formed in cell membranes due to alcohol consumption, has gained prominence. PEth has been analyzed in whole blood with liquid chromatography-tandem mass spectrometry (LC-MS/MS) as two isomers: 18:1 (fatty acid:double bond) and 18:2 (fatty acid:double bond). We have developed an LC-MS/MS method that can detect these isomers in 4.5 minutes at concentrations of 10-500 ng/mL with excellent linearity and precision. In contrast to most LC-MS/MS methods where phospholipids are removed to minimize ion suppression, this method requires retaining phospholipids because the analyte, PEth, is a type of phospholipid. This was achieved through a ‘reverse extraction’ approach using phospholipid removal cartridges. Subsequently, the extracted samples were separated in an Agilent Protoshell 120 HPLC column on a Vanquish UHPLC instrument using 1 mM ammonium formate with 0.2% formic acid in water as Solution A and 50/50 isopropanol/acetonitrile as Solution B. The separation is performed starting at 80% Solution B (rest is Solution A) followed by a gradient elution where Solution B was increased to 95% over 2.3 minutes. Ions were analyzed using a ThermoFisher TSQ Endura MS/MS instrument in the negative mode. Additionally, commercially available whole blood was found to contain detectable levels of PEth in random lots, rendering them unsuitable for use as blank blood in the preparation of calibrators and quality controls. Instead, synthetic was used for this purpose. Ion suppression was observed, and various strategies were attempted to minimize it, including adjusting additives in the mobile phase, extending LC separation times, among others. More details on the impact of these adjustments and their effectiveness in improving the method’s performance will be presented.
Changes in platelet transfusion thresholds utilized in the NICU since PlaNeT-2
Authors: Hasan Rida , Upadhyay Kirtikumar , Breit Nathan , Tsang Hamilton , Hess John , Asif Maryam , Pagano Monica 
Published: October 2024

Abstract

Background
Use of prophylactic platelet transfusions in the neonatal setting is a common practice due to concerns about bleeding despite evidence of a developmental mismatch with transfusion of adult platelets. The Platelets for Neonatal Transfusion-Study 2 (PlaNeT-2) published in January 2019 showed that using higher platelet thresholds (50 x 109/L) was associated with more adverse outcomes and no improvement in bleeding events compared to lower platelet thresholds (25 x 109/L). Following this trial, implementation of a lower threshold was provider dependent as there were no changes made to the neonatal intensive care unit (NICU) institutional policies. Our goal was to determine if there were any overall changes in individual clinical practice with thresholds used for platelet transfusions in our NICU and number of transfusions per patient.

Methods
Our center has a 52 bed level IV NICU that specializes in care for critical and/or preterm infants. We performed a retrospective review from 2017 to 2023 of patients admitted in the NICU who received platelet transfusion and had an available platelet count in the 12 hours prior to the transfusion.

Results
There were 272 platelet transfusions given to 107 unique NICU patients who had platelet counts available. There were 66 transfusions given to 32 patients pre-PlaNeT-2 (2017-2018) and 206 transfusions given to 75 patients post-PlaNeT-2 (2019-2023). The average platelet count pre-PLaNeT-2(2017 to 2018) was 43 x 109/L (SD±24 x 109/L) and post-PLaNeT-2 (2019 to 2023) was 36 x 109/L (SD±21 x 109/L, p=0.03). Prior to PLaNeT-2, 26% of platelet transfusions were for platelet counts ≤ 25 x 109/L and 35% were for platelet counts between 51 to 100x9/L. Post PLaNeT-2 the percentages changed to 35% and 21% respectively. The percentage of transfusions occurring for platelet counts between 26 to ≤50 x 109/L was 38% and 44%, respectively.
The number of platelet transfusions per year did not decrease significantly after the PLaNeT-2 trial (average of 33/year versus 41/year, p=0.5). The number of transfusions per patient before and after PLaNeT-2 did not change (2.1 transfusions per patient versus 2.6 transfusions per patient, p=0.3).

Conclusion
Platelet thresholds utilized in our center decreased after the PLaNeT-2 trial with less transfusions occurring for platelet counts greater than 50 x 109/L. Although thresholds decreased, the number of transfusions per year and the number of transfusions per patient did not significantly change. Further studies are needed to understand changes in patient outcomes and platelet utilization in the NICU.
Performance of Gram Stain Characteristics for Predicting Culture Growth in Lower Respiratory Specimens Across Multiple Institutions
Authors: Spies Nicholas , Chamberland Robin , Rhoads Daniel , Wallace Meghan , Bussian David , Buchan Blake , Burnham Carey-Ann , Yarbrough Melanie 
Published: October 2024

Abstract
Data from Gram stains performed on clinical specimens can guide clinical decision-making and downstream laboratory workflows. However, the extent to which Gram stain results predict gold-standard culture results, and the generalizability of this performance across sites, warrants additional exploration. We sought to discover which Gram stain features would be associated with clinically significant culture growth (“growth”). To do so, 574 remnant specimens from the lower respiratory tract collected from four institutions were analyzed. A standardized Gram stain and culture protocol was performed across all study sites. Final reference organism identification was assigned using MALDI-ToF MS, and clinically significant growth was defined as any organism reported to the medical record. Gram stains with mixed microorganisms detected were considered concordant if ≥2 organisms of different Gram stain appearances were identified in culture. Growth was observed in 53% of specimens, including 25% of specimens in which Gram stain demonstrated no organisms. The 95% confidence intervals (CI) of the site-wise growth rates were 30-42%, 42-52%, 51-67%, and 60-77% (p &lt; 0.001 by Chi-squared). By Gram stain morphology, 53-69% of specimens with Gram-positive cocci grew any clinically significant bacteria, while 31-49% grew a concordant Gram-positive cocci. 65-85% of specimens with Gram-negative bacilli grew any clinically significant bacteria, while 59-74% grew a concordant Gram-negative bacilli. 63-83% of specimens with mixed microorganisms grew 2 or more clinically relevant bacteria of differing Gram stain morphologies. Specimens from bronchoalveolar lavages (BALs) were less likely to have growth (30-42%) than endotracheal aspirates (66-77%). Polymorphonuclear leukocyte (PMNs) abundance was associated with culture growth, with Gram stains showing no, rare, few, moderate, or abundant PMNs demonstrating growth 28, 44, 52, 60, and 67% of the time (p &lt; 0.001). Specimens with None and few squamous epithelial cells on Gram stain demonstrated growth 38-52% and 59-68% of the time (p &lt; 0.001). No significant difference in growth was observed for red blood cells. Organism abundance on Gram stain was significantly associated with culture growth, with growth rate CIs being 20-32% for no organisms, 44-62% for rare, 58-77% for few, 70-93% for moderate, and 81-95% for abundant organisms seen. The extent to which these associations could guide the development of decision rules or machine learning models that demonstrate acceptable negative predictive value for pathogen growth to alleviate the operational burden of low-utility cultures is an exciting future direction to explore.
Concordance Between Platelet Expression Assay (PEA) and Serotonin Release Assay (SRA) in Heparin-Induced Thrombocytopenia: Implications for Diagnostic Accuracy and Laboratory Test Utilization
Authors: Grewal Sarah K , Hanson Laura , Choe Yeon-Whan , Barrette Eileen , Lee Kathi , Maier Cheryl 
Published: October 2024

Abstract

Introduction
The Serotonin Release Assay (SRA) serves as the gold standard functional assay for heparin-induced thrombocytopenia (HIT) diagnosis at our institution. The present turnaround time (TAT) for SRA results ranges from 4-5 days, occasionally extending beyond 8 days, given outsourcing to a reference laboratory. Recently, an alternative functional assay, the Platelet Expression Assay (PEA), has been introduced. The PEA detects platelet activation through p-selectin expression by flow cytometry, with a reported sensitivity of 100% and specificity of 95% for HIT. The PEA is also a send-out test, and thus has similar TAT and cost profiles as the SRA, but with potential to transition into an in-house assay in the future. Herein, we sought to determine the concordance of the PEA and SRA for the detection and diagnosis of HIT within our patient population.

Methods
A retrospective review was conducted on all cases in which a HIT ELISA (screening immunoassay) was performed by the Special Coagulation Laboratory during a one-year period. A 4Ts score was calculated if not already documented by chart review. In a subset analysis, specimens with a positive screening ELISA were concurrently tested by both SRA and PEA to compare performance.

Results
Of the 1341 cases, a 4Ts score was calculated and found to be less than 4 for 30-50% of cases. A total of 490 cases were sent out for SRA, of which 49 (10%) returned positive results. In a subset analysis of 20 samples from 14 patients, all with positive ELISAs, results revealed concordance between the SRA and PEA for all but one sample (19/20, 95%). Specifically, three successive samples from one patient had [negative, negative, positive] results for SRA, and [negative, positive, positive] results for PEA, respectively. This discordant result suggests a potential difference in sensitivity between the two assays. Further subset analysis focused on an additional 13 samples from 10 patients, all with SRA-positive results. Only 8 samples were concordant (8/13, 62%). One case showed a negative PEA and presumed false positive SRA, given the clinical history, while 4 samples showed likely true positive SRA results, given strength of ELISA results and 4Ts score, but were negative by PEA.

Conclusions
The lack of adherence to the 4Ts score algorithm to drive laboratory test utilization resulted in unnecessary ELISA and functional assay testing, potentially resulting in increased costs and false positive results. This result underscores the need for vigilance in assessing and refining diagnostic strategies. Additionally, our discordant findings in the subset analysis underscore the importance of evaluating the concordance and discordance between SRA and PEA results, providing insights into the diagnostic accuracy and potential limitations of each assay in identifying HIT.
A personalized and systematically designed adherence intervention improves photoprotection in adults with xeroderma pigmentosum (XP): results of the XPAND randomized controlled trial
Authors: Walburn Jessica , Norton Sam , Sarkany Robert , Canfield Martha , Sainsbury Kirby , McCrone Paul , Araújo-Soares Vera , Morgan Myfanwy , Boadu Janette , Foster Lesley , Heydenreich Jakob , Mander Adrian P , Sniehotta Falko F , Wulf Hans Christian , Weinman John 
Published: October 2024

Abstract

Background
Poor adherence to photoprotection in xeroderma pigmentosum (XP) increases morbidity and shortens lifespan due to skin cancers.

Objectives
To test a highly personalized intervention (XPAND) to reduce the dose of ultraviolet radiation (UVR) reaching the face in adults with XP, designed using known psychosocial determinants of poor photoprotection.

Methods
A two-arm parallel group randomized controlled trial, including patients with suboptimal photoprotection to receive XPAND or a delayed-intervention control arm that received XPAND the following year. XPAND comprises seven 1 : 1 sessions targeting photoprotection barriers (e.g. misconceptions about UVR) supported by personalized text messages, activity sheets and educational materials incorporating behaviour change techniques. The primary outcome, mean daily UVR dose to face across 21 days in June–July 2018, was calculated by combining UVR exposure at the wrist with a face photoprotection activity diary. Secondary outcomes were UVR dose to face across 21 days in August 2018, time spent outside, photoprotective measures used outside, mood, automaticity and confidence to photoprotect. Financial costs and quality-adjusted life years (QALYs) were calculated.

Results
Sixteen patients were randomized; 13 provided sufficient data for primary outcome analysis. The XPAND group (n = 8) had lower mean daily UVR dose to face [0.03 standard error of difference (SED) (SD 0.02)] compared with controls (n = 7) [0.43 SED (SD 0.17)] (adjusted difference = –0.25, P &lt; 0.001, Hedge’s g = 2.21) at the June 2018 assessment. No significant between-group differences were observed in time spent outside, photoprotection outside, mood or confidence. The delayed-intervention control showed improvements in UVR dose to face (adjusted difference = –0.05; Hedge’s g = –0.1), time outside (adjusted difference = –69.9; Hedge’s g = –0.28) and photoprotection (adjusted difference = –0.23, Hedge’s g = 0.45) after receiving XPAND (June 2019 assessment). XPAND was associated with lower treatment costs [–£2642; 95% confidence interval (CI) –£8715 to £3873] and fewer QALYs (–0.0141; 95% CI –0.0369 to 0.0028).

Conclusions
XPAND was associated with a lower UVR dose to face in patients with XP and was cost-effective.
Intrinsic functional defects in B cells of patients with NFKB2 mutations
Authors: Min Qing , Li Yaxuan , Wu Xuzhe , Yu Meiping , Ying Wenjing , Zhou Qinhua , Hou Jia , Sun Bijun , Hui Xiaoying , Dong Lulu , Meng Xin , Zhang Hai , Hu Ziying , Feng Xiaoqian , Sun Jinqiao , Wang Wenjie , Wang Xiaochuan , Wang Ji-Yang 
Published: October 2024

Abstract
Mutations in the human nuclear factor-κB2 gene (NFKB2) are associated with common variable immunodeficiency (CVID) or combined immunodeficiency diseases (CID), characterized by B-cell lymphopenia, hypogammaglobulinemia, and T-cell dysfunction. This study investigated whether B cells with NFKB2 mutations exhibit intrinsic impairments in activation, class-switch recombination, and differentiation. We analyzed five patients from four unrelated families with CVID, each carrying a heterozygous NFKB2 mutation: P1 (C.2595_2614del, p.A867Gfs*12), P2 (C.2597G &gt; A, p.S866N), P3 (C.2540dupT, p.R848Efs*38), and P4 and P5 (C.2570_2571insCAGCACA, p.A860Qfs*28). The patients with frameshift mutations (P1, P3, P4, and P5) exhibited truncated proteins detectable in their peripheral blood mononuclear cells, while P2 had a missense mutation. All identified mutations disrupted the processing of p100 into the active p52 form, resulting in NF-κB2 loss of function and IκBδ gain of function. Clinically, P1, P2, and P3 exhibited B-cell lymphopenia, and all five patients presented with hypogammaglobulinemia. Notably, P2 exhibited a markedly low B-cell count, associated with increased proportions of memory B and IgD−CD27− double-negative B cells. In vitro experiments with naïve B cells from P1 and P4 demonstrated decreased survival, impaired activation, and reduced differentiation into CD27+IgD− cells and plasmablasts, while class-switch recombination was unaffected. These findings reveal novel B-cell intrinsic functional defects in patients with NFKB2 mutations.
Emergency department use of a high-sensitivity point-of-care troponin assay reduces length of stay: an implementation study preliminary report
Authors: Pickering John W , Joyce Laura R , Florkowski Christopher M , Buchan Vanessa , Hamill Laura , Than Martin P 
Published: October 2024

Abstract

Aims
Point-of-care (POC) high-sensitivity troponin (hs-cTn) assays within a clinical pathway may safely reduce length of stay (LoS) for patients presenting to the emergency department (ED) with possible acute myocardial infarction (AMI). In this early report, we present the first evaluation of a POC hs-cTn in real-life care.

Methods and results
In adult patients presenting to ED investigated for possible AMI, we compared the LoS in patients assessed with a troponin in the 8 weeks before (usual-care phase) and the 8 weeks following introduction of the Siemens Atellica VTLi POC hs-cTnI for decision-making (intervention phase). The VTLi replaced the laboratory (Beckman Coulter) assay as the default hs-cTn test within the clinical pathway. This was the only change to the pathway process. The safety outcome was first event AMI or cardiac death within 30 days. There were 2376 presentations in the usual-care phase with 188 individuals with AMI and 2392 in the intervention phase with 198 AMI. In the intervention phase, there was a mean (95% CI) reduction in LoS of 32 min (22–41 min) compared with the usual-care phase. This represents 21.4 fewer patient-hours in the ED each day (1196 in the 8-week period). In both phases, the pathway correctly identified all cases of AMI at index attendance. There were four follow-up events (two usual-care, two intervention) within 30 days.

Conclusion
The deployment of a hs-cTn POC analyser into a large ED safely reduced length of stay. If translatable to other EDs, this could represent an important advancement to patient care.

Trial registration
Australia New Zealand Clinical Trials Registry, No. ACTRN12619001189112.